Additional diagnoses of Echinococcus multilocularis in eastern chipmunks (Tamias striatus) from southern Ontario - results from ongoing surveillance for E. multilocularis in intermediate hosts in Ontario, Canada.

Echinococcus multilocularis, a cestode with zoonotic potential, is now known to have a high prevalence in wild canid definitive hosts of southern Ontario. The distribution of E. multilocularis across this region in red foxes (Vulpes vulpes) and coyotes (Canis latrans) is widespread yet heterogenous. In contrast, confirmed diagnoses of E. multilocularis in wild free-ranging intermediate hosts within Ontario are currently limited to a single eastern chipmunk (Tamias striatus). These findings prompted ongoing surveillance efforts in intermediate host species, primarily rodents. Our report describes the results of passive surveillance through wildlife carcass submissions to the Canadian Wildlife Health Cooperative (CWHC) and targeted active sampling of small mammal species from 2018 to 2023; a second and third eastern chipmunk were found to be infected with E. multilocularis. However, these were the only occurrences from surveillance efforts which collectively totaled 510 rodents and other small mammals. Continued surveillance for E. multilocularis in intermediate hosts is of high importance in light of the recent emergence of this parasite in Ontario.

Genomic and transcriptomic characterization of delta SARS-CoV-2 infection in free-ranging white-tailed deer (Odocoileus virginianus).

White-tailed deer (WTD) are susceptible to SARS-CoV-2 and represent an important species for surveillance. Samples from WTD (n = 258) collected in November 2021 from Québec, Canada were analyzed for SARS-CoV-2 RNA. We employed viral genomics and host transcriptomics to further characterize infection and investigate host response. We detected Delta SARS-CoV-2 (B.1.617.2) in WTD from the Estrie region; sequences clustered with human sequences from October 2021 from Vermont, USA, which borders this region. Mutations in the S-gene and a deletion in ORF8 were detected. Host expression patterns in SARS-CoV-2 infected WTD were associated with the innate immune response, including signaling pathways related to anti-viral, pro- and anti-inflammatory signaling, and host damage. We found limited correlation between genes associated with innate immune response from human and WTD nasal samples, suggesting differences in responses to SARS-CoV-2 infection. Our findings provide preliminary insights into host response to SARS-CoV-2 infection in naturally infected WTD.

Factors associated with Giardia infection in dogs in southern Ontario, Canada.

Information concerning risk factors associated with Giardia infection in dogs in southern Ontario, Canada, is currently lacking. This study therefore aimed to identify risk factors for Giardia infection in dogs that visit off-leash dog parks in southern Ontario. From May-November 2018, fecal samples were collected from 466 dogs in 12 off-leash dog parks in the Niagara and Hamilton regions of Ontario. A survey that asked questions pertaining to travel history (i.e., area of residence, locations and regions visited in the previous 6 months), basic medical history (i.e., spay/neuter status, veterinary visits, use of deworming medication), consumption of a raw diet, and the physical (i.e., age, sex, breed) and behavioral characteristics (i.e., off-leash activities, hunting activities) of each dog sampled was administered to the respective owner. All fecal samples were examined with the Giardia plate ELISA (IDEXX Laboratories) for parasite antigen. Multivariable logistic regression analyses were conducted on the survey data to investigate putative risk factors for Giardia infection. Overall, 11.8% (95% CI: 9.2-15.1%) of samples tested positive for Giardia antigen. Results from the multivariable logistic regression analyses identified an interaction between dog age and spay/neuter status that was significantly associated with Giardia infection. The odds of infection were greater in intact as compared to neutered adult dogs (OR: 3.6, 95% CI: 1.7-7.9, p = 0.001), and in neutered juvenile dogs as compared to neutered adults (OR: 5.2, 95% CI: 2.2-12.2, p < 0.001). The results provide veterinarians with evidence-based information for identifying dogs at greatest risk of Giardia infection in southern Ontario.

Divergent SARS-CoV-2 variant emerges in white-tailed deer with deer-to-human transmission.

