Biofilms 101 Remediation Strategies in Meat Processing

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1 Biofilms 101 Remediation Strategies in Meat Processing Jeremy M. Adler Director of Technology and Innovation American Meat Science Association April 28, 2016 Webinar

2 Biofilms How does Pseudomonas know where it is, in relation to the human body? J.W. Costerton Costerton, JW. The Bioflim Primer. Springer, 2007.

3 Bacteria Growth Planktonic bacteria Bacteria in suspension Traditional usage/perception Sesile bacteria Bacteria in a biofilm Non traditional usage/perception Bacteria prefer growth in a biofilm!

4 Biofilm Growth Cycle BIOFILM

5 Biofilms Examples Not always visible Best visible examples are usually found in piping Water transfer pipe Drain (Courtesy of Montana State University Center for Biofilm Engineering)

6 Listeria Monocytogenes Biofilms

7 Biofilm Formation Increases Bacterial Survival Biofilm is a bacteria s personal protection equipment! Protects from chemical exposure Shields from chemical contact Inactivates chemical compounds Helps resist physical removal Aids in attachment Unifies colony Insulates from heat exposure Protects from dehydration Helps bacteria remain viable

8 Environmental Biofilms in Meat Processing Major area of concern for food processors especially for ready to eat items in wet processing areas Typically tolerate to conventional cleaning and sanitation processes Frequent culprit where persistent bacterial contamination is a problem Very common in drains Specialty products required for removal

9 Environmental Biofilms in Meat Processing Biofilms are a leading source of bacteria in the food plant environment because of the presence of moisture Many of the problem bacteria such as Listeria and Salmonella readily produce biofilm protected colonies Not removing the biofilm means that colonies continue to produce more bacteria, releasing them into the plant Most general cleaning and sanitation products will not penetrate or detach the sturdy polysaccharide biofilm matrix.

10 Recovered Bacteria (log CFU/cm 2 ) E. Coli O157:H7 Growth on Stainless Steel Stainless steel coupons incubated in sterile and unsterile beef grinder washings Total Aerobic Bacteria (Unsterile washings) E. coli O157:H7 (Sterile washings) E. coli O157:H7 in the presence of competing flora (Unsterile washings) Adler, J.M., PhD Thesis, Colorado State University.

11 Strength of attachment (Sr) E. coli O157:H7 attachment to stainless steel Stainless steel coupons incubated in sterile and unsterile beef grinder washings 0.4 Day Day 1 Day 2 Day E. coli O157:H7 with other flora E. coli O157:H7 Adler, J.M., PhD Thesis, Colorado State University.

12 Recent Data from the USDA

13 Cleaning and Sanitation An ounce of prevention is worth a pound of cure! Ben Franklin

14 Mechanisms of Cleaning Cleaning and Sanitation 101 Primary Hydrolysis Breaking compounds apart by reacting with water Dissolution Dissolving soils into a liquid Displacement/Dispersion Lifting of a soil into a liquid suspension Emulsification Dispersing oil in water Supplemental Peptizing Breaking soil into smaller pieces Chelation Complexing ions into water Buffering Stabilizing the ph of a cleaner for optimal performance

15 Cleaning and Sanitation Biofilm Removal Soils Visible Any substance that is found on a surface but is not an intended part of that surface and therefore is unwanted. Dirt, mineral deposits, protein, carbohydrates, fats, rust. Soils harbor, and provide food for, microorganisms Removed with the right cleaner Invisible Microorganisms Inactivated with the right sanitizer

16 Cleaning and Sanitation Biofilm Removal Cleaners Chemicals are selected to remove soils. Selected for the specific soil, soil adhesion and substrate to be cleaned. Alkaline Cleaners: ph > 9 Dissolve proteins, sugars, and starches Dispersion and saponification fats Acid Cleaners: ph < 2 Dissolve proteins, sugars, starches, and minerals Solvents Dissolve fats, greases, and oils Oxidizing Agents Hydrolyze proteins

17 Cleaning and Sanitation Biofilm Removal Cleaners often need additional energy to work effectively Scrubbing/Mechanical action Manual scrubbing during washing reduced the recovery of bacteria. Scrubbing increased the passing scores of ATP testing. Farrell et al Journal of Food Protection 61:

18 Fat Removal Cleaning and Sanitation Biofilm Removal Cleaners often need additional energy to work effectively Temperature 100% 80% 60% 40% Water Cleaner 20% 0% Temperature (C)

19 CFU/15 liters air Cleaning and Sanitation Biofilm Removal Cleaners often need additional energy to work effectively Pressure AIRBORNE MICROORGANISMS AS DETERMINED USING AN AIR SAMPLER bar 20 bar :00 1:15 1:30 1:45 2:00 2:15 2:30 2:45 Time (hrs) after rinsing that the samples were obtained Banner et al Food Microbiology and Food Safety into the Next Millennium.

