Reducing Campylobacter jejuni Colonization of Poultry via Vaccination

Campylobacter jejuni is a leading bacterial cause of human gastrointestinal disease worldwide. While C. jejuni is a commensal organism in chickens, case-studies have demonstrated a link between infection with C. jejuni and the consumption of foods that have been cross-contaminated with raw or undercooked poultry. We hypothesized that vaccination of chickens with C. jejuni surface-exposed colonization proteins (SECPs) would reduce the ability of C. jejuni to colonize chickens, thereby reducing the contamination of poultry products at the retail level and potentially providing a safer food product for consumers. To test our hypothesis, we injected chickens with recombinant C. jejuni peptides from CadF, FlaA, FlpA, CmeC, and a CadF-FlaA-FlpA fusion protein. Seven days following challenge, chickens were necropsied and cecal contents were serially diluted and plated to determine the number of C. jejuni per gram of material. The sera from the chickens were also analyzed to determine the concentration and specificity of antibodies reactive against the C. jejuni SECPs. Vaccination of chickens with the CadF, FlaA, and FlpA peptides resulted in a reduction in the number of C. jejuni in the ceca compared to the non-vaccinated C. jejuni-challenged group. The greatest reduction in C. jejuni colonization was observed in chickens injected with the FlaA, FlpA, or CadF-FlaA-FlpA fusion proteins. Vaccination of chickens with different SECPs resulted in the production of C. jejuni-specific IgY antibodies. In summary, we show that the vaccination of poultry with individual C. jejuni SECPs or a combination of SECPs provides protection of chickens from C. jejuni colonization.     

Molecular Epidemiology of Campylobacter jejuni in a Geographically Isolated Country with a Uniquely Structured Poultry Industry

