Investigation of Campylobacter colonization in three Australian commercial free-range broiler farms.
ABSTRACT: Campylobacter spp. contaminated poultry products are strongly associated with foodborne illnesses worldwide. Development of effective management strategies to reduce contamination by Campylobacter spp. requires an improved understanding of the numerous factors that drive these contamination processes. Currently, chicken farms are using more free-range chicken meat production systems in response to consumer preferences. However, Campylobacter spp. colonization has rarely been investigated on free-range broiler farms. The present study investigated the temporal and environmental factors influencing Campylobacter spp. colonization of free-range broilers as well as potential sources and genetic diversity of Campylobacter jejuni (C. jejuni) and Campylobacter coli (C. coli) in commercial free-range broiler farms. Genetic linkages among the isolates were analyzed using flaA amplicon analysis. Campylobacter coli was first detected in fecal samples of a commercial free-range broiler flock on day 10 of rearing. Multiple genotypes of C. jejuni and C. coli were identified in this study. The farm environment was identified as a potential source of C. jejuni and C. coli colonization of free-range broilers. The dominant Campylobacter genotype varied between free-range broiler farms over time, with C. jejuni being the most frequently isolated species. These findings enhance the understanding of C. jejuni and C. coli colonization in free-range broiler farms and could inform the development of more effective intervention strategies to help control this important foodborne pathogen.
Project description: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.
Project description:BACKGROUND:Understanding potential risks of multi-drug resistant (MDR) pathogens from the booming poultry sector is a crucial public health concern. Campylobacter spp. are among the most important zoonotic pathogens associated with MDR infections in poultry and human. This study systematically examined potential risks and associated socio-environmental factors of MDR Campylobacter spp. in poultry farms and live bird markets (LBMs) of Bangladesh. METHODS:Microbial culture and PCR-based methods were applied to examine the occurrence and MDR patterns of Campylobacter spp. in potential sources (n =?224) at 7 hatcheries, 9 broiler farms and 4 LBMs in three sub-districts. Antimicrobial residues in broiler meat and liver samples (n =?50) were detected by advanced chromatographic techniques. A questionnaire based cross-sectional survey was conducted on socio-environmental factors. RESULTS:Overall, 32% (71/ 224) samples were found contaminated with Campylobacter spp. In poultry farms, Campylobacter spp. was primarily found in cloacal swab (21/49, 43%), followed by drinking water (8/24, 33%), and meat (8/28, 29%) samples of broilers. Remarkably, at LBMs, Campylobacter spp. was detected in higher prevalence (p <?0.05) in broiler meat (14/26, 54%), which could be related (p <?0.01) to bacterial contamination of drinking water (11/21, 52%) and floor (9/21, 43%). Campylobacter isolates, one from each of 71 positive samples, were differentiated into Campylobacter jejuni (66%) and Campylobacter coli (34%). Alarmingly, 49 and 42% strains of C. jejuni and C. coli, respectively, were observed as MDR, i.e., resistant to three or more antimicrobials, including, tetracycline, amoxicillin, streptomycin, fluoroquinolones, and macrolides. Residual antimicrobials (oxytetracycline, ciprofloxacin and enrofloxacin) were detected in majority of broiler liver (79%) and meat (62%) samples, among which 33 and 19%, respectively, had concentration above acceptable limit. Inadequate personal and environmental hygiene, unscrupulously use of antimicrobials, improper waste disposal, and lack of health surveillance were distinguishable risk factors, with local diversity and compound influences on MDR pathogens. CONCLUSION:Potential contamination sources and anthropogenic factors associated with the alarming occurrence of MDR Campylobacter, noted in this study, would aid in developing interventions to minimize the increasing risks of poultry-associated MDR pathogens under 'One Health' banner that includes poultry, human and environment perspectives.
Project description:Campylobacter infections sourced mainly to poultry products, are the most important bacterial foodborne zoonoses worldwide. No effective measures to control these infections in broiler production exist to date. Here, we used passive immunization with hyperimmune egg yolks to confer broad protection of broilers against Campylobacter infection. Two novel vaccines, a bacterin of thirteen Campylobacter jejuni (C. jejuni) and C. coli strains and a subunit vaccine of six immunodominant Campylobacter antigens, were used for the immunization of layers, resulting in high and prolonged levels of specific immunoglobulin Y (IgY) in the hens' yolks. In the first in vivo trial, yolks (sham, bacterin or subunit vaccine derived) were administered prophylactically in the broiler feed. Both the bacterin- and subunit vaccine-induced IgY significantly reduced the number of Campylobacter-colonized broilers. In the second in vivo trial, the yolks were administered therapeutically during three days before euthanasia. The bacterin IgY resulted in a significant decrease in C. jejuni counts per infected bird. The hyperimmune yolks showed strong reactivity to a broad representation of C. jejuni and C. coli clonal complexes. These results indicate that passive immunization with hyperimmune yolks, especially bacterin derived, offers possibilities to control Campylobacter colonization in poultry.
