-by- Jovana Kovačević Food Spoilage: “Stinkies”, “Slimies” and Biofilms October 22, 2009

10/22/20092 Outline Spoilage of Food oSpoilage Signs oMeat, Fish, Dairy oWhat do laboratory tests tell you about spoilage? Biofilms oProperties and formation oWhy are they a problem? How do we deal with spoilage and biofilms?

10/22/20093 Food Spoilage Undesirable deterioration of food quality that may result in changes in odor, taste and appearance of food.

10/22/20094 Food Spoilage Types of spoilage:  Microbial:  Bacteria, yeasts, molds  Non-microbial  Foreign materials, enzymes Changes not necessarily harmful

10/22/20095 Microbial Food Spoilage Each food unique microbial environment  unique spoilage agents for each Why do microorganisms spoil our food?  food composition ideal source of nutrients for microbes  availability of water and oxygen  suitable pH Factors that affect spoilage:  Oxygen, temperature, pH, a w, humidity

10/22/20096 Spoilage organisms Similar microflora emerge in different foods under the same conditions Affected by the: oType of food oProcessing oPreservation oStorage conditions Domination of specific spoilage microbes: oInitial counts present oNutrient composition oChemical and physical parameters

10/22/20097 Spoilage Signs Odor:  Breakdown of proteins – putrefaction e.g. “rotten egg” smell Sliminess Discoloration  Mold on bread, blue and green mold on citrus fruit and cheese

10/22/20098 Spoilage Signs… Souring  Production of acid e.g. sour milk from production of lactic acid Gas formation  Meat becomes spongy  Swollen or bubbling packages and cans

10/22/20099 Food Types: Based on rate of spoilage 1) Highly perishable  meat  fruit  milk  vegetables  eggs 2) Semi perishable  potatoes  nuts 3) Stable  rice  flour  dry beans WET DRY

10/22/ Inadequate storage temperatures Prolonged storage times Improper ventilation Cross contamination Excessive delays between receiving and storing Inadequate food safety standards Common Causes of Food Spoilage

10/22/ Meat Spoilage

10/22/ Meat spoilage… Cutting board contamination Conveyor belts Temperature Delay between storage and distribution Fecal contamination from intestines

10/22/ Meat Spoilage Continued Food Types of Spoilage Spoilage Microorganisms MEAT Fresh Putrefaction Clostridium, Pseudomonas, Proteus, Alcaligenes, Chromobacterium Souring Chromobacterium, Lactobacillus, Pseudomonas Cured Mouldy Penicillium, Aspergillus, Rhizopus Souring Pseudomonas, Micrococcus, Bacillus Greening Lactobacillus spp., Streptococcus, Pediococcus Slimy Leuconostoc Vacuum Packed Souring Greening Lactobacillus, Carnobacterium, Leuconostoc PoultryOdor, Slime Pseudomonas, Alcaligenes, Xanthomonas

10/22/ Meat Spoilage Spoilage microflora is dominated by:  Pseudomonas spp.  P. fragi, P. lundensis and P. fluorescens  Acinetobacter  Psychrobacter immobilis When surface is left to dry:  Micrococci  Yeasts  Molds

10/22/ Meat Spoilage Vacuum – packed:  Lactic acid bacteria  Enterobacteriaceae  Brochothrix thermosphacta (occasionally)  Clostridia  Shewanella putrefaciens (meat with ↑ pH)

10/22/ Fish and Eggs Spoilage

10/22/ Fish and Eggs Spoilage Continued Fish  Polluted waters  Transportation vehicles, boxes Poultry and Eggs  Human contact  Penetration by bacteria  Cracks TEMPERATURE PHYSICAL DAMAGE INTRINSIC FACTORS

10/22/ Fish & Eggs Spoilage Continued Food Types of Spoilage Spoilage Microorganisms FISH Discoloration Pseudomonas Putrefaction Chromobacterium, Halobacterium, Micrococcus EGGS Green rot Pseudomonas Colorless rot Pseudomonas, Alcaligenes, Chromobacterium Black rot Coliforms Fungal rot Proteus, Penicillium, Mucor

10/22/ Spoilage of Milk and Dairy Products

10/22/ Spoilage of Milk and Dairy Products Continued FoodTypes of SpoilageSpoilage Microorganisms DAIRY MILK (pasteurized) BitternessPseudomonas spp. SouringLactobacillus thermophilus Sweet curdlingBacillus cereus CHEESE Green discoloration Penicillium Green to black discoloration Cladosporium Black discoloration Candida Sliminess (high pH) Pseudomonas spp. “Gassy” cheese Coliforms, LAB, Clostridia

10/22/ Spoilage of Fruits and Vegetables

10/22/ Spoilage of Fruits and Vegetables Food Types of Spoilage Spoilage Microorganisms FRESH FRUITS AND VEGETABLES Bacterial soft rotErwinia carotovera, Pseudomonas spp. Gray mould rotBotryitis cinerea Rhizopus soft rotRhizopus nigrican Blue mould rotPenicillium italicum Black mould rotAspergillus niger, Alternaria Sliminess and Souring Saprophytic bacteria

10/22/ Examples of Food Quality Tests Meat & Fish: pH for vacuum packaging (< 6.0) Visual inspection for leakage / color change  Faulty seals, pin-holes Sensory test Aerobic plate count at 30°C  Indication of hygiene at packaging  Expected refrigerated shelf-life  ↑ APC = improper cleaning of equipment, unacceptable time/temperature history of raw products

10/22/ Testing for Microbial Quality Total Viable Count Enterobacteriacea Coliforms Escherichia coli Enterococci Staphilococcus aureus Bacillus spp. Clostridium perfringens Lactic acid bacteria Pseudomonas species Yeasts and molds

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10/22/ Examples of Food Submission Forms

10/22/ Total Viable Count Total Viable Count also known as:  Heterotrophic plate count (HPC)  Aerobic plate count (APC)  Total plate count (TPC)  Aerobic colony count (ACC)  Aerobic mesophilic count Purpose: The aerobic plate count gives the total number of bacteria able to grow in an oxygenated or aerobic environment. Cooked foods should have little to no bacteria present Result provides a guide for shelf-life of foods  The higher the count: prolonged storage, and/or inadequate cooking. Note: Not a useful indicator for fermented foods or fresh ready-to-eat raw fruits and vegetables.

