博士（獣医学）, 北海道大学

バイオロギング

コウモリ

マールブルグウイルス

表面糖蛋白質

抗体

エボラウイルス

出芽

フィロウイルス

ライフサイエンス, 獣医学

博士課程, 国際感染症学院

2019年01月, 8th NSV-Jシンポジウム, ベスト・オーラル・プレゼンテーション賞
ザンビアのコウモリにおけるウイルス保有調査
梶原将大;Changula Katendi;宮本洋子;邱永晋;播磨勇人;Hang’ombe Bernard;Mweene Aaron;髙田礼人

2019年01月, 8th NSV-Jシンポジウム, トラベルアワード
ザンビアのコウモリにおける ウイルス保有調査
梶原将大;Changula Katendi;宮本洋子;邱永晋;播磨勇人;Hang’ombe Bernard;Mweene Aaron;髙田礼人

2011年09月, The 3rd International Young Researcher Seminar for Zoonosis Control, Best Poster Presentation
An H5N1 highly pathogenic avian influenza virus that invaded Japan through waterfowl migration
Masahiro Kajihara;Keita Matsuno;Edgar Simulundu;Mieko Muramatsu;Osamu Noyori;Rashid Manzoor;Eri Nakayama;Manabu Igarashi;Masatoshi Okamatsu;Yoshihiro Sakoda;Hiroshi Kida;Ayato Takada

Molecular diversity of tick-associated Rickettsia species in multiple provinces of Zambia.
Yongjin Qiu; Martin C Simuunza; Masahiro Kajihara; Joseph Ndebe; Yoshiki Eto; Akina Mori-Kajihara; Edgar Simulundu; Walter Muleya; Hayato Harima; Katendi Changula; Ngonda Saasa; Penjaninge Kapila; Ladslav Moonga; Evans Mulenga; Andrew Nalishuwa Mukubesa; Bernard Mudenda Hang'ombe; Ayato Takada; Nariaki Nonaka; Hirofumi Sawa; Ryo Nakao
The Journal of veterinary medical science, 2026年06月22日, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）, Ticks harbor diverse Rickettsia spp., including several spotted fever group rickettsiae of veterinary and public health importance. However, information on tick-associated Rickettsia spp. in Zambia remains geographically limited. In this retrospective molecular survey, we examined 1,587 individual ticks collected from vegetation and domestic animals in eight provinces of Zambia during 2016-2018. Ticks were identified morphologically and screened for Rickettsia spp. by PCR targeting the citrate synthase gene (gltA). Representative gltA positive samples were further characterized by sequencing of ompA, ompB, htrA, sca4, and the 16S rRNA gene. In total, 204 ticks (12.9%) were tested positive for gltA and detected in all eight provinces examined. Sequencing of gltA revealed 26 sequence variants, of which 25 clustered within the spotted fever group and one belonged to transitional group rickettsiae. Multilocus phylogenetic analysis identified sequence variants related to recognized or putative tick-borne rickettsiae, including R. africae, R. conorii, R. massiliae, R. aeschlimannii, Candidatus R. jingxinensis, Candidatus R. rhabdomydis, and R. hoogstraalii. These findings indicate that ticks in Zambia harbor genetically diverse Rickettsia lineages and provide a geographically broad molecular baseline for future surveillance, isolation attempts, and genomic characterization of tick-borne rickettsiae in the region.

Characterization of inner capsid σA protein as a virulence factor of the pteropine orthoreovirus.
Hayato Harima; Michihito Sasaki; Takuma Ariizumi; Nijiho Kawaguchi; Hiroko Kobayashi; Kittiya Intaruck; Takeshi Kobayashi; Takahiro Kawagishi; Yuta Kanai; Naganori Nao; Yongjin Qiu; Masahiro Kajihara; Yasuko Orba; Naoto Ito; Kanako Ishihara; Masayuki Saijo; William W Hall; Bernard M Hang'ombe; Hirofumi Sawa
PLoS pathogens, 22, 5, e1014252, 2026年05月26日, ［国際誌］
英語, 研究論文（学術雑誌）, Pteropine orthoreovirus (PRV) is an emerging zoonotic virus that causes pneumonia in humans. We previously isolated the PRV strain Nachunsulwe-57 (N57) from a Zambian fruit bat and demonstrated its low virulence in laboratory mice. Here, we have attempted to identify factors responsible for differences in the virulence between strain N57 and the human-derived clinical strain Miyazaki-Bali/2007 (MB). Characterization of the virulence of recombinant monoreassortant PRVs derived from highly virulent MB and low virulent N57 strains in mice revealed that compared with wild-type (WT) MB, MB-based monoreassortants carrying the L1, S1, or S2 segment from N57 exhibited attenuated virulence. Among these, the monoreassortants carrying the S1 or S2 segment exhibited reduced viral loads and reduced cytokine gene expression levels in the lungs. Genetic mapping of virulence determinants using the reciprocal monoreassortant viruses with increased virulence demonstrated that N57-based monoreassortants carrying the S1 or S2 segment of MB exhibited enhanced virulence, resulting in lower survival rate compared with WT N57. Unlike the S1 segment, the functions of the S2 segment in pathogenesis are unclear. Thus, we further investigated the functional region of the inner-capsid σA protein encoded by the S2 segment. Notably, Ser-46 of σA was identified as a key amino acid determinant of PRV virulence and is present in strains derived from humans, monkeys, and bat flies, but not those identified from bats (their natural host). Collectively, these findings demonstrate that PRV σA is one factor that regulates virulence, and that σA Ser-46 may be related to potential interspecies transmission events from bats.

Seroprevalence of Crimean-Congo Hemorrhagic Fever Virus Infection in Humans and Domestic Ruminants, Democratic Republic of the Congo
Boniface Pongombo Lombe; Yannick Munyeku-Bazitama; Gracia Kashitu-Mujinga; Patrick Kakoni Mukadi; Honoré Mampasi; Curé Georges Mbuyi Tshilenge; Marguerite Pemba Manwana; Noël Mwayakala; Niclette Bibi Zenga; Patient Okitale-Talunda; Elisabeth Pukuta-Simbu; Suguru Taga; Gaston Sefu Amzati; Masahiro Kajihara; Pierre Nsele Mutantu; Jean-Jacques Muyembe-Tamfum; Steve Ahuka-Mundeke; Justin Masumu; Sheila Makiala-Mandanda; Ayato Takada
Emerging Infectious Diseases, 32, 4, 543, 552, Centers for Disease Control and Prevention (CDC), 2026年04月, ［査読有り］
英語, 研究論文（学術雑誌）

Detection of carbapenem resistance among third-generation cephalosporin-resistant Enterobacterales from small-scale poultry farms in peri-urban Lusaka, Zambia
Situmbeko J. Nasilele; Misheck Shawa; Harvey K. Kamboyi; Bruno S. J. Phiri; Tapiwa Lundu; Mike Nundwe; Angela Lungu; Ladslav Moonga; Shohei Ogata; Masahiro Kajihara; Hirofumi Sawa; Yasuhiko Suzuki; Hideaki Higashi; Ntombi B. Mudenda; Mudenda B. Hang’ombe; Kaampwe Muzandu
Access Microbiology, 8, 1, 543, 552, Microbiology Society, 2026年01月01日, ［査読有り］
英語, 研究論文（学術雑誌）, Background. Carbapenem and third-generation cephalosporin (3GC) resistance among Gram-negative bacteria poses a serious threat to human and animal health. This study aimed to identify and characterize carbapenem- and 3GC-resistant Enterobacterales isolated from poultry in Lusaka Province, Zambia.

Methods. Ninety pooled cloacal samples were collected from market-ready broiler chickens in the Chongwe and Chilanga districts of Lusaka Province. The isolates were screened for 3GC and carbapenem resistance using the disc diffusion and broth microdilution methods. PCR and Sanger sequencing were performed for species identification and detection of β-lactamase-encoding ( bla ) genes, including bla _CTX-M , bla _TEM , bla _OXA-1 and bla _SHV . Hierarchical clustering was used to assess phenotypic and genotypic relationships.

Results. A total of 83 3GC-resistant Gram-negative isolates were recovered, of which 12% were also carbapenem resistant. Escherichia coli was the most prevalent species, followed by Klebsiella pneumoniae and Enterobacter spp., then Pseudomonas aeruginosa, other Pseudomonas spp., Acinetobacter baumannii , Citrobacter freundii and Aeromonas caviae . Multidrug resistance occurred in 84.3% of the isolates, with the highest resistance to ampicillin, tetracycline and co-trimoxazole. Overall, 80.7% of the isolates harboured at least one of the four tested bla genes, with bla _CTX-M and bla _TEM being the most common. Hierarchical clustering revealed that isolates from both districts shared similar phenotypic and genotypic resistance patterns.

Conclusions. The presence of multidrug- and carbapenem-resistant Enterobacterales from poultry highlights the emergence of carbapenem resistance in Zambia’s food production sector. The detection of imipenem-resistant isolates indicates the potential for transmission of resistance genes between animals and humans. These findings underscore the need for prudent antimicrobial use, strengthened stewardship and a One Health surveillance approach to contain the spread of carbapenem resistance genes.

Presence or Emergence of Canine Leishmaniasis, Malawi
Boniface Chikufenji; Kyoko Hayashida; Yasuyuki Goto; Tatsuki Sugi; Chizu Sanjoba; Chrispin Njala; Inga McDermott; Frederic Lohr; Dagmar Mayer; Shohei Ogata; Masahiro Kajihara; Naganori Nao; Ryo Nakao; Laston Chimaliro; Donales Kapira; Janelisa Musaya; Junya Yamagishi; Elisha Chatanga
Emerging Infectious Diseases, 32, 1, Centers for Disease Control and Prevention (CDC), 2026年01月
研究論文（学術雑誌）

Oita virus rediscovered after 50 years: isolation of genetically conserved strains from bats in Southern Japan
Saiko Sawai; Kittiya Intaruck; Sho Sata; Mana Esaki; Kimitake Funakoshi; Shin Murakami; Keita Matsuno; Ryo Nakao; Kazunori Kimitsuki; Takaaki Yahiro; Akira Nishizono; Naganori Nao; Yasuko Orba; Ayato Takada; Masahiro Kajihara; Makoto Ozawa; Kosuke Okuya
Microbiology Spectrum, American Society for Microbiology, 2025年11月11日
研究論文（学術雑誌）, ABSTRACT

Bats are natural reservoirs of numerous viruses, including members of the Rhabdoviridae family. Oita virus (OITV), classified within the genus Ledantevirus , was first isolated from the blood of a Japanese horseshoe bat ( Rhinolophus cornutus ) in the Oita Prefecture, Japan, in 1972. However, since then, no other OITV isolates have been reported. In this study, we isolated two OITV strains—OITV 321/2022 and OITV 326/2022—from oral swab samples of R. cornutus collected in the Katano Cave, Kagoshima Prefecture, Japan. Genetic analysis revealed >98% nucleotide identity with the previous isolate, OITV 296/1972. Phylogenetic analysis confirmed their classification within the Ledantevirus subgroup C. We assessed viral growth kinetics using 17 cell lines from diverse animal species, including bats, rodents, humans, and arthropods. OITV 321/2022 showed efficient replication in primate-derived, bat-derived, and human-derived cells, but not in tick or mosquito cell lines. Experimental infection of BALB/c mice demonstrated that OITV 321/2022 induced no overt clinical signs via either the intracerebral or intranasal routes. However, systemic infection and viral replication in the brain and lungs were observed by intracerebral inoculation. Intranasal inoculation with OITV 321/2022 and OITV 296/1972 resulted in transient pulmonary infection without systemic dissemination. This study provides evidence of OITV circulation in wild bat populations over 50 years, highlighting the need for continued surveillance of bat-associated rhabdoviruses in Japan.

IMPORTANCE

Bats are known reservoirs of zoonotic viruses, and their proximity to humans raises concerns regarding zoonotic risks. We report the first isolation of the Oita virus (OITV), a bat-associated ledantevirus, that has circulated for over 50 years. Unlike a previous blood-derived isolate, our isolates were obtained from oral swabs, suggesting their potential for respiratory transmission. OITV could infect a wide range of mammalian cells, including human-derived cells, and induce systemic infection in mice without clinical symptoms. These findings indicate that OITV possesses a broad host tropism and may circulate among microbats through the respiratory tract. Although the pathogenicity of the newly isolated strain appears to be attenuated compared with that of a historical brain-passaged strain, its ability to replicate in human cells underscores its potential zoonotic relevance, necessitating active surveillance and functional characterization of bat-derived rhabdoviruses to better assess emerging infectious disease threats.

A high prevalence of dogs seropositive to Leishmania in Zambia.
Herman M Chambaro; Kyoko Hayashida; Lavel C Moonga; Misheck Shawa; Walter Muleya; Joseph Chizimu; David Squarre; Tatsuki Sugi; Junya Yamagishi; Shohei Ogata; Masahiro Kajihara; Hirofumi Sawa; Chizu Sanjoba; Enala T Mwase; Roma Chilengi; Gilbert H Munsaka; Kelvin L Sarenje; Namwiinga R Mulunda; Mable M Mutengo; Boniface Namangala; Yasuyuki Goto
Parasitology international, 108, 103081, 103081, 2025年10月, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）, Domestic dogs are the main reservoir of Leishmania infantum, a causative agent of visceral leishmaniasis in humans. Although the disease is widespread in the world, the burden of visceral or any other disease form of leishmaniasis is poorly documented in Zambia, largely due to lack of surveillance. Recently, three cases of autochthonous canine leishmaniasis (CanL) were reported in Zambia following decades of presumed disease absence. This finding intimated probable disease emergence, raising the need for the identification of infection foci. Thus, in this study, we conducted the first mass serological survey for Leishmania infections in domestic dogs from two densely populated urban areas of Zambia in July 2022. In some of the study sites, seropositivity was up to ∼17 %, suggesting probable presence of Leishmania transmission hot spots. Moreover, on follow-up surveys of seropositive dogs, presence of antileishmanial antibodies was a risk factor for dog survival (relative risk = 7.9; odds ratio = 42.5). Our findings implies that Leishmania infection can be a health issue in domestic dogs in Zambia. Considering the risk of zoonotic transmission, the need for improved disease diagnosis and surveillance in both humans, dogs and sand fly vectors is highlighted in Zambia.

In silico characterization of chromosomally integrated blaCTX-M genes among clinical Enterobacteriaceae in Africa: insights from whole-genome analysis
Misheck Shawa; Herman Chambaro; Harvey K. Kamboyi; Clement Sulwe; Joseph Y. Chizimu; Situmbeko J. Nasilele; Shohei Ogata; Mulemba Samutela; Tuvshinzaya Zorigt; Steward Mudenda; Manyando Simbotwe; Mwamba Nsofwa; Jedidiah Chanda; Freeman Chabala; Mike Nundwe; Joseph Ndebe; Msangwa Sinjani; Kyoko Hayashida; Naganori Nao; Roma Chilengi; Hirofumi Sawa; Yasuhiko Suzuki; Bernard Hang'ombe; Masahiro Kajihara; Hideaki Higashi
Frontiers in Microbiology, 16, Frontiers Media SA, 2025年09月12日
研究論文（学術雑誌）, Antimicrobial resistance (AMR) mediated by extended-spectrum β-lactamases (ESBLs) is a growing global concern, particularly among Enterobacteriaceae. The CTX-M-type ESBLs, encoded by the bla_CTX-M gene, are of significant public health importance due to their high prevalence and broad geographic distribution. Typically located on plasmids and often co-occurring with other AMR genes, bla_CTX-M contributes to multidrug resistance (MDR). However, increasing evidence suggests secondary chromosomal integration of bla_CTX-M, sometimes alongside other resistance determinants. The extent and implications of this mechanism remain poorly characterized, especially in Africa, where genomic surveillance is limited. In this study, we retrieved 295 chromosomal sequences of Enterobacteriaceae of African origin from the GenBank and performed in silico predictions of bla_CTX-M and other AMR genes. bla_CTX-M-carrying sequences were further characterized by in silico multilocus sequence typing and genome annotation. Chromosomal insertions were identified through alignment with reference genomes. Overall, 47 of 295 sequences (15.9%) harbored the bla_CTX-M gene, with the highest prevalence in Klebsiella pneumoniae (29/157, 18.5%), followed by Escherichia coli (13/72, 18.1%), Enterobacter spp. (4/38, 10.5%), and Shigella spp. (1/12, 8.3%). The most common allele was bla_CTX-M-15 (31/47, 66.0%), followed by bla_CTX-M-14 (12/47, 25.5%), bla_CTX-M-55 (3/47, 6.4%), and bla_CTX-M-27 (1/27, 3.7%). Co-occurrence of bla_CTX-M with additional AMR genes was frequently observed, with integration events often associated with mobile genetic elements such as ISEcp1 and IS26. Notably, strains from the same hospital setting were phylogenetically related and shared sequence types and AMR gene profiles, suggesting local clonal dissemination. These findings reveal a notable presence of chromosomally integrated bla_CTX-M among African Enterobacteriaceae, frequently in association with other resistance genes, thereby facilitating stable MDR propagation independent of plasmid maintenance. This evolutionary adaptation may have significant implications for the persistence and spread of MDR in clinical settings.

Mosquito-borne alphaviruses in Zambia: Isolation and characterization of Eilat and Sindbis viruses.
Chadwic De'Sean Mears; Koshiro Tabata; Takuma Ariizumi; Bernard M Hang'ombe; Yongjin Qiu; Hayato Harima; Masahiro Kajihara; William W Hall; Michihito Sasaki; Hirofumi Sawa; Yasuko Orba
Virus research, 358, 199604, 199604, 2025年08月, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）, Alphaviruses in the family Togaviridae include zoonotic arthropod-borne viruses, including Sindbis virus (SINV), chikungunya virus, as well as insect-specific viruses such as Eilat virus (EILV). Previous investigations of alphaviruses in Zambia have identified a novel insect-specific alphavirus, Mwinilunga alphavirus in mosquitoes. Further ongoing surveillance resulted in the isolation of EILV and SINV for the first time in Zambia. Here, these alphaviruses were characterized in terms of growth kinetics in cells, and molecular phylogenetic relatedness to other alphaviruses. Zambian EILV (strain zmq19_M44) exhibited a close phylogenetic relationship with other insect-specific alphaviruses and shared a close nucleotide identity to those of EILV isolate (90.4 %) and Mwinilunga alphavirus (75.5 %). EILV zmq19_M44 attained a saturating titer in C6/36 cells at 6-8-days post infection but was unable to replicate in mammalian cells. Phylogenetic analysis revealed the Zambian SINV (strain zmq17_M115) belongs in Clade D of SINV Genotype 1 along with the Kenyan isolate BONI 584 from Central Africa. The growth of the SINV zmq17_M115 was comparable to that of the prototype SINV strain AR339 in mammalian cells but was statistically different in insect cells. Our findings will contribute to public health measures for the control of alphaviral diseases in Zambia.

Occurrence of cefotaxime-resistant and beta-lactamase-producing Enterobacterales in poultry from small-scale farms in the Copperbelt province of Zambia.
Situmbeko J Nasilele; Misheck Shawa; Harvey K Kamboyi; Bruno S J Phiri; Herman Chambaro; Tapiwa Lundu; Mike Nundwe; Angela Lungu; Ladslav Moonga; Masahiro Kajihara; Hirofumi Sawa; Yasuhiko Suzuki; Hideaki Higashi; Amon Siame; Ntombi B Mudenda; Mudenda B Hang'ombe; Kaampwe Muzandu
JAC-antimicrobial resistance, 7, 4, dlaf125, 2025年08月, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）, BACKGROUND: The inappropriate use of antimicrobials in poultry farming is associated with the emergence of antimicrobial-resistant Enterobacterales. This cross-sectional study aimed to identify various cefotaxime-resistant and β-lactamase-producing Enterobacterales and characterize their antimicrobial resistance profiles. METHODS: Pooled cloacal and meat samples collected from market-ready broiler chickens in Kitwe and Ndola districts of Zambia were screened for cefotaxime-resistant Enterobacterales. The samples were inoculated on MacConkey agar supplemented with 1 mg/L cefotaxime. The cefotaxime-resistant isolates were further subjected to antimicrobial susceptibility tests. Further, the isolated cefotaxime-resistant Enterobacterales were analysed for bla CTX-M, bla TEM, bla OXA-1-like and bla SHV genes using PCR and Sanger sequencing. RESULTS: From a total of 114 pooled samples, 81 (71.1%) cefotaxime-resistant Gram-negative strains were isolated. These were dominated by Escherichia coli (77.8%) followed by Klebsiella pneumoniae (6.2%), Pseudomonas spp. (6.2%), Acinetobacter baumannii (4.9%), Pseudomonas aeruginosa (2.5%), Enterobacter spp. (1.2%) and Comamonas aquatica (1.2%). Furthermore, 64.2% of the 81 isolates exhibited multidrug resistance with high resistance (>64%) to ampicillin, co-trimoxazole and tetracycline. The results also showed that 66.7% of the isolates harboured at least one of the four tested bla genes (bla CTX-M, bla TEM, bla OXA-1-like and bla SHV), with the commonest being bla CTX-M (58%) and bla TEM (45.7%). CONCLUSIONS: The study revealed a high prevalence of cefotaxime-resistant Enterobacterales and multidrug resistance involving medically important antibiotics. Four bla genes (bla CTX-M, bla TEM, bla OXA-1-like and bla SHV) were identified. Our results highlight the need to strengthen antimicrobial stewardship programmes and optimize antimicrobial use in poultry farming.

Genetic diversity of sarbecoviruses isolated from microbats in Southern Japan.
Kosuke Okuya; Sho Sata; Mana Esaki; Isshu Kojima; Masahiro Kajihara; Shin Murakami; Takuji Sakata; Kimitake Funakoshi; Makoto Ozawa
Virology journal, 22, 1, 226, 226, 2025年07月08日, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, BACKGROUND: Bat-derived sarbecoviruses, members of the Betacoronavirus genus, such as severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), pose a zoonotic threat. While sarbecoviruses have been isolated from bats in eastern Japan, their distribution and diversity in southern Japan remain poorly understood. We investigated the genetic characteristics and zoonotic potential of bat-derived sarbecoviruses from Rhinolophus cornutus in the Kagoshima and Kumamoto Prefectures, Japan. METHODS: We collected 133 oral swab samples from R. cornutus between 2022 and 2025. The samples were inoculated into various Vero cell derivatives to isolate sarbecoviruses. The virus genomes were sequenced and subjected to phylogenetic analyses. Viral replication was assessed in vitro. Cross-species potential was evaluated in vivo through experimental infection in Syrian hamsters. RESULTS: Six and three sarbecoviruses from the Kagoshima and Kumamoto Prefectures, respectively, were isolated. Genetic analyses revealed substantial diversity among our isolates (95.98–99.95% nucleotide identity). Phylogenetic analyses revealed clustering of the isolates separately from those found in eastern Japan, indicating regional genetic divergence among bat populations. Our isolates exhibited deletions in the receptor-binding domain of the spike protein. These deletions attenuate the viral binding affinity to human angiotensin-converting enzyme 2 (ACE2) receptors. This suggests a limited capacity of our isolates to bind human ACE2. Our isolates exclusively replicated in Vero cells expressing R. cornutus-derived ACE2 (RcACE2). Syrian hamsters, a model for SARS-CoV-2 infection, were not susceptible to infection from our isolates, suggesting limited potential for cross-species transmission. CONCLUSIONS: We characterized genetically diverse sarbecoviruses circulating in R. cornutus populations in southern Japan. Although the isolates appear to have a low immediate zoonotic risk, they likely represent only a small fraction of the sarbecoviruses circulating among bats. Given the potential presence of viruses with greater zoonotic capacity, continuous surveillance is critical for early detection of emerging threats and advancing our understanding of coronavirus ecology in bat populations.

Development of a PCR-dipstick DNA chromatography-based tool for the detection of CTX-M- and TEM-producing Escherichia coli and Klebsiella pneumoniae isolated from patients in Kafue and Katete districts of Zambia.
Misheck Shawa; Kyoko Hayashida; Naganori Nao; Atmika Paudel; Harvey Kamboyi; Herman Chambaro; Malala Mulavu; Cynthia Sipho Khumalo; Mike Nundwe; Tuvshinzaya Zorigt; Chinatsu Nakamura; Yongjin Qiu; Naoko Kawai; Maisa Kasanga; Joseph Chizimu; Lavel Moonga; Joseph Ndebe; Manyando Simbotwe; Shohei Ogata; Mulemba Samutela; Chie Nakajima; Roma Chilengi; Mable Mutengo; Masahiro Kajihara; Hirofumi Sawa; Bernard Hang'ombe; Yasuhiko Suzuki; Hideaki Higashi
BMC infectious diseases, 25, 1, 541, 541, 2025年04月16日, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）, In Zambia, 40% of clinical Gram-negative bacteria are either Escherichia coli or Klebsiella pneumoniae, with a high third-generation cephalosporin (3GC) resistance prevalence. Therefore, 3GC resistance surveillance is a crucial indicator for guiding focused intervention policies. However, the lack of genotypic diagnostic tools limits the ability to elucidate trends, especially in peri-urban and rural areas of developing countries. This study aimed to develop a rapid, cost-effective tool for the genotypic surveillance of 3GC resistance. Here, 900 stool samples collected from patients in Kafue (peri-urban, n = 400) and Katete (rural, n = 500) districts of Zambia were used for bacterial isolation on MacConkey agar supplemented with 1 μg/ml cefotaxime. Isolated 3GC-resistant strains were characterized by sequencing the 16S rRNA gene and screening for blaCTX-M and blaTEM genes using single polymerase chain reaction (PCR). Furthermore, selected 3GC-resistant strains were subjected to whole-genome sequencing (WGS) using MiSeq/HiSeq (n = 34) and MinION (n = 1). Using the data from this and other previous studies, we developed a rapid PCR-dipstick DNA chromatography-based tool for detecting blaCTX-M, blaTEM, E. coli-specific yaiO, and K. pneumoniae-specific khe genes. The prevalence of isolated 3GC resistant strains was 15.4% (139/900), dominated by E. coli (102/139, 73.4%). On PCR, the blaCTX-M gene was detected in 72.7% (101/139) of the isolates, while blaTEM was found in 46.8% (65/139) of the strains. The developed tool displayed a high level of agreement with WGS and single PCR/Sanger sequencing, with sensitivity and specificity ≥ 95% and Kappa ≥ 0.95 for each of the four target genes. We envisage that the simplicity and adaptability of this tool will be a significant advantage for the surveillance of 3GC resistance in Zambia and elsewhere.

Evidence of multiple bacterial, viral, and parasitic infectious disease agents in Mastomys natalensis rodents in riverine areas in selected parts of Zambia.
Samuel Munalula Munjita; Annie Kalonda; Benjamin Mubemba; Manu Vanaerschot; Cristina Tato; Lusajo Mwakibete; John Tembo; Simbarashe Chitanga; Katendi Changula; Masahiro Kajihara; Walter Muleya; Hirofumi Sawa; Ayato Takada; Matthew Bates; Sody Munsaka; Edgar Simulundu
Infection ecology & epidemiology, 15, 1, 2441537, 2441537, 2025年, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）, BACKGROUND: Infectious disease agents pose significant threats to humans, wildlife, and livestock, with rodents carrying a third of these agents, many linked to human diseases. However, the range of pathogens in rodents and the hotspots for disease remain poorly understood. AIM: This study evaluated the prevalence of viral, bacterial, and parasitic pathogens in Mastomys natalensis rodents in riverine and non-riverine areas in selected districts in Zambia. METHODS: The study applied metagenomic next generation sequencing (mNGS). Tissues analysed included semen, foetal tissues, and blood-rich organs (liver, spleen, kidneys, and lungs). A multivariate logistic regression model explored the relationship between pathogen presence and host or ecological factors. RESULTS: A total of 182 rodents were captured, and 14 pathogens were detected in 10.4% of the samples (19/182). Detected organisms included zoonoses (Klebsiella michiganensis, Klebsiella pneumoniae, Salmonella enterica, and Bartonella elizabethae); Emerging zoonoses (Elizabethkingia miricola, Klebsiella variicola, Bartonella tribocorum, and Cardiovirus B); among others (Eimeria papillata etc). Riverine areas showed higher odds of pathogen presence (OR = 8.45; p < 0.001; 95% CI: 3.07-23.26). CONCLUSION: These results suggest that M. natalensis harbours multiple infectious agents with zoonotic potential, and riverine regions may be key hotspots for rodent-borne pathogens in Zambia.

