Parallel evolution of Varroa resistance in honey bees: a common mechanism across continents?

The near-globally distributed ecto-parasitic mite of the Apis mellifera honeybee, Varroa destructor, has formed a lethal association with Deformed wing virus, a once rare and benign RNA virus. In concert, the two have killed millions of wild and managed colonies, particularly across the Northern Hemisphere, forcing the need for regular acaricide application to ensure colony survival. However, despite the short association (in evolutionary terms), a small but increasing number of A. mellifera populations across the globe have been surviving many years without any mite control methods. This long-term survival, or Varroa resistance, is consistently associated with the same suite of traits (recapping, brood removal and reduced mite reproduction) irrespective of location. Here we conduct an analysis of data extracted from 60 papers to illustrate how these traits connect together to explain decades of mite resistance data. We have potentially a unified understanding of natural Varroa resistance that will help the global industry achieve widespread miticide-free beekeeping and indicate how different honeybee populations across four continents have resolved a recent threat using the same suite of behaviours.

qPCR assays with dual-labeled probes for genotyping honey bee variants associated with varroa resistance

Background The varroa mite is one of the main causes of honey bee mortality. An important mechanism by which honey bees increase their resistance against this mite is the expression of suppressed mite reproduction. This trait describes the physiological inability of mites to produce viable offspring and was found associated with eight genomic variants in previous research. Results This paper presents the development and validation of high-throughput qPCR assays with dual-labeled probes for discriminating these eight single-nucleotide variants. Amplicon sequences used for assay validation revealed additional variants in the primer/probe binding sites in four out of the eight assays. As for two of these the additional variants interfered with the genotyping outcome supplementary primers and/or probes were developed. Inclusion of these primers and probes in the assay mixes allowed for the correct genotyping of all eight variants of interest within our bee population. Conclusion These outcomes underline the importance of checking for interfering variants in designing qPCR assays. Ultimately, the availability of this assay allows genotyping for the suppressed mite reproduction trait and paves the way for marker assisted selection in breeding programs.

Reproductive success of the parasitic mite (Varroa destructor) is lower in honeybee colonies that target infested cells with recapping

Cell recapping is a behavioural trait of honeybees (Apis mellifera) where cells with developing pupae are uncapped, inspected, and then recapped, without removing the pupae. The ectoparasitic mite Varroa destructor, unarguably the most destructive pest in apiculture world-wide, invades the cells of developing pupae to feed and reproduce. Honeybees that target mite infested cells with this behaviour may disrupt the reproductive cycle of the mite. Hence, cell recapping has been associated with colony-level declines in mite reproduction. In this study we compared the colony-level efficacy of cell recapping (how often infested cells are recapped) to the average mite fecundity in A. mellifera. Our study populations, known to be adapted to V. destructor, were from Avignon, France, Gotland, Sweden, and Oslo, Norway, and were compared to geographically similar, treated control colonies. The results show that colonies with a higher recapping efficacy also have a lower average mite reproductive success. This pattern was likely driven by the adapted populations as they had the largest proportion of highly-targeted cell recapping. The consistent presence of this trait in mite-resistant and mite-susceptible colonies with varying degrees of expression may make it a good proxy trait for selective breeding on a large scale.

Elevated recapping behaviour and reduced Varroa destructor reproduction in natural Varroa resistant Apis mellifera honey bees from the UK

Varroa destructor mites remain a major threat to Apis mellifera honey bees, yet many populations across the world have naturally evolved survivorship to infestation. Here, we investigated the roles of recapping and mite reproduction in natural Varroa resistant (NVR) colonies in the UK. Recapping frequency was higher in NVR colonies and targeted mite-infested cells in which the recapped diameters were larger. Mite reproduction was lower in NVR colonies due to increased offspring mortality, although recapping is unlikely the primary mechanism responsible. In an additional small experiment, infested brood removal was immediately present in naïve colonies, and recapping increased rapidly following initial mite exposure. Targeted recapping behaviour is a common trait in NVR colonies and may provide a useful indicator for mite resistance. In addition, reduced mite reproduction is a key resistance mechanism in NVR colonies in the UK, as also found in Europe, S. Africa, Brazil and Mexico.

qPCR Assays with Dual-Labeled Probes for Genotyping Honey bee Variants Associated with Varroa Resistance

Background: The varroa mite is one of the main causes of honey bee mortality. An important mechanism by which honey bees increase their resistance against this mite is the expression of suppressed mite reproduction. This trait describes the physiological inability of mites to produce viable offspring and was found associated with eight genomic variants in previous research. Results: This paper presents the development and validation of high-throughput qPCR assays with dual-labeled probes for discriminating these eight single-nucleotide variants. Within the sequenced samples, additional variants were detected in the primer/probe binding sites in four out of the eight variants of interest. As for two of these the additional variants interfered with the genotyping outcome supplementary primers and/or probes were developed. Inclusion of these primers and probes in the assay mixes allowed for the correct genotyping of all eight variants of interest.Conclusion: These outcomes underline the importance of checking for interfering variants in designing qPCR assays. Ultimately, the availability of this assay allows genotyping for the suppressed mite reproduction trait and paves the way for marker assisted selection in breeding programs.

