Associations between Heat-Stable (O) and Heat-Labile (HL) Serogroup Antigens of Campylobacter jejuni: Evidence for Interstrain Relationships within Three O/HL Serovars

Abstract

A comparative examination of the heat-stable (O) and heat-labile (HL) serogrouping results for 9,024 sporadic human isolates of Campylobacter jejuni revealed conserved associations between specific O and HL antigens (O/HL serovars). Forty-nine percent of the isolates which grouped for both O and HL antigens belonged to one of three serovars: O 4 complex/HL 1 (17.9%), O 1/HL 2 (16.8%), or O 50/HL 7 (14.5%). Other common serovars were O 2/HL 4 (8.3%), O 6/HL 6 (8.1%), O 53/HL 11 (4.5%), O 19/HL 17 (3.3%), O 5/HL 9 (3.3%), O 9/HL 9 (3.2%), and O 23/HL 5 (3.1%). These 10 serovars accounted for 83.1% of the serogroupable isolates. A large number of strains (41.3%) could be typed by only one of the two methods or could not be serogrouped (11%). Strains belonging to three serovars, O 2/HL 4, O 50/HL 7, and O 23/HL 5, were further characterized by combining data from expressed features (O/HL serogroups, phage groups, and biotypes) with restriction fragment length polymorphism genotypes. These polyphasic data demonstrated that within each serovar, individual isolates showed substantial conservation of both genomic and phenotypic characteristics. The essentially clonal nature of the three serovars confirmed the potential of combined O and HL serogrouping as a practical and phylogenetically valid method for investigating the epidemiology of sporadic C. jejuni infection.

The gram-negative spirillum Campylobacter jejuni is a primary etiologic agent of human gastroenteritis. Most campylobacter infections appear to be sporadic rather than outbreak associated, and in a majority of cases, the original source of infection cannot be determined (10). Case-control studies (4, 17) have identified several risk factors associated with campylobacteriosis, including consumption of untreated water or milk, foreign travel, and the handling or consumption of raw or undercooked poultry. Other significant but unrecognized sources of infection may also exist.

A better understanding of the epidemiology of sporadic campylobacter infection is essential for the development of intervention measures to reduce human exposure to this pathogen. The subtyping of campylobacter isolates has been valuable in the epidemiological investigation of point source campylobacter outbreaks (15, 31) and provides a means for monitoring the transmission of C. jejuni from environmental reservoirs and animal hosts into the human food chain (27). Numerous phenotyping methods for C. jejuni have been described, all of which, to some degree, fulfill the requirements of stability, reproducibility, and discrimination necessary for epidemiological analysis. However, inconsistent strain associations occur when different phenotypic techniques are used to examine identical sets of isolates (30). The development of campylobacter genotyping methods has not substantially resolved these anomalies (1, 9, 28), although some uniformity between genetic and phenotypic analyses for the most common O serogroups of C. jejuni has been reported (14, 18, 29). Progress towards methodological standardization (39) and increased awareness of the mechanisms responsible for variation in subtyping characteristics (12) may improve the quality of epidemiological information derived from the use of multiple campylobacter typing schemes.

In this report, we describe serovars of C. jejuni identified by a comparison of heat-stable (O) and heat-labile (HL) serogrouping results from over 9,000 sporadic human campylobacter isolates. An integrated analysis of expressed features (O and HL serogroups, phage groups, and biotypes) and restriction fragment length polymorphism (RFLP) genotypes was used to produce polyphasic strain profiles (41) for 52 strains from three common O/HL serovars of C. jejuni. The results of this comparative study revealed a number of novel features relevant to the epidemiology and population biology of this important enteropathogen.

MATERIALS AND METHODS

Bacterial strains and culture media.

Campylobacter strains for serogrouping were submitted from 23 United Kingdom (UK) laboratories over the period of 1988 to 1992. All strains were random, sporadic human fecal isolates submitted as part of a UK passive campylobacter surveillance program. Serogroup and RFLP data were also included in this study from 124 systemic C. jejuni isolates (19) and 14 human isolates and 1 bovine isolate from C. Nicol (Enteric Reference Laboratory, Porirua, New Zealand). O and HL serogroup reference cultures were provided by J. L. Penner (33 strains) and H. Lior (12 strains), including C. jejuni NCTC 11168, which was the HL 4 strain.

