Publications by authors named "Ken Maeda"

146 Publications

Histopathological Characterization of Cases of Spontaneous Fatal Feline Severe Fever with Thrombocytopenia Syndrome, Japan.

Emerg Infect Dis 2021 Apr;27(4):1068-1076

Severe fever with thrombocytopenia syndrome (SFTS) is an emerging tickborne infectious disease caused by SFTS virus (SFTSV). We report 7 cases of spontaneous fatal SFTS in felines. Necropsies revealed characteristic lesions, including necrotizing lymphadenitis in 5 cases and necrotizing splenitis and SFTSV-positive blastic lymphocytes in all cases. We detected hemorrhagic lesions in the gastrointestinal tract in 6 cases and lungs in 3 cases, suggesting a more severe clinical course of SFTS in felids than in humans. We noted necrotic or ulcerative foci in the gastrointestinal tract in 3 cases, the lung in 2 cases, and the liver in 4 cases. We clarified that blastic lymphocytes are predominant targets of SFTSV and involved in induction of necrotic foci. We also found that thymic epithelial cells were additional targets of SFTSV. These results provide insights for diagnosing feline SFTS during pathological examination and demonstrate the similarity of feline and human SFTS cases.
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http://dx.doi.org/10.3201/eid2704.204148DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8007303PMC
April 2021

Detection and molecular characterization of Babesia sp. in wild boar (Sus scrofa) from western Japan.

Ticks Tick Borne Dis 2021 Feb 27;12(4):101695. Epub 2021 Feb 27.

Transboundary Animal Diseases Research Center, Joint Faculty of Veterinary Medicine, Kagoshima University, 1-21-24 Korimoto, Kagoshima, 890-0065, Japan; Joint Graduate School of Veterinary Medicine, Kagoshima University, 1-21-24 Korimoto, Kagoshima, 890-0065, Japan; Laboratory of Zoonotic Diseases, Faculty of Applied Biosciences, Gifu University, 1-1 Yanagido, Gifu, 501-1193, Japan. Electronic address:

Wild animals often act as reservoirs of tick-borne Babesia and Theileria spp., which cause piroplasmosis. Therefore, epidemiological investigations about the distribution of these parasites in wild animals are important for evaluating the transmission risk to humans and livestock. In this study, we surveyed Babesia and Theileria spp. infecting wild boar (Sus scrofa) in Kagoshima and Yamaguchi prefectures and Tsushima island, which are all in western Japan, and performed molecular genetic analyses on the samples. DNA was extracted from either blood or liver samples of wild boar captured in Kagoshima prefecture in 2015, 2016, and 2018 and from blood samples from wild boar captured in Yamaguchi prefecture in 2013-2015 and Tsushima island in 2018. PCR screening for the partial 18S ribosomal RNA gene (18S rRNA) of both Babesia and Theileria spp. in wild boar revealed that 63.9 % (140 of 219 samples) were positive. Sequencing of all positive samples revealed that they were all the same Babesia species. Subsequent phylogenetic analyses showed that the parasite is closely related to Babesia sp. previously detected in the hard tick, Amblyomma testudinarium in Kagoshima, and further analyses suggested that this species is genetically related to Babesia gibsoni. On the other hand, no Theileria were detected in any of the samples. In summary, we observed a high prevalence of B. gibsoni-like Babesia sp. in wild boar in western regions of Japan. The host range, distribution, pathogenicity, and life cycle of this protozoan should be further evaluated.
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http://dx.doi.org/10.1016/j.ttbdis.2021.101695DOI Listing
February 2021

Assessment of SARS-CoV-2 infectivity of upper respiratory specimens from COVID-19 patients by virus isolation using VeroE6/TMPRSS2 cells.

BMJ Open Respir Res 2021 02;8(1)

Department of Veterinary Science, NIID, Shinjuku-ku, Tokyo, Japan.

Background: An outbreak of novel coronavirus (SARS-CoV-2)-associated respiratory infectious diseases (COVID-19) emerged in 2019 and has spread rapidly in humans around the world. The demonstration of in vitro infectiousness of respiratory specimens is an informative surrogate for SARS-CoV-2 transmission from patients with COVID-19; accordingly, viral isolation assays in cell culture are an important aspect of laboratory diagnostics for COVID-19.

Methods: We developed a simple and rapid protocol for isolating SARS-CoV-2 from respiratory specimens using VeroE6/TMPRSS2 cells, a cell line that is highly susceptible to the virus. We also investigated a correlation between isolation of SARS-CoV-2 and viral load detected by real-time RT-PCR (rRT-PCR) using N2 primer/probe set that has been developed for testing of COVID-19 in Japan.

Results: The SARS-CoV-2 isolation protocol did not require blind passage of inoculated cells and yielded the results of viral isolation within 7 days after inoculation. Specimens with cycle threshold (Ct) values of <20.2, determined by rRT-PCR, were predicted to be isolation-positive. On the other hand, 6.9% of specimens with Ct values >35 were virus isolation-positive, indicating that low viral loads (high Ct values) in upper respiratory specimens do not always indicate no risk of containing transmissible virus.

Conclusion: In combination with rRT-PCR, the SARS-CoV-2 isolation protocol provides a means for assessing the potential risk of transmissible virus in upper respiratory specimens.
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http://dx.doi.org/10.1136/bmjresp-2020-000830DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7907832PMC
February 2021

A serological survey and characterization of Getah virus in domestic pigs in Thailand, 2017-2018.

Transbound Emerg Dis 2021 Feb 22. Epub 2021 Feb 22.

