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Goals & Objective: Goal 2
Objective 2.2

Obj 2.2: Screening populations and commercial queens for genetic diversity

(Sheppard)

Rationale and significance
While QTL mapping offers great promise for screening bees for inclusion into breeding programs and queen producing operations, the number of genes that need to be identified is potentially large because resistance to each pest is likely a consequence of the effects of many different genes. Traditional breeding programs offer an opportunity for immediate relief. Honey bees suffer from multiple parasites and pathogens; however, to date, breeding programs have concentrated on resistance to one trait to the exclusion of others. Prior to establishing a breeding program that simultaneously selects for two or more traits, it is necessary to know the heritability of these traits and the genetic correlations between them. Previous work has demonstrated positive or neutral phenotypic correlations between three desirable colony-level traits: general hygienic behavior, Varroa sensitive hygiene (VSH) and honey production (Strange and Calderone, in press). These traits are major mechanisms of resistance to the principal problems affecting bees, including Varroa destructor (VSH assay) and American foulbrood and chalkbrood (general hygienic behavior assay). These findings suggest that these traits can be incorporated into a single breeding program. 

Genetic diversity of managed honey bee populations in the U.S. has declined over the past decade. Commercial queen propagation in the U.S. has focused on the production of large numbers of saleable queens from a limited number of queen mothers. The ratio of daughter queens to mothers produced annually averages over 1500:1 (Schiff and Sheppard 1995, 1996). This constitutes a genetic bottleneck and reduces the genetic variation available for breeding purposes. At the colony level, inbreeding has serious fitness costs, and conversely, high diversity within a colony from multiple matings has positive effects (Tarpy and Seeley 2006). We will confront the erosion of genetic diversity by measuring sequence diversity across populations and identifying practices that increase diversity. 

Expected outcomes 
To:

  1. determine heritability values and genetic correlations between important colony-level traits,
  2. provide foundation stock for model, multi-trait breeding program,
  3. develop molecular methods for identifying beneficial germplasm for incorporating into domestic queen producing and breeding programs, and
  4. identify novel sources of genetic diversity for incorporating into domestic queen producing and breeding programs. 
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Summary Statement for Goal 2

Workers in this goal have focused on improving breeding methods, identifying bee stocks with measurable resistance to Varroa mites, Nosema ceranae, and the virus IAPV, and improving the genetic diversity of bee stocks available to U.S. beekeepers. Greg Hunt’s lab developed an assay that correlates the proportion of mites removed from adult bees to the proportion of mites on bottom screens showing bite marks. Lab assays to screen bees for genetic resistance to N. ceranae were terminated following logistical difficulties and mounting evidence from this CAP that pathogenicity of N. ceranae is comparably low. Similar assays screening bees for resistance to IAPV showed only slight improvements in survivorship in a comparably resistant line. The return on effort has been much greater in the sub-objectives focused on identifying genes that confer resistance to Varroa mite. One putative quantitative trait locus (QTL) influencing grooming behavior and several putative QTLs influencing Varroa Sensitive Hygiene have been found, and work for year 4 will focus on confirming these QTLs for mite resistance and performing expression studies and RNAi experiments on the candidate genes identified.

In 2010, the Sheppard lab collected and imported semen from the Caucasian honey bee (A. m. caucasica) from the Caucasus Mountains of European Georgia and the Italian honey bee (A. m. ligustica) from Bologna, Italy. Germplasm was transported under USDA-APHIS permit to Washington State where it was used for instrumental insemination of honey bee queens. These queens were then maintained over the winter of 2010-2011 and some were provided to western queen producers for propagation in 2011. This work will continue in 2011 with semen imports from multiple locations in Slovenia (A. m. carnica) and Georgia (A. m. caucasica). In late 2011 and again in 2012, multiple successive queens will be produced from these stocks, resulting in nearly pure subspecies representatives for distribution to queen propagators to supply U.S. beekeepers. Sheppard’s group has been studying improved methods of cryopreserving sperm. Room temperature storage of semen for 45 days is possible with no appreciable loss in sperm viability, and future plans include studies to increase this room temperature storage life to 6 months and subsequently cryopreserve semen that was initially stored at room temperature. This would greatly facilitate collection efforts toward the development of a functional honey bee genetic repository by reducing the need to transport equipment and liquid nitrogen in the field during collecting trips abroad.

Progress

Increasing genetic diversity Sheppard and collaborators continued importation, screening and release of honey bee germplasm for breeding purposes. In 2010, we collected and imported germplasm (semen) from two honey bee subspecies previously introduced to the US in the 19th century. These were the Caucasian honey bee (A. m. caucasica) from the Caucasus Mountains of Georgia and the Italian honey bee (A. m. ligustica) from Bologna, Italy. Germplasm was transported under USDAAPHIS permit to Washington State where it was used for instrumental insemination of honey bee queens. These queens were then maintained over the winter of 2010-2011 and some were provided to western queen producers for propagation in 2011. Virgin queen progeny (50% Caucasian) from the 2010 importation will be used for insemination with 2011 - collected germplasm from other regions of the Caucasus Mountains. This is a noteworthy achievement, as the “Caucasian” strain of honey bees was present in the US for about 100 years following early introductions in the late 1800s, but has not been commercially available in the US as a distinct strain for the past 20 years. Caucasian honey bees are generally conservative of winter stores, making them well-suited to colder temperate areas and to overwinter with less feed inputs.

Improved breeding
In 2010 Sheppard and his student applied newly developed cryogenic methods to rear sequential generations of queens from a cryopreserved source of honey bee semen. Prior to this development, collected honey bee semen had a limited useful working life of around 2 weeks, limiting genetic contributions from imported semen to the target breeding population to 50% per importation event (typically one importation per year). However, using cryogenic preservation, multiple successive generations of queens can be reared per year using the cryopreserved semen sources. Therefore, reconstruction of desirable genetic stocks that would normally take 6 years of importation and breeding can now be achieved in a single year (6 queen backcrosses per season results in >98% genetic contribution from the source population). Virgin queens inseminated with cryopreserved semen that produce a majority of worker brood can be used as “queen mothers.” The queen mothers can be used for the production of subsequent queens that can then be instrumentally or naturally mated.

Methodology, data and analysis of results to date are shared in an annual report to USDA.

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Further Background Information

Documentation of CAP progress is available through the following sources:

  1. Bee Health, an eXention initiative for peer-reviewed scientific recommendations
  2. Colony Collapse Disorder Progress Report for 2009
  3. Breeding Bees for Resistance to Parasites and Diseases
  4. Genetic Toolkits for Bee Health
  5. Laying Groundwork for a Sustainable Market of Genetically-Improved Queens

Updated July 22, 2011.

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