Host-Parasite Co-Evolution in Real-Time: Changes in Honey Bee Resistance Mechanisms and Mite Reproductive Strategies

Co-evolution is a major driving force shaping the outcome of host-parasite interactions over time. After host shifts, the lack of co-evolution can have a drastic impact on novel host populations. Nevertheless, it is known that Western honey bee (Apismellifera) populations can cope with host-shifted ectoparasitic mites (Varroa destructor) by means of natural selection. However, adaptive phenotypic traits of the parasites and temporal variations in host resistance behavior are poorly understood. Here, we show that mites made adaptive shifts in reproductive strategy when associated with resistant hosts and that host resistance traits can change over time. In a fully-crossed field experiment, worker brood cells of local adapted and non-adapted (control) A.mellifera host colonies were infested with mites originating from both types of host colonies. Then, mite reproduction as well as recapping of cells and removal of infested brood (i.e., Varroa Sensitive Hygiene, VSH) by host workers were investigated and compared to data from the same groups of host colonies three years earlier. The data suggest adaptive shifts in mite reproductive strategies, because mites from adapted hosts have higher probabilities of reproduction, but lower fecundity, when infesting their associated hosts than mites in treated colonies. The results confirm that adapted hosts can reduce mite reproductive success. However, neither recapping of cells nor VSH were significantly expressed, even though the latter was significantly expressed in this adapted population three years earlier. This suggests temporal variation in the expression of adaptive host traits. It also appears as if mechanisms not investigated here were responsible for the reduced mite reproduction in the adapted hosts. In conclusion, a holistic view including mite adaptations and studies of the same parasite/host populations over time appears overdue to finally understand the mechanisms enabling survival of V.destructor-infested honey bee host colonies.

Geographical Distribution and Selection of European Honey Bees Resistant to Varroa destructor

Developing resistance to the varroa mite in honey bees is a major goal for apicultural science and practice, the development of selection strategies and the availability of resistant stock. Here we present an extended literature review and survey of resistant populations and selection programs in the EU and elsewhere, including expert interviews. We illustrate the practical experiences of scientists, beekeepers, and breeders in search of resistant bees. We describe numerous resistant populations surviving without acaricide treatments, most of which developed under natural infestation pressure. Their common characteristics: reduced brood development; limited mite population growth; and low mite reproduction, may cause conflict with the interests of commercial beekeeping. Since environmental factors affect varroa mite resistance, particular honey bee strains must be evaluated under different local conditions and colony management. The resistance traits of grooming, hygienic behavior and mite reproduction, together with simple testing of mite population development and colony survival, are significant in recent selection programs. Advanced breeding techniques and genetic and physiological selection tools will be essential in the future. Despite huge demand, there is no well-established market for resistant stock in Europe. Moreover, reliable experience or experimental evidence regarding the resistance of stocks under different environmental and management conditions is still lacking.

How the Infestation Level of Varroa destructor Affects the Distribution Pattern of Multi-Infested Cells in Worker Brood of Apis mellifera

The mite Varroa destructor, the main ectoparasite of honey bees, is a threat to apiculture worldwide. Understanding the ecological interactions between Varroa and honeybees is fundamental for reducing mite impact in apiaries. This work assesses bee colonies with various Varroa infestation levels in apiaries to determine: (1) the relationship between multi-infested brood cells and brood infestation level, (2) the damage caused by Varroa to parasitized honey bee pupae, and (3) mite reproduction rate at various infestation levels. Data were collected from 19 worker brood combs, each from a different colony, ranging from 160 to 1725 (mean = 706) sealed cells per comb. Mite distribution was aggregated, ranging from about 2% to 74% infested cells per comb. The percentage of cells invaded by one, two, three, four, or more than four foundress mites, as a function of infestation level, was estimated by five highly significant (p < 0.0001) second-degree polynomial regression equations. The correction factors found could increase the precision of prediction models. Varroa fertility and adult bee longevity decreased as multi-infestation levels increased, and the implications of this relationship are discussed. Finally, these findings could improve sampling methods and the timing of mite treatments in apiaries, thus favoring sustainable management strategies.

Evaluation of Traits for the Selection of Apis Mellifera for Resistance against Varroa Destructor

Infestation with Varroa destructor is a serious cause of bee colony (Apis mellifera) losses on a global level. However, the presence of untreated survivor populations in many different regions supports the idea that selection for resistance can be successful. As colony survival is difficult or impossible to measure, differences in mite infestation levels and tests for specific behavioral traits are used for selective breeding for Varroa resistance. In this paper we looked into different definitions of mite infestation and linked these with brood hygiene (pin test), brood recapping and suppressed mite reproduction. We based our analyses on datasets of Apis mellifera carnica from three countries: Austria (147 records), Croatia (135) and Germany (207). We concluded that bee infestation in summer, adjusted for the level of natural mite fall in spring, is a suitable trait in the breeding objective, and also suggested including brood infestation rate and the increase rate of bee infestation in summer. Repeatability for bee infestation rate was about 0.55, for cells opened in pin test about 0.33, for recapping 0.35 and for suppressed mite reproduction (SMR) virtually zero. Although in most cases we observed correlations with the expected sign between infestation parameters and behavioral traits, the values were generally low (<0.2) and often not significantly different from zero.

Evaluation of Suppressed Mite Reproduction (SMR) Reveals Potential for Varroa Resistance in European Honey Bees (Apis mellifera L.)

In the fight against the Varroa destructor mite, selective breeding of honey bee (Apis mellifera L.) populations that are resistant to the parasitic mite stands as a sustainable solution. Selection initiatives indicate that using the suppressed mite reproduction (SMR) trait as a selection criterion is a suitable tool to breed such resistant bee populations. We conducted a large European experiment to evaluate the SMR trait in different populations of honey bees spread over 13 different countries, and representing different honey bee genotypes with their local mite parasites. The first goal was to standardize and validate the SMR evaluation method, and then to compare the SMR trait between the different populations. Simulation results indicate that it is necessary to examine at least 35 single-infested cells to reliably estimate the SMR score of any given colony. Several colonies from our dataset display high SMR scores indicating that this trait is present within the European honey bee populations. The trait is highly variable between colonies and some countries, but no major differences could be identified between countries for a given genotype, or between genotypes in different countries. This study shows the potential to increase selective breeding efforts of V. destructor resistant populations.