Faculty of Veterinary Technology, Kasetsart University, Bangkok, Thailand.

Getah virus (GETV) is a mosquito-borne RNA virus belonging to the family Togaviridae, genus Alphavirus. GETV infection causes diarrhoea and death in piglets, and reproductive failure and abortion in sows. This study conducted a serological survey of GETV infection among domestic pig populations in Thailand. ELISA was used to analyse 1,188 pig serum samples collected from 11 provinces of Thailand during 2017-2018, with 23.1% of the samples being positive for anti-GETV antibodies. The positive ratio of anti-GETV antibodies was significantly higher in nursery (67.9%) and older stages (84.5%) of pigs than in finishing stage (14.2%). Furthermore, we successfully isolated GETV from one pig serum, designated as GETV strain GETV/SW/Thailand/2017, and determined the complete genome sequence (11,689 nt). Phylogenetic analysis demonstrated that our isolate was different from the recent GETV group spreading among pig populations in East Asia and formed a cluster with two GETV strains, namely YN12031 (China, 2015) and LEIV16275Mar (Far-East Russia, 2007). We concluded that two different GETV groups are currently spreading among pig populations in Asian countries.
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http://dx.doi.org/10.1111/tbed.14042DOI Listing
February 2021

Pathological Characteristics of a Patient with Severe Fever with Thrombocytopenia Syndrome (SFTS) Infected with SFTS Virus through a Sick Cat's Bite.

Viruses 2021 01 29;13(2). Epub 2021 Jan 29.

Department of Virology 1, National Institute of Infectious Diseases, 4-7-1 Gakuen, Musashimurayama City, Tokyo 208-0011, Japan.

A woman in her fifties showed symptoms of fever, loss of appetite, vomiting, and general fatigue 2 days after she was bitten by a sick cat, which had later died, in Yamaguchi prefecture, western Japan, in June 2016. She subsequently died of multiorgan failure, and an autopsy was performed to determine the cause of death. However, the etiological pathogens were not quickly identified. The pathological features of the patient were retrospectively re-examined, and the pathology of the regional lymph node at the site of the cat bite was found to show necrotizing lymphadenitis with hemophagocytosis. The pathological features were noted to be similar to those of patients reported to have severe fever with thrombocytopenia syndrome (SFTS). Therefore, the lymph node section was retrospectively tested immunohistochemically, revealing the presence of the SFTS virus (SFTSV) antigen. The sick cat showed similar symptoms and laboratory findings similar to those shown in human SFTS cases. The patient had no history of tick bites, and did not have skin lesions suggestive of these. She had not undertaken any outdoor activities. It is highly possible that the patient was infected with SFTSV through the sick cat's bite. If a patient gets sick in an SFTS-endemic region after being bitten by a cat, SFTS should be considered in the differential diagnosis.
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http://dx.doi.org/10.3390/v13020204DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7912689PMC
January 2021

Seroprevalence of Severe Fever with Thrombocytopenia Syndrome Virus in Small-Animal Veterinarians and Nurses in the Japanese Prefecture with the Highest Case Load.

Viruses 2021 02 2;13(2). Epub 2021 Feb 2.

Center for Animal Disease Control, University of Miyazaki, Miyazaki 889-2192, Japan.

Severe fever with thrombocytopenia syndrome virus (SFTSV) is the causative agent of SFTS, an emerging tick-borne disease in East Asia, and is maintained in enzootic cycles involving ticks and a range of wild animal hosts. Direct transmission of SFTSV from cats and dogs to humans has been identified in Japan, suggesting that veterinarians and veterinary nurses involved in small-animal practice are at occupational risk of SFTSV infection. To characterize this risk, we performed a sero-epidemiological survey in small-animal-practice workers and healthy blood donors in Miyazaki prefecture, which is the prefecture with the highest per capita number of recorded cases of SFTS in Japan. Three small-animal-practice workers were identified as seropositive by ELISA, but one had a negative neutralization-test result and so was finally determined to be seronegative, giving a seropositive rate of 2.2% (2 of 90), which was significantly higher than that in healthy blood donors (0%, 0 of 1000; < 0.05). The seroprevalence identified here in small-animal-practice workers was slightly higher than that previously reported in other high-risk workers engaged in agriculture and forestry in Japan. Thus, enhancement of small-animal-practice workers' awareness of biosafety at animal hospitals is necessary for control of SFTSV.
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http://dx.doi.org/10.3390/v13020229DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7912989PMC
February 2021

Diagnostic system for the detection of severe fever with thrombocytopenia syndrome virus RNA from suspected infected animals.

PLoS One 2021 28;16(1):e0238671. Epub 2021 Jan 28.

Department of Veterinary Science, National Institute of Infectious Diseases, Shinjuku, Tokyo, Japan.

Background: Severe fever with thrombocytopenia syndrome virus (SFTSV) causes severe hemorrhagic fever in humans and cats. Clinical symptoms of SFTS-infected cats resemble those of SFTS patients, whereas SFTS-contracted cats have high levels of viral RNA loads in the serum and body fluids. Due to the risk of direct infection from SFTS-infected cats to human, it is important to diagnose SFTS-suspected animals. In this study, a reverse transcription polymerase chain reaction (RT-PCR) was newly developed to diagnose SFTS-suspected animals without non-specific reactions.

