Campylobacter jejuni Strains Compete for Colonization in Broiler Chicks

Campylobacter jejuni isolates possess multiple adhesive proteins termed adhesins, which promote the organism's attachment to epithelial cells. Based on the proposal that one or more adhesins are shared among C. jejuni isolates, we hypothesized that C. jejuni strains would compete for intestinal and cecal colonization in broiler chicks. To test this hypothesis, we selected two C. jejuni strains with unique SmaI pulsed-field gel electrophoresis macrorestriction profiles and generated one nalidixic acid-resistant strain (the F38011 Nal^r strain) and one streptomycin-resistant strain (the 02-833L Str^r strain). In vitro binding assays revealed that the C. jejuni F38011 Nal^r and 02-833L Str^r strains adhered to LMH chicken hepatocellular carcinoma epithelial cells and that neither strain influenced the binding potential of the other strain at low inoculation doses. However, an increase in the dose of the C. jejuni 02-833L Str^r strain relative to that of the C. jejuni F38011 Nal^r strain competitively inhibited the binding of the C. jejuni F38011 Nal^r strain to LMH cells in a dose-dependent fashion. Similarly, the C. jejuni 02-833L Str^r strain was found to significantly reduce the efficiency of intestinal and cecal colonization by the C. jejuni F38011 Nal^r strain in broiler chickens. Based on the number of bacteria recovered from the ceca, the maximum number of bacteria that can colonize the digestive tracts of chickens may be limited by host constraints. Collectively, these data support the hypothesis that C. jejuni strains compete for colonization in chicks and suggest that it may be possible to design novel intervention strategies for reducing the level at which C. jejuni colonizes the cecum.     

Dissemination of fluoroquinolone-resistant Campylobacter spp. within an integrated commercial poultry production system

While characterizing the intestinal bacterial community of broiler chickens, we detected epsilon-proteobacterial DNA in the ilea of 3-day-old commercial broiler chicks (J. Lu, U. Idris, B. Harmon, C. Hofacre, J. J. Maurer, and M. D. Lee, Appl. Environ. Microbiol. 69:6816-6824, 2003). The sequences exhibited high levels of similarity to Campylobacter jejuni and Campylobacter coli sequences, suggesting that chickens can carry Campylobacter at a very young age. Campylobacter sp. was detected by PCR in all samples collected from the ilea of chicks that were 3 to 49 days old; however, it was detected only in the cecal contents of chickens that were at least 21 days old. In order to determine whether the presence of Campylobacter DNA in young chicks was due to ingestion of the bacteria in food or water, we obtained commercial broiler hatching eggs, which were incubated in a research facility until the chicks hatched. DNA sequencing of the amplicons resulting from Campylobacter-specific 16S PCR performed with the ileal, cecal, and yolk contents of the day-of-hatching chicks revealed that Campylobacter DNA was present before the chicks consumed food or water. The 16S rRNA sequences exhibited 99% similarity to C. jejuni and C. coli sequences and 95 to 98% similarity to sequences of other thermophilic Campylobacter species, such as C. lari and C. upsaliensis. The presence of C. coli DNA was detected by specific PCR in the samples from chicks obtained from a commercial hatchery; however, no Campylobacter was detected by culturing. In order to determine whether the same strains of bacteria were present in multiple levels of the integrator, we cultured Campylobacter sp. from a flock of broiler breeders and their 6-week-old progeny that resided on a commercial broiler farm. The broiler breeders had been given fluoroquinolone antibiotics, and we sought to determine whether the same fluoroquinolone-resistant strain was present in their progeny. The isolates were typed by pulsed-field gel electrophoresis, which confirmed that the parental and progeny flocks contained the same strain of fluoroquinolone-resistant C. coli. These data indicate that resistant C. coli can be present in multiple levels of an integrated poultry system and demonstrated that molecular techniques or more sensitive culture methods may be necessary to detect early colonization by Campylobacter in broiler chicks.     

