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

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 have been standardized or reported in the U.S. Here we describe 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 for recovery of environmental Salmonella and an inoculated fluorescent S. Typhimurium strain (ca. 30 CFU/L). One-liter samples (n=20) were subjected to recovery and enrichment by 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, DEUF was compared to SM at two different laboratory locations 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.

Resources in this dataset:

• Resource Title: Data Dictionary File Name: Data Dictionary AgCommons 3 9 2023.xlsx Resource Description: List of abbreviations used in data set.

• Resource Title: Water quality and rainfall data for surface water collected from MA04 site File Name: Water quality rainfall data Ag Data Commons Phase 1 Sharma 3 22 2023.xlsx Resource Description: Water quality data for surface water collected from MA04 site - surface water used for inoculation of fluorescent Salmonella Typhimurium on five different dates (Biological Replicates 1 -5) in September / October 2021 - this is termed Phase 1 in the manuscript.

• Resource Title: Phase 1 Comparison of recovery methods of fluorescent Salmonella Typhimurium using three different recovery methods File Name: Phase 1 BW SM VMMS comparison data 9 1 2022.xlsx Resource Description: Recovery methods of fluorescent Salmonella Typhimurium at four different laboratory locations (A,B,C,D) using three different recovery methods (BW, SM, VMMS).

• Resource Title: Recovery of naturally present (environmental Salmonella) from surface water recovered by three different methods File Name: Phase 1 Environmental Salmonella recovery 9 1 2023.xlsx Resource Description: Recovery of naturally present (environmental Salmonella) from surface water recovered at four different laboratory locations by three different methods (BW, SM, VMMS).

• Resource Title: Phase 2 Recovery of fluorescent Salmonella Typhimurium by SM and DEUF at two different locations File Name: Phase 2 DEUF vs SM bioball data 9 1 2022.xlsx Resource Description: Recovery of fluorescent Salmonella Typhimurium by SM and DEUF Methods using surface water from different laboratory locations (B and D)

• Resource Title: Recovery of fluorescent Salmonella Typhimurium from refrigerated surface water File Name: Refrigeration Data Summary Tabl.xlsx Resource Description: Recovery of fluorescent Salmonella Typhimurium from surface water refrigerated for 7 days before recovery by BW, SM, or VMMS.