20 Cleaning and Sanitation Biofilm Removal Cleaners often need additional energy to work effectively Chemical energy Chemical reactions provide the energy to break up soils Peroxyacetic acid = Acetic acid + water Foaming Only chemical available to do work is at the surface interface Insure 360 contact Increases cleaner to soil contact time

21 Effective Cleaning is 99% of the sanitation job. The remaining 1% is the job of sanitizers

22 Cleaning and Sanitation Biofilm Removal Sanitizers No Rinse Sanitizer 5-log reduction of suspended bacteria in 30 seconds Nonfood Contact surfaces 3-log reduction of suspended bacteria in 30 seconds Definition (EPA, FDA) is based on suspended bacteria Definition is NOT based on bacteria in a biofilm Label explains proper usage

23 log CFU/cm Cleaning and Sanitation E. coli O157:H7 in a biofilm 8 1 min exposure 10 min exposure 7 8 Beef-grinder washings fold diluted TSB DL 0 0 Water PAOAQUAT SH Water PAOA QUAT SH Water Sanitizer PAOA QUAT SH Adler, J.M., PhD Thesis, Colorado State University.

24 Sanitizing a Unclean Surface Quaternary Ammonium Compounds Salmonella inoculated into almond dust treated with Quat Quat sanitizers cannot penetrate into soils Residual water allowed growth in dry particulate matter Wen-Xian et al. Journal of Food Science 75:M7-M13.

25 Cleaning and Sanitation Targeting Biofilms Treat biofilm as a soil Cleaning removes and inactivates biofilm Sanitizer alone won t inactivate biofilm Soiled surfaces aid in bacterial attachment Assure sanitary design of facility and equipment Niches and harborage points need to be eliminated in order to eliminate biofilm. Address bacterial accumulation during production Sanitizing belt sprays during production Sanitize and/or clean during production breaks

26 Recovered Bacteria (log CFU/cm 2 ) Sanitizing Belt Spray During Production Peroxyacetic acid 3 A 2 B 1 0 C C Detection Limit No treatment PAA, Trt 1 PAA, Trt 2 Down process

27 Cleaning and Sanitation Remediation Targeting Biofilms Disinfect Zones 1 3 Zone 1: Product contact surfaces Zone 2: Non-product contact surfaces close to Zone 1 Zone 3: Non-product contact surfaces in processing area Follow written procedures What is available works if applied correctly Use a peroxide based cleaner, disinfectant, or sanitizer PAA boosted caustic cleaner Hydrogen peroxide followed by caustic (CIP) Hydrogen peroxide and quaternary ammonium (PerQuat) PAA

28 Biofilm Control Chemistry We know that oxidizers and acidic chemicals are effective in penetrating, breaking down and destroying biofilms These are the most important products to control biofilm growth Chlorinated Alkaline Cleaner: At proper alkalinity and chlorine content for optimum cleaning Hydrogen peroxide additive for Soft Metal Safe Cleaning at proper concentration Acid Cleaners for removal of inorganic buildups that may entrap biofilms Acidic Sanitizers: Need to use at disinfection levels Acidic quat, Acid based iodine, Anionic acid sanitizers Peroxyacetic Sanitizers Sterilex Ultra-Kleen: (Peroxide/Quat Disinfectant/Cleaner)

29 Cleaning and Sanitation Novel treatments Phage Enzymes Microbial exclusion/metabolite molecules New treatments must consider the following: Plant environment (temperature, water etc.) ph of applied cleaners and sanitizers Oxidizing potential of cleaners and sanitizers C&S time resources (Simoes, et al A review of current and emergent biofilm control strategies. Food Sci. and Tech. 43: )

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