Methodology/principle Findings: Four primer sets were newly designed from consensus sequences constructed from 108 strains of SFTSV. A RT-PCR with these four primer sets successfully and specifically detected four clades of SFTSV. Their limits of detection are 1-10 copies/reaction. Using this RT-PCR, 5 cat cases among 56 SFTS-suspected animal cases were diagnosed as SFTS. From these cats, IgM or IgG against SFTSV were detected by enzyme-linked immunosorbent assay (ELISA), but not neutralizing antibodies by plaque reduction neutralization titer (PRNT) test. This phenomenon is similar to those of fatal SFTS patients.

Conclusion/significance: This newly developed RT-PCR could detect SFTSV RNA of several clades and from SFTS-suspected animals. In addition to ELISA and PRNT test, the useful laboratory diagnosis systems of SFTS-suspected animals has been made in this study.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0238671PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7842937PMC
April 2021

New canine parvovirus 2a infection in an imported Asian small-clawed otter (Aonyx cinereus) in Japan.

J Vet Med Sci 2021 Apr 21;83(3):507-511. Epub 2021 Jan 21.

Laboratory of Veterinary Pathology, Faculty of Veterinary Medicine, Okayama University of Science, 1-3 Ikoino-oka, Imabari-shi, Ehime 794-8555, Japan.

Post-import from the Republic of Indonesia to Japan in 2017, two juvenile, captive bred Asian small-clawed otters (Aonyx cinereus) exhibited gastrointestinal symptoms, including vomiting, diarrhea, and hematemesis, and died. One of them was examined postmortem. Microscopically, the small intestinal mucosa was necrotic with crypts lined by regenerating large epithelial cells. A gastric cardiac mucosal ulcerative lesion containing fungal yeasts and pseudohyphae morphologically indicated Candida spp. The lymph nodes exhibited marked lymphoid depletion. Canine parvovirus 2 (CPV-2) was isolated from an oral swab, and virus protein 2 (VP2) gene sequencing revealed new CPV-2a. To our knowledge, this is the first new CPV-2a infection report in Asian small-clawed otters. This infection should be considered in gastrointestinal symptom-related cases in this species.
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http://dx.doi.org/10.1292/jvms.20-0480DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8025432PMC
April 2021

Dispersal history of Miniopterus fuliginosus bats and their associated viruses in east Asia.

PLoS One 2021 14;16(1):e0244006. Epub 2021 Jan 14.

Laboratory of Animal Morphology, Graduate School of Bioagricultural Sciences, Nagoya University, Nagoya, Japan.

In this study, we examined the role of the eastern bent-winged bat (Miniopterus fuliginosus) in the dispersion of bat adenovirus and bat alphacoronavirus in east Asia, considering their gene flows and divergence times (based on deep-sequencing data), using bat fecal guano samples. Bats in China moved to Jeju Island and/or Taiwan in the last 20,000 years via the Korean Peninsula and/or Japan. The phylogenies of host mitochondrial D-loop DNA was not significantly congruent with those of bat adenovirus (m2XY = 0.07, p = 0.08), and bat alphacoronavirus (m2XY = 0.48, p = 0.20). We estimate that the first divergence time of bats carrying bat adenovirus in five caves studied (designated as K1, K2, JJ, N2, and F3) occurred approximately 3.17 million years ago. In contrast, the first divergence time of bat adenovirus among bats in the 5 caves was estimated to be approximately 224.32 years ago. The first divergence time of bats in caves CH, JJ, WY, N2, F1, F2, and F3 harboring bat alphacoronavirus was estimated to be 1.59 million years ago. The first divergence time of bat alphacoronavirus among the 7 caves was estimated to be approximately 2,596.92 years ago. The origin of bat adenovirus remains unclear, whereas our findings suggest that bat alphacoronavirus originated in Japan. Surprisingly, bat adenovirus and bat alphacoronavirus appeared to diverge substantially over the last 100 years, even though our gene-flow data indicate that the eastern bent-winged bat serves as an important natural reservoir of both viruses.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0244006PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7808576PMC
April 2021

Virulence of Subspecies Biovar and Phenotypic Change during Serial Passages on Artificial Media.

Microorganisms 2020 Nov 27;8(12). Epub 2020 Nov 27.

Department of Veterinary Science, National Institute of Infectious Diseases, Toyama 1-23-1, Shinjuku, Tokyo 162-8640, Japan.

() is the etiological agent of the zoonotic disease tularemia. subspecies biovar has rarely been isolated in Japan and is considered to have moderate virulence, although the biological properties of fresh isolates have not been analyzed in detail. Here, we analyzed the virulence of two strains of subspecies biovar (NVF1 and KU-1) and their phenotypic stability during serial passages in Eugon chocolate agar (ECA) and Chamberlain's chemically defined medium (CDM) based agar (CDMA). C57BL/6 mice intradermally inoculated with 10 colony-forming units of NVF1 or KU-1 died within 9 days, with a median time to death of 7.5 and 7 days, respectively. Both NVF1 and KU-1 strains passaged on ECA 10 times had comparable virulence prior to passaging, whereas strains passaged on ECA 20 times and on CDMA 50 times were attenuated. Attenuated strains had decreased viability in 0.01% HO and lower intracellular growth rates, suggesting both properties are important for virulence. Additionally, passage on ECA of the KU-1 strains altered lipopolysaccharide antigenicity and bacterial susceptibility to β-lactam antibiotics. Our data demonstrate strain virulence in Japan and contribute to understanding phenotypic differences between natural and laboratory environments.
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http://dx.doi.org/10.3390/microorganisms8121881DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7760542PMC
November 2020

Complete Genome Sequences of Two Strains of Francisella tularensis subsp. bv. japonica.

