Phage typing combined with pulsed-field gel electrophoresis and random amplified polymorphic DNA increases discrimination in the epidemiological analysis of Salmonella enteritidis strains

Phage typing (PT) combined with pulsed-field gel electrophoresis (PFGE) and a random amplified polymorphic DNA (RAPD) fingerprinting method was used to characterize Salmonella enteritidis strains. Twenty-four epidemiologically unrelated isolates, sampled from diverse ecological niches and fifteen isolates from four well-defined outbreaks of foodborne gastroenteritis, were studied. Seven phage types, with a predominance of PT 4 (63% of isolates), were observed when analysing the epidemiologically unrelated group. PT 4 was detected in all of the ecological niches studied, including food and fecally polluted river and beach water. The discriminatory power for phage typing, the average probability that the typing system will assign a different type to two unrelated strains randomly sampled in the microbial population, was 0.62. Ten PFGE pattern types were obtained with Xba I restriction endonuclease enzyme among the unrelated isolates; thirteen isolates belonged to PFGE pattern type 1 and the rest of the PFGE types were assigned to one or two isolates. The Dice coefficient clustered the similarities of the PFGE patterns between 80–100%. PFGE showed a discriminatory power of 0.72. Five clearly distinct RAPD patterns were observed with the OPS-19 oligonucleotide, but the discrimination obtained was low (0.46). The combination of the three typing methods increased the number of types to seventeen, giving high discrimination (0.92). Seven of the isolates recovered from various ecological niches belonged to the combination PT 4/PFGE 1/RAPD A and other combinations were unique or included only two strains. The four epidemiologically well-defined foodborne outbreaks were associated with the PT 4 phage type. In two of the outbreaks, other phage types (PT 7a and RDNC) were also observed in two isolates. Most of the isolates belonging to the foodborne outbreaks had an identical PFGE pattern (PFGE pattern type 1), but a difference in a restriction band was observed in an isolate belonging to an outbreak. Two RAPD patterns were observed in the outbreaks; RAPD pattern type A was detected in three of the four outbreaks. When the combined typing method was applied to the study, high concordance was observed and most of the outbreak strains belonged to the combination PT 4/PFGE 1/RAPD A. It is concluded that the combination of phage type with PFGE and RAPD provides a powerful discriminatory tool for the epidemiological analysis of unrelated and related strains of S. enteritidis.