Morphometric, histopathological and molecular findings of Macracanthorhynchus hirudinaceus infection in wild boar (Sus scrofa) from continental Italy.

Although Macracanthorhynchus hirudinaceus is a neglected acanthocephalan of suids occasionally responsible for severe infections in humans, the spread of wild boar (Sus scrofa) populations in Europe could promote the circulation. Herein, we report the first morphometric, histological and molecular characterization of a severe M. hirudinaceus infection in a boar from continental Italy. The boar's intestine displayed granulomatous enteritis due to 24 helminths (14 females, 10 males), identified as adults of M. hirudinaceus by a combined morphometric/molecular approach. The phylogenetic analysis of the cox1 gene revealed a close relationship of the M. hirudinaceus sequence type found herein with those from Hungary and insular Italy. The high haplotype diversity and low nucleotide diversity of M. hirudinaceus specimens would suggest its rapid demographic expansion in the Mediterranean basin. More research is needed to assess the presence of M. hirudinaceus in susceptible beetle species and the role of boars in the epidemiology of infection.

Quantifying the direct and indirect effects of sika deer (Cervus nippon) on the prevalence of infection with Rickettsia in questing Haemaphysalis megaspinosa: A field experimental study.

Sika deer (Cervus nippon) are important hosts for all life stages of Haemaphysalis megaspinosa, a suspected Rickettsia vector. Because some Rickettsia are unlikely to be amplified by deer in Japan, the presence of deer may decrease the prevalence of Rickettsia infection in questing H. megaspinosa. As sika deer decrease vegetation cover and height and thereby indirectly cause changes in the abundance of other hosts, including reservoirs of Rickettsia, the prevalence of Rickettsia infection in questing ticks can also change. We investigated these possible effects of deer on the prevalence of infection with Rickettsia in questing ticks in a field experiment in which deer density was manipulated at three fenced sites: a deer enclosure (Deer-enclosed site); a deer enclosure where deer had been present until 2015 and only indirect effects remained (Indirect effect site); and a deer exclosure in place since 2004 (Deer-exclosed site). Density of questing nymphs and the prevalence of infection with Rickettsia sp. 1 in questing nymphs at each site were compared from 2018 to 2020. The nymph density at the Deer-exclosed site did not significantly differ from that at the Indirect effect site, suggesting that the deer herbivory did not affect the nymph density by reducing vegetation and increasing the abundance of other host mammals. However, the prevalence of infection with Rickettsia sp. 1 in questing nymphs was higher at the Deer-exclosed site than at the Deer-enclosed site, possibly because ticks utilized alternative hosts when deer were absent. The difference in Rickettsia sp. 1 prevalence between the Indirect effect and Deer-exclosed sites was comparable to that between the Indirect effect and Deer-enclosed sites, indicating that the indirect effects of deer were as strong as the direct effects. Examining the indirect effects of ecosystem engineers in the study of tick-borne diseases may be more important than previously recognized.

Molecular characterization of oriental eyeworm (Thelazia callipaeda) detected from raccoon (Procyon lotor) and Japanese raccoon dog (Nyctereutes viverrinus) in Kanto region, Japan.

BACKGROUND: The oriental eyeworm Thelazia callipaeda (Spirurida: Thelaziidae) is an emerging parasitic ocular nematode of carnivores and humans. In domestic animals and humans, the infection causes varying degrees of inflammation and lacrimation, and wild carnivores represent an important reservoir. In this study we examined the infection status and molecular characterization of T. callipaeda in two urban carnivores, raccoons Procyon lotor and wild Japanese raccoon dogs Nyctereutes viverrinus, in the Kanto region of Japan. METHODS: From January 2020 to December 2021, 193 carcasses including 178 raccoons and 15 raccoon dogs were examined for the presence of worms in the eye. The worms from infected animals (one worm per host) were morphologically identified as T. callipaeda. Worms (1-5 worms per host) were subjected to genetic analysis using mitochondrial cytochrome c oxidase subunit I gene sequences. RESULTS: The prevalence of T. callipaeda in raccoons and Japanese raccoon dogs was 20.2% (36/178) and 13.3% (2/15), respectively. The cox1 sequences from 56 worms from 38 animals revealed three haplotypes (h9, h10, and h12). Analysis of multiple worms for five raccoons showed co-infection of two different haplotypes (h9 and h10) in a single host. Comparing our data with published sequences, three sequences obtained from raccoons and raccoon dogs shared the same haplotypes as those reported in humans, dogs, and cats in Japan. CONCLUSIONS: Our findings show a high prevalence of T. callipaeda in raccoons, suggesting that this invasive carnivore species serves as an important natural reservoir of T. callipaeda in the Kanto region of Japan, an area with the highest human population of the country.

