Evaluating approved and alternative treatments against an oxytetracycline-resistant bacterium responsible for European foulbrood disease in honey bees

Abstract

European foulbrood (EFB) is a disease of honey bee larvae caused by Melissococcus plutonius. In North America, oxytetracycline (OTC) is approved to combat EFB disease though tylosin (TYL) and lincomycin (LMC) are also registered for use against American foulbrood disease. Herein, we report and characterize an OTC-resistant M. plutonius isolate from British Columbia, Canada, providing an antimicrobial sensitivity to the three approved antibiotics and studying their abilities to alter larval survival in an in vitro infection model. Specifically, we investigated OTC, TYL, and LMC as potential treatment options for EFB disease using laboratory-reared larvae infected with M. plutonius. The utility of the three antibiotics were compared through an experimental design that either mimicked metaphylaxis or antimicrobial intervention. At varying concentrations, all three antibiotics prevented clinical signs of EFB disease following infection with M. plutonius 2019BC1 in vitro. This included treatment with 100 μg/mL of OTC, a concentration that was ~ 3× the minimum inhibitory concentration measured to inhibit the strain in nutrient broth. Additionally, we noted high larval mortality in groups treated with doses of OTC corresponding to ~ 30× the dose required to eliminate bacterial growth in vitro. In contrast, TYL and LMC were not toxic to larvae at concentrations that exceed field use. As we continue to investigate antimicrobial resistance (AMR) profiles of M. plutonius from known EFB outbreaks, we expect a range of AMR phenotypes, reiterating the importance of expanding current therapeutic options along with alternative management practices to suppress this disease.

Are fungicides a driver of European foulbrood disease in honey bee colonies pollinating blueberries?

“…This observation may be explained by direct bacterial inhibition of the fungicide products on M. plutonius. While the antimicrobial properties of royal jelly have been previously reported to decrease M. plutonius viability in the diet (Takamatsu et al, 2017;Vezeteu et al, 2017;Floyd et al, 2020;Masood et al, 2022), the potential bactericidal effects of these fungicidal products on M. plutonius is unknown, and an area that warrants further investigation.…”

“…Melissococcus plutonius isolate 2019BC1 (Wood et al, 2020;Masood et al, 2022) was used to infect honey bee larvae in vitro. To prepare this isolate for larval infection, 100 μl of thawed liquid culture of 2019BC1, previously stored at −80°C in 20% glycerol, was inoculated into 100 ml of KSBHI liquid media (brain heart infusion, supplemented with 0.15 M KH 2 PO 4 , and 1% soluble starch) and incubated at 37°C under microaerophilic conditions (Arai et al, 2012) for 48 h, shaking at 200 rpm.…”

“…Honey bee pollination is crucial to blueberry production in North America, contributing 90% of the value of Canada's blueberry crops each year (Government of Canada, 2018). Unfortunately, blueberry growers face a shortage of pollination services, in part due to a reported increased incidence of European foulbrood (EFB) disease in blueberry pollinating honey bee colonies (Wardell, 1982;Guarna et al, 2019;Olmstead et al, 2019;Thebeau et al, 2022). The negative economic consequences of EFB outbreaks include lost honey, reduced pollination service revenue, and increased treatment and colony replacement costs (Laate et al, 2020), thereby threatening the continued profitability of the blueberry and beekeeping industries and calling for scientific investigation of the predisposing factors for this disease.…”

“…Previously, colonies with EFB were observed to spontaneously recover when stressors such as inadequate food and water resources were alleviated (Forsgren, 2010); however, recent outbreaks of EFB associated with blueberry pollination have been described as refractory to traditional management practices (Olmstead et al, 2019;Laate et al, 2020;Thebeau et al, 2022). Proposed causes of the increased clinical severity of EFB include highly virulent (Djukic et al, 2018;Grossar et al, 2020;Thebeau et al, 2022) or antimicrobial-resistant strains of M. plutonius (Masood et al, 2022), and environmental factors such as the poor nutritional quality of blueberry pollen (Wardell, 1982;Olmstead et al, 2019); however, the role of pesticide exposure as a predisposing factor for EFB during blueberry pollination has been incompletely explored. For example, several studies have investigated the risk of pesticides on survival of honey bee brood through larval exposure to unrealistically high concentration of active ingredients (Mussen et al, 2004;Wade et al, 2019).…”

Sacbrood viruses and select Lake Sinai virus variants dominated Apis mellifera colonies symptomatic for European foulbrood

“…Honey bees infected with EFB can exhibit high mortality, but disease monitoring is complicated by M. plutonius being present in both symptomatic and asymptomatic colonies (9, 10). Currently, the only approved treatment in the US for EFB is the antibiotic Oxytetracycline, for which antimicrobial-resistant strains have been detected in North American honey bee colonies (11). Not only are better treatments urgently needed, but improved fundamental knowledge of the disease and its progression, in addition to the onset of secondary infections is also necessary to manage this disease.…”