Microbiol Resour Announc 2020 Nov 5;9(45). Epub 2020 Nov 5.

Department of Veterinary Science, National Institute of Infectious Diseases, Tokyo, Japan.

, a highly infectious bacterium, is the etiological agent of the zoonotic disease tularemia. It is widely distributed in the Northern Hemisphere, including Japan. Here, we have determined the complete genome sequences of two strains of subsp. bv. japonica isolated from hares in 2008 and 2009.
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http://dx.doi.org/10.1128/MRA.01127-20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7645672PMC
November 2020

Detection and phylogenetic analysis of Bartonella species from bat flies on eastern bent-wing bats (Miniopterus fuliginosus) in Japan.

Comp Immunol Microbiol Infect Dis 2020 Dec 25;73:101570. Epub 2020 Oct 25.

Laboratory of Veterinary Public Health, Department of Veterinary Medicine, College of Bioresource Sciences, Nihon University, 1866 Kameino, Fujisawa, Kanagawa, 252-0880, Japan. Electronic address:

We examined Bartonella prevalence in 281 bat flies collected from 114 eastern bent-wing bats (Miniopterus fuliginosus) in Japan and phylogenetically analyzed with other bat fly and bat strains. The bat flies were identified as Penicilidia jenynsii (PJ; n = 45), Nycteribia allotopa (NA; n = 157), and novel Nycteribia species (NS; n = 79). Bartonella DNAs were detected in 31.7 % (89/281) of bat flies by PCR targeting the citrate synthase (gltA) gene. The prevalence of Bartonella DNA among the bat flies was 47.1 % (74/157) in NA, 15.2 % (12/79) in NS, and 6.7 % (3/45) in PJ. Bartonella bacteria were also isolated from two NA and one NS. A phylogenetic analysis of the gltA sequences revealed that bat fly-associated strains were classified into three lineages and the same lineages of Bartonella were commonly detected from both Nycteribia bat flies and Miniopterus bats. These results suggest that Nycteribia bat flies are potential vectors for transmitting Bartonella among Miniopterus bats.
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http://dx.doi.org/10.1016/j.cimid.2020.101570DOI Listing
December 2020

Surveillance of Shiga toxin-producing Escherichia coli and Campylobacter spp. in wild Japanese deer (Cervus nippon) and boar (Sus scrofa).

J Vet Med Sci 2020 Sep 13;82(9):1287-1294. Epub 2020 Jul 13.

Laboratory of Veterinary Public Health, Department of Veterinary Medicine, Faculty of Agriculture, Kagoshima University, 1-21-24 Korimoto, Kagoshima, Kagoshima 890-0065, Japan.

Increasing game meat consumption in Japan requires the dissemination of safety information regarding the presence of human pathogens in game animals. Health information regarding the suitability of these animals as a meat source is not widely available. In this study, we aimed to evaluate the safety of game meat and detect potential human pathogens in wild deer (Cervus nippon) and boar (Sus scrofa) in Japan. Fecal samples from 305 wild deer and 248 boars of Yamaguchi, Kagoshima, and Tochigi prefectures collected monthly for 2 years were examined for the prevalence of Shiga toxin-producing Escherichia coli (STEC) and Campylobacter spp. STEC was isolated from 51 deer consistently throughout the year and from three boars; O-antigen genotype O146, the expression of stx2b, and eaeA absence (n=33) were the major characteristics of our STEC isolates. Other serotypes included the medically important O157, stx2b or stx2c, and eaeA-positive (n=4) and O26, stx1a, and eaeA-positive strains (n=1). Campylobacter spp. were isolated from 17 deer and 31 boars. Campylobacter hyointestinalis was the most common species isolated from 17 deer and 25 boars, whereas Campylobacter lanienae and Campylobacter coli were isolated from three and two boars, respectively. Seasonal trends for the isolation of these bacteria were not significant. This study demonstrates that wild game animals carry human pathogens; therefore, detailed knowledge of the safe handling of game meat is needed to prevent foodborne infections.
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http://dx.doi.org/10.1292/jvms.19-0265DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7538328PMC
September 2020

Molecular detection of tick-borne protozoan parasites in sika deer (Cervus nippon) from western regions of Japan.

Parasitol Int 2020 Dec 19;79:102161. Epub 2020 Jun 19.

Laboratory of Veterinary Parasitic Diseases, Department of Veterinary Sciences, Faculty of Agriculture, University of Miyazaki, 1-1 Gakuen-Kibanadai-Nishi, Miyazaki 889-2192, Japan; Center for Animal Disease Control, University of Miyazaki, 1-1 Gakuen-Kibanadai-Nishi, Miyazaki 889-2192, Japan.

The sika deer (Cervus nippon) is one of the most common species of wildlife in Japan. This study aimed to reveal the prevalence of tick-borne protozoan parasites in wild sika deer living in western Japan. We used nested polymerase chain reaction (PCR) to detect the 18S rRNA gene of tick-borne apicomplexan parasites (Babesia, Theileria, and Hepatozoon spp.) from 276 blood and liver samples from sika deer captured in the Yamaguchi, Oita, Kagoshima, Okayama, Ehime, Kochi, and Tokushima Prefectures. In total, 259 samples (259/276; 93.8%) tested positive in the nested PCR screening. Gene sequencing revealed that 99.6% (258/259) of positive samples contained Theileria sp. (sika 1), while Theileria sp. (sika 2), another Theileria species, was detected in only 3 samples. We also found that one sample from a sika deer captured in Kagoshima contained the gene of an unidentified Babesia sp. related to Babesia sp. Kh-Hj42, which was previously collected from tick in western Siberia. In conclusion, we found a high prevalence of piroplasms in sika deer from western Japan, and DNA analysis revealed that Theileria sp. (sika 1) had the highest infection rate.
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http://dx.doi.org/10.1016/j.parint.2020.102161DOI Listing
December 2020

Molecular evidence for vaccine-induced canine distemper virus and canine adenovirus 2 coinfection in a fennec fox.

