Differentiation of Staphylococcus spp. by terminal-restriction fragment length polymorphism analysis of glyceraldehyde-3-phosphate dehydrogenase-encoding gene.
Classical phenotypic and biochemical testing do not lead to correct identification of the distinct Staphylococcus species. Therefore, the aim of our study was to develop a method for the reliable and accurate determination of distinct Staphylococcus species. In the present study, the 931-934-bp partial sequences of the glyceraldehyde-3-phosphate dehydrogenase-encoding (gap) gene of 28 validly described Staphylococcus species were amplified and sequenced. By using the respective sequence information we performed a terminal-restriction fragment length polymorphism (T-RFLP) analysis. For T-RFLP the partial gap gene was amplified with double-fluorescently labelled primers and digested with the restriction enzymes DdeI, BspHI and TaqI. Distinctive T-RFLP patterns were rendered by the use of capillary electrophoresis with laser-induced fluorescence detection. This molecular method allowed us to identify all 28 Staphylococcus species with high specificity. This was validated by analysis of 34 Staphylococcus epidermidis and 28 Staphylococcus haemolyticus isolates. These results demonstrate the feasibility and applicability of the T-RFLP method based on the partial gap gene sequences for rapid and accurate species identification.
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F
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Otto-von-Guericke University, Institute of Medical Microbiology, Leipziger Str. 44, D-39120 Magdeburg, Germany.
Ghebremedhin
B
B
König
W
W
König
B
B
D001483:Base Sequence; D017931:DNA Primers; D005798:Genes, Bacterial; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000235:genetics*; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000379:methods; D012150:Polymorphism, Restriction Fragment Length; D015183:Restriction Mapping / Q000379:methods; D013210:Staphylococcus / Q000145:classification* / Q000201:enzymology / Q000235:genetics
D017931:DNA Primers; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases