Nosema ceranae Infections in Honey Bees (Apis mellifera) Treated with Pre/Probiotics and Impacts on Colonies in the Field.

Alternatives to the antibiotic fumagillin for the control of Nosema ceranae, a gut parasite of the honey bee, are needed. The prebiotics eugenol, chitosan, and naringenin and the probiotic Protexin® (Enterococcus faecium) provided in sugar syrup or protein patty either in spring or fall were evaluated for their effects on N. ceranae infection, colony population, honey yield and winter survivorship using field colonies. In the first year, spring treatments with eugenol, naringenin, and Protexin® significantly reduced N. ceranae infection and increased honey production, while Protexin® also increased adult bee populations and chitosan was ineffective. Fall treatments increased survivorship and decreased N. ceranae infection the following spring. In the second year, selected compounds were further tested with a larger number of colonies per treatment and only protein patty used in the spring and sugar syrup in the fall. Protexin® and naringenin significantly decreased N. ceranae infections and increased the population of adult bees after spring treatment, but did not affect honey yields. There were no differences between treatments for colony winter mortality, but surviving colonies that had been treated with Protexin® and naringenin were significantly more populated and had lower N. ceranae spore counts than control, non-treated colonies. Protexin® and naringenin were the most promising candidates for controlling N. ceranae and promoting honey bee populations, warranting further investigation. Future research should investigate the optimal colony dose and treatment frequency to maximize colony health.

Selective Breeding for Low and High Varroa destructor Growth in Honey Bee (Apis mellifera) Colonies: Initial Results of Two Generations.

After two years of bidirectional selection for low and high rates of Varroa destructor population growth (LVG and HVG, respectively) in honey bee (Apis mellifera) colonies in Ontario, Canada, significant differences between the two genotypes were observed. LVG colonies had V. destructor population increases over the summer of 1.7 fold compared to 9.6 fold for HVG colonies by Generation 2. Additionally, HVG colonies had significantly higher mite infestation rates in adult bees compared to LVG colonies for both selected generations. DWV prevalence and levels were significantly higher in HVG colonies than in LVG colonies in Generation 1 but not in Generation 2. Winter mortality rates of Generation 1 colonies were significantly different at 26% and 14% for the HVG and LVG genotypes, respectively. The results of this study thus far indicate that selection for LVG may result in colonies with lower V. destructor infestation rates, lower prevalence, and levels of DWV and higher colony winter survivorship. Future work will focus on determining what mechanisms are responsible for the genotypic differences, estimating genetic parameters, and molecular analyses of the genotypes to identify candidate genes associated with resistance to V. destructor and DWV that could potentially be used for marker-assisted selection.

Evaluation of Dry and Wet Formulations of Oxalic Acid, Thymol, and Oregano Oil for Varroa Mite (Acari: Varroidae) Control in Honey Bee (Hymenoptera: Apidae) Colonies.

The efficacy and safety of dry and wet formulations of three nonsynthetic compounds, oxalic acid (OA), thymol (T), and oregano oil (OO), for the control of Varroa destructor Anderson and Trueman infestations in honey bee (Apis mellifera Linnaeus) colonies were determined. The treatments were OA in dust, OA diluted in glycerin solvent embedded in a towel, T in dust, T in glycerin solvent and towel, OO in dry microcapsules, OO in glycerin solvent and towel, and the control. The treatments were applied weekly for 4 wk during the fall season. The rates of acaricide efficacy, weekly mite fall, bee mortality, colony survivorship, and strength, were determined for each of the treatments. All formulations, with the exemption of OO microcapsules, were effective at controlling infestations of V. destructor. The most effective formulations were T dust (96.6%), T glycerin (92.4%), and OA glycerin (79%). More than 85% of the mites were killed during the first 2 wk of treatment with T formulations, compared to less than 30% for the OA glycerin formulation. The lowest efficacy rate was for OO microcapsules (21.3%), and the only treatment that significantly increased bee mortality was OA glycerin. The rates of winter colony survival and honey bee populations were related to the varroacidal efficacy of the formulations. The implications of these findings are discussed.

