MICROBIAL FOOD SPOILAGE

Introduction Microbial Food Spoilage Factors Affecting Spoilage Microflora Microbial Spoilage How Does It Manifest Itself? Foods Affected By Microbial Spoilage Spoilage Microorganisms Some of The More Important Moulds Responsible For Post-Harvest Diseases of Fruits Genera of Fungi Causing Important Spoilage of Fresh And Stored Vegetables Yeast Spoilage of Fresh Fruits And Vegetables Bacteria That Cause Soft Rot of Vegetables And Fruit Monitoring And Detection of Microbial Spoilage Microbial Indicators of Product Quality Prevention of Microbial Spoilage

CONTENTS:

INTRODUCTION Food spoilage can be defined as any sensory change (tactile, visual, olefactory or flavour) which the consumer considers to be unacceptable. Spoilage may occur at any stage along the food chain. Spoilage may arise from: - insect damage - physical damage (bruising, freezing, drying, etc) - indigenous enzyme activity in the animal or plant tissue - chemical changes (usually involving oxygen) Spoilage is therefore complex, involving physical, chemical, biochemical and biological changes. MICROBIAL FOOD SPOILAGE By bacteria, yeasts and moulds: Bacteria Gram -ve Cocci Rods Non-spore Formers Gram +ve Spore Formers Lactic Acid Bacteria

Coliform/ Pseudomonas Micrococcus Bacillus Lactobacillus Enteric Acinetobacter Brocothrix Clostridium Streptococcus Citrobacter Aeromonas Leuconostoc Serratia Alcaligenes Pediococcus Proteus Moraxella Escherichia Altermonas Enterobacter Flavobacterium Erwinia Klebsiella Hafnia Yeasts: Candida, Saccharomyces, Zygosaccharomyces, Torulospora, Rhodotolura, Pichia, Moulds: Aspergillus, Mucor, Penecillium, Rhizopus, Geotrichum, Botrytis, Cladosporium, Byssochlamys

FACTORS AFFECTING SPOILAGE MICROFLORA 1. Initial microbial flora 2. Food type/intrinsic properties: - pH - Aw (water activity) - nutrient content - antimicrobial constituents - Eh-Redox potential (oxidation-reduction potential) - biological structures. 3. Processing and storage conditions: - heating - acidification - reduced water activity - use of preservatives - chilled storage - storage atmosphere - combination of (above) treatments In general terms, organisms tolerant of/resistant to the preservation mechanisms are those that will cause subsequent spoilage i.e. PRESERVATION MECHANISM ORGANISMS OF SIGNIFICANCE (TOLERANT) Heat processing Thermodurics : clostridia, bacilli, spores Pasteurisation Thermodurics: streptococci, lactobacilli, spores Chilled Storage Psychrotrophs: pseudomonads, Enterobacteriaceae, lactic (c. 5C) acid bacteria, some micrococci, flavobacteria Most yeasts incl. Candida, Rhodotolura, Torulopsis Most moulds Frozen Storage None grow, but Gram-positives and spores survive (-18C) Dried Foods Osmophilic yeasts and moulds, e.g. Saccharomyces rouxii, (<25% water, Aw 0.6) Aspergillus spp. Intermediate Moisture Foods Osmophilic yeasts and moulds (<50% water, Aw <0.85) Staphylococci Salt Osmophiles, micrococci, staphylococci Sugar Osmophiles Vacuum packing Anaerobes, microaerophiles e.g. clostridia, Enterobacteriaceae, lactobacilli, streptococci, some bacilli Some yeasts Carbon dioxide Lactobacilli, Brocothrix thermosphacta Preservatives - benzoate - sorbate - nitrites/nitrates Yeasts, e.g. Zygosaccharomyces bailii Some Escherichia coli

Lactobacillus acidophilus

- sulphur dioxide - chlorine - alcohol Irradiation Acid

Zygosaccharomyces spp. e.g. bysporus Pseudomonas fragi Saccharomyces cerevisiae Sporeformers, Micrococcus radiodurans, streptococci Moulds (Candida)
Most yeasts and moulds Lactic acid bacteria

MICROBIAL SPOILAGE HOW DOES IT MANIFEST ITSELF? 1. Visible growth e.g. moulds, often produce large, pigmented colonies (visible bacterial and yeast colonies are less common) 2. Gas production 3. Slime 4. Diffusable pigment and enzymes which may produce softening and rotting (proteolysis) 5. Off-odours 6. Off-flavours There are a wide range of metabolites produced during microbial spoilage, including: Alcohols Sulphur compounds Ketones Hydrocarbons Fluorescent pigments Organic acids Esters Carbonyls Diamines