Bacteriophage influence Campylobacter jejuni types populating broiler chickens

The characteristics that allow one Campylobacter jejuni genotype to succeed over another under the influence of bacteriophage predation have been examined in experimental broiler chickens following the observation that this succession appeared to occur in naturally colonized broiler chicken flocks. Examination of three C. jejuni strains from a single flock indicated that horizontal transfer of at least 112 kb of genomic DNA from strain F2C10 (bacteriophage sensitive) to strain F2E1 (bacteriophage insensitive) had created strain F2E3. Transfer of this DNA was associated with acquisition of sensitivity to 6 of 25 lytic bacteriophage isolated from the same flock. All strains tested were capable of colonizing broiler chickens but cocolonization revealed that the bacteriophage sensitive strains F2E3 and F2C10 had a competitive advantage over the bacteriophage insensitive strain F2E1. With the addition of lytic bacteriophage the situation was completely reversed, with F2E1 dominating. The inability to replicate bacteriophage is associated with a significant fitness cost that renders the insensitive strain competitive only in the presence of bacteriophage. We demonstrate that interstrain recombination in vivo can generate genome diversity in C. jejuni and that bacteriophage predation is a strong selective pressure that influences the relative success of emergent strains in broiler chickens.     

The Campylobacter jejuni response regulator, CbrR, modulates sodium deoxycholate resistance and chicken colonization

Two-component regulatory systems play a major role in the physiological response of bacteria to environmental stimuli. Such systems are composed of a sensor histidine kinase and a response regulator whose ultimate function is to affect the expression of target genes. Response regulator mutants of Campylobacter jejuni strain F38011 were screened for sensitivity to sodium deoxycholate. A mutation in Cj0643, which encodes a response regulator with no obvious cognate histidine kinase, resulted in an absence of growth on plates containing a subinhibitory concentration of sodium deoxcholate (1%, wt/vol). In broth cultures containing 0.05% (wt/vol) sodium deoxycholate, growth of the mutant was significantly inhibited compared to growth of the C. jejuni F38011 wild-type strain. Complementation of the C. jejuni cbrR mutant in trans restored growth in both broth and plate cultures supplemented with sodium deoxycholate. Based on the phenotype displayed by its mutation, we designated the gene corresponding to Cj0643 as cbrR (Campylobacter bile resistance regulator). While the MICs of a variety of bile salts and other detergents for the C. jejuni cbrR mutant were lower, no difference was noted in its sensitivity to antibiotics or osmolarity. Finally, chicken colonization studies demonstrated that the C. jejuni cbrR mutant had a reduced ability to colonize compared to the wild-type strain. These data support previous findings that bile resistance contributes to colonization of chickens and establish that the response regulator, CbrR, modulates resistance to bile salts in C. jejuni.     

Reduction of Campylobacter jejuni colonization of chicks by cecum-colonizing bacteria producing anti-C. jejuni metabolites.

Cecum-colonizing bacteria were isolated from Campylobacter jejuni-free White Leghorn (Gallus domesticus) laying hens and screened for the ability to produce anti-C. jejuni metabolites. Nine isolates were obtained that possessed this characteristic. The peroral administration of the nine isolates as a mixture (ca. 10(9) per chick) to 1-day-old chicks was followed 1 week later by peroral inoculation of Campylobacter jejuni (ca. 10(9) per chick) to determine if the cecal isolates could protect chicks from colonization by campylobacters. The nine-strain mixture of cecal bacteria provided from 41 to 85% protection from C. jejuni colonization. The protective bacteria were reduced to a mixture of three strains on the basis of their ability to utilize mucin as a sole substrate for growth. These strains included Klebsiella pneumoniae 23, Citrobacter diversus 22, and Escherichia coli (O13:H-) 25. Four feeding trials with this three-strain mixture provided from 43 to 100% (average, 78%) protection from C. jejuni colonization. The dominant cecal bacterium of chicks treated with the three-strain mixture was consistently E. coli O13:H-. Similarly, three trials with only E. coli 25 used as the protective bacterium resulted in 49 to 72% (average, 59%) protection from C. jejuni colonization, with E. coli O13:H- being the dominant cecal bacterium in all cases. Although not completely effective, E. coli 25 substantially reduced the incidence of C. jejuni colonization of chicks. For all trials, fewer C. jejuni were present in the ceca of colonized chicks receiving the protective bacteria before exposure to C. jejuni than in chicks receiving only C. jejuni.(ABSTRACT TRUNCATED AT 250 WORDS)     