In New Zealand the number of campylobacteriosis notifications increased markedly between 2000 and 2007. Notably, this country''s poultry supply is different than that of many developed countries as the fresh and frozen poultry available at retail are exclusively of domestic origin. To examine the possible link between human cases and poultry, a sentinel surveillance site was established to study the molecular epidemiology of Campylobacter jejuni over a 3-year period from 2005 to 2008 using multilocus sequence typing. Studies showed that 60.1 to 81.4% of retail poultry carcasses from the major suppliers were contaminated with C. jejuni. Differences were detected in the probability and level of contamination and the relative frequency of genotypes for individual poultry suppliers and humans. Some carcasses were contaminated with isolates belonging to more than one sequence type (ST), and there was evidence of both ubiquitous and supplier-associated strains, an epidemiological pattern not recognized yet in other countries. The common poultry STs were also common in human clinical cases, providing evidence that poultry is a major contributor to human infection. Both internationally rare genotypes, such as ST-3069 and ST-474, and common genotypes, such as ST-45 and ST-48, were identified in this study. The dominant human sequence type in New Zealand, ST-474, was found almost exclusively in isolates from one poultry supplier, which provided evidence that C. jejuni has a distinctive molecular epidemiology in this country. These results may be due in part to New Zealand''s geographical isolation and its uniquely structured poultry industry.Campylobacteriosis is a leading enteric zoonosis in the developed world, and the majority of cases are caused by Campylobacter jejuni (22). Poultry sources are suspected to be the major source of human infection with C. jejuni (22, 43, 49), and this conclusion is supported by high levels of contamination of poultry and the detection of identical C. jejuni genotypes in human cases and poultry samples (25). However, C. jejuni can be isolated from a variety of sources, including ruminants (17) and environmental water (41). Due to the complex epidemiology of this pathogen, there is still uncertainty about the relative contributions of individual pathways to the human disease burden (8). In New Zealand the number of notified campylobacteriosis cases increased markedly in the last decade, peaking in 2006 at a total of 15,873 notified cases (422.4 cases per 100,000 population) (1) and costing the economy an estimated US$32 million annually (42). Although there are a number of factors that may have contributed to this increase, it has been noted that the rise in campylobacteriosis cases coincided with a marked increase in the sale and consumption of fresh poultry between 1992 and 2005, while the sale of frozen poultry remained relatively static (1).New Zealand has one of the highest enteric infectious disease rates in industrialized countries (26), and the high ratio of domestic production animals to humans and the frequent use of rural water supplies in New Zealand have been postulated to be underlying causes (9). In addition, this country''s poultry industry is uniquely structured; it is almost entirely focused on the domestic market, and no raw, fresh or frozen, poultry products are imported because of biosecurity threats. Due to its geographical isolation and tight border controls, New Zealand has remained free of poultry diseases endemic in other countries, such as diseases caused by Salmonella enterica serovar Enteritidis PT4 and S. enterica serovar Typhimurium DT 104, Newcastle disease, and infectious bursal disease.The production of poultry meat in New Zealand is highly integrated; only three companies supply 90% of the chicken meat, which represents 95% of the poultry meat consumed. The remaining 5% of poultry meat consumed includes species such as turkey and duck. The chicken processors own or control most stages of production, processing, and distribution. One of the three dominant companies has one processing plant that distributes nationwide, and one company has multiple plants that tend to be more localized in their distribution, except when they make specialty products, which are distributed nationally. The other companies distribute primarily locally. The broilers are commonly barn raised, not free range, and animal welfare standards require a maximum stocking density of 38 kg (live weight) of broiler chickens per m². There are approximately 160 broiler farms in a number of specific areas of New Zealand. These farms are usually located near the slaughterhouses that they supply.To enable regulators to implement food safety programs to reduce human campylobacteriosis, there has been great interest in understanding the importance and epidemiology of C. jejuni in the New Zealand poultry production system. A sentinel surveillance site was therefore established in the Manawatu region to quantify the contributions of different sources, including poultry suppliers, to the human disease burden and to study the molecular epidemiology of C. jejuni (33, 34). Isolates were typed using multilocus sequence typing (MLST), a method that has major advantages over other methods of genotyping when the long-term epidemiology of a disease is studied. However, other methods may be more appropriate in other settings, such as outbreak investigations (21), where a higher degree of discrimination may be required. MLST offers a large web-based archive of isolates from many sources and countries: the Campylobacter PubMLST database (10). Sequence typing by MLST is now internationally recognized as a valuable approach for national and international epidemiological characterization and source tracking of major pathogenic microorganisms, such as C. jejuni (46, 48).The use of integrated surveillance across human, domestic animal, and wildlife populations has been identified as a key component of strategies aimed at prevention and control of emerging pathogens, particularly when the population dynamics of multihost pathogens are poorly understood (52). At the Manawatu surveillance site, samples from human clinical cases, animal-derived food products, and the environment were gathered in a defined geographical area of New Zealand over a 3-year period (19, 34) and genotyped using MLST (11). The resulting data set contained a total of 969 typed samples, 502 of which were from human cases. The temporal and spatial scale of the data allowed us to obtain a more in-depth understanding of local transmission dynamics compared with the results of previous research (43, 50). The application of novel risk attribution approaches to these data previously identified poultry as the major contributor to the human disease burden, with widely varying contributions from different suppliers (33).In this study we extended the findings of risk attribution and epidemiological studies of human cases (33, 34). Data from the sentinel surveillance site were used to study the epidemiology of C. jejuni for the individual producers that comprise the poultry sector in New Zealand and to better understand the contributions of the producers to the human campylobacteriosis burden. Our study included an investigation of both the probability of contamination and the level of contamination of poultry carcasses and a study of human and poultry MLST sequence types (STs). The resulting data were compared to make inferences about the epidemiology of C. jejuni in the New Zealand poultry industry and to identify determinants for the high number of human cases attributed to this food source.     

Genomic Characterization of Campylobacter jejuni Strain M1

Campylobacter jejuni strain M1 (laboratory designation 99/308) is a rarely documented case of direct transmission of C. jejuni from chicken to a person, resulting in enteritis. We have sequenced the genome of C. jejuni strain M1, and compared this to 12 other C. jejuni sequenced genomes currently publicly available. Compared to these, M1 is closest to strain 81116. Based on the 13 genome sequences, we have identified the C. jejuni pan-genome, as well as the core genome, the auxiliary genes, and genes unique between strains M1 and 81116. The pan-genome contains 2,427 gene families, whilst the core genome comprised 1,295 gene families, or about two-thirds of the gene content of the average of the sequenced C. jejuni genomes. Various comparison and visualization tools were applied to the 13 C. jejuni genome sequences, including a species pan- and core genome plot, a BLAST Matrix and a BLAST Atlas. Trees based on 16S rRNA sequences and on the total gene families in each genome are presented. The findings are discussed in the background of the proven virulence potential of M1.     