Flavour, aroma and textural changes on spoilage: Flavour/Aroma Food Problem Meat, eggs, fish Nitrogenous (e.g. bad eggs) Chemical Cause Trimethylamines, ammonia, H2S Organisms Involved

Souring

Dairy, vacuum packed meats, beer, wine

Acids: acetic, citric, butryic

Pseudomonads Acinetobacters Moxarellas Clostridia lactic, Lactic acid bacteria

Brochothrix thermosphacta Bacillus sp.

Butyric acid bacteria Acetobacter spp. Yeasts

Alcoholic

Mustiness Pig-sty Garlic

Fruit juices Mayonnaise-dressed salads Bread, Cake Vegetables Various

Ethanol

Fruity Potato-like

Meat Meat, eggs, milk

Chloroanisoles P-cresole, indole, skatole Bis (methylthio)methane Trimethylarsine Esters of short chain fatty acids 2-methoxy-3-isopropyl Pyrazine Chemical/Biochemical Cause Polysaccharide production Polysaccharides (gas production) Pectinases Cellulase Xylanase Acid production

Clostridia Unknown

Erwinia sp.

Moulds

Pseudomonas fragi
Pseudomonads

Texture Problem Slime

Food e.g. meats confectionery Bread, milk Milk Hard cheese Fruit and vegetables

Organisms Involved

Ropiness Bittiness Holes Softening/rotting

Pseudomonas fragi Leuconostoc mesentroides Bacillus subtilis Alcaligenes sp. Bacillus cereus
Coliforms Erwinia, Clostridia, Yeasts, Moulds Lactic acid bacteria

Curdling

Milk Meat

Visual Spoilage: Problem Gas formation Holes Bubbles Fisheyes Bloaters Surface growth Moulds Surface Discolouration/growth Cloudiness Discolouration Pigments - flourescent - pink Red spot Browning Blackening Greening Food Cheese Cottage cheese, coleslaw, etc Olives Cucumbers Many Potato salad, etc Beetroot Beverages/brines Organism Coliforms Yeasts, lactic acid bacteria

Fungi

Pichia membranaefacien Bacillus spp.

Yeasts

Meats/egg Sauerkraut Cheese Brined vegetable Dairy products Meats

Pseudomonas sp. Rhodotorula sp. Lactobacillus plantarum Lactobacillus brevis Pseudomonas nigrifaciens Lactobacillus viridescens

FOODS AFFECTED BY MICROBIAL SPOILAGE Sources of spoilage microorganisms include: - Soil and water - Plants and plant products - Air and dust - Animal hides - Gastro intestinal tract of animals and humans - Food handlers - Food utensils and processing equipment Generally, spoilage is most rapid in proteinaceous chilled fresh foods (e.g red meats, poultry, fish and dairy products). Due to: - Highly nutritious - High moisture content/Aw - Relatively neutral pH Other fresh foods that may suffer significant spoilage are fruit and vegetables - Fruit = spoilt by yeast and moulds, as the low pH prevents bacterial growth - Vegetables = subject to both bacterial and fungal spoilage Suitably processed (and packaged) or ambient stable foods with a low moisture content will be expected to show little sign of microbial spoilage. As regards the latter, will be limited to yeasts or moulds.