Evidence of human exposure and associated risk factors to Rift Valley fever in selected districts of Central and Western Zambia.
Chilufya C Kasongamulilo; Michelo Syakalima; Ngondo Saasa; Henson Kainga; Girja S Pandey; Andrew N Mukubesa; Innocent Mwape; Masahiro Kajihara; Ayato Takada; Martin Simuunza
PloS one, 20, 1, e0309288, 2025年, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）, Rift Valley fever (RVF) is an important viral zoonotic disease that not only affects ruminants but causes serious morbidity and mortality in humans. In humans, its symptoms range from mild flu-like signs to a severe form such as retinal damage, meningoencephalitis to haemorrhagic fever. In this study, 202 human serum samples were collected from central and western parts of Zambia and tested for RVF-specific antibodies using a commercially available ELISA kit. A pre-tested semi-structured questionnaire was used to collect data for determining hypothesised risk factors for exposure to RVF. The participants enrolled in this study, were only those who are at high risk of RVF infection and were in close contact with animals and animal products. The study revealed an overall seropositivity of 9.90%, occupationally distributed as 16.67% among slaughter house workers, 14.41% among livestock farmers, and 0% among the others (i.e., students, butchery, and farm workers). The prevalence was highest (19.23%) in Sesheke district found in the western part of Zambia, while Chisamba district in central Zambia had the lowest prevalence (1.41). Movement of animals in search of greener pastures was identified as a risk factor to being RVF-seropositive. This suggests that there was silent circulation of the virus in the interepidemic period in the study areas. Therefore, this study recommends that public education of livestock farmers, public health workers, slaughterhouse workers, communities, livestock and veterinary staff needs to be enhanced to increase awareness and preparedness for RVF outbreak in Zambia.

First identification and whole genome characterization of rotavirus C in pigs in Zambia.
Hayato Harima; Yongjin Qiu; Michihito Sasaki; Joseph Ndebe; Kapila Penjaninge; Edgar Simulundu; Masahiro Kajihara; Aiko Ohnuma; Keita Matsuno; Naganori Nao; Yasuko Orba; Ayato Takada; Kanako Ishihara; William W Hall; Bernard M Hang'ombe; Hirofumi Sawa
Virology, 603, 110385, 110385, 2024年12月31日, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）, Rotavirus C (RVC) causes acute gastroenteritis in neonatal piglets. Despite the clinical importance of RVC infection, the distribution and prevalence in pig populations in most African countries remains unknown. In this study, we identified RVC in Zambian pigs by metagenomic analysis. The full genome sequence of the RVC revealed two different VP4 sequences, implying that two different RVC strains (ZP18-77-c1 and ZP18-77-c2) were present in the same sample. Genetic analyses demonstrated that all segments of ZP18-77-c1 and ZP18-77-c2 showed high nucleotide sequence identities (87.7-94.5%) to known porcine RVC strains, and ZP18-77-c1 and ZP18-77-c2 strains were assigned to genotype constellations, G1-P[4]/P[14]-I13-R5-C5-M1-A7-N9-T10-E5-H1. We further screened RVC genomes among pig feces collected in Zambia (n = 147) by RT-qPCR, and 78 samples (53.1%) were positive. This study demonstrated the first full genome sequence of African RVC strains with a relatively high prevalence of RVC infection in the pig populations in Zambia.

Development of a seroepidemiological tool for bat-borne and shrew-borne hantaviruses and its application using samples from Zambia
Rakiiya Sikatarii Sarii; Masahiro Kajihara; Zuoxing Wei; Sithumini M. W. Lokpathirage; Devinda S. Muthusinghe; Akina Mori-Kajihara; Katendi Changula; Yongjin Qiu; Joseph Ndebe; Bernard M. Hang’ombe; Fuka Kikuchi; Ai Hayashi; Motoi Suzuki; Hajime Kamiya; Satoru Arai; Ayato Takada; Kumiko Yoshimatsu
PLOS Neglected Tropical Diseases, 18, 11, e0012669, e0012669, Public Library of Science (PLoS), 2024年11月21日, ［査読有り］, ［国際共著］, ［国際誌］
研究論文（学術雑誌）, Background

Rodent-borne orthohantaviruses are the causative agents of hemorrhagic fever with renal syndrome and hantavirus pulmonary syndrome. Apart from the classic rodent-borne hantaviruses, numerous species of hantaviruses have been identified in shrews and bats; however, their antigenicity and pathogenicity are unknown. This study focused on developing a serological method to detect antibodies against bat- and shrew-borne hantaviruses.

Methodology/Principal findings

Five bat-borne (Brno, Dakrong, Quezon, Robina, and Xuan Song) and 6 shrew-borne (Asama, Altai, Cao Bang, Nova, Seewis, and Thottapalayam) viruses were selected based on the phylogenetic differences in their N proteins. The recombinant N (rN) proteins of these viruses were expressed as antigens in Vero E6 and 293T cell lines using the pCAGGS/MCS vector. Antisera against the Nus-tagged rN fusion proteins of these viruses (mouse anti-Brno, Dakrong, Quezon, Robina, Xuan Song, Asama, Cao Bang, and Nova, while rabbit anti-Altai, Seewis and Thottapalayam) were also generated. Antigenic cross-reactivity was examined in antisera and rN-expressing Vero E6 cells. The rN proteins of almost all the tested viruses, except for the Quezon and Robina viruses, showed independent antigenicity. For serological screening of bat samples, 5 rNs of the bat-borne viruses were expressed together in a single transfection protocol. Similarly, 6 rNs of shrew-borne viruses were expressed. Reactivities of the mixed antigen system were also examined across the singly transfected Vero cell lines to ensure that all antigens were expressed. Using these antigens, bat serum samples collected from Zambia were screened using the indirect immunofluorescence antibody test (IFAT). Selected positive samples were individually tested for the respective antigens by IFAT and western blot assays using rN-expressing 293T cell lysates. Of the 1,764 bat serum samples tested, 11.4% and 17.4% were positive for bat and shrew mixed antigens, respectively. These samples showed positive reactions to the Brno, Dakrong, Quezon, Xuan Son, Robina, Asama, Altai, Cao Bang, or Thottapalayam virus antigens.

Conclusions/Significance

These observations suggest that the mixed-antigen screening system is useful for serological screening For Orthohantavirus infections and that bats in Zambia are likely exposed to not only bat-borne hantaviruses but also to shrew-borne hantaviruses.

A first report of rotavirus B from Zambian pigs leading to the discovery of a novel VP4 genotype P[9].
Hayato Harima; Yongjin Qiu; Michihito Sasaki; Joseph Ndebe; Kapila Penjaninge; Edgar Simulundu; Masahiro Kajihara; Aiko Ohnuma; Keita Matsuno; Naganori Nao; Yasuko Orba; Ayato Takada; Kanako Ishihara; William W Hall; Bernard M Hang'ombe; Hirofumi Sawa
Virology journal, 21, 1, 263, 263, 2024年10月24日, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）, BACKGROUND: Rotavirus B (RVB) causes diarrhea in humans and pigs. Although various RVB strains were identified in humans and various animals globally, little is known about the epidemiology RVB infection in Africa. In this study, we attempted to examine the prevalence of RVB infection in pig populations in Zambia. METHODS: Metagenomic analyses were conducted on pig feces collected in Zambia to detect double stranded RNA viruses, including RVB. To clarify the prevalence of RVB infection in pig populations in Zambia, 147 fecal samples were screened for the RVB detection by RT-qPCR. Full genome sequence of a detected RVB was determined by Sanger sequencing and genetically analyzed. RESULTS: The metagenomic analyses revealed that RVB sequence reads and contigs of RVB were detected from one fecal sample collected from pigs in Zambia. RT-qPCR screening detected RVB genomes in 36.7% (54/147) of fecal samples. Among 54 positive samples, 13 were positive in non-diarrheal samples (n = 48, 27.1%) and 41 in diarrheal samples (n = 99, 41.4%). Genetic analyses demonstrated that all the segments of ZP18-18, except for VP4, had high nucleotide sequence identities (80.6-92.6%) with all other known RVB strains detected in pigs. In contrast, the VP4 sequence of ZP18-18 was highly divergent from other RVB strains (< 64.6% identities) and formed a distinct lineage in the phylogenetic tree. Notably, the VP8 subunit of the VP4 showed remarkably low amino acid identities (33.3%) to those of known RVB strains, indicating that the VP8 subunit of ZP18-18 was unique among RVB strains. According to the whole genome classification for RVB, ZP18-18 was assigned to a genotype constellation, G18-P[9]-I12-R4-C4-M4-A8-N10-T5-E4-H7 with the newly established VP4 genotype P[9]. CONCLUSIONS: This current study updates the geographical distribution and the genetic diversity of RVB. Given the lack of information regarding RVB in Africa, further RVB surveillance is required to assess the potential risk to humans and animals.

Stability of Bas-Congo virus neutralising antibodies in serum samples during long-term storage-Authors' reply.
Yannick Munyeku-Bazitama; Patient Okitale-Talunda; Takanari Hattori; Takeshi Saito; Boniface Pongombo Lombe; Hiroko Miyamoto; Akina Mori-Kajihara; Masahiro Kajihara; Agathe Bikupe Nkoy; Augustin Tshibwabwa Twabela; Justin Masumu; Steve Ahuka-Mundeke; Jean-Jacques Muyembe-Tamfum; Manabu Igarashi; Eun-Sil Park; Shigeru Morikawa; Sheila Makiala-Mandanda; Ayato Takada
The Lancet. Microbe, 5, 10, 100917, 100917, 2024年10月, ［国際誌］
英語

A wide distribution of Beiji nairoviruses and related viruses in Ixodes ticks in Japan.
Mai Kishimoto; Yukari Itakura; Koshiro Tabata; Rika Komagome; Hiroki Yamaguchi; Kohei Ogasawara; Ryo Nakao; Yongjin Qiu; Kozue Sato; Hiroki Kawabata; Masahiro Kajihara; Naota Monma; Junji Seto; Asako Shigeno; Masayuki Horie; Michihito Sasaki; William W Hall; Hirofumi Sawa; Yasuko Orba; Keita Matsuno
Ticks and tick-borne diseases, 15, 6, 102380, 102380, 2024年07月11日, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, Beiji nairovirus (BJNV), in the family Nairoviridae, the order Bunyavirales, was recently reported as a causative agent of an emerging tick-borne zoonotic infection in China. This study investigated the prevalence of BJNV in ticks in Japan. Screening of over 2,000 ticks from multiple regions revealed a widespread distribution of BJNV and BJNV-related viruses in Japan, particularly in the northern island, and in other high altitude areas with exclusive occurrence of Ixodes ticks. Phylogenetic analysis identified three distinct groups of nairoviruses in ticks in Japan: BJNV, Yichun nairovirus (YCNV) and a newly identified Mikuni nairovirus (MKNV). BJNV and YCNV variants identified in ticks in Japan exhibited high nucleotide sequence identities to those in China and Russia with evidence of non-monophyletic evolution among BJNVs, suggesting multiple cross-border transmission events of BJNV between the Eurasian continent and Japan. Whole genome sequencing of BJNV and MKNV revealed a unique GA-rich region in the S segment, the significance of which remains to be determined. In conclusion, the present study has shown a wide distribution and diversity of BJNV-related nairoviruses in Ixodes ticks in Japan and has identified unique genomic structures. The findings demonstrate the significance of BJNV as well as related viruses in Japan and highlight the necessity of monitoring emerging nairovirus infections and their potential risks to public health.

Characterization of human tibrovirus envelope glycoproteins
Yannick Munyeku-Bazitama; Takeshi Saito; Takanari Hattori; Hiroko Miyamoto; Boniface Pongombo Lombe; Akina Mori-Kajihara; Masahiro Kajihara; Jean-Jacques Muyembe-Tamfum; Manabu Igarashi; Eun-sil Park; Shigeru Morikawa; Sheila Makiala-Mandanda; Ayato Takada
Journal of Virology, American Society for Microbiology, 2024年07月02日, ［査読有り］
研究論文（学術雑誌）, ABSTRACT

Tibroviruses are novel rhabdoviruses detected in humans, cattle, and arthropods. Four tibroviruses are known to infect humans: Bas-Congo virus (BASV), Ekpoma virus 1 (EKV-1), Ekpoma virus 2, and Mundri virus. However, since none of them has been isolated, their biological properties are largely unknown. We aimed to characterize the human tibrovirus glycoprotein (G), which likely plays a pivotal role in viral tropism and pathogenicity. Human tibrovirus Gs were found to share some primary structures and display 14 conserved cysteine residues, although their overall amino acid homology was low (29%–48%). Multiple potential glycosylation sites were found on the G molecules, and endoglycosidase H- and peptide-N-glycosidase F-sensitive glycosylation was confirmed. AlphaFold-predicted three-dimensional (3D) structures of human tibrovirus Gs were overall similar. Membrane fusion mediated by these tibrovirus Gs was induced by acidic pH. The low pH-induced conformational change that triggers fusion was reversible. Virus-like particles (VLPs) were produced by transient expression of Gs in cultured cells and used to produce mouse antisera. Using vesicular stomatitis Indiana virus pseudotyped with Gs, we found that the antisera to the respective tibrovirus VLPs showed limited cross-neutralizing activity. It was also found that human C-type lectins and T-cell immunoglobulin mucin 1 acted as attachment factors for G-mediated entry into cells. Interestingly, BASV-G showed the highest ability to utilize these molecules. The viruses infected a wide range of cell lines with preferential tropism for human-derived cells whereas the preference of EKV-1 was unique compared with the other human tibroviruses. These findings provide fundamental information to understand the biological properties of the human tibroviruses.

IMPORTANCE

Human tibroviruses are poorly characterized emerging rhabdoviruses associated with either asymptomatic infection or severe disease with a case fatality rate as high as 60% in humans. However, the extent and burden of human infection as well as factors behind differences in infection outcomes are largely unknown. In this study, we characterized human tibrovirus glycoproteins, which play a key role in virus-host interactions, mainly focusing on their structural and antigenic differences and cellular tropism. Our results provide critical information for understanding the biological properties of these novel viruses and for developing appropriate preparedness interventions such as diagnostic tools, vaccines, and effective therapies.

Detection of Human Adenovirus and Rotavirus in Wastewater in Lusaka, Zambia: Demonstrating the Utility of Environmental Surveillance for the Community
Ngonda Saasa; Ethel M’kandawire; Joseph Ndebe; Mulenga Mwenda; Fred Chimpukutu; Andrew Nalishuwa Mukubesa; Fred Njobvu; Doreen Mainza Shempela; Jay Sikalima; Carol Chiyesu; Bruce Muvwanga; Sarah M. Nampokolwe; Clement Sulwe; Thokozile Khondiwa; Todd Jennings; Ameck Kamanga; Edgar Simulundu; Conceptor Mulube; Wizaso Mwasinga; Jalaimo Mumeka; John Simwanza; Patrick Sakubita; Otridah Kapona; Chilufya Susan Aneta Mulenga; Musole Chipoya; Kunda Musonda; Nathan Kapata; Nyambe Sinyange; Muzala Kapina; Joyce Siwila; Misheck Shawa; Masahiro Kajihara; Ayato Takada; Hirofumi Sawa; Simulyamana A. Choonga; Roma Chilengi; Earnest Muyunda; King S. Nalubamba; Bernard M. Hang’ombe
Pathogens, 13, 6, 486, 486, MDPI AG, 2024年06月07日, ［査読有り］
英語, 研究論文（学術雑誌）, Enteric infections due to viral pathogens are a major public health concern. Detecting the risk areas requires a strong surveillance system for pathogenic viruses in sources such as wastewater. Towards building an environmental surveillance system in Zambia, we aimed to identify group A rotavirus (RVA) and human adenovirus (HAdV) in wastewater. Convenient sampling was conducted at four study sites every Tuesday for five consecutive weeks. The research team focused on three different methods of viral concentration to determine the suitability in terms of cost and applicability for a regular surveillance system: the bag-mediated filtration system (BMFS), polyethylene glycol-based (PEG) precipitation, and skimmed milk (SM) flocculation. We screened 20 wastewater samples for HAdV and RVA using quantitative polymerase chain reaction (qPCR) and conventional polymerase chain reaction (cPCR). Of the 20 samples tested using qPCR, 18/20 (90%) tested positive for HAdV and 14/20 (70%) tested positive for RVA. For the genetic sequencing, qPCR positives were subjected to cPCR, of which 12 positives were successfully amplified. The human adenovirus was identified with a nucleotide identity range of 98.48% to 99.53% compared with the reference genome from GenBank. The BMFS and SM flocculation were the most consistent viral concentration methods for HAdV and RVA, respectively. A statistical analysis of the positives showed that viral positivity differed by site (p < 0.001). SM and PEG may be the most appropriate options in resource-limited settings such as Zambia due to the lower costs associated with these concentration methods. The demonstration of HAdV and RVA detection in wastewater suggests the presence of the pathogens in the communities under study and the need to establish a routine wastewater surveillance system for the identification of pathogens.

Antimicrobial Use Survey and Detection of ESBL-Escherichia coli in Commercial and Medium-/Small-Scale Poultry Farms in Selected Districts of Zambia
Taona Sinyawa; Misheck Shawa; Geoffrey M. Muuka; Fusya Goma; Paul Fandamu; Joseph Yamweka Chizimu; Cynthia Sipho Khumalo; Malala Mulavu; Masuzyo Ngoma; Herman Moses Chambaro; Harvey Kakoma Kamboyi; Masahiro Kajihara; Hirofumi Sawa; Yasuhiko Suzuki; Hideaki Higashi; Geoffrey Mainda; Musso Munyeme; John Bwalya Muma; Christian Owusu Nyantakyi; Beverly Egyir; Bernard Mudenda Hang’ombe
Antibiotics, 13, 5, 467, 467, MDPI AG, 2024年05月20日, ［査読有り］
英語, 研究論文（学術雑誌）, Antimicrobial resistance (AMR) among Escherichia coli from food animals is a rising problem, and heavy antimicrobial use in poultry is a contributing factor. In Zambia, studies linking poultry-associated AMR and antibiotic use (AMU) are rare. This study aimed to investigate commercial and medium-/small-scale poultry farmers’ usage of antimicrobials based on a questionnaire survey in ten districts of Zambia. In addition, the study characterized extended-spectrum β-lactamase (ESBL)-producing E. coli isolates obtained from poultry in the same districts. Data regarding knowledge and usage of antimicrobials were collected from commercial and medium-/small-scale poultry farmers using a pre-tested structured questionnaire. At the same time, cloacal samples were collected and analyzed. One hundred and fifty E. coli isolates were tested for antimicrobial susceptibility using eight antibiotic classes. The isolates were further screened for ESBL production by streaking them on cefotaxime (CTX)-supplemented MacConkey agar, then subjecting them to sequencing on a NextSeq. The questionnaire survey showed that more medium-/small-scale than commercial poultry farmers used antimicrobials (OR = 7.70, 95% CI = 2.88–20.61) but less prescriptions (OR = 0.02, 95% CI = 0.00–0.08). Susceptibility testing revealed that resistance was highest to ampicillin (128/148, 86.5%) and tetracycline (101/136, 74.3%) and that the prevalence of multidrug resistance (MDR) (28/30, 93.3%) was high. Whole-genome sequencing (WGS) of eight (8/30, 26.7%) isolates with CTX Minimum Inhibitory Concentration (MIC) ≥ 4 µg/mL revealed the presence of ESBL-encoding genes blaCTX-M-14, blaCTX-M-55, and blaTEM. WGS also detected other AMR genes for quinolones, aminoglycosides, phenicols, tetracycline, macrolides, and folate-pathway antagonists. Altogether, the questionnaire survey results showed a higher proportion of AMU and lower prescription usage among medium-/small-scale farmers. In addition, our results emphasize the circulation of ESBL-producing E. coli strains with associated MDR. It is critical to educate farmers about AMR risks and to encourage responsible usage of antimicrobials. Furthermore, there is a need to strengthen regulations limiting access to antimicrobials. Finally, there is a need to establish a one health system to guide public health response.

Detection of Extended-Spectrum Beta-Lactamase (ESBL)-Producing Enterobacteriaceae from Diseased Broiler Chickens in Lusaka District, Zambia.
Chikwanda Chileshe; Misheck Shawa; Nelson Phiri; Joseph Ndebe; Cynthia Sipho Khumalo; Chie Nakajima; Masahiro Kajihara; Hideaki Higashi; Hirofumi Sawa; Yasuhiko Suzuki; Walter Muleya; Bernard Mudenda Hang'ombe
Antibiotics (Basel, Switzerland), 13, 3, 2024年03月15日, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, Poultry products in Zambia form an integral part of the human diet in many households, as they are cheap and easy to produce. The burden of poultry diseases has, however, remained a major challenge. Growing consumer demand for poultry products in Zambia has resulted in non-prudent antimicrobial use on farms, intending to prevent and treat poultry diseases for growth optimisation and maximising profits. This cross-sectional study aimed to identify the different types of bacteria causing diseases in chickens in Lusaka and to detect the extended-spectrum lactamase (ESBL)-encoding genes. We collected 215 samples from 91 diseased chickens at three post-mortem facilities and screened them for Gram-negative bacteria. Of these samples, 103 tested positive for various clinically relevant Enterobacteriaceae, including Enterobacter (43/103, 41.7%), Escherichia coli (20/103, 19.4%), Salmonella (10/103, 9.7%), and Shigella (8/103, 7.8%). Other isolated bacteria included Yersinia, Morganella, Proteus, and Klebsiella, which accounted for 21.4%. E. coli, Enterobacter, Salmonella, and Shigella were subjected to antimicrobial susceptibility testing. The results revealed that E. coli, Enterobacter, and Shigella were highly resistant to tetracycline, ampicillin, amoxicillin, and trimethoprim-sulfamethoxazole, while Salmonella showed complete susceptibility to all tested antibiotics. The observed resistance patterns correlated with antimicrobial usage estimated from sales data from a large-scale wholesale and retail company. Six (6/14, 42.9%) E. coli isolates tested positive for blaCTX-M, whilst eight (8/14, 57.1%) Enterobacter samples tested positive for blaTEM. Interestingly, four (4/6, 66.7%) of the E. coli isolates carrying blaCTX-M-positive strains were also positive for blaTEM. Sanger sequencing of the PCR products revealed that five (5/6, 83.3%) of the abovementioned isolates possessed the blaCTX-M-15 allele. The results suggest the presence of potentially pathogenic ESBL-producing Enterobacteriaceae in poultry, threatening public health.

Seroprevalence of Bas-Congo virus in Mangala, Democratic Republic of the Congo: a population-based cross-sectional study
Yannick Munyeku-Bazitama; Patient Okitale-Talunda; Takanari Hattori; Takeshi Saito; Boniface Pongombo Lombe; Hiroko Miyamoto; Akina Mori-Kajihara; Masahiro Kajihara; Agathe Bikupe Nkoy; Augustin Tshibwabwa Twabela; Justin Masumu; Steve Ahuka-Mundeke; Jean-Jacques Muyembe-Tamfum; Manabu Igarashi; Eun-sil Park; Shigeru Morikawa; Sheila Makiala-Mandanda; Ayato Takada
The Lancet Microbe, Elsevier BV, 2024年03月, ［査読有り］
英語, 研究論文（学術雑誌）

Phylogenetic Analysis of Newcastle Disease Virus Isolated from Poultry in Live Bird Markets and Wild Waterfowl in Zambia.
Annie Kalonda; Ngonda Saasa; Masahiro Kajihara; Naganori Nao; Ladslav Moonga; Joseph Ndebe; Akina Mori-Kajihara; Andrew Nalishuwa Mukubesa; Yoshihiro Sakoda; Hirofumi Sawa; Ayato Takada; Edgar Simulundu
Microorganisms, 12, 2, 2024年02月08日, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, Poultry production is essential to the economy and livelihood of many rural Zambian households. However, the industry is threatened by infectious diseases, particularly Newcastle disease virus (NDV) infection. Therefore, this study employed next-generation sequencing to characterise six NDV isolates from poultry in Zambia's live bird markets (LBMs) and wild waterfowl. Four NDV isolates were detected from 410 faecal samples collected from chickens in LBMs in Lusaka and two from 2851 wild birds from Lochinvar National Park. Phylogenetic analysis revealed that the four NDVs from LBM clustered in genotype VII and sub-genotype VII.2 were closely related to viruses previously isolated in Zambia and other Southern African countries, suggesting possible local and regional transboundary circulation of the virus. In contrast, the two isolates from wild birds belonged to class I viruses, genotype 1, and were closely related to isolates from Europe and Asia, suggesting the possible introduction of these viruses from Eurasia, likely through wild bird migration. The fusion gene cleavage site motif for all LBM-associated isolates was 112RRQKR|F117, indicating that the viruses are virulent, while the isolates from wild waterfowl had the typical 112ERQER|L117 avirulent motif. This study demonstrates the circulation of virulent NDV strains in LBMs and has, for the first time, characterised NDV from wild birds in Zambia. The study further provides the first whole genomes of NDV sub-genotype VII.2 and genotype 1 from Zambia and stresses the importance of surveillance and molecular analysis for monitoring the circulation of NDV genotypes and viral evolution.

The First Isolation and Characterization of Bat Jeilongviruses in Japan.
Sho Sata; Isshu Kojima; Mana Esaki; Kimitake Funakoshi; Masahiro Kajihara; Shinji Hirano; Shin Murakami; Kazuo Miyazaki; Makoto Ozawa; Kosuke Okuya
Transboundary and emerging diseases, 2024, 5530007, 5530007, 2024年, ［国際誌］
英語, 研究論文（学術雑誌）, Bats represent natural reservoirs of several paramyxoviruses, raising concerns about the potential for these viruses to cause cross-species infections. In this study, we isolated two jeilongviruses belonging to the family Paramyxoviridae from oral swab samples of the Eastern bent-wing bat (Miniopterus fuliginosus) and Far Eastern myotis bat (Myotis bombinus) in Kagoshima Prefecture, Japan. Notably, this is the first report isolating bat paramyxoviruses in Japan. Genomic analyses revealed a high identity between Kagoshima isolates (PMV/Bat35 and PMV/Bat111) and jeilongvirus B16-40, previously isolated from a Schreiber's bent-wing bat (Miniopterus schreibersii) in South Korea in 2016. PMV/Bat35 infected and replicated in a range of cell lines derived from different animal species, although the level of syncytium formation varied among cell lines. Animal experiments revealed that Syrian hamsters inoculated intranasally with PMV/Bat35 did not exhibit clinical symptoms or significant weight loss. Nevertheless, viral genes were detected in the lungs and tracheas of Syrian hamsters on 2- and 5-day postinfection (dpi). Importantly, neutralizing antibodies against PMV/Bat35 developed in hamsters on 14 dpi. These results suggest that bat jeilongviruses can cross the species barriers. Our findings highlight the critical importance of ongoing monitoring and characterization of viruses circulating in bat populations to assess the risk of zoonotic outbreaks.

Trends, patterns and relationship of antimicrobial use and resistance in bacterial isolates tested between 2015-2020 in a national referral hospital of Zambia.
Misheck Shawa; Atmika Paudel; Herman Chambaro; Harvey Kamboyi; Ruth Nakazwe; Luke Alutuli; Tuvshinzaya Zorigt; Taona Sinyawa; Mulemba Samutela; Joseph Chizimu; Manyando Simbotwe; Kyoko Hayashida; Naganori Nao; Masahiro Kajihara; Yoshikazu Furuta; Yasuhiko Suzuki; Hirofumi Sawa; Bernard Hang'ombe; Hideaki Higashi
PloS one, 19, 4, e0302053, 2024年, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, Increased antimicrobial resistance (AMR) among bacteria underscores the need to strengthen AMR surveillance and promote data-based prescribing. To evaluate trends and associations between antimicrobial usage (AMU) and AMR, we explored a dataset of 34,672 bacterial isolates collected between 2015 and 2020 from clinical samples at the University Teaching Hospital (UTH) in Lusaka, Zambia. The most frequently isolated species were Escherichia coli (4,986/34,672; 14.4%), Staphylococcus aureus (3,941/34,672; 11.4%), and Klebsiella pneumoniae (3,796/34,672; 10.9%). Of the 16 drugs (eight classes) tested, only amikacin and imipenem showed good (> 50%) antimicrobial activity against both E. coli and K. pneumoniae, while nitrofurantoin was effective only in E. coli. Furthermore, 38.8% (1,934/4,980) of E. coli and 52.4% (2,079/3,791) of K. pneumoniae isolates displayed multidrug resistance (MDR) patterns on antimicrobial susceptibility tests. Among S. aureus isolates, 44.6% (973/2,181) were classified as methicillin-resistant (MRSA). Notably, all the MRSA exhibited MDR patterns. The annual hospital AMR rates varied over time, while there was a weak positive relationship (r = 0.38, 95% CI = 0.11-0.60) between the monthly use of third-generation cephalosporins (3GCs) and 3GC resistance among Enterobacterales. Overall, the results revealed high AMR rates that fluctuated over time, with a weak positive relationship between 3GC use and resistance. To our knowledge, this is the first report to evaluate the association between AMU and AMR in Zambia. Our results highlight the need to strengthen antimicrobial stewardship programs and optimize AMU in hospital settings.

