Zabrodski MW, Epp T, Wilson G, Moshynskyy I, Sharafi M, Reitsma L, Castano Ospina M, DeBruyne JE, Wentzell A, Wood SC, Kozii IV, Klein CD, Thebeau J, Sobchishin L, Ruzzini AC, Simko E. "Establishment of apiary-level risk of American foulbrood through the detection of Paenibacillus larvae spores in pooled, extracted honey in Saskatchewan." Scientific Reports. 2022 May 25;12(1):8848. doi: 10.1038/s41598-022-12856-4. 

Abstract

Paenibacillus larvae, the causative agent of American foulbrood (AFB), produces spores that may be detectable within honey. We analyzed the spore content of pooled, extracted honey from 52 large-scale (L) and 64 small-scale (S) Saskatchewan beekeepers over a two-year period (2019–2020). Our objectives were: (i) establish reliable prognostic reference ranges for spore concentrations in extracted honey to determine future AFB risk at the apiary level; (ii) identify management practices as targets for mitigation of risk. P. larvae spores were detected in 753 of 1476 samples (51%). Beekeepers were stratified into low (< 2 spores/gram), moderate (2- < 100 spores/gram), and high (≥ 100 spores/gram) risk categories. Of forty-nine L beekeepers sampled in 2019, those that reported AFB in 2020 included 0/26 low, 3/18 moderate, and 3/5 high risk. Of twenty-seven L beekeepers sampled in 2020, those that reported AFB in 2021 included 0/11 low, 2/14 moderate, and 1/2 high risk. Predictive modelling included indoor overwintering of hives, purchase of used equipment, movement of honey-producing colonies between apiaries, beekeeper demographic, and antimicrobial use as risk category predictors. Saskatchewan beekeepers with fewer than 2 spores/gram in extracted honey that avoid high risk activities may be considered at low risk of AFB the following year.

Masood, F., Thebeau, J.M., Cloet, A. et al. Evaluating approved and alternative treatments against an oxytetracycline-resistant bacterium responsible for European foulbrood disease in honey bees. Sci Rep 12, 5906 (2022). https://doi.org/10.1038/s41598-022-09796-4

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.

Zabrodski MW, DeBruyne JE, Wilson G, Moshynskyy I, Sharafi M, Wood SC, Kozii IV, Thebeau J, Klein CD, Medici de Mattos I, Sobchishin L, Epp T, Ruzzini AC, Simko E. "Comparison of individual hive and apiary-level sample types for spores of Paenibacillus larvae in Saskatchewan honey bee operations." PLoS One. 2022 Feb 7;17(2):e0263602. doi: 10.1371/journal.pone.0263602. 

Abstract

Kozii IK, Wood SC, Koziy RV, Simko E. 2022. "Histomorphological description of the reproductive system in mated honey bee queens." Journal of Apicultural Research, 61:1, 114-126, DOI: 10.1080/00218839.2021.1900636

The accelerating decline of pollinator populations has become increasingly concerning in the last two decades. Honey bees are economically important pollinators and a model species to evaluate pollinator health. The decline in queen quality is one of the frequently reported causes associated with honey bee losses worldwide. Histopathology is an essential tool used for diagnostics and research in mammalian species and may provide insight into the histomorphological basis of a decline in queen quality. Thus, the purpose of this study was to summarize the previously described normal morphology of the entire reproductive tract of mated honey bee queens and to illustrate it by high quality photomicrographs. Accordingly, reproductive tracts of one-year old mated honey bee queens were processed for histology, serially sectioned, stained with hematoxylin and eosin, and used for the capture of high quality histological photomicrographs of the entire reproductive tract. This study illustrates the microscopic morphology of the queen reproductive tract which may facilitate further investigations of the cause and pathogenesis of the recent decline in reproductive fitness of honey bees using histopathology.

Kozii IV, Barnsley S, Silva MCBD, Wood SC, Klein CD, de Mattos IM, Zabrodski MW, Silva RCM, Fabela CIO, Guillemin L, Dvylyuk I, Ferrari MCO, Simko E. "Reproductive fitness of honey bee queens exposed to thiamethoxam during development." Veterinary Pathology. 2021 Nov;58(6):1107-1118. doi: 10.1177/03009858211031845. Epub 2021 Jul 16. 