Reduction of Campylobacter jejuni colonization of chicks by cecum-colonizing bacteria producing anti-C. jejuni metabolites.

Cecum-colonizing bacteria were isolated from Campylobacter jejuni-free White Leghorn (Gallus domesticus) laying hens and screened for the ability to produce anti-C. jejuni metabolites. Nine isolates were obtained that possessed this characteristic. The peroral administration of the nine isolates as a mixture (ca. 10(9) per chick) to 1-day-old chicks was followed 1 week later by peroral inoculation of Campylobacter jejuni (ca. 10(9) per chick) to determine if the cecal isolates could protect chicks from colonization by campylobacters. The nine-strain mixture of cecal bacteria provided from 41 to 85% protection from C. jejuni colonization. The protective bacteria were reduced to a mixture of three strains on the basis of their ability to utilize mucin as a sole substrate for growth. These strains included Klebsiella pneumoniae 23, Citrobacter diversus 22, and Escherichia coli (O13:H-) 25. Four feeding trials with this three-strain mixture provided from 43 to 100% (average, 78%) protection from C. jejuni colonization. The dominant cecal bacterium of chicks treated with the three-strain mixture was consistently E. coli O13:H-. Similarly, three trials with only E. coli 25 used as the protective bacterium resulted in 49 to 72% (average, 59%) protection from C. jejuni colonization, with E. coli O13:H- being the dominant cecal bacterium in all cases. Although not completely effective, E. coli 25 substantially reduced the incidence of C. jejuni colonization of chicks. For all trials, fewer C. jejuni were present in the ceca of colonized chicks receiving the protective bacteria before exposure to C. jejuni than in chicks receiving only C. jejuni.(ABSTRACT TRUNCATED AT 250 WORDS)     

Bacteriophage therapy to reduce Campylobacter jejuni colonization of broiler chickens

Colonization of broiler chickens by the enteric pathogen Campylobacter jejuni is widespread and difficult to prevent. Bacteriophage therapy is one possible means by which this colonization could be controlled, thus limiting the entry of campylobacters into the human food chain. Prior to evaluating the efficacy of phage therapy, experimental models of Campylobacter colonization of broiler chickens were established by using low-passage C. jejuni isolates HPC5 and GIIC8 from United Kingdom broiler flocks. The screening of 53 lytic bacteriophage isolates against a panel of 50 Campylobacter isolates from broiler chickens and 80 strains isolated after human infection identified two phage candidates with broad host lysis. These phages, CP8 and CP34, were orally administered in antacid suspension, at different dosages, to 25-day-old broiler chickens experimentally colonized with the C. jejuni broiler isolates. Phage treatment of C. jejuni-colonized birds resulted in Campylobacter counts falling between 0.5 and 5 log10 CFU/g of cecal contents compared to untreated controls over a 5-day period postadministration. These reductions were dependent on the phage-Campylobacter combination, the dose of phage applied, and the time elapsed after administration. Campylobacters resistant to bacteriophage infection were recovered from phage-treated chickens at a frequency of <4%. These resistant types were compromised in their ability to colonize experimental chickens and rapidly reverted to a phage-sensitive phenotype in vivo. The selection of appropriate phage and their dose optimization are key elements for the success of phage therapy to reduce campylobacters in broiler chickens.     

Microbial ecology of Campylobacter jejuni in a United Kingdom chicken supply chain: intermittent common source, vertical transmission, and amplification by flock propagation.

