Epidemiological Survey of Tick-Borne Encephalitis Virus Infection in Wild Animals on Hokkaido and Honshu Islands, Title Japan
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Epidemiological survey of tick-borne encephalitis virus infection in wild animals on Hokkaido and Honshu islands, Title Japan Jamsransuren, Dulamjav; Yoshii, Kentaro; Kariwa, Hiroaki; Asakawa, Mitsuhiko; Okuda, Kei; Fujii, Kei; Fukumoto, Author(s) Shinya; Umemiya-Shirafuji, Rika; Sasaki, Motoki; Matsumoto, Kotaro; Yamaguchi, Emi; Ogawa, Haruko; Imai, Kunitoshi Citation Japanese Journal of Veterinary Research, 67(2), 163-172 Issue Date 2019-05 DOI 10.14943/jjvr.67.2.163 Doc URL http://hdl.handle.net/2115/74797 Type bulletin (article) File Information p163-172 Kunitoshi Imai.pdf Instructions for use Hokkaido University Collection of Scholarly and Academic Papers : HUSCAP Japanese Journal of Veterinary Research 67(2): 163-172, 2019 REGULAR PAPER Experimental Research Epidemiological survey of tick-borne encephalitis virus infection in wild animals on Hokkaido and Honshu islands, Japan Dulamjav. Jamsransuren1), Kentaro Yoshii2), Hiroaki Kariwa2), Mitsuhiko Asakawa3), Kei Okuda4), Kei Fujii5), Shinya Fukumoto6), Rika Umemiya-Shirafuji6), Motoki Sasaki1), Kotaro Matsumoto1), Emi Yamaguchi1), Haruko Ogawa1) and Kunitoshi Imai1,＊) 1) Department of Veterinary Medicine, Obihiro University of Agriculture and Veterinary Medicine, Inada Nishi 2-11, Obihiro, Hokkaido 080-8555, Japan 2) Faculty of Veterinary Medicine, Hokkaido University, Kita 18 Nishi 9, Kita-ku, Sapporo, Hokkaido 060-0818, Japan 3) School of Veterinary Medicine, Rakuno Gakuen University, 582 Bunkyodai-Midorimachi, Ebetsu, Hokkaido 069-8501, Japan 4) Faculty of Agriculture, Tokyo University of Agriculture and Technology, Saiwai-cho 3-5-8, Fuchu, Tokyo 183-8509, Japan 5) Animal Research Center, Hokkaido Research Organization, Shintoku-cho, Hokkaido 081-0038, Japan 6) National Research Center for Protozoan Diseases, Obihiro University of Agriculture and Veterinary Medicine, Inada Nishi 2-11, Obihiro, Hokkaido 080-8555, Japan Received for publication, October 3, 2018; accepted, November 14, 2018 Abstract The first human case of tick-borne encephalitis (TBE) in Japan was recorded in southern Hokkaido in 1993 and was followed by four further cases in southern, central, and northern Hokkaido during 2016- 2018. However, the distribution of TBE virus (TBEV) foci in Japan is unclear. Therefore, here, we serologically examined raccoons (Procyon lotor), sika deer (Cervus nippon), and wild boars (Sus scrofa) as sentinels of TBEV infection in Hokkaido and in Fukushima and Tochigi Prefectures in Honshu. A total of 1,649 serum samples collected between 2003 and 2018 were screened by enzyme-linked immunosorbent assay using subviral particles and confirmed using the virus neutralization test. In raccoons, the seroprevalence of TBEV was 5.9% (39/662 samples) in central Hokkaido in 2003-2005 and 0.8% (3/368 samples) in eastern Hokkaido in 2010-2018, revealing the presence of TBEV foci in these areas. In addition, 0.5% (2/414) of deer sampled in eastern Hokkaido in 2010-2017 and 2.4% (1/42) of deer sampled in Tochigi Prefecture in 2016-2018 were seropositive. On Honshu, seropositive rodents have previously been detected only in Shimane Prefecture. Therefore, the detection of seropositive animals in Tochigi Prefecture may indicate the widespread distribution of TBEV foci throughout Japan. TBEV and viral genes were not detected in 507 ticks collected in the same area of eastern Hokkaido where seropositive animals were found, reemphasizing the value of using serological examination of wild animals as a tool for revealing unknown TBE risk areas. Our findings also indicate that raccoons may be particularly useful sentinels. Key Words: Deer, Raccoon, Sentinel, Seroprevalence, Tick-borne encephalitis (TBE) *Corresponding author: Kunitoshi Imai, Department of Veterinary Medicine, Obihiro University of Agriculture and Veterinary Medicine, Inada Nishi 2-11, Obihiro, Hokkaido 080-8555, Japan E-mail: [email protected] doi: 10.14943/jjvr.67.2.163 164 TBEV seroprevalence in wild animals Introduction during 2016-201726) and a fifth patient was reported in northern Hokkaido in 2018 (http// Tick-borne encephalitis (TBE) is a tick- www.city.asahikawa.hokkaido.jp/kurashi/135/ transmitted viral infection of the human central 136/150/ d064126.html; in Japanese), suggesting nervous system that occurs in many Eurasian the widespread distribution of TBEV in Hokkaido. countries. The TBE virus (TBEV) belongs to the The location of southern, central, and northern genus Flavivirus within the family Flaviviridae Hokkaido areas is indicated in Fig. 1. Therefore, and is phylogenetically classified into three the examination and clarification of whether subtypes: the Western or European subtype, TBEV is endemic in other parts of Hokkaido such the Siberian subtype, and the Far-Eastern as eastern Hokkaido (Fig. 1) is urgently required subtype18,20,26). to prevent the further spread of this disease. The epidemiology of TBE differs between Outside Hokkaido, Shimane Prefecture in Honshu countries according to the ecology of TBEV3,5,20). (the main island of Japan) is