Development of an Immunochromatography Assay to Detect Marburg Virus and Ravn Virus
Katendi Changula; Masahiro Kajihara; Shino Muramatsu; Koji Hiraoka; Toru Yamaguchi; Yoko Yago; Daisuke Kato; Hiroko Miyamoto; Akina Mori-Kajihara; Asako Shigeno; Reiko Yoshida; Corey W. Henderson; Andrea Marzi; Ayato Takada
Viruses, 15, 12, 2349, 2349, MDPI AG, 2023年11月29日, ［査読有り］
英語, 研究論文（学術雑誌）, The recent outbreaks of Marburg virus disease (MVD) in Guinea, Ghana, Equatorial Guinea, and Tanzania, none of which had reported previous outbreaks, imply increasing risks of spillover of the causative viruses, Marburg virus (MARV) and Ravn virus (RAVV), from their natural host animals. These outbreaks have emphasized the need for the development of rapid diagnostic tests for this disease. Using monoclonal antibodies specific to the viral nucleoprotein, we developed an immunochromatography (IC) assay for the rapid diagnosis of MVD. The IC assay was found to be capable of detecting approximately 102−4 50% tissue culture infectious dose (TCID50)/test of MARV and RAVV in the infected culture supernatants. We further confirmed that the IC assay could detect the MARV and RAVV antigens in the serum samples from experimentally infected nonhuman primates. These results indicate that the IC assay to detect MARV can be a useful tool for the rapid point-of-care diagnosis of MVD.

Expanding diversity of bunyaviruses identified in mosquitoes.
Yasuko Orba; Yusuf Eshimutu Abu; Herman M Chambaro; Tapiwa Lundu; Walter Muleya; Yuki Eshita; Yongjin Qiu; Hayato Harima; Masahiro Kajihara; Akina Mori-Kajihara; Keita Matsuno; Michihito Sasaki; William W Hall; Bernard M Hang'ombe; Hirofumi Sawa
Scientific reports, 13, 1, 18165, 18165, 2023年10月24日, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, Mosquitoes interact with various organisms in the environment, and female mosquitoes in particular serve as vectors that directly transmit a number of microorganisms to humans and animals by blood-sucking. Comprehensive analysis of mosquito-borne viruses has led to the understanding of the existence of diverse viral species and to the identification of zoonotic arboviruses responsible for significant outbreaks and epidemics. In the present study on mosquito-borne bunyaviruses we employed a broad-spectrum RT-PCR approach and identified eighteen different additional species in the Phenuiviridae family and also a number of related but unclassified bunyaviruses in mosquitoes collected in Zambia. The entire RNA genome segments of the newly identified viruses were further analyzed by RNA sequencing with a ribonuclease R (RNase R) treatment to reduce host-derived RNAs and enrich viral RNAs, taking advantage of the dsRNA panhandle structure of the bunyavirus genome. All three or four genome segments were identified in eight bunyavirus species. Furthermore, L segments of three different novel viruses related to the Leishbunyaviridae were found in mosquitoes together with genes from the suspected host, the Crithidia parasite. In summary, our virus detection approach using a combination of broad-spectrum RT-PCR and RNA sequencing analysis with a simple virus enrichment method allowed the discovery of novel bunyaviruses. The diversity of bunyaviruses is still expanding and studies on this will allow a better understanding of the ecology of hematophagous mosquitoes.

Prevalence and Genomic Characterization of Rotavirus A from Domestic Pigs in Zambia: Evidence for Possible Porcine–Human Interspecies Transmission
Joseph Ndebe; Hayato Harima; Herman Moses Chambaro; Michihito Sasaki; Junya Yamagishi; Annie Kalonda; Misheck Shawa; Yongjin Qiu; Masahiro Kajihara; Ayato Takada; Hirofumi Sawa; Ngonda Saasa; Edgar Simulundu
Pathogens, 12, 10, 1199, 1199, MDPI AG, 2023年09月27日, ［査読有り］
英語, 研究論文（学術雑誌）, Rotavirus is a major cause of diarrhea globally in animals and young children under 5 years old. Here, molecular detection and genetic characterization of porcine rotavirus in smallholder and commercial pig farms in the Lusaka Province of Zambia were conducted. Screening of 148 stool samples by RT-PCR targeting the VP6 gene revealed a prevalence of 22.9% (34/148). Further testing of VP6-positive samples with VP7-specific primers produced 12 positives, which were then Sanger-sequenced. BLASTn of the VP7 positives showed sequence similarity to porcine and human rotavirus strains with identities ranging from 87.5% to 97.1%. By next-generation sequencing, the full-length genetic constellation of the representative strains RVA/pig-wt/ZMB/LSK0137 and RVA/pig-wt/ZMB/LSK0147 were determined. Genotyping of these strains revealed a known Wa-like genetic backbone, and their genetic constellations were G4-P[6]-I5-R1-C1-M1-A8-N1-T1-E1-H1 and G9-P[13]-I5-R1-C1-M1-A8-N1-T1-E1-H1, respectively. Phylogenetic analysis revealed that these two viruses might have their ancestral origin from pigs, though some of their gene segments were related to human strains. The study shows evidence of reassortment and possible interspecies transmission between pigs and humans in Zambia. Therefore, the “One Health” surveillance approach for rotavirus A in animals and humans is recommended to inform the design of effective control measures.

Surveillance, Isolation, and Genetic Characterization of Bat Herpesviruses in Zambia.
Hayato Harima; Yongjin Qiu; Junya Yamagishi; Masahiro Kajihara; Katendi Changula; Kosuke Okuya; Mao Isono; Tomoyuki Yamaguchi; Hirohito Ogawa; Naganori Nao; Michihito Sasaki; Edgar Simulundu; Aaron S Mweene; Hirofumi Sawa; Kanako Ishihara; Bernard M Hang'ombe; Ayato Takada
Viruses, 15, 6, 2023年06月13日, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, Bats are of significant interest as reservoirs for various zoonotic viruses with high diversity. During the past two decades, many herpesviruses have been identified in various bats worldwide by genetic approaches, whereas there have been few reports on the isolation of infectious herpesviruses. Herein, we report the prevalence of herpesvirus infection of bats captured in Zambia and genetic characterization of novel gammaherpesviruses isolated from striped leaf-nosed bats (Macronycteris vittatus). By our PCR screening, herpesvirus DNA polymerase (DPOL) genes were detected in 29.2% (7/24) of Egyptian fruit bats (Rousettus aegyptiacus), 78.1% (82/105) of Macronycteris vittatus, and one Sundevall's roundleaf bat (Hipposideros caffer) in Zambia. Phylogenetic analyses of the detected partial DPOL genes revealed that the Zambian bat herpesviruses were divided into seven betaherpesvirus groups and five gammaherpesvirus groups. Two infectious strains of a novel gammaherpesvirus, tentatively named Macronycteris gammaherpesvirus 1 (MaGHV1), were successfully isolated from Macronycteris vittatus bats, and their complete genomes were sequenced. The genome of MaGHV1 encoded 79 open reading frames, and phylogenic analyses of the DNA polymerase and glycoprotein B demonstrated that MaGHV1 formed an independent lineage sharing a common origin with other bat-derived gammaherpesviruses. Our findings provide new information regarding the genetic diversity of herpesviruses maintained in African bats.

Head-to-head comparison of diagnostic accuracy of four Ebola virus disease rapid diagnostic tests versus GeneXpert® in eastern Democratic Republic of the Congo outbreaks: a prospective observational study
Daniel Mukadi-Bamuleka; Junior Bulabula-Penge; Bart K.M. Jacobs; Anja De Weggheleire; François Edidi-Atani; Fabrice Mambu-Mbika; Anaïs Legand; John D. Klena; Peter N. Fonjungo; Placide Mbala-Kingebeni; Sheila Makiala-Mandanda; Masahiro Kajihara; Ayato Takada; Joel M. Montgomery; Pierre Formenty; Jean-Jacques Muyembe-Tamfum; Kevin K. Ariën; Johan van Griensven; Steve Ahuka-Mundeke; Hgo Kavunga-Membo; Elie Ishara-Nshombo; Stijn Roge; Noella Mulopo-Mukanya; Espérance Tsiwedi-Tsilabia; Emile Muhindo-Milonde; Marie-Anne Kavira-Muhindo; Maria E. Morales-Betoulle; Antoine Nkuba-Ndaye
eBioMedicine, 91, 104568, 104568, Elsevier BV, 2023年05月, ［査読有り］
英語, 研究論文（学術雑誌）

Surveillance and Phylogenetic Characterisation of Avian Influenza Viruses Isolated from Wild Waterfowl in Zambia in 2015, 2020, and 2021
Annie Kalonda; Ngonda Saasa; Masahiro Kajihara; Naganori Nao; Ladislav Moonga; Joseph Ndebe; Akina Mori-Kajihara; Andrew Nalishuwa Mukubesa; Mulemba Samutela; Samuel Munjita; Yoshihiro Sakoda; Hirofumi Sawa; Ayato Takada; Edgar Simulundu
Transboundary and Emerging Diseases, 2023, 1, 16, Hindawi Limited, 2023年03月01日, ［査読有り］
英語, 研究論文（学術雑誌）, In recent years, the southern African region has experienced repeated incursions of highly pathogenic avian influenza viruses (HPAIVs), with wild migratory birds being implicated in the spread. To understand the profile of avian influenza viruses (AIVs) circulating in Zambia, we surveyed wild waterfowl for AIVs and phylogenetically characterised the isolates detected in 2015, 2020, and 2021. A total of 2,851 faecal samples of wild waterfowl were collected from Lochinvar National Park in the Southern Province of Zambia. During the study period, 85 (3.0%) low pathogenicity AIVs belonging to various subtypes were isolated, with H2N9, H8N4, and H10N8 being reported for the first time in avian species in Africa. The majority of the isolates were detected from glossy ibis (order Pelecaniformes) making it the first report of AIV from these birds in Zambia. Phylogenetic analysis of all eight gene segments of the 30 full genomes obtained in this study revealed that all the isolates belonged to the Eurasian lineage with their closest relatives being viruses isolated from wild and/or domestic birds in Bangladesh, Belgium, Egypt, Georgia, Mongolia, the Netherlands, and South Africa. Additionally, the Zambian viruses were grouped into distinct clusters based on the year of isolation. While no notifiable AIVs of the H5 or H7 subtypes were detected in wild birds in Zambia, viral internal protein genes of some viruses were closely related to H7 low pathogenicity AIVs. This study shows that periodically, a considerable diversity of AIV subtypes are introduced into the Zambian ecosystem by wild migratory waterfowl. The findings highlight the importance of continuous surveillance and monitoring of AIVs in wild waterfowl, including birds traditionally not considered to be major AIV reservoirs, for a better understanding of the eco-epidemiology and evolutionary dynamics of AIVs in Africa.

Detection of Old and New World Relapsing Fever Borreliae in Ornithodoros Ticks Collected from Warthog Burrows in Zambia.
Yongjin Qiu; Herman M Chambaro; Kozue Sato; David Squarre; Edgar Simulundu; Masahiro Kajihara; Katendi Changula; Manyando Simbotwe; Hayato Harima; Joseph Ndebe; Ladslav Moonga; Ryo Nakao; Ayato Takada; Bernard Mudenda Hang'ombe; Hirofumi Sawa; Hiroki Kawabata
Microorganisms, 11, 1, 2023年01月12日, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, Relapsing fever (RF) is an arthropod-borne disease caused by Borrelia spirochete, which is one of the major public health concerns in endemic regions including Africa. However, information on Borrelia&nbsp;spirochetes is limited in Zambia. Here, we investigate the Borrelia&nbsp;spirochetes harbored by Ornithodoros ticks in Zambian National Parks. We analyzed 182 DNA samples pooled from 886 Ornithodoros ticks. Of these, 43 tested positive, and their sequence revealed that the ticks harbored both Old and New World RF borreliae. This research presents the first evidence of Old-World RF borreliae in Zambia. The New World RF borreliae detected herein differed from the Candidatus Borrelia fainii previously reported in Zambia and were closely related to the pathogenic Borrelia sp. VS4 identified in Tanzania. Additionally, Borrelia theileri was recently reported in Zambia. Hence, at least four different Borrelia species occur in Zambia, and the organisms causing relapsing fever there might be more complex than previously thought. We empirically confirmed that real-time PCR with TaqMan minor groove binder probes accurately and simultaneously detected both Old and New World RF. In this manner, they could facilitate quantitative analyses of both types of RF borreliae. Subsequent investigations should endeavor to isolate the aforementioned Borrelia spp. and perform serosurveys on patients with RF.

Current status and molecular epidemiology of rabies virus from different hosts and regions in Malawi.
Henson Kainga; Elisha Chatanga; Marvin Collen Phonera; John Pilate Kothowa; Precious Dzimbiri; Gladson Kamwendo; Malala Mulavu; Cynthia Sipho Khumalo; Katendi Changula; Herman Chambaro; Hayato Harima; Masahiro Kajihara; Kholiwe Mkandawire; Patrick Chikungwa; Julius Chulu; Gilson Njunga; Simbarashe Chitanga; Benjamin Mubemba; Michihito Sasaki; Yasuko Orba; Yongjin Qiu; Junya Yamagishi; Edgar Simulundu; Ayato Takada; Boniface Namangala; Hirofumi Sawa; Walter Muleya
Archives of virology, 168, 2, 61, 61, 2023年01月12日, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, Although rabies is endemic in Malawi, there have been no studies in which rabies virus was systematically investigated and characterized in multiple animal hosts in that country. In order to provide molecular epidemiological data on rabies virus in Malawi, 683 suspected rabies case reports from 2008 to 2021 were examined, and 46 (dog = 40, cow = 5, and cat = 1) viable rabies-positive brain samples archived at the Central Veterinary Laboratory (CVL), Lilongwe, Malawi, were analyzed genetically. The results showed an increase in the submission of brain samples from 2008 to 2010, with the highest number of submissions observed in 2020. Of the 683 case reports analyzed for the period under review, 38.1% (260/683) (CI: 34.44 - 41.84) were confirmed by direct fluorescent antibody test. Among the confirmed cases, 65.4% (170/260) (CI: 59.23 - 71.09) were canine rabies. Further, phylogenetic analysis revealed that sequences from different animal hosts clustered together within the Africa 1b lineage, suggesting that the strains circulating in livestock are similar to those in domestic dogs. This finding supports the hypothesis that canine rabies is spilling over to livestock and emphasizes the need for further studies to provide data for effective control of rabies in Malawi.

Isolation and Characterization of Distinct Rotavirus A in Bat and Rodent Hosts.
Mai Kishimoto; Masahiro Kajihara; Koshiro Tabata; Yukari Itakura; Shinsuke Toba; Seiya Ozono; Yuko Sato; Tadaki Suzuki; Naoto Ito; Katendi Changula; Yongjin Qiu; Akina Mori-Kajihara; Yoshiki Eto; Hayato Harima; Daniel Mwizabi; Bernard M Hang'ombe; William W Hall; Ayato Takada; Yasuko Orba; Hirofumi Sawa; Michihito Sasaki
Journal of virology, 97, 1, e0145522, 2023年01月12日, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, Rotavirus A (RVA) causes diarrheal disease in humans and various animals. Recent studies have identified bat and rodent RVAs with evidence of zoonotic transmission and genome reassortment. However, the virological properties of bat and rodent RVAs with currently identified genotypes still need to be better clarified. Here, we performed virus isolation-based screening for RVA in animal specimens and isolated RVAs (representative strains: 16-06 and MpR12) from Egyptian fruit bat and Natal multimammate mouse collected in Zambia. Whole-genome sequencing and phylogenetic analysis revealed that the genotypes of bat RVA 16-06 were identical to that of RVA BATp39 strain from the Kenyan fruit bat, which has not yet been characterized. Moreover, all segments of rodent RVA MpR12 were highly divergent and assigned to novel genotypes, but RVA MpR12 was phylogenetically closer to bat RVAs than to other rodent RVAs, indicating a unique evolutionary history. We further investigated the virological properties of the isolated RVAs. In brief, we found that 16-06 entered cells by binding to sialic acids on the cell surface, while MpR12 entered in a sialic acid-independent manner. Experimental inoculation of suckling mice with 16-06 and MpR12 revealed that these RVAs are causative agents of diarrhea. Moreover, 16-06 and MpR12 demonstrated an ability to infect and replicate in a 3D-reconstructed primary human intestinal epithelium with comparable efficiency to the human RVA. Taken together, our results detail the unique genetic and virological features of bat and rodent RVAs and demonstrate the need for further investigation of their zoonotic potential. IMPORTANCE Recent advances in nucleotide sequence detection methods have enabled the detection of RVA genomes from various animals. These studies have discovered multiple divergent RVAs and have resulted in proposals for the genetic classification of novel genotypes. However, most of these RVAs have been identified via dsRNA viral genomes and not from infectious viruses, and their virological properties, such as cell/host tropisms, transmissibility, and pathogenicity, are unclear and remain to be clarified. Here, we successfully isolated RVAs with novel genome constellations from three bats and one rodent in Zambia. In addition to whole-genome sequencing, the isolated RVAs were characterized by glycan-binding affinity, pathogenicity in mice, and infectivity to the human gut using a 3D culture of primary intestinal epithelium. Our study reveals the first virological properties of bat and rodent RVAs with high genetic diversity and unique evolutional history and provides basic knowledge to begin estimating the potential of zoonotic transmission.

Susceptibility of common dabbling and diving duck species to clade 2.3.2.1 H5N1 high pathogenicity avian influenza virus: an experimental infection study
Kosuke SODA; Yukiko TOMIOKA; Tatsufumi USUI; Hiroichi OZAKI; Hiroshi ITO; Yasuko NAGAI; Naoki YAMAMOTO; Masatoshi OKAMATSU; Norikazu ISODA; Masahiro KAJIHARA; Yoshihiro SAKODA; Ayato TAKADA; Toshihiro ITO
Journal of Veterinary Medical Science, 85, 9, 942, 949, Japanese Society of Veterinary Science, 2023年, ［査読有り］
研究論文（学術雑誌）

Field-deployable multiplex detection method of SARS-CoV-2 and influenza virus using loop-mediated isothermal amplification and DNA chromatography.
Kyoko Hayashida; Alejandro Garcia; Lavel Chinyama Moonga; Tatsuki Sugi; Kodera Takuya; Mitsuo Kawase; Fumihiro Kodama; Atsushi Nagasaka; Nobuhisa Ishiguro; Ayato Takada; Masahiro Kajihara; Naganori Nao; Masashi Shingai; Hiroshi Kida; Yasuhiko Suzuki; William W Hall; Hirofumi Sawa; Junya Yamagishi
PloS one, 18, 5, e0285861, 2023年, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, A novel multiplex loop-mediated isothermal amplification (LAMP) method combined with DNA chromatography was developed for the simultaneous detection of three important respiratory disease-causing viruses: severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), influenza A virus, and influenza B virus. Amplification was performed at a constant temperature, and a positive result was confirmed by a visible colored band. An in-house drying protocol with trehalose was used to prepare the dried format multiplex LAMP test. Using this dried multiplex LAMP test, the analytical sensitivity was determined to be 100 copies for each viral target and 100-1000 copies for the simultaneous detection of mixed targets. The multiplex LAMP system was validated using clinical COVID-19 specimens and compared with the real-time qRT-PCR method as a reference test. The determined sensitivity of the multiplex LAMP system for SARS-CoV-2 was 71% (95% CI: 0.62-0.79) for cycle threshold (Ct) ≤ 35 samples and 61% (95% CI: 0.53-0.69) for Ct ≤40 samples. The specificity was 99% (95%CI: 0.92-1.00) for Ct ≤35 samples and 100% (95%CI: 0.92-1.00) for the Ct ≤40 samples. The developed simple, rapid, low-cost, and laboratory-free multiplex LAMP system for the two major important respiratory viral diseases, COVID-19 and influenza, is a promising field-deployable diagnosis tool for the possible future 'twindemic, ' especially in resource-limited settings.

Seroprevalence and Associated Risk Factors of Rift Valley Fever in Livestock from Three Ecological Zones of Malawi.
Henson Kainga; Marvin Collen Phonera; Elisha Chatanga; Simegnew Adugna Kallu; Prudence Mpundu; Mulemba Samutela; Herman Moses Chambaro; Masahiro Kajihara; Doreen Mainza Shempela; Jay Sikalima; Walter Muleya; Misheck Shawa; Julius Chulu; Gilson Njunga; Martin Simuunza; Ayato Takada; Hirofumi Sawa; Edgar Simulundu; Ngonda Saasa
Pathogens (Basel, Switzerland), 11, 11, 2022年11月14日, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, The epidemiology of Rift Valley fever (RVF) is poorly understood in Malawi. Here, a cross-sectional study was conducted (March-June 2020) to investigate the seroprevalence and potential risk factors of RVF virus (RVFV) in cattle, goats, and sheep in three ecological zones of Malawi. A total of 1523 serum samples were tested for anti-RVFV IgG and IgM antibodies by ELISA. Additionally, a questionnaire survey was used to assess potential RVF risk factors. The overall seroprevalence was 17.14% (261/1523; 95% CI = 15.33-19.11) for individual livestock and 33.24% (120/361; 95% CI = 28.18-38.11) for the livestock herd. Seroprevalence was significantly high in sheep (25.68%, 95% CI = 19.31-33.26) compared with cattle (21.35%, 95% CI = 18.74-24.22) and goats (7.72%, 95% CI = 5.72-10.34), (p = 0.047). At the individual livestock level, the risk was elevated in female livestock (OR: 1.74, 95% CI = 1.08-12.82) (p = 0.016), while at the herd level, areas receiving approximately 1001-1500 mm of rainfall (OR: 2.47, 95% CI = 1.14-5.37) (p = 0.022), areas of rainfall amount greater than approximately 1600 mm (OR: 2.239, 95% CI = 1.07-8.82) (p = 0.023), and mixed species herds (OR: 10.410, 95% CI = 3.04-35.59) (p = 0.001), were significant risk factors. The detection of IgM antibodies confirmed active circulation of RVFV in Malawi. Therefore, monitoring of RVF in animals, humans, and vectors using a "One Health" approach, along with community sensitization among the high-risk populations, could help mitigate the threat posed by this zoonotic disease in Malawi.

Single Nucleotide Variants of the Human TIM-1 IgV Domain with Reduced Ability to Promote Viral Entry into Cells
Takanari Hattori; Takeshi Saito; Hiroko Miyamoto; Masahiro Kajihara; Manabu Igarashi; Ayato Takada
Viruses, 14, 10, 2124, 2124, MDPI AG, 2022年09月26日, ［査読有り］
英語, 研究論文（学術雑誌）, Human T-cell immunoglobulin mucin 1 (hTIM-1) is known to promote cellular entry of enveloped viruses. Previous studies suggested that the polymorphisms of hTIM-1 affected its function. Here, we analyzed single nucleotide variants (SNVs) of hTIM-1 to determine their ability to promote cellular entry of viruses using pseudotyped vesicular stomatitis Indiana virus (VSIV). We obtained hTIM-1 sequences from a public database (Ensembl genome browser) and identified 35 missense SNVs in 3 loops of the hTIM-1 immunoglobulin variable (IgV) domain, which had been reported to interact with the Ebola virus glycoprotein (GP) and phosphatidylserine (PS) in the viral envelope. HEK293T cells transiently expressing wildtype hTIM-1 or its SNV mutants were infected with VSIVs pseudotyped with filovirus or arenavirus GPs, and their infectivities were compared. Eleven of the thirty-five SNV substitutions reduced the efficiency of hTIM-1-mediated entry of pseudotyped VSIVs. These SNV substitutions were found not only around the PS-binding pocket but also in other regions of the molecule. Taken together, our findings suggest that some SNVs of the hTIM-1 IgV domain have impaired ability to interact with PS and/or viral GPs in the viral envelope, which may affect the hTIM-1 function to promote viral entry into cells.

Genomic Surveillance of SARS-CoV-2 in the Southern Province of Zambia: Detection and Characterization of Alpha, Beta, Delta, and Omicron Variants of Concern.
Ben Katowa; Annie Kalonda; Benjamin Mubemba; Japhet Matoba; Doreen Mainza Shempela; Jay Sikalima; Boniface Kabungo; Katendi Changula; Simbarashe Chitanga; Mpanga Kasonde; Otridah Kapona; Nathan Kapata; Kunda Musonda; Mwaka Monze; John Tembo; Matthew Bates; Alimuddin Zumla; Catherine G Sutcliffe; Masahiro Kajihara; Junya Yamagishi; Ayato Takada; Hirofumi Sawa; Roma Chilengi; Victor Mukonka; Walter Muleya; Edgar Simulundu
Viruses, 14, 9, 2022年08月24日, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants of concern (VOCs) have significantly impacted the global epidemiology of the pandemic. From December 2020 to April 2022, we conducted genomic surveillance of SARS-CoV-2 in the Southern Province of Zambia, a region that shares international borders with Botswana, Namibia, and Zimbabwe and is a major tourist destination. Genetic analysis of 40 SARS-CoV-2 whole genomes revealed the circulation of Alpha (B.1.1.7), Beta (B.1.351), Delta (AY.116), and multiple Omicron subvariants with the BA.1 subvariant being predominant. Whereas Beta, Delta, and Omicron variants were associated with the second, third, and fourth pandemic waves, respectively, the Alpha variant was not associated with any wave in the country. Phylogenetic analysis showed evidence of local transmission and possible multiple introductions of SARS-CoV-2 VOCs in Zambia from different European and African countries. Across the 40 genomes analysed, a total of 292 mutations were observed, including 182 missense mutations, 66 synonymous mutations, 23 deletions, 9 insertions, 1 stop codon, and 11 mutations in the non-coding region. This study stresses the need for the continued monitoring of SARS-CoV-2 circulation in Zambia, particularly in strategically positioned regions such as the Southern Province which could be at increased risk of introduction of novel VOCs.

Human ACE2 Genetic Polymorphism Affecting SARS-CoV and SARS-CoV-2 Entry into Cells
Takanari Hattori; Takeshi Saito; Kosuke Okuya; Yuji Takahashi; Hiroko Miyamoto; Masahiro Kajihara; Manabu Igarashi; Ayato Takada
Microbiology Spectrum, American Society for Microbiology, 2022年07月11日, ［査読有り］
英語, 研究論文（学術雑誌）, SARS-CoV and SARS-CoV-2 are known to cause severe pneumonia in humans. The S protein of these CoVs binds to the ACE2 molecule on the plasma membrane and mediates virus entry into cells.

Field performance of three Ebola rapid diagnostic tests used during the 2018–20 outbreak in the eastern Democratic Republic of the Congo: a retrospective, multicentre observational study
Daniel Mukadi-Bamuleka; Junior Bulabula-Penge; Anja De Weggheleire; Bart K M Jacobs; François Edidi-Atani; Fabrice Mambu-Mbika; Placide Mbala-Kingebeni; Sheila Makiala-Mandanda; Martin Faye; Cheick T Diagne; Moussa M Diagne; Oumar Faye; Masahiro Kajihara; Ousmane Faye; Ayato Takada; Amadou A Sall; Jean-Jacques Muyembe-Tamfum; Johan van Griensven; Kevin K Ariën; Steve Ahuka-Mundeke
The Lancet Infectious Diseases, 22, 6, 891, 900, Elsevier BV, 2022年06月, ［査読有り］
英語, 研究論文（学術雑誌）

Detection of Tick-Borne Bacterial and Protozoan Pathogens in Ticks from the Zambia-Angola Border.
Yongjin Qiu; Martin Simuunza; Masahiro Kajihara; Joseph Ndebe; Ngonda Saasa; Penjani Kapila; Hayato Furumoto; Alice C C Lau; Ryo Nakao; Ayato Takada; Hirofumi Sawa
Pathogens (Basel, Switzerland), 11, 5, 2022年05月10日, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, Tick-borne diseases (TBDs), including emerging and re-emerging zoonoses, are of public health importance worldwide; however, TBDs tend to be overlooked, especially in countries with fewer resources, such as Zambia and Angola. Here, we investigated Rickettsia, Anaplasmataceae, and Apicomplexan pathogens in 59 and 96 adult ticks collected from dogs and cattle, respectively, in Shangombo, a town at the Zambia-Angola border. We detected Richkettsia africae and Rickettsia aeschilimannii in 15.6% of Amblyomma variegatum and 41.7% of Hyalomma truncatum ticks, respectively. Ehrlichia minasensis was detected in 18.8% of H. truncatum, and Candidatus Midichloria mitochondrii was determined in Hyalomma marginatum. We also detected Babesia caballi and Theileria velifera in A. variegatum ticks with a 4.4% and 6.7% prevalence, respectively. In addition, Hepatozoon canis was detected in 6.5% of Rhipicephalus lunulatus and 4.3% of Rhipicephalus sanguineus. Coinfection of R. aeshilimannii and E. minasensis were observed in 4.2% of H. truncatum. This is the first report of Ca. M. mitochondrii and E. minasensis, and the second report of B. caballi, in the country. Rickettsia africae and R. aeschlimannii are pathogenic to humans, and E. minasensis, B. caballi, T. velifera, and H. canis are pathogenic to animals. Therefore, individuals, clinicians, veterinarians, and pet owners should be aware of the distribution of these pathogens in the area.

