Parasitological survey on wild carnivora in north-western Tohoku, Japan.

In the winter of 1997-1998, we collected parasitological data from 60 wild carnivora in the north-western part of Tohoku region, Japan. These included 7 foxes (Vulpes vulpes japonica), 20 raccoon dogs (Nyctereutes procyonoides viverrinus), 29 martens (Martes melampus melampus), 3 weasels (two Mustela sibirica itatsi and one M. nivalis namiyei), and one Japanese badger (Meles meles anakuma). Roundworms (Toxocara canis in foxes and Toxocara tanuki in raccoon dogs), hookworms (Ancylostoma kusimaense and Arthrostoma miyazakiense) and Molineus sp. in the small intestine were the most prevalent in foxes and raccoon dogs. In martens, Aonchotheca putorii in the stomach, Concinnum ten in the pancreatic duct, Molineus sp. and Euryhelmis costaricensis in the small intestine were the most prevalent. Collected parasites include some new helminth species for this region or Japan; the strobilar stage of Taenia polyacantha from foxes, Pygidliopsis summa from a raccoon dog, Eucoleus aerophilus, A. putorii, and Soholiphyme baturini from martens.     

Epizootiological survey of Trichinella spp. infection in carnivores, rodents and insectivores in Hokkaido, Japan

In order to evaluate the present epidemiological situation of Trichinella infection in wild animals in Hokkaido, Japan, red foxes (Vulpes vulpes), raccoon dogs (Nyctereutes procyonoides) , brown bears (Ursus arctos) , martens (Martes melampus), rodents and insectivores captured in Hokkaido were examined for muscle larvae by the artificial digestion method from 2000 to 2006. Foxes (44/319, 13.8%), raccoon dogs (6/77, 7.8%) and brown bears (4/126, 3.2%) were found to be infected with Trichinella larvae and all other animal species evaluated were negative. Multiplex PCR and DNA sequencing revealed that larvae from a fox captured in Otofuke, in south-eastern Hokkaido, were T. nativa, and larvae from 27 animals including 21 foxes, 2 raccoon dogs and 4 brown bears captured in western Hokkaido were Trichinella T9.     

Mitochondrial DNA variation in the Japanese marten Martes melampus and Japanese sable, Martes zibellina

Genetic relationship among Japanese sables, Martes zibellina and the introduced Japanese martens, Martes melampus in northern Japan was revealed by analyzing a 521-524bp DNA sequence from the cytochrome b (112bp)/transfer RNA-threonine (67bp)/tRNA-proline (65bp) and control region (277-280bp) of the mitochondrial genome. Intraspecific differences in sequences of M. zibellina and M. melampus (3.8-15.0% and 1.9-16.4%, respectively) were similar to interspecific differences between these two species (5.8-16.6%). Comparison of sequence data exhibited five haplotypes of M. melampus and four haplotypes of M. zibellina, which clustered into two groups (clusters-A and-B). Cluster-A included two haplotypes of M. melampus and two haplotypes of M. zibellina, whereas cluster-B included three haplotypes of M. melampus and two haplotypes of M. zibellina. Results of this study lead three possible explanations. Firstly, past hybridization between M. zibellina and M. melampus might have occurred. Secondary, these two species might have similar heteroplasmy of mtDNA. Thirdly, these haplotypes might have come from nuclear genome. Although further intensive studies are needed to make a conclusion, detection of hybridization with the Japanese marten are occurred or not is quite important to conserve the Japanese sable.     

