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Whole genome sequencing (WGS) is a widely available, inexpensive means of providing a wealth of information about an organism’s diversity and evolution. However, WGS for many pathogenic bacteria remain limited because they are difficult, slow and/or dangerous to culture. To avoid culturing, metagenomic sequencing can be performed directly on samples, but the sequencing effort required to characterize low frequency organisms can be expensive. Recently developed methods for selective whole genome amplification (SWGA) can enrich target DNA to provide efficient sequencing. We amplified Coxiella burnetii (a bacterial select agent and human/livestock pathogen) from 3 three environmental samples that were overwhelmed with host DNA. The 68- to 147-fold enrichment of the bacterial sequences provided enough genome coverage for SNP analyses and phylogenetic placement. SWGA is a valuable tool for the study of difficult-to-culture organisms and has the potential to facilitate high-throughput population characterizations as well as targeted epidemiological or forensic investigations.

In eastern North America, the blacklegged tick, Ixodes scapularis, is the vector for Borrelia burgdorferi sensu stricto and Anaplasma phagocytophilum, the causal agents for human and canine Lyme disease and anaplasmosis, respectively. The extensive range expansion of I. scapularis in Ontario is a growing veterinary and public health concern. However, there is limited information on the risk factors associated with I. scapularis carriage on dogs. Within an emerging area for Lyme disease risk in southeastern Ontario, we identified the tick species carried by dogs; determined the prevalence of B. burgdorferi and A. phagocytophilum in I. scapularis; and examined associations between pet demographics, travel history, and geographical location and the odds of: a dog carrying I. scapularis relative to other tick species (i.e., case-case design), and a removed I. scapularis being infected with B. burgdorferi. Seven species of ticks were collected from 543 companion dogs at 20 participating veterinary hospitals from April to December 2015. Ixodes scapularis were detected on 85.6% of parasitized dogs, and 7.5% of these dogs were carrying at least one B. burgdorferi-positive tick. Based on a multivariable logistic regression model, the odds of I. scapularis infestation relative to other tick species was significantly higher in fall and spring compared to summer, with closer proximity to Lake Ontario, in female compared to male dogs, in dogs weighing over 30 kg compared to lighter dogs, and in dogs that had not visited a farm in the 7 days prior to tick removal. Based on univariable exact logistic regression models, the odds of B. burgdorferi-positive I. scapularis carriage relative to B. burgdorferi-negative I. scapularis were significantly higher for dogs that traveled in the 14 days prior to tick removal, for those dogs under 1 year of age, and for those weighing less than 10 kg. This case-case study provides information for veterinarians and public health practitioners to help protect dogs and their owners from Lyme disease in southeastern Ontario.

Lyme disease is an emerging public health threat in Ontario, Canada due to ongoing range expansion of the tick vector, Ixodes scapularis. Tick density is an important predictor of human Lyme disease risk and is typically measured using active tick surveillance via drag sampling, which is time and resource-intensive. New cost-effective tools are needed to augment current surveillance activities. Our objective was to evaluate the ability of a maximum entropy (Maxent) species distribution model to predict I. scapularis density in three regions of Ontario - Ottawa, Kingston, and southern Ontario - in order to determine its utility in predicting the public health risk of Lyme disease. Ticks were collected via drag sampling at 60 sites across the three regions. Model-predicted habitat suitability was calculated from a previously constructed Maxent model as the mean predicted habitat suitability within a 1-km radius of each site. Spearman's correlation coefficient was used to quantify the continuous relationship between model-predicted habitat suitability and tick density, and negative binomial regression was used to quantify the relationship between tick density and model-predicated habitat suitability. Spearman's correlation coefficients for the full study area, Kingston region, and Ottawa region were 0.517, 0.707, and 0.537, respectively, indicating a moderate positive relationship and ability of the model to predict tick density. Regression analysis further demonstrated a significant positive association between tick density and model-predicted habitat suitability (p0.001). Using a dichotomized measure of model-predicted habitat suitability, the incidence rate ratio - the ratio of ticks per m