J Vet Diagn Invest 2020 Jul 19;32(4):598-603. Epub 2020 Jun 19.

Den-en-chofu Animal Hospital, Ota-ku, Tokyo, Japan (Tamukai).

A 61-d-old fennec fox (), 11 d after receiving a multivalent, modified-live virus vaccine containing canine distemper virus (CDV), canine adenovirus 2 (CAdV-2), parainfluenza virus, parvovirus, and canine coronavirus, developed oculonasal discharge, and subsequently convulsions, and hemoptysis, and died. Microscopic changes in the cerebrum were evident, including neuronal degeneration and necrosis; intracytoplasmic eosinophilic inclusion bodies were observed in astrocytes. CDV was detected in the brain tissue by immunohistochemistry. Pulmonary lesions of multifocal necrotizing bronchopneumonia had Cowdry type A intranuclear inclusions in the bronchial epithelial cells. Electron microscopy revealed crystalline arrays of adenovirus-like particles within the intranuclear inclusions. Additionally, the hemagglutinin gene of CDV and the CAdV-2 DNA polymerase gene were detected in the fennec fox; sequence analysis showed 100% identity with those of the vaccine strain viruses. To our knowledge, vaccine-induced CDV and CAdV-2 coinfections using molecular analysis have not been reported previously. Therefore, vaccine strains should be considered prior to CDV vaccination in nondomestic carnivores.
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http://dx.doi.org/10.1177/1040638720934809DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7438639PMC
July 2020

16S rRNA Gene Amplicon Sequence Data from Feces of Five Species of Wild Animals in Japan.

Microbiol Resour Announc 2020 May 28;9(22). Epub 2020 May 28.

Research Center for Food Safety, Graduate School of Agricultural and Life Sciences, The University of Tokyo, Tokyo, Japan

We report 16S rRNA amplicon sequence data from feces from 58 wild boars, 60 feral raccoons, 9 wild Japanese badgers, 21 wild masked palm civets, and 8 wild raccoon dogs in Japan. The predominant bacterial taxa in the fecal microbiota were similar in part but varied among the animal species.
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http://dx.doi.org/10.1128/MRA.00368-20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7256260PMC
May 2020

16S rRNA Gene Amplicon Sequence Data from Feces of Wild Deer (Cervus nippon) in Japan.

Microbiol Resour Announc 2020 May 28;9(22). Epub 2020 May 28.

Research Center for Food Safety, Graduate School of Agricultural and Life Sciences, The University of Tokyo, Tokyo, Japan

We report 16S rRNA amplicon sequence data from feces of 109 wild deer in Japan. The dominant bacterial taxa in fecal microbiota of wild deer hunted between village and mountainous areas and those living on Miyajima Island and in Nara Park were similar but differed in abundance.
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http://dx.doi.org/10.1128/MRA.00346-20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7256258PMC
May 2020

Mosquito-borne viruses, insect-specific flaviviruses (family Flaviviridae, genus Flavivirus), Banna virus (family Reoviridae, genus Seadornavirus), Bogor virus (unassigned member of family Permutotetraviridae), and alphamesoniviruses 2 and 3 (family Mesoniviridae, genus Alphamesonivirus) isolated from Indonesian mosquitoes.

J Vet Med Sci 2020 Jul 25;82(7):1030-1041. Epub 2020 May 25.

Laboratory of Veterinary Microbiology, Joint Faculty of Veterinary Medicine, Yamaguchi University, 1677-1 Yoshida, Yamaguchi 753-8515, Japan.

Mosquitoes transmit many kinds of arboviruses (arthropod-borne viruses), and numerous arboviral diseases have become serious problems in Indonesia. In this study, we conducted surveillance of mosquito-borne viruses at several sites in Indonesia during 2016-2018 for risk assessment of arbovirus infection and analysis of virus biodiversity in mosquito populations. We collected 10,015 mosquitoes comprising at least 11 species from 4 genera. Major collected mosquito species were Culex quinquefasciatus, Aedes albopictus, Culex tritaeniorhynchus, Aedes aegypti, and Armigeres subalbatus. The collected mosquitoes were divided into 285 pools and used for virus isolation using two mammalian cell lines, Vero and BHK-21, and one mosquito cell line, C6/36. Seventy-two pools showed clear cytopathic effects only in C6/36 cells. Using RT-PCR and next-generation sequencing approaches, these isolates were identified as insect flaviviruses (family Flaviviridae, genus Flavivirus), Banna virus (family Reoviridae, genus Seadornavirus), new permutotetravirus (designed as Bogor virus) (family Permutotetraviridae, genus Alphapermutotetravirus), and alphamesoniviruses 2 and 3 (family Mesoniviridae, genus Alphamesonivirus). We believed that this large surveillance of mosquitoes and mosquito-borne viruses provides basic information for the prevention and control of emerging and re-emerging arboviral diseases.
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http://dx.doi.org/10.1292/jvms.20-0261DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7399325PMC
July 2020

Role of pattern recognition receptors and interferon-beta in protecting bat cell lines from encephalomyocarditis virus and Japanese encephalitis virus infection.

