Use of Bacteroidales Microbial Source Tracking To Monitor Fecal Contamination in Fresh Produce Production

Abstract

In recent decades, fresh and minimally processed produce items have been associated with an increasing proportion of foodborne illnesses. Most pathogens associated with fresh produce are enteric (fecal) in origin, and contamination can occur anywhere along the farm-to-fork chain. Microbial source tracking (MST) is a tool developed in the environmental microbiology field to identify and quantify the dominant source(s) of fecal contamination. This study investigated the utility of an MST method based on Bacteroidales 16S rRNA gene sequences as a means of identifying potential fecal contamination, and its source, in the fresh produce production environment. The method was applied to rinses of fresh produce, source and irrigation waters, and harvester hand rinses collected over the course of 1 year from nine farms (growing tomatoes, jalapeño peppers, and cantaloupe) in Northern Mexico. Of 174 samples, 39% were positive for a universal Bacteroidales marker (AllBac), including 66% of samples from cantaloupe farms (3.6 log 10 genome equivalence copies [GEC]/100 ml), 31% of samples from tomato farms (1.7 log 10 GEC/100 ml), and 18% of samples from jalapeño farms (1.5 log 10 GEC/100 ml). Of 68 AllBac-positive samples, 46% were positive for one of three human-specific markers, and none were positive for a bovine-specific marker. There was no statistically significant correlation between Bacteroidales and generic Escherichia coli across all samples. This study provides evidence that Bacteroidales markers may serve as alternative indicators for fecal contamination in fresh produce production, allowing for determination of both general contamination and that derived from the human host.

Efficacy of two hygiene methods to reduce soil and microbial contamination on farmworker hands during harvest

“…Remaining sample rinsate was stored at 4 C for no more than 72 h and reprocessed by filtration and plating for cases in which colony counts were inconsistent or outside of assay detection limits. The final sample rinsate volumes processed ranged from 0.01 ml to 50 ml (Ravaliya et al, 2014).…”

“…A second aliquot was sent overnight on ice to the Department of Food, Bioprocessing, and Nutrition Sciences, North Carolina State University for microbial source tracking analysis. Detection and quantification of the universal and human-specific 16S rDNA markers for Bacteroidales were performed using the AllBac and BFD primers and probes as described (Ravaliya et al, 2014). The remaining sample was processed as follows to detect and enumerate coliforms, generic E. coli, and Enterococcus (three common, non-pathogenic bacteria) hereafter called "indicator bacteria" as described (Heredia et al, 2015).…”

“…Food handlers who practice poor hand hygiene may increase the risk of microbial transmission to produce (Altekruse, Timbo, Mowbray, Bean, & Potter, 1998;Bryan, 1988;Michaels et al, 2004). Our group has shown that microbial indicators on produce are associated with those on agricultural worker hands, suggesting that hand contamination contributes to the final microbial profile and load on produce (Lickness, 2014;Ravaliya et al, 2014).…”

An environmental survey of surface waters using mitochondrial DNA from human, bovine and porcine origin as fecal source tracking markers

“…In parallel, we used the Bacteroidales HF183 marker, a commonly used FST marker for human fecal contamination (Ahmed et al, 2012;Chase et al, 2012;Edge et al, 2010;Fremaux et al, 2009;Green et al, 2014;Harwood et al, 2014;Marti et al, 2013;Ravaliya et al, 2014;Sauer et al, 2011;Schill and Mathes, 2008;Stoeckel et al, 2011;Tambalo et al, 2012b;Vuong et al, 2013), for assessing the robustness of using mtDNA as a human FST marker. Quantitative PCR was used to measure the concentration of human and bovine mtDNA along with the HF183 marker.…”

Inactivation of Escherichia Coli O157:h7 on Green Bell Pepper by Chlorine Dioxide Simulating Packinghouses Process

“…In the United States, every year occurs 9.4 million episodes of foodborne illnesses, 55,961 hospitalizations and 1,351 deaths; much of these cases, reported as outbreaks, have been related to the consumption of contaminated fruits and vegetables, (Scallan et al, 2011;CDC, 2006;CDC, 2008;CDC, 2013). Several studies using microbial source tracking indicate that cross-contamination offers to bacteria, parasites and virus a pathway to reach fruits and vegetables (Beuchat, 1996;Ravaliya et al, 2014;Harwood et al, 2014). Microbial contamination throughout the production chain may occur as a consequence of inadequate or scarce sanitation process during production chain, resulting in diverse punctual contamination sources, especially when Good Agricultural and Manufacturing Practices are not strictly followed (Cummings et Pathogenic bacteria, such as Escherichia coli O157:H7 has been responsible for at least 18 foodborne outbreaks in the last ten years in the United States, and nine of these were related to vegetables (CDC, 2016) causing economic losses up to $271 million dollars per year (Hoffmann et al, 2015).…”