DNAse-positive strains of Campylobacter jejuni degrade their chromosomal DNA during standard preparative procedures before pulsed-field gel electrophoresis (PFGE). A simple method for inactivation of this DNAse activity is described. Formaldehyde fixation of the bacterial cells resulted in the preservation of the DNA in a state suitable for restriction digestion and subsequent electrophoretic analysis.

We describe a molecular subtyping scheme for two principal O (heat-stable [HS]) serotypes of Campylobacter jejuni, HS1 and the HS4 complex. A 16S rRNA gene-specific probe confirmed that almost all the C. jejuni strains had three copies of this gene, and strains could be assigned with complete typeability to 1 of 16 combined (PstI and HaeIII) 16S ribotypes. Macrorestriction profiles (mrps) consisting of up to 10 SmaI fragments from ϳ40 to ϳ480 kbp were resolved by pulsed-field gel electrophoresis (PFGE). There were 11 mrps among the HS1 strains and 9 mrps among HS4 strains which corresponded to valid types-they occurred in multiple isolates, hosts, places, and times. There were 14 additional single-strain mrp fingerprints in HS1 and 20 in HS4. PFGE exhibited complete typeability when formaldehyde fixation of cells was employed, and PFGE was generally more differential than ribotyping. The data presented elucidate a high-resolution genotypic subtyping scheme for these common subspecific phenotypes of C. jejuni, which is both coherent and efficient for epidemiological purposes.

Campylobacter coli strains from clinical and other sources were examined in terms of O (heat-stabile; HS) serotype and by several molecular typing techniques. Restriction fragment length polymorphism (RFLP) around the three 16S rRNA genes revealed 10 variants, none found in Campylobacter jejuni. RFLP analysis of a polymerase chain reaction amplicon generated from the flagellin gene (flaA) yielded 11 polymorphism groups, some of them linked to HS serotypes. Enlarged flaA genes, contributing three further polymorphisms, were detected in strains isolated from fresh water. Restriction of the genome with SmaI and pulsed-field gel electrophoresis was the most discriminatory typing method, detecting 33 macrorestriction profiles that subtyped within HS serotypes. The coincidence of HS serotype and the three genotypic markers identified clonal lines of evolutionary and epidemiologic significance.

SU'MMARYFlagellin gene sequence polymorphisms were used to discriminate amongst 77 strains of Carnpylobacter jejuni from sporadic and outbreak-associated human enteric infections, and from chickens, sheep and calves. The results were assessed in relation to Lior biotyping and serotyping (Penner somatic antigens). Eight DNA PCR-RFLP patterns (genotypes) were identified by analysis of HinfI fragment length polymorphisms in flagellin gene (flaA) polymerase chain reaction (PCR) products. One genotype (F-1) was a feature of 55 % of strains. Strains within the genotypes were heterogeneous with respect to somatic antigens with 12 serogroups represented amongst the C. jejuni isolates offla A type F-1. Serogroups Pen 1, 2 and 23 were the commonest (45 %) amongst the 20 different serogroups represented. Several unique clusters of isolates with diverse biotypes were defined, and one cluster (F-7/Pen 23) contained epidemiologically implicated outbreak strains as well as sheep and calf isolates. We conclude that HinfI fia A typing is reproducible and offers high typability, and its combination with serogrouping provides a novel approach to characterizing isolates of C. jejuni with improved discrimination.