Project Number: 5070-12130-006-22-N
Project Type: Non-Funded Cooperative Agreement
Start Date: May 15, 2020
End Date: May 14, 2023
1. Determine distribution and prevalence of various clades of cryptic E. coli in the US. 2. Evaluate the fate of enteric members of cryptic E. coli vs. real E. coli in aquatic environment.
Objective 1. Task 1: Fecal and water sample collections: Suitable sampling sites that represent specific dominant land uses will be identified. Ten fecal samples will be collected in Missouri and Virginia from each of the host species considered as the major sources of fecal pollution, including human (wastewater treatment plant), livestock (cattle, chicken, pig, and turkey), wild animals (deer and raccoon), and waterfowls (duck and goose). At least 20 water samples (4 samples at 5 locations) will be collected in Missouri and Virginia respectively, from the drainage area of each type of land uses, including at least urban, forest, cropland (without manure as the fertilizer), and grassland. Task 2: Isolations and identification of presumptive E. coli using the standard methods: Cooperator will isolate presumptive E. coli from the aforementioned fecal and water samples using the EPA Method 1603 for water samples and a modified procedure for fecal samples. Task 3: DNA extraction from the presumptive E. coli isolates: Cooperator will use polymerase chain reaction (PCR) assay for the identification. Task 4: PCR screening of cryptic E. coli strains in the presumptive E. coli. Cooperator will test the DNA of each strain of the aforementioned presumptive E. coli isolates for the presence of cryptic E. coli using a PCR assay. Task 5: Data analyses for the studies of Objective 1: The concentration of cryptic E. coli will be determined for each water sample. The mean cryptic E. coli concentrations at the four sampling sites will be analyzed as a function of dominant land use using one-way analysis of variance (ANOVA). Land use will be categorized by the dominant land use: cropland, forest, grassland, and urban. A fifth sampling point will be characteristic of either mixed land use in the drainage area, or of another land use. Objective 2: Evaluate the fate of enteric members of cryptic E. coli vs. real E. coli in aquatic environment. The effects of temperature, pH, and UV radiation on E. coli versus cryptic E. coli will be determined in a water microcosm. The ARS PI and Cooperator as well as other team members will have monthly meetings to discuss research and data analysis methods, interpretation of results, and related studies reported in the literature. The ARS PI will be part of the Master's student committee.”