Abstract

The productivity and survival of honey bee (Apis mellifera) colonies depend on queen bee health. Colony-level neonicotinoid exposure has negative effects on reproductive fitness of honey bee queens. However, it is unclear if the observed effects are a direct outcome of neonicotinoid toxicity or result from suboptimal care of developing queens by exposed workers. The aim of this study was to evaluate larval survival, reproductive fitness, and histopathology of honey bee queens exposed to incremental doses (0, 5, 50 ng) of the neonicotinoid thiamethoxam (THI) applied directly to individual late larvae (7 days post-oviposition) of queens. The 5 ng dose represents a calculated high environmental level of exposure for honey bee queen larvae. Morphometric evaluation revealed that the total area of mandibular gland epithelium in queens exposed to 5 and 50 ng THI was reduced by 14% (P = .12) and 25% (P = .001), respectively. Decreased mandibular gland size may alter pheromone production, which could in part explain previously observed negative effects of THI on the reproductive fitness of queens. We also found that late larval exposure to THI reduced larval and pupal survival and decreased sperm viability in mated queens. These changes may interfere with queen development and reproductive longevity.

Klein CD, Kozii IV, Wood SC, Koziy RV, Zabrodski MW, Dvylyuk I, de Mattos IM, Moshynskyy I, Honaramooz A, Simko E. "Testicular changes of honey bee drones, Apis mellifera (Hymenoptera: Apidae), during sexual maturation." Journal of Insect Science. 2021 Nov 1;21(6):3. doi: 10.1093/jisesa/ieab049. 

Abstract

The normal developmental anatomy and histology of the reproductive tract of the honey bee drone, Apis mellifera (Linnaeus, 1758), has been well documented. The post-emergence maturation changes of the accessory glands are likewise well understood, but the normal histological changes of the testicle undergoing physiologic atrophy are not well characterized. To address this knowledge gap, herein we describe the anatomy and sequential histological stages of normal testicular atrophy of drones sampled daily from emergence to sexual maturity in the spring (June) and early summer (July). Testicular histological changes during maturation are characterized by the following stages: I) conclusion of spermiogenesis; II) evacuation of spermatodesms from tubular lumens; III) progressive follicular cell atrophy, and IV) complete atrophy and collapse of testicular parenchyma. Tubular changes occur in a basilar to apical direction where segments closer to the vas deferens are histologically more mature than corresponding apical segments. In addition, the rate of testicular maturation was found to change with seasonal progression. This description of physiologic testicular atrophy should be useful for future studies investigating potential pathological effects of stressors on drone testes during sexual maturation.

Wood SC, Chalifour JC, Kozii IV, Medici de Mattos I, Klein CD, Zabrodski MW, Moshynskyy I, Guarna MM, Wolf Veiga P, Epp T, Simko E. "In vitro effects of pesticides on European foulbrood in honeybee larvae." Insects. 2020 Apr 17;11(4):252. doi: 10.3390/insects11040252. 

Abstract

Neonicotinoid and fungicide exposure has been linked to immunosuppression and increased susceptibility to disease in honeybees (Apis mellifera). European foulbrood, caused by the bacterium Melissococcus plutonius, is a disease of honeybee larvae which causes economic hardship for commercial beekeepers, in particular those whose colonies pollinate blueberries. We report for the first time in Canada, an atypical variant of M. plutonius isolated from a blueberry-pollinating colony. With this isolate, we used an in vitro larval infection system to study the effects of pesticide exposure on the development of European foulbrood disease. Pesticide doses tested were excessive (thiamethoxam and pyrimethanil) or maximal field-relevant (propiconazole and boscalid). We found that chronic exposure to the combination of thiamethoxam and propiconazole significantly decreased the survival of larvae infected with M. plutonius, while larvae chronically exposed to thiamethoxam and/or boscalid or pyrimethanil did not experience significant increases in mortality from M. plutonius infection in vitro. Based on these results, individual, calculated field-realistic residues of thiamethoxam and/or boscalid or pyrimethanil are unlikely to increase mortality from European foulbrood disease in honeybee worker brood, while the effects of field-relevant exposure to thiamethoxam and propiconazole on larval mortality from European foulbrood warrant further study.

Zabrodski MW, Wilson G, Moshynskyy I, Wentzell A, Wood SC, Klein CD, Kozii IV, de Mattos IM, Epp T, Simko E. "Investigation of clinical outbreaks of American foulbrood in honey-bee operations in Saskatchewan." Canadian Veterinary Journal. 2020 Oct;61(10):1055-1059. PMID: 33012819; PMCID: PMC7488369.