Nematodes from raccoon dogs (Nyctereutes procyonoides viverrinus) introduced recently on Yakushima Island, Japan

Nematodes were collected from 14 raccoon dogs (Nyctereutes procyonoides viverrinus) on Yakushima Island, Kagoshima Prefecture (Kyushu), Japan, that were the progeny of recently introduced animals to this World Natural Heritage Area, probably within the last 2 decades. Seven nematode species (Toxocara tanuki, Ancylostoma kusimaense, Arthrostoma miyazakiense, Strongyloides planiceps, Trichuris vulpis, Gongylonema sp., and Dirofilaria immitis) were collected from the raccoon dogs. Two nematode species (Molineus legerae and Strongyloides martis) were collected from 2 Japanese weasels (Mustela itatsi) indigenous to this island that were killed by traffic accidents. Considering that foxes and other carnivores, except for weasels, dogs and cats, are not distributed on this island, most of soil-borne nematodes such as roundworms, hookworms, and threadworms in raccoon dogs should have been introduced by the original population from the mainland Japan, and the infection hence maintained at a high level by new generations. Recovery of the third-stage larvae of Gongylonema sp. from the esophagus of raccoon dogs was remarkably frequent (79%), suggesting that the animals might vigorously take insect hosts. Although S. martis have never been recorded from weasels in Japan until this study, this threadworm might be prevalent in a variety of mustelids in Eurasia.     

Juvenile bird acanthocephalans recovered incidentally from raccoon dogs (Nyctereutes procyonoides viverrinus) on Yakushima Island, Japan

Helminth parasites were collected from 14 raccoon dogs (Nyctereutes procyonoides viverrinus) and 2 Japanese weasels (Mustela itatsi sho) on Yakushima Island, Kagoshima Prefecture (Kyushu), Japan. In addition to trematodes and nematodes reported separately, everted cystacanths of 3 bird acanthocephalans (Mediorhynchus robustus in 5 animals, Porrorchis leibyi in 2 animals, and Sphaerirostris lanceoides in 1 animal) were collected from the raccoon dogs introduced recently on this World Natural Heritage Area. A cystacanth of P. leibyi was collected also from a weasel indigenous to this island that were killed by traffic accidents. Recovery of everted cystacanths of M. robustus from the intestine of raccoon dogs was remarkably frequent (36%), suggesting that this acanthocephalan should be highly prevalent in passerine hosts on Yakushima Island, and the animals might vigorously take insect hosts or paratenic hosts. Furthermore, the present report is the second on M. robustus after its first record in Japan (Aichi Prefecture, Honshu) as M. garruli from a Eurasian jay (Garrulus glandarius japonicus) by Yamaguti in 1939. Porrorchis leibyi has never been recorded in Japan.     

Epizootiological investigations of canine distemper virus in free-ranging carnivores from Germany

Canine distemper virus (CDV) infects a broad range of carnivores. To assess whether wild carnivores may play a role in the epidemiology of CDV in domestic dogs in Germany, the seroprevalence of CDV was determined. In sera from red foxes (30 of 591 (5%)) and stone martens (2 of 10 (20%)) antiviral antibodies were detected using a neutralization assay, whereas sera of raccoons, two mink, one pine marten and one raccoon dog were negative. In foxes, there was a significantly higher prevalence in urban and suburban compared to rural regions. When testing lung and spleen tissue samples (fox, badger, stone marten, polecat, raccoon dog) 13 of 253 (5.1%) foxes, 2 of 13 (15.4%) stone martens and 2 of 6 (33%) badgers were virus positive using RT-PCR. Phylogenetic analysis based on partial sequences of the F gene revealed a distinct relatedness to canine CDV isolates. Together, the data support the concept of transmission of CDV between domestic dogs and wild carnivores.     

Trichinella spiralis in wild animals, cats, mice, rats and farmed fur animals in Finland

A total of 1 399 samples of wild animals, cats, mice, rats and fur animals were examined for Trichinella larvae during the period 1.1.1982-30.6.1984. Samples were obtained both from the problem area, were Trichinella larvae had been found in pigs, and for comparison from the rest of Finland. The frequency of the infection in wild carnivores, badgers, pine martens, raccoon dogs, foxes and wild mink was significantly higher in the problem area than in the rest of Finland.     

