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ARS Home » Plains Area » Clay Center, Nebraska » U.S. Meat Animal Research Center » Meat Safety and Quality » Research » Research Project #439758

Research Project: Effects of Deep Cleaning Sanitation on Biofilms and Pathogens

Location: Meat Safety and Quality

Project Number: 3040-42000-021-005-T
Project Type: Trust Fund Cooperative Agreement

Start Date: Mar 1, 2021
End Date: Jun 1, 2022

Evidence suggests biofilms may play a role in high event periods (HEPs) of pathogen detection in beef trimmings. HEP strains of E. coli O157:H7 and Salmonella have been found to form stronger biofilms and resist sanitizers to a greater extent than control strains from other sources. Further, our recent research using biofilms recovered from floor drains at a processor experiencing an increased number of events, has shown these biofilms recruited and protected E. coli O157:H7 to a significantly greater extent than biofilms recovered from drains at a control plant not experiencing similar problems. One of the actions taken to address events of E. coli O157:H7 is a deep cleaning and sanitizing of the processing plant environment. Sanitizers such as DECON7 are often used to disrupt any biofilms present during these enhanced sanitation protocols. Deep cleaning sanitation is also a part of routine food safety systems that many large processors preform annually or semiannually. Anecdotal evidence provides mixed results on the efficacy of deep cleaning. Reports range from “everything was great after sanitation” to “I think we made it mad, and events increased”. Reports in the literature are scant with regards to effects of sanitizing strategies on naturally occurring biofilms. Therefore, this study proposes to directly examine the effects of deep sanitizing on processing plant floor drain biofilms before and after a DECON7 treatment, then follow the treated drains over time to monitor how the biofilms recover, and interact with pathogens (E. coli O157:H7, non-O157 STEC, and Salmonella). The specific objectives are: 1. Identify the immediate effects of deep sanitization on processing plant biofilms. 2. Examine biofilm recovery following deep sanitization of processing plant. 3. Determine the points at which biofilms are most likely to start recruiting and protecting pathogens.

We have identified large processors willing to permit sample collection, and a pilot set of samples has already been collected. Samples will be collected from 12 floor drains per visit. Drains will span from hot scale to fabrication, with 2-3 drains sampled from each area: hot box, cooler, and fabrication, and include samples from trim storage and grind rooms where present. Each sample will be examined for general bacterial counts (APC, mesophiles, psychrophiles, etc…) and its biofilm forming ability measured. The community structure of each biofilm sample will be determined through 16S metagenomic sequencing and analysis (pathogen groups may be filtered out of the analysis if requested by cooperating processor). Each sample will be examined for its ability to recruit and protect co-inoculated strains of E. coli O157:H7 (3 strains pooled), non-O157 STEC (2 strains each O-group pooled), and Salmonella (3 strains pooled) in mixed biofilms. The influence of the pathogen on the biofilm community will be examined (16S metagenomics) on a subset of 6 drain types (4 for each pathogen). We have extensive experience preforming these sorts of assays and will follow established protocols (Wang 2020). Three processing plants in total will be examined with 8 total sample visits planned (2-3 days pre-sanitization, 2-3 days post-sanitization, then 1, 2, 4, 8, 12, and 16 weeks post-sanitization) with the same drains visited as frequently as possible. We have encountered during the pilot sample collection that some drains may be inaccessible for a variety of reasons so will select a suitable substitute drain if necessary. In short, 3 plants x 12 drains x 8 time points = 288 samples for analysis of ACP, EBC, E. coli/coliforms, mesophiles, psychrofiles by PetriFilm. Each sample used in a total of 9 biofilm biofilm assays: alone, with E. coli O157:H7, non-O157 STEC (n=6) and Salmonella. 16S metagenomic analysis of the 288 samples + select pathogen/biofilm assays (n=72) for a total of 360 metagenomic samples.