Evaluation of methods for subtyping Campylobacter jejuni during an outbreak involving a food handler

Abstract

In October 1998, the Centers for Disease Control and Prevention (CDC) assisted in an investigation of an outbreak of campylobacteriosis at a school in Salina, Kansas. Twenty-two isolates were submitted from the Kansas state public health laboratory to CDC, 9 associated with the outbreak and 13 epidemiologically unrelated sporadic isolates. Pulsed-field gel electrophoresis (PFGE) using SmaI and SalI was initially used to validate the epidemiologic data. We then tested the ability of other subtyping techniques to distinguish the outbreak-associated isolates from unrelated sporadic isolates. The methods employed were somatic O serotyping, PCR-restriction fragment length polymorphism (RFLP) analysis of flaA, DNA sequence analysis of 582 bp of flaA that included the short variable region (SVR), and sequencing of the entire flaA gene. PFGE was the most discriminatory technique, yielding 11 SmaI and 10 SalI restriction profiles. All outbreak isolates were indistinguishable by PFGE, somatic O serotyping, and sequencing of the 582-bp region of the flaA gene. fla typing by PCR-RFLP grouped one sporadic isolate with the outbreak strain. Analysis of the DNA sequence of a 582-bp segment of flaA produced strain groupings similar to that generated by PCR-RFLP but further differentiated two flaA PCR-RFLP types (with a 1-bp difference in the 582-bp region). Two sporadic strains were distinct by flaA PCR-RFLP but differed only by a single base substitution in the 582-bp region. The entire flaA gene was sequenced from strains differing by a single base pair in the 582-bp region, and the data revealed that additional discrimination may in some cases be obtained by sequencing outside the SVR. PFGE was superior to all other typing methods tested for strain discrimination; it was crucial for understanding the Kansas outbreak and, when SmaI was used, provided adequate discrimination between unrelated isolates.

FIG. 1

DdeI flaA PCR-RFLP patterns of Kansas C. jejuni strains. Lanes: 1, 100-bp ladder marker; 2, D5475; 3, D5478; 4, D5481; 5, D5488; 6, D5490; 7, D5492; 8, D5495.

FIG. 2

(a) PFGE restriction profiles of SmaI-digested DNA of Kansas C. jejuni strains. Lanes: 2, D5482; 3, D5487; 4, D5488; 5, D5481; 6, D5490; 7, D5491; 9, D5492; 10, D5493; 11, D5494; 12, D5495; 13, D5497; 1, 8, and 14, 48.5-kb DNA ladder. (b) PFGE restriction profiles of SalI-digested DNA of Kansas C. jejuni strains. Lanes: 2, D5482; 3, D5487; 4, D5481; 5, D5488; 6, D5490; 8, D5491; 9, D5493; 10, D5494; 11, D5495; 12, D5497; 1, 7, and 13, 48.5-kb DNA ladder.