PCR detection of Neospora caninum, Toxoplasma gondii and Encephalitozoon cuniculi in brains of wild carnivores

Neospora caninum, Toxoplasma gondii and Encephalitozoon cuniculi are important pathogens with affinity to the central nervous system of many animals. 240 brains of wild carnivores were examined by PCR-based diagnosis. The presence of N. caninum DNA was confirmed in 4.61% (7/152) red foxes (Vulpes vulpes). DNA of T. gondii was found in 4.92% (3/61) martens (Martes sp.) and in 1.32% (2/152) red foxes. DNA of E. cuniculi was determined in 3.28% (2/61) martens and in one examined European otter (Lutra lutra). There were no co-infections found. These results provide the first evidence of E. cuniculi in the European otter, the first report of N. caninum in foxes in the Czech Republic and confirm the presence of T. gondii in wild carnivores in the Czech Republic.     

Paragonimus westermani and some rare intestinal trematodes recovered from raccoon dogs (Nyctereutes procyonoides viverrinus) introduced recently on Yakushima Island, Japan

Helminth parasites were collected from 9 raccoon dogs (Nyctereutes procyonoides viverrinus) and 2 Japanese weasels (Mustela itatsi sho) on Yakushima Island, Japan. The former carnivore was introduced to this World Natural Heritage Area presumably within the last two decades, expanding its population thence, although detailed process(es) of the introduction is unknown. The collected trematodes from raccoon dogs included the triploid form of Paragonimus westermani, Brachylaima tokudai, Maritrema eroliae, and Pseudocryptotropa sp. Simultaneously, Paragonimus ohirai was found in one weasel killed by a traffic accident. Although the triploid form of P. westermani and P. ohirai are known to be distributed in some river-mouth areas of Yakushima Island based on previous surveys on crab hosts, natural infection was detected for the first time in wild final hosts. Particularly, the raccoon dog infected with P. westermani was caught in a mountainous area, distant from human residence or river-mouth areas. Although it is possible that the infected raccoon dog moved from a river-mouth area endemic with P. westermani after infection, the alternative scenario remains to be pursued; the endemic area of this zoonosis is expanding along with the recent expansion of raccoon dogs or feral cats (Felis catus), that became prevalent recently on this island including the mountainous areas. Maritrema eroliae taking a variety of shorebirds as its natural final hosts, and a minute trematode, Pseudocryptotropa sp., taking unknown natural final host(s) were recorded for the first time in raccoon dogs.     

Neospora caninum antibodies in wild carnivores from Spain

Serum samples from 251 wild carnivores from different regions of Spain were tested for antibodies to Neospora caninum by the commercial competitive screening enzyme linked immunosorbent assay (c-ELISA) and confirmed by Neospora agglutination test (NAT) and/or by indirect fluorescent antibody test (IFAT). Samples with antibodies detected by at least two serological tests were considered seropositive. Antibodies to N. caninum were found in 3.2% of 95 red foxes (Vulpes vulpes); in 21.4% of 28 wolves (Canis lupus); in 12.0% of 25 Iberian lynx (Lynx pardinus); in 16.7% of 6 European wildcats (Felis silvestris); in 6.4% of 31 Eurasian badgers (Meles meles); in 21.4% of 14 stone martens (Martes foina); in 66.7% of 3 pine martens (M. martes) and in 50% of 2 polecats (Mustela putorius). Antibodies to N. caninum in common genets (Genetta genetta) and Egyptian mongooses (Herpestes ichneumon) were only observed by c-ELISA but were not confirmed by IFAT and/or NAT. No antibodies were detected in 5 Eurasian otters (Lutra lutra) by any technique. Statistically significant differences were observed among species and among geographical areas. The highest seroprevalence of N. caninum infection was observed in the Cantabric Coastal region characterized by high humidity. To our knowledge, this is the first report of antibodies to N. caninum in free ranging wild carnivores, other than wild canids, in Europe. The existence of a possible sylvatic cycle could have important implications in both sylvatic and domestic cycles since they might influence the prevalence of infection in cattle farms in those areas.     

貉细小病毒LN10-1株分离鉴定及其免疫原性研究

为研制貉细小病毒性肠炎疫苗,筛选出针对貉细小病毒性肠炎免疫原性好、安全高效的疫苗备选株,应用CRFK细胞从辽宁省发病貉的粪便中分离病毒,并通过形态学、血清学、分子生物学、动物回归及免疫接种等方法对分离株进行鉴定。鉴定结果表明成功分离出1株貉细小病毒,命名为LN10-1株。其VP2基因核苷酸序列与猕猴源猫泛白细胞综合征病毒株(BJ-22/2008/CHN株)相似性高达99.7%。VP2蛋白上决定宿主范围的2个氨基酸位点发生了突变。VP2基因种系发生分析显示,LN10-1株位于猫泛白细胞综合征病毒(Feline panleukopenia virus,FPLV)、蓝狐细小病毒(Blue fox parvovirus,BFPV)、水貂肠炎病毒(Mink enteritis virus,MEV)组成的食肉类动物细小病毒聚类分支与由犬细小病毒(Canine parvovirus,CPV)组成的聚类分支。由LN10-1株制备的灭活疫苗免疫结果显示,接种28d细小病毒中和抗体滴度可达到1∶256以上。推测LN10-1株可能正处于FPLV与CPV进化的中间状态,或是CPV适应新宿主(貉)而形成的一种新病毒,可以作为针对貉细小病毒性肠炎灭活疫苗的候选株。     

Evidence for widespread Leishmania infantum infection among wild carnivores in L. infantum periendemic northern Spain

Leishmania spp. infection was investigated in tissue samples of wild carnivores from the Spanish Basque Country (BC), by PCR and DNA sequencing. The region is at the northern periphery of Leishmania infantum endemic Iberian Peninsula and infection in the dog (reservoir) or other species has not been previously reported. Leishmania kinetoplast DNA was detected by real-time PCR (rtPCR) in 28% (44/156) of animals. Specifically, in 26% of Eurasian badgers (n = 53), 29% of foxes (n = 48), 29% of stone martens (n = 21) and in 25–50% of less numerous species including genets, wild cats, pole cats, European mink and weasels. Infected animals particularly badgers, were most prevalent in the southernmost province of the BC (Araba) in areas dominated by arable land. Subsequent amplification and sequencing of a fragment of the rRNA internal transcribed spacer 2 (ITS2) from a subset of rtPCR positives samples confirmed the species as L. infantum, showing a high sequence homogeneity with ITS2 sequences of L. infantum from dogs and humans from southern Spain. In summary, this study reports for the first time L. infantum infection in wild carnivores from the BC including in stone martens, pole cats and minks in which infection has not been previously described. It supports the need to study infection in dogs and people in this region and is an example of the value of infection surveillance in wildlife to assess potential risks in the domestic environment and their role in spreading infections in non-endemic areas.     

Survey of Cryptosporidium spp. and Giardia spp. infections in various animals at a zoo in Japan.

A total of 284 fecal samples of 89 species (43 mammalian species and 46 avian species) were examined for Cryptosporidium oocysts and Giardia cysts from 1999 to 2002. Each sample was collected at the zoo located at Osaka in Japan and examined by microscopy after performing the sucrose flotation method and by two immunofluorescent assay kits for detection of Cryptosporidium oocysts and Giardia cysts. Cryptosporidium spp. was found only in a raccoon dog (Nyctereutes procyonoides), and Giardia spp. was detected in a mandarin duck (Aix galericulata) and two ruddy shelducks (Tadorna ferruginea). In this study, the prevalences of these parasites were found to be low. However, these results suggested that the infected animals could serve as a source of contamination for surface water. This is the first report about the survey of Cryptosporidium spp. and Giardia spp. at a zoo in Japan.     

