Genotypic and phenotypic properties of cattle-associated Campylobacter and their implications to public health in the USA

Abstract

Since cattle are a major source of food and the cattle industry engages people from farms to processing plants and meat markets, it is conceivable that beef-products contaminated with Campylobacter spp. would pose a significant public health concern. To better understand the epidemiology of cattle-associated Campylobacter spp. in the USA, we characterized the prevalence, genotypic and phenotypic properties of these pathogens. Campylobacter were detected in 181 (19.2%) out of 944 fecal samples. Specifically, 71 C. jejuni, 132 C. coli, and 10 other Campylobacter spp. were identified. The prevalence of Campylobacter varied regionally and was significantly (P<0.05) higher in fecal samples collected from the South (32.8%) as compared to those from the North (14.8%), Midwest (15.83%), and East (12%). 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. In addition, 13 new C. jejuni and two C. coli sequence types (STs) were detected by Multi Locus Sequence Typing (MLST). C. jejuni associated with clinically human health important sequence type, ST-61 which was not previously reported in the USA, was identified in the present study. Most frequently observed clonal complexes (CC) were CC ST-21, CC ST-42, and CC ST-61, which are also common in humans. Further, the cattle associated C. jejuni strains showed varying invasion and intracellular survival capacity; however, C. coli strains showed a lower invasion and intracellular survival potential compared to C. jejuni strains. Furthermore, many cattle associated Campylobacter isolates showed resistance to several antimicrobials including ciprofloxacin, erythromycin, and gentamicin. Taken together, our results highlight the importance of cattle as a potential reservoir for clinically important Campylobacter.

Figure 1. Dendrogram showing the Pulsed-field gel electrophoresis patterns for SmaI restricted cattle-associated C. jejuni (A) and C. coli (B).

Similarity analysis was performed using the Dice coefficient, and clustering was performed by the unweighted pair-group method with arithmetic averages UPGMA (optimization, 1% and position tolerance, 1.5%). Clustering cut-off was 75% similarity, which was represented by dashed line. Numbers on bootstraps represent Cophenetic correlations. N = North, S = South, M = Midwest, and E = East. Cj = C. jejuni, CC = C. coli.

Figure 2. Antimicrobial resistance of Campylobacter isolated from cattle.

C. jejuni (A) and C. coli (B) isolates obtained from four different locations across the US. Percentage of isolates resistant to different antimicrobials used in this study is shown. C. jejuni 81–176 and C. coli ATCC 33559 strains were used for quality control.

Figure 3. Invasion and intracellular survival of cattle C. jejuni isolates in INT407 cells.

A. CFU ml⁻¹ representing the number of the internalized bacteria which could be retrieved after treatment of cells with gentamicin. B. Intracellular survival of C. jejuni isolates in INT407cells. CFU ml⁻¹ representing the numbers of internalized bacteria retrieved after 24 h of incubation following the gentamicin treatment. The INT407 were infected with 1∶100 MOI of C. jejuni strains. C. jejuni 81–176 and NCTC11168 were used as controls. The detection limit of the assay is represented by the dashed line. Each bar represents the mean ± SE of three independent experiments done in duplicates for each sample (P<0.01).

Figure 4. Invasion and intracellular survival of cattle C. coli isolates in INT407 cells.

A. CFU ml⁻¹ representing the number of the internalized bacteria which could be retrieved after treatment of cells with gentamicin. B. Intracellular survival of C. coli isolates in INT407cells. CFU ml⁻¹ representing the numbers of internalized bacteria retrieved after 24 h of incubation following the gentamicin treatment. The INT407 cells were infected with 1∶100 MOI of C. coli strains. C. coli (ATCC 33559) and C. jejuni NCTC11168 were used as controls. The detection limit of the assay is represented by the dashed line. Each bar represents the mean ± SE of three independent experiments done in duplicates for each sample (P<0.01).