10/22/ Enterobacteriaceae A family of Gram-negative, facultatively anaerobic, rod-shaped bacteria that do not form endospores and ferment variety of sugars Includes:  Pathogenic microbes: e.g. pathogenic E. coli, Proteus, Salmonella, Shigella, Yersinia, Enterobacter, Serratia, Klebsiella  Non-pathogenic microbes: e.g. Citrobacter, Erwinia, Hafnia

10/22/ Coliforms and E. coli Coliforms:  Total coliforms  Fecal coliforms: o Found in human and animal intestinal wastes o More precise indicator of the presence of sewage contamination than total coliforms  Four groups of fecal coliforms: oEnterobacter oKlebsiella o Escherichia oCitrobacter

10/22/ Escherichia coli Best indicator of fecal sanitation problems Characterized by the fermentation of glucose and lactose Found  In the gut of warm blooded mammals  In the gut of insects and reptiles  As natural soil microbial flora

Sanitary Quality Guidelines Food Group Indicator Test Result (CFU/g) SatisfactoryUnsatisfactory Aerobic Colony Count Canned foods & Cooked hot-held food < 5> 5 Cooked chilled food no handling < 100> 1000 Cooked chilled food with handling & Preserved foods < 100,000> 1,000,000 Long shelf life fish products, meat products, fruit and vegetable products < 1,000,000> 100,000,000 Fermented foods & Fresh fruits and vegetables Not applicable Total Coliform Count All foods except fresh fruits & vegetables, or foods containing them <100> 1000 Fecal Coliform Count All foods except fresh fruits & vegetables, or foods containing them < 3> 3 Escherichia coli Count All foods < 3> 3

10/22/ Aerobic Plate Count: Environmental Swabs Guidelines for interpretation of bacterial counts from swabs and sponges as means of monitoring the cleanliness of surfaces. Interpretation CFU Count Per Area Swabbed Log Conversion Counts on Surface* (based on 25 cm 2 surface area sampled) Clean< 45 CFU< 1.65< 5 CFU / cm 2 Contaminated140 to 260 CFU2.15 – 2.41~ 5 to 10 CFU / cm 2 Very Contaminated> 260 CFU> 2.41> 10 CFU / cm 2 ** Values will vary depending on size of the surface area sampled. To calculate the number of bacteria per cm 2 take the total CFU and divide by the surface area swabbed.

10/22/ Common Spoilage Microorganisms Shewanella putrefaciens Pseudomonas spp. oGram negative oRod shaped oMotile oAerobic oNon-spore forming oBiofilm formers gpta/interfaces_bacteries_aliments_surfaces_solides

Pseudomonas aeruginosa

10/22/ Biofilms “Variety of microorganisms arranged in a complex relationship to one another and embedded in a mass of extracellular polysaccharides of their own making” (

Biofilms Continued Communities of bacteria adhering to:  Environmental surfaces  Living tissues  Indwelling medical devices  Industrial or potable water system piping  Natural aquatic systems  Food processing facilities: oFloors, waste water pipes, bends in pipes, rubber seals, conveyor belts, stainless steel surfaces (Kumar and Anand, 1998) FEMS Microbiology Letters. 228:

10/22/ Environmental Biofilms Sewage treatment bioreactors Water pipes Legume root nodules Dental units Contact lenses Termite, ruminant digestion Fermentations

10/22/ Why do bacteria form biofilms? Protection from the environment oSanitizers oAntibiotics Nutrient availability and metabolic cooperativity Acquisition of new genetic traits

10/22/ Biofilm formation

10/22/ Biofilm formation Figure 2. Formation and development of gram-negative microorganisms’ biofilm. Adapted from Microbial Biofilms: from ecology to molecular genetics by Davey and O’Toole (2000). Microbiol. Molec. Bio. Rev. 64(4):

10/22/ Control of Microbial Spoilage Good manufacturing practices Cleaning and sanitation  Processing environment  Equipment Handling with minimal physical damage Washing (in some case) Proper storage temperature Rapid movement of food through processing plant Treatments…  preservation

10/22/ Temperature based: oChilling oRefrigeration oFreezing oPasteurization oCanning Water activity reduction oDehydration oAddition of salts and sugars Irradiation Modified Atmosphere Packaging Fermentation Treatment with chemicals o Sorbic acid, phenylphenates, diphenyl and iodophors, fumigation with sulfur-containing dusts Food preservation

10/22/ References 1. Davey and O’Toole (2000). Microbial Biofilms: from ecology to molecular genetics. Microbiol. Molec. Bio. Rev. 64(4): Forsythe, S.J, and P.R. Hayes Food Hygiene Microbiology and HACCP. 3 rd Ed. pp Aspen Publishers Inc., Maryland. 3. ICMSF Microorganisms in Foods. Microbial Ecology of Food Commodities. 2 nd Ed. pp Kluwer Academic/Plenum Publishers, New York, NY. 4. Marsh, E.J., H. Luo, H. Wang A three-tiered approach to differentiate Listeria monocytogenes biofilm-forming abilities. FEMS Microbiology Letters. 228: gpta/interfaces_bacteries_aliments_surfaces_solides

10/22/ Thank you!