Select phytochemicals reduce Campylobacter jejuni in postharvest poultry and modulate the virulence attributes of C. jejuni

Authors: WAGLE, BASANTA - University Of Arkansas; Donoghue, Ann - Annie; Jesudhasan, Palmy

Submitted to: Frontiers in Microbiology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 7/20/2021
Publication Date: 8/12/2021
Citation: Wagle, B., Donoghue, A.M., Jesudhasan, P. 2021. Select phytochemicals reduce Campylobacter jejuni in postharvest poultry and modulate the virulence attributes of C. jejuni. Frontiers in Microbiology. 12. Article 725087. https://doi.org/10.3389/fmicb.2021.725087.
DOI: https://doi.org/10.3389/fmicb.2021.725087

Interpretive Summary: Campylobacter species are a leading cause of foodborne diarrheal illness in humans. Mainly, consuming undercooked poultry meat or handling Campylobacter-contaminated poultry products is the cause of disease in humans. Current interventions employed to reduce Campylobacter in live chickens provide limited protection, indicating an alternative approach. This study investigated the efficacy of phytochemicals, namely turmeric, curcumin, allyl sulfide, garlic, and ginger oil, to reduce Campylobacter in poultry. Two experiments were conducted on the thigh skin of the chicken. The first experiment was performed using a lower dose (0.25%) of phytochemicals, whereas a higher dose (0.5%) was used in the second experiment. Samples were challenged with Campylobacter jejuni and allowed to adhere for 30 min. Then, skin samples were dipped into their respective pre-chilled treatment solutions at 4° C for an hour and plated on agar to enumerate C. jejuni. The underlying mechanisms of action were investigated in cell culture, quorum sensing (cell to cell communication), and gene expression analyses. We found that the counts of C. jejuni were reduced with garlic or ginger oil at a concentration of 0.25% and 0.5%. These compounds were as effective as 220 ppm of peracetic acid (industry controls). The selected phytochemicals (except curcumin) reduced the adhesion of C. jejuni to chicken embryo cells. In addition, all the phytochemicals reduced the quorum sensing of C. jejuni. The cell viability test revealed that the cells treated with 0.25% of phytochemicals had damage cell membranes indicating this as a mechanism that phytochemicals use to kill C. jejuni. These studies strongly support that using the phytochemicals mentioned above in the post-harvest poultry would reduce C. jejuni in poultry meat and further reduce the incidence of human C. jejuni infection.

Technical Abstract: Consumption or handling of poultry and poultry products contaminated with Campylobacter species are a leading cause of foodborne illness in humans. Current strategies employed to reduce Campylobacter in live chickens provide inconsistent results indicating the need for an alternative approach. This study investigated the efficacy of phytochemicals namely turmeric, curcumin, allyl sulfide, garlic, and ginger oil to reduce Campylobacter jejuni in postharvest poultry and sought to delineate the underlying mechanisms of action. Two experiments were conducted on the thigh skin of the chicken and each experiment repeated twice. The first experiment used a lower dose (0.25%) of phytochemicals and the second experiment used a higher dose (0.5%). Samples were inoculated with 50 µL (~107 CFU/sample) of C. jejuni strain S-8 and allowed to adhere for 30 min. Skin samples were dipped into their respective pre-chilled treatment solutions at 4°C for an hour to simulate chilling tank treatment, followed by plating to enumerate C. jejuni (n=3 samples/treatment/trial). The underlying mechanisms of action was investigated using subinhibitory concentration (SIC) in adhesion, quorum sensing, and gene expression analyses. Adhesion assay was conducted on the monolayers of ATCC CRL-1590 chicken embryo cells challenged with C. jejuni (multiplicity of infection 10:1), incubated in the presence or absence of phytochemicals for 1.5 h followed by plating to enumerate adhered C. jejuni. Vibrio harveyi bioluminescence and LIVE/Dead BacLight™ bacterial viability assays investigated the effects of phytochemicals on quorum sensing and cell viability, respectively. In addition, droplet digital PCR determined the gene expression analyses of C. jejuni exposed to phytochemicals. The bacterial counts were logarithmically transformed, and data were analyzed by GraphPad version Prism 9. The counts of C. jejuni were reduced by 1.0-1.5 Log CFU/sample with garlic or ginger oil at a concentration of 0.25% and 0.5% (p<0.05). These compounds were as effective as 220 ppm of peracetic acid (industry controls). The select phytochemicals (except curcumin) reduced the adhesion of C. jejuni to chicken embryo cells (p<0.05). In addition, all the phytochemicals at SIC reduced quorum sensing of C. jejuni (p<0.05). The cell viability test revealed that cells treated with 0.25% of phytochemicals had compromised cell membranes indicating this as a mechanism that phytochemicals use to damage/kill C. jejuni. This study strongly supports that the application of phytochemicals in postharvest poultry would significantly reduce C. jejuni in poultry meat and further reduce the incidence of human C. jejuni infection.