The degree of variation was then

compared among the different loci, and three were found to have the greatest detection power for identifying A. apis haplotypes. The described loci can help to resolve strain differences and population genetic structures, to elucidate host–pathogen interaction and to test evolutionary hypotheses for the world’s most important pollinator: the honey bee and one of its most common pathogens. The parasite and pathogen pressure on honey bees is high because of both their eusocial lifestyle, which facilitates horizontal transfer between nest mates, and the close relatedness among nest mates. The fungus Ascosphaera apis is a common pathogen in honey bee colonies worldwide, causing chalkbrood disease (see Aronstein & Murray, 2010). This pathogen affects honey bee larvae, BYL719 which become infected upon ingestion of A. apis ascospores Akt inhibitor (Gilliam and Vandenberg, 1997). Honey bee larvae have a closed hindgut during most of their development where ingested ascospores germinate, and subsequently the hyphae penetrate the gut wall, entering the hemocoel into an environment that is scarce of other microorganisms with which they might compete for the easily accessible nutrients. If the fungus overcomes the host’s immune responses, the hyphae expand and will eventually

kill and mummify the infected larva. All members of the genus Ascosphaera live in association with social or solitary bees, some as saprophytes on

larval cAMP debris, fecal matter, or pollen provisions. Several species have similar life histories and pathologies that are comparable to A. apis, but infect solitary bees instead of honey bees (Skou, 1972, 1988; Bissett, 1988; Anderson et al., 1998). In addition to A. apis, Ascosphaera aggregata is also of economic importance, causing fatal infections in alfalfa leafcutting bees, especially when these bees are kept in dense populations for pollination service in alfalfa seed production systems (Pitts-Singer, 2008). A better understanding of the competitive interactions between A. apis strains and their bee hosts will aid disease control efforts (James, 2008). However, first, we must be able to differentiate between different strains or haplotypes. The internal transcribed spacer (ITS) region of the nuclear ribosomal repeat unit is the locus most often used for molecular species identification and subgeneric phylogenetic inference within the fungal kingdom (Nilsson et al., 2008). The ITS region has been used to study the genetic relationships of species within Ascosphaera (Anderson et al., 1998) and is also the locus used for development of species-specific primers (James & Skinner, 2005; Murray et al., 2005). The intraspecific variability of the ITS region, however, seems to be limited, with no sequence difference between A. apis isolates (Anderson et al., 1998). A lack of intraspecific variation in the ITS sequences were likewise found in A.