Biochem Biophys Res Commun 2020 06 23;527(1):1-7. Epub 2020 Apr 23.

Laboratory of Animal Morphology, Graduate School of Bioagricultural Sciences, Nagoya University, Nagoya, Japan. Electronic address:

Bats are potential natural hosts of Encephalomyocarditis virus (EMCV) and Japanese encephalitis virus (JEV). Bats appear to have some unique features in their innate immune system that inhibit viral replication causing limited clinical symptoms, and thus, contributing to the virus spill over to humans. Here, kidney epithelial cell lines derived from four bat species (Pteropus dasymallus, Rousettus leschenaultii, Rhinolophus ferrumequinum, and Miniopterus fuliginosus) and two non-bat species (Homo sapiens and Mesocricetus auratus) were infected with EMCV and JEV. The replication of EMCV and JEV was lower in the bat cell lines derived from R. leschenaultii, R. ferrumequinum, and M. fuliginosus with a higher expression level of pattern recognition receptors (PRRs) (TLR3, RIG-I, and MDA5) and interferon-beta (IFN-β) than that in the non-bat cell lines and a bat cell line derived from P. dasymallus. The knockdown of TLR3, RIG-I, and MDA5 in Rhinolophus bat cell line using antisense RNA oligonucleotide led to decrease IFN-β expression and increased viral replication. These results suggest that TLR3, RIG-I, and MDA5 are important for antiviral response against EMCV and JEV in Rhinolophus bats.
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http://dx.doi.org/10.1016/j.bbrc.2020.04.060DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7177169PMC
June 2020

sp. nov. isolated from the feline oral cavity.

Int J Syst Evol Microbiol 2020 May;70(5):3355-3360

Department of Veterinary Science, National Institute of Infectious Diseases, Tokyo 162-8640, Japan.

Four strains, KC07070, KC07105, 11 025B-8C and 11 026B-8-C, were isolated from the oral cavity of cats in 2007 or 2011 in Japan. These strains were Gram-stain-negative rods, exhibited gliding motility, grew in air with 5 % CO and showed catalase and oxidase activity. The sequences of 16S rRNA genes of the four strains were 100 % identical. Additionally, the sequences of 16S rRNA genes of KC07070 had identity to those of the type strains of (97.7 %), (97.8 %) and (97.4 %) and 91.2-93.8% identity to those of other species of the genus . The major cellular fatty acids of KC07070 were iso-C (56.2 %) and summed feature 11 (14.9 %). The G+C content of the DNA from KC07070 was 35.6 mol%, and the genome size was 2.88 Mbp. KC07070 had digital DNA-DNA hybridization (dDDH) values of 26.2-27.6% and average nucleotide identity (ANI) values of 75.4-83.3 % to the type strains of the closest relatives, , and . These results of phylogenetic analysis of 16S rRNA gene sequence, cellular fatty acids compositions and dDDH and ANI values indicate that strain KC07070 represents a novel species, for which we propose the name sp. nov., with type strain KC07070 (=JCM 32681=DSM 107251).
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http://dx.doi.org/10.1099/ijsem.0.004176DOI Listing
May 2020

The first discovery of severe fever with thrombocytopenia syndrome virus in Taiwan.

Emerg Microbes Infect 2020 10;9(1):148-151. Epub 2020 Jan 10.

College of Veterinary Medicine, National Chung Hsing University, Taichung, Taiwan.

Severe fever with thrombocytopenia syndrome (SFTS), an emerging tick-borne zoonosis, has been rapidly spread in many Asian counties since 2010, which raises the great concern in East Asia. Nevertheless, the infection status of SFTS in Taiwan remains unclear. To investigate the existence of SFTSV in Taiwan, a total of 151 serum samples collected from 31 sheep, 63 bovine and 57 dogs were enrolled this study. Furthermore, 360 adult female were also included. One-step RT-nested PCR and IgG ELISA were conducted to test SFTSV specific RNA and antibodies, respectively. The result provided the first evidence of the existence of SFTSV RNA and antibodies in ruminants and ticks in Taiwan.
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http://dx.doi.org/10.1080/22221751.2019.1710436DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6968498PMC
September 2020

Distribution of Japanese Encephalitis Virus, Japan and Southeast Asia, 2016-2018.

Emerg Infect Dis 2020 01;26(1):125-128

During 2016-2018, we conducted surveillance for Japanese encephalitis virus (JEV) in mosquitoes and pigs in Japan, Thailand, the Philippines, and Indonesia. Phylogenetic analyses demonstrated that our isolates (genotypes Ia, Ib, III, IV) were related to JEV isolates obtained from the same regions many years ago. Indigenous JEV strains persist in Asia.
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http://dx.doi.org/10.3201/eid2601.190235DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6924880PMC
January 2020

Effective methods for the inactivation of Francisella tularensis.

PLoS One 2019 14;14(11):e0225177. Epub 2019 Nov 14.

Department of Veterinary Science, National Institute of Infectious Diseases, Tokyo Japan.