Gargling with povidone iodine has a short-term inhibitory effect on SARS-CoV-2 in patients with COVID-19
T. Seikai; A. Takada; A. Hasebe; M. Kajihara; K. Okuya; T. Sekiguchi (Yamada); W. Kakuguchi; S. Konno; Y. Ohiro
Journal of Hospital Infection, 123, 179, 181, Elsevier BV, 2022年05月, ［査読有り］
英語, 研究論文（学術雑誌）

Susceptibility of herons (family: Ardeidae) to clade 2.3.2.1 H5N1 subtype high pathogenicity avian influenza virus.
Kosuke Soda; Yukiko Tomioka; Tatsufumi Usui; Yukiko Uno; Yasuko Nagai; Hiroshi Ito; Takahiro Hiono; Tomokazu Tamura; Masatoshi Okamatsu; Masahiro Kajihara; Naganori Nao; Yoshihiro Sakoda; Ayato Takada; Toshihiro Ito
Avian pathology : journal of the W.V.P.A, 51, 2, 146, 153, 2022年04月, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, The pathogenicity of the H5 subtype high pathogenicity avian influenza viruses (HPAIVs) in Ardeidae bird species has not been investigated yet, despite the increasing infections reported. Therefore, the present study aimed to examine the susceptibility of the Ardeidae species, which had already been reported to be susceptible to HPAIVs, to a clade 2.3.2.1 H5N1 HPAIV. Juvenile herons (four grey herons, one intermediate egret, two little egrets, and three black-crowned night herons) were intranasally inoculated with 106 50% egg infectious dose of the virus and observed for 10 days. Two of the four grey herons showed lethargy and conjunctivitis; among them, one died at 6 days post-inoculation (dpi). The viruses were transmitted to the other two cohoused naïve grey herons. Some little egrets and black-crowned night herons showing neurological disorders died at 4-5 dpi; these birds mainly shed the virus via the oral route. The viruses predominantly replicated in the brains of birds that died of infection. Seroconversion was observed in most surviving birds, except some black-crowned night herons. These results demonstrate that most Ardeidae species are susceptible to H5 HPAIVs, sometimes with lethal effects. Herons are mostly colonial and often share habitats with Anseriformes, natural hosts of influenza A viruses; therefore, the risks of cluster infection and contribution to viral dissemination should be continuously evaluated. RESEARCH HIGHLIGHTSClade 2.3.2.1 H5N1 HPAIV causes lethal infections in Ardeidae sp.Viruses are transmitted among grey herons.Some herons with HPAIV showed conjunctivitis or neurological symptoms.HPAIV systemically replicated in herons tissues.

Mapping of Antibody Epitopes on the Crimean-Congo Hemorrhagic Fever Virus Nucleoprotein.
Boniface Pongombo Lombe; Takeshi Saito; Hiroko Miyamoto; Akina Mori-Kajihara; Masahiro Kajihara; Masayuki Saijo; Justin Masumu; Takanari Hattori; Manabu Igarashi; Ayato Takada
Viruses, 14, 3, 2022年03月06日, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, Crimean-Congo hemorrhagic fever virus (CCHFV), a nairovirus, is a tick-borne zoonotic virus that causes hemorrhagic fever in humans. The CCHFV nucleoprotein (NP) is the antigen most used for serological screening of CCHFV infection in animals and humans. To gain insights into antibody epitopes on the NP molecule, we produced recombinant chimeric NPs between CCHFV and Nairobi sheep disease virus (NSDV), which is another nairovirus, and tested rabbit and mouse antisera/immune ascites, anti-NP monoclonal antibodies, and CCHFV-infected animal/human sera for their reactivities to the NP antigens. We found that the amino acids at positions 161-320 might include dominant epitopes recognized by anti-CCHFV IgG antibodies, whereas cross-reactivity between anti-CCHFV and anti-NSDV antibodies was limited. Their binding capacities were further tested using a series of synthetic peptides whose sequences were derived from CCHFV NP. IgG antibodies in CCHFV-infected monkeys and patients were reactive to some of the synthetic peptide antigens (e.g., amino acid residues at positions 131-150 and 211-230). Only a few peptides were recognized by IgG antibodies in the anti-NSDV serum. These results provide useful information to improve NP-based antibody detection assays as well as antigen detection tests relying on anti-NP monoclonal antibodies.

Molecular Mechanisms Underlying the Cellular Entry and Host Range Restriction of Lujo Virus.
Takeshi Saito; Takanari Hattori; Kosuke Okuya; Rashid Manzoor; Hiroko Miyamoto; Masahiro Kajihara; Ayato Takada
mBio, e0306021, 2022年02月15日, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, Like other human-pathogenic arenaviruses, Lujo virus (LUJV) is a causative agent of viral hemorrhagic fever in humans. LUJV infects humans with high mortality rates, but the susceptibilities of other animal species and the molecular determinants of its host specificity remain unknown. We found that mouse- and hamster-derived cell lines (NIH 3T3 and BHK, respectively) were less susceptible to a replication-incompetent recombinant vesicular stomatitis virus (Indiana) pseudotyped with the LUJV glycoprotein (GP) (VSVΔG*-LUJV/GP) than were human-derived cell lines (HEK293T and Huh7). To determine the cellular factors involved in the differential susceptibilities between the human and mouse cell lines, we focused on the CD63 molecule, which is required for pH-activated GP-mediated membrane fusion during LUJV entry into host cells. The exogenous introduction of human CD63, but not mouse or hamster CD63, into BHK cells significantly increased susceptibility to VSVΔG*-LUJV/GP. Using chimeric human-mouse CD63 proteins, we found that the amino acid residues at positions 141 to 150 in the large extracellular loop (LEL) region of CD63 were important for the cellular entry of VSVΔG*-LUJV/GP. By site-directed mutagenesis, we further determined that a phenylalanine at position 143 in human CD63 was the key residue for efficient membrane fusion and VSVΔG*-LUJV/GP infection. Our data suggest that the interaction of LUJV GP with the LEL region of CD63 is essential for cell susceptibility to LUJV, thus providing new insights into the molecular mechanisms underlying the cellular entry of LUJV and the host range restriction of this virus. IMPORTANCE Lujo virus (LUJV) infects humans with high mortality rates, but the host range of LUJV remains unknown. We found that rodent-derived cell lines were less susceptible to LUJV infection than were human-derived cell lines, and the differential susceptibilities were determined by the difference of CD63, the intercellular receptor of LUJV. We further identified an amino acid residue on human CD63 important for efficient LUJV infection. These results suggest that the interaction between LUJV glycoprotein and CD63 is one of the important factors determining the host range of LUJV. Our findings on the CD63-regulated susceptibilities of the cell lines to LUJV infection provide important information for the development of anti-LUJV drugs as well as the identification of natural hosts of LUJV. Importantly, our data support a concept explaining the molecular mechanism underlying viral tropisms controlled by endosomal receptors.

Current knowledge of vector-borne zoonotic pathogens in Zambia: A clarion call to scaling-up "One Health" research in the wake of emerging and re-emerging infectious diseases.
Benjamin Mubemba; Monicah M Mburu; Katendi Changula; Walter Muleya; Lavel C Moonga; Herman M Chambaro; Masahiro Kajihara; Yongjin Qiu; Yasuko Orba; Kyoko Hayashida; Catherine G Sutcliffe; Douglas E Norris; Philip E Thuma; Phillimon Ndubani; Simbarashe Chitanga; Hirofumi Sawa; Ayato Takada; Edgar Simulundu
PLoS neglected tropical diseases, 16, 2, e0010193, 2022年02月, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, BACKGROUND: Although vector-borne zoonotic diseases are a major public health threat globally, they are usually neglected, especially among resource-constrained countries, including those in sub-Saharan Africa. This scoping review examined the current knowledge and identified research gaps of vector-borne zoonotic pathogens in Zambia. METHODS AND FINDINGS: Major scientific databases (Web of Science, PubMed, Scopus, Google Scholar, CABI, Scientific Information Database (SID)) were searched for articles describing vector-borne (mosquitoes, ticks, fleas and tsetse flies) zoonotic pathogens in Zambia. Several mosquito-borne arboviruses have been reported including Yellow fever, Ntaya, Mayaro, Dengue, Zika, West Nile, Chikungunya, Sindbis, and Rift Valley fever viruses. Flea-borne zoonotic pathogens reported include Yersinia pestis and Rickettsia felis. Trypanosoma sp. was the only tsetse fly-borne pathogen identified. Further, tick-borne zoonotic pathogens reported included Crimean-Congo Haemorrhagic fever virus, Rickettsia sp., Anaplasma sp., Ehrlichia sp., Borrelia sp., and Coxiella burnetii. CONCLUSIONS: This study revealed the presence of many vector-borne zoonotic pathogens circulating in vectors and animals in Zambia. Though reports of human clinical cases were limited, several serological studies provided considerable evidence of zoonotic transmission of vector-borne pathogens in humans. However, the disease burden in humans attributable to vector-borne zoonotic infections could not be ascertained from the available reports and this precludes the formulation of national policies that could help in the control and mitigation of the impact of these diseases in Zambia. Therefore, there is an urgent need to scale-up "One Health" research in emerging and re-emerging infectious diseases to enable the country to prepare for future epidemics, including pandemics.

Seroprevalence and Risk Factors of Crimean-Congo Hemorrhagic Fever in Cattle of Smallholder Farmers in Central Malawi.
Marvin Collen Phonera; Martin Chitolongo Simuunza; Henson Kainga; Joseph Ndebe; Mwelwa Chembensofu; Elisha Chatanga; Setiala Kanyanda; Katendi Changula; Walter Muleya; Benjamin Mubemba; Simbarashe Chitanga; Masahiro Kajihara; Hirofumi Sawa; Gilson Njunga; Ayato Takada; Edgar Simulundu
Pathogens (Basel, Switzerland), 10, 12, 2021年12月10日, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, Crimean-Congo hemorrhagic fever virus (CCHFV) is endemic in Africa, Asia, and Eastern Europe where it circulates among animals and ticks causing sporadic outbreaks in humans. Although CCHF is endemic in sub-Saharan Africa, epidemiological information is lacking in many countries, including Malawi. To assess the risk of CCHF in Malawi, we conducted an epidemiological study in cattle reared by smallholder livestock farmers in central Malawi. A cross-sectional study was conducted in April 2020 involving seven districts, four from Kasungu and three from Lilongwe Agriculture Development Divisions. A structured questionnaire was administered to farmers to obtain demographic, animal management, and ecological risk factors data. Sera were collected from randomly selected cattle and screened for CCHF virus (CCHFV) specific antibodies using a commercial ELISA kit. Ticks were collected from cattle and classified morphologically to species level. An overall CCHFV seropositivity rate of 46.9% (n = 416; 95% CI: 42.0-51.8%) was observed. The seropositivity was significantly associated with the age of cattle (p < 0.001), sex (p < 0.001), presence of ticks in herds (p = 0.01), district (p = 0.025), and type of grazing lands (p = 0.013). Five species of ticks were identified, including Hyalomma truncatum, a known vector of CCHFV. Ticks of the species Hyalomma truncatum were not detected in two districts with the highest seroprevalence for CCHF and vector competency must be further explored in the study area. To our knowledge, this is the first report of serologic evidence of the presence of CCHV among smallholder cattle in central Malawi. This study emphasizes the need for continued monitoring of CCHFV infection among livestock, ticks, and humans for the development of data-based risk mitigation strategies.

Structural Requirements in the Hemagglutinin Cleavage Site-Coding RNA Region for the Generation of Highly Pathogenic Avian Influenza Virus.
Yurie Kida; Kosuke Okuya; Takeshi Saito; Junya Yamagishi; Aiko Ohnuma; Takanari Hattori; Hiroko Miyamoto; Rashid Manzoor; Reiko Yoshida; Naganori Nao; Masahiro Kajihara; Tokiko Watanabe; Ayato Takada
Pathogens (Basel, Switzerland), 10, 12, 2021年12月09日, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, Highly pathogenic avian influenza viruses (HPAIVs) with H5 and H7 hemagglutinin (HA) subtypes are derived from their low pathogenic counterparts following the acquisition of multiple basic amino acids in their HA cleavage site. It has been suggested that consecutive adenine residues and a stem-loop structure in the viral RNA region that encodes the cleavage site are essential for the acquisition of the polybasic cleavage site. By using a reporter assay to detect non-templated nucleotide insertions, we found that insertions more frequently occurred in the RNA region (29 nucleotide-length) encoding the cleavage site of an H5 HA gene that was predicted to have a stem-loop structure containing consecutive adenines than in a mutated corresponding RNA region that had a disrupted loop structure with fewer adenines. In virus particles generated by using reverse genetics, nucleotide insertions that created additional codons for basic amino acids were found in the RNA region encoding the cleavage site of an H5 HA gene but not in the mutated RNA region. We confirmed the presence of virus clones with the ability to replicate without trypsin in a plaque assay and to cause lethal infection in chicks. These results demonstrate that the stem-loop structure containing consecutive adenines in HA genes is a key molecular determinant for the emergence of H5 HPAIVs.

Influenza A and D Viruses in Non-Human Mammalian Hosts in Africa: A Systematic Review and Meta-Analysis.
Annie Kalonda; Marvin Phonera; Ngonda Saasa; Masahiro Kajihara; Catherine G Sutcliffe; Hirofumi Sawa; Ayato Takada; Edgar Simulundu
Viruses, 13, 12, 2021年12月02日, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, We conducted a systematic review and meta-analysis to investigate the prevalence and current knowledge of influenza A virus (IAV) and influenza D virus (IDV) in non-human mammalian hosts in Africa. PubMed, Google Scholar, Wiley Online Library and World Organisation for Animal Health (OIE-WAHIS) were searched for studies on IAV and IDV from 2000 to 2020. Pooled prevalence and seroprevalences were estimated using the quality effects meta-analysis model. The estimated pooled prevalence and seroprevalence of IAV in pigs in Africa was 1.6% (95% CI: 0-5%) and 14.9% (95% CI: 5-28%), respectively. The seroprevalence of IDV was 87.2% (95% CI: 24-100%) in camels, 9.3% (95% CI: 0-24%) in cattle, 2.2% (95% CI: 0-4%) in small ruminants and 0.0% (95% CI: 0-2%) in pigs. In pigs, H1N1 and H1N1pdm09 IAVs were commonly detected. Notably, the highly pathogenic H5N1 virus was also detected in pigs. Other subtypes detected serologically and/or virologically included H3N8 and H7N7 in equids, H1N1, and H3N8 and H5N1 in dogs and cats. Furthermore, various wildlife animals were exposed to different IAV subtypes. For prudent mitigation of influenza epizootics and possible human infections, influenza surveillance efforts in Africa should not neglect non-human mammalian hosts. The impact of IAV and IDV in non-human mammalian hosts in Africa deserves further investigation.

Hepatitis E virus infection in pigs: a first report from Zambia.
Herman M Chambaro; Michihito Sasaki; Walter Muleya; Masahiro Kajihara; Misheck Shawa; Kabemba E Mwape; Hayato Harima; Yongjin Qiu; William W Hall; Paul Fandamu; David Squarre; Edgar Simulundu; Hirofumi Sawa; Yasuko Orba
Emerging microbes & infections, 10, 1, 2169, 2172, 2021年12月, ［査読有り］, ［国際誌］
英語, While evidence suggests presence of HEV infection in humans in Zambia, currently, there is no information on its occurrence in domestic pigs. Here, we investigated the presence of HEV antibodies and genome in domestic pigs in Zambia. Sera (n = 484) from domestic pigs were screened for antibodies against HEV by ELISA while genome detection in fecal (n = 25) and liver (n = 100) samples from slaughter pigs was conducted using nested RT-PCR assay. Overall, seroprevalence was 47.7% (231/484) while zoonotic genotype 3 HEV RNA was detected in 16.0% (20/125) of slaughtered pigs. This is the first report to highlight occurrence of HEV infection in domestic pigs in Zambia. This finding suggests possible contamination of the pork supply chain. Moreover, there is a potential risk of zoonotic transmission of HEV to abattoir workers, pig farmers and handlers.

Functional Importance of Hydrophobic Patches on the Ebola Virus VP35 IFN-Inhibitory Domain.
Nodoka Kasajima; Keita Matsuno; Hiroko Miyamoto; Masahiro Kajihara; Manabu Igarashi; Ayato Takada
Viruses, 13, 11, 2021年11月20日, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, Viral protein 35 (VP35) of Ebola virus (EBOV) is a multifunctional protein that mainly acts as a viral polymerase cofactor and an interferon antagonist. VP35 interacts with the viral nucleoprotein (NP) and double-stranded RNA for viral RNA transcription/replication and inhibition of type I interferon (IFN) production, respectively. The C-terminal portion of VP35, which is termed the IFN-inhibitory domain (IID), is important for both functions. To further identify critical regions in this domain, we analyzed the physical properties of the surface of VP35 IID, focusing on hydrophobic patches, which are expected to be functional sites that are involved in interactions with other molecules. Based on the known structural information of VP35 IID, three hydrophobic patches were identified on its surface and their biological importance was investigated using minigenome and IFN-β promoter-reporter assays. Site-directed mutagenesis revealed that some of the amino acid substitutions that were predicted to disrupt the hydrophobicity of the patches significantly decreased the efficiency of viral genome replication/transcription due to reduced interaction with NP, suggesting that the hydrophobic patches might be critical for the formation of a replication complex through the interaction with NP. It was also found that the hydrophobic patches were involved in the IFN-inhibitory function of VP35. These results highlight the importance of hydrophobic patches on the surface of EBOV VP35 IID and also indicate that patch analysis is useful for the identification of amino acid residues that directly contribute to protein functions.

Serological and molecular epidemiological study on swine influenza in Zambia.
Hayato Harima; Kosuke Okuya; Masahiro Kajihara; Hirohito Ogawa; Edgar Simulundu; Eugene Bwalya; Yongjin Qiu; Akina Mori-Kajihara; Musso Munyeme; Yoshihiro Sakoda; Takehiko Saito; Bernard M Hang'ombe; Hirofumi Sawa; Aaron S Mweene; Ayato Takada
Transboundary and emerging diseases, 69, 4, e931-e943, 2021年11月01日, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, Influenza A viruses (IAVs) cause highly contagious respiratory diseases in humans and animals. In 2009, a swine-origin pandemic H1N1 IAV, designated A(H1N1)pdm09 virus, spread worldwide, and has since frequently been introduced into pig populations. Since novel reassortant IAVs with pandemic potential may emerge in pigs, surveillance for IAV in pigs is therefore necessary not only for the pig industry but also for public health. However, epidemiological information on IAV infection of pigs in Africa remains sparse. In this study, we collected 246 serum and 605 nasal swab samples from pigs in Zambia during the years 2011-2018. Serological analyses revealed that 49% and 32% of the sera collected in 2011 were positive for hemagglutination-inhibition (HI) and neutralizing antibodies against A(H1N1)pdm09 virus, respectively, whereas less than 5.3% of sera collected during the following period (2012-2018) were positive in both serological tests. The positive rate and the neutralization titres to A(H1N1)pdm09 virus were higher than those to classical swine H1N1 and H1N2 IAVs. On the other hand, the positive rate for swine H3N2 IAV was very low in the pig population in Zambia in 2011-2018 (5.3% and 0% in HI and neutralization tests, respectively). From nasal swab samples, we isolated one H3N2 and eight H1N1 IAV strains with an isolation rate of 1.5%. Phylogenetic analyses of all eight gene segments revealed that the isolated IAVs were closely related to human IAV strains belonging to A(H1N1)pdm09 and seasonal H3N2 lineages. Our findings indicate that reverse zoonotic transmission from humans to pigs occurred during the study period in Zambia and highlight the need for continued surveillance to monitor the status of IAVs circulating in swine populations in Africa.

Attenuated infection by a Pteropine orthoreovirus isolated from an Egyptian fruit bat in Zambia.
Hayato Harima; Michihito Sasaki; Yasuko Orba; Kosuke Okuya; Yongjin Qiu; Christida E Wastika; Katendi Changula; Masahiro Kajihara; Edgar Simulundu; Tomoyuki Yamaguchi; Yoshiki Eto; Akina Mori-Kajihara; Akihiko Sato; Satoshi Taniguchi; Ayato Takada; Masayuki Saijo; Bernard M Hang'ombe; Hirofumi Sawa
PLoS neglected tropical diseases, 15, 9, e0009768, 2021年09月, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）, BACKGROUND: Pteropine orthoreovirus (PRV) is an emerging bat-borne zoonotic virus that causes severe respiratory illness in humans. Although PRVs have been identified in fruit bats and humans in Australia and Asia, little is known about the prevalence of PRV infection in Africa. Therefore, this study performed an PRV surveillance in fruit bats in Zambia. METHODS: Egyptian fruit bats (Rousettus aegyptiacus, n = 47) and straw-colored fruit bats (Eidolon helvum, n = 33) captured in Zambia in 2017-2018 were screened for PRV infection using RT-PCR and serum neutralization tests. The complete genome sequence of an isolated PRV strain was determined by next generation sequencing and subjected to BLAST and phylogenetic analyses. Replication capacity and pathogenicity of the strain were investigated using Vero E6 cell cultures and BALB/c mice, respectively. RESULTS: An PRV strain, tentatively named Nachunsulwe-57, was isolated from one Egyptian fruit bat. Serological assays demonstrated that 98% of sera (69/70) collected from Egyptian fruit bats (n = 37) and straw-colored fruit bats (n = 33) had neutralizing antibodies against PRV. Genetic analyses revealed that all 10 genome segments of Nachunsulwe-57 were closely related to a bat-derived Kasama strain found in Uganda. Nachunsulwe-57 showed less efficiency in viral growth and lower pathogenicity in mice than another PRV strain, Miyazaki-Bali/2007, isolated from a patient. CONCLUSIONS: A high proportion of Egyptian fruit bats and straw-colored fruit bats were found to be seropositive to PRV in Zambia. Importantly, a new PRV strain (Nachunsulwe-57) was isolated from an Egyptian fruit bat in Zambia, which had relatively weak pathogenicity in mice. Taken together, our findings provide new epidemiological insights about PRV infection in bats and indicate the first isolation of an PRV strain that may have low pathogenicity to humans.

Mosquito-Borne Viral Pathogens Detected in Zambia: A Systematic Review.
Rachel Milomba Velu; Geoffrey Kwenda; Liyali Libonda; Caroline Cleopatra Chisenga; Bumbangi Nsoni Flavien; Obvious Nchimunya Chilyabanyama; Michelo Simunyandi; Samuel Bosomprah; Nicholus Chintu Sande; Katendi Changula; Walter Muleya; Monicah Mirai Mburu; Benjamin Mubemba; Simbarashe Chitanga; John Tembo; Matthew Bates; Nathan Kapata; Yasuko Orba; Masahiro Kajihara; Ayato Takada; Hirofumi Sawa; Roma Chilengi; Edgar Simulundu
Pathogens (Basel, Switzerland), 10, 8, 2021年08月10日, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）, Emerging and re-emerging mosquito-borne viral diseases are a threat to global health. This systematic review aimed to investigate the available evidence of mosquito-borne viral pathogens reported in Zambia. A search of literature was conducted in PubMed and Google Scholar for articles published from 1 January 1930 to 30 June 2020 using a combination of keywords. Eight mosquito-borne viruses belonging to three families, Togaviridae, Flaviviridae and Phenuiviridae were reported. Three viruses (Chikungunya virus, Mayaro virus, Mwinilunga virus) were reported among the togaviruses whilst four (dengue virus, West Nile virus, yellow fever virus, Zika virus) were among the flavivirus and only one virus, Rift Valley fever virus, was reported in the Phenuiviridae family. The majority of these mosquito-borne viruses were reported in Western and North-Western provinces. Aedes and Culex species were the main mosquito-borne viral vectors reported. Farming, fishing, movement of people and rain patterns were among factors associated with mosquito-borne viral infection in Zambia. Better diagnostic methods, such as the use of molecular tools, to detect the viruses in potential vectors, humans, and animals, including the recognition of arboviral risk zones and how the viruses circulate, are important for improved surveillance and design of effective prevention and control measures.

Screening of tick-borne pathogens in argasid ticks in Zambia: Expansion of the geographic distribution of Rickettsia lusitaniae and Rickettsia hoogstraalii and detection of putative novel Anaplasma species.
Yongjin Qiu; Martin Simuunza; Masahiro Kajihara; Herman Chambaro; Hayato Harima; Yoshiki Eto; Edgar Simulundu; David Squarre; Shiho Torii; Ayato Takada; Bernard Mudenda Hang'ombe; Hirofumi Sawa; Chihiro Sugimoto; Ryo Nakao
Ticks and tick-borne diseases, 12, 4, 101720, 101720, 2021年07月, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）, Ticks (Ixodidae and Argasidae) are important arthropod vectors of various pathogens that cause human and animal infectious diseases. Many previously published studies on tick-borne pathogens focused on those transmitted by ixodid ticks. Although there are increasing reports of viral pathogens associated with argasid ticks, information on bacterial pathogens they transmit is scarce. The aim of this molecular study was to detect and characterize Rickettsia and Anaplasmataceae in three different argasid tick species, Ornithodoros faini, Ornithodoros moubata, and Argas walkerae collected in Zambia. Rickettsia hoogstraalii and Rickettsia lusitaniae were detected in 77 % (77/100) of Ar. walkerae and 10 % (5/50) of O. faini, respectively. All O. moubata pool samples (n = 124) were negative for rickettsial infections. Anaplasmataceae were detected in 63 % (63/100) of Ar. walkerae and in 82.2 % (102/124) of O. moubata pools, but not in O. faini. Phylogenetic analysis based on the concatenated sequences of 16S rRNA and groEL genes revealed that Anaplasma spp. detected in the present study were distinct from previously validated Anaplasma species, indicating that the current knowledge on the diversity and vector range of Anaplasma spp. is incomplete. Our findings highlight new geographical records of R. lusitaniae and R. hoogstraalii and confirm that the wide geographic distribution of these species includes the African continent. The data presented here increase our knowledge on argasid tick-borne bacteria and contribute toward understanding their epidemiology.

Molecular Detection and Genotyping of Coxiella-Like Endosymbionts in Ticks Collected from Animals and Vegetation in Zambia.
Toshiya Kobayashi; Elisha Chatanga; Yongjin Qiu; Martin Simuunza; Masahiro Kajihara; Bernard Mudenda Hang'ombe; Yoshiki Eto; Ngonda Saasa; Akina Mori-Kajihara; Edgar Simulundu; Ayato Takada; Hirofumi Sawa; Ken Katakura; Nariaki Nonaka; Ryo Nakao
Pathogens (Basel, Switzerland), 10, 6, 2021年06月21日, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）, Ticks are obligate ectoparasites as they require to feed on their host blood during some or all stages of their life cycle. In addition to the pathogens that ticks harbor and transmit to vertebrate hosts, they also harbor other seemingly nonpathogenic microorganisms including nutritional mutualistic symbionts. Tick nutritional mutualistic symbionts play important roles in the physiology of the host ticks as they are involved in tick reproduction and growth through the supply of B vitamins as well as in pathogen maintenance and propagation. Coxiella-like endosymbionts (CLEs) are the most widespread endosymbionts exclusively reported in ticks. Although CLEs have been investigated in ticks in other parts of the world, there is no report of their investigation in ticks in Zambia. To investigate the occurrence of CLEs, their maintenance, and association with host ticks in Zambia, 175 ticks belonging to six genera, namely Amblyomma, Argas, Haemaphysalis, Hyalomma, Ornithodoros, and Rhipicephalus, were screened for CLEs, followed by characterization of CLEs by multi-locus sequence typing of the five Coxiella housekeeping genes (dnaK, groEL, rpoB, 16S rRNA, and 23S rRNA). The results showed that 45.7% (n = 80) were positive for CLEs. The comparison of the tick 16S rDNA phylogenetic tree with that of the CLEs concatenated sequences showed that there was a strong correlation between the topology of the trees. The results suggest that most of the CLEs have evolved within tick species, supporting the vertical transmission phenomenon. However, the negative results for CLE in some ticks warrants further investigations of other endosymbionts that the ticks in Zambia may also harbor.