Fifty-two isolates associated with three C. jejuni serovars (O 2/HL 4, O 50/HL 7, and O 23/HL 5) were further characterized by phage group, biotype, and RFLP. These strains included one bovine, four environmental, and eight serogroup reference strains. The remainder were sporadic fecal or systemic human isolates originating from 17 UK regional centers.

Cultures were stored at −70°C in brain heart infusion broth (Unipath Ltd.) containing 15% glycerol. Bacteria were grown on Columbia agar (Unipath Ltd.) supplemented with 5% (vol/vol) whole horse blood for 48 h at 37°C under microaerophilic conditions (5% CO₂, 3% H₂, 85% N₂, 7% O₂) in a Variable Atmosphere Incubator (Don Whitley Scientific Ltd.).

Strain characterization.

Serogrouping of 9,024 sporadic human isolates of C. jejuni was carried out at Manchester Public Health Laboratory over the period of 1988 to 1992 by using both the Penner (O) and Lior (HL) systems in accordance with standard methods (22, 33, 34). The panel of available typing sera tested for 33 O antigens associated with C. jejuni and a restricted set of 12 of the most common HL serogroups (HL types 1, 2, 4 to 9, 11, 17, 20, and 21). Frequent cross-reactions occurred between O antigens 4, 13, 16, 43, and 50, and strains agglutinating with any of these five antisera were classified within the O 4 complex. The program DataEase (DataEase International, Inc.) was used for the comparison of O and HL serogroup determinants, and multiple isolates from known outbreaks were excluded from the analysis. Significant associations (P < 0.001) between specific O and HL antigens were calculated by chi-square analysis. Identification to the species level, biotyping, and phage typing were performed at the Preston Public Health Laboratory by previously described methods (8, 35). The methods used for nucleic acid extraction, restriction endonuclease digestion, and RFLP analysis of C. jejuni have been published elsewhere (18).

RESULTS

Serovars of C. jejuni are characterized by associations between specific O and HL antigens.

Typing results for both the O and HL serogroup antigens were obtained for 4,294 (47.6%) of the 9,024 isolates tested. Thirteen of the 33 available O antisera, including the five cross-reacting epitopes of the O 4 complex, grouped 3,845 (89.5%) of these strains and 6,222 (69.0%) of the total number of isolates. The remaining 20 O antisera grouped 449 (10.5%) of the strains with both O and HL results, and 1,454 (16.1%) of the total number of isolates (data not shown). Three hundred forty-three isolates (3.8%) were typeable by the HL system but not by the O system, while a much larger proportion (3,382; 37.5%) were typed by O but not HL antisera. There were 999 strains (11.0%) which could not be typed by either scheme.

A comparison of the most frequently reported O serogroups and their corresponding HL results (Table 1) provided evidence of strong nonrandom (P <0.001) associations or linkage between specific O and HL serogroups. These serovar-like combinations were most marked in the O 6, O 23, and O 53 strains, which associated almost exclusively with HL 6, HL 5, and HL 11 antigens, respectively. Similarly, within the three most common O serogroups, O 1 and O 2 strains showed strong linkage to the HL 2 and HL 4 antigens. Three serovars, O 1/HL 2, O 4/HL 1, and O 50/HL 7, accounted for 1,856 (43.2%) of the 4,294 strains, and 10 serovars represented 3,128 (72.8%) of these isolates.

TABLE 1.

Serogrouping results for 8,010 (88.7%) of 9,024 sporadic human C. jejuni isolates obtained with 17 HS and 12 HL antisera^a

O serogroup	No. of isolates in HL serogroup:													Total
	1	2	4	5	6	7	8	9	11	17	20	21	NT
1	81d	630c	3	1	2	9	1	3	2	8	0	14	244	998
2	10	13	314c	2	0	4	6	3	3	19	3	83	725	1,105
4	678c	33	8	4	5	548b,c	1	5	2	69	9	23	537	1,922
5	5	13	2	1	1	0	1	125c	36	7	0	3	216	410
6	1	0	1	0	303c	0	1	1	15	0	1	3	28	354
9	4	9	0	2	0	2	5	120c	23	9	1	16	223	414
11	4	5	0	0	1	4	80d	3	1	22	0	27	263c	410
15	3	0	1	1	0	2	1	3	9	1	0	2	139c	162
18	1	1	1	0	0	0	1	0	7	0	1	0	139c	151
19	2	1	1	0	0	22	1	1	0	124c	18	0	45	215
21	0	1	1	0	2	1	3	2	2	4	0	0	172c	188
23	0	0	2	117c	4	0	0	0	0	2	4	1	10	140
53	1	0	0	0	1	1	1	5	169c	1	1	0	19	199
NT	34	26	14	17	10	16	22	52	68	21	15	48	999	8,010

