Molecular Differentiation of Four Species of Oropsylla (Siphonaptera: Ceratophyllidae) Using PCR-Based Single Strand Conformation Polymorphism Analyses and DNA Sequencing
| Autor: | Todd K. Shury, Jessica T. Thoroughgood, Terry D. Galloway, Jeffery E. Lane, James S. Armstrong, L. Robbin Lindsay, Brandon White, Neil B. Chilton, Clare A. Anstead |
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| Rok vydání: | 2020 |
| Předmět: |
0106 biological sciences
Sequence analysis animal diseases 030231 tropical medicine 010603 evolutionary biology 01 natural sciences Polymerase Chain Reaction DNA sequencing law.invention 03 medical and health sciences 0302 clinical medicine law Animals Polymerase chain reaction Genetics Polymorphism Genetic General Veterinary biology Single-strand conformation polymorphism Sequence Analysis DNA Ribosomal RNA bacterial infections and mycoses biology.organism_classification DNA extraction Ceratophyllidae Infectious Diseases Genetic marker Insect Science Siphonaptera Parasitology |
| Zdroj: | Journal of medical entomology. 58(1) |
| ISSN: | 1938-2928 |
| Popis: | It is often difficult to distinguish morphologically between closely related species of fleas (Siphonaptera). Morphological identification of fleas often requires microscopic examination of internal structures in specimens cleared using caustic solutions. This process degrades DNA and/or inhibits DNA extraction from specimens, which limits molecular-based studies on individual fleas and their microbiomes. Our objective was to distinguish between Oropsylla rupestris (Jordan), Oropsylla tuberculata (Baker), Oropsylla bruneri (Baker), and Oropsylla labis (Jordan & Rothschild) (Ceratophyllidae) using PCR-based single strand conformation polymorphism (SSCP) analyses and DNA sequencing. A 446 bp region of the nuclear 28S ribosomal RNA (rRNA) gene was used as the genetic marker. The results obtained for 36 reference specimens (i.e., fleas that were morphologically identified to species) revealed no intraspecific variation in DNA sequence, whereas the DNA sequences of the four species of Oropsylla differed from one another at two to six nucleotide positions. Each flea species also had a unique SSCP banding pattern. SSCP analyses were then used to identify another 84 fleas that had not been identified morphologically. DNA sequencing data confirmed the species identity of fleas subjected to SSCP. This demonstrates that PCR-SSCP combined with DNA sequencing of the 28S rRNA gene is a very effective approach for the delineation of four closely related species of flea. |
| Databáze: | OpenAIRE |
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