Francisella tularensis (F. tularensis) is highly pathogenic to humans and must be handled under biosafety level 3 conditions. Samples used for the diagnosis and experimental analysis must be completely inactivated, although methods for the inactivation of F. tularensis are limited. In this study, effective methods for the inactivation of F. tularensis SCHU P9 and five other strains were determined by comparisons of colony-forming units between treated and control samples. The results showed that F. tularensis SCHU P9 was denatured by heat treatment (94°C for 3 min and 56°C for 30 min), filtration with a 0.22 μm filter, and the use of various solutions (i.e. >70% ethanol, methanol, acetone, and 4% paraformaldehyde). F. tularensis SCHU P9 remained viable after treatment with 50% ethanol for 1 min, filtration with a 0.45 μm filter, and treatments with detergents (i.e. 1% lithium dodecyl sulfate buffer, 1% Triton X-100 and 1% Nonidet P-40) at 4°C for 24 h. Additionally, F. tularensis SCHU P9 suspended in fetal bovine serum in plastic tubes was highly resistant to ultraviolet radiation compared to suspensions in water and chemically defined medium. The methods for inactivation of F. tularensis SCHU P9 was applicable to the other five strains of F. tularensis. The data presented in this study could be useful for the establishment of guidelines and standard operating procedures (SOP) to inactivate the contaminated samples in not only F. tularensis but also other bacteria.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0225177PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6855423PMC
March 2020

First record of Larsonella pumilus (Teleostei: Gobiidae) from Japan, with phylogenetic placement of the genus Larsonella.

Zootaxa 2019 Nov 11;4695(4):zootaxa.4695.4.4. Epub 2019 Nov 11.

Okinawa Churashima Foundation Research Center, 888 Ishikawa, Motobu-cho, Okinawa 905-0206, Japan..

During a survey of deep-sea fauna, using a Remotely Operated Vehicle, a single specimen (21.6 mm in standard length) of Larsonella pumilus (Larson Hoese, 1980) was collected at a depth of 214 m off the coast of Okinawa Island, Japan. It represents the first record of this species from Japan. The collection site was far deeper than previous reports for this species. This suggests that the main habitat of L. pumilus is deeper than previously recognized and it may explain the paucity of records of this species. As the previously available morphological description of L. pumilus was based on only a single specimen (holotype), this new specimen is described herein. Its morphology corresponds closely to the original description of the holotype, except that faint melanophores are arranged radially around the eyes and scattered on the trunk and the fins. Mitochondrial genome sequences of L. pumilus and 19 related species demonstrate close relationships between L. pumilus and the genus Priolepis. These data also indicate that the genus Priolepis is not monophyletic.
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http://dx.doi.org/10.11646/zootaxa.4695.4.4DOI Listing
November 2019

Antiviral effect of sinefungin on in vitro growth of feline herpesvirus type 1.

J Antibiot (Tokyo) 2019 12 18;72(12):981-985. Epub 2019 Sep 18.

Joint Faculty of Veterinary Medicine, Yamaguchi University, 1677-1 Yoshida, Yamaguchi, 753-8515, Japan.

Feline herpesvirus type 1 (FHV-1) causes a potentially fatal disease in cats. Through the use of virus inhibition and cytotoxicity assays, sinefungin, a nucleoside antibiotic, was assessed for its potential to inhibit the growth of FHV-1. Sinefungin inhibited in vitro growth of FHV-1 most significantly over other animal viruses, such as feline infectious peritonitis virus, equine herpesvirus, pseudorabies virus and feline calicivirus. Our results revealed that sinefungin specifically suppressed the replication of FHV-1 after its adsorption to the host feline kidney cells in a dose-dependent manner without obvious cytotoxicity to the host cells. This antibiotic can potentially offer a highly effective treatment for animals infected with FHV-1, providing alternative medication to currently available antiviral therapies.
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http://dx.doi.org/10.1038/s41429-019-0234-4DOI Listing
December 2019

Natural severe fever with thrombocytopenia syndrome virus infection in domestic cats in Japan.

Vet Microbiol 2019 Sep 20;236:108346. Epub 2019 Jul 20.

Laboratory of Veterinary Microbiology, Joint Faculty of Veterinary Medicine, Yamaguchi University, 1677-1 Yoshida, Yamaguchi, 753-8511, Japan.

Severe fever with thrombocytopenia syndrome (SFTS) is a recently discovered emerging infectious disease. A zoonotic disease with a high fatality rate in human beings, clinical information on SFTS virus (SFTSV) infection in animals is important. Since 2017, we have diagnosed 24 client-owned cats living in western Japan with SFTS, by genetic and serological testing. In this study, we characterized the clinical features of SFTS in cats and their associated risk factors, by evaluating the clinical parameters retrospectively. A phylogenetic analysis on SFTSV was also conducted. There were no obvious tendencies in age or sex, outdoor cats were commonly at risk of SFTSV infection. All infected cats showed acute onset of clinical signs including anorexia and lethargy, while 68.2% of the cats showed fever and 41.7% showed vomiting. The case fatality rate was 62.5%. Thrombocytopenia, leukopenia, and elevated serum total bilirubin, serum amyloid A, and creatinine phosphokinase concentration were the characteristic findings in the first clinical blood examination. Phylogenic analysis revealed that regional clustered viruses infect both humans and cats. For pet owners and animal hospitals, SFTS in small animals could be an important public health issue.
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http://dx.doi.org/10.1016/j.vetmic.2019.06.019DOI Listing
September 2019

Severe Fever with Thrombocytopenia Syndrome Phlebovirus causes lethal viral hemorrhagic fever in cats.

Sci Rep 2019 08 19;9(1):11990. Epub 2019 Aug 19.

Department of Veterinary Science, National Institute of Infectious Diseases, Tokyo, Japan.