Genetic Diversity and Quinolone Resistance in Campylobacter jejuni Isolates from Poultry in Senegal

We used the multilocus sequence typing (MLST) method to evaluate the genetic diversity of 46 Campylobacter jejuni isolates from chickens and to determine the link between quinolone resistance and sequence type (ST). There were a total of 16 ST genotypes, and the majority of them belonged to seven clonal complexes previously identified by using isolates from human disease. The ST-353 complex was the most common complex, whereas the ST-21, ST-42, ST-52, and ST-257 complexes were less well represented. The resistance phenotype varied for each ST, and the Thr-86-Ile substitution in the GyrA protein was the predominant mechanism of resistance to quinolone. Nine of the 14 isolates having the Thr-86-Ile substitution belonged to the ST-353 complex. MLST showed that the emergence of quinolone resistance is not related to the diffusion of a unique clone and that there is no link between ST genotype and quinolone resistance. Based on silent mutations, different variants of the gyrA gene were shown to exist for the same ST. These data provide useful information for understanding the epidemiology of C. jejuni in Senegal.     

Arsenic Resistance in Campylobacter spp. Isolated from Retail Poultry Products

Organoarsenicals are commonly used for growth promotion in U.S. poultry production. Susceptibilities to arsenite, arsenate, and the organoarsenical roxarsone were measured in 251 Campylobacter isolates from conventional and antimicrobial-free retail poultry products. Isolates from conventional poultry products had significantly higher roxarsone MICs (z = 8.22; P < 0.0001).     

Campylobacter fetus Subspecies jejuni (Vibrio fetus) from Commercially Processed Poultry

Three isolates of the human and animal pathogen Campylobacter fetus ss. jejuni (Vibrio fetus) were obtained from 165 poultry meat samples purchased from local retail stores.     

Genomic investigation of phenotypic variation in Campylobacter jejuni flagellin.

Western blots of whole-cell sonicates of 10 different clones of a faecal isolate of Campylobacter jejuni 533 detected the expression of flagella antigens of either 59 or 62 kDa. Other antigenic proteins appeared identical both in the parent and all the clones. The mechanism for this phenotypic variation was studied using Southern blotting with a flagellin-specific gene probe and products of a polymerase chain reaction (PCR) using flagellin-gene primers. Restriction-enzyme digestion and Southern blotting did not detect any genomic rearrangements in the flagellin genes of the different phenotypes nor did restriction-enzyme analysis of the PCR products.     

Genomic Insights into the Convergence and Pathogenicity Factors of Campylobacter jejuni and Campylobacter coli Species

