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Detection of a Microsporidian, Nosema apis, Via Quantitative Real-time PCR
Bombus terrestris has played an important role in the pollination in agricultural fields for the alternatives in colony collapsing in the honeybee. Recently, some pathogens or parasites such as viruses, bacteria, mites have been discovered in B. terristris, which affects its life span and fecundity. In order to detect a microsporidian, Nosema apis. in the field population, we collected honeybees and isolated genomic DNA. PCR primers specific for 16S ribosomal RNA (16S rRNA) were synthesized and applied to gene amplification for cloning and quantitative real-time PCR (qRT-PCR). The amplified gene was cloned and sequenced to confirm the 16S rRNA gene. qRT-PCR analysis showed the detection limit of 16S rRNA of Nosema apis was approximately 0.5 ng/μl genomic DNA. This result suggests that detection via qRT-PCR can be applied for the diagnosis of pathogen infection.
