Campylobacteriosis is recognized as a leading bacterial cause of human gastro-enteritis in both developed and developing countries. Though it has been known for a long time that poultry is the main source of Campylobacter infection, recent studies have suggested that cattle and sheep are potential emerging sources for human infections. Cattle can contribute to human infections through several routes of transmission such as direct contact, environmental contamination, and ingestion of contaminated food and milk. C. jejuni and C. coli usually colonize cattle without showing clear symptoms. Some studies have reported that 80% of livestock herds and 40–60% of individual animals may shed Campylobacter. However, the direct and indirect contributions of cattle and sheep in spreading of human infections are still under investigation. In addition, the sources of Campylobacter and its routes of transmission at dairy farms have not been clearly established. Several studies have also indicated that wild birds may play a potential role in the epidemiology of human campylobacteriosis by transmitting Campylobacter from animals to humans. In our study we hypothesized that dairy cattle and feedlot beef cattle act as potential sources of human campylobacteriosis. Further, the prevalence, and genotypic relatedness of C. jejuni and C. Coli isolated from dairy and beef cattle are different depending on the region where they originate. We also hypothesized that wild birds’ reservoirs may spread Campylobacter species to dairy farm environments which can be transmitted to dairy cattle and humans. Thus, both wild birds and dairy cattle play important role in human infections. For testing our hypothesis, in the first study, we determined the prevalence, genotypic and phenotypic properties of these pathogens. Further, characterization of the public health relevance of the cattle-associated Campylobacter by determining their invasion and intracellular potential to human intestinal cells (INT407), as well as their antimicrobial resistance profiles. Fecal samples were collected from beef and cull-cows at slaughter plants from four different representative geographical locations in the US. to determine the rate of Campylobacter infection. Our results showed that the prevalence of Campylobacter varied regionally and was significantly (P<0.05) higher in fecal samples collected from the South as compared to North, East and Midwest. Pulsed Field Gel Electrophoresis (PFGE) analysis showed that C. jejuni and C. coli isolates were genotypically diverse and certain genotypes were shared across two or more of the geographic locations. Further, MLST analysis demonstrated that the cattle associated C. jejuni strains harbored sequence types that were commonly shared in human cases and also showed varying invasion and intracellular survival capacity to human intestinal cells. Furthermore, many cattle associated Campylobacter isolates showed resistance to several antimicrobials including ciprofloxacin, erythromycin, and gentamicin. Conclusively, our results highlight the importance of cattle as an important reservoir of Campylobacter spp clinically important to human.
In the second study, we investigated the occurrence of the invasion associated marker (iam) in C. jejuni isolated from cattle in order to determine the contribution of this reservoir to human infections with invasive Campylobacter. Additionally, we assessed iam’s contribution to the colonization of multiple hosts by characterizing the potential of iam-containing cattle isolates for chicken colonization and human intestinal cell invasion. Simultaneous RAPD typing and iam-specific PCR analysis of cattle and human-associated C. jejuni isolates showed that the prevalence of iam in cattle C. jejuni is relatively lower as compared to isolates occurring in humans and chickens. In addition, iam was polymorphic and certain alleles occured in cattle isolates that were capable of colonizing and invading chickens and human intestinal cells, respectively. However, the iam did not appear to contribute to the invasion, intracellular survival, and antimicrobial resistance potential of cattle-associated C. jejuni. We propose that the screening of the iam as a virulence determinant for epidemiological purposes in C. jejuni must be carefully considered.
In the third study, we determined the prevalence, genotypic, and phenotypic properties of Campylobacter that were isolated from paired fecal samples of dairy cattle and starlings (Sturnus vulgaris) in Northeastern Ohio. Our results showed that the prevalence of C. jejuni in birds was significantly (P<0.01) higher than that in dairy cattle. Further, PFGE analysis showed that C. jejuni were mostly genetically diverse and host restricted. However, there were several shared genotypes between dairy and starlings isolates. Furthermore, clonal complexes (CC) ST-45, and ST-21 were frequently shared between dairy and starlings by MLST analysis, which were also commonly found in humans. However, unique starlings’ isolates associated STs such as CC ST-177 and ST-682 as well as CC ST-42 that was restricted to dairy isolates were detected. Further, two new STs were detected in C. jejuni isolated from dairy cattle. Interestingly, cattle and starlings Campylobacter showed high resistance to multiple antimicrobials including ciprofloxacin, erythromycin, and gentamicin. In conclusion, our results highlight starlings as potential reservoirs for C. jejuni and may play important role in the epidemiology of clinically important C. jejuni in dairy cattle population.
In the last study, we conducted a pilot study to understand the pathogenesis of C. jejuni induced abortion in sheep. Campylobacter jejuni has been recently shown to be increasingly associated with sheep abortion; however, it is still not known which circumstances and factors contribute to emergence of virulent C.jejuni strains that can cause abortion in sheep. We determined the genotypic and phenotypic properties of ovine and bovine abortion associated Campylobacter isolates which were acquired from the diagnostic laboratories of the Ohio Department of Agriculture (ODA). Further analyses included invasion and intracellular survival potentials in human INT407 cells, along with in-vivo experiment in pregnant ewes. PFGE analysis showed that the fingerprints of ovine and bovine abortion C. jejuni were identical. Furthermore, by MLST analysis, ovine and bovine abortion isolates were classified as (Sequence Type) ST-8 which belonged to Clonal Complex (CC) ST-21 which is also commonly found in humans. Additionally, the ovine and bovine abortion associated C. jejuni strains showed varying invasion and intracellular survival capacity to human intestinal INT407 cells; however, one isolate showed a significantly higher invasion potential compared to other strains (P<0.01) which matched that of C. jejuni 81-1176, a hyper-invasive strain. Furthermore, the ovine and bovine abortion associated Campylobacter isolates showed resistance to several antimicrobials including ciprofloxacin, erythromycin, and gentamicin. Abortion-like symptoms and pathological lesions including suppurative necrotizing placentitis, suppurative and necrotic endometritis, along with occasional lymphadenitis and hepatitis were detected in pregnant ewes after inoculation with the abortion-associated C. jejuni strains. Taken together, our results showed that these C. jejuni isolates are capable of causing abortion in sheep, which indicated that the virulence characters of abortion inducing C. jejuni can be better studied in a natural host, sheep. Since this clone of C. jejuni share genotypic similarities with clones that exist in human population, it may have zoonotic potential.
Our study fills the knowledge gap of prevelance, distribution, genotypic diversity and molecular epidemiology of C.jejuni and C.coli strains from cattle (beef and dairy) as well as from European Starlings that exists particularly in the US. The results from our study would enhance preharvest efforts to establish effective control and preventive measures, which interm limite the public health impact of these pathogens. Further, the pathogenesis studies of ovine and bovine abortion-associated C. jejuni in pregnant ewes would provide a better understanding of the pathogenic mechanisms used by these pathogens and may identify novel targets to develop control strategies.