Severe fever with thrombocytopenia syndrome (SFTS) is an emerging hemorrhagic fever caused by the SFTS phlebovirus (SFTSV). SFTS patients were first reported in China, followed by Japan and South Korea. In 2017, cats were diagnosed with SFTS for the first time, suggesting that these animals are susceptible to SFTSV. To confirm whether or not cats were indeed susceptible to SFTSV, animal subjects were experimentally infected with SFTSV. Four of the six cats infected with the SPL010 strain of SFTSV died, all showing similar or more severe symptoms than human SFTS patients, such as a fever, leukocytopenia, thrombocytopenia, weight loss, anorexia, jaundice and depression. High levels of SFTSV RNA loads were detected in the serum, eye swab, saliva, rectal swab and urine, indicating a risk of direct human infection from SFTS-infected animals. Histopathologically, acute necrotizing lymphadenitis and hemophagocytosis were prominent in the lymph nodes and spleen. Severe hemorrhaging was observed throughout the gastrointestinal tract. B cell lineage cells with MUM-1 and CD20, but not Pax-5 in the lesions were predominantly infected with SFTSV. The present study demonstrated that cats were highly susceptible to SFTSV. The risk of direct infection from SFTS-infected cats to humans should therefore be considered.
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http://dx.doi.org/10.1038/s41598-019-48317-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6700174PMC
August 2019

Establishment of a Virulent Full-Length cDNA Clone for Type I Feline Coronavirus Strain C3663.

J Virol 2019 11 15;93(21). Epub 2019 Oct 15.

Laboratory of Clinical Research on Infectious Diseases, Research Institute for Microbial Diseases, Osaka University, Osaka, Japan

Feline infectious peritonitis (FIP) is one of the most important infectious diseases in cats and is caused by feline coronavirus (FCoV). Tissue culture-adapted type I FCoV shows reduced FIP induction in experimental infections, which complicates the understanding of FIP pathogenesis caused by type I FCoV. We previously found that the type I FCoV strain C3663 efficiently induces FIP in specific-pathogen-free cats through the naturally infectious route. In this study, we employed a bacterial artificial chromosome-based reverse genetics system to gain more insights into FIP caused by the C3633 strain. We successfully generated recombinant virus (rC3663) from Fcwf-4 cells transfected with infectious cDNA that showed growth kinetics similar to those shown by the parental virus. Next, we constructed a reporter C3663 virus carrying the nanoluciferase (Nluc) gene to measure viral replication with high sensitivity. The inhibitory effects of different compounds against rC3663-Nluc could be measured within 24 h postinfection. Furthermore, we found that A72 cells derived from canine fibroblasts permitted FCoV replication without apparent cytopathic effects. Thus, our reporter virus is useful for uncovering the infectivity of type I FCoV in different cell lines, including canine-derived cells. Surprisingly, we uncovered aberrant viral RNA transcription of rC3663 in A72 cells. Overall, we succeeded in obtaining infectious cDNA clones derived from type I FCoV that retained its virulence. Our recombinant FCoVs are powerful tools for increasing our understanding of the viral life cycle and pathogenesis of FIP-inducing type I FCoV. Feline coronavirus (FCoV) is one of the most significant coronaviruses, because this virus induces feline infectious peritonitis (FIP), which is a lethal disease in cats. Tissue culture-adapted type I FCoV often loses pathogenicity, which complicates research on type I FCoV-induced feline infectious peritonitis (FIP). Since we previously found that type I FCoV strain C3663 efficiently induces FIP in specific-pathogen-free cats, we established a reverse genetics system for the C3663 strain to obtain recombinant viruses in the present study. By using a reporter C3663 virus, we were able to examine the inhibitory effect of 68 compounds on C3663 replication in Fcwf-4 cells and infectivity in a canine-derived cell line. Interestingly, one canine cell line, A72, permitted FCoV replication but with low efficiency and aberrant viral gene expression.
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http://dx.doi.org/10.1128/JVI.01208-19DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6803248PMC
November 2019

Risk assessment for hepatitis E virus infection from domestic pigs introduced into an experimental animal facility in a medical school.

J Vet Med Sci 2019 Aug 8;81(8):1191-1196. Epub 2019 Jul 8.

Department of Virology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, 2-5-1 Shikata-cho, Kita-ku, Okayama 700-8558, Japan.

Hepatitis E virus (HEV) is known to cause zoonotic infections from pigs, wild boars and deer. Domestic pigs have been used as an experimental animal model in medical research and training; however, the risks of HEV infection from pigs during animal experiments are largely unknown. Here, we retrospectively investigated the seroprevalence and detection rates of viral RNA in 73 domestic pigs (average 34.5 kg) introduced into an animal experimental facility in a medical school during 2012-2016. We detected anti-HEV immunoglobulin G antibodies in 24 of 73 plasma samples (32.9%), though none of the samples were positive for viral RNA. Plasma samples of 18 pigs were sequentially monitored and were classified into four patterns: sustained positive (5 pigs), sustained negative (5 pigs), conversion to positive (6 pigs) and conversion to negative (2 pigs). HEV genomes were detected in 2 of 4 liver samples from pigs that were transported from the same farm during 2016-2017. Two viral sequences of the overlapping open reading frame (ORF) 2/3 region (97 bp) were identical and phylogenetically fell into genotype 3. A 459-bp length of the ORF2 region of an amplified fragment from a pig transported in 2017 was clustered with the wbJYG1 isolate (subgenotype 3b) with 91.5% (420/459 bp) nucleotide identity. Based on our results, we suggest that domestic pigs introduced into animal facilities carry a potential risk of HEV infection to researchers, trainees and facility staff. Continuous surveillance and precautions are important to prevent HEV infection in animal facilities.
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http://dx.doi.org/10.1292/jvms.19-0086DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6715933PMC
August 2019