Whether or not bacteria form coherent evolutionary groups via means of genetic exchange and, hence, elicit distinct species boundaries remains an unsettled issue. A recent report implied that not only may the former be true but also, in fact, the clearly distinct Campylobacter jejuni and Campylobacter coli species may be converging as a consequence of increased interspecies gene flow fostered, presumably, by the recent invasion of an overlapping ecological niche (S. K. Sheppard, N. D. McCarthy, D. Falush, and M. C. Maiden, Science 320:237-239, 2008). We have reanalyzed the Campylobacter multilocus sequence typing database used in the previous study and found that the number of interspecies gene transfer events may actually be too infrequent to account, unequivocally, for species convergence. For instance, only 1 to 2% of the 4,507 Campylobacter isolates examined appeared to have imported gene alleles from another Campylobacter species. Furthermore, by analyzing the available Campylobacter genomic sequences, we show that although there seems to be a slightly higher number of exchanged genes between C. jejuni and C. coli relative to other comparable species (∼10% versus 2 to 3% of the total genes in the genome, respectively), the function and spatial distribution in the genome of the exchanged genes are far from random, and hence, inconsistent with the species convergence hypothesis. In fact, the exchanged genes appear to be limited to a few environmentally selected cellular functions. Accordingly, these genes may represent important pathogenic determinants of pathogenic Campylobacter, and convergence of (any) two bacterial species remains to be seen.High-throughput sequencing studies during the last decade have revealed that bacterial genomes are much more diverse and “fluid” than previously anticipated (14, 31). This genomic fluidity is primarily attributable to the great pervasiveness and promiscuity of horizontal gene transfer (HGT) in the bacterial world (5, 17). Nonetheless, evidence of any two distinct bacterial species or lineages merging due to directed (as opposed to promiscuous) interspecies genetic exchange was reported, probably for the first time ever, by the recent study of Sheppard et al. (26). Species convergence, if occurring, has major theoretical implications for the bacterial species concept (reviewed extensively elsewhere [9, 10, 14, 24, 30]) and important practical consequences for accurate identification of bacterial pathogens in the clinical setting.Sheppard and colleagues reported that as many as ∼18.6% of the unique alleles of housekeeping genes found in Campylobacter coli isolates may have been recently imported (through HGT) from a close relative, Campylobacter jejuni (26). The results were based on the analysis of 4,507 Campylobacter spp. isolates, which were genotyped at seven genes (loci), available though the Campylobacter multilocus sequence typing (MLST) database (4). In brief, the 4,507 genotyped isolates contained a total of 2,917 unique sequence types (STs). A unique ST represents the concatenated sequence of the seven genes present in the genome of an isolate and contains a unique sequence (allele) for at least one of the seven genes when compared against any other unique ST in the database (different isolates may be characterized by the same ST). The unique STs were assigned to either C. coli or C. jejuni species by using the program STRUCTURE (6). Neighbor-joining phylogenetic trees of all available unique alleles for each individual gene were subsequently built. Instances where the ST assignment to a species differed from the assignment of an individual gene sequence, which comprised the ST, were attributed to interspecies transfer of the gene, and the number of such instances was reported (26).Here, we have reevaluated the available Campylobacter MLST data set and show that the predominant STs, i.e., the STs characterizing >98% of the isolates, do not contain imported alleles and, hence, do not support the species convergence hypothesis. In agreement with these findings, analyses of the available Campylobacter genomic sequences indicate that the interspecies genetic exchange is limited and heavily biased toward a few genes under positive selection. In fact, housekeeping genes (such as those used in MLST) were found to be exchanged between the two species only in (rare) hitchhiking events associated with the horizontal transfer of adaptive genes. Accordingly, a clear species boundary between the C. jejuni and C. coli species is evident and it is unlikely that this boundary is being eroded.     

Genomic characterization of the Guillain-Barre syndrome-associated Campylobacter jejuni ICDCCJ07001 Isolate

Campylobacter jejuni ICDCCJ07001 (HS:41, ST2993) was isolated from a Guillain-Barré syndrome (GBS) patient during a 36-case GBS outbreak triggered by C. jejuni infections in north China in 2007. Sequence analysis revealed that the ICDCCJ07001 genome consisted of 1,664,840 base pairs (bp) and one tetracycline resistance plasmid of 44,084 bp. The GC content was 59.29% and 1,579 and 37 CDSs were identified on the chromosome and plasmid, respectively. The ICDCCJ07001 genome was compared to C. jejuni subsp. jejuni strains 81-176, 81116, NCTC11168, RM1221 and C. jejuni subsp. doylei 269.97. The length and organization of ICDCCJ07001 was similar to that of NCTC11168, 81-176 and 81-116 except that CMLP1 had a reverse orientation in strain ICDCCJ07001. Comparative genomic analyses were also carried out between GBS-associated C. jejuni strains. Thirteen common genes were present in four GBS-associated strains and 9 genes mapped to the LOS cluster and the ICDCCJ07001_pTet (44 kb) plasmid was mosaic in structure. Thirty-seven predicted CDS in ICDCCJ07001_pTet were homologous to genes present in three virulence-associated plasmids in Campylobacter: 81-176_pTet, pCC31 and 81-176_pVir. Comparative analysis of virulence loci and virulence-associated genes indicated that the LOS biosynthesis loci of ICDCCJ07001 belonged to type A, previously reported to be associated with cases of GBS. The polysaccharide capsular biosynthesis (CPS) loci and the flagella modification (FM) loci of ICDCCJ07001 were similar to corresponding sequences of strain 260.94 of similar serotype as strain ICDCCJ07001. Other virulence-associated genes including cadF, peb1, jlpA, cdt and ciaB were conserved between the C. jejuni strains examined.     