Increased Risk of Infection with Severe Fever with Thrombocytopenia Virus among Animal Populations on Tsushima Island, Japan, Including an Endangered Species, Tsushima Leopard Cats.

To investigate the seroprevalence of severe fever with thrombocytopenia syndrome (SFTS) among wild and companion animals on Tsushima Island, Japan, SFTS virus (SFTSV)-specific ELISA and virus-neutralizing tests were conducted on 50 wild boars, 71 Sika deer, 84 dogs, 323 domestic cats, and 6 Tsushima leopard cats. In total, 1 wild boar (1.8%), 2 dogs (2.4%), 7 domestic cats (2.2%), and 1 Tsushima leopard cat (16.7%) were positive for anti-SFTSV antibodies. Among the 11 positive animals, 10 were collected after 2019, and all were found on the southern part of the island. SFTSV, thus far, seems to be circulating within a limited area of Tsushima Island. To protect humans and animals, including endangered Tsushima leopard cats, from SFTSV infection, countermeasures are needed to prevent the spread of SFTSV on Tsushima Island.

Body Length and Craniometrics of Non-Native Raccoons in Two Regions in Middle Japan during Early Invasion Stages.

Although the genetic distribution of introduced raccoons (Procyon lotor) in recent years is well known, few studies have examined their morphometrics, especially the relationships between sex and age in the introduced populations. The aim of this study was to describe the morphological characteristics of raccoons from parts of eastern and western Japan during their early invasion stages, focusing on the relationships between body length and the principal component of craniometrics, with region, sex, age class, body length, and body mass index using a regression model. The body length increased more in males than females and in the older age class, supporting the association with intrasexual selection and competition for food resources. Positive relationships for body length and body mass index were found in craniometric analyses, particularly regarding cranial size components, in addition to age class for both sexes, while cranial size also differed between regions for females. The relationship between body length and craniometrics was inconsistent with that of subspecies originating in North America. Given the sympatric distribution of haplotypes of multiple subspecies without reproductive-isolating barriers in North America and in several introduced areas, hybridization must have occurred prior to the introduction or naturalization of this species.

Mapping the Potential Distribution of Ticks in the Western Kanto Region, Japan: Predictions Based on Land-Use, Climate, and Wildlife.

BACKGROUND: Tick distributions have changed rapidly with changes in human activity, land-use patterns, climate, and wildlife distributions over the last few decades. METHODS: To estimate potential distributions of ticks, we conducted a tick survey at 134 locations in western Kanto, Japan. We estimated the potential distributions of six tick species (Amblyomma testudinarium Koch, 1844; Haemaphysalis flava Neumann, 1897; Haemaphysalis kitaokai Hoogstraal, 1969; Haemaphysalis longicornis Neumann, 1901; Haemaphysalis megaspinosa Saito, 1969; and Ixodes ovatus Neumann, 1899) using MaxEnt modeling based on climate patterns, land-use patterns, and the distributions of five common wildlife species: sika deer (Cervus nippon Temminck, 1838), wild boar (Sus scrofa Linnaeus, 1758), raccoon (Procyon lotor Linnaeus, 1758), Japanese raccoon dog (Nyctereutes procyonoides Gray, 1834), and masked palm civet (Paguma larvata C.E.H. Smith, 1827)). RESULTS: We collected 24,546 individuals of four genera and 16 tick species. Our models indicated that forest connectivity contributed to the distributions of six tick species and that raccoon distribution contributed to five tick species. Other than that, sika deer distribution contributed to H. kitaokai, and wild boar distribution, bamboo forest, and warm winter climate contributed specifically to A. testudinarium. CONCLUSIONS: Based on these results, the dispersal of some tick species toward residential areas and expanded distributions can be explained by the distribution of raccoons and by forest connectivity.