the only other area Forest hematophagous ticks primarily of the that has been shown to be TBEV-endemic based genus Ixodes, such as Ixodes ricinus (I. ricinus) on a serological survey of wild rodents25). and I. persulcatus, are vectors of TBEV and However, the presence of TBEV-endemic areas in transmit the virus to competent hosts during other parts of Honshu has not been sufficiently their blood-feeding, allowing it to circulate in the examined, and few epidemiological studies have environment. Wild rodents are reservoir hosts been conducted in other parts of Japan. of TBEV and develop a long-lasting viremia, Indicator hosts such as cervid species and allowing them to infect other vector ticks and Eurasian wild boars (Sus scrofa) have been used maintaining natural foci. By contrast, larger as sentinels of the virus in TBEV-endemic forest animals, such as cervid species and wild European countries to monitor the distribution boars (Sus scrofa), and domestic animals are and abundance of vector ticks2,12). Very little is indicator hosts for TBEV transmission, being known regarding TBEV infection in these sentinel infested by vast numbers of vector ticks and thus species in Japan, where sika deer (Cervus nippon) facilitating transmission of the virus to new and wild boars are overabundant in many areas and circulating it through their habitats regions, although the latter are not present in but playing no role in transmitting the virus to Hokkaido9,14,21). However, a recent serological new ticks, probably due to them developing only survey did indicate that a small number of brief viremia with low viral titers. Once infected, Hokkaido sika deer (C. n. yesoensis) collected in these animals produce antibodies to TBEV, northern Hokkaido during 2013 and 2014 tested which are probably maintained over a longer positive for TBEV24). period1,4,7,8,12,15). Raccoons (Procyon lotor), which are native to TBE is a notifiable disease in Japan. The first North America, are now widely distributed in human case of TBE was recorded in southern European countries where TBEV is endemic6,17). Hokkaido (the northernmost island of Japan) in However, there have been no reports of TBEV 199322), and it was subsequently revealed that infection in raccoons in these countries. The this region was a TBEV-endemic area based on raccoon is also an invasive species in Japan, the isolation of TBEV-FE strains from I. ovatus where it has rapidly spread since the 1970s to ticks, rodents, and dogs, as well as serological become widely distributed throughout the evidence in rodents, dogs, and horses 22,23,25). country11). However, it is unclear whether raccoons Although there were no further records of TBE carry TBEV-infected ticks as indicator hosts or for a long period, three confirmed cases were play a role as reservoir hosts. reported in southern and central Hokkaido In this study, we investigated the prevalence Dulamjav. Jamsransuren et al. 165 Fig. 1. Location of the sampling sites on Hokkaido and Honshu islands, Japan. The types of wild animals and ticks that were collected are indicated in parentheses. Hokkaido is divided into four areas (central, southern, northern, and eastern Hokkaido). of TBEV in raccoons and Hokkaido sika deer in Tick collection: A total of 420 I. ovatus and 87 I. central and eastern Hokkaido using serological persulcatus adults were collected by flagging examinations and in ticks in eastern Hokkaido in forested areas in Tokachi District, eastern using virological examinations to define TBEV- Hokkaido, in 2018 (Table 1 and Fig. 1). Following endemic areas. In addition, we serologically morphological identification of the species and examined Honshu sika deer (C. n. centralis), wild sex of the ticks, they were pooled into groups of boars, and raccoons captured in Fukushima and 5-10 ticks according to species, sex, and sampling Tochigi Prefectures in Honshu to identify other location. Each pooled sample was homogenized TBEV-endemic areas. Sampling locations, year, in 1 ml of virus transfer medium containing and numbers are described in Table 1 and Fig. 1. Dulbecco’s Modified Eagle’s Medium (DMEM; Nissui, Tokyo, Japan) supplemented with 10% bovine serum albumin (BSA), penicillin (1,000 Materials and Methods units/ml), streptomycin (1 mg/ml), gentamycin (100 μg/ml), and amphotericin B (20 μg/ml). The Serum collection: A total of 1,649 serum samples homogenates were centrifuged, and the resulting from 1,072 raccoons in Hokkaido and Fukushima supernatants were collected and used for virus Prefectures, from 519 sika deer in Hokkaido, and gene detection, as described below. Fukushima, and Tochigi Prefectures, and from 58 wild boars in Fukushima and Tochigi Serological assays: To detect the presence of Prefectures collected during 2003-2018 were TBEV antibodies, all of the sera were first tested investigated (Table 1 and Fig. 1). The detailed by enzyme-linked immunosorbent assay using animal numbers captured in each sampling recombinant TBEV subviral particles with location (district) are provided in Table 1. Prior Strep-tag (Strep-SP ELISA), as previously to testing, all of the sera were inactivated at described13). Briefly, antigens were prepared 56°C for 30 min.