Evidence for a Genetically Stable Strain of Campylobacter jejuni

The genetic stability of selected epidemiologically linked strains of Campylobacter jejuni during outbreak situations was investigated by using subtyping techniques. Strains isolated from geographically related chicken flock outbreaks in 1998 and from a human outbreak in 1981 were investigated. There was little similarity in the strains obtained from the different chicken flock outbreaks; however, the strains from each of three chicken outbreaks, including strains isolated from various environments, were identical as determined by fla typing, amplified fragment length polymorphism (AFLP) analysis, and pulsed-field gel electrophoresis, which confirmed the genetic stability of these strains during the short time courses of chicken flock outbreaks. The human outbreak samples were compared with strain 81116, which originated from the same outbreak but has since undergone innumerable laboratory passages. Two main AFLP profiles were recognized from this outbreak, which confirmed the serotyping results obtained at the time of the outbreak. The major type isolated from this outbreak (serotype P6:L6) was exemplified by strain 81116. Despite the long existence of strain 81116 as a laboratory strain, the AFLP profile of this strain was identical to the profiles of all the other historical P6:L6 strains from the outbreak, indicating that the genotype has remained stable for almost 20 years. Interestingly, the AFLP profiles of the P6:L6 group of strains from the human outbreak and the strains from one of the recent chicken outbreaks were also identical. This similarity suggests that some clones of C. jejuni remain genetically stable in completely different environments over long periods of time and considerable geographical distances.     

Evidence for a genetically stable strain of Campylobacter jejuni

The genetic stability of selected epidemiologically linked strains of Campylobacter jejuni during outbreak situations was investigated by using subtyping techniques. Strains isolated from geographically related chicken flock outbreaks in 1998 and from a human outbreak in 1981 were investigated. There was little similarity in the strains obtained from the different chicken flock outbreaks; however, the strains from each of three chicken outbreaks, including strains isolated from various environments, were identical as determined by fla typing, amplified fragment length polymorphism (AFLP) analysis, and pulsed-field gel electrophoresis, which confirmed the genetic stability of these strains during the short time courses of chicken flock outbreaks. The human outbreak samples were compared with strain 81116, which originated from the same outbreak but has since undergone innumerable laboratory passages. Two main AFLP profiles were recognized from this outbreak, which confirmed the serotyping results obtained at the time of the outbreak. The major type isolated from this outbreak (serotype P6:L6) was exemplified by strain 81116. Despite the long existence of strain 81116 as a laboratory strain, the AFLP profile of this strain was identical to the profiles of all the other historical P6:L6 strains from the outbreak, indicating that the genotype has remained stable for almost 20 years. Interestingly, the AFLP profiles of the P6:L6 group of strains from the human outbreak and the strains from one of the recent chicken outbreaks were also identical. This similarity suggests that some clones of C. jejuni remain genetically stable in completely different environments over long periods of time and considerable geographical distances.     

Phenotypic and Genotypic Characterizations of Campylobacter jejuni Isolated from the Broiler Meat Production Process

A set of C. jejuni isolates of different origins and flaA-genotypes obtained throughout the broiler meat production chain was tested in this study for a possible correlation of their origin, phylogenetic relationship, and phenotypic properties. Interestingly, the results showed a correlation of the origin and the phylogenetic relationship between the C. jejuni isolates and their ability to form biofilm, but not in their ability to survive at -18, 5, 20, and 48?°C. Two strains, a broiler cloacae isolate and a broiler fillet isolate, were unable to develop biofilm, while most of the C. jejuni isolates originating from meat and surfaces of the slaughterhouse readily formed biofilms after both 24, 48, and 72?h. Interestingly, these biofilm-forming strains were closely related. Furthermore, two strains that were isolated after disinfection developed significantly more biofilms after 24?h of incubation than the remaining strains. A comparative genomic analysis using DNA microarrays showed that the gene contents of strains that efficiently formed biofilms were different from those that did not. The study suggests that biofilm formation might be a lineage specific property, allowing C. jejuni to both survive environmental stress at the slaughterhouse and to attach to the surface of meat.     

Isolation of Campylobacter jejuni from groundwater

A pollution event which occurred at a spring in the Arnside area of Cumbria provided an opportunity to investigate whether Campylobacter jejuni could be detected in groundwater. Hydrological evidence suggested that the source of contamination was a dairy farm situated within the hydrological catchment of the polluted spring. The microbiological quality of the polluted spring was monitored during intervals over the following 12 months and compared with others in the area. Campylobacter jejuni was isolated by filter enrichment of 500 ml and 100 ml filtered volumes of groundwater. It was not isolated in the absence of faecal indicator species. Some strains of Camp. jejuni from water had identical biotypes to strains isolated from the dairy herd. This paper reports the first isolation of Camp. jejuni from groundwater using cultural methods and supports the theory that groundwater may be a vehicle for Campylobacter transmission.     