SPOILAGE MICROORGANISMS Gram -ve Rod Shaped Bacteria The most common spoilage microorganisms of fresh chilled produce Min. growth temperatures are often 0-3C and they grow rapidly at 5-10C Pseudomonas is the most common. - They are widely distributed in the environment and may contaminate from many sources - Often may proliferate on inadequately cleaned surfaces - Able to utilise a wide range of materials as substrates for growth - May multiple relatively rapidly at chill temperatures and hence may come to dominate the microflora of foods stored at low temperatures - Spoilage caused by: Production of lipase or protease enzymes that result in food rot, putrid odours (sulphurous and nitrogenous compounds), off flavours and rancidity Production of extracellular slime Development of visible, often pigmented growth (in thermally processed products with extended shelf life) extremely heat resistant enzymes may cause long-term defects Other Gramve rod-shaped bacteria may also grow at chill temperatures and have been shown to contribute to the spoilage of chilled red meat, cured meats, poultry, fish, shellfish, milk and dairy products, e.g. Acinetobacter, Aeromonas, Alcaligenes, Alteromonas, Flavobacterium, Moraxella and Archromobacter. [Also Vibrio spp, which are halophilic (salt loving) and therefore may cause spoilage of sea fish and cured meats.] Overall, the group is not heat-resistant and can be readily removed by mild thermal treatments. Coliform/Enteric Bacteria Generally slower-growing at chill temperatures than Pseudomonas spp. but become more significant than Pseudomonas as the temperature rises above 5C (dominating between 815C) But many strains are psychrotrophic have been isolated from vacuum packed meats, poultry, cured meats, milk, dairy and egg products Widely disseminated in the environment, including animals. Associated with animals, slaughter and dressing Typical spoilage species are Citrobacter, Enterobacter, Escherichia, Klebsiella, Proteus, Serratia, Hafnia and Erwinia. Spoilage characterised by - the production of gas - acid - slime - rope - bitter flavours - pigmented colonies - unclean/medicinal/faecal odours Coliforms are very susceptible to heat. Their presence in heat processed foods is indicative of inadequate processing or post-process contamination. Gram-Positive Sporeforming Bacteria Of particular significance due to their ability to produce heat-resistant spores which can survive many heating/pasteurisation processes and germinate under suitable conditions and grow in foods. They are therefore seen in the spoilage of canned foods.

In general, growth tends to be slower than that of the Gram negative. Minimum growth temperatures are often 0-5C, although growth is often slow below 8C. Genera of importance are Bacillus and Clostridium species. Common in the environment; spores may survive for considerable periods. Common spoilage is production of large quantities of gas which may result in pack or product blowing. Bacillus spp. largely aerobic, some psychrotrophic strains (e.g. 5C or less) in milk; produce enzymes which result in sweet curdling and bitty cream B. subtilis ropy spoilage of bread Some Bacillus spp. are acid-tolerant and spoil pickles (blackening by Bacillus nigrificans and softening by pectolytic strains) Alicyclobacillus (acid tolerant sporeformer) has been identified as causing spoilage of aseptically packaged fruit juice (particularly orange) with a disinfectant or antiseptic taint. Is a thermophile (growth only seems to occur over 25C) spores survive pasteurisation and later germinate and gown under suitable conditions.

Lactic Acid Bacteria Spoil foods by the fermentation of sugars to form lactic acid. Results in drop in pH and offflavours (souring) Tend to grow slowly at chill temperatures and are easily out competed by other microorganisms Generally present in low numbers but in warm countries or where temperatures are poorly controlled can cause problems. Also known to grow under conditions of vacuum and modified atmosphere. Constitute major spoilage microorganisms of vacuum packed meat and poultry (where Pseudomonas spp. are inhibited) May also spoil cured or fermented meat products where growth of other spoilage microorganisms is inhibited or prevented More acid tolerant than other spoilage bacteria (down to pH 3.6). Typical genera are Lactobacillus, Streptococcus, Leuconostoc (main cause of souring) and Pediococcus. NB Some lactic acid bacteria are used in food manufacture (cheese, yoghurts, fermented meats). May also produce antimicrobial compounds. Gram Positive Bacteria Brocothrix thermosphacta Gram +ve rod, may occasionally be present on fresh meats. Modified atmosphere packaging etc may allow Br. thermosphacta to dominate the microflora. - Has been isolated from frankfurters, fermented and fresh sausages and cured meats. - Spoils with low numbers (000s usually compared with 105+ normally with other spoilage microorganisms) - In some products (e.g. vacuum packed sliced meats) may generate objectionable and pungent cheesy odour Micrococcus spp. able to grow in the present of salt - Can spoil bacon, producing slime, souring or pigmented growth - Often predominate in freshly collected milk and as many strains are thermoduric can survive pasteurisation and so cause spoilage Yeasts and Moulds Not normally found on freshly collected food materials (except some plants) but are extremely common in the environment and can cause contamination through airborne transmission. Slow growers in comparison to bacteria and so are often out-competed