A study of Campylobacter jejuni on a broiler chicken farm between 1989 and 1994 gave an estimated isolation rate of 27% (3,304 of 12,233) from a 0.9% sample of 1.44 million broiler chickens from six to eight sheds over 32 consecutive rearing flocks comprising 251 broiler shed flocks. During the study, C. jejuni was found in 35.5% of the 251 shed flocks but only 9.2% (23 of 251) had Campylobacter isolates in successive flocks, with 9 of those 23 sheds having the same serotype between consecutive flocks, indicating a low level of transmission between flocks. Analysis of a systematic sample of 484 of 3,304 (14.6%) C. jejuni isolates showed that 85% were of 10 serotype complexes but 58% were of 3 serotype complexes, indicating a high degree of strain similarity throughout the entire study. The three commonest types were detected in 8 of 32 flocks during the 5-year study period, suggesting an intermittent common external Campylobacter source. This hypothesis was tested by a retrospective cohort analysis of C. jejuni rates and types by reference to hatchery supplier of the 1-day-old chicks. Isolation rates of C. jejuni and frequency distribution of types were determined in 6-week-old broiler chickens identified by the hatchery supplying the original chicks. The isolation rate of C. jejuni in broilers, supplied by hatchery A, was 17.6%, compared to 42.9% (P < 0.0001) for broilers reared from chicks supplied by hatchery B. In two instances, when both hatcheries were used to stock the same farm flock, Campylobacter isolates were found only in those sheds with chicks supplied by hatchery B. Thus, the frequency distribution of Campylobacter types for chickens supplied by the two hatcheries over the 5-year period showed marked dissimilarity. These findings suggest that the isolation rate and type of Campylobacter isolates in broiler chickens was associated with the hatchery supplying chicks. The lack of diversity of types and the intermittent high positivity of sheds is evidence for a common source of C. jejuni introduced by vertical transmission rather than contamination at the hatchery or during transportation.     

Effect of Campylobacter-specific maternal antibodies on Campylobacter jejuni colonization in young chickens

Using laboratory challenge experiments, we examined whether Campylobacter-specific maternal antibody (MAB) plays a protective role in young chickens, which are usually free of Campylobacter under natural production conditions. Kinetics of C. jejuni colonization were compared by infecting 3-day-old broiler chicks, which were naturally positive for Campylobacter-specific MAB, and 21-day-old broilers, which were negative for Campylobacter-specific MAB. The onset of colonization occurred much sooner in birds challenged at the age of 21 days than it did in the birds inoculated at 3 days of age, which suggested a possible involvement of specific MAB in the delay of colonization. To further examine this possibility, specific-pathogen-free layer chickens were raised under laboratory conditions with or without Campylobacter infection, and their 3-day-old progenies with (MAB(+)) or without (MAB(-)) Campylobacter-specific MAB were orally challenged with C. jejuni. Significant decreases in the percentage of colonized chickens were observed in the MAB(+) group during the first week compared with the MAB(-) group. These results indicate that Campylobacter-specific MAB plays a partial role in protecting young chickens against colonization by C. jejuni. Presence of MAB in young chickens did not seem to affect the development of systemic immune response following infection with C. jejuni. However, active immune responses to Campylobacter occurred earlier and more strongly in birds infected at 21 days of age than those infected at 3 days of age. Clearance of Campylobacter infection was also observed in chickens infected at 21 days of age. Taken together, these findings (i) indicate that anti-Campylobacter MAB contributes to the lack of Campylobacter infection in young broiler chickens in natural environments and (ii) provide further evidence supporting the feasibility of development of immunization-based approaches for control of Campylobacter infection in poultry.     

