Effect of post-harvest interventions on surficial carrot bacterial community dynamics, pathogen survival, and antibiotic resistance

Abstract

Strategies to mitigate antibiotic-resistant bacteria (ARB), including human pathogens, on raw vegetables are needed to reduce incidence of resistant infections. The objective of this research was to determine the effectiveness of standard post-harvest interventions, sanitizer washing and cold storage, to reduce ARB, including antibiotic resistant strains of the human pathogen E. coli O15:H7 and a common spoilage bacterium Pseudomonas, on raw carrots. To provide a background inoculum representing potential pre-harvest carryover of ARB, carrots were dip-inoculated in dairy cow manure compost slurry and further inoculated with known ARB. Inoculated carrots were washed with one of three treatments: sodium hypochlorite (50 ppm free chlorine), peroxyacetic acid (40 ppm peroxyacetic acid; 11.2% hydrogen peroxide), tap water (no sanitizer), or no washing (control). Washed carrots were air dried, packaged and stored at 10 °C for 7d or 2 °C for up to 60 d. Enumeration was performed using total heterotrophic plate counts (HPC), HPCs on antibiotic-containing media ("ARBs"), E. coli O157:H7, and Pseudomonas sp. immediately after washing (0 d) and after 7 d of storage. In addition to the cultured bacteria, changes to the surficial carrot microbiota were profiled by sequencing bacterial 16S rRNA gene amplicons to determine the effect of sanitizer wash, storage temperature, and time of storage (0, 1, 7, 14 and 60 d). Storage temperature, addition of a sanitizer during wash, and duration of storage significantly affected the bacterial microbiota (Wilcoxon, p < 0.05). Inclusion of either sanitizer in the wash water significantly reduced the log CFU/g of E. coli O157:H7 and Pseudomonas sp., as well as HPCs enumerated on cefotaxime- (10 μg/ml), sulfamethoxazole- (100 μg/ml), or tetracycline (3 μg/ml) supplemented media compared to the unwashed control (ANOVA, p < 0.05). However, no significant reductions to bacteria resistant to vancomycin or clindamycin occurred after washing and storage. Members of the Proteobactetria, Firmicutes, Actinobacteria, Planctomycetes, and Acidobacteria comprised the bacterial carrot microbiota. The diversity of the carrot microbiota was significantly affected by the temperature of storage and by extended storage (60 d), when spoilage began to occur. There were no significant differences to the relative abundance of bacterial groups associated with the type of sanitizer used for washing. Results of this study indicate that inclusion of a sanitizer in wash water, followed by storage at 2 °C, might be an effective strategy to prevent re-growth of pathogenic E. coli O157:H7 and reduce levels of bacteria resistant to certain antibiotics on carrots.

Keywords: Escherichia coli O157; Low temperature storage; Microbiota; Produce; Pseudomonas; Sanitizer.