Ecological traps and boosters of ixodid ticks: The differing ecological roles of two sympatric introduced mammals.

The raccoon (Procyon lotor) and masked palm civet (Paguma larvata) are introduced species in Japan and have become abundant in human-inhabited environments. We surveyed tick infestations and tick ingestion by introduced raccoons and masked palm civets captured in Hayama, Kanagawa Prefecture, Japan between November 2018 and January 2020. We collected ticks from the body surface of animals and tick capitula from the gastrointestinal contents. We collected 18,357 ticks identified as Haemaphysalis flava, Haemaphysalis megaspinosa, Haemaphysalis longicornis, Ixodes ovatus, Ixodes tanuki, and Amblyomma testudinarium from 58 of 60 raccoons and 152 ticks, identified as H. flava and I. tanuki, from 16 of 41 masked palm civets. Furthermore, we obtained 16 capitula from 12 % of raccoons and 106 capitula from 63 % of masked palm civets. Raccoons harbored a greater number of ticks (all stages of H. flava and adult I. tanuki) compared with masked palmed civets, whereas the latter species ingested a greater number of nymphal and larval ticks. The results of this study extend our understanding of the ecological roles of two introduced wildlife species. The raccoon may act as an ecological booster, thereby increasing the success rate of bloodmeals and reproduction in ticks. In contrast, the masked palm civet may act as an ecological trap by effectively grooming to remove ticks and prevent bloodmeals.

First glimpse into the origin and spread of the Asian longhorned tick, Haemaphysalis longicornis, in the United States.

Established populations of Asian longhorned ticks (ALT), Haemaphysalis longicornis, were first identified in the United States (US) in 2017 by sequencing the mitochondrial cytochrome c oxidase subunit I (cox1) 'barcoding' locus followed by morphological confirmation. Subsequent investigations detected ALT infestations in 12, mostly eastern, US states. To gain information on the origin and spread of US ALT, we (1) sequenced cox1 from ALT populations across 9 US states and (2) obtained cox1 sequences from potential source populations [China, Japan and Republic of Korea (ROK) as well as Australia, New Zealand and the Kingdom of Tonga (KOT)] both by sequencing and by downloading publicly available sequences in NCBI GenBank. Additionally, we conducted epidemiological investigations of properties near its initial detection locale in Hunterdon County, NJ, as well as a broader risk analysis for importation of ectoparasites into the area. In eastern Asian populations (China/Japan/ROK), we detected 35 cox1 haplotypes that neatly clustered into two clades with known bisexual versus parthenogenetic phenotypes. In Australia/New Zealand/KOT, we detected 10 cox1 haplotypes all falling within the parthenogenetic cluster. In the United States, we detected three differentially distributed cox1 haplotypes from the parthenogenetic cluster, supporting phenotypic evidence that US ALT are parthenogenetic. While none of the source populations examined had all three US cox1 haplotypes, a phylogeographic network analysis supports a northeast Asian source for the US populations. Within the United States, epidemiological investigations indicate ALT can be moved long distances by human transport of animals, such as horses and dogs, with smaller scale movements on wildlife. These results have relevant implications for efforts aimed at minimizing the spread of ALT in the United States and preventing additional exotic tick introductions.

First record of infestation of a pet sloth in Japan with the exotic tick Amblyomma geayi (Acari: Ixodidae).

On December 11, 2018, a single unengorged adult tick was found on the body surface of the trunk of an imported wild-caught Linnaeus's two-toed sloth (Choloepus didactylus) during a routine health check in an animal clinic in Tokyo, Japan. The tick was identified as Amblyomma geayi based on the morphological and molecular characteristics. This is the first case of the introduction of an Amblyomma species to Japan via an imported pet sloth. The present study highlights the current loopholes in Japan's regulatory system for animal imports.

Molecular Characterization of Haemaphysalis Species and a Molecular Genetic Key for the Identification of Haemaphysalis of North America.

