Aerotolerance and Multi-Locus Sequence Typing of Campylobacter jejuni Isolated from Commercial Broiler Processing Plants

Abstract

Campylobacter jejuni is one of the leading causes of acute diarrhea in the United States. Despite being a microaerophilic pathogen, C. jejuni continues to endure within the domain of food production, especially in poultry processing. Recent research on aerotolerance indicates that close monitoring of this pathogen is necessary. A total of 40 C. jejuni isolates previously obtained from commercial broiler processing plants were analyzed for aerotolerance and genetic diversity. In addition, the effect of aerotolerance and storage time (days) on the survival of C. jejuni on broiler drumsticks at refrigeration (4 °C) and freezing conditions (-20 °C) was also evaluated. Out of 40 isolates, 25 (62.5%) were aero-sensitive (AS), 10 (25%) were intermediately aerotolerant (IAT), and 5 (12.5%) were hyper aerotolerant (HAT). The isolates belonged to four clonal complexes (CCs) and six sequence types, with the majority of isolates assigned to the CC-353 clonal complex. C. jejuni counts were reduced by 0.40 log CFU/g after 7 days at 4 °C and by 1.50 log CFU/g after 14 days at -20 °C, respectively, irrespective of aerotolerance (p < 0.001). At both refrigeration (p < 0.013) and freezing (p < 0.001), HAT showed greater reductions as compared to AS and IAT. These findings suggest that both refrigeration and freezing reduce C. jejuni counts.

Keywords: aerotolerance; clonal complex; freezing; refrigeration; survival.

Figure 1

Average Log CFU/ml of C. jejuni isolates at 24 h when incubated micro-aerobically at 42 °C in BHI broth with shaking at 200 rpm. The results show the mean and standard deviation of the three independent replicates.

Figure 2

Average Log CFU/ml of aero-sensitive C. jejuni isolates at 0 h, 12 h, and 24 h time points in an aerobic condition. The results show the mean and standard deviation of the three independent replicates. These isolates did not survive 12 h and 24 h of aerobic exposure.

Figure 3

Average Log CFU/ml of intermediate aerotolerant C. jejuni isolates at 0 h, 12 h, and 24 h time points in an aerobic condition. These isolates did not survive 24 h of aerobic exposure. The results show the mean and standard deviation of the three independent replicates.

Figure 4

Average Log CFU/ml of hyper aerotolerant C. jejuni isolates at 0 h, 12 h, and 24 h time points in an aerobic condition. The results show the mean and standard deviation of the three independent replicates.

Figure 5

Frequency distribution of C. jejuni sequence type (ST) and clonal complexes (CC) among isolates with different aerotolerance levels. AS = Aero-sensitive; IAT = intermediate aerotolerant; HAT = Hyper aerotolerant; UN = Unassigned, not assigned to any clonal complexes.

Figure 6

Dendrogram showing phylogenetic relationship between sequence type and isolates whose sequence type was novel. ST = sequence type. ST–12437, ST–12438, and ST–12439 are the new sequence types assigned by the Campylobacter PubMLST database. Evolutionary distances were computed using the p-distance method using the UPGMA method.

Figure 7

Reduction in C. jejuni counts when stored at 4 °C for 7 days (p < 0.001). Means with different letters are statistically different. The error bar represents the pooled standard error of the means.

Figure 8

Reduction in C. jejuni counts when stored at −20 °C for 14 days (p < 0.001). Means with different letters are statistically different. The error bar represents the pooled standard error of the means.