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Background: Species of Trichinella are globally important foodborne parasites infecting a number of domestic and wild vertebrates, including humans. Free-ranging carnivores can act as sentinel species for detection of Trichinella spp. Knowledge of the epidemiology of these parasites may help prevent Trichinella spp. infections in northern Canadian animals and people. Previous research on Trichinella spp. in wildlife from Yukon did not identify risk factors associated with infection, or the diversity and identity of species of Trichinella in regional circulation, based on geographically extensive sampling with large sample sizes., Methods: In a cross-sectional study, we determined the prevalence, infection intensity, risk factors, and species or genotypes of Trichinella in wolverine (Gulo gulo) in two regions of Yukon, Canada, from 2013-2017. A double separatory funnel digestion method followed by mutiplex PCR and PCR-RFLP were used to recover and identify species of Trichinella, respectively., Results: We found larvae of Trichinella in the tongues of 78% (95% CI 73-82) of 338 wolverine sampled. The odds of adult (≥ 2 years) and yearling (1 year) wolverine being Trichinella spp.-positive were four and two times higher, respectively, compared to juveniles (<1 year). The odds of Trichinella spp. presence were three times higher in wolverine from southeast than northwest Yukon. The mean intensity of infection was 22.6 ± 39 (SD, range 0.1-295) larvae per gram. Trichinella T6 was the predominant genotype (76%), followed by T. nativa (8%); mixed infections with Trichinella T6 and T. nativa (12%) were observed. In addition, T. spiralis was detected in one wolverine. Out of 22 isolates initially identified as T. nativa in multiplex PCR, 14 were analyzed by PCR-RFLP to distinguish them from T. chanchalensis, a recently discovered cryptic species, which cannot be distinguished from the T. nativa on multiplex PCR. Ten isolates were identified either as T. chanchalensis alone (n = 7), or mixed infection with T. chanchalensis and T. nativa (n = 2) or T. chanchalensis and Trichinella T6 (n = 1)]., Conclusions: Wolverine hosted high prevalence, high larval intensity, and multiple species of Trichinella, likely due to their scavenging habits, apex position in the food chain, and wide home range. Wolverine (especially adult males) should be considered as a sentinel species for surveys for Trichinella spp. across their distributional range.

Understanding parasite diversity and distribution is essential in managing the potential impact of parasitic diseases in animals and people. Imperfect diagnostic methods, however, may conceal cryptic species. Here, we report the discovery and phylogeography of a previously unrecognized species of Trichinella in wolverine (Gulo gulo) from northwestern Canada that was indistinguishable from T. nativa using the standard multiplex PCR assay based on the expansion segment 5 (ESV) of ribosomal DNA. The novel genotype, designated as T13, was discovered when sequencing the mitochondrial genome. Phylogenetic analyses of the mitochondrial genome and of 15 concatenated single copy orthologs of nuclear DNA indicated a common ancestor for the encapsulated clade is shared by a subclade containing Trichinella spiralis and Trichinella nelsoni, and a subclade containing T13 and remaining taxa: T12 + (T2 + T6) + [(T5 + T9) + (T3 + T8)]. Of 95 individual hosts from 12 species of mammalian carnivores from northwestern Canada from which larvae were identified as T. nativa on multiplex PCR, only wolverines were infected with T13 (14 of 42 individuals). These infections were single or mixed with T. nativa and/or T6. Visual examination and motility testing confirmed that T13 is encapsulated and likely freeze-tolerant. We developed a new Polymerase Chain Reaction-Restriction Fragment Length Polymorphism which unequivocally distinguishes between T13 and T. nativa. We propose Trichinella chanchalensis n. sp. for T13, based on significant genetic divergence from other species of Trichinella and broad-based sampling of the Trichinella genome. Exploration of Alaskan and Siberian isolates may contribute to further resolution of a phylogeographically complex history for species of Trichinella across Beringia, including Trichinella chanchalensis n. sp. (T13)., (Published by Elsevier Ltd.)

Trichinella is an important zoonotic parasite found in a range of wildlife species harvested for food and fur in Canada. We compared larval intensity from tongue and diaphragm, the best predilection sites in other animal species, from naturally infected, wild wolverines (Gulo gulo) (n = 95). Muscle larvae of Trichinella spp. were recovered by the pepsin/HCl artificial digestion method (gold standard) using double separatory funnels, and species were identified using multiplex PCR. Prevalence was 83% (79/95). Of those positive for Trichinella spp. (n = 79), 76 (96.2%) were detected in both tissues, 2 (2.5%) were positive only on diaphragm, and 1 (1.3%) only on tongue. A total of 62 of 79 wolverines (78.5%) had higher larval burden in tongue than in diaphragm, whereas 17 wolverines (21.5%) had higher larval burden in diaphragm. The predilection site (higher larval burden) of Trichinella spp. larvae did not vary significantly between juvenile and adult wolverines (P = 0.2), between male and female wolverines (P = 0.9), and among wolverines classified as having low and high larval intensities overall (P = 0.2). Trichinella T6 was the predominant genotype (63 of 79; 80%), followed by T. nativa (T2) (6 of 79; 8%). Mixed infections of T2 and T6 were observed in 9 of 79 (12%) wolverines. Larval intensity of Trichinella T6 was higher in tongues than diaphragms. No statement can be made for T2 due to insufficient T2 positive samples. In conclusion, tongues are a better site for sampling than diaphragms in future surveys of Trichinella larval intensity in wolverines; however, either tissue is suitable for prevalence studies., (Copyright © 2018. Published by Elsevier B.V.)