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The internal spacer region (ISR) between the 16S and 23S rRNA genes of Campylobacter was investigated by PCR fragment length typing and DNA sequencing of clinical and chicken wild-type isolates. PCR fragment length typing showed one fragment of 859 nt in length for the 12 strains of Campylobacter coli investigated. Thirty-six of the Campylobacter jejuni subsp. jejuni strains possessed one fragment, which varied in size between 727 and 802 nt. Three strains showed two fragments between 501 and 923 nt. Strains of C. jejuni subsp. doylei, Campylobacter lari and Campylobacter upsaliensis possessed one or two fragments with lengths different from those of C. coli and C. jejuni subsp. jejuni. DNA sequences were obtained from 54 nt downstream of rrs up to rrl of four strains of C. coli, eight strains of C. jejuni subsp. jejuni, and one strain each of C. jejuni subsp. doylei and C. lari, selected to represent the different biotypes of Campylobacter. ISR lengths determined by PCR fragment length typing and DNA sequencing corresponded for 12 strains. For two strains of C. coli, PCR fragment length typing underestimated ISR lengths by 159 and 193 nt, probably related to incomplete resolution of the distal helical structures, which were not fully denatured during PAGE. For the 14 strains and the published C. jejuni subsp. jejuni sequence, the first 206-211 nt were conserved and included the two tRNA genes in the characteristic tRNA(Ala) to tRNA(Ile) order separated by a short 8-9 nt spacer region. Within the region downstream of tRNA(Ile) conserved regions were identified which allowed a separation of C. lari from C. coli and C. jejuni but not separation of C. coli from C. jejuni. The 69-282 nt longer variable regions in C. coli strains allowed separation of this species from C. jejuni, confirming results obtained by PCR typing. Certain nucleic acid positions in variable regions were related to the Lior biotypes. Sequence information from ISRs of more strains is needed to ascertain if separation of species and biotypes will be possible for diagnostic purposes.

Differentiation of Campylobacter coli and C. jejuni by length and DNA sequence of the 16S-23S rRNA internal spacer region