Using clotted, pelleted blood samples for direct molecular detection of Bartonella spp. in small mammal wildlife surveillance studies.

OBJECTIVE: Bartonella are emerging bacterial zoonotic pathogens. Utilization of clotted blood samples for surveillance of these bacteria in wildlife has begun to supersede the use of tissues; however, the efficacy of these samples has not been fully investigated. Our objective was to compare the efficacy of spleen and blood samples for DNA extraction and direct detection of Bartonella spp. via qPCR. In addition, we present a protocol for improved DNA extraction from clotted, pelleted (i.e., centrifuged) blood samples obtained from wild small mammals. RESULTS: DNA concentrations from kit-extracted blood clot samples were low and A260/A280 absorbance ratios indicated high impurity. Kit-based DNA extraction of spleen samples was efficient and produced ample DNA concentrations of good quality. We developed an in-house extraction method for the blood clots which resulted in apposite DNA quality when compared to spleen samples extracted via MagMAX DNA Ultra 2.0 kit. We detected Bartonella in 9/30 (30.0%) kit-extracted spleen DNA samples and 11/30 (36.7%) in-house-extracted blood clot samples using PCR. Our results suggest that kit-based methods may be less suitable for DNA extraction from blood clots, and that blood clot samples may be superior to tissues for Bartonella detection.

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.