Seasonality of Nosema ceranae Infections and Their Relationship with Honey Bee Populations, Food Stores, and Survivorship in a North American Region.

Nosema ceranae is an emerging pathogen of the western honey bee (Apis mellifera L.), and thus its seasonality and impact on bee colonies is not sufficiently documented for North America. This study was conducted to determine the infection intensity, prevalence, and viability of N. ceranae in >200 honey bee colonies during spring, summer, and fall, in a North American region. We also determined the relationship of N. ceranae infections with colony populations, food stores, bee survivorship, and overwinter colony mortality. The highest rates of N. ceranae infection, prevalence, and spore viability were found in the spring and summer, while the lowest were recorded in the fall. N. ceranae spore viability was significantly correlated with its prevalence and infection intensity in bees. Threshold to high levels of N. ceranae infections (>1,000,000 spores/bee) were significantly associated with reduced bee populations and food stores in colonies. Furthermore, worker bee survivorship was significantly reduced by N. ceranae infections, although no association between N. ceranae and winter colony mortality was found. It is concluded that N. ceranae infections are highest in spring and summer and may be detrimental to honey bee populations and colony productivity. Our results support the notion that treatment is justified when infections of N. ceranae exceed 1,000,000 spores/bee.

Continuous release of oregano oil effectively and safely controls Varroa destructor infestations in honey bee colonies in a northern climate.

The ectoparasitic mite Varroa destructor is responsible for the death of millions of honey bee (Apis mellifera) colonies worldwide. Testing potential miticide compounds with different delivery methods that effectively control V. destructor and have low toxicity for honey bees is crucial to manage this parasite in hives. We determined the varroacide efficacy of three natural compounds delivered to hives with three application methods over a 4-week period. Oxalic acid in a sucrose solution was applied impregnated in cardboard (T1). A mixture of oregano and clove oils in an ethanol-gelatin solution was applied impregnated in absorbent pads (T2). Oregano oil alone was delivered using electric vaporizers (T3) to test the hypothesis that continuous release of miticides increases the varroacidal efficacy of essential oils. The varroa mite control rates for treatments T1-T3 were 76.5 ± 7.11, 57.8 ± 12.79 and 97.4 ± 0.68%, respectively, and there were no differences for bee mortality between control and treatments 1 and 3. Additionally, most mites were killed in the first 2 weeks in T3 colonies compared to the last 2 weeks in colonies of the other treatments. These results demonstrate the importance of continuously releasing natural miticides to achieve safe and high rates of mite control in hives. They also show that oregano oil may be an effective miticide against V. destructor infestations in colonies.

Viability and infectivity of fresh and cryopreserved Nosema ceranae spores.

The microsporidium fungus Nosema ceranae is an intracellular parasite that infects the midgut of the honey bee, Apis mellifera. A major limitation of research on N. ceranae is that the fungus is non-culturable and thus studying it depends on the seasonal availability of Nosema spores. Also, spore viability and infectivity can vary considerably, and thus there is a need for reliable methods for determining those traits. This study examined different conditions for N. ceranae spore cryopreservation at -70°C, assessing spore viability and infectivity. Viability was determined by a staining procedure counting total spores numbers with bright field microscopy and un-viable spore numbers with the fluorescent dye, propidium iodide. Spore infectivity was determined with a dilution inoculation assay. Infectivity was dependent on the inoculum dose for the proportion of bees with detectable Nosema infections based on the number of spores per bee at 18days after inoculation; 4000 spores per bee or higher were needed to get approx. 100% of the inoculated bees infected. The median infective dose (ID50) was 149 spores per bee, and the minimum dose capable of causing a detectable infection was 1.28 spores. The proportion of N. ceranae infected bees correlated significantly with the number of spores per bee (r=0.98, P<0.0001). N. ceranae spores cryopreserved in water or 10% glycerol did not differ in viability compared to fresh spores, but lost infectivity when inoculated into bees. This study shows that while cryopreservation of N. ceranae spores can preserve viability, the spores can have reduced infectivity.