Dissemination of Fluoroquinolone-Resistant Campylobacter spp. within an Integrated Commercial Poultry Production System

While characterizing the intestinal bacterial community of broiler chickens, we detected -proteobacterial DNA in the ilea of 3-day-old commercial broiler chicks (J. Lu, U. Idris, B. Harmon, C. Hofacre, J. J. Maurer, and M. D. Lee, Appl. Environ. Microbiol. 69:6816-6824, 2003). The sequences exhibited high levels of similarity to Campylobacter jejuni and Campylobacter coli sequences, suggesting that chickens can carry Campylobacter at a very young age. Campylobacter sp. was detected by PCR in all samples collected from the ilea of chicks that were 3 to 49 days old; however, it was detected only in the cecal contents of chickens that were at least 21 days old. In order to determine whether the presence of Campylobacter DNA in young chicks was due to ingestion of the bacteria in food or water, we obtained commercial broiler hatching eggs, which were incubated in a research facility until the chicks hatched. DNA sequencing of the amplicons resulting from Campylobacter-specific 16S PCR performed with the ileal, cecal, and yolk contents of the day-of-hatching chicks revealed that Campylobacter DNA was present before the chicks consumed food or water. The 16S rRNA sequences exhibited 99% similarity to C. jejuni and C. coli sequences and 95 to 98% similarity to sequences of other thermophilic Campylobacter species, such as C. lari and C. upsaliensis. The presence of C. coli DNA was detected by specific PCR in the samples from chicks obtained from a commercial hatchery; however, no Campylobacter was detected by culturing. In order to determine whether the same strains of bacteria were present in multiple levels of the integrator, we cultured Campylobacter sp. from a flock of broiler breeders and their 6-week-old progeny that resided on a commercial broiler farm. The broiler breeders had been given fluoroquinolone antibiotics, and we sought to determine whether the same fluoroquinolone-resistant strain was present in their progeny. The isolates were typed by pulsed-field gel electrophoresis, which confirmed that the parental and progeny flocks contained the same strain of fluoroquinolone-resistant C. coli. These data indicate that resistant C. coli can be present in multiple levels of an integrated poultry system and demonstrated that molecular techniques or more sensitive culture methods may be necessary to detect early colonization by Campylobacter in broiler chicks.     

Production of organic acids by probiotic lactobacilli can be used to reduce pathogen load in poultry

Probiotic Lactobacillus can be used to reduce the colonization of pathogenic bacteria in food animals, and therefore reduce the risk of foodborne illness to consumers. As a model system, we examined the mechanism of protection conferred by Lactobacillus species to inhibit C. jejuni growth in vitro and reduce colonization in broiler chickens. Possible mechanisms for the reduction of pathogens by lactobacilli include: 1) stimulation of adaptive immunity; 2) alteration of the cecal microbiome; and, 3) production of inhibitory metabolites, such as organic acids. The Lactobacillus species produced lactic acid at concentrations sufficient to kill C. jejuni in vitro. We determined that lactic acid produced by Lactobacillus disrupted the membrane of C. jejuni, as judged by biophotonics. The spectral features obtained using Fourier-transform infrared (FT-IR) and Raman spectroscopy techniques were used to accurately predict bacterial viability and differentiate C. jejuni samples according to lactic acid treatment. FT-IR spectral features of C. jejuni and Lactobacillus grown in co-culture revealed that the metabolism was dominated by Lactobacillus prior to the killing of C. jejuni. Based on our results, the development of future competitive exclusion strategies should include the evaluation of organic acid production.     

Defined competitive exclusion cultures in the prevention of enteropathogen colonisation in poultry and swine