The Effect of Lactic Acid Sprays on Campylobacter jejuni Inoculated onto Poultry Carcasses

Spraying poultry carcasses with 1 % lactic acid 10 min after inoculation with Campylobacter jejuni, resulted in a significant reduction in the number of the bacteria after 4 h at 4°C. Some of the inoculated cells, however, survived for at least 144 h. Spraying 10 min after inoculation with 2% lactic acid, totally eliminated all inoculated C. jejuni within 24 h. On the other hand, spraying 24 h after inoculation, with either 1 % or 2 % lactic acid did not eliminate all the bacteria. Inoculated C. jejuni on poultry carcasses not sprayed with lactic acid, survived at 4°C throughout the sampling period (up to 144 h) and showed little tendency to decrease in number even when the carcasses started to deteriorate. Resident Campylobacters on poultry carcasses were significantly reduced by the lactic acid treatment. Frozen and thawed chickens appeared to show a graying of the skins immediately after spraying with lactic acid, slightly stronger with 2 % lactic acid, but the colour reverted to normal after 24 h. We were not able to observe any colour change on the fresh broiler chickens after lactic acid treatment. Our results indicated that lactic acid had a significant bactericidal effect on C. jejuni on both naturally and artificially contaminated poultry carcasses. This effect, however, became manifest only several hours after acid treatment.     

Evidence for a system of general protein glycosylation in Campylobacter jejuni

A genetic locus from Campylobacter jejuni 81-176 (O:23, 36) has been characterized that appears to be involved in glycosylation of multiple proteins, including flagellin. The lipopolysaccharide (LPS) core of Escherichia coli DH5alpha containing some of these genes is modified such that it becomes immunoreactive with O:23 and O:36 antisera and loses reactivity with the lectin wheat germ agglutinin (WGA). Site-specific mutation of one of these genes in the E. coli host causes loss of O:23 and O:36 antibody reactivity and restores reactivity with WGA. However, site-specific mutation of each of the seven genes in 81-176 failed to show any detectable changes in LPS. Multiple proteins from various cellular fractions of each mutant showed altered reactivity by Western blot analyses using O:23 and O:36 antisera. The changes in protein antigenicity could be restored in one of the mutants by the presence of the corresponding wild-type allele in trans on a shuttle vector. Flagellin, which is known to be a glycoprotein, was one of the proteins that showed altered reactivity with O:23 and O:36 antiserum in the mutants. Chemical deglycosylation of protein fractions from the 81-176 wild type suggests that the other proteins with altered antigenicity in the mutants are also glycosylated.     

Genomic variations define divergence of water/wildlife-associated Campylobacter jejuni niche specialists from common clonal complexes

Although the major food-borne pathogen Campylobacter jejuni has been isolated from diverse animal, human and environmental sources, our knowledge of genomic diversity in C. jejuni is based exclusively on human or human food-chain-associated isolates. Studies employing multilocus sequence typing have indicated that some clonal complexes are more commonly associated with particular sources. Using comparative genomic hybridization on a collection of 80 isolates representing diverse sources and clonal complexes, we identified a separate clade comprising a group of water/wildlife isolates of C. jejuni with multilocus sequence types uncharacteristic of human food-chain-associated isolates. By genome sequencing one representative of this diverse group (C. jejuni 1336), and a representative of the bank-vole niche specialist ST-3704 (C. jejuni 414), we identified deletions of genomic regions normally carried by human food-chain-associated C. jejuni. Several of the deleted regions included genes implicated in chicken colonization or in virulence. Novel genomic insertions contributing to the accessory genomes of strains 1336 and 414 were identified. Comparative analysis using PCR assays indicated that novel regions were common but not ubiquitous among the water/wildlife group of isolates, indicating further genomic diversity among this group, whereas all ST-3704 isolates carried the same novel accessory regions. While strain 1336 was able to colonize chicks, strain 414 was not, suggesting that regions specifically absent from the genome of strain 414 may play an important role in this common route of Campylobacter infection of humans. We suggest that the genomic divergence observed constitutes evidence of adaptation leading to niche specialization.     