^aFive cross-reacting antigens (O 4, O 13, O 16, O 43, and O 50) were expressed by strains in the HS 4 serogroup. One thousand three hundred forty-two isolates (14.8%) were NT by the O system. The remaining 1,014 strains (11.2%) were characterized by using the following 16 additional O antisera: O 3, 151 strains; O 7, 9; O 8, 134; O 10, 197; O 17, 107; O 29, 43; O 35, 73; O 38, 7; O 40, 31; O 41, 24; O 42, 7; O 44, 26; O 45, 21; O 55, 121; O 58, 9; O 60, 23. 

^bO 4 complex strains associated with the HL 7 antigen frequently expressed the cross-reacting O 50 antigen. This strain cluster is referred to in the text as serovar O 50/HL 7. 

^cSignificant (P < 0.001) conserved association between specific O and HL antisera. 

^dSecondary antigen association. 

Strains within the O 4 complex were associated with two HL antigens, HL 1 and HL 7. Most O 4 complex strains of serogroup HL 7 expressed the O 50 epitope in various combinations with the other cross-reacting antigens. These isolates were placed within serovar O 50/HL 7, a defined lineage within the O 4 complex strains characterized by the O 50 and HL 7 serogroup reference strains (see Table 3). However, reactions with the O 50 antigen were not exclusive to this serovar but were also found with many other O 4 complex strains.

TABLE 3.

Strain details and typing characteristics for 18 random C. jejuni isolates associated with serovar O 50/HL 7^a

Laboratory no.	Date of isolation	Region	Source	Biotype	Phage group	RFLP type	16S ribotype
O 50b	1983	Toronto	S/G	6020	NT	8	4
O 65	NKc	NK	S/G	6000	55	8	4
HL 7	1981	Ottawa	S/G	6000	44	8	4
B/16023b	27/07/84	Manchester	S	6000	44	8	4
C/23502b	01/10/93	NZ CDSCd	B	6000	55	8	4
C/19584e	05/08/92	Poole	H	6000	55	8	4
C/13383f	07/03/91	Southampton	H	6000	44	8	4
C/12279	07/12/90	Blairmore	H	ND	55	8	4
B/1338	08/07/87	Cardiff	S	6000	44	8	4
B/2841	14/12/88	Glasgow	S	6000	55	8	4
P/3183	10/04/91	Aberdeen	E	6000	55	8	4
P/3187	10/04/91	Eastbourne	H	6000	44	8	4
C/14000	03/05/91	Dumfries	H	6000	44	8	4
P/3827	23/05/91	Bath	H	6000	44	8	4
B/8794	18/03/86	Manchester	S	6010	55	8	4
B/8817	02/07/90	Manchester	S	6010	44	8	4
P/3758	20/05/91	Poole	H	6000	52	4	1
P/2587	14/02/91	Edinburgh	H	6222	69	5	31

^aSee Table 2, footnote a. 

^bNot Lior typed (HL ND) or NT by available Lior antisera. 

^cNK, not known. 

^dNew Zealand bovine isolate. 

^eExpressed HL 20, an antigen atypical of this serovar. 

^fExpressed HL 1, an antigen atypical of this serovar. 

Within each O serogroup, a varying but often high proportion of isolates scored an HL nontypeable (HL NT) reaction. The Lior typing scheme recognizes in excess of 160 HL antigens for C. jejuni and C. coli, and only 12 of the most common ones were tested for in this study. For serogroups O 15, O 18, and O 21, greater than 95% of the strains were HL NT (Table 1), and these groups presumably agglutinate with individual homologous HL antisera that were not available in our panel. Some O serogroups did show a strong association with single HL antigens but also had a high proportion of HL NT results. For example, 28% of serogroup O 2 strains were serovar O 2/HL 4 and 65% were O 2/HL NT, while for O 11 isolates, 19.4% were serovar O 11/HL 8 and 64% were O 11/HL NT. An indeterminate but substantial proportion of these HL NT results may be due to factors such as antigenic loss or masking and autoagglutination, as well as the use of a restricted panel of HL antisera. Nontypeability was also experienced with the Penner system, but to a lesser extent (Table 1, row 14).