A competitive exclusion culture (CE) containing a mixture of 29 different bacterial isolates obtained from the cecae of broiler chickens was developed utilizing continuous-flow culture techniques. This culture (CF3) has been efficacious in controlling gut colonization by enteropathogens in both experimentally infected broilers and under commercial field conditions. In day-old broiler chicks provided CF3, and challenged with 10,000 CFU Salmonella typhimurium greater than a 99% reduction in Salmonella cecal colonization levels was observed compared to control chicks. Similarly, CF3 has also been shown to protect experimentally infected broiler chicks from cecal colonization by S. enteritidis (Phage types 4 and 13), S. gallinarum, Listeria Monocytogenes, and Escherichia coli O157:H7. A commercial product was developed from CF3 and is sold under the tradename PREEMPT™. In a Food and Drug Administration approved, double blinded, pivotal field trial, chicks treated with PREEMPTT™ had significantly fewer salmonellae than untreated chicks at end-of-growout. This product is the first of its kind available to the U.S. poultry industry. Using similar technology a product has also been developed that decreases shedding of salmonellae in neonate and weaned pigs, and also has been shown to reduce mortality associated with enteropathogens in young pigs both in the laboratory and in a commercial swine herd. This revised version was published online in August 2006 with corrections to the Cover Date.     

Genotyping of Campylobacter jejuni strains from Danish broiler chickens by restriction fragment length polymorphism of the LPS gene cluster

AIMS: To apply and evaluate LG (LPS genes) genotyping, which is a genotyping method based on a cluster of genes involved in the synthesis of surface lipopolysaccharides (LPS) in Campylobacter species, for typing of Campylobacter jejuni isolates obtained from Danish broiler chickens. Furthermore, the LG genotyping method was used to study the genetic stability of four C. jejuni strains after gastrointestinal passage through experimentally infected chickens. METHODS AND RESULTS: In the present study, the LG genotyping method was modified with respect to the restriction enzymes used. To validate the method, 63 Penner serotype reference strains and 107 C. jejuni chicken isolates, representing the most common Penner serotypes of C. jejuni in Danish poultry, were selected for typing. The method was successfully used for typing all isolates and the LG genotype profiles were reproducible. There were no changes in the LG genotype of the C. jejuni strains obtained after experimental passage through chickens. CONCLUSIONS: All C. jejuni strains obtained from broiler chickens were typeable by the LG genotyping method. Application of the RsaI restriction enzyme improved the method in terms of ease and consistency of analyses and increase of discriminatory power. SIGNIFICANCE AND IMPACT OF THE STUDY: The LG genotyping method is a valuable tool for typing C. jejuni isolates obtained from poultry. However, the association between Penner serotyping based on passive haemagglutination of heat-stable antigens and LG genotyping was low when applied to poultry isolates. This is in contrast to previous studies on isolates of human origin that reported a high correlation between results obtained by the two typing methods (Shi et al. 2002).     

Cecal Propionic Acid as a Biological Indicator of the Early Establishment of a Microbial Ecosystem Inhibitory toSalmonellain Chicks

Experiments were conducted to evaluate the effects of different competitive exclusion (CE) cultures on the concentration of cecal propionic acid in 3-day-old broiler chicks, and the correlation between cecal propionic acid concentration and protection againstSalmonellacolonization. CE cultures that significantly (P < 0.05) increased cecal propionic acid in 3-day-old chicks decreased (P < 0.05) cecalSalmonellacolonization in 10-day-old chicks compared with the untreated controls. CE cultures that failed to significantly (P > 0.05) increase cecal propionic acid concentrations in 3-day-old chicks failed to protect (P > 0.05) against cecalSalmonellacolonization in 10-day-old chicks compared with untreated controls. A significant (P < .05) correlation (−.88) was found between cecal propionic acid concentration in 3-day-old chicks and cecalSalmonellacolonization in 10-day-old chicks.     

