Application of single-strand conformation polymorphism and denaturing gradient gel electrophoresis for fla sequence typing of Campylobacter jejuni.
Campylobacter jejuni is a frequent cause of bacterial gastroenteritis in humans all over the world. Several molecular typing methods are used to study the epidemiology of Campylobacter spp. infections. The aim of the present study was to investigate the application of single-strand conformation polymorphism (SSCP) and denaturing gradient gel electrophoresis (DGGE) analysis as rapid primary subtyping methods for C. jejuni. A variable fragment from the 3' end of the flaA to the 3' end of the intergenic region, separating the flaA and flaB genes, was subjected to SSCP and DGGE analysis. A total of 48 clinical C. jejuni isolates, 49 C. jejuni strains isolated from poultry, 2 strains isolated from ducks and 1 strain isolated from a pheasant were assigned to 24 distinct SSCP patterns. Sequence analysis of the respective DNA fragments revealed that every different fla sequence type could be distinguished by SSCP. DGGE proved to be equally discriminatory. Both methods can be applied as primary subtyping methods, because pulsed-field gel electrophoresis (PFGE) and amplified fragment length polymorphism (AFLP) analysis further differentiated isolates belonging to the same fla sequence types.
Hein
Ingeborg
I
Institute of Milk Hygiene, Milk Technology, and Food Science, University of Veterinary Medicine, Veterinaerplatz 1, 1210, Vienna, Austria. Ingeborg.Hein@vu-wien.ac.at
Mach
Robert L
RL
Farnleitner
Andreas H
AH
Wagner
Martin
M
D015373:Bacterial Typing Techniques / Q000379:methods*; D001483:Base Sequence; D016123:Campylobacter jejuni / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification; D004269:DNA, Bacterial / Q000032:analysis* / Q000737:chemistry; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods*; D005408:Flagellin / Q000032:analysis / Q000235:genetics; D005798:Genes, Bacterial; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D012150:Polymorphism, Restriction Fragment Length; D018807:Polymorphism, Single-Stranded Conformational; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA
D004269:DNA, Bacterial; D005408:Flagellin; C067138:flaA protein, bacteria; C416532:flaB flagellin