Mange caused by Sarcoptes scabiei (Acari: Sarcoptidae) in wild raccoon dogs, Nyctereutes procyonoides, in Kanagawa Prefecture, Japan

Parasitological and histopathological examinations were performed in 25 raccoon dogs (Nyctereutes procyonoides) obtained in Kanagawa Prefecture, Japan, all of which were found to be heavily infected with Sarcoptes scabiei. The mites detected on these raccoon dogs were morphologically indistinguishable from the human species, and no Demodex mites were detected. Histopathological examinations showed prominent hyperkeratosis and acanthosis with eczema, and numerous burrows containing mites were observed in the epidermis. The enzootic dermatitis of wild raccoon dogs in recent years was clearly demonstrated to be caused by S. scabiei in the present study.     

Ovicidal and larvicidal activity in vitro of Spigelia anthelmia Linn. extracts on Haemonchus contortus

The prevalence of antibodies to Neospora caninum was examined in six wild Artiodactyla species, and in five wild Carnivora species from Kenya. Blood sera (104 wild ungulates from Marula Estates (MEs), and 31 wild carnivores from Masai-Mara reserve and from other wildlife areas in northern and Southern Kenya), were screened using a Neospora agglutination test (NAT), with a twofold dilution (1:40-1:320 titres). Presence of NAT antibodies to N. caninun is reported here for the first time in zebra (Equus burchelli), eland (Taurotragus oryx), African buffalo (Syncerus caffer), Thompson gazelle (Gazella thompsoni), impala (Aepyceros melampus), warthog (Phacochoerus aethiopicus), spotted hyena (Crocuta crocuta) and in free-ranging cheetah (Acinonyx jubatus). At 1:80 dilution, prevalence was 61.5% in eland, 58.5% in zebra, 19.2% in Thompson gazelle, 33.3% in warthog, 50% in African buffalo, 30% in lion (Panthera leo), 20% in cheetah, and 33.3% in spotted hyena. Antibodies up to 1:320 titre were detected in eland (38.4%), zebra (19.5%), Thompson gazelle (3.8%) and lion (5%). Amongst herbivores, sero-prevalence was significantly (P<0.05) higher, at all dilutions, in "grazer/digger" species (e.g. eland and zebra) than in non-"grazer/digger" species (e.g. impala and Thompson gazelle). No antibodies to N. caninum were found in two leopards (Panthera pardus) and one serval (Felis serval). Our results indicates a steady presence of N. caninum in wild mammals from Kenya. The hypothesis of a sylvatic cycle of N. caninum could be suggested, but more data are needed to verify the hypothesis, as to evaluate the role of N. caninum infection on the dynamics of wild animals population in the study area.     

Epidemiology of sarcoptic mange in free-ranging raccoon dogs (Nyctereutes procyonoides) in Yokohama,Japan

Free-ranging raccoon dogs (Nyctereutes procyonoides) from Nogeyama Zoological Gardens, Kanazawa Zoological Gardens, and Yokohama Zoological Gardens frequently rescued dogs having Sarcoptes scabiei infestation. However, the epidemiology of S. scabiei infestation has not yet been elucidated. In the present study, we investigated the epidemiology of S. scabiei infestation in raccoon dogs and its influence on the population of masked palm civets in Yokohama, Japan. We examined records of raccoon dog rescue between 1981 and 2010 and classified the dogs into the following 4 categories on the basis of the reason for rescue: dogs with S. scabiei infestation, scabies-infested dogs involved in car accidents, uninfested dogs involved in car accidents, and other reasons for rescue. We found that the number of dogs rescued due to car accidents and other reasons increased from 1989 onwards, and an S. scabiei outbreak was recorded since 1993. The infestation spread from the southern to the northern regions of Yokohama. The total number of raccoon dogs rescued annually peaked in 1995 and declined thereafter. The number of masked palm civets (Paguma larvata) rescued gradually increased with a decline in the number of raccoon dogs rescued. In the present study, we revealed the epidemiology of S. scabiei infestation in the raccoon dog. The outbreak might be induced by the increased population density, and the infestation spread immediately from the southern to the northern regions of Yokohama since 1993. Further, the population of masked palm civets may have increased due to the decrease in the population of the raccoon dog.     