Immunization Coverage and Antibody Retention against Rabies in Domestic Dogs in Lusaka District, Zambia.
Chiho Kaneko; Michihito Sasaki; Ryosuke Omori; Ryo Nakao; Chikako Kataoka-Nakamura; Ladslav Moonga; Joseph Ndebe; Walter Muleya; Edgar Simulundu; Bernard M Hang'ombe; George Dautu; Masahiro Kajihara; Akina Mori-Kajihara; Yongjin Qiu; Naoto Ito; Herman M Chambaro; Chihiro Sugimoto; Hideaki Higashi; Ayato Takada; Hirofumi Sawa; Aaron S Mweene; Norikazu Isoda
Pathogens (Basel, Switzerland), 10, 6, 2021年06月11日, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）, Rabies remains endemic in Zambia. Despite conducting canine vaccinations in Lusaka district, the vaccination coverage and actual seropositivity in the dog population in Lusaka district are rarely evaluated. This study estimated the seropositivity-based immunization coverage in the owned dog population in Lusaka district using the expanded program on immunization cluster survey method. The time-series trend of neutralizing antibodies against rabies in vaccinated dogs was also evaluated. Of 366 dogs in 200 dog-owning households in Lusaka district, blood samples were collected successfully from 251 dogs. In the sampled dogs, 42.2% (106/251) had an antibody titer ≥0.5 IU/mL. When the 115 dogs whose blood was not collected were assumed to be seronegative, the minimum immunization coverage in Lusaka district's owned dog population was estimated at 29.0% (95% confidence interval: 22.4-35.5). It was also found that a single vaccination with certified vaccines is capable of inducing protective levels of antibodies. In contrast, higher antibody titers were observed in multiple-vaccinated dogs than in single-vaccinated dogs, coupled with the observation of a decline in antibody titer over time. These results suggest the importance of continuous booster immunization to maintain herd immunity and provide useful information to plan mass vaccination against rabies in Zambia.

Serologic and molecular evidence for circulation of Crimean-Congo hemorrhagic fever virus in ticks and cattle in Zambia.
Masahiro Kajihara; Martin Simuunza; Ngonda Saasa; George Dautu; Akina Mori-Kajihara; Yongjin Qiu; Ryo Nakao; Yoshiki Eto; Hayato Furumoto; Bernard M Hang'ombe; Yasuko Orba; Hirofumi Sawa; Edgar Simulundu; Shuetsu Fukushi; Shigeru Morikawa; Masayuki Saijo; Jiro Arikawa; Swithine Kabilika; Mwaka Monze; Victor Mukonka; Aaron Mweene; Ayato Takada; Kumiko Yoshimatsu
PLoS neglected tropical diseases, 15, 6, e0009452, 2021年06月, ［査読有り］, ［筆頭著者］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）, Crimean-Congo hemorrhagic fever (CCHF) is a tick-borne zoonosis with a high case fatality rate in humans. Although the disease is widely found in Africa, Europe, and Asia, the distribution and genetic diversity of CCHF virus (CCHFV) are poorly understood in African countries. To assess the risks of CCHF in Zambia, where CCHF has never been reported, epidemiologic studies in cattle and ticks were conducted. Through an indirect immunofluorescence assay, CCHFV nucleoprotein-specific serum IgG was detected in 8.4% (88/1,047) of cattle. Among 290 Hyalomma ticks, the principal vector of CCHFV, the viral genome was detected in 11 ticks. Phylogenetic analyses of the CCHFV S and M genome segments revealed that one of the detected viruses was a genetic reassortant between African and Asian strains. This study provides compelling evidence for the presence of CCHFV in Zambia and its transmission to vertebrate hosts.

Domestic dog demographics and estimates of canine vaccination coverage in a rural area of Zambia for the elimination of rabies.
Chiho Kaneko; Ryosuke Omori; Michihito Sasaki; Chikako Kataoka-Nakamura; Edgar Simulundu; Walter Muleya; Ladslav Moonga; Joseph Ndebe; Bernard M Hang'ombe; George Dautu; Yongjin Qiu; Ryo Nakao; Masahiro Kajihara; Akina Mori-Kajihara; Herman M Chambaro; Hideaki Higashi; Chihiro Sugimoto; Hirofumi Sawa; Aaron S Mweene; Ayato Takada; Norikazu Isoda
PLoS neglected tropical diseases, 15, 4, e0009222, 2021年04月, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）, BACKGROUND: An estimated 75% or more of the human rabies cases in Africa occur in rural settings, which underscores the importance of rabies control in these areas. Understanding dog demographics can help design strategies for rabies control and plan and conduct canine mass vaccination campaigns effectively in African countries. METHODOLOGY/PRINCIPAL FINDINGS: A cross-sectional survey was conducted to investigate domestic dog demographics in Kalambabakali, in the rural Mazabuka District of Zambia. The population of ownerless dogs and the total achievable vaccination coverage among the total dog population was estimated using the capture-recapture-based Bayesian model by conducting a canine mass vaccination campaign. This study revealed that 29% of the domestic dog population was under one year old, and 57.7% of those were under three months old and thus were not eligible for the canine rabies vaccination in Zambia. The population growth was estimated at 15% per annum based on the cross-sectional household survey. The population of ownerless dogs was estimated to be small, with an ownerless-to-owned-dog ratio of 0.01-0.06 in the target zones. The achieved overall vaccination coverage from the first mass vaccination was estimated 19.8-51.6%. This low coverage was principally attributed to the owners' lack of information, unavailability, and dog-handling difficulties. The follow-up mass vaccination campaign achieved an overall coverage of 54.8-76.2%. CONCLUSIONS/SIGNIFICANCE: This paper indicates the potential for controlling canine rabies through mass vaccination in rural Zambia. Rabies education and responsible dog ownership are required to achieve high and sustainable vaccination coverage. Our findings also propose including puppies below three months old in the target population for rabies vaccination and emphasize that securing an annual enforcement of canine mass vaccination that reaches 70% coverage in the dog population is necessary to maintain protective herd immunity.

Prevalence and genetic diversity of Shibuyunji virus, a novel tick-borne phlebovirus identified in Zambia.
Edgar Simulundu; Saidon Mbambara; Herman M Chambaro; Karen Sichibalo; Masahiro Kajihara; King S Nalubamba; Hirofumi Sawa; Ayato Takada; Katendi Changula; Simbarashe Chitanga
Archives of virology, 166, 3, 915, 919, 2021年03月, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）, Tick-borne pathogens are an emerging public health threat worldwide. However, information on tick-borne viruses is scanty in sub-Saharan Africa. Here, by RT-PCR, 363 ticks (Amblyomma, Hyalomma and Rhipicephalus) in the Namwala and Livingstone districts of Zambia were screened for tick-borne phleboviruses (TBPVs). TBPVs (L gene) were detected in 19 (5.2%) Rhipicephalus ticks in Namwala. All the detected TBPVs were Shibuyunji viruses. Phylogenetically, they were closely related to American dog tick phlebovirus. This study highlights the possible role of Rhipicephalus ticks as the main host of Shibuyunji virus and suggests that these viruses may be present outside the area where they were initially discovered.

An African tick flavivirus forming an independent clade exhibits unique exoribonuclease-resistant RNA structures in the genomic 3'-untranslated region.
Hayato Harima; Yasuko Orba; Shiho Torii; Yongjin Qiu; Masahiro Kajihara; Yoshiki Eto; Naoya Matsuta; Bernard M Hang'ombe; Yuki Eshita; Kentaro Uemura; Keita Matsuno; Michihito Sasaki; Kentaro Yoshii; Ryo Nakao; William W Hall; Ayato Takada; Takashi Abe; Michael T Wolfinger; Martin Simuunza; Hirofumi Sawa
Scientific reports, 11, 1, 4883, 4883, 2021年03月01日, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）, Tick-borne flaviviruses (TBFVs) infect mammalian hosts through tick bites and can cause various serious illnesses, such as encephalitis and hemorrhagic fevers, both in humans and animals. Despite their importance to public health, there is limited epidemiological information on TBFV infection in Africa. Herein, we report that a novel flavivirus, Mpulungu flavivirus (MPFV), was discovered in a Rhipicephalus muhsamae tick in Zambia. MPFV was found to be genetically related to Ngoye virus detected in ticks in Senegal, and these viruses formed a unique lineage in the genus Flavivirus. Analyses of dinucleotide contents of flaviviruses indicated that MPFV was similar to those of other TBFVs with a typical vertebrate genome signature, suggesting that MPFV may infect vertebrate hosts. Bioinformatic analyses of the secondary structures in the 3'-untranslated regions (UTRs) revealed that MPFV exhibited unique exoribonuclease-resistant RNA (xrRNA) structures. Utilizing biochemical approaches, we clarified that two xrRNA structures of MPFV in the 3'-UTR could prevent exoribonuclease activity. In summary, our findings provide new information regarding the geographical distribution of TBFV and xrRNA structures in the 3'-UTR of flaviviruses.

Detection of B.1.351 SARS-CoV-2 Variant Strain - Zambia, December 2020.
Mulenga Mwenda; Ngonda Saasa; Nyambe Sinyange; George Busby; Peter J Chipimo; Jason Hendry; Otridah Kapona; Samuel Yingst; Jonas Z Hines; Peter Minchella; Edgar Simulundu; Katendi Changula; King Shimumbo Nalubamba; Hirofumi Sawa; Masahiro Kajihara; Junya Yamagishi; Muzala Kapin'a; Nathan Kapata; Sombo Fwoloshi; Paul Zulu; Lloyd B Mulenga; Simon Agolory; Victor Mukonka; Daniel J Bridges
MMWR. Morbidity and mortality weekly report, 70, 8, 280, 282, 2021年02月26日, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）, The first laboratory-confirmed cases of coronavirus disease 2019 (COVID-19), the illness caused by SARS-CoV-2, in Zambia were detected in March 2020 (1). Beginning in July, the number of confirmed cases began to increase rapidly, first peaking during July-August, and then declining in September and October (Figure). After 3 months of relatively low case counts, COVID-19 cases began rapidly rising throughout the country in mid-December. On December 18, 2020, South Africa published the genome of a SARS-CoV-2 variant strain with several mutations that affect the spike protein (2). The variant included a mutation (N501Y) associated with increased transmissibility.†,§ SARS-CoV-2 lineages with this mutation have rapidly expanded geographically.¶,** The variant strain (PANGO [Phylogenetic Assignment of Named Global Outbreak] lineage B.1.351††) was first detected in the Eastern Cape Province of South Africa from specimens collected in early August, spread within South Africa, and appears to have displaced the majority of other SARS-CoV-2 lineages circulating in that country (2). As of January 10, 2021, eight countries had reported cases with the B.1.351 variant. In Zambia, the average number of daily confirmed COVID-19 cases increased 16-fold, from 44 cases during December 1-10 to 700 during January 1-10, after detection of the B.1.351 variant in specimens collected during December 16-23. Zambia is a southern African country that shares substantial commerce and tourism linkages with South Africa, which might have contributed to the transmission of the B.1.351 variant between the two countries.

Purification of Crimean-Congo hemorrhagic fever virus nucleoprotein and its utility for serological diagnosis.
Boniface Pongombo Lombe; Hiroko Miyamoto; Takeshi Saito; Reiko Yoshida; Rashid Manzoor; Masahiro Kajihara; Masayuki Shimojima; Shuetsu Fukushi; Shigeru Morikawa; Tomoki Yoshikawa; Takeshi Kurosu; Masayuki Saijo; Qing Tang; Justin Masumu; David Hawman; Heinz Feldmann; Ayato Takada
Scientific reports, 11, 1, 2324, 2324, 2021年01月27日, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）, Crimean-Congo hemorrhagic fever virus (CCHFV) causes a zoonotic disease, Crimean-Congo hemorrhagic fever (CCHF) endemic in Africa, Asia, the Middle East, and Southeastern Europe. However, the prevalence of CCHF is not monitored in most of the endemic countries due to limited availability of diagnostic assays and biosafety regulations required for handling infectious CCHFV. In this study, we established a protocol to purify the recombinant CCHFV nucleoprotein (NP), which is antigenically highly conserved among multiple lineages/clades of CCHFVs and investigated its utility in an enzyme-linked immunosorbent assay (ELISA) to detect CCHFV-specific antibodies. The NP gene was cloned into the pCAGGS mammalian expression plasmid and human embryonic kidney 293 T cells were transfected with the plasmid. The expressed NP molecule was purified from the cell lysate using cesium-chloride gradient centrifugation. Purified NP was used as the antigen for the ELISA to detect anti-CCHFV IgG. Using the CCHFV NP-based ELISA, we efficiently detected CCHFV-specific IgG in anti-NP rabbit antiserum and CCHFV-infected monkey serum. When compared to the commercially available Blackbox CCHFV IgG ELISA kit, our assay showed equivalent performance in detecting CCHFV-specific IgG in human sera. These results demonstrate the usefulness of our CCHFV NP-based ELISA for seroepidemiological studies.

First COVID-19 case in Zambia - Comparative phylogenomic analyses of SARS-CoV-2 detected in African countries.
Edgar Simulundu; Francis Mupeta; Pascalina Chanda-Kapata; Ngonda Saasa; Katendi Changula; Walter Muleya; Simbarashe Chitanga; Miniva Mwanza; Paul Simusika; Herman Chambaro; Benjamin Mubemba; Masahiro Kajihara; Duncan Chanda; Lloyd Mulenga; Sombo Fwoloshi; Aaron Lunda Shibemba; Fred Kapaya; Paul Zulu; Kunda Musonda; Mwaka Monze; Nyambe Sinyange; Mazyanga L Mazaba; Muzala Kapin'a; Peter J Chipimo; Raymond Hamoonga; Davie Simwaba; William Ngosa; Albertina N Morales; Nkomba Kayeyi; John Tembo; Mathew Bates; Yasuko Orba; Hirofumi Sawa; Ayato Takada; King S Nalubamba; Kennedy Malama; Victor Mukonka; Alimuddin Zumla; Nathan Kapata
International journal of infectious diseases : IJID : official publication of the International Society for Infectious Diseases, 102, 455, 459, 2021年01月, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）, Since its first discovery in December 2019 in Wuhan, China, COVID-19, caused by the novel coronavirus SARS-CoV-2, has spread rapidly worldwide. While African countries were relatively spared initially, the initial low incidence of COVID-19 cases was not sustained for long due to continuing travel links between China, Europe and Africa. In preparation, Zambia had applied a multisectoral national epidemic disease surveillance and response system resulting in the identification of the first case within 48 h of the individual entering the country by air travel from a trip to France. Contact tracing showed that SARS-CoV-2 infection was contained within the patient's household, with no further spread to attending health care workers or community members. Phylogenomic analysis of the patient's SARS-CoV-2 strain showed that it belonged to lineage B.1.1., sharing the last common ancestor with SARS-CoV-2 strains recovered from South Africa. At the African continental level, our analysis showed that B.1 and B.1.1 lineages appear to be predominant in Africa. Whole genome sequence analysis should be part of all surveillance and case detection activities in order to monitor the origin and evolution of SARS-CoV-2 lineages across Africa.

A biaryl sulfonamide derivative as a novel inhibitor of filovirus infection.
Mao Isono; Wakako Furuyama; Makoto Kuroda; Tatsunari Kondoh; Manabu Igarashi; Masahiro Kajihara; Reiko Yoshida; Rashid Manzoor; Kosuke Okuya; Hiroko Miyamoto; Heinz Feldmann; Andrea Marzi; Masahiro Sakaitani; Asuka Nanbo; Ayato Takada
Antiviral research, 183, 104932, 104932, 2020年11月, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）, Ebolaviruses and marburgviruses, members of the family Filoviridae, are known to cause fatal diseases often associated with hemorrhagic fever. Recent outbreaks of Ebola virus disease in West African countries and the Democratic Republic of the Congo have made clear the urgent need for the development of therapeutics and vaccines against filoviruses. Using replication-incompetent vesicular stomatitis virus (VSV) pseudotyped with the Ebola virus (EBOV) envelope glycoprotein (GP), we screened a chemical compound library to obtain new drug candidates that inhibit filoviral entry into target cells. We discovered a biaryl sulfonamide derivative that suppressed in vitro infection mediated by GPs derived from all known human-pathogenic filoviruses. To determine the inhibitory mechanism of the compound, we monitored each entry step (attachment, internalization, and membrane fusion) using lipophilic tracer-labeled ebolavirus-like particles and found that the compound efficiently blocked fusion between the viral envelope and the endosomal membrane during cellular entry. However, the compound did not block the interaction of GP with the Niemann-Pick C1 protein, which is believed to be the receptor of filoviruses. Using replication-competent VSVs pseudotyped with EBOV GP, we selected escape mutants and identified two EBOV GP amino acid residues (positions 47 and 66) important for the interaction with this compound. Interestingly, these amino acid residues were located at the base region of the GP trimer, suggesting that the compound might interfere with the GP conformational change required for membrane fusion. These results suggest that this biaryl sulfonamide derivative is a novel fusion inhibitor and a possible drug candidate for the development of a pan-filovirus therapeutic.

Receptor-Mediated Host Cell Preference of a Bat-Derived Filovirus, Lloviu Virus.
Yoshihiro Takadate; Rashid Manzoor; Takeshi Saito; Yurie Kida; Junki Maruyama; Tatsunari Kondoh; Hiroko Miyamoto; Hirohito Ogawa; Masahiro Kajihara; Manabu Igarashi; Ayato Takada
Microorganisms, 8, 10, 2020年10月05日, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, Lloviu virus (LLOV), a bat-derived filovirus that is phylogenetically distinct from human pathogenic filoviruses such as Ebola virus (EBOV) and Marburg virus (MARV), was discovered in Europe. However, since infectious LLOV has never been isolated, the biological properties of this virus remain poorly understood. We found that vesicular stomatitis virus (VSV) pseudotyped with the glycoprotein (GP) of LLOV (VSV-LLOV) showed higher infectivity in one bat (Miniopterus sp.)-derived cell line than in the other bat-derived cell lines tested, which was distinct from the tropism of VSV pseudotyped with EBOV (VSV-EBOV) and MARV GPs. We then focused on the interaction between GP and Niemann-Pick C1 (NPC1) protein, one of the cellular receptors of filoviruses. We introduced the Miniopterus bat and human NPC1 genes into NPC1-knockout Vero E6 cells and their susceptibilities to the viruses were compared. The cell line expressing the bat NPC1 showed higher susceptibility to VSV-LLOV than that expressing human NPC1, whereas the opposite preference was seen for VSV-EBOV. Using a site-directed mutagenesis approach, amino acid residues involved in the differential tropism were identified in the NPC1 and GP molecules. Our results suggest that the interaction between GP and NPC1 is an important factor in the tropism of LLOV to a particular bat species.

Avian Influenza Viruses Detected in Birds in Sub-Saharan Africa: A Systematic Review.
Annie Kalonda; Ngonda Saasa; Panji Nkhoma; Masahiro Kajihara; Hirofumi Sawa; Ayato Takada; Edgar Simulundu
Viruses, 12, 9, 2020年09月07日, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）, In the recent past, sub-Saharan Africa has not escaped the devastating effects of avian influenza virus (AIV) in poultry and wild birds. This systematic review describes the prevalence, spatiotemporal distribution, and virus subtypes detected in domestic and wild birds for the past two decades (2000-2019). We collected data from three electronic databases, PubMed, SpringerLink electronic journals and African Journals Online, using the Preferred Reporting Items for Systematic reviews and Meta-Analyses protocol. A total of 1656 articles were reviewed, from which 68 were selected. An overall prevalence of 3.0% AIV in birds was observed. The prevalence varied between regions and ranged from 1.1% to 7.1%. The Kruskal-Wallis and Wilcoxon signed-rank sum test showed no significant difference in the prevalence of AIV across regions, χ2(3) = 5.237, p = 0.1553 and seasons, T = 820, z = -1.244, p = 0.2136. Nineteen hemagglutinin/neuraminidase subtype combinations were detected during the reviewed period, with southern Africa recording more diverse AIV subtypes than other regions. The most detected subtype was H5N1, followed by H9N2, H5N2, H5N8 and H6N2. Whilst these predominant subtypes were mostly detected in domestic poultry, H1N6, H3N6, H4N6, H4N8, H9N1 and H11N9 were exclusively detected in wild birds. Meanwhile, H5N1, H5N2 and H5N8 were detected in both wild and domestic birds suggesting circulation of these subtypes among wild and domestic birds. Our findings provide critical information on the eco-epidemiology of AIVs that can be used to improve surveillance strategies for the prevention and control of avian influenza in sub-Saharan Africa.

Characterization of mammalian orthoreoviruses isolated from faeces of pigs in Zambia.
Hayato Harima; Michihito Sasaki; Masahiro Kajihara; Gabriel Gonzalez; Edgar Simulundu; Eugene C Bwalya; Yongjin Qiu; Kosuke Okuya; Mao Isono; Yasuko Orba; Ayato Takada; Bernard M Hang'ombe; Aaron S Mweene; Hirofumi Sawa
The Journal of general virology, 101, 10, 1027, 1036, 2020年07月24日, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）, Mammalian orthoreovirus (MRV) has been identified in humans, livestock and wild animals; this wide host range allows individual MRV to transmit into multiple species. Although several interspecies transmission and genetic reassortment events of MRVs among humans, livestock and wildlife have been reported, the genetic diversity and geographic distribution of MRVs in Africa are poorly understood. In this study, we report the first isolation and characterization of MRVs circulating in a pig population in Zambia. In our screening, MRV genomes were detected in 19.7 % (29/147) of faecal samples collected from pigs by reverse transcription PCR. Three infectious MRV strains (MRV-85, MRV-96 and MRV-117) were successfully isolated, and their complete genomes were sequenced. Recombination analyses based on the complete genome sequences of the isolated MRVs demonstrated that MRV-96 shared the S3 segment with a different MRV isolated from bats, and that the L1 and M3 segments of MRV-117 originated from bat and human MRVs, respectively. Our results suggest that the isolated MRVs emerged through genetic reassortment events with interspecies transmission. Given the lack of information regarding MRVs in Africa, further surveillance of MRVs circulating among humans, domestic animals and wildlife is required to assess potential risk for humans and animals.

Comparative Analyses of the Antiviral Activities of IgG and IgA Antibodies to Influenza A Virus M2 Protein.
Kosuke Okuya; Nao Eguchi; Rashid Manzoor; Reiko Yoshida; Shinji Saito; Tadaki Suzuki; Michihito Sasaki; Takeshi Saito; Yurie Kida; Akina Mori-Kajihara; Hiroko Miyamoto; Osamu Ichii; Masahiro Kajihara; Hideaki Higashi; Ayato Takada
Viruses, 12, 7, 2020年07月20日, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, The influenza A virus (IAV) matrix-2 (M2) protein is an antigenically conserved viral envelope protein that plays an important role in virus budding together with another envelope protein, hemagglutinin (HA). An M2-specific mouse monoclonal IgG antibody, rM2ss23, which binds to the ectodomain of the M2 protein, has been shown to be a non-neutralizing antibody, but inhibits plaque formation of IAV strains. In this study, we generated chimeric rM2ss23 (ch-rM2ss23) IgG and IgA antibodies with the same variable region and compared their antiviral activities. Using gel chromatography, ch-rM2ss23 IgA were divided into three antibody subsets: monomeric IgA (m-IgA), dimeric IgA (d-IgA), and trimeric and tetrameric IgA (t/q-IgA). We found that t/q-IgA had a significantly higher capacity to reduce the plaque size of IAVs than IgG and m-IgA, most likely due to the decreased number of progeny virus particles produced from infected cells. Interestingly, HA-M2 colocalization was remarkably reduced on the infected cell surface in the presence of ch-rM2ss23 antibodies. These results indicate that anti-M2 polymeric IgA restricts IAV budding more efficiently than IgG and suggest a role of anti-M2 IgA in cross-protective immunity to IAVs.

Characterization of non-typhoid Salmonellae isolated from domestic animals and wildlife from selected areas of Zambia
Charles M. Mubita; Bwalya J. Muma; King Nalubamba; Girja Shanker Pandey; Kenny Samui; Musso Munyeme; Kajihara Masahiro; Yongjin Qiu; Ngonda Saasa; Bernard M. Hang'ombe
Scientific African, 8, e00345, e00345, Elsevier BV, 2020年07月, ［査読有り］, ［国際共著］, ［国際誌］
研究論文（学術雑誌）

Isolation of Candidatus Bartonella rousetti and Other Bat-associated Bartonellae from Bats and Their Flies in Zambia.
Yongjin Qiu; Masahiro Kajihara; Ryo Nakao; Evans Mulenga; Hayato Harima; Bernard Mudenda Hang'ombe; Yoshiki Eto; Katendi Changula; Daniel Mwizabi; Hirofumi Sawa; Hideaki Higashi; Aaron Mweene; Ayato Takada; Martin Simuunza; Chihiro Sugimoto
Pathogens (Basel, Switzerland), 9, 6, 2020年06月13日, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）, Bat-associated bartonellae, including Bartonella mayotimonensis and Candidatus Bartonella rousetti, were recently identified as emerging and potential zoonotic agents, respectively. However, there is no report of bat-associated bartonellae in Zambia. Thus, we aimed to isolate and characterize Bartonella spp. from bats and bat flies captured in Zambia by culturing and PCR. Overall, Bartonella spp. were isolated from six out of 36 bats (16.7%), while Bartonella DNA was detected in nine out of 19 bat flies (47.3%). Subsequent characterization using a sequence of five different genes revealed that three isolates obtained from Egyptian fruit bats (Rousettus aegyptiacus) were Ca. B. rousetti. The isolates obtained from insectivorous bats (Macronycteris vittatus) were divided into two previously unclassified bat-associated bartonellae. A phylogenetic analysis of the six genotypes of Bartonella gltA sequences from nine pathogen-positive bat flies revealed that three genotypes belonged to the same clades as bat-associated bartonellae, including Ca. B. rousetti. The other three genotypes represented arthropod-associated bartonellae, which have previously been isolated only from ectoparasites. We demonstrated that Ca. B. rousetti is maintained between bats (R. aegyptiacus) and bat flies in Zambia. Continuous surveillance of Bartonella spp. in bats and serological surveys in humans in Africa are warranted to evaluate the public health importance of bat-associated bartonellae.

Evidence for exposure of asymptomatic domestic pigs to African Swine fever virus during an inter-epidemic period in Zambia.
Herman M Chambaro; Michihito Sasaki; Yona Sinkala; Gabriel Gonzalez; David Squarre; Paul Fandamu; Caesar Lubaba; Liywalii Mataa; Misheck Shawa; Kabemba E Mwape; Sarah Gabriël; Mwelwa Chembensofu; Michael J Carr; William W Hall; Yongjin Qiu; Masahiro Kajihara; Ayato Takada; Yasuko Orba; Edgar Simulundu; Hirofumi Sawa
Transboundary and emerging diseases, 67, 6, 2741, 2752, 2020年05月20日, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）, African swine fever (ASF) causes persistent outbreaks in endemic and non-endemic regions in Zambia. However, the epidemiology of the disease is poorly understood, particularly during the inter-epidemic periods. We conducted surveillance for ASF in asymptomatic domestic pigs and soft ticks in selected Zambian provinces. Whilst serum samples (n=1,134) were collected from crossbred pigs from all study sites between 2014 and 2017, whole blood (n=300) was collected from both crossbred and indigenous pigs in Eastern Province (EP) in 2017. Soft ticks were collected from Mosi-oa-Tunya National Park in Southern Province (SP) in 2019. Sera were screened for antibodies against ASF by ELISA while genome detection in whole blood and soft ticks was conducted by PCR. Ticks were identified morphologically and by phylogenetic analysis of the 16S rRNA gene. Seroprevalence was highest in EP (50.9%, 95% CI [47.0 - 54.9]) compared to significantly lower rates in SP (2.9%, 95% CI [1.6 - 5.1]). No antibodies to ASFV were detected in Lusaka Province. In EP, the prevalence of ASFV genome was 11.7% (35/300), significantly higher (OR = 6.2, 95% CI [2.4 - 16.6]) in indigenous pigs compared to crossbred pigs. The pooled prevalence of ASFV genome in ticks was 11.0%, 95% CI [8.5-13.9]. Free-range husbandry system was the only factor that was significantly associated with seropositive (p < 0.0001, OR = 39.3) and PCR positive results (p < 0.001, OR = 5.7). Phylogenetically, based on the p72 gene, ASFV from Ornithodoros moubata ticks detected in this study belonged to genotype I, but they separated into two distinct clusters. Besides confirming ASF endemicity in EP and the presence of ASFV-infected ticks in SP, these results provide evidence for exposure of domestic pigs to ASFV in non-endemic regions during the inter-epidemic period.