Endothermic heart rate response in broiler and White Leghorn chicks (Gallus gallus domesticus) during the first two days of post-hatch life

The embryonic modal value of heart rate (MHR) differs between broiler and White Leghorn chickens, but the initial development of cholinergic chronotropic control of embryonic heart rate (HR) does not. Thus, we hypothesized that hatchling MHR should also differ between broiler and White Leghorn strains, while the development of a physiological regulation, such as the endothermic HR response, should not be different between hatchlings of the two strains. To test this, we measured the response of HR and cloaca temperature (Tb) to alteration of ambient temperature (Ta); i.e., 35 degrees C-25 degrees C-35 degrees C, in four groups of hatchlings on Days 0 and 1 post-hatch. Fertile eggs of both strains with similar mass were incubated simultaneously in the same incubator. Eggs of broiler chickens hatched approximately 7 h earlier than White Leghorn chicken eggs. Chick mass at hatching was identical in both strains, but diverged during 2 days after hatching. Tb measured at the initial Ta of 35 degrees C was identical in both strains. MHR at the same Ta was approximately 30 bpm lower in broiler chicks than in White Leghorn chicks, but the difference was reversed to that observed in the embryos. The endothermic HR response was advanced by approximately 1 day in broiler chicks compared with White Leghorn chicks. As a result, eggs of similar mass in both strains produced chicks with similar mass and Tb at hatching, but during 2 days of post-hatch life their masses diverged and regulation of the endothermic HR response developed earlier in broiler than in White Leghorn hatchlings. This physiological heterochrony between strains is most likely due to genetic selection for fast growth in broiler chickens.     

Prevalence, antigenic specificity, and bactericidal activity of poultry anti-Campylobacter maternal antibodies

Poultry are considered the major reservoir for Campylobacter jejuni, a leading bacterial cause of human food-borne diarrhea. To understand the ecology of C. jejuni and develop strategies to control C. jejuni infection in the animal reservoir, we initiated studies to examine the potential role of anti-Campylobacter maternal antibodies in protecting young broiler chickens from infection by C. jejuni. Using an enzyme-linked immunosorbent assay (ELISA), the prevalence of anti-C. jejuni antibodies in breeder chickens, egg yolks, and broilers from multiple flocks of different farms were examined. High levels of antibodies to the organism were detected in serum samples of breeder chickens and in egg yolk contents. To determine the dynamics of anti-Campylobacter maternal antibody transferred from yolks to hatchlings, serum samples collected from five broiler flocks at weekly intervals from 1 to 28 or 42 days of age were also examined by ELISA. Sera from the 1-day and 7-day-old chicks showed high titers of antibodies to C. jejuni. Thereafter, antibody titers decreased substantially and were not detected during the third and fourth weeks of age. The disappearance of anti-Campylobacter maternal antibodies during 3 to 4 weeks of age coincides with the appearance of C. jejuni infections observed in many broiler chicken flocks. As shown by immunoblotting, the maternally derived antibodies recognized multiple membrane proteins of C. jejuni ranging from 19 to 107 kDa. Moreover, in vitro serum bactericidal assays showed that anti-Campylobacter maternal antibodies were active in antibody-dependent complement-mediated killing of C. jejuni. Together, these results highlight the widespread presence of functional anti-Campylobacter antibodies in the poultry production system and provide a strong rationale for further investigation of the potential role of anti-C. jejuni maternal antibodies in protecting young chickens from infection by C. jejuni.     

Dynamics of Dual Infection with Campylobacter jejuni Strains in Chickens Reveals Distinct Strain-to-Strain Variation in Infection Ecology

Although multiple genotypes of Campylobacter jejuni may be isolated from the same commercial broiler flock, little is known about the infection dynamics of different genotypes within individuals or their colonization sites within the gut. Single experimental infections with C. jejuni M1 (sequence type 137, clonal complex 45) and C. jejuni 13126 (sequence type 21, clonal complex 21) revealed that 13126 colonized the ceca at significantly higher levels. The dissemination and colonization sites of the two C. jejuni strains then were examined in an experimental broiler flock. Two 33-day-old broiler chickens were infected with M1 and two with 13126, and 15 birds were left unchallenged. Cloacal swabs were taken postinfection to determine the colonization and shedding of each strain. By 2 days postinfection (dpi), 8/19 birds were shedding M1 whereas none were shedding 13126. At 8 dpi, all birds were shedding both strains. At 18 dpi, liver and cecal levels of each isolate were quantified, while in 10 birds they also were quantified at nine sites throughout the gastrointestinal (GI) tract. 13126 was found throughout the GI tract, while M1 was largely restricted to the ceca and colon. The livers of 7/19 birds were culture positive for 13126 only. These data show that 13126 has a distinctly different infection biology than strain M1. It showed slower colonization of the lower GI tract but was more invasive and able to colonize at a high level throughout the GI tract. The finding that C. jejuni strains have markedly different infection ecologies within the chicken has implications for control in the poultry industry and suggests that the contamination risk of edible tissues is dependent on the isolate involved.     