Strains of serogroups O 5 and O 9 showed concordance in the results for all 12 HL antisera, with strong linkage to the HL 9 antigen (30 and 28%, respectively) and to HL NT (53% each). The reasons for these similarities are unknown. Serogroup reference strains O 5 and HL 9 had identical biotypes and similar RFLP genotypes (18) but appeared to be unrelated to the O 9 serostrain.

Strains within three serovars of C. jejuni have many features in common.

The polyphasic strain characteristic (Preston biotype, phage group, 16S ribotype, and E3CJC2 RFLP type) of 52 random isolates of C. jejuni belonging to three serovars, O 2/HL 4, O 50/HL 7, and O 23/HL 5, are listed in Tables 2 to 4. These strains originated from different geographical areas within the UK and abroad and were isolated from a variety of sources over the period of 1980 to 1993.

TABLE 2.

Strain details and typing characteristics for 19 random C. jejuni isolates associated with serovar O 2/HL 4^a

Laboratory no.	Date of isolation	Region	Source	Biotype	Phage group	RFLP type	16S ribotype
O 2	1980	Toronto	S/G	6004	52	2	1
HL 4b	1981	London	S/G	6010	52	2	1
P/3618	10/05/91	Manchester	H	6004	52	2	1
C/4537	21/07/89	Birmingham	S	6004	52	2	1
C/12950	30/01/91	Birmingham	H	6004	52	2	1
C/12646	09/01/91	Hereford	H	6004	52	2	1
P/2361	24/01/91	Southampton	H	6004	75	2	1
C/4533	21/07/89	Birmingham	S	6004	NT	2	1
C/18508	17/05/92	Newcastle	H	6010	52	2	1
P/2987	16/03/91	Southampton	H	6010	52	2	1
P/2164	09/01/91	Norwich	H	6014	52	2	1
P/1962	13/12/90	Edinburgh	H	6014	69	2	1
P/2248	17/01/91	Aberdeen	H	6020	52	2	1
B/23383	11/10/84	Liverpool	S	6020	52	2	1
P/3018	14/03/91	Aberdeen	H	6024	52	2	1
P/2399	28/01/91	Cambridge	H	6024	52	2	1
P/3666	13/05/91	Norwich	H	6034	52	2	1
P/2255	01/17/91	Aberdeen	H	6124	69	1	1
P/3546	05/02/91	Southampton	H	6172	125	5	1

^aStrains belonging to three O/HL serovars of C. jejuni were characterized by biotyping, phage grouping, 16S ribotyping, and RFLP typing by using the E3CJC2 probe. Strain sources: B, bovine; E, environmental; H, human; S, systemic; S/G, serogroup reference culture. 

^bStrain HL 4 is NCTC 11168. 

TABLE 4.

Strain details and typing characteristics for 15 random C. jejuni isolates associated with serovar O 23/HL 5^a

Laboratory no.	Date of isolation	Region	Source	Biotype	Phage group	RFLP type	16S ribotype
O 36	1983	Toronto	S/G	6002	NT	9	13
HL 5	1981	Ottawa	S/G	6000	NT	9	13
C/13871	18/04/91	Aberdeen	H	6000	69	9	13
C/12632	09/01/91	Aberdeen	H	6004	69	9	13
C/12784	23/1/91	Moray	E	6004	69	9	13
C/13710	27/3/91	Turriff	E	6004	69	9	13
C/12267	07/12/90	Aberdeen	H	6006	69	9	13
C/18509	17/06/92	Newcastle	H	6006	69	9	13
C/19785	21/08/92	Kettering	H	6006	NT	9	13
C/13613	20/3/91	Aberdeen	E	6674	NT	9	13
C/13461	11/03/91	Hereford	H	6006	69	NDb	ND
C/14195	23/05/91	Bath	H	6004	69	ND	ND
C/13918	24/04/91	Dumfries	H	6044	69	ND	ND
O 23c	1980	Toronto	S/G	6020	NT	14	10
B/1916c	17/11/87	Worcester	S	6135	76	36	2

^aSee Table 2, footnote a. 

^bND, not done. 

^cSee Table 3, footnote b. 