A potential role of non-neutralizing IgA antibodies in cross-protective immunity against influenza A viruses of multiple hemagglutinin subtypes.
Kosuke Okuya; Reiko Yoshida; Rashid Manzoor; Shinji Saito; Tadaki Suzuki; Michihito Sasaki; Takeshi Saito; Yurie Kida; Akina Mori-Kajihara; Tatsunari Kondoh; Masahiro Sato; Masahiro Kajihara; Hiroko Miyamoto; Osamu Ichii; Hideaki Higashi; Ayato Takada
Journal of virology, 94, 12, 2020年04月08日, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, IgA antibodies on mucosal surfaces are known to play an important role in protection from influenza A virus (IAV) infection and are believed to be more potent than IgG for cross-protective immunity against IAVs of multiple hemagglutinin (HA) subtypes. However, in general, neutralizing antibodies specific to HA are principally HA subtype-specific. Here we focus on non-neutralizing but broadly cross-reactive HA-specific IgA antibodies. Recombinant IgG, monomeric IgA (mIgA), and polymeric secretory IgA (pSIgA) antibodies were generated based on the sequence of a mouse anti-HA monoclonal antibody (MAb) 5A5 that had no neutralizing activity but showed broad binding capacity to multiple HA subtypes. While confirming that there was no neutralizing activity of the recombinant MAbs against IAV strains A/Puerto Rico/8/1934 (H1N1), A/Adachi/2/1957 (H2N2), A/Hong Kong/483/1997 (H5N1), A/shearwater/South Australia/1/1972 (H6N5), A/duck/England/1/1956 (H11N6), and A/duck/Alberta/60/1976 (H12N5), we found that pSIgA, but not mIgA and IgG, significantly reduced budding and release of most of the viruses from infected cells. Electron microscopy demonstrated that pSIgA deposited newly produced virus particles on the surfaces of infected cells, most likely due to tethering of virus particles. Furthermore, we found that pSIgA showed significantly higher activity to reduce plaque sizes of the viruses than IgG and mIgA. These results suggest that non-neutralizing pSIgA reactive to multiple HA subtypes may play a role in inter-subtype cross-protective immunity against IAVs.Importance Mucosal immunity represented by pSIgA plays important roles in protection from IAV infection. Furthermore, IAV HA-specific pSIgA antibodies are thought to contribute to cross-protective immunity against multiple IAV subtypes. However, the mechanisms by which pSIgA exerts such versatile antiviral activity are not fully understood. In this study, we generated broadly cross-reactive recombinant IgG and pSIgA having the same antigen-recognition site and compared their antiviral activities in vitro These recombinant antibodies did not show "classical" neutralizing activity, whereas pSIgA, but not IgG, significantly inhibited the production of progeny virus particles from infected cells. Plaque formation was also significantly reduced by pSIgA, but not IgG. These effects were seen in infection with IAVs of several different HA subtypes. Based on our findings, we propose an antibody-mediated host defense mechanism by which mucosal immunity may contribute to broad cross-protection from IAVs of multiple HA subtypes, including viruses with pandemic potential.

West Nile Virus in Farmed Crocodiles, Zambia, 2019.
Edgar Simulundu; Kunda Ndashe; Herman M Chambaro; David Squarre; Paul Michael Reilly; Simbarashe Chitanga; Katendi Changula; Andrew N Mukubesa; Joseph Ndebe; John Tembo; Nathan Kapata; Matthew Bates; Yona Sinkala; Bernard M Hang'ombe; King S Nalubamba; Masahiro Kajihara; Michihito Sasaki; Yasuko Orba; Ayato Takada; Hirofumi Sawa
Emerging infectious diseases, 26, 4, 811, 814, 2020年04月, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）, We detected West Nile virus (WNV) nucleic acid in crocodiles (Crocodylus niloticus) in Zambia. Phylogenetically, the virus belonged to lineage 1a, which is predominant in the Northern Hemisphere. These data provide evidence that WNV is circulating in crocodiles in Africa and increases the risk for animal and human transmission.

Genetic and Biological Diversity of Porcine Sapeloviruses Prevailing in Zambia.
Hayato Harima; Masahiro Kajihara; Edgar Simulundu; Eugene Bwalya; Yongjin Qiu; Mao Isono; Kosuke Okuya; Gabriel Gonzalez; Junya Yamagishi; Bernard M Hang'ombe; Hirofumi Sawa; Aaron S Mweene; Ayato Takada
Viruses, 12, 2, 2020年02月05日, ［査読有り］, ［責任著者］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）, Porcine sapelovirus (PSV) has been detected worldwide in pig populations. Although PSV causes various symptoms such as encephalomyelitis, diarrhea, and pneumonia in pigs, the economic impact of PSV infection remains to be determined. However, information on the distribution and genetic diversity of PSV is quite limited, particularly in Africa. In this study, we investigated the prevalence of PSV infection in Zambia and characterized the isolated PSVs genetically and biologically. We screened 147 fecal samples collected in 2018 and found that the prevalences of PSV infection in suckling pigs and fattening pigs were high (36.2% and 94.0%, respectively). Phylogenetic analyses revealed that the Zambian PSVs were divided into three different lineages (Lineages 1-3) in the clade consisting of Chinese strains. The Zambian PSVs belonging to Lineages 2 and 3 replicated more efficiently than those belonging to Lineage 1 in Vero E6 and BHK cells. Bioinformatic analyses revealed that genetic recombination events had occurred and the recombination breakpoints were located in the L and 2A genes. Our results indicated that at least two biologically distinct PSVs could be circulating in the Zambian pig population and that genetic recombination played a role in the evolution of PSVs.

Detection of novel orthoreovirus genomes in shrew (Crocidura hirta) and fruit bat (Rousettus aegyptiacus).
Hayato Harima; Michihito Sasaki; Masahiro Kajihara; Akina Mori-Kajihara; Bernard M Hang'ombe; Katendi Changula; Yasuko Orba; Hirohito Ogawa; Martin Simuunza; Reiko Yoshida; Aaron Mweene; Ayato Takada; Hirofumi Sawa
The Journal of veterinary medical science, 82, 2, 162, 167, 2020年02月04日, ［査読有り］, ［国際共著］, ［国内誌］
英語, 研究論文（学術雑誌）, Orthoreoviruses have been indentified in several mammals, however, there is no information about orthoreoviruses in shrews. In this study, we screened wild animals in Zambia, including shrews, rodents, and bats for the detection of orthoreoviruses. Two orthoreovirus RNA genomes were detected from a shrew intestinal-contents (1/24) and a bat colon (1/96) sample by reverse-transcription (RT)-PCR targeting the RNA-dependent RNA polymerase gene of orthoreoviruses. Phylogenetic analyses revealed that each of the identified orthoreoviruses formed a distinct branch among members of the Orthoreovirus genus. This is the first report that shrews are susceptible to orthoreovirus infection. Our results suggest the existence of undiscovered orthoreoviruses in shrews and provide important information about the genetic diversity of orthoreoviruses.

Niemann-Pick C1 Heterogeneity of Bat Cells Controls Filovirus Tropism.
Yoshihiro Takadate; Tatsunari Kondoh; Manabu Igarashi; Junki Maruyama; Rashid Manzoor; Hirohito Ogawa; Masahiro Kajihara; Wakako Furuyama; Masahiro Sato; Hiroko Miyamoto; Reiko Yoshida; Terence E Hill; Alexander N Freiberg; Heinz Feldmann; Andrea Marzi; Ayato Takada
Cell reports, 30, 2, 308, 319, 2020年01月14日, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）, Fruit bats are suspected to be natural hosts of filoviruses, including Ebola virus (EBOV) and Marburg virus (MARV). Interestingly, however, previous studies suggest that these viruses have different tropisms depending on the bat species. Here, we show a molecular basis underlying the host-range restriction of filoviruses. We find that bat-derived cell lines FBKT1 and ZFBK13-76E show preferential susceptibility to EBOV and MARV, respectively, whereas the other bat cell lines tested are similarly infected with both viruses. In FBKT1 and ZFBK13-76E, unique amino acid (aa) sequences are found in the Niemann-Pick C1 (NPC1) protein, one of the cellular receptors interacting with the filovirus glycoprotein (GP). These aa residues, as well as a few aa differences between EBOV and MARV GPs, are crucial for the differential susceptibility to filoviruses. Taken together, our findings indicate that the heterogeneity of bat NPC1 orthologs is an important factor controlling filovirus species-specific host tropism.

Seroprevalence of Rift Valley fever in cattle of smallholder farmers in Kwilu Province in the Democratic Republic of Congo.
Abdel-Amir Dib Halawi; Ngonda Saasa; Boniface Lombe Pongombo; Masahiro Kajihara; Herman Moses Chambaro; Mutambel Hity; Hirofumi Sawa; Ayato Takada; Aaron S Mweene; Luamba Lua Nsembo; Edgar Simulundu
Tropical animal health and production, 51, 8, 2619, 2627, 2019年11月, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）, Rift Valley fever (RVF) is a zoonotic mosquito-borne disease caused by RVF virus (RVFV) that causes abortions and high mortalities in livestock and is also associated with acute and fatal disease in humans. In the Democratic Republic of Congo (DRC), information on the epidemiology of RVF is limited, particularly among cattle reared by smallholder farmers. This cross-sectional study was conducted to investigate the seroprevalence of RVF in cattle raised by smallholder farmers in Kwilu Province of DRC, which has not yet reported an RVF epidemic. A total of 677 cattle sera were collected from four territories and tested for anti-RVFV antibodies using immunofluorescent assay and enzyme-linked immunosorbent assay. The overall seroprevalence of anti-RVFV IgG was 6.5% (44/677) (95% CI 4.81-8.7). There was a statistically significant difference in the seroprevalence among the territories (χ2 = 28.79, p < 0.001). Territory seroprevalences were as follows: Idiofa 14.08% (95% CI 9.78-19.76), Bulungu 4.14% (95% CI 1.83-8.68), Gungu 3.21% (95% CI 1.41-6.78), and Masi-Manimba 1.19% (95% CI 0.06-7.37). Seroprevalence differed significantly among age categories (p = 0.0017) and ecosystem (p < 0.001). The seroprevalence of animals aged between 1 and 2 years was 20.0% (95% CI 8.4-39.13) and was higher than group aged <1 year, between 2 and 3 years, and > 3 years. Forest area (18.92% (95% CI 12.35-27.7)) had higher seropositivity than savannah area (4.06% (95% CI 2.65-6.12)). Sex difference was not significant (χ2 = 0.14, p = 0.704). These findings indicate that cattle in Kwilu Province had been exposed to RVFV, which represents a significant risk for both livestock and human health.

Molecular detection and characterization of genotype 1 bovine leukemia virus from beef cattle in the traditional sector in Zambia.
Mundia M Phiri; Evans Kaimoyo; Katendi Changula; Isaac Silwamba; Herman M Chambaro; Penjaninge Kapila; Masahiro Kajihara; Martin Simuunza; John Bwalya Muma; Girja S Pandey; Ayato Takada; Aaron S Mweene; Simbarashe Chitanga; Edgar Simulundu
Archives of virology, 164, 10, 2531, 2536, 2019年10月, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）, Whilst bovine leukemia virus (BLV) causes considerable economic losses to the dairy industry worldwide, information on its molecular epidemiology and economic impact in beef cattle is limited. Here, blood from 880 animals from Zambia's major cattle-rearing provinces was screened for BLV by nested PCR. Positive pools were sequenced and phylogenetically analyzed. The estimated pooled prevalence was 2.1%. All strains belonged to genotype 1 and formed a distinct phylogenetic cluster. The study suggests circulation of genotype 1 BLV in beef cattle in these regions. This is the first report on molecular detection and characterization of BLV from beef cattle in Africa.

Genetic diversity of rabies virus in different host species and geographic regions of Zambia and Zimbabwe.
Muleya W; Chambaro HM; Sasaki M; Gwenhure LF; Mwenechanya R; Kajihara M; Saasa N; Mupila Z; Mori-Kajihara A; Qiu Y; Kangwa E; Mweene A; Namangala B; Takada A; Sawa H
Virus genes, 55, 5, 713, 719, 2019年10月, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）, Rabies is endemic in Zambia and Zimbabwe. The previously investigated strains of rabies virus in central Zambia belong to the Africa 1b lineage, with similar circulating virus strains found in the various tested hosts and regions. However, prior work assessed only limited regions and host species. Thus, this study aimed to more comprehensively determine the genetic diversity of rabies virus across regions of Zambia and Zimbabwe. RNA (n = 76) was extracted from positive direct fluorescent antibody test brain tissues from dog, cow, goat, cat, pig, human, and jackal collected from Zambia and Zimbabwe. The amplicons of the nucleoprotein and glycoprotein genes were obtained from all examined samples by nested RT-PCR and subsequently sequenced. A phylogenetic analysis of the N gene confirmed that all the endemic strains of rabies virus in Zambia and Zimbabwe belong to the Africa 1b lineage. The obtained viral gene sequences were phylogenetically divided into two clusters. Cluster II comprised only Zambian strains. In contrast, cluster I comprised both Zambia and Zimbabwe strains, with strains from Zimbabwe forming a distinct lineage from Zambian strains, implying viral genetic divergence due to geographical barriers. However, no evidence of clustering based on host or region was observed, implying the circulation of similar virus strains occurs in different hosts and regions of Zambia and Zimbabwe. The clustering of rabies virus strains from jackals with those from domestic animals provides evidence of similar virus strains circulating in both wildlife and domestic animals, and that the jackal might be one of the potential reservoirs of rabies virus infection. In this study, no strains circulating in Zimbabwe were detected in Zambia.

Marburgvirus in Egyptian Fruit Bats, Zambia.
Kajihara M; Hang'ombe BM; Changula K; Harima H; Isono M; Okuya K; Yoshida R; Mori-Kajihara A; Eto Y; Orba Y; Ogawa H; Qiu Y; Sawa H; Simulundu E; Mwizabi D; Munyeme M; Squarre D; Mukonka V; Mweene A; Takada A
Emerging infectious diseases, 25, 8, 1577, 1580, 2019年08月, ［査読有り］, ［筆頭著者］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）, We detected Marburg virus genome in Egyptian fruit bats (Rousettus aegyptiacus) captured in Zambia in September 2018. The virus was closely related phylogenetically to the viruses that previously caused Marburg outbreaks in the Democratic Republic of the Congo. This finding demonstrates that Zambia is at risk for Marburg virus disease.

Clinical Evaluation of QuickNaviTM-Ebola in the 2018 Outbreak of Ebola Virus Disease in the Democratic Republic of the Congo.
Sheila Makiala; Daniel Mukadi; Anja De Weggheleire; Shino Muramatsu; Daisuke Kato; Koichi Inano; Fumio Gondaira; Masahiro Kajihara; Reiko Yoshida; Katendi Changula; Aaron Mweene; Placide Mbala-Kingebeni; Jean-Jacques Muyembe-Tamfum; Justin Masumu; Steve Ahuka; Ayato Takada
Viruses, 11, 7, 2019年06月28日, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）, The recent large outbreaks of Ebola virus disease (EVD) in West Africa and the Democratic Republic of the Congo (DRC) have highlighted the need for rapid diagnostic tests to control this disease. In this study, we clinically evaluated a previously developed immunochromatography-based kit, QuickNaviTM-Ebola. During the 2018 outbreaks in DRC, 928 blood samples from EVD-suspected cases were tested with QuickNaviTM-Ebola and the WHO-approved GeneXpert. The sensitivity and specificity of QuickNaviTM-Ebola, estimated by comparing it to GeneXpert-confirmed cases, were 85% (68/80) and 99.8% (846/848), respectively. These results indicate the practical reliability of QuickNaviTM-Ebola for point-of-care diagnosis of EVD.

Human Borreliosis Caused by a New World Relapsing Fever Borrelia-like Organism in the Old World.
Yongjin Qiu; Ryo Nakao; Bernard Mudenda Hang'ombe; Kozue Sato; Masahiro Kajihara; Sharon Kanchela; Katendi Changula; Yoshiki Eto; Joseph Ndebe; Michihito Sasaki; May June Thu; Ayato Takada; Hirohumi Sawa; Chihiro Sugimoto; Hiroki Kawabata
Clinical infectious diseases : an official publication of the Infectious Diseases Society of America, 69, 1, 107, 112, 2019年06月18日, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）, BACKGROUND: Relapsing fever is an infectious disease previously neglected in Africa, which imposes a large public health burden in the country. We aimed to investigate and report on a case of relapsing fever borreliosis in Zambia. METHODS: A previously unknown Borrelia species was isolated from the blood of a febrile patient. Investigations of the presumptive vector ticks and natural hosts for the Borrelia species were conducted by culture isolation and/or DNA detection by Borrelia-specific polymerase chain reaction. Using culture isolates from the patient and bat specimens, genetic characterization was performed by multilocus sequence analysis based on the draft genome sequences. RESULTS: The febrile patient was diagnosed with relapsing fever. The isolated Borrelia species was frequently detected in Ornithodoros faini (n = 20/50 [40%]) and bats (n = 64/237 [27%]). Multilocus sequence analysis based on a draft genome sequence revealed that the Borrelia species isolates from the patient and presumptive reservoir host (bats) formed a monophyletic lineage that clustered with relapsing fever borreliae found in the United States. CONCLUSIONS: A febrile illness caused by a Borrelia species that was treatable with erythromycin was identified in Zambia. This is the first study to report on relapsing fever Borrelia in Zambia and suggesting the likely natural reservoir hosts of the isolated Borrelia species. Interestingly, the isolated Borrelia species was more closely related to New World relapsing fever borreliae, despite being detected in the Afrotropic ecozone.

Molecular characterization and phylogenetic analysis of Trypanosoma spp. isolated from striped leaf-nosed bat (Hipposideros vittatus) in Zambia.
Qiu Y; Simuunza M; Mwizabi D; Changula K; Harima H; Takada A; Kajihara M; Takadate Y; Nakao R; Kawabata H; Sugimoto C; Mweene A; Sawa H; Eto Y; Mudenda Hang’ombe B; Hayashida K; Yoshida R; Mori-Kajihara A; Ndebe J
International Journal for Parasitology, 9, 234, 238, 2019年04月, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）, Bat trypanosomes consist of more than 30 trypanosome species from over 70 species of bats. Recent studies suggest that bats play a role in disseminating trypanosomes from African continent to the terrestrial mammals both in the Afrotropic-Palearctic Ecozones and Nearctic Ecozone. However, the diversity, distribution, and evolution of bat trypanosomes are still unclear. To better understand their evolution, more genetic data of bat trypanosomes from a variety of locations are required. During a survey of Borrelia spp. of bats inhabiting a cave in Zambia, we observed flagellate parasites from 5 of 43 hemocultures. Sequence and phylogenetic analyses of the glycosomal glyceraldehyde 3-phosphate dehydrogenase gene (gGAPDH; 572 bp) and the 18S ribosomal RNA gene (18S rRNA gene; 1,079-1,091 bp) revealed that all were Trypanosoma spp. belonged to the Trypanosoma cruzi clade. Three and two of them exhibited the similarity with T. conorhini and T. dionisii, respectively. The present study provides the first genetic data on Trypanosoma spp. of bats inhabiting Zambia.

Diversity of spotted fever group rickettsiae and their association with host ticks in Japan.
Thu MJ; Qiu Y; Matsuno K; Kajihara M; Mori-Kajihara A; Omori R; Monma N; Chiba K; Seto J; Gokuden M; Andoh M; Oosako H; Katakura K; Takada A; Sugimoto C; Isoda N; Nakao R
Scientific reports, 9, 1, 1500, 1500, 2019年02月06日, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）, Spotted fever group (SFG) rickettsiae are obligate intracellular Gram-negative bacteria mainly associated with ticks. In Japan, several hundred cases of Japanese spotted fever, caused by Rickettsia japonica, are reported annually. Other Rickettsia species are also known to exist in ixodid ticks; however, their phylogenetic position and pathogenic potential are poorly understood. We conducted a nationwide cross-sectional survey on questing ticks to understand the overall diversity of SFG rickettsiae in Japan. Out of 2,189 individuals (19 tick species in 4 genera), 373 (17.0%) samples were positive for Rickettsia spp. as ascertained by real-time PCR amplification of the citrate synthase gene (gltA). Conventional PCR and sequencing analyses of gltA indicated the presence of 15 different genotypes of SFG rickettsiae. Based on the analysis of five additional genes, we characterised five Rickettsia species; R. asiatica, R. helvetica, R. monacensis (formerly reported as Rickettsia sp. In56 in Japan), R. tamurae, and Candidatus R. tarasevichiae and several unclassified SFG rickettsiae. We also found a strong association between rickettsial genotypes and their host tick species, while there was little association between rickettsial genotypes and their geographical origins. These observations suggested that most of the SFG rickettsiae have a limited host range and are maintained in certain tick species in the natural environment.

Generation of bat-derived influenza viruses and their reassortants.
Sato M; Maruyama J; Kondoh T; Nao N; Miyamoto H; Takadate Y; Furuyama W; Kajihara M; Ogawa H; Manzoor R; Yoshida R; Igarashi M; Takada A
Scientific reports, 9, 1, 1158, 1158, 2019年02月, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, Two novel influenza A virus-like genomes were detected in fruit bats in Central and South America. However, the biological properties of these bat-derived influenza viruses (BatIVs) are still largely unknown since infectious viral particles have never been isolated from the infected host species. In this study, a reverse genetics approach was used to generate infectious BatIV particles entirely from plasmids encoding full-length sequences in eight gene segments. We inoculated BatIV particles into various cell cultures including bat-derived cell lines and found that BatIVs infected particular bat-derived cells efficiently but not the other cell lines tested. Reassortant viruses between the two BatIVs were also successfully generated and their replication in the susceptible bat cell lines was confirmed. These findings suggest a limited host range and reassortment potential of BatIVs in nature, providing fundamental information for understanding of the ecology of BatIVs.

Infection of newly identified phleboviruses in ticks and wild animals in Hokkaido, Japan indicating tick-borne life cycles.
Shiho Torii; Keita Matsuno; Yongjin Qiu; Akina Mori-Kajihara; Masahiro Kajihara; Ryo Nakao; Naganori Nao; Katsunori Okazaki; Mariko Sashika; Takahiro Hiono; Masatoshi Okamatsu; Yoshihiro Sakoda; Hideki Ebihara; Ayato Takada; Hirofumi Sawa
Ticks and tick-borne diseases, 10, 2, 328, 335, 2019年02月, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, Recent discoveries of tick-borne pathogens have raised public health concerns on tick-borne infectious diseases and emphasize the need to assess potential risks of unrecognized tick-borne pathogens. First, to determine the existence of tick-borne phleboviruses (TBPVs), genetic surveillance of phleboviruses in ticks was conducted mainly in Hokkaido, the northernmost island in Japan from 2013 to 2015. Genes of two TBPVs, previously reported as Mukawa virus (MKWV) and a newly identified relative of MKWV, Kuriyama virus (KURV), were detected and the viruses were isolated from Ixodes persulcatus collected in Hokkaido, but not in I. persulcatus collected from other areas of Japan. These viruses were phylogenetically and antigenically similar to each other. Next, to investigate the infection of MKWV in mammals, serum samples from wildlife captured in Hokkaido from 2007 to 2011 were used for serological screening. Neutralizing antibodies against MKWV were detected in both Yezo-deer (Cervus nippon yesoensis) (2/50) and raccoons (Procyon lotor) (16/64). However, no infectious MKWV was recovered from laboratory mice in experimental infections, though viral RNAs were detected in their tissues. Thus, MKWV and KURV may maintain tick-mammalian life cycles in Hokkaido, suggesting their potential as causative agents of tick-borne diseases in mammals.

Seroprevalence of Filovirus Infection of Rousettus aegyptiacus Bats in Zambia.
Katendi Changula; Masahiro Kajihara; Akina Mori-Kajihara; Yoshiki Eto; Hiroko Miyamoto; Reiko Yoshida; Asako Shigeno; Bernard Hang'ombe; Yongjin Qiu; Daniel Mwizabi; David Squarre; Joseph Ndebe; Hirohito Ogawa; Hayato Harima; Edgar Simulundu; Ladslav Moonga; Penjaninge Kapila; Wakako Furuyama; Tatsunari Kondoh; Masahiro Sato; Yoshihiro Takadate; Chiho Kaneko; Ryo Nakao; Victor Mukonka; Aaron Mweene; Ayato Takada
The Journal of infectious diseases, 218, suppl_5, S312-S317, 2018年11月22日, ［査読有り］, ［筆頭著者］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）, Bats are suspected to play important roles in the ecology of filoviruses, including ebolaviruses and marburgviruses. A cave-dwelling fruit bat, Rousettus aegyptiacus, has been shown to be a reservoir of marburgviruses. Using an enzyme-linked immunosorbent assay with the viral glycoprotein antigen, we detected immunoglobulin G antibodies specific to multiple filoviruses in 158 of 290 serum samples of R aegyptiacus bats captured in Zambia during the years 2014-2017. In particular, 43.8% of the bats were seropositive to marburgvirus, supporting the notion that this bat species continuously maintains marburgviruses as a reservoir. Of note, distinct peaks of seropositive rates were repeatedly observed at the beginning of rainy seasons, suggesting seasonality of the presence of newly infected individuals in this bat population. These data highlight the need for continued monitoring of filovirus infection in this bat species even in countries where filovirus diseases have not been reported.

Identification of group A rotaviruses from Zambian fruit bats provides evidence for long-distance dispersal events in Africa
Michihito Sasaki; Masahiro Kajihara; Katendi Changula; Akina Mori-Kajihara; Hirohito Ogawa; Bernard M. Hang'ombe; Aaron S. Mweene; Martin Simuunza; Reiko Yoshida; Michael Carr; Yasuko Orba; Ayato Takada; Hirofumi Sawa
Infection, Genetics and Evolution, 63, 104, 109, Elsevier B.V., 2018年09月01日, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）

First isolation of West Nile virus in Zambia from mosquitoes.
Orba Y; Hang'ombe BM; Mweene AS; Wada Y; Anindita PD; Phongphaew W; Qiu Y; Kajihara M; Mori-Kajihara A; Eto Y; Sasaki M; Hall WW; Eshita Y; Sawa H
Transboundary and emerging diseases, 65, 4, 933, 938, 2018年08月, ［査読有り］, ［国際共著］

The Unique Phylogenetic Position of a Novel Tick-Borne Phlebovirus Ensures an Ixodid Origin of the Genus Phlebovirus.
Keita Matsuno; Masahiro Kajihara; Ryo Nakao; Naganori Nao; Akina Mori-Kajihara; Mieko Muramatsu; Yongjin Qiu; Shiho Torii; Manabu Igarashi; Nodoka Kasajima; Keita Mizuma; Kentaro Yoshii; Hirofumi Sawa; Chihiro Sugimoto; Ayato Takada; Hideki Ebihara
mSphere, 3, 3, 2018年06月27日, ［査読有り］, ［筆頭著者］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）, The recent emergence of novel tick-borne RNA viruses has complicated the epidemiological landscape of tick-borne infectious diseases, posing a significant challenge to public health systems that seek to counteract tick-borne diseases. The identification of two novel tick-borne phleboviruses (TBPVs), severe fever with thrombocytopenia syndrome virus (SFTSV) and Heartland virus (HRTV), as causative agents of severe illness in humans has accelerated the investigation and discoveries of novel TBPVs. In the present study, we isolated a novel TBPV designated Mukawa virus (MKWV) from host-questing Ixodes persulcatus females captured in Japan. Genetic characterization revealed that MKWV is a member of the genus Phlebovirus in the family Phenuiviridae Interestingly, MKWV is genetically distinct from other known TBPVs and shares a most recent common ancestor with mosquito/sandfly-borne (insect-borne) phleboviruses. Despite its genetic similarity to insect-borne phleboviruses, the molecular footprints of its viral proteins and its biological characteristics define MKWV as a tick-borne virus that can be transmitted to mammals. A phylogenetic ancestral-state reconstruction for arthropod vectors of phleboviruses including MKWV based on viral L segment sequences indicated that ticks likely harbored ancestral phleboviruses that evolved into both the tick-borne and MKWV/insect-borne phlebovirus lineages. Overall, our findings suggest that most of the phlebovirus evolution has occurred in hard ticks to generate divergent viruses, which may provide a seminal foundation for understanding the mechanisms underlying the evolution and emergence of pathogenic phleboviruses, such as Rift Valley fever virus and SFTSV/HRTV.IMPORTANCE The emergence of novel tick-borne RNA viruses causing severe illness in humans has complicated the epidemiological landscape of tick-borne diseases, requiring further investigation to safeguard public health. In the present study, we discovered a novel tick-borne phlebovirus from Ixodes persulcatus ticks in Japan. While its viral RNA genome sequences were similar to those of mosquito/sandfly-borne viruses, molecular and biological footprints confirmed that this is a tick-borne virus. The unique evolutionary position of the virus allowed us to estimate the ancestral phlebovirus vector, which was likely a hard tick. Our findings may provide a better understanding of the evolution and emergence of phleboviruses associated with emerging infectious diseases, such as severe fever with thrombocytopenia syndrome (SFTS) and Heartland virus disease.

Monoclonal Antibody Cocktail Protects Hamsters From Lethal Marburg Virus Infection.
Marzi A; Haddock E; Kajihara M; Feldmann H; Takada A
The Journal of infectious diseases, 218, suppl_5, S662-S665, 2018年06月, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）, Marburg virus (MARV), family Filoviridae, causes Marburg hemorrhagic fever (MHF) in humans and nonhuman primates with case fatality rates of up to 90%. There is no approved therapeutic for MHF, yet several experimental approaches have been evaluated in preclinical studies including small interfering RNA and monoclonal antibody (mAb) treatment. In this study we attempted to improve the therapeutic efficacy of the neutralizing mAb M4 by combining treatment with 1 or 2 of blocking but nonneutralizing mAbs 126-15 and 127-8. We found that single-dose treatment early after infection with the neutralizing mAb M4 or any of the mAb combinations resulted in similar protection in the MARV hamster model. However, a single-dose treatment with the cocktail of all 3 mAbs provided the best protection in delayed treatment, with 67%-100% of the animals surviving a lethal challenge depending on the time of treatment. This study identified a new promising mAb cocktail as a therapeutic option for MHF.

