OCENA PRZYDATNOŚCI WYBRANYCH METOD GENOTYPOWYCH I FENOTYPOWYCH DO WEWNĄTRZGATUNKOWEGO RÓŻNICOWANIA PAŁECZEK CAMPYLOBACTER JEJUNI ORAZ CAMPYLOBACTER COLI IZOLOWANYCH OD LUDZI. II. PRZYDATNOŚĆ METOD PFGE, ERIC-PCR, PCR-FLA-A-RFLP I MLST
[EVALUATION OF GENOTYPIC AND PHENOTYPIC METHODS FOR THE DIFFERENTIATION OF CAMPYLOBACTER JEJUNI AND CAMPYLOBACTER COLI CLINICAL ISOLATES FROM POLAND. II. PFGE, ERIC-PCR, PCR-FLAA-RFLP AND MLST]

Streszczenie.

Oceniono przydatność metod PFGE, ERIC-PCR oraz PCR-flaA-RFLP do wewnątrzgatunkowego różnicowania izolatów pałeczek C. jejuni C. coli wyosobnionych od ludzi ze sporadycznych zakażeń oraz z ognisk zakażeń pokarmowych. Izolaty z ognisk zakażeń pokarmowych dodatkowo analizowano metodą MLST. Wykazano, że spośród zastosowanych genotypowych metod typowania najwyższą wartością indeksu różnicowania izolatów pałeczek C. jejuni charakteryzowała się metoda PFGE i ERIC-PCR. Stwierdzono, że wszystkie oceniane genotypowe metody mogą być wykorzystywane do potwierdzania pokrewieństwa izolatów pałeczek C. jejuni i C. coli pochodzących od osób z poszczególnych ognisk zakażenia tymi drobnoustrojami.

Abstract.

Campylobacter jejuni and Campylobacter coli are leading cause of bacterial gastroenteritis in many developed countries. We compared pulsed-field gel electrophoresis (PFGE), enterobacterial repetitive intergenic consensus (ERIC-PCR) and restriction fragment length polymorphisms of the flagellin gene (PCR-flaA-RFLP) for distinguishing 128 C. jejuni isolates and 17 C. coli clinical isolates
isolated between 2006 and 2007 in Poland. We also analysed seven isolates from three C. jejuni family
outbreaks and three isolates from one C. coli family outbreak. These isolates were also analysed
by multilocus sequence typing (MLST). PFGE and PCR-flaA-RFLP were performed as described by CAMPYNET. The methods were evaluated and compared on the basis of their abilities to identify outbreaks isolates and discriminate between unrelated isolates (with D index). PFGE was found to be the most discriminatory method (D=0,999), followed by ERIC-PCR (D=0,997) and PCR-flaA-RFLP (D=0,912). PFGE and ERIC-PCR clearly divided C. jejuni and C. coli into two clusters. PFGE, ERIC-PCR and PCR-flaA-RFLP distinguished 117, 107 and 18 distinct profiles, respectively, among 128 C. jejuni isolates. Among 17 C. coli isolates, 15 PFGE and ERIC-PCR, and 7 PCR-flaA-RFLP profiles were found. All the methods identified the outbreaks strains. MLST analysis showed that C.
jejuni isolates obtained from three outbreaks belonged to three new 3847, 3848 and 3849 STs. We found isolates with the indistinguishable patterns in each method which were obtained from humans from the same region and related in time that potentially represent common-source outbreaks. We also found isolates with the indistinguishable patterns in each method that were obtained from humans from different part of Poland. This is the first report of using MLST, ERIC-PCR and PCR-flaA-RFLP methods to distinguish C. jejuni and C. coli clinical isolates in Poland. Our results demonstrate that
all typing methods evaluated in this study are highly discriminatory and useful to investigate Campylobacter
outbreaks in Poland. Our results also show that the genetic diversity of polish C. jejuni and C. coli isolates is very high.