Wildlife reservoirs of broad-host-range viruses have the potential to enable evolution of viral variants that can emerge to infect humans. In North America, there is phylogenomic evidence of continual transmission of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) from humans to white-tailed deer (Odocoileus virginianus) through unknown means, but no evidence of transmission from deer to humans. We carried out an observational surveillance study in Ontario, Canada during November and December 2021 (n = 300 deer) and identified a highly divergent lineage of SARS-CoV-2 in white-tailed deer (B.1.641). This lineage is one of the most divergent SARS-CoV-2 lineages identified so far, with 76 mutations (including 37 previously associated with non-human mammalian hosts). From a set of five complete and two partial deer-derived viral genomes we applied phylogenomic, recombination, selection and mutation spectrum analyses, which provided evidence for evolution and transmission in deer and a shared ancestry with mink-derived virus. Our analysis also revealed an epidemiologically linked human infection. Taken together, our findings provide evidence for sustained evolution of SARS-CoV-2 in white-tailed deer and of deer-to-human transmission.

Surface and Air Contamination With Severe Acute Respiratory Syndrome Coronavirus 2 From Hospitalized Coronavirus Disease 2019 Patients in Toronto, Canada, March-May 2020.

BACKGROUND: We determined the burden of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in air and on surfaces in rooms of patients hospitalized with coronavirus disease 2019 (COVID-19) and investigated patient characteristics associated with SARS-CoV-2 environmental contamination. METHODS: Nasopharyngeal swabs, surface, and air samples were collected from the rooms of 78 inpatients with COVID-19 at 6 acute care hospitals in Toronto from March to May 2020. Samples were tested for SARS-CoV-2 ribonucleic acid (RNA), cultured to determine potential infectivity, and whole viral genomes were sequenced. Association between patient factors and detection of SARS-CoV-2 RNA in surface samples were investigated. RESULTS: Severe acute respiratory syndrome coronavirus 2 RNA was detected from surfaces (125 of 474 samples; 42 of 78 patients) and air (3 of 146 samples; 3 of 45 patients); 17% (6 of 36) of surface samples from 3 patients yielded viable virus. Viral sequences from nasopharyngeal and surface samples clustered by patient. Multivariable analysis indicated hypoxia at admission, polymerase chain reaction-positive nasopharyngeal swab (cycle threshold of ≤30) on or after surface sampling date, higher Charlson comorbidity score, and shorter time from onset of illness to sampling date were significantly associated with detection of SARS-CoV-2 RNA in surface samples. CONCLUSIONS: The infrequent recovery of infectious SARS-CoV-2 virus from the environment suggests that the risk to healthcare workers from air and near-patient surfaces in acute care hospital wards is likely limited.

SARS-CoV-2 wildlife surveillance in Ontario and Québec.

Background: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the virus responsible for the coronavirus disease 2019 pandemic, is capable of infecting a variety of wildlife species. Wildlife living in close contact with humans are at an increased risk of SARS-CoV-2 exposure and, if infected, have the potential to become a reservoir for the pathogen, making control and management more difficult. The objective of this study is to conduct SARS-CoV-2 surveillance in urban wildlife from Ontario and Québec, increasing our knowledge of the epidemiology of the virus and our chances of detecting spillover from humans into wildlife. Methods: Using a One Health approach, we leveraged activities of existing research, surveillance and rehabilitation programs among multiple agencies to collect samples from 776 animals from 17 different wildlife species between June 2020 and May 2021. Samples from all animals were tested for the presence of SARS-CoV-2 viral ribonucleic acid, and a subset of samples from 219 animals across three species (raccoons, Procyon lotor; striped skunks, Mephitis mephitis; and mink, Neovison vison) were also tested for the presence of neutralizing antibodies. Results: No evidence of SARS-CoV-2 viral ribonucleic acid or neutralizing antibodies was detected in any of the tested samples. Conclusion: Although we were unable to identify positive SARS-CoV-2 cases in wildlife, continued research and surveillance activities are critical to better understand the rapidly changing landscape of susceptible animal species. Collaboration between academic, public and animal health sectors should include experts from relevant fields to build coordinated surveillance and response capacity.

Prevalence of intestinal parasites in dogs in southern Ontario, Canada, based on fecal samples tested using sucrose double centrifugation and Fecal Dx® tests.

In southern Ontario, Canada, there is a lack of information concerning the prevalence of intestinal parasites in dogs. As such, this study aimed to characterize the prevalence of intestinal parasites in dogs visiting off-leash parks in the region using sucrose double centrifugation and Fecal Dx® tests. Additionally, data obtained via the sucrose double centrifugation method were used to evaluate the performance of the Fecal Dx® tests. Fecal samples were collected from 466 dogs aged ≥6 months from May to November 2018 (mean age = 3.7 years). Overall, eleven intestinal parasites were identified using sucrose double centrifugation. Roundworm eggs (Toxocara canis and Baylisascaris procyonis), hookworm eggs (Ancylostoma caninum and Uncinaria stenocephala), and whipworm eggs (Trichuris vulpis) were identified in 1.07% (95% confidence interval [CI] 0.38-2.56%), 5.79% (95% CI 3.85-8.31%), and 5.15% (95% CI 3.33-7.57) of samples, respectively. Using the Fecal Dx® tests, 1.07% (95% CI 0.38-2.56%), 4.29% (95% CI 2.64-6.55%), and 2.15% (95% CI 1.03-3.91) of the samples tested positive for roundworm, hookworm, and whipworm antigen, respectively. To assess the level of agreement between the Fecal Dx® tests and sucrose double centrifugation, three methods were used. Cohen's kappa indicated a fair-to-moderate level of agreement between Fecal Dx® tests and sucrose double centrifugation. In contrast, the prevalence-adjusted bias-adjusted kappa and Gwet's first-order agreement coefficient indicated almost perfect agreement between these tests, ranging from 0.87 to 0.99 among the parasites examined. This study provides valuable information on the prevalence of intestinal parasites in mature dogs in southern Ontario that will help guide parasite control recommendations for dogs in this region.

Investigation of the occurrence of Angiostrongylus vasorum in coyotes in southern Ontario, Canada.

In North America, the only endemic focus for Angiostrongylus vasorum (French heartworm) was historically thought to occur in the southeastern part of the island of Newfoundland. However, reports of A. vasorum infection in wild canids in West Virginia, USA, and Nova Scotia, Canada, suggest the introduction of the parasite to mainland North America. We screened for A. vasorum in coyotes from across southern Ontario. Additionally, we evaluated the performance of ELISAs for detection of circulating A. vasorum antigen (Ag-ELISA) and antibodies against A. vasorum (Ab-ELISA) designed for use in sera or blood of foxes for use with coyotes in this region. Autopsies were performed on 397 coyotes, and lung tissue extract prepared from each carcass was tested via both ELISAs. The sensitivity and specificity for both tests were estimated in the absence of a gold standard using a 2-test single population Bayesian model; sensitivity and specificity priors were based on the performance of the assays in foxes in Switzerland. Eight coyotes tested positive for A. vasorum antigen; no animal was antibody positive. The estimated sensitivity and specificity of the Ag-ELISA were 90.8% (95% credible interval [CrI]: 83.8-95.6%) and 95.5% (95% CrI: 93.4-97.2%), respectively. For the Ab-ELISA, the estimated sensitivity and specificity were 41.9% (95% CrI: 32.1-51.9%) and 98.0% (95% CrI: 96.3-99.0%), respectively. Based on these findings and negative postmortem data for the same animals, there is insufficient evidence to suggest the presence of A. vasorum in southern Ontario coyotes.

Evaluation of the prevalence of Echinococcus multilocularis in dogs that visit off-leash dog parks in southern Ontario, Canada.

