# Inoculum growth impacts Salmonella and Shiga-toxin producing Escherichia coli resilience on wheat grain

## TL;DR

This study shows how different ways of growing bacteria affect their survival on wheat grain and how tempering treatments can reduce their presence.

## Contribution

The study introduces a systematic comparison of five inoculum growth methods for Salmonella and STEC on wheat grain.

## Key findings

- Acid-adapted STEC showed increased desiccation tolerance compared to other methods.
- Broth-grown pathogens were better modeled by the Weibull model, while others followed a log-linear pattern.
- Tempering with sodium bisulfate achieved the highest pathogen reduction (1.96 log CFU/g).

## Abstract

Previous studies have shown that using a surface growth method for preparing inoculum impacted pathogen population stability and inactivation kinetics. Here, we quantified the desiccation survival and responses to tempering treatment of Salmonella and Shiga-toxin producing Escherichia coli (STEC) on wheat grain when grown using five different methods: broth, lawn-aerobic, lawn-anaerobic, lawn-acid-adapted, and lawn-low inoculum. Three strains of Salmonella and STEC each were individually inoculated onto wheat grain, conditioned to 0.45 aw and stored up to 24 weeks. Pathogen survival curves on grain were different when inoculated with a lawn-grown culture compared to broth-grown. Acid adaptation of STEC led to increased (P < 0.001) tolerance to desiccation, while no change in Salmonella desiccation tolerance was observed. While most survival curves were better described by the log-linear model, survival of pathogens from broth-grown inoculum was better characterized by the Weibull model. Five-strain cocktails of each species were inoculated onto wheat, stored for 1, 2, 7, 28, and 84 days, followed by tempering for 18 h with three chemical interventions: water (control), 800 ppm chlorine, 5% lactic acid (LA) + 26.6% NaCl and 5% sodium bisulfate (SBS). These treatments led to different (P < 0.001) average reductions of 0.52, 1.04, 1.43, and 1.96 log CFU/g, respectively. The length of storage and inoculum growth method affected (P < 0.001) pathogen survival during tempering. In general, pathogens inoculated on grain survived better during tempering when inoculum was prepared by acid-adapted ≥ lawn-aerobic ≥ low inoculum > lawn-anaerobic = broth.

Outbreaks linked to wheat flour increased interest in evaluating pathogen survival kinetics. With minimal information on how foodborne pathogens contaminate wheat grain, the “worst-case scenario” should be identified to characterize pathogen survival kinetics on grain and be used to assess the effectiveness of food safety interventions. Using an antimicrobial solution during wheat tempering, an existing unit operation where grain is exposed to water prior to milling into flour can be a cost-effective way to mitigate the risk of foodborne pathogens. The lack of consistent inoculum preparation methods makes it difficult to compare results across studies evaluating tempering treatments. We assessed five different inoculum growth methods to quantify pathogen survival during desiccation and long-term storage and pathogen inactivation efficacy of several existing tempering solutions. In addition, these data provide insights on statistically important parameters to consider for low-moisture food challenge study experimental design, such as inoculum growth, inoculation level, and pathogen adaptation.