Within each serovar, most isolates demonstrated strong conservation between the various expressed (phenotypic) and chromosomal (genetic) markers. Serovar O 2/HL 4 isolates (Table 2) were RFLP/ribotype 2/1 and associated strongly with phage group 52. Variable metronidazole resistance, 5-fluorouracil production, and elaboration of DNase characterized the biotypes (types 6004, 6010, 6014, 6020, and 6024). Isolates of serovar O 50/HL 7 (Table 3) were RFLP/ribotype 8/4 and expressed either very closely related phage group 44 or 55. This serovar was usually fully sensitive in resistotype tests and enzymatically unreactive (biotype 6000) but occasionally showed metronidazole or 5-fluorouracil resistance (biotypes 6010 and 6020).

Serovar O 23/HL 5 strains (Table 4) were characteristically RFLP/ribotype 9/13, phage group 69, and showed variable production of either hydrogen sulfide (H₂S) or DNase (biotypes 6000, 6002, 6004, and 6006). Phenotype characteristics of three O 23/HL 5 isolates (C/13461, C/14159, and C/13918) have been included in this table, although RFLP typing of these strains was not done. The evident relationships between the serogroup, phage group, and biotype characteristics of these three strains and those of other isolates of this serovar provide substantive evidence of their clonal identity, even without recourse to genomic analysis.

Atypical variation of both expressed and genotypic characteristics occurs within serovars.

The cross-reacting O 23 and O 36 antigens are almost identical serologically and biochemically (7), and it was the latter serotype strain which showed the serovar characteristics. The O 23 serogroup reference strain and isolate B/1916 (Table 4) were clearly unrelated to other strains of this serovar. Although both possessed the O 23 antigen, the HL antigens of these strains were atypical, demonstrating the low predictive value of inferring interstrain relationships from O serogroup data alone. However, even within true O/HL serovars, atypical strain variation occurred. Two serovar O 2/HL 4 strains, P/2255 and P/3546 (Table 2), and two serovar O 50/HL 7 strains, P/3758 and P/2587 (Table 3), possessed E3CJC2/16S RFLP types, phage groups, and biotypes that were mostly dissimilar from those of the other strains of these serovars. More limited variation was seen with isolates C/13383 and C/19584 (Table 3), which shared the distinctive features of serovar O 50/HL 7 strains, except that the expected HL 7 antigen had been replaced with HL 1 and HL 20 epitopes, respectively.

DISCUSSION

A number of different phenotypic approaches to strain identification have been employed in attempts to resolve the epidemiology of sporadic campylobacter infection. The two most common systems involve serogrouping based on either O lipopolysaccharide or HL surface-associated antigens. In this report, we show that the O and HL epitopes exhibit strong nonrandom associations and that many UK strains can be allocated to specific serovars. A recent report has also demonstrated O/HL serogroup associations for sporadic campylobacter isolates from the United States (32), where the patterns of serovar occurrence are broadly similar to those in the UK. These data suggest that common prevalent strains may cause a majority of campylobacter infections in both of these developed nations. The similarities between certain North American serogroup reference strains and sporadic UK isolates (18) (Tables 2 to 4) provide further support for this hypothesis.

Evidence of clonal relationships within three serovars indicates that linkage between the O and HL epitopes may represent a practical epidemiological marker. It is not known why specific O and HL antigens occur in stable, preferred associations, nor why only a small number of serovars account for a majority of isolates from human infections. The possession of specific surface-expressed serogroup characteristics may confer a general fitness phenotype, enabling colonization of a wide range of hosts. Alternatively, certain antigenic combinations may act as particularly effective adhesins, enhance invasiveness, affect complement binding and serum resistance, or otherwise function as virulence factors.

The three serovars analyzed for polyphasic markers show conservation of phage group and, to a lesser extent, biotype characteristics. The coincidence of specific phage groups within serovars may be the result of linkage, with phage adsorption characteristics directly influenced by the epitope configuration of either the lipopolysaccharide or flagellar antigens. Both somatic, lipopolysaccharide-dependent phage adsorption and serogroup-specific flagellotropic phages have been demonstrated in other species (16, 26). However, there are no wholly exclusive associations between any phage group and any given O/HL serovar, indicating that phage lysis patterns do vary independently of serogroup. For example, phage group 69 is prevalent among serovar O 23/HL 5 strains but also includes two strains of serovar O 2/HL 4 and one serovar O 50/HL 7 strain.