Abstract

Four outbreaks of American foulbrood were investigated in honey-bee operations in Saskatchewan during the summer of 2019. Clinical signs were confirmed by the Saskatchewan Provincial Specialist in Apiculture and the causative agent was cultured and identified through matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS). Evaluation of management practices revealed off-label metaphylactic use of oxytetracycline in 3 of 4 operations and a discontinuation of antibiotic use in the fourth. Recent regulatory changes regarding access to medically important antimicrobials has provided an opportunity for veterinarians to promote evidence-based use of antimicrobials in apiculture while safe-guarding the health of commercial honeybee populations and the economic viability of their producers.

Wood SC, de Mattos IM, Kozii IV, Klein CD, Dvylyuk I, Folkes CDA, de Carvalho Macedo Silva R, Moshynskyy I, Epp T, Simko E. 2020. "Effects of chronic dietary thiamethoxam and prothioconazole exposure on Apis mellifera worker adults and brood." Pest. Manag. Sci., 76: 85-94. https://doi.org/10.1002/ps.5501

Abstract

Koziy RV, Wood SC, Kozii IV, van Rensburg CJ, Moshynskyy I, Dvylyuk I, Simko E. "Deformed wing virus infection in honey bees (Apis mellifera L.)." Vet Pathol. 2019 Jul;56(4):636-641. doi: 10.1177/0300985819834617. Epub 2019 Mar 11. PMID: 30857499.

Abstract

Deformed wing virus (DWV) is a single-stranded RNA virus of honey bees (Apis mellifera L.) transmitted by the parasitic mite Varroa destructor. Although DWV represents a major threat to honey bee health worldwide, the pathological basis of DWV infection is not well documented. The objective of this study was to investigate clinicopathological and histological aspects of natural DWV infection in honey bee workers. Emergence of worker honey bees was observed in 5 colonies that were clinically affected with DWV and the newly emerged bees were collected for histopathology. DWV-affected bees were 2 times slower to emerge and had 30% higher mortality compared to clinically normal bees. Hypopharyngeal glands in bees with DWV were hypoplastic, with fewer intracytoplasmic secretory vesicles; cells affected by apoptosis were observed more frequently. Mandibular glands were hypoplastic and were lined by cuboidal epithelium in severely affected bees compared to tall columnar epithelium in nonaffected bees. The DWV load was on average 1.7 × 10⁶ times higher (P < .001) in the severely affected workers compared to aged-matched sister honey bee workers that were not affected by deformed wing disease based on gross examination. Thus, DWV infection is associated with prolonged emergence, increased mortality during emergence, and hypoplasia of hypopharyngeal and mandibular glands in newly emerged worker honey bees in addition to previously reported deformed wing abnormalities.

Wood SC, Kozii IV, Medici de Mattos I, de Carvalho Macedo Silva R, Klein CD, Dvylyuk I, Moshynskyy I, Epp T, Simko E. "Chronic high-dose neonicotinoid exposure decreases overwinter survival of Apis mellifera L." Insects. 2019 Dec 31;11(1):30. doi: 10.3390/insects11010030. 

Abstract

Overwinter colony mortality is an ongoing challenge for North American beekeepers. During winter, honey bee colonies rely on stored honey and beebread, which is frequently contaminated with the neonicotinoid insecticides clothianidin and thiamethoxam. To determine whether neonicotinoid exposure affects overwinter survival of Apis mellifera L., we chronically exposed overwintering field colonies and winter workers in the laboratory to thiamethoxam or clothianidin at different concentrations and monitored survival and feed consumption. We also investigated the sublethal effects of chronic thiamethoxam exposure on colony pathogen load, queen quality, and colony temperature regulation. Under field conditions, high doses of thiamethoxam significantly increased overwinter mortality compared to controls, with field-realistic doses of thiamethoxam showing no significant effect on colony overwinter survival. Under laboratory conditions, chronic neonicotinoid exposure significantly decreased survival of winter workers relative to negative control at all doses tested. Chronic high-dose thiamethoxam exposure was not shown to impact pathogen load or queen quality, and field-realistic concentrations of thiamethoxam did not affect colony temperature homeostasis. Taken together, these results demonstrate that chronic environmental neonicotinoid exposure significantly decreases survival of winter workers in the laboratory, but only chronic high-dose thiamethoxam significantly decreases overwinter survival of colonies in the field.

Wood SC, Kozii IV, Koziy RV, Epp T, Simko E. "Comparative chronic toxicity of three neonicotinoids on New Zealand packaged honey bees." PLoS One. 2018 Jan 2;13(1):e0190517. doi: 10.1371/journal.pone.0190517. 

Abstract