Prior to 2012, Echinococcus multilocularis was not known to occur in any host in Ontario, Canada. However, since that year, five cases of alveolar echinococcosis have been diagnosed in dogs that resided at the western end of Lake Ontario. In addition, E. multilocularis has been shown to be a common infection in wild canids (i.e. coyotes and foxes) across southern Ontario with a high-risk infection cluster in the area surrounding the western shores of Lake Ontario and northern shores of Lake Erie. In regions endemic for E. multilocularis, dog ownership is considered a risk factor for human alveolar echinococcosis. A study was therefore carried out to determine the prevalence of E. multilocularis intestinal infections in dogs within the high-risk infection cluster. From May to November 2018, faecal samples were collected from 477 dogs aged ≥6 months that visited 12 off-leash dog parks in the Halton, Hamilton and Niagara public health units. Faecal samples were analysed via a magnetic capture probe DNA extraction and real-time PCR method for E. multilocularis DNA. Overall, 0% (97.5% CI: 0%-0.80%) of samples tested positive. This result informs preventive recommendations for E. multilocularis infections in dogs in this region.

Evaluation of the SNAP® 4Dx® plus test for the detection of Dirofilaria immitis antigen and characterization of exposure to tick-borne pathogens in wild canids in southern Ontario.

Population-level surveys for Dirofilaria immitis in wild canids typically rely on identification of the parasite at necropsy. More recently, some studies have employed the use of the SNAP® 4Dx® Plus test. However, since the assay was designed for use with domestic dogs it needs to be validated for use with wild canids for accurate interpretation of results. We therefore evaluated the performance of the SNAP® 4Dx® Plus test for detection of D. immitis in wild canids in southern Ontario. Overall, 199 wild canid carcasses were collected from across the region and assessed for the presence of D. immitis parasites at necropsy; ten were infected. Lung tissue extract (LE) and thoracic fluid filter paper extract (TFE) prepared from each wild canid were tested via the SNAP® 4Dx® Plus test, which simultaneously tests for the presence of D. immitis antigen and antibodies to Borrelia burgdorferi, Anaplasma spp., and Ehrlichia spp. The prevalence adjusted bias adjusted kappa (PABAK) and Gwet's first-order-agreement coefficient (AC1) were used to assess the level of agreement between sample pairs. The PABAK and AC1 between LF and TFE applied to the SNAP® 4Dx® Plus test and the necropsy-confirmed D. immitis status indicated a very high level of agreement for all sample pairs. Compared to necropsy-confirmed D. immitis status, the estimated sensitivity and specificity of the SNAP® 4Dx® Plus test for D. immitis antigen in LE was 80 % (95 % CI 44.4-97.5%) and 98.9 % (95 % CI 96.2-99.9%), respectively. For the TFE, the sensitivity was 70 % (95 % CI 34.8-93.3%) and the specificity was 97.9 % (95 % CI 94.7-99.4%). With respect to the tick-borne pathogen components, 1.5 % (3/199; 95 % CI 0.3-4.5%) of wild canids tested positive for B. burgdorferi antibody; 1.0 % (2/199; 95 % CI 0-3.8%) of LE samples were positive and 0.5 % (1/199; 95 % CI 0-3.1%) of TFE samples were positive. No samples tested positive for antibody to Anaplasma spp. (95 % CI 0-2.3%) or Ehrlichia spp. (95 % CI 0-2.3%). Collectively, the results suggest that the SNAP® 4Dx® Plus test may be a suitable test for use with LE and TFE for the detection of D. immitis antigen in wild canids from southern Ontario.

Factors associated with Echinococcus multilocularis infection in coyotes in southern Ontario.

Echinococcus multilocularis was recently reported in wild canids across southern Ontario, a newly recognized endemic area in Canada. In such areas, a comprehensive understanding of factors associated with infection in definitive hosts (wild canids) is critical for mitigating risk of transmission to humans. However, little is known about the transmission dynamics of the parasite in definitive hosts for this region. A study was therefore carried out to investigate the association of host-level (sex, body condition), environmental (southern Ontario region, land cover), temporal (season, hunting season, calendar year) and extraneous factors (submitter type) with E. multilocularis infection in coyotes in southern Ontario. Between November 2015 and March 2017, 416 coyotes were collected from across the region as part of a study that investigated the prevalence and distribution of the parasite in wild canids; approximately 24% of coyotes were positive for E. multilocularis. Associations between infection and factors of interest were assessed via a mixed-effects logistic regression model with a random intercept for submitter to account for clustering. Coyotes with poor body condition were at greater odds of E. multilocularis infection than those in good condition (odds ratio [OR] 2.14; 95% CI: 1.08-4.26; p = .030). A negative association was observed between infection in coyotes and the proportion of natural land in a coyote's estimated home range (OR: 0.67; 95% CI: 0.52-0.85; p = .001). Coyotes from the western region of southern Ontario had lower odds of infection compared to coyotes from the central region (OR: 0.26; 95% CI: 0.12-0.55; p < .001). These results can be used to help guide future public health prevention strategies for human alveolar echinococcosis.

