Toward the Adoption of Loop-Mediated Isothermal Amplification for Salmonella Screening at the National Antimicrobial Resistance Monitoring System's Retail Meat Sites

Abstract

The National Antimicrobial Resistance Monitoring System (NARMS) is a One Health program in the United States that collects data on antimicrobial resistance in enteric bacteria from humans, animals, and the environment. Salmonella is a major pathogen tracked by the NARMS retail meat arm but currently lacks a uniform screening method. We evaluated a loop-mediated isothermal amplification (LAMP) assay for the rapid screening of Salmonella from 69 NARMS retail meat and poultry samples. All samples were processed side by side for culture isolation using two protocols, one from NARMS and the other one described in the U.S. Food and Drug Administration's Bacteriological Analytical Manual (BAM). Overall, 10 (14.5%) samples screened positive by the Salmonella LAMP assay. Of those, six were culture-confirmed by the NARMS protocol and six by the BAM method with overlap on four samples. No Salmonella isolates were recovered from samples that screened negative with LAMP. These results suggested 100% sensitivity for LAMP in reference to culture. Antimicrobial susceptibility testing and whole-genome sequencing analysis confirmed identities of these isolates. Using the BAM protocol, all Salmonella isolates were recovered from samples undergoing Rappaport-Vassiliadis medium selective enrichment and presumptive colonies (n = 130) were dominated by Hafnia alvei (44.6%), Proteus mirabilis (22.3%), and Morganella morganii (9.9%) based on matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. This method comparison study clearly demonstrated the benefit of a rapid, robust, and highly sensitive molecular screening method in streamlining the laboratory workflow. Fourteen NARMS retail meat sites further verified the performance of this assay using a portion of their routine samples, reporting an overall specificity of 98.8% and sensitivity of 90%. As of July 2022, the vast majority of NARMS retail meat sites have adopted the Salmonella LAMP assay for rapid screening of Salmonella in all samples.

Keywords: LAMP; NARMS; Salmonella; antimicrobial resistance; monitoring; screening.

FIG. 1.

A schematic diagram of the design and workflow of this method comparison study. *Maryland site used the 3M Molecular Detection Assay 2—Salmonella as a screening method. BAP, blood agar plate; BPW, buffered peptone water; BS, bismuth sulfite agar; CVM, U.S. Food and Drug Administration’s Center for Veterinary Medicine; HE, Hektoen enteric agar; LAMP, loop-mediated isothermal amplification; LIA, lysine iron agar slant; MALDI, matrix-assisted laser desorption/ionization; NARMS, National Antimicrobial Resistance Monitoring System; RV, Rappaport-Vassiliadis medium; RVR10, Rappaport-Vassiliadis R10 broth; TSI, triple sugar iron slant; TT, tetrathionate broth; XLD, xylose lysine desoxycholate agar; XLT4, xylose lysine Tergitol 4 agar.