Haemaphysalis longicornis (Acari: Ixodidae), the Asian longhorned tick, is native to East Asia, but has become established in Australia and New Zealand, and more recently in the United States. In North America, there are other native Haemaphysalis species that share similar morphological characteristics and can be difficult to identify if the specimen is damaged. The goal of this study was to develop a cost-effective and rapid molecular diagnostic assay to differentiate between exotic and native Haemaphysalis species to aid in ongoing surveillance of H. longicornis within the United States and help prevent misidentification. We demonstrated that restriction fragment length polymorphisms (RFLPs) targeting the 16S ribosomal RNA and the cytochrome c oxidase subunit I (COI) can be used to differentiate H. longicornis from the other Haemaphysalis species found in North America. Furthermore, we show that this RFLP assay can be applied to Haemaphysalis species endemic to other regions of the world for the rapid identification of damaged specimens. The work presented in this study can serve as the foundation for region specific PCR-RFLP keys for Haemaphysalis and other tick species and can be further applied to other morphometrically challenging taxa.

Effects of introduced sika deer (Cervus nippon) and population control activity on the distribution of Haemaphysalis ticks in an island environment.

The effects of introduced mammal species on the ecology of parasites are often under investigated. The sika deeer, Cervus nippon, is host species of many hard ticks. We collected 8348 ticks on an island where sika deer were introduced. The most representative species was Haemaphysalis megaspinosa (n = 4198; 50.3%), followed by H. longicornis (n = 1945; 23.3%), H. cornigera (n = 1179; 14.1%), H. flava (n = 713; 8.5%), Ixodes turdus (n = 289; 3.7%), I. granulatus (n = 22; 0.3%), and H. hystricis (n = 2; <0.1%) on an island where sika deer were introduced. H. megaspinosa and H. hystricis have not previously been recorded on the Izu islands. The high abundance of H. megaspinosa indicated that the tick species may have been introduced with the sika deer. Furthermore, H. megaspinosa larvae were more abundant at collection sites 21-40 days after sika deer were caught by foot snare traps indicate that engorged female of this tick species were forced to drop off in a very limited area near the foot snare trap. This represented a risk for hunters and people associated with wildlife control visiting the area.

Infestation of introduced raccoons (Procyon lotor) with indigenous ixodid ticks on the Miura Peninsula, Kanagawa Prefecture, Japan.

Since the raccoon (Procyon lotor) was introduced to Japan, studies have established that they are infested with native Japanese tick species. However, the quantity of ticks infesting raccoons is unknown. We conducted a survey of ticks on invasive raccoons captured on the Miura Peninsula, Kanagawa Prefecture, Japan, from April 2015 through June 2016 to determine the species of ticks and to quantify the intensity of tick infestation in order to obtain basal information related to the ecology of host-parasite relationships among indigenous tick species and an alien mammalian species. We collected and identified 15,931 ticks of two genera and six species, namely, Haemaphysalis flava, H. megaspinosa, H. longicornis, H. japonica, Ixodes ovatus, and I. tanuki, from 100 out of 115 raccoons. The dominant tick species was H. flava (96.8%) and individuals were mainly adults. Seasonal patterns of infestation intensity of adults and nymphs peaked in the autumn and winter and decreasing in the late spring and summer, May to August, while larvae peaked in August. Our results indicated that host-parasite relationships between invasive raccoons and Japanese tick species, especially H. flava, were established in Kanagawa Prefecture. The ticks infest invasive raccoons for their blood-meal and also for overwintering. The results of this study extend our understanding of the ecology of tick-borne diseases.

Epizootic of sarcoptic mange in raccoon dogs (Nyctereutes procyonoides) in relation to population density.

To examine outbreaks of mange in raccoon dogs (Nyctereutes procyonoides) with respect to population density, we analyzed camera trap videos, and isolated mites from raccoon dog carcasses. In a camera trapping survey, we categorized the skin condition of raccoon dogs, and used a number of independent videos to calculate the relative abundance index (RAI). The RAI of raccoon dogs with alopecia increased following an increase in the RAI of those without alopecia. Among 27 raccoon dog carcasses, 12 showed mange-compatible skin lesions. Sarcoptes scabiei was isolated from 11 of these raccoon dogs, indicating that sarcoptic mange was endemic in our study area. Therefore, a high relative population density may be a factor underlying epizootics of sarcoptic mange in raccoon dogs.