A comparison of methods to detect low levels of Salmonella enterica in surface waters to support antimicrobial resistance surveillance efforts performed in multiple laboratories

Authors: KRAFT, AUTUMN - Oak Ridge Institute For Science And Education (ORISE); Wells, James - Jim; Frye, Jonathan; Ibekwe, Abasiofiok - Mark; Durso, Lisa; Hiott, Lari; East, Cheryl - Roberts; MCCONN, BETTY - Oak Ridge Institute For Science And Education (ORISE); FRANKLIN, ALISON - Us Environmental Protection Agency (EPA); BOCZEK, LAURA - Us Environmental Protection Agency (EPA); GARLAND, JAY - Us Environmental Protection Agency (EPA); KABRERA, CLAUDINE - Food And Drug Administration(FDA); MCDERMOTT, PARICK - Food And Drug Administration(FDA); OTTESEN, ANDREA - Food And Drug Administration(FDA); ZHENG, JIE - Food And Drug Administration(FDA); Cook, Kimberly - Kim; Sharma, Manan

Submitted to: Science of the Total Environment
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 9/16/2023
Publication Date: 9/28/2023
Citation: Kraft, A., Wells, J., Frye, J.G., Ibekwe, A.M., Durso, L.M., Hiott, L.M., East, C.L., Mcconn, B., Franklin, A.M., Boczek, L.A., Garland, J.L., Kabrera, C., Mcdermott, P., Ottesen, A.R., Zheng, J., Cook, K.L., Sharma, M. 2023. A comparison of methods to detect low levels of Salmonella enterica in surface waters to support antimicrobial resistance surveillance efforts performed in multiple laboratories . Science of the Total Environment. https://doi.org/10.1016/j.scitotenv.2023.167189.
DOI: https://doi.org/10.1016/j.scitotenv.2023.167189

Interpretive Summary: Antibiotic-resistant Salmonella enterica is an emerging issue in food safety. One route of contamination of antibiotic resistant Salmonella enterica onto fruits and vegetables is through agricultural or irrigation water. Recovery of antibiotic-resistant Salmonella in water is also important for other U.S. government surveillance programs, like the National Antimicrobial Resistance Monitoring and Surveillance (NARMS) program. To date, there is not a specific or standardized method used to recovery Salmonella from surface water or agricultural irrigation water. Our objective here was to compare several different microbiological methods to recover low levels of Salmonella from surface water to identify a single method that could be used a national surveillance of rivers and streams. Research here was conducted in two phases. In phase 1, three methods were compared to recover low levels of fluorescent Salmonella Typhimurium from inoculated surface water – Bulk Water (BW), vertical Modified Moore Swab (VMMS), and the modified Standard Method 9260.B2 (SM). Sixty water samples were compared across four USDA ARS laboratories on five different sites. Statistical analysis revealed that laboratory location impacted the recovery of S. Typhimurium more than the method or the date (replicate) of the experiment. However, SM recovered S. Typhimurium from 75% (15/20) of water samples while BW (12/20) and VMMS (12/20) recovered Salmonella from 60% of samples. In phase 2, dead end ultrafiltration (DEUF) was compared to SM at two different locations and 40 samples were analyzed. SM recovered Salmonella from 100% (20/20) of samples while DEUF recovered Salmonella from 95% (19/20) samples. Research presented here shows that the modified standard method 9260.B2 (SM) consistently recovered low levels of Salmonella from inoculated water samples and should be prioritized for Salmonella recovery from surface water in laboratory settings. This research benefits other federal agencies (FDA, CDC, and EPA) involved in NARMS surveillance, as well as farmers who can utilize these methods to improve their surveillance of agricultural water.

Technical Abstract: Identifying and developing effective and sensitive detection methods for antimicrobial resistant Salmonella enterica from surface water is a goal of the U.S. National Antimicrobial Resistance Monitoring System (NARMS). No specific microbiological methods used in surveillance efforts for Salmonella enterica or antimicrobial resistant S. enterica in water, has been standardized or reported in the U.S. Here we use a multi-laboratory evaluation of four methods - bulk water enrichment (BW), vertical Modified Moore Swab (VMMS), modified Standard Method 9260.B3 (SM), and dead-end ultrafiltration (DEUF) - to recover S. enterica from surface water. In Phase 1, one-liter volumes of surface water (n=60) were collected from the same site in Fall 2021 on five different dates. Water was shipped and analyzed at four different USDA ARS laboratories (A, B, C, and D) for recovery of environmentally present Salmonella and an inoculated fluorescent S. Typhimurium strain (ca. 30 CFU/L). One-liter samples (n=20) were subjected to either BW, VMMS, or SM. Overall, fluorescent S. Typhimurium and environmental Salmonella spp. were recovered from 65% (39/60) and 45% (27/60) of water samples, respectively. SM, VMMS, and BW recovered fluorescent S. Typhimurium from 75%, 60% and 60% of inoculated samples, respectively. Analysis by Chi-squared test determined that laboratory location had a significant (p < 0.05) effect on recovery compared to method or date of water collection. In Phase 2, Dead-end ultrafiltration (DEUF) was compared to SM at laboratory locations B and D to recover fluorescent S. Typhimurium (30 CFU/L) from 1-L samples. SM and DEUF recovered S. Typhimurium from 100% (20/20) and 95% (19/20) of inoculated water samples, respectively; laboratory location nor recovery method (p> 0.05) affected S. Typhimurium recovery. Results indicate that SM method consistently recovered low levels of Salmonella from inoculated water samples and should be prioritized for Salmonella recovery from surface water in laboratory settings.