Improved Sampling and Analytical Methods for Testing Agricultural Water for Pathogens, Surrogates and Source Tracking Indicators

<p>Technical Abstract: The new Produce Safety and Preventive Controls for Human Food Rules established under the Food Safety Modernization Act (FSMA) identify monitoring frequencies and Escherichia coli concentrations for characterizing agricultural water quality. In addition to monitoring for E. coli, risk-based collection and testing of agricultural source water for pathogens and alternative water quality surrogates can provide farm operators with a better understanding of the quality of water used in crop production. In this project, an emerging water sampling technique—ultrafiltration (UF)—will be used to collect large-volume irrigation water samples from three farms in Georgia to investigate the benefits of collecting large-volume samples for pathogens and alternative microbial water quality parameters. UF and grab samples will be collected at least every month for one year from the primary irrigation pond at each of the three farms. Precipitation-impacted samples will be collected for four rain events to enhance the comparison of large- versus small-volume collection procedures and evaluate this risk-based sampling approach. UF and grab samples will be analyzed for Salmonella, Cryptosporidium, E. coli O157:H7, pathogen surrogates, and indicators of fecal contamination [including “microbial source tracking” (MST) analytes], which can identify potential sources of fecal contamination contributing to elevated microbial levels. Controlled laboratory experiments will also be conducted to determine the effectiveness of UF for recovering Salmonella, Cryptosporidium and Cyclospora cayatenensis from agricultural water.</p>

<p>This two-year project is a collaboration between the Centers for Disease Control and Prevention (CDC), University of Georgia (UGA), and Emory University (Emory). UGA staff will collect UF and grab samples from three farms in the area of Tifton, Georgia to enable statistical comparison of microbial detection rates using small-volume and large-volume sampling. UGA staff will provide the samples to CDC and Emory staff, who will process and analyze the samples at CDC in Atlanta, Georgia. Samples will be analyzed for E. coli, enterococci, F+ coliphages, and Salmonella using quantitative culture techniques. Salmonella will also be serotyped to identify potential host-specific serovars. Cryptosporidium will be analyzed by real-time PCR and sequencing to identify Cryptosporidium genotypes and subtypes for risk characterization (potential human infection risks) and fecal source tracking (identification of human and non-human animal hosts in the watershed of the farms). E. coli O157:H7 will be analyzed by real-time PCR and isothermal amplification, in conjunction with propidium monoazide (PMA) to detect the presence of viable cells. Molecular microbial source tracking analytes will include human-specific analytes [Bacteroides (HF183 human marker), human polyomaviruses BK and JC (HpYVs), and Methanobrevibacter smithii ], and mitochondrial markers for domestic animals (dogs, cats), wildlife (birds and deer), and livestock (cow, chicken, horse, pig). </p>

<p>This project will result in large-volume water sample collection and testing protocols to enable sensitive detection of pathogens, pathogen surrogates, and fecal source tracking analytes. Produce industry operators and technical consultants can use these tools to better characterize the microbial quality of irrigation water. This project will also provide an evidence base for evaluating seasonal and precipitation-related sampling as risk-based approaches for agricultural water quality monitoring.</p>