Differentiation of Ascosphaera apis isolates by rep-PCR fingerprinting and determination of chalkbrood incidence in Argentinean honey samples

SUMMARY

Ascosphaera apis, the causative agent of chalkbrood disease in honey bee larvae occurs throughout the world and is found in many beekeeping areas of Argentina. Microbiological analysis of 394 honey samples from Buenos Aires province from three years of sampling (1999–2001) yielded 51 positive cases (13% incidence). Eighty-four isolates of A. apis from Argentina and Chile isolated from diseased larvae and/or honey samples were characterized on the basis of DNA fingerprints using the repetitive-sequence-based polymerase chain reaction technique (rep-PCR) with BOX, REP, and ERIC sequence-specific primers. Computer-assisted analysis of combined fingerprints distinguished six groups of patterns, designated A, B, C, D, E and F. Pattern C was the most prevalent and suggests a limited diversity in the populations of A. apis from Argentina and Chile. The results demonstrated the usefulness of rep-PCR genomic fingerprinting to characterize populations of A. apis. In addition, a simple and efficient protocol for the extraction of total fungal genomic DNA was developed that was appropriate for simultaneous and cost-efficient processing of many samples.

Keywords:

• Ascosphaera apis
• chalkbrood
• Apis mellifera
• honey bees
• rep-PCR
• DNA fingerprinting
• honey