Evaluation of 10 methods to distinguish epidemic-associated Campylobacter strains

Abstract

We compared four phenotypic and six genotypic methods for distinguishing Campylobacter jejuni strains from animals and humans involved in four epidemics. Based on a comparison with epidemiologic data, the methods that correctly identified all strains in three milkborne outbreaks and one waterborne outbreak were heat-stable and heat-labile serotyping; multilocus enzyme electrophoresis (MEE); DNA restriction endonuclease analysis with BgIII, XhoI, PvuII, or PstI; and Southern blot and hybridization of PvuII-and PstI-digested DNA with Escherichia coli 16S and 23S rRNA (ribotyping). Biotyping, phage typing, plasmid analysis, and probing of BglII and XhoI DNA digests with C. jejuni 16S rRNA genes failed to correctly separate one or more strains. MEE, restriction endonuclease analysis, and ribotyping were the most sensitive methods and identified nine types among the 22 strains. These methods were also capable of further distinguishing strains within the same serotype. Data from MEE were also analyzed to calculate genetic relatedness among strains. Serotyping was the most discriminating phenotypic method, with eight and seven types distinguished by the heat-stable and heat-labile methods, respectively. MEE and ribotyping had several advantages over the other methods because they measure relatively stable and significant chromosomal differences and are applicable to other species and genera. These methods, however, are complex and not easily quantified; they are currently limited to specialized laboratories. When antisera are available, serotyping appears to be an effective and more practical approach to the identification of epidemic-related strains.

Scientific Opinion on Quantification of the risk posed by broiler meat to human campylobacteriosis in the EU

“…Serotyping, based on heat-labile antigens (HL) (Lior et al, 1982) or heat-stable antigens (HS) (Penner and Hennessy, 1980), and phagetyping (Grajewski et al, 1985;Frost et al, 1999;Patton et al, 1991) have been the main phenotypic methods applied separately or in combination. As well as being technically demanding, poor and variable typeability (the proportion of isolates for which a subtype can be determined) is inherent to the serotyping methods applied to C. jejuni .…”

“…This may, in part, be due to the geographical and temporal restriction of the sample used to define the origin panel of isolates for the most widely applied Penner scheme (Penner and Hennessy, 1980), which limits the diversity indexed among these reference serotypes. Increasing the number of phages used in phagetyping schemes has improved the typeability and discrimination offered by this approach Patton et al, 1991). The combination of phagetyping with serotyping improves discrimination and became the UK reference laboratory approach .…”

Restriction fragment length polymorphism analysis of the flaA gene of Campylobacter jejuni for subtyping human, animal and poultry isolates

“…Most of the C. jejuni strains examined were human isolates, (133) with other isolates from cattle (27), sheep (17), poultry (6), and dogs (4). Most £aA-types Table 2 Serotypes and EP £a-types of the C. jejuni strains studied…”

“…Campylobacter jejuni is a Gram-negative microaerobic bacterium that is an important agent of human diarrhoea throughout the world [1^3]. To improve the general understanding of the epidemiology of such infections, in particular to establish sources of outbreaks, various strain typing methods have been developed and applied over the past decade [4,5]. The approaches used included internationally recognised serotyping schemes based on heat stable (HS) antigens and heat labile (HL) antigens, as well as phage typing and biotyping (resistotyping).…”

“…There is an increasing trend in many areas of clinical microbiology to adopt molecular ¢ngerprinting (genotyping) methods such as ribotyping, pulsed ¢eld gel electrophoresis (PFGE) and PCR-based pro¢ling, as alternatives to phenotyping for discriminating between microorganisms [6,7]. Evaluations of such methods for C. jejuni have been reported [4,5,8] and although no consensus has been reached, £agellin gene pro¢ling has emerged from work done at a number of centres as a method suitable for rapidly investigating large numbers of C. jejuni [9^17]. However, there is currently no generally accepted protocol for £a-typing of C. jejuni with most investigators using RFLP analysis protocols based either on DdeI digestion [9], on HinfI and DdeI digestions [13,17], or on combined EcoRI/PstI digestion [10,16].…”