Traditionally considered an agent affecting domestic dogs, canine distemper virus (CDV) is now well known for an ability to infect a broad range of hosts. In Ontario, domestic dogs are routinely vaccinated and clinical disease attributed to CDV infection in this population is infrequent. CDV has been regularly documented in Ontario wildlife spanning at least 4 decades however, the molecular identity of circulating CDV strains is currently unknown. Our objective was to investigate the molecular identities of and genetic relationships between CDV detected in wild and domestic animals from Canada, across multiple host species and over time. Samples were opportunistically collected from submissions to the Ontario-Nunavut node of the Canadian Wildlife Health Cooperative and the Animal Health Laboratory in Guelph, Ontario. RT-PCR was used to confirm CDV diagnosis, and the hemagglutinin gene was sequenced. Phylogenetic relationships were inferred, and the geographic distribution of clades was visualized using a geographic information system . Phenetic relationships between sequences were investigated with a median joining network analysis and through mixed multivariable linear regression. CDV sequences from ten wild and domestic species were characterized into seven lineages, that overlapped geographically and temporally. The predominant lineage circulating in Ontario wildlife, denoted Canada-1, has not been previously described to the authors knowledge. Our analysis indicates that the Canada-1 lineage is most genetically similar to America-1 sequences, however according to current methodology represents a distinct lineage. Multiple co-circulating CDV lineages were also identified, and raccoons appear to play an important role in the maintenance and transmission of these heterogeneous lineages in Ontario. This study also confirmed the presence of CDV from a lineage not found to be circulating in Ontario wildlife, in a domestic dog imported into Ontario from South America. Therefore, travel and the trade of animals may be an important avenue for the introduction of novel CDV lineages. It remains unclear whether and to what extent the genetic heterogeneity identified poses a risk to the efficacy of current vaccines. Increasing viral activity and continued antigenic drift resulting in partial protection or vaccine failure remains a concern.

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.

Wild birds that forage around livestock facilities have been implicated as vectors of antimicrobial resistant organisms. Although antimicrobial resistant bacteria have been isolated from European starlings (Sturnus vulgaris), their role in the dissemination of antimicrobial resistant elements in livestock facilities needs further investigation. To determine whether on-farm starling density and other factors were associated with the presence of cefotaxime and ciprofloxacin resistant E. coli among dairy cows in Ohio, bovine fecal pats from 150 farms were tested for the presence of cefotaxime and ciprofloxacin resistant E. coli. Each farm was visited twice (during the summer and fall of 2007-2009). Multi-level logistic regression models with a random intercept to account for fecal pats collected within a specific visit to a farm were used to assess the associations. The percentage of samples with cefotaxime and ciprofloxacin resistant E. coli was 13.4% and 13.6%, respectively. The percentage of farms having at least one sample testing positive for cefotaxime and ciprofloxacin resistant E. coli was 56.7% and 48.7%, respectively. The odds of detecting cefotaxime and ciprofloxacin resistant E. coli in the samples was significantly higher in 2007 compared to 2008 and 2009, in fall compared to summer, and from farms closer than 60km to starling night roost sites compared to the farms further than 60km. The presence of starlings during the day had a negative association with the likelihood of detecting cefotaxime resistant E. coli. Presence of calves also had a negative association with the likelihood of detecting both cefotaxime and ciprofloxacin resistant E. coli. European starlings might play a role in the dissemination of antimicrobial resistant agents in livestock facilities related to their daily population movements rather than the specific density of birds on farm during the day.

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.

Raccoon latrines represent sites of potential infection by the zoonotic parasite Baylisascaris procyonis for wildlife and humans. Our objective was to determine the prevalence of B. procyonis at raccoon latrine sites in southern Ontario. Thirty raccoon latrines were sampled between June – July 2018; multiple scats were collected and homogenized to form a representative sample of each latrine. To determine the presence or absence of B. procyonis eggs in each sample, we used the Cornell-Wisconsin centrifugal floatation technique. Twenty-three percent (7/30) of homogenized samples tested positive for B. procyonis. Eggs per gram of feces ranged from 1 to 388 (median = 1.28, IQR = 0.32–232.5). Baylisascaris procyonis positive latrines were found in conservation areas heavily used by people, which may represent a possible source of exposure for humans in these areas.

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.