Prevalence and distribution of Dirofilaria immitis infection in wild canids in southern Ontario.

Wild canids represent a potential reservoir host for Dirofilaria immitis infection in dogs in Ontario. Since wild canids are not protected by chemoprophylaxis, understanding the epidemiology of D. immmitis in these populations may help elucidate the background risk of infection for dogs. The aim of this study was to determine the prevalence and distribution of D. immitis infection in wild canids across southern Ontario. From February 2016 to March 2017, 290 wild canid carcasses (273 coyotes and 17 foxes) were collected from across the region and assessed for the presence of D. immitis at the time of necropsy. Overall, D. immitis infection was identified in 4.8% (95% CI 2.8-8.0%) of these wild canid carcasses. Among coyotes, 5.1% (95% CI 3.0-8.5%) were positive; no evidence of D. immitis was found in the 17 foxes. Dirofilaria immitis infections in wild canids were detected in two regions of southern Ontario: 12 of the 14 D. immitis infections were detected in the south-western region and two were detected in the eastern region. Our findings provide preliminary insights into the prevalence and geographical distribution of D. immitis in coyotes and foxes in southern Ontario.

Echinococcus multilocularis Infection, Southern Ontario, Canada.

Alveolar echinococcosis, the disease caused by infection with the intermediate stage of the Echinococcus multilocularis tapeworm, is typically fatal in humans and dogs when left untreated. Since 2012, alveolar echinococcosis has been diagnosed in 5 dogs, 3 lemurs, and 1 chipmunk in southern Ontario, Canada, a region previously considered free of these tapeworms. Because of human and animal health concerns, we estimated prevalence of infection in wild canids across southern Ontario. During 2015-2017, we collected fecal samples from 460 wild canids (416 coyotes, 44 foxes) during postmortem examination and analyzed them by using a semiautomated magnetic capture probe DNA extraction and real-time PCR method for E. multilocularis DNA. Surprisingly, 23% (95% CI 20%-27%) of samples tested positive. By using a spatial scan test, we identified an infection cluster (relative risk 2.26; p = 0.002) in the western-central region of the province. The cluster encompasses areas of dense human population, suggesting zoonotic transmission.

Echinococcus multilocularis in a wild free-living eastern chipmunk (Tamias striatus) in Southern Ontario: A case report and subsequent field study of wild small mammals.

Southern Ontario has recently been identified as a risk area for Echinococcus multilocularis, based on surveys of foxes (Vulpes vulpes) and coyotes (Canis latrans) which act as definitive hosts of the parasite. In this manuscript, we describe the first detection of E. multilocularis in an eastern chipmunk (Tamias striatus) in North America. This case, submitted to the Canadian Wildlife Health Cooperative (CWHC) in August 2016 as part of ongoing wildlife disease surveillance activities, represents the first report of alveolar echinococcosis in a wild, free living, intermediate host from southern Ontario, providing further evidence of an established sylvatic cycle of E. multilocularis in this region. The finding prompted a field investigation to identify additional cases of alveolar echinococcosis in small mammals in the summer of 2017. Echinococcus multilocularis was not detected in any of the 196 small mammals submitted to the CWHC from across southern Ontario or in any of the 43 small mammals trapped in the area where the infected chipmunk was found. However, given the suspected low prevalence and patchy distribution of E. multilocularis in small mammals, our negative results do not preclude the established presence of the parasite. This case emphasizes the importance of passive surveillance networks for monitoring new and emerging diseases in wildlife populations.