No abstract

Studies were conducted into the formation and physiological state of coccoid cells of a strain of the human and animal pathogen Campylobacter jejuni. It was found that growth phase and the presence of chloramphenicol did not affect the rate of shape transformation from spiral to coccoid, while nutrient limitation, aeration of the medium and the presence of free-radical scavengers had profound effects. Coccoid cells were found to reduce the tetrazolium salts INT (2-(p-iodophenyl)-3-(p-nitrophenyl)-5-phenyl tetrazolium chloride) and CTC (5-cyano-2,3-ditolyl tetrazolium chloride) to their respective formazans and this was linked to cellular respiration. However, respiring coccoid cells could not sustain their existence in prolonged adverse conditions, and it was concluded that they represent a degenerative stage rather than a dormant state of the organism.

Amplified-fragment length polymorphism (AFLP) analysis is the name given to a genotypic technique in which adapter oligonucleotides are ligated to restriction enzyme fragments and then used as target sites for primers in a PCR amplification process. The amplified fragments are electrophoretically separated to give strain-specific band profiles. We have developed a single-enzyme approach that did not require costly equipment or reagents for the fingerprinting of strains ofHelicobacter pylori. The method was assessed with 46 isolates of H. pylori from 28 patients, and the results were compared with those from other genotypic tests. The AFLP profiles derived from HindIII fragments differentiated strains ofH. pylori from unrelated individuals and confirmed the common origin of strains in some family members. AFLP analysis was also applied to investigate persistent infection following antibiotic therapy. Overall, the modified technique was relatively rapid and technically simple yet gave reproducible and discriminatory results. AFLP analysis samples variation throughout the genome and is a valuable addition to the existing genotypic fingerprinting methods for H. pylori.

A 1151‐bp amplicon containing the hippuricase (hipO) gene was obtained from 118strains of Campylobacter jejuni and double‐digested with AluI and DdeI togive five different PCR‐RFLP patterns. Most strains had the six‐banded profile predicted fromsequence data. Lack of polymorphisms within the hipO gene indicated it was highlyconserved amongst strains of Camp. jejuni, and that RFLP analysis provided only lowdiscrimination as an epidemiological typing method. Detection of hipO by PCR provided auseful test for confirmatory identification of Camp. jejuni.

High resolution molecular subtyping was applied to Campylobacter jejuni Penner heatstable (HS) 11 isolates from human infections and other sources. Strains were genotyped by restriction fragment length polymorphism analysis involving PCR-based flagellin gene CflaA) profiling with HinfI and DdeI, and pulsed-field gel electrophoretic (PFGE) profiling with SmaI and KpnI. Fla-genes of the strains were highly conserved as most (95%) had the same fla-profile. PFGE analysis of SmaI digests was more discriminatory with 15 profile subtypes identified, although 36% of isolates had a common profile. The study showed that strains of C. jejuni HS11, unlike those of HSl and the HS4 complex, were relatively homogeneous at the genomic level and that high resolution molecular techniques were essential for detailed epidemiological subtyping.