Comparison of 2-day-old and 14-day-old chicken colonization models for Campylobacter jejuni

In this study, we compared two types of chicken infection models for Campylobacter jejuni in terms of infectious dose required to colonize the chickens and the susceptibility of chickens of different ages to persistent colonization by C. jejuni. In one model, chickens at day 2 posthatching were used, and in the other, 14-day-old chickens were used. The minimum C. jejuni cell number required to colonize 14-day-old chickens was 5 x 10(4) cells, and that for 2-day-old chickens was 5 x 10(3). The ability of various C. jejuni strains to colonize the chicken gastrointestinal tract was the same in both models.     

Commonality and biosynthesis of the O-methyl phosphoramidate capsule modification in Campylobacter jejuni

In this study we investigated the commonality and biosynthesis of the O-methyl phosphoramidate (MeOPN) group found on the capsular polysaccharide (CPS) of Campylobacter jejuni. High resolution magic angle spinning NMR spectroscopy was used as a rapid, high throughput means to examine multiple isolates, analyze the cecal contents of colonized chickens, and screen a library of CPS mutants for the presence of MeOPN. Sixty eight percent of C. jejuni strains were found to express the MeOPN with a high prevalence among isolates from enteritis, Guillain Barré, and Miller-Fisher syndrome patients. In contrast, MeOPN was not observed for any of the Campylobacter coli strains examined. The MeOPN was detected on C. jejuni retrieved from cecal contents of colonized chickens demonstrating that the modification is expressed by bacteria inhabiting the avian gastrointestinal tract. In C. jejuni 11168H, the cj1415-cj1418 cluster was shown to be involved in the biosynthesis of MeOPN. Genetic complementation studies and NMR/mass spectrometric analyses of CPS from this strain also revealed that cj1421 and cj1422 encode MeOPN transferases. Cj1421 adds the MeOPN to C-3 of the beta-d-GalfNAc residue, whereas Cj1422 transfers the MeOPN to C-4 of D-glycero-alpha-L-gluco-heptopyranose. CPS produced by the 11168H strain was found to be extensively modified with variable MeOPN, methyl, ethanolamine, and N-glycerol groups. These findings establish the importance of the MeOPN as a diagnostic marker and therapeutic target for C. jejuni and set the groundwork for future studies aimed at the detailed elucidation of the MeOPN biosynthetic pathway.     

Characterization of Campylobacter jejuni biofilms under defined growth conditions

Campylobacter jejuni is a major cause of human diarrheal disease in many industrialized countries and is a source of public health and economic burden. C. jejuni, present as normal flora in the intestinal tract of commercial broiler chickens and other livestock, is probably the main source of human infections. The presence of C. jejuni in biofilms found in animal production watering systems may play a role in the colonization of these animals. We have determined that C. jejuni can form biofilms on a variety of abiotic surfaces commonly used in watering systems, such as acrylonitrile butadiene styrene and polyvinyl chloride plastics. Furthermore, C. jejuni biofilm formation was inhibited by growth in nutrient-rich media or high osmolarity, and thermophilic and microaerophilic conditions enhanced biofilm formation. Thus, nutritional and environmental conditions affect the formation of C. jejuni biofilms. Both flagella and quorum sensing appear to be required for maximal biofilm formation, as C. jejuni flaAB and luxS mutants were significantly reduced in their ability to form biofilms compared to the wild-type strain.