Discovery of Mwinilunga alphavirus: A novel alphavirus in Culex mosquitoes in Zambia.
Shiho Torii; Yasuko Orba; Bernard M Hang'ombe; Aaron S Mweene; Yuji Wada; Paulina D Anindita; Wallaya Phongphaew; Yongjin Qiu; Masahiro Kajihara; Akina Mori-Kajihara; Yoshiki Eto; Hayato Harima; Michihito Sasaki; Michael Carr; William W Hall; Yuki Eshita; Takashi Abe; Hirofumi Sawa
Virus research, 250, 31, 36, 2018年05月02日, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）, Mosquito-borne alphaviruses are disseminated globally and cause febrile illness in humans and animals. Since the prevalence and diversity of alphaviruses has not been previously investigated in Zambia, reverse transcription PCR was employed as a broad-spectrum approach for the detection of alphaviruses in mosquitoes. From 552 mosquito pools, a novel alphavirus, tentatively named Mwinilunga alphavirus (MWAV), was discovered from a single Culex quinquefasciatus mosquito pool. The full genome of MWAV was subsequently determined, and pairwise comparisons demonstrated that MWAV represented a new alphavirus species. Phylogenetic analyses and a linear discriminant analysis based on the dinucleotide ratios in various virus sequences indicated that MWAV is related to a mosquito-specific alphavirus distinct from other known mosquito-borne alphaviruses due to its inability to replicate in vertebrate cell lines. Further analyses of these novel alphaviruses will help to facilitate a greater understanding of the molecular determinants of host range restriction and the evolutionary relationships of alphaviruses.

Tick-borne haemoparasites and Anaplasmataceae in domestic dogs in Zambia.
Yongjin Qiu; Chiho Kaneko; Masahiro Kajihara; Saasa Ngonda; Edgar Simulundu; Walter Muleya; May June Thu; Mudenda Bernard Hang'ombe; Ken Katakura; Ayato Takada; Hirofumi Sawa; Martin Simuunza; Ryo Nakao
Ticks and tick-borne diseases, 9, 4, 988, 995, 2018年05月, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）, Tick-borne diseases (TBDs), including emerging and re-emerging infectious diseases, are important threats to human and animal health worldwide. Indeed, the number of reported human and animal infectious cases of novel TBD agents has increased in recent decades. However, TBDs tend to be neglected, especially in resource-limited countries that often have limited diagnostic capacity. The aim of this molecular survey was to detect and characterise tick-borne pathogens (Babesia, Theileria, and Hepatozoon parasites and Anaplasmataceae bacteria) in domestic dogs in Zambia. In total, 247 canine peripheral blood samples were collected in Lusaka, Mazabuka, Monze, and Shangombo. Conventional PCR to detect the selected pathogens was performed using DNA extracted from canine blood. One hundred eleven samples were positive for protozoa and 5 were positive for Anaplasmataceae. Sequencing of thirty-five randomly selected protozoa-positive samples revealed the presence of Babesia rossi, Babesia vogeli, and Hepatozoon canis 18S rDNA. Based on these sequences, a multiplex PCR system was developed to yield PCR products with different amplicons, the size of which depended on the parasite species; thus, each species could be identified without the need for sequence analysis. Approximately 40% of dogs were positive for H. canis. In particular, the positive rate (75.2%) of H. canis infection was significantly higher in Shangombo than in other sampling sites. Multiplex PCR assay detected B. rossi and B. vogeli infections in five and seven dogs, respectively, indicating that this approach is useful for detecting parasites with low prevalence. Sequencing analysis of gltA and groEL genes of Anaplasmataceae revealed that two and one dogs in Lusaka were infected with Anaplasma platys and Ehrlichia canis, respectively. The data indicated that Zambian dogs were infected with multiple tick-borne pathogens such as H. canis, B. rossi, B. vogeli, A. platys, E. canis and uncharacterized Ehrlichia sp. Since some of these parasites are zoonotic, concerted efforts are needed to raise awareness of, and control, these tick-borne pathogens.

Expression of a Recombinant Nucleocapsid Protein of Rift Valley Fever Virus in Vero Cells as an Immunofluorescence Antigen and Its Use for Serosurveillance in Traditional Cattle Herds in Zambia
Ngonda Saasa; Masahiro Kajihara; George Dautu; Akina Mori-Kajihara; Shuetsu Fukushi; Yona Sinkala; Shigeru Morikawa; Aaron Mweene; Ayato Takada; Kumiko Yoshimatsu; Jiro Arikawa
Vector-Borne and Zoonotic Diseases, 18, 5, 273, 277, Mary Ann Liebert Inc., 2018年05月01日, ［査読有り］, ［筆頭著者］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）

Co-circulation of multiple genotypes of African swine fever viruses among domestic pigs in Zambia (2013-2015).
Simulundu E; Chambaro HM; Sinkala Y; Kajihara M; Ogawa H; Mori A; Ndebe J; Dautu G; Mataa L; Lubaba CH; Simuntala C; Fandamu P; Simuunza M; Pandey GS; Samui KL; Misinzo G; Takada A; Mweene AS
Transboundary and emerging diseases, 65, 1, 114, 122, 2018年02月, ［査読有り］, ［国際共著］, ［国際誌］

First molecular detection and genetic characterization of Coxiella burnetii in Zambian dogs and rodents.
Simbarashe Chitanga; Edgar Simulundu; Martin C Simuunza; Katendi Changula; Yongjin Qiu; Masahiro Kajihara; Ryo Nakao; Michelo Syakalima; Ayato Takada; Aaron S Mweene; Samson Mukaratirwa; Bernard M Hang'ombe
Parasites & vectors, 11, 1, 40, 40, 2018年01月17日, ［査読有り］, ［国際共著］, ［国際誌］
英語, Coxiella burnetii, the causative agent of Q fever, is a zoonotic pathogen associated with sylvatic or domestic transmission cycles, with rodents being suspected to link the two transmission cycles. Infection and subsequent disease in humans has historically been associated with contact with infected livestock, especially sheep. However, recently there have been reports of Q fever outbreaks associated with contact with infected rodents and dogs. Studies exploring the potential role of these animal hosts in the epidemiology of Q fever in many developing countries in Africa are very limited. This study aimed to determine the potential role of rodents and dogs in the epidemiological cycle of C. burnetti in Zambia. Using pathogen-specific polymerase chain reaction assays targeting the 16S rRNA gene, C. burnetii was detected for the first time in 45% of rodents (9/20), in one shrew and in 10% of domestic dogs (15/150) screened in Zambia. Phylogenetic characterization of six samples based on the isocitrate synthase gene revealed that the strains were similar to a group of isolates from chronic human Q fever patients, goats and rodents reported in multiple continents. Considering the close proximity of domestic dogs and rodents to humans, especially in resource-limited communities, the presence of C. burnetii in these animals could be of significant public health importance. It is thus important to determine the burden of Q fever in humans in such resource-limited communities where there is close contact between humans, rodents and dogs.

Molecular detection and characterization of zoonotic Anaplasma species in domestic dogs in Lusaka, Zambia
Pipina A. Vlahakis; Simbarashe Chitanga; Martin C. Simuunza; Edgar Simulundu; Yongjin Qiu; Katendi Changula; Herman M. Chambaro; Masahiro Kajihara; Ryo Nakao; Ayato Takada; Aaron S. Mweene
TICKS AND TICK-BORNE DISEASES, 9, 1, 39, 43, 2018年01月, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）

Characterization of a Novel Bat Adenovirus Isolated from Straw-Colored Fruit Bat (Eidolon helvum).
Hirohito Ogawa; Masahiro Kajihara; Naganori Nao; Asako Shigeno; Daisuke Fujikura; Bernard M Hang'ombe; Aaron S Mweene; Alisheke Mutemwa; David Squarre; Masao Yamada; Hideaki Higashi; Hirofumi Sawa; Ayato Takada
Viruses, 9, 12, 2017年12月04日, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）, Bats are important reservoirs for emerging zoonotic viruses. For extensive surveys of potential pathogens in straw-colored fruit bats (Eidolon helvum) in Zambia, a total of 107 spleen samples of E. helvum in 2006 were inoculated onto Vero E6 cells. The cell culture inoculated with one of the samples (ZFB06-106) exhibited remarkable cytopathic changes. Based on the ultrastructural property in negative staining and cross-reactivity in immunofluorescence assays, the virus was suspected to be an adenovirus, and tentatively named E. helvum adenovirus 06-106 (EhAdV 06-106). Analysis of the full-length genome of 30,134 bp, determined by next-generation sequencing, showed the presence of 28 open reading frames. Phylogenetic analyses confirmed that EhAdV 06-106 represented a novel bat adenovirus species in the genus Mastadenovirus. The virus shared similar characteristics of low G + C contents with recently isolated members of species Bat mastadenoviruses E, F and G, from which EhAdV 06-106 diverged by more than 15% based on the distance matrix analysis of DNA polymerase amino acid sequences. According to the taxonomic criteria, we propose the tentative new species name "Bat mastadenovirus H". Because EhAdV 06-106 exhibited a wide in vitro cell tropism, the virus might have a potential risk as an emerging virus through cross-species transmission.

Putative endogenous filovirus VP35-like protein potentially functions as an IFN antagonist but not a polymerase cofactor
Tatsunari Kondoh; Rashid Manzoor; Naganori Nao; Junki Maruyama; Wakako Furuyama; Hiroko Miyamoto; Asako Shigeno; Makoto Kuroda; Keita Matsuno; Daisuke Fujikura; Masahiro Kajihara; Reiko Yoshida; Manabu Igarashi; Ayato Takada
PLOS ONE, 12, 10, e0186450, 2017年10月, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）

The Epidemiology of African Swine Fever in "Nonendemic" Regions of Zambia (1989-2015): Implications for Disease Prevention and Control
Edgar Simulundu; Caesar H. Lubaba; Juanita van Heerden; Masahiro Kajihara; Liywalii Mataa; Herman Moses Chambaro; Yona Sinkala; Samuel Munalula Munjita; Hetron Mweemba Munang'andu; King Shimumbo Nalubamba; Kenny Samui; Girja Shanker Pandey; Ayato Takada; Aaron S. Mweene
VIRUSES-BASEL, 9, 9, 2017年09月, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）

Genetic characterization of orf virus associated with an outbreak of severe orf in goats at a farm in Lusaka, Zambia (2015)
Edgar Simulundu; Nandi Mtine; Thoko F. Kapalamula; Masahiro Kajihara; Yongjin Qiu; James Ngoma; Victor Zulu; Geoffrey Kwenda; Chrispin Chisanga; Isaac K. Phiri; Ayato Takada; Aaron S. Mweene
ARCHIVES OF VIROLOGY, 162, 8, 2363, 2367, 2017年08月, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）

Clinical and subclinical bovine leukemia virus infection in a dairy cattle herd in Zambia
Girja S. Pandey; Edgar Simulundu; Danstan Mwiinga; Kenny L. Samui; Aaron S. Mweene; Masahiro Kajihara; Alfred Mangani; Racheal Mwenda; Joseph Ndebe; Satoru Konnai; Ayato Takada
ARCHIVES OF VIROLOGY, 162, 4, 1051, 1056, 2017年04月, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）

Putative RNA viral sequences detected in an Ixodes scapularis-derived cell line
Ryo Nakao; Keita Matsuno; Yongjin Qiu; Junki Marilyama; Nao Eguchi; Naganori Nao; Masahiro Kajihara; Kentaro Yoshii; Hirofumi Sawa; Ayato Takada; Chihiro Sugimoto
TICKS AND TICK-BORNE DISEASES, 8, 1, 103, 111, 2017年, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）

Genetic Predisposition To Acquire a Polybasic Cleavage Site for Highly Pathogenic Avian Influenza Virus Hemagglutinin
Naganori Nao; Junya Yamagishi; Hiroko Miyamoto; Manabu Igarashi; Rashid Manzoor; Aiko Ohnuma; Yoshimi Tsuda; Wakako Furuyama; Asako Shigeno; Masahiro Kajihara; Noriko Kishida; Reiko Yoshida; Ayato Takada
MBIO, 8, 1, 2017年01月, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）

Development of an Immunochromatography Assay (QuickNavi-Ebola) to Detect Multiple Species of Ebolaviruses
Reiko Yoshida; Shino Muramatsu; Hiroshi Akita; Yuji Saito; Miwa Kuwahara; Daisuke Kato; Katendi Changula; Hiroko Miyamoto; Masahiro Kajihara; Rashid Manzoor; Wakako Furuyama; Andrea Marzi; Heinz Feldmann; Aaron Mweene; Justin Masumu; Jimmy Kapeteshi; Jean-Jacques Muyembe-Tamfum; Ayato Takada
JOURNAL OF INFECTIOUS DISEASES, 214, suppl 3, S185, S191, 2016年10月, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）

Seroepidemiological Prevalence of Multiple Species of Filoviruses in Fruit Bats (Eidolon helvum) Migrating in Africa
Hirohito Ogawa; Hiroko Miyamoto; Eri Nakayama; Reiko Yoshida; Ichiro Nakamura; Hirofumi Sawa; Akihiro Ishii; Yuka Thomas; Emiko Nakagawa; Keita Matsuno; Masahiro Kajihara; Junki Maruyama; Naganori Nao; Mieko Muramatsu; Makoto Kuroda; Edgar Simulundu; Katendi Changula; Bernard Hang'ombe; Boniface Namangala; Andrew Nambota; Jackson Katampi; Manabu Igarashi; Kimihito Ito; Heinz Feldmann; Chihiro Sugimoto; Ladslav Moonga; Aaron Mweene; Ayato Takada
JOURNAL OF INFECTIOUS DISEASES, 212, S101, S108, 2015年10月, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）

A Single Amino Acid in the M1 Protein Responsible for the Different Pathogenic Potentials of H5N1 Highly Pathogenic Avian Influenza Virus Strains
Naganori Nao; Masahiro Kajihara; Rashid Manzoor; Junki Maruyama; Reiko Yoshida; Mieko Muramatsu; Hiroko Miyamoto; Manabu Igarashi; Nao Eguchi; Masahiro Sato; Tatsunari Kondoh; Masatoshi Okamatsu; Yoshihiro Sakoda; Hiroshi Kida; Ayato Takada
PLOS ONE, 10, 9, e0137989, 2015年09月, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）

Interaction between TIM-1 and NPC1 Is Important for Cellular Entry of Ebola Virus
Makoto Kuroda; Daisuke Fujikura; Asuka Nanbo; Andrea Marzi; Osamu Noyori; Masahiro Kajihara; Junki Maruyama; Keita Matsuno; Hiroko Miyamoto; Reiko Yoshida; Heinz Feldmann; Ayato Takada
JOURNAL OF VIROLOGY, 89, 12, 6481, 6493, 2015年06月, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）

Pathological and molecular diagnosis of the 2013 African swine fever outbreak in Lusaka, Zambia
John Yabe; Pharaoh Hamambulu; Edgar Simulundu; Hirohito Ogawa; Masahiro Kajihara; Akina Mori-Kajihara; Katendi Changula-Chitanga; Max Mwase; Mutinta Mweemba-Muwowo; Herman Moses Chambaro; Liywalii Mataa; Bernard Hang'ombe; Bonniface Namangala; Paul Fandamu; Hirofumi Sawa; Ayato Takada; Hideaki Higashi; Aaron Simanyengwe Mweene
TROPICAL ANIMAL HEALTH AND PRODUCTION, 47, 2, 459, 463, 2015年02月, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）

Comprehensive Molecular Detection of Tick-Borne Phleboviruses Leads to the Retrospective Identification of Taxonomically Unassigned Bunyaviruses and the Discovery of a Novel Member of the Genus Phlebovirus
Keita Matsuno; Carla Weisend; Masahiro Kajihara; Colette Matysiak; Brandi N. Williamson; Martin Simuunza; Aaron S. Mweene; Ayato Takada; Robert B. Tesh; Hideki Ebihara
JOURNAL OF VIROLOGY, 89, 1, 594, 604, 2015年01月, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）

Host Cell Factors Involved in Filovirus Infection
Masahiro Kajihara; Ayato Takada
Current Tropical Medicine Reports, 2, 1, 30, 40, 2015年, ［査読有り］, ［筆頭著者］, ［国際誌］
英語, 研究論文（学術雑誌）

A polymorphism of the TIM-1 IgV domain: Implications for the susceptibility to filovirus infection
Makoto Kuroda; Daisuke Fujikura; Osamu Noyori; Masahiro Kajihara; Junki Maruyama; Hiroko Miyamoto; Reiko Yoshida; Ayato Takada
BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS, 455, 3-4, 223, 228, 2014年12月, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）

The zoonotic potential of avian influenza viruses isolated from wild waterfowl in Zambia
Edgar Simulundu; Naganori Nao; John Yabe; Nilton A. Muto; Thami Sithebe; Hirofumi Sawa; Rashid Manzoor; Masahiro Kajihara; Mieko Muramatsu; Akihiro Ishii; Hirohito Ogawa; Aaron S. Mweene; Ayato Takada
ARCHIVES OF VIROLOGY, 159, 10, 2633, 2640, 2014年10月, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）

Ebola and Marburg virus diseases in Africa: Increased risk of outbreaks in previously unaffected areas?
Katendi Changula; Masahiro Kajihara; Aaron S. Mweene; Ayato Takada
MICROBIOLOGY AND IMMUNOLOGY, 58, 9, 483, 491, 2014年09月, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）

Heat Shock Protein 70 Modulates Influenza A Virus Polymerase Activity
Rashid Manzoor; Kazumichi Kuroda; Reiko Yoshida; Yoshimi Tsuda; Daisuke Fujikura; Hiroko Miyamoto; Masahiro Kajihara; Hiroshi Kida; Ayato Takada
JOURNAL OF BIOLOGICAL CHEMISTRY, 289, 11, 7599, 7614, 2014年03月, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）

Characterization of the Envelope Glycoprotein of a Novel Filovirus, Lloviu Virus
Junki Maruyama; Hiroko Miyamoto; Masahiro Kajihara; Hirohito Ogawa; Ken Maeda; Yoshihiro Sakoda; Reiko Yoshida; Ayato Takada
JOURNAL OF VIROLOGY, 88, 1, 99, 109, 2014年01月, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）

Comparison of Antiviral Activity between IgA and IgG Specific to Influenza Virus Hemagglutinin: Increased Potential of IgA for Heterosubtypic Immunity
Mieko Muramatsu; Reiko Yoshida; Ayaka Yokoyama; Hiroko Miyamoto; Masahiro Kajihara; Junki Maruyama; Naganori Nao; Rashid Manzoor; Ayato Takada
PLOS ONE, 9, 1, e85582, 2014年01月, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）

Differential potential for envelope glycoprotein-mediated steric shielding of host cell surface proteins among filoviruses
Osamu Noyori; Keita Matsuno; Masahiro Kajihara; Eri Nakayama; Manabu Igarashi; Makoto Kuroda; Norikazu Isoda; Reiko Yoshida; Ayato Takada
VIROLOGY, 446, 1-2, 152, 161, 2013年11月, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）

Mapping of conserved and species-specific antibody epitopes on the Ebola virus nucleoprotein
Katendi Changula; Reiko Yoshida; Osamu Noyori; Andrea Marzi; Hiroko Miyamoto; Mari Ishijima; Ayaka Yokoyama; Masahiro Kajihara; Heinz Feldmann; Aaron S. Mweene; Ayato Takada
Virus Research, 176, 1-2, 83, 90, 2013年09月, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）

Heterosubtypic Antiviral Activity of Hemagglutinin-Specific Antibodies Induced by Intranasal Immunization with Inactivated Influenza Viruses in Mice
Mieko Muramatsu; Reiko Yoshida; Hiroko Miyamoto; Daisuke Tomabechi; Masahiro Kajihara; Junki Maruyama; Takashi Kimura; Rashid Manzoor; Kimihito Ito; Ayato Takada
PLOS ONE, 8, 8, e71534, 2013年08月, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）

Novel mutations in Marburg virus glycoprotein associated with viral evasion from antibody mediated immune pressure
Masahiro Kajihara; Eri Nakayama; Andrea Marzi; Manabu Igarashi; Heinz Feldmann; Ayato Takada
JOURNAL OF GENERAL VIROLOGY, 94, Pt 4, 876, 883, 2013年04月, ［査読有り］, ［筆頭著者］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）

The PB2, PA, HA, NP, and NS genes of a highly pathogenic avian influenza virus A/whooper swan/Mongolia/3/2005 (H5N1) are responsible for pathogenicity in ducks
Masahiro Kajihara; Yoshihiro Sakoda; Kosuke Soda; Kenji Minari; Masatoshi Okamatsu; Ayato Takada; Hiroshi Kida
VIROLOGY JOURNAL, 10, 45, 45, 2013年02月, ［査読有り］, ［筆頭著者］, ［国際誌］
英語, 研究論文（学術雑誌）

Inhibition of Marburg Virus Budding by Nonneutralizing Antibodies to the Envelope Glycoprotein
Masahiro Kajihara; Andrea Marzi; Eri Nakayama; Takeshi Noda; Makoto Kuroda; Rashid Manzoor; Keita Matsuno; Heinz Feldmann; Reiko Yoshida; Yoshihiro Kawaoka; Ayato Takada
JOURNAL OF VIROLOGY, 86, 24, 13467, 13474, 2012年12月, ［査読有り］, ［筆頭著者］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）

致死的経過をとったアデノウイルス37型による新生児重症肺炎の1例
津曲俊太郎; 工藤絵理子; 佐々木大輔; 簗詰紀子; 縄手満; 吉岡幹朗; 鹿野高明; 高橋豊; 直亨則; 鈴木昭; 佐多徹太郎; 梶原将大
日本小児科学会雑誌, 116, 5, 940, 2012年05月01日, ［査読有り］, ［国内誌］
日本語

An H5N1 highly pathogenic avian influenza virus that invaded Japan through waterfowl migration
Masahiro Kajihara; Keita Matsuno; Edgar Simulundu; Mieko Muramatsu; Osamu Noyori; Rashid Manzoor; Eri Nakayama; Manabu Igarashi; Daisuke Tomabechi; Reiko Yoshida; Masatoshi Okamatsu; Yoshihiro Sakoda; Kimihito Ito; Hiroshi Kida; Ayato Takada
JAPANESE JOURNAL OF VETERINARY RESEARCH, 59, 2-3, 89, 100, 2011年08月, ［査読有り］, ［筆頭著者］, ［国内誌］
英語, 研究論文（学術雑誌）

A vaccine prepared from a non-pathogenic H5N1 influenza virus strain from the influenza virus library conferred protective immunity to chickens against the challenge with antigenically drifted highly pathogenic avian influenza virus
Rozanah Asmah Abdul Samad; Naoki Nomura; Yoshimi Tsuda; Rashid Manzoor; Masahiro Kajihara; Daisuke Tomabechi; Takashi Sasaki; Norihide Kokumai; Toshiaki Ohgitani; Masatoshi Okamatsu; Ayato Takada; Yoshihiro Sakoda; Hiroshi Kida
JAPANESE JOURNAL OF VETERINARY RESEARCH, 59, 1, 23, 29, 2011年02月, ［査読有り］, ［国際共著］, ［国内誌］
英語, 研究論文（学術雑誌）

Detection of highly pathogenic avian influenza virus infection in vaccinated chicken flocks by monitoring antibodies against non-structural protein 1 (NS1)
Natsumi Takeyama; Kenji Minari; Masahiro Kajihara; Norikazu Isoda; Ryuichi Sakamoto; Takashi Sasaki; Norihide Kokumai; Noriyasu Takikawa; Rikiya Shiraishi; Masaji Mase; Junko Hagiwara; Toshiaki Kodama; Takashi Imamura; Masashi Sakaguchi; Toshiaki Ohgitani; Akira Sawata; Masatoshi Okamatsu; Masatake Muramatsu; Kenji Tsukamoto; Zhifeng Lin; Kotaro Tuchiya; Yoshihiro Sakoda; Hiroshi Kida
VETERINARY MICROBIOLOGY, 147, 3-4, 283, 291, 2011年01月, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）

Characterization of H5N1 highly pathogenic avian influenza virus strains isolated from migratory waterfowl in Mongolia on the way back from the southern Asia to their northern territory
Yoshihiro Sakoda; Sengee Sugar; Damdinjav Batchluun; Tseren-Ochir Erdene-Ochir; Masatoshi Okamatsu; Norikazu Isoda; Kosuke Soda; Hiroki Takakuwa; Yoshimi Tsuda; Naoki Yamamoto; Noriko Kishida; Keita Matsuno; Eri Nakayama; Masahiro Kajihara; Ayaka Yokoyama; Ayato Takada; Ruuragchaa Sodnomdarjaa; Hiroshi Kida
VIROLOGY, 406, 1, 88, 94, 2010年10月, ［査読有り］, ［国際共著］, ［国際誌］
英語, 研究論文（学術雑誌）

Development of vaccine strains of H5 and H7 influenza viruses
Kosuke Soda; Yoshihiro Sakoda; Norikazu Isoda; Masahiro Kajihara; Yoshinari Haraguchi; Hitomi Shibuya; Hiromi Yoshida; Takashi Sasaki; Ryuichi Sakamoto; Kazue Saijo; Junko Hagiwara; Hiroshi Kida
JAPANESE JOURNAL OF VETERINARY RESEARCH, 55, 2-3, 93, 98, 2008年01月, ［査読有り］, ［国内誌］
英語, 研究論文（学術雑誌）

ダニが媒介するウイルス感染症 海外で注目される疾患 ―クリミヤ・コンゴ出血熱, ポワッサンウイルス感染症, その他の新興ダニ媒介性ウイルス感染症
梶原 将大, 臨床と微生物, 51, 4, 47, 52, 2024年07月, ［招待有り］, ［筆頭著者, 責任著者］
日本語, 記事・総説・解説・論説等（学術雑誌）

コウモリ由来およびトガリネズミ由来ハンタウイルスの組換えN抗原を用いたコウモリ抗体検出系の確立
KODUA Emmanuel; 梶原将大; 梶原将大; 危卓行; 危卓行; LOKPATHIRAGE Sithumini M. W.; LOKPATHIRAGE Sithumini M. W.; MUTHUSINGHE Devinda; MUTHUSINGHE Devinda; 森(梶原)亜紀奈; 邱永晋; CHANGULA Katendi; HANGOMBE Bernard M; 菊池風花; 神谷元; 新井智; 高田礼人; 吉松組子; 吉松組子, 日本ウイルス学会学術集会プログラム・予稿集(Web), 71st, 2024年

人獣共通感染症対策に資する活動
澤洋文; 澤洋文; 澤洋文; 田畑耕史郎; 板倉友香里; 梶原将大; 林田京子; 松野啓太; 直亨則; 大場靖子; 大場靖子; 佐々木道仁, 日本化学療法学会雑誌, 72, 5, 2024年

コウモリ由来オルソレオウイルスの病原性発現機構の解明
播磨勇人; 佐々木道仁; 有泉拓馬; 川口虹穂; 小林広子; KITTIYA Intaruck; 小林剛; 川岸崇裕; 梶原将大; 大場靖子; 伊藤直人; 石原加奈子; HANG’OMBE Bernard; 澤洋文, 日本獣医学会学術集会講演要旨集, 167th, 2024年

株間ゲノム比較によるネルソンベイオルソレオウイルスの新規病原性因子の同定
播磨勇人; 佐々木道仁; 有泉拓馬; 川口虹穂; 小林広子; KITTIYA Intaruck; 小林剛; 川岸崇裕; 金井祐太; 直亨則; 邱永晋; 梶原将大; 大場靖子; 伊藤直人; 石原加奈子; HANGOMBE Bernard M; 澤洋文, 日本ウイルス学会学術集会プログラム・予稿集(Web), 71st, 2024年

鹿児島県内のコキクガシラコウモリから分離されたラブドウイルス科Oita virusの性状解析
澤井祭子; 佐多翔; 江さき真南; 船越公威; 梶原将大; 村上晋; 君付和範; 八尋隆明; 西園晃; 君付和範; 八尋隆明; 西園晃; 小澤真; 奥谷公亮, 日本獣医学会学術集会講演要旨集, 167th, 2024年

ザンビアにおけるブタインフルエンザの疫学調査
播磨 勇人; 奥谷 公亮; 梶原 将大; 小川 寛人; Simulundu Edgar; Bwalya Eugene; 邱 永晋; 迫田 義博; 西藤 岳彦; Hang' ombe Bernard; 澤 洋文; Mweene Aaron; 高田 礼人, 日本獣医学会学術集会講演要旨集, 166回, 154, 154, 2023年09月
(公社)日本獣医学会, 日本語