Genomic Diversity of Campylobacter coli and Campylobacter jejuni Isolates Recovered from Free-Range Broiler Farms and Comparison with Isolates of Various Origins

In many industrialized countries, the incidence of campylobacteriosis exceeds that of salmonellosis. Campylobacter bacteria are transmitted to humans mainly in food, especially poultry meat products. Total prevention of Campylobacter colonization in broiler flocks is the best way to reduce (or eliminate) the contamination of poultry products. The aim of this study was to establish the sources and routes of contamination of broilers at the farm level. Molecular typing methods (DNA macrorestriction pulsed-field gel electrophoresis and analysis of gene polymorphism by PCR-restriction fragment length polymorphism) were used to characterize isolates collected from seven broiler farms. The relative genomic diversity of Campylobacter coli and Campylobacter jejuni was determined. Analysis of the similarity among 116 defined genotypes was used to determine clusters within the two species. Furthermore, evidence of recombination suggested that there were genomic rearrangements within the Campylobacter populations. Recovery of related clusters from different broiler farms showed that some Campylobacter strains might be specifically adapted to poultry. Analysis of the Campylobacter cluster distribution on three broiler farms showed that soil in the area around the poultry house was a potential source of Campylobacter contamination. The broilers were infected by Campylobacter spp. between days 15 and 36 during rearing, and the type of contamination changed during the rearing period. A study of the effect of sanitary barriers showed that the chickens stayed Campylobacter spp. free until they had access to the open area. They were then rapidly colonized by the Campylobacter strains isolated from the soil.     

Genomic analysis of the diversity,antimicrobial resistance and virulence potential of clinical Campylobacter jejuni and Campylobacter coli strains from Chile

Campylobacter jejuni and Campylobacter coli are the leading cause of human gastroenteritis in the industrialized world and an emerging threat in developing countries. The incidence of campylobacteriosis in South America is greatly underestimated, mostly due to the lack of adequate diagnostic methods. Accordingly, there is limited genomic and epidemiological data from this region. In the present study, we performed a genome-wide analysis of the genetic diversity, virulence, and antimicrobial resistance of the largest collection of clinical C. jejuni and C. coli strains from Chile available to date (n = 81), collected in 2017–2019 in Santiago, Chile. This culture collection accounts for more than one third of the available genome sequences from South American clinical strains. cgMLST analysis identified high genetic diversity as well as 13 novel STs and alleles in both C. jejuni and C. coli. Pangenome and virulome analyses showed a differential distribution of virulence factors, including both plasmid and chromosomally encoded T6SSs and T4SSs. Resistome analysis predicted widespread resistance to fluoroquinolones, but low rates of erythromycin resistance. This study provides valuable genomic and epidemiological data and highlights the need for further genomic epidemiology studies in Chile and other South American countries to better understand molecular epidemiology and antimicrobial resistance of this emerging intestinal pathogen.     

Genetic Diversity and Antibiotic Resistance Patterns in a Campylobacter Population Isolated from Poultry Farms in Switzerland

The diversity and genetic interrelation of Campylobacter jejuni and C. coli isolated from Swiss poultry were assessed by three independent typing methods. Samples were derived prior to slaughter from 100 randomly selected flocks (five birds per flock) raised on three different farm types. The observed flock prevalence was 54% in total, with 50% for conventional and 69% for free-range farms. Birds held on farms with a confined roaming area had the lowest prevalence of 37%. Campylobacter isolates were characterized by amplified fragment length polymorphism (AFLP), restriction fragment length polymorphism of flaA PCR fragments (flaA-RFLP), and disk diffusion testing for eight antimicrobial agents that are commonly used in veterinary or human medicine in Switzerland. Analysis of the genotypic results indicates that the Campylobacter population in Swiss poultry is genetically highly diverse. Nevertheless, occasionally, isolates with identical or nearly identical characteristics were isolated from different farms or farm types in different locations. Genetic typing by AFLP and flaA-RFLP was found to be complementary. The majority of isolates (67%) were susceptible to all tested antibiotics; however, single, double, and triple resistances were observed in 7%, 23%, and 2% of the strains, respectively. There was no correlation between genotype and antibiotic resistance. Surprisingly, sulfonamide resistance was frequently found together with streptomycin resistance. Our findings illustrate the results of common genetic exchange in the studied bacterial population.