Canine distemper virus infection among wildlife before and after the epidemic

In 2007–2008, a canine distemper virus (CDV) epidemic occurred among wild animals in Wakayama Prefecture, Japan, and many mammals, including the wild boar and deer, were infected. In this study, CDV prevalence among wild animals was surveyed before and after the epidemic. At first, an enzyme-linked immunosorbent assay (ELISA) with horseradish peroxidase-conjugated protein A/G was established to detect CDV antibodies in many mammalian species. This established ELISA was available for testing dogs, raccoons and raccoon dogs as well as virus-neutralization test. Next, a serological survey of wild mammalians was conducted, and it was indicated that many wild mammalians, particularly raccoons, were infected with CDV during the epidemic, but few were infected before and after the epidemic. On the other hand, many raccoon dogs died during the epidemic, but CDV remained prevalent in the remaining population, and a small epidemic occurred in raccoon dogs in 2012–2013. These results indicated that the epidemic of 2007–2008 may have been intensified by transmission to raccoons.     

Prevalence of Dirofilaria immitis infection in living raccoon dogs assessed by hematological examination

The prevalence of Dirofilaria immitis in free-ranging raccoon dogs (Nyctereutes procyonoides viverrinus) was examined in the southeast region of Kanagawa Prefecture, Japan, using a rapid immunomigration (RIM) test kit. Between April 2007 and March 2010, we examined 108 raccoon dogs rescued and housed by the Kanazawa Zoological Garden. D. immitis infection was found in 8 (7.4%) raccoon dogs. This is the first report to reveal the prevalence of D. immitis infection in living raccoon dogs. The prevalence of the infection was lower than previously reported values obtained on postmortem examination. One reason might be that the present study included young raccoon dogs infected with immature worms. Significant high-risk areas of D. immitis infection in the raccoon dogs were not observed.     

Twenty year experience of the oral rabies vaccine SAG2 in wildlife: a global review

The SAG2 vaccine (RABIGEN® SAG2) is a modified live attenuated rabies virus vaccine, selected from the SAD Bern strain in a two-step process of amino acid mutation using neutralizing monoclonal antibodies. The strain is genetically stable and does not spread in vivo or induce a persistent infection. Its absence of residual pathogenicity was extensively demonstrated in multiple target and non target species (such as wild carnivores and rodent species), including non-human primates. The efficacy of SAG2 baits was demonstrated according to the EU requirements for the red fox and raccoon dog. The use of safe and potent rabies vaccines such as SAG2 largely contributed to the elimination of rabies in Estonia, France, Italy and Switzerland. Importantly, these countries were declared free of rabies after few years of oral vaccination campaigns with SAG2 baits distributed with an appropriate strategy. The excellent tolerance of the SAG2 vaccine has been confirmed in the field since its first use in 1993. No safety issues have been reported, and in particular no vaccine-induced rabies cases were diagnosed, after the distribution of more than 20 million SAG2 baits in Europe.     

Tick infestation of sika deer (Cervus nippon) in the western part of Yamaguchi Prefecture,Japan

Ticks were collected from 94 sika deer (Cervus nippon) hunted in the western part of Yamaguchi Prefecture, Japan from August to November 1999, and March to July 2000. Haemaphysalis longicornis and H. yeni were the dominant species from April to August, while H. flava and H. megaspinosa were dominant in October, November and March. This is the first report of H. yeni in the mainland of Japan. Small numbers of H. kitaokai, Amblyomma testudinarium and Ixodes ovatus were also recorded.