Project description:The small-scale mobile poultry-processing unit (MPPU) produced raw poultry products are of particular food safety concern due to exemption of USDA poultry products inspection act. Limited studies reported the microbial quality and safety of MPPU-processed poultry carcasses. This study evaluated the <i>Salmonella</i> and <i>Campylobacter</i> prevalence in broiler ceca and on MPPU-processed carcasses and efficacy of commercial antimicrobials against <i>Campylobacter jejuni</i> on broilers. In study I, straight-run Hubbard?×?Cobb broilers (147) were reared for 38?days on clean-shavings (CS, 75) or built-up-litter (BUL, 72) and processed at an MPPU. Aerobic plate counts (APCs), coliforms, <i>Escherichia coli</i>, and yeast/molds (Y/M) of carcasses were analyzed on petrifilms. Ceca and carcass samples underwent microbial analyses for <i>Salmonella</i> and <i>Campylobacter</i> spp. using the modified USDA method and confirmed by API-20e test (<i>Salmonella</i>), latex agglutination immunoassay (<i>Campylobacter</i>), and Gram staining (<i>Campylobacter</i>). Quantitative polymerase chain reaction (CadF gene) identified the prevalence of <i>C. jejuni</i> and <i>Campylobacter coli</i> in ceca and on carcasses. In study II, fresh chilled broiler carcasses were spot inoculated with <i>C. jejuni</i> (4.5?log<sub>10</sub>?CFU/mL) and then undipped, or dipped into peroxyacetic acid (PAA) (1,000?ppm), lactic acid (5%), lactic and citric acid blend (2.5%), sodium hypochlorite (69?ppm), or a H<sub>2</sub>O<sub>2</sub>-PAA mix (SaniDate<sup>®</sup> 5.0, 0.25%) for 30?s. Surviving <i>C. jejuni</i> was recovered onto Brucella agar. APCs, coliforms, and <i>E. coli</i> populations were similar (<i>P</i>?>?0.05) on CS and BUL carcasses. Carcasses of broilers raised on BUL contained a greater (<i>P</i>?<?0.05) Y/M population (2.2?log<sub>10</sub>?CFU/mL) than those reared on CS (1.8?log<sub>10</sub>?CFU/mL). <i>Salmonella</i> was not detected in any ceca samples, whereas 2.8% of the carcasses from BUL were present with <i>Salmonella</i>. Prevalence of <i>Campylobacter</i> spp., <i>C. jejuni</i> was lower (<i>P</i>?<?0.05), and <i>C. coli</i> was similar (<i>P</i>?>?0.05) in CS-treated ceca than BUL samples. Prevalence of <i>Campylobacter</i> spp., <i>C. jejuni</i>, and <i>C. coli</i> was not different (<i>P</i>?>?0.05) on CS- and BUL-treated carcasses. All antimicrobials reduced <i>C. jejuni</i> by 1.2-2.0?log CFU/mL on carcasses compared with controls. Hence, raising broilers on CS and applying post-chilling antimicrobial treatment can reduce <i>Salmonella</i> and <i>Campylobacter</i> on MPPU-processed broiler carcasses.
Project description:Campylobacter infections are among the most prevalent foodborne infections in humans, resulting in a massive disease burden worldwide. Broilers have been identified as the major source of campylobacteriosis and reducing Campylobacter loads in the broiler caeca has been proposed as an effective measure to decrease the number of infections in humans. Failure of current methods to control Campylobacter in broilers stresses the urgency to develop novel mitigation measures. We obtained six nanobodies with a broad specificity, that recognize strains belonging to the two most relevant species, Campylobacter jejuni and Campylobacter coli. The target of the nanobodies was identified as the major outer membrane protein, a porin that contributes to bacterial virulence and viability. Multimerization of the nanobodies led to agglutination of C. jejuni cells, which may affect colonization in the chicken gut. These Campylobacter-specific nanobodies may be useful to develop a strategy for preserving chickens from Campylobacter colonization.
Project description:Poultry origin Campylobacter is considered as one of the leading causal agents of human foodborne illness. This study was conducted to estimate the occurrence, molecular identification, and antimicrobial resistance (AMR) of Campylobacter species from the broiler farms in Bangladesh. Samples (352) were collected from 32 farms and comprised of 128 cloacal swab, 64 feed, 64 drinking water, 64 attendants' hand rinsed water, and 32 whole carcasses. All samples were tested for the presence of Campylobacter via cultural, biochemical, and PCR. The AMR was determined via the disc diffusion method. An overall occurrence of Campylobacter spp. was estimated as 26.4%. The level of Campylobacter contamination was found to be higher in conventional farms (36.4%) than the good practice farms (16.5%) including all sample categories (p = 0.000). Of 93 isolates, 67.74% and 32.26% were confirmed as C. jejuni and C. coli respectively, of which 34.92% C. jejuni, and 30% C. coli were shown to be multidrug-resistant. A significant occurrence of Campylobacter contamination in broiler farms with multidrug resistant patterns might be cogitated as serious food safety and public health concern linking to poultry food chain. A risk reduction approach through good farming practices targeting the prudent use of antimicrobials for broiler production is thus necessitated.
