Discrimination between house dust mite Dermatophagoides pteronyssinus and D. farinae by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP)

Abstract

Problem/Background : House dust mites, Dermatophagoides pteronyssinus and D. farinae are the most common among all mite species distributed worldwide. They are also the most common sources of allergens causing asthma and other allergic diseases. Epidemiological studies of these house dust mites are based on morphological identification. This procedure faces some difficulties, including requirements of expertise to identify the mites and inability to discriminate immature stages. Therefore, we applied the molecular method for discriminating between the two most common house dust mites, D. pteronyssinus and D. farinae.

Objective : To demonstrate the usage of the cytochrome oxidase I (COI) gene for discriminating between D. pteronyssinus and D. farinae by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP).

Design : Descriptive study.

Setting : Department of Parasitology, Faculty of Medicine, Chulalongkorn University

Methods : The house dust mites; D. pteronyssinus and D. farinae, were maintained at Faculty of Agricultural Technology, King Mongkut’s Institute of Technology Ladkrabang, Bangkok, Thailand. The DNA extraction was amplified for COI gene by PCR technique. The DNA sequences were compared with GenBank database. As for PCR-RFLP analysis, the PCR products were digested in separate reaction with AluI.

Results : PCR yielded 1,559 and 1,562 bps for the COI gene of D. pteronyssinus and D. farinae respectively. AluI restriction digestions of the PCR amplicons produce different RFLP patterns between these house dust mite species.

Conclusions : This technique provides accurate identification of the house dust mites which can be used for epidemiological surveys, and therefore to perform diagnostic testing and for prescribing immunotherapy more efficiency.