The biotype markers are more heterogeneous than the phage groups within conserved cell lines. Selective pressure on individual strains as a result of human or veterinary antibiotic usage may account for some of the resistotype variability within serovars. Selection may be particularly important if only single point mutations are necessary to alter any of the five antibiotic resistance characteristics of the Preston biotyping scheme. Plasmid- or phage-mediated transfer of resistance determinants can cause independent variation in resistotypes, and plasmid carriage may also influence phage susceptibility (40). The relatively labile nature of markers from both the Preston biotype and phage type schemes results in their low correlation with clonality (2) and with other strain markers (30), although their high discrimination indices substantiate their usefulness for primary epidemiological investigations.

Clonal associations have been well characterized within several somatic (O-antigen) and flagellar (H-antigen) serovars of the gut pathogen Salmonella enterica (36, 37). Similarly, the polyphasic analysis of strains from three C. jejuni O/HL serovars shows that these groups represent cell lines of common descent. For each serovar, strains sharing genetic and phenotypic characteristics have been isolated from a variety of sources, at different time periods, and from unrelated geographical locations within the UK and abroad. A recent study using flagellin gene typing (25) showed that allelic variation at the flaA locus for common O and HL serogroups of C. jejuni in the United States segregates substantially in accordance with the serovar associations reported here. This provides further evidence for clonal relationships within O/HL serovars. Two serovars which could not be identified in our study due to the use of a restricted set of HL antisera are evident from these data (25). Serogroup O 15 shows clear linkage to HL 13 (flaA type 30), and serogroup O 11 is linked to HL 40 (flaA type 27). The subsidiary association of flaA type 27 with a second O 11 serovar, O 11/HL 8, is also supported by our results (Table 1, row 7).

The clonal groupings identified here embrace strains which do show some differences in either genetic or phenotypic characteristics. Such variable isolates still have a lineage in common, and strains which show broad overall identity but limited variations in phenotype or genotype may more accurately be described as forming a clone complex or clonal subgrouping (3). A similar descriptive was proposed in a recent pulsed-field gel electrophoresis study of HS 1 isolates of C. jejuni (29) to classify strains which showed minor RFLPs with otherwise identical macrorestriction profiles.

As with S. enterica, there is evidence of polyphyletic variation within C. jejuni serovars. Two isolates of serovar O 2/HL 4 (Table 2, strains P/2255 and P/3546) and two of serovar O 50/HL 7 (Table 3, strains P/3758 and P/2578) appear otherwise to be unrelated to the clonal strains of these serovars. Major intragenomic rearrangements of DNA (11, 20) or the chromosomal integration of lysogenic phage (21) could generate this type of atypical restriction pattern diversity within serovars. However, horizontal transfer of serogroup determinants between strains of different genetic backgrounds has been proposed as the primary mechanism for polyphyletic variation in Salmonella (36, 37) and may also be significant in Campylobacter (6, 42). Lateral gene transfer could also account for the replacement of the serovar-characteristic HL 7 antigen with HL 1 and HL 20 epitopes in strains C/13383 and C/19584 (Table 3). Lastly, the possibility of convergent evolution of similar antigens in different cell lines cannot be excluded. Interestingly, the relationship between the O 50 and O 65 serogroup strains reported here (Table 3) is borne out by the results of macrorestriction profile data for these two isolates (13). Evidently, strains with O antigens that do not serologically cross-react can be genotypically closely related.

A recent study of campylobacter population genetics using multilocus enzyme electrophoresis predicted a moderately high frequency of intraspecific recombination for C. jejuni and C. coli, implying that these species should not be clonal (5). However, a clonal population structure has recently been proposed for C. coli (38), and in this report, we provide evidence that serovars represent lineages of common descent in C. jejuni. The apparent absence of genetic barriers to intraspecific recombination in C. jejuni and C. coli (5) suggests that the provenance of such clonal groupings could be recent, possibly due to the rapid proliferation and spread of specific pathogenic strains (23). Large-scale point source outbreaks (24) are one mechanism by which this type of spread could occur. Additionally, modern intensive methods of animal husbandry have been strongly implicated in the recent emergence of several zoonotic pathogens.

In this study, we have demonstrated that many human isolates of C. jejuni show strong associations between specific O and HL antigens and that most strains within three common serovars share genetic and phenotypic characteristics. Further investigations are necessary to characterize the extent of clonal relationships within these and other campylobacter serovars. Where genotypic identities are substantially conserved, rapid and simple phenotypic methods may be the most appropriate means for subtyping within serovars. We conclude that serogrouping with combined O and HL antigens represents both a practical and a phenologically valid method for the epidemiological typing of sporadic isolates of C. jejuni.