Project description:The genetic diversity of Campylobacter jejuni and Campylobacter coliisolates from commercial broiler farms was examined by multilocus sequence typing (MLST), with an assessment of the impact of the sample type and laboratory method on the genotypes of Campylobacter isolated. A total of 645C. jejuniand 106C. coli isolates were obtained from 32 flocks and 17 farms, with 47 sequence types (STs) identified. The Campylobacter jejuniisolates obtained by different sampling approaches and laboratory methods were very similar, with the same STs identified at similar frequencies, and had no major effect on the genetic profile of Campylobacter population in broiler flocks at the farm level. ForC. coli, the results were more equivocal. While some STs were widely distributed within and among farms and flocks, analysis of molecular variance (AMOVA) revealed a high degree of genetic diversity among farms forC. jejuni, where farm effects accounted for 70.5% of variance, and among flocks from the same farm (9.9% of variance for C. jejuni and 64.1% forC. coli). These results show the complexity of the population structure of Campylobacterin broiler production and that commercial broiler farms provide an ecological niche for a wide diversity of genotypes. The genetic diversity of C. jejuni isolates among broiler farms should be taken into account when designing studies to understand Campylobacter populations in broiler production and the impact of interventions. We provide evidence that supports synthesis of studies on C. jejuni populations even when laboratory and sampling methods are not identical.
Project description:Biosecurity seems to be the most promising tool for <i>Campylobacter</i> control on poultry farms. A longitudinal molecular epidemiological study was performed during two production cycles, in which the broilers, the poultry house, and the environment of 10 (mixed) broiler farms were monitored weekly. Cecal droppings from the second production cycle were also used for 16S metabarcoding to study the differences in the microbiota of colonized and uncolonized flocks. Results showed that 3 out of 10 farms were positive for <i>Campylobacter</i> in the first production cycle, and 4 out of 10 were positive in the second. Broilers became colonized at the earliest when they were four weeks old. The majority of the flocks (57%) became colonized after partial depopulation. Before colonization of the flocks, <i>Campylobacter</i> was rarely detected in the environment, but it was frequently isolated from cattle and swine. Although these animals appeared to be consistent carriers of <i>Campylobacter</i>, molecular typing revealed that they were not the source of flock colonization. In accordance with previous reports, this study suggests that partial depopulation appears to be an important risk factor for <i>Campylobacter</i> introduction into the broiler house. Metabarcoding indicated that two <i>Campylobacter</i>-free flocks carried high relative abundances of <i>Megamonas</i> in their ceca, suggesting potential competition with <i>Campylobacter</i>.
Project description: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.
Project description:Contaminated poultry meat is considered to be the main source of human infection with Campylobacter spp., a pathogen that asymptomatically colonizes broiler chickens during fattening and contaminates carcasses during slaughter. To prevent or reduce the colonization of broiler flocks with Campylobacter spp., applying different organic acids, especially in combinations, via feed or drinking water seems to be a promising approach. However, only very few combinations of organic acids have been tested for their antibacterial efficacy against Campylobacter spp. Therefore, the in vitro susceptibility of 30 Campylobacter spp. isolates (20 C. jejuni and ten C. coli) to ten organic acids and ten combinations was determined. The testing of minimum inhibitory concentration (MIC) values was performed at pH 6.0 and 7.3 by using the broth microdilution method and included the following organic acids: Caprylic acid, sorbic acid, caproic acid, benzoic acid, ascorbic acid, propionic acid, acetic acid, formic acid, fumaric acid and tartaric acid and combinations thereof. The lowest MIC values were seen for caprylic acid (MIC range at pH 7.3: 0.5-2 mmol/L) and sorbic acid (MIC range at pH 7.3: 1-4 mmol/L). One to two dilution steps lower MIC values were determined at the lower pH value of 6.0. Furthermore, ten combinations consisting of three to five organic acids were developed. In addition to the tested antibacterial activity, other criteria were included such as approval as feed additives, reported synergistic effects and chemical properties. For nine of ten combinations, the MIC90 values of the organic acids decreased 1.25- to 241.5-fold compared to the MIC90 values for the individual substances. Furthermore, nine of ten combinations exhibited synergistic activities against two or more of the tested C. jejuni and C. coli isolates. A combination of caprylic acid, sorbic acid and caproic acid exhibited synergistic activities against the largest number of Campylobacter spp. isolates (six C. jejuni and four C. coli) with fractional inhibitory concentration (FIC) indices (?FIC) ranging from 0.33 to 1.42. This study shows in vitro synergistic activities of different organic acids in combinations against the major Campylobacter species and could therefore be a promising basis for reducing Campylobacter spp. in vivo.