マールブルグウイルスおよびエジプトルーセットオオコウモリの生態学的調査
梶原将大; 藤岡慧明; CHANGULA Katendi; 邱永晋; 梶原亜紀奈; 手嶋優風; NDEBE Joseph; MOONGA Ladslav; 直亨則; 直亨則; 尾針由真; 林田京子; HANG’OMBE Bernard M; 川端寛樹; 澤洋文; 澤洋文; 澤洋文; 飛龍志津子; 高田礼人; 高田礼人, 日本ウイルス学会学術集会プログラム・予稿集(Web), 70th, 2023年

ザンビアのコウモリが保有する多様なヘルペスウイルスの遺伝学的性状の解明
播磨勇人; 邱永晋; 山岸潤也; 梶原将大; CHANGULA Katendi; 佐々木道仁; MWEENE Aaron S; 澤洋文; 石原加奈子; HANG’OMBE Bernard M; 高田礼人, 日本ウイルス学会学術集会プログラム・予稿集(Web), 70th, 2023年

DR.CongoにおけるBas-Congoウイルス関連急性出血熱発生後14年間のMangala住民間の高血清有病率【JST・京大機械翻訳】|||
MUNYEKU Yannick Bazitama; MUNYEKU Yannick Bazitama; MUNYEKU Yannick Bazitama; HATTORI Takanari; SAITO Takeshi; OKITALE Patient Talunda; OKITALE Patient Talunda; MIYAMOTO Hiroko; KAJIHARA Masahiro; MASUMU Justin; MASUMU Justin; MASUMU Justin; AHUKA Steve Mundeke; AHUKA Steve Mundeke; MUYEMBE Jean-Jacques Tamfum; MUYEMBE Jean-Jacques Tamfum; MAKIALA Sheila Mandanda; MAKIALA Sheila Mandanda; IGARASHI Manabu; TAKADA Ayato, 日本ウイルス学会学術集会プログラム・予稿集(Web), 70th, 2023年

ザンビアのエジプトルーセットオオコウモリおよびマストミスから分離されたロタウイルスAの性状解析
岸本麻衣; 岸本麻衣; 梶原将大; 高田礼人; 大園誠也; 鈴木忠樹; 伊藤直人; 大場靖子; 澤洋文; 澤洋文; 佐々木道仁, 日本獣医学会学術集会講演要旨集, 166th, 2023年

ザンビアのコウモリにおけるネルソンベイオルソレオウイルスの保有調査および分離ウイルスの病原性比較解析
播磨 勇人; 佐々木 道仁; 大場 靖子; 邱 永晋; Changula Katendi; 梶原 将大; Simulundu Edgar; 谷口 怜; 高田 礼人; 西條 政幸; Hang'ombe Bernard M.; 澤 洋文, 日本獣医学会学術集会講演要旨集, 165回, [F1P, 09], 2022年09月
(公社)日本獣医学会, 日本語

ザンビアとガボンにおけるCOVID-19疫学の協力
山岸潤也; 邱永晋; 梶原将大; 林田京子; 杉達紀; 澤洋文; 阿部遥; 安田二郎, 日本ウイルス学会学術集会プログラム・予稿集(Web), 69th, 2022年

ウイルス分離先行型スクリーニングにより分離された野生動物由来ロタウイルスAの性状解析
岸本麻衣; 梶原将大; 高田礼人; 伊藤直人; 大場靖子; 澤洋文; 佐々木道仁, 日本ウイルス学会学術集会プログラム・予稿集(Web), 69th, 2022年

ザンビアにおけるクリミア・コンゴ出血熱の疫学調査
梶原将大; SIMUUNZA Martin; SAASA Ngonda; DAUTU George; 森亜紀奈; 邱永晋; 中尾亮; HANG’OMBE Bernard; HANG’OMBE Bernard; 澤洋文; 澤洋文; SIMULUNDU Edgar; 福士秀悦; 森川茂; 西條政幸; 有川二郎; 高田礼人; 高田礼人; 吉松組子, 日本ウイルス学会学術集会プログラム・予稿集(Web), 69th, 2022年

T-cell immunoglobulin mucin1の一塩基変異がウイルスの細胞侵入へ与える影響
服部貴成; 斎藤健; 高館佳弘; 笠島和; 梶原亜紀奈; 宮本洋子; 梶原将大; 五十嵐学; 高田礼人, 日本ウイルス学会学術集会プログラム・予稿集(Web), 69th, 2022年

特集 新型ウイルスの最新情報と対応 新たな問題となりうるコウモリ･節足動物媒介性ウイルス
梶原 将大; 大場 靖子; 高田 礼人, Pharma Medica, 39, 1, 43, 47, 2021年01月20日, ［招待有り］, ［筆頭著者］
メディカルレビュー社, 日本語, 記事・総説・解説・論説等（学術雑誌）

ザンビアのエジプトルーセットオオコウモリから単離したネルソンベイオルソレオウイルスの性状解析
播磨勇人; 佐々木道仁; 大場靖子; 邱永晋; チャングラカテンディ; 梶原将大; シムルンドゥエドガー; 谷口怜; 高田礼人; 西條政幸; ハンゴンべバーナード; 澤洋文, 日本ウイルス学会学術集会プログラム・予稿集(Web), 68th, 2021年

ヒトアンジオテンシン変換酵素2受容体における一塩基多型とSARS-CoV-2に対する細胞感受性【JST・京大機械翻訳】|||
HATTORI Takanari; SAITO Takeshi; TAKADATE Yoshihiro; OKUYA Kosuke; KASAJIMA Nodoka; KAJIHARA Akina; MIYAMOTO Hiroko; KAJIHARA Masahiro; TAKADA Ayato, 日本ウイルス学会学術集会プログラム・予稿集(Web), 68th, 2021年

Niemann-Pick C1のループ構造由来環状ペプチドによるフィロウイルス侵入阻害効果
高橋侑嗣; 高舘佳弘; 宮本洋子; 梶原将大; 五十嵐学; 高田礼人, 日本ウイルス学会学術集会プログラム・予稿集(Web), 68th, 2021年

ザンビアのブタにおける消化管ウイルスの疫学調査
播磨勇人; 梶原将大; 佐々木道仁; SIMULUNDU Edgar; BWALYA Eugene; 邱永晋; 奥谷公亮; 大場靖子; HANG’OMBE Bernard M; MWEENE Aaron S; 高田礼人; 澤洋文, 日本獣医学会学術集会講演要旨集, 164th (CD-ROM), 2021年

ザンビアで採集したオオコウモリから単離したオルソレオウイルスの解析
播磨勇人; 佐々木道仁; 大場靖子; QIU Yongjin; 梶原将大; 谷口怜; 高田礼人; 西條政幸; 澤洋文, 日本分子生物学会年会プログラム・要旨集(Web), 44th, 2021年

マダニ中のフレボウイルスの遺伝子系統解析に基づく性状推定
松野 啓太; 中尾 亮; 梶原 将大; 下田 宙; 海老原 秀喜; 高田 礼人; 前田 健; 岡松 正敏; 迫田 義博, 日本獣医学会学術集会講演要旨集, 162回, 400, 400, 2019年08月
(公社)日本獣医学会, 日本語

エボラウイルスの細胞侵入阻害薬の探索
磯野真央; 古山若呼; 黒田誠; 近藤達成; 五十嵐学; 梶原将大; 吉田玲子; マンズール ラシッド; 奥谷公亮; 宮本洋子; フェルドマン ハインツ; マルジ アンドレア; 堺谷政弘; 高田礼人, 日本ウイルス学会学術集会プログラム・予稿集(Web), 67th, 2019年

ザンビアのエジプトルーセットオオコウモリから検出されたマールブルグウイルスの遺伝学的解析
梶原将大; CHANGULA Katendi; HANG’OMBE Bernard; 播磨勇人; 宮本洋子; 衛藤芳樹; 奥谷公亮; 磯野真央; 吉田玲子; 邱永晋; 森(梶原)亜紀奈; 大場靖子; 澤洋文; 澤洋文; 小川寛仁; SIMULUNDU Edgar; SQUARRE David; MUKONKA Victor; MWEENE Aaron; 高田礼人; 高田礼人, 日本ウイルス学会学術集会プログラム・予稿集(Web), 67th, 2019年

ザンビアのダニで発見した新規フラビウイルスはフラビウイルス属の中で独特な系統を形成するNgoye virusに遺伝的に密接に関係している
播磨勇人; 鳥居志保; 邱永晋; 梶原将大; 衛藤芳樹; 大場靖子; 江下優樹; ハンゴンベ バーナード; 好井健太朗; シムンザ マーティン; 高田礼人; 澤洋文, 日本ウイルス学会学術集会プログラム・予稿集(Web), 67th, 2019年

臨床ウイルス分野の海外協力 アフリカを舞台にしたフィロウイルス研究―自然宿主探索と診断法開発―
梶原将大; 高田礼人; 高田礼人, 臨床とウイルス, 45, 1, 32‐40, 2017年03月31日, ［招待有り］, ［筆頭著者］, ［国内誌］
日本語

ザンビア共和国に飛来するストローオオコウモリ(Eidolon helvum)から分離されたアデノウイルスの性状解析と分子疫学的解析
小川寛人; 小川寛人; 小川寛人; 梶原将大; 直亨則; 丸山隼輝; 上野恵介; 石井秋宏; 石井秋宏; 藤倉大輔; HANG’OMBE Bernard; AARON Mweene; 山田雅夫; 東秀明; 東秀明; 澤洋文; 澤洋文; 高田礼人; 高田礼人, 日本獣医学会学術集会講演要旨集, 159th, 376, 376, 2016年08月30日, ［査読有り］, ［国際共著］, ［国内誌］
(公社)日本獣医学会, 日本語, 研究発表ペーパー・要旨（全国大会，その他学術会議）

北海道で分離された新規ダニ媒介性フレボウイルスの性状解析
鳥居志保; 鳥居志保; 松野啓太; 松野啓太; 中尾亮; 邱永晋; 梶原将大; 直亨則; 村松美笑子; 海老原秀喜; 高田礼人; 高田礼人, 日本分子生物学会年会プログラム・要旨集(Web), 39th, ROMBUNNO.1LBA‐083 (WEB ONLY), 2016年, ［国際共著］, ［国内誌］
日本語, 研究発表ペーパー・要旨（全国大会，その他学術会議）

RT‐PCR法による新規ダニ媒介性フレボウイルス遺伝子の探索
松野啓太; 松野啓太; 下田宙; 鳥居志保; 邱永晋; 中尾亮; 梶原将大; 岡松正敏; 迫田義博; 迫田義博; 奥村敦; 奥村敦; 高田礼人; 高田礼人; 前田健; 海老原秀喜, 日本獣医学会学術集会講演要旨集, 158th, 344, 344, 2015年08月30日, ［国際共著］, ［国内誌］
(公社)日本獣医学会, 日本語, 研究発表ペーパー・要旨（全国大会，その他学術会議）

新興・再興感染症と感染症対策 A.ウイルス フィロウイルス感染症
梶原将大; 高田礼人, 生体の科学, 66, 4, 296, 300, 2015年08月15日, ［招待有り］, ［筆頭著者］, ［国内誌］
日本語, 記事・総説・解説・論説等（学術雑誌）

ラボレポート-留学編- 「虎穴に入らずんば虎児を得ず！〜アフリカでのウイルス研究生活 −Samora Machel School of Veterinary Meicine, The University of Zambia」
梶原 将大, 実験医学, 33, 8, 1344, 1346, 2015年04月, ［招待有り］, ［筆頭著者］, ［国内誌］
日本語, 記事・総説・解説・論説等（学術雑誌）

フィロウイルスの細胞侵入におけるTIM‐1とNPC1相互作用の解析
黒田誠; 藤倉大輔; 南保明日香; 野依修; 梶原将大; 丸山隼輝; 宮本洋子; 吉田玲子; 高田礼人, 日本ウイルス学会学術集会プログラム・抄録集, 62nd, 174, 2014年10月31日, ［国内誌］
日本語, 研究発表ペーパー・要旨（全国大会，その他学術会議）

北海道産マダニからの新規フレボウイルスの検出
中尾亮; 梶原将大; 邱永晋; 森亜紀奈; 直亨則; 村松美笑子; 好井健太朗; 苅和宏明; 澤洋文; 杉本千尋; 高田礼人, 日本獣医学会学術集会講演要旨集, 157th, 451, 2014年08月11日, ［国内誌］
日本語, 研究発表ペーパー・要旨（全国大会，その他学術会議）

A型インフルエンザウイルスヘマグルチニン特異的IgA抗体の亜型間交差抗ウイルス活性
村松美笑子; 吉田玲子; 横山文香; 宮本洋子; 梶原将大; 丸山隼輝; 直享則; MANZOOR Rashid; 高田礼人, 日本ウイルス学会学術集会プログラム・抄録集, 62nd, 2014年

マダニ媒介性フレボウイルスの検出
中尾亮; 梶原将大; 松野啓太; 邱永晋; 森亜紀奈; 海老原秀喜; 高田礼人; 杉本千尋, 大原綜合病院年報, 53, 69, 2013年12月15日, ［国際共著］, ［国内誌］
日本語, 機関テクニカルレポート，技術報告書，プレプリント等

フィロウイルス感受性に及ぼすTIM‐1多型の影響
黒田誠; 藤倉大輔; 野依修; 中山絵里; 梶原将大; 丸山隼輝; 宮本洋子; 吉田玲子; 高田礼人, 日本ウイルス学会学術集会プログラム・抄録集, 61st, 163, 2013年10月29日, ［国内誌］
日本語, 研究発表ペーパー・要旨（全国大会，その他学術会議）

2010年に稚内で分離された2株の高病原性鳥インフルエンザウイルスの病原性比較
直亨則; 梶原将大; 丸山隼輝; 木村享史; MANZOOR Rashid; 村松美笑子; 岡松正敏; 宮本洋子; 吉田玲子; 迫田義博; 喜田宏; 高田礼人, 日本ウイルス学会学術集会プログラム・抄録集, 61st, 150, 2013年10月29日, ［国内誌］
日本語, 研究発表ペーパー・要旨（全国大会，その他学術会議）

感染症“死の病原体”に前線で挑むサイエンス フィロウイルスの生態
梶原将大; 小川寛人; 高田礼人, 実験医学, 31, 19, 2013年, ［招待有り］
日本語, 記事・総説・解説・論説等（学術雑誌）

フィロウイルス糖蛋白質による宿主細胞表面分子の立体的遮蔽現象の解析
野依修; 松野啓太; 梶原将大; 中山絵里; 五十嵐学; 磯田典和; 吉田玲子; 高田礼人, 日本ウイルス学会学術集会プログラム・抄録集, 60th, 198, 2012年10月31日, ［国内誌］
日本語, 研究発表ペーパー・要旨（全国大会，その他学術会議）

新生児に致死的肺炎を引き起こしたアデノウイルスの解析
直亨則; 津曲俊太郎; 吉岡幹郎; 梶原将大; 鈴木昭; 高橋豊; 高田礼人, 日本ウイルス学会北海道支部夏季シンポジウム講演抄録, 46th, 10, 2012年, ［国内誌］
日本語, 研究発表ペーパー・要旨（全国大会，その他学術会議）

動物インフルエンザのグローバルサーベイランスと全ての亜型ウイルスライブラリーの構築
梶原将大; 梶原将大; 迫田義博; 迫田義博; 伊藤壽啓; 高田礼人; 岸田典子; 五十嵐学; 磯田典和; 磯田典和; 曽田公輔; 曽田公輔; 喜田宏; 喜田宏; 喜田宏, 日本ウイルス学会学術集会プログラム・抄録集, 55th, 216, 2007年10月01日, ［国内誌］
日本語, 研究発表ペーパー・要旨（全国大会，その他学術会議）

動物インフルエンザのグローバルサーベイランスと全ての亜型ウイルスライブラリーの構築
梶原将大; 梶原将大; 迫田義博; 迫田義博; 伊藤壽啓; 高田礼人; 伊藤公人; 岸田典子; 五十嵐学; 磯田典和; 磯田典和; 曽田公輔; 曽田公輔; WEBSTER R.G; 喜田宏; 喜田宏; 喜田宏, 日本獣医学会学術集会講演要旨集, 144th, 91, 2007年08月27日, ［国内誌］
日本語, 研究発表ペーパー・要旨（全国大会，その他学術会議）

識別図鑑日本のコウモリ
梶原将大, 日本のコウモリ研究最前線③「コウモリが持つウイルスを追跡する」
文一総合出版, 2023年10月, 9784829972472, 239p, 日本語, ［分担執筆］

ザンビアを知るための55章
島田, 周平; 大山, 修一, コウモリが宿すウイルスを探る.
明石書店, 2020年08月, 9784750350622, 370p, 日本語, 一般書・啓蒙書, ［国際共著］, ［分担執筆］

アフリカにおける出血熱ウイルスの疫学調査
梶原将大
令和5年度 新興再興感染症制御学シンポジウム, 2023年12月08日, 日本語, 公開講演，セミナー，チュートリアル，講習，講義等
［招待講演］

ウイルスの生態を解き明かす
梶原将大
第31回先端科学移動大学2023 in 旭川, 日本語, 公開講演，セミナー，チュートリアル，講習，講義等
2023年11月17日 - 2023年11月18日, ［招待講演］

コウモリがもつウイルスの生態学
梶原将大
第61回 文科省と附置研センターとの定例ランチミーティング, 2023年10月13日, 日本語, 公開講演，セミナー，チュートリアル，講習，講義等
［招待講演］

アフリカのコウモリがもつウイルスの話
梶原将大
ラブラボ2023, 2023年08月04日, 日本語, 公開講演，セミナー，チュートリアル，講習，講義等
［招待講演］

Surveillance of Zoonoses in Zambia
Masahiro Kajihara
Africa symposium "Acceleration of the malaria elimination in tropical Africa: the post―COVID―19 pandemic challenges", 2022年12月02日, 英語, 口頭発表（招待・特別）
［招待講演］

アフリカのコウモリがもつウイルスの話
梶原将大
第31回先端科学移動大学2022 in 釧路, 日本語, 公開講演，セミナー，チュートリアル，講習，講義等
2022年11月18日 - 2022年11月19日, ［招待講演］

アフリカのコウモリが保有するフィロウイルス・ナイロウイルス
梶原将大
第165回日本獣医学会学術集会, 2022年09月06日, 日本語, シンポジウム・ワークショップパネル（指名）
2022年09月06日 - 2022年09月08日, ［招待講演］

Filovirus in Egyptian fruit bats in Zambia
梶原将大
日本哺乳類学会2021年度大会, 2021年08月29日, 日本語, シンポジウム・ワークショップパネル（指名）
2021年08月28日 - 2021年08月31日, ［招待講演］

アフリカでのウイルスハンティング −出血熱ウイルスから未知のウイルスまで−
梶原将大
第19回みちのくウイルス塾, 2020年07月23日, 日本語, 公開講演，セミナー，チュートリアル，講習，講義等
2020年07月23日 - 2020年07月23日, ［招待講演］

Surveillance of Viral Zoonoses in Africa
Mweene A; Kajihara M; Inano K; Takada A
Prince Mahidol Award Conference, 2018年01月29日, 英語, シンポジウム・ワークショップパネル（指名）
［招待講演］

保全医学演習, 2024年, 博士後期課程, 獣医学院

保全医学演習, 2024年, 博士後期課程, 国際感染症学院

音響サイバー空間を利用したコウモリの階層ナビゲーションの理解
科学研究費助成事業
2021年09月10日 - 2026年03月31日
飛龍 志津子; 小林 耕太; 福井 大; 小川 宏人; 梶原 将大; 藤岡 慧明
日本学術振興会, 学術変革領域研究(A), 同志社大学, 21H05295

アフリカのコウモリが保有する病原性未知の新規コロナウイルスに関する研究
科学研究費助成事業
2022年04月01日 - 2025年03月31日
梶原 将大
日本学術振興会, 基盤研究(C), 北海道大学, 22K05981

マダニに潜む病原体のグローバルプロファイリング
科学研究費助成事業
2019年04月01日 - 2023年03月31日
松野 啓太; 山口 宏樹; 大場 靖子; 梶原 将大
本年度は主に、昨年度までに検出あるいは分離培養に成功したマダニ由来ウイルスについての性状解析を実施した。さらに、昨年度に報告したマダニ咬傷後に発熱を呈した患者から検出されたオルソナイロウイルス（Yezoウイルス)が分布していると考え られる北海道において、このウイルスを対象とした性状解析ならびに野外調査を実施した。なお、Yezoウイルスの調査については、他研究費との連携により効率的に展開した。また、マダニからオルソナイロウイルスを検出する系を樹立し、実際に北海道のマダニから複数のオルソナイロウイルスを検出した。本年度は新型コロナウ イルス感染拡大の影響で、海外でのマダニ採集が引き続き困難になったため、研究協力者からマダニを送付してもらうなどの代替策を進め、採集地域が限定されることを防いだ。さらに、実験室内で人為的にウイルスを接種したマダニを用いた実験モデルからのウイルス検出・分離培養方法検証を進め、状況が改善した際に野外調査を精力的に進められるよう準備を行った。 マダニから分離培養された各微生物については、遺伝情報解読と病原性解析の2つの性状比較解析を実施し、マダニから検出された各微生物は、サンガーシークエンサーと次世代シークエンサーを併用して遺伝子配列を解読した。さらに、これまでに収集あるいは分離培養したウイルスについて、実験動物を用いて病原性の検討を実施した。これらの結果を用いて、微生物のプロファイリングに用いることができる基礎的データを収集し、各微生物について塩基配列およびアミノ酸配列に基づいて病原性を推定する方法の検討を開始した。
日本学術振興会, 基盤研究(B), 北海道大学, 19H03112

回帰熱を主とした節足動物媒介性感染症に関する総合的対策のための研究
新興・再興感染症研究基盤創生事業（海外拠点活用研究領域）
2020年07月 - 2023年03月
川端寛樹; 邱永晋; 中尾亮; 梶原将大; 飛龍志津子
国立研究開発法人日本医療研究開発機構（AMED), 22wm0225016h0103

アフリカのコウモリから分離された新規パラミクソウイルスの研究
科学研究費助成事業
2018年04月01日 - 2021年03月31日
梶原 将大
本事業ではザンビアの食果コウモリから分離したバットムンプスウイルス (bMuV) の研究を行なった。全ゲノム解析の結果、bMuVはムンプスウイルスと塩基配列で77%の相同性を有していた。また、21%のコウモリからbMuVゲノムが検出された。脾臓、肺、腎臓、結腸、唾液腺から高頻度にウイルス遺伝子が検出されたため、唾液、尿、便等を介するウイルス伝播が疑われた。bMuVをマウス、ラット、ハムスターに実験的に接種した。いずれの個体も明確な臨床症状は示さなかったが、bMuV特異抗体の産生および臓器でのウイルス増殖が確認されたことから、bMuVはコウモリ以外の哺乳動物に対して感染性を有することがわかった。
日本学術振興会, 若手研究, 北海道大学, 18K15163

新規ダニ媒介性フレボウイルスの探索とSFTSウイルスの病原性獲得メカニズム
科学研究費助成事業
2016年04月01日 - 2019年03月31日
松野 啓太; 梶原 将大; 海老原 秀喜
重症熱性血小板減少症(SFTS)ウイルスは、ダニ媒介性フレボウイルスの中でもヒトに対する病原性が高く、日本を含む東アジアで問題となっている。本研究では、SFTSウイルスに近縁なダニ媒介性フレボウイルスを比較解析することで、SFTSウイルスがヒトに対する病原性を獲得するに至った進化の過程を解き明かすことを目的とした。日本国内外の様々な地域でマダニ中から複数の新規フレボウイルスを発見し、野生動物がこれらのウイルスに感染していることを明らかにした。また、ダニ媒介性フレボウイルスはマダニと共進化しており、病原性獲得にNSsと呼ばれるタンパク質遺伝子を獲得することが重要である可能性を明らかにした。
日本学術振興会, 若手研究(B), 北海道大学, 16K18791

ザンビアにおける潜在的病原ウイルスに関する調査研究
科学研究費助成事業
2015年04月01日 - 2019年03月31日
梶原 将大
本研究はザンビアにおける疫学調査により既知・未知のウイルスを探索し、その生態を理解すると共に、公衆衛生的なリスク評価を目的に実施した。研究期間にザンビアにおいてマダニ2,327匹およびコウモリ919頭を捕獲し解析に供した。ヒトに対する病原性を有するウイルスとして、イボマダニからはクリミア・コンゴ出血熱ウイルス (CCHFV) が、エジプトルーセットオオコウモリからはマールブルグウイルスの遺伝子が検出された。また、マダニからは遺伝的に異なる8つのダニ媒介性フレボウイルスが検出され、一部のウイルスは新規ウイルスであると思われる。また、ケンショウコウモリからはコウモリムンプスウイルスが分離された。
日本学術振興会, 若手研究(B), 北海道大学, 15K18778

抗体によるウイルスの出芽阻害メカニズムに関する研究
科学研究費助成事業
2013年08月30日 - 2015年03月31日
梶原 将大
過去の研究を通してウイルスに対する出芽阻害抗体を得たが、阻害機序や防御免疫への寄与は不明であった。本研究を通して、鼻腔内免疫によりインフルエンザウイルスに対する放出阻害抗体が誘導されることがわかった。また、誘導されたIgA抗体を介したウイルス粒子の架橋により、新生ウイルスが細胞上に繋留される事が示唆された。このウイルス放出阻害が鼻腔内免疫による亜型間交差防御の機序である可能性が示唆された。また、出芽阻害抗体の有する防御効果をマールブルグ熱の齧歯類動物モデルを用いて検討した。その結果、出芽阻害抗体投与群において予後の改善が見られた。よって、出芽阻害抗体が生体の感染防御に寄与するものと考えられる。
日本学術振興会, 研究活動スタート支援, 北海道大学, 25892002

ウイルスの病原性と感染制御に関する研究
科学研究費助成事業
2010年 - 2012年
梶原 将大
フィロウイルス(Marburg virus(MARV)およびEbola virus(EBOV))はヒトを含む霊長類に致死的な出血熱を引き起こす。フィロウイルスはその粒子表面に一種類のみ糖蛋白質(GP)を有する。よって、GPはウイルスの粒子形成、感染性、増殖性、組織指向性および宿主特異性、さらには病原性に関与する重要な因子である。それと同時に、中和抗体の唯一の標的であると考えられる。これまでMARVの感染性を効果的に抑制する抗体の報告は無く、抗体によるMARVに対する感染防御メカニズムはほとんど不明であった。昨年度までの研究結果から、MARVのGP特異的モノクローナル抗体であるAGP127-8およびMGP72-17がMARVの感染性を抑制することが明らかとなっていた。そこで、今年度はこれら抗体によるMARVの感染抑制メカニズムの解明を目的として以下の実験を行った。
最初に、MARVのGPをまとったシュードタイプ豚水疱性口炎ウイルスを用いてAGPI27-8およびMGP72-17抗体の中和活性を評価した。これら抗体と反応させたウイルスを細胞に接種したところ、感染価の減少は確認されなかったことから、AGP127-8およびMGP72-17抗体は従来の中和活性、すなわちウイルスの細胞侵入を阻害しないことがわかった。一方で、これら抗体のウイルス様粒子(VLP)形成への影響を評価したところ、抗体存在下においてVLPの放出が顕著に抑制されることがわかった。さらに、本物のMARVを用いて同様に試験したところ、AGP127-8およびMGP72-17抗体はMARVの細胞への感染を抑制しないものの、MARVの細胞からの出芽を阻害することがわかった。電子顕微鏡を用いて、出芽阻害抗体存在下におけるVLPおよびVLP産生細胞を形態学的に解析したところ、抗体非存在下ではVLPが細胞から効率よく出芽する一方で、抗体存在下では重度に絡まったVLPが細胞表面に堆積する様子が観察された。また、AGP127-8抗体の抗原結合部位(Fab)のみでVLPの形成は阻害されなかったため、AGPl27-8およびMGP72-17抗体はその分子中に複数の抗原結合部位を有することにより、GP分子間に複雑な架橋を形成し、VLPの凝集を引き起こしたと考えられる。これら抗体はGP1のfurin開裂部位近傍にエピトープを有することから、furin開裂により露出するエピトープを含む領域がMARVの出芽に対して補助的な働きを有する可能性が示唆された。
本研究は、従来の中和試験では評価されない抗体もMARV感染における防御免疫に寄与する可能性を示すものである。また、AGPI27-8およびMGP72-17抗体はマールブルグ出血熱に対する抗体治療に応用できる可能性が考えられる。本研究の成績は抗体を用いたフィロウイルスの予防・治療法の開発に対して新たな知見を与えるものである。
日本学術振興会, 特別研究員奨励費, 北海道大学, 10J05529