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The microsporidian pathogens Vairimorpha apisand V. ceranaeare known to cause intestinal infection in honey bees and are associated with decreased colony productivity and colony loss. The widely accepted method for determining Vairimorphacolony infection level for risk assessment and antibiotic treatment is based on spore counts of 60 pooled worker bees using light microscopy. Given that honey bee colonies consist of as many as 1,000 times more individuals, the number of bees collected for Vairimorphadetection may significantly impact the estimated colony infection level, especially in the case of uneven distribution of high- and low-infected individuals within a hive. Hence, we compared the frequency and severity of Vairimorphainfection in individual bees to pooled samples of 60, 120, and 180 bees, as well as compared the Vairimorphaspp. prevalence in pooled samples of 60 and 180 bees. Overall, we did not find significant differences in spore counts in pooled samples containing incremental numbers of bees, although we observed that, in less-infected colonies, a low frequency of highly infected individuals influenced the estimated colony infection level. Moreover, Vairimorphaspp. prevalence did not differ significantly among the pooled bee samples tested. Increasing the number of pooled bees from the recommended 60 bees to 180 bees did not yield a more accurate representation of colony infection level for highly infected colonies, but the clinical importance of a low frequency of highly infected individuals in less-infected colonies needs to be addressed in future studies.

American foulbrood (AFB) is an infectious disease of honey bee brood caused by the endospore-forming bacterium Paenibacillus larvae. P. larvaespores are resilient in the environment, thus colonies with clinical signs of AFB are often destroyed by burning to eradicate the causative agent. To prevent outbreaks of AFB, oxytetracycline metaphylaxis is widely used in North America, resulting in sustained selective pressure for oxytetracycline resistance in P. larvae. To determine if antimicrobial resistance (AMR) is present among P. larvaeisolates from commercial beekeeping operations in Saskatchewan, Canada, we performed antimicrobial susceptibility testing of 718 P. larvaesamples cultured from pooled, extracted honey collected from 52 beekeepers over a 2-y period, 2019 and 2020. We found that 65 of 718 (9%) P. larvaesamples collected from 8 beekeepers were resistant to oxytetracycline with minimum inhibitory concentration (MIC) values of 64–256 µg/mL. Eight of 718 (1%) samples from 4 beekeepers had intermediate resistance to oxytetracycline (MIC: 4–8 µg/mL). Susceptibility testing for tylosin and lincomycin indicated that P. larvaein Saskatchewan continue to be susceptible to these antimicrobials (tylosin MIC: <1 µg/mL, lincomycin MIC: ≤2 µg/mL). Most oxytetracycline-resistant P. larvaesamples were identified in northeastern Saskatchewan. Whole-genome sequence analysis identified the P. larvae–specific plasmid pMA67with tetracycline-resistance gene tet(L) in 9 of 11 oxytetracycline-resistant P. larvaeisolates sequenced. Our results highlight the advantage of using pooled, extracted honey as a surveillance tool for monitoring AMR in P. larvae.