Seldom responsible for the spoilage of fresh proteinaceous material Yeasts and moulds more resistant to low temperature, low pH, lower Aw values and the presence of preservatives than bacteria. Most are not heat resistant (except mould Byssochlamys may produce relatively heatresistant ascospores) Chilled foods subject to spoilage include fruit and vegetables, unprotected meat carcasses, yoghurt, cheese, fermented meat products and salads If carcass surface dries out, this inhibits bacteria and permits mould growth (visible mycelial growth described as whiskers, black spot or white spot). Chill foods of low pH inhibit bacteria but may allow mould/yeasts to develop (yoghurt, fruit/mayonnaise-based salads, fermented meat products, fruit juices, etc). Species involved are detailed above. Fungal spoilage may be characterised by - Highly visible, often pigmented growth - Slime - Fermentation of sugars to form acid, gas or alcohol - Off odours/off flavours Yeasts and moulds may be added as part of food manufacture, e.g. Penicillium camembertii

SOME OF THE MORE IMPORTANT MOULDS RESPONSIBLE FOR POST-HARVEST DISEASES OF FRUITS Genus Spoilage Problems Brown to black spots on apples, stone fruits1 and figs; stem-end and black rot of citrus fruits [ 1Stone fruits include cherries, peaches, nectarines, apricots and
plums]

Alternaria

Aspergillus Botryodiplodia Botrytis Cladosporium Colletotrichum Diplodia Fusarium Geotrichum Gloeosporium Monilinia Mucor Nigrospora Penicillium Phomopsis Phytophthora Rhizopus Trichoderma

Black rot on peaches, nectarines, apricots, citrus fruits and figs Cushion or crown rot of bananas; ripe rot of pawpaws Grey mould rot of apples, pears, raspberries, strawberries, grapes, figs, blueberries, citrus and stone fruits Restricted rot with grey black core on stone fruits, olive-green growth on raspberries, black rot on grapes and spotting of figs. Brown to black spots (anthracnose) on citrus fruits, avocados, mangoes, pawpaws and papayas Stem-end rot of citrus fruit, avocados, mangoes and papayas; watery, tanbrown rot of peaches Brown rot of citrus fruit and pineapple; soft rot of figs Sour rot of citrus fruits and peaches Anthracnose, black rot and lesion rot of bananas; eye rot of pome fruits2
[2 Pome fruits include apples, pears and quince]

Brown rot of stone fruits Soft rot of strawberries, pears Soft, watery (squirter) rot of the pulp of bananas Blue and green mould rots of citrus fruits; blue mould rot of apples, grapes, pears, stone fruits and figs; brown rot of pineapples Stem-end rot of citrus fruits and avocados Brown rot of apples and citrus fruits, leathery rot of strawberries Watery, soft rot of apples, pears, stone fruits, grapes, strawberries, avocados and figs Cocoa-brown to green rot of citrus fruits

GENERA OF FUNGI CAUSING IMPORTANT SPOILAGE OF FRESH AND STORED VEGETABLES Examples of Commodities Most Affected Most vegetables especially carrot, lettuce, celery, cabbage Most vegetables. Especially carrot, lettuce, legumes, Brassica spp. Legumes, carrot, Brassica spp. Tomato, cucumber, asparagus, potato Tomato, potato, carrot Tomato, potato, beetroot Cucumber, legumes, potato Onion, Brassica spp. Tomato, Brassica spp Genus Type of Spoilage Grey mould rot

Botrytis

Sclerotinia

Watery soft rot

Rhizopus Fusarium Phytophthora Phoma Pythium Peronospora Alternaria

Soft rot Dry rots Brown rots (blight) Dry brown, black rots Cottony leak Downy mildews Firm, black rots

YEAST SPOILAGE OF FRESH FRUITS AND VEGETABLES Dates

Figs

Strawberries Tomatoes

Saccharomyces spp. Hanseniaspora valbvensis Candida quilliermandii Hanseniaspora uvarum (Klorkera apiculata) Hanseniaspora valbvensis Torulopsis stellata Saccharomyces cerevisiae Candida krusei Kloekera apiculata Hanseniaspora uvarum (K. apiculata) Pichia kluvveri Nematospora corvli Nematospora corvli

Legumes Coffee berries Citrus fruits Nuts Pineapple Rhubarb

Candida spp. Trichosporon cutaneum

YEAST SPOILAGE OF PROCESSED FRUITS AND VEGETABLES a. Salt-brined Vegetables Sauerkraut

Discolouration

Cucumbers

Bloaters

Olives

Surface films Softening

b. Acetic Acid Preserves Onions Gherkins Red beetroot Red cabbage Piccalilli Sauces c. Mayonnaise-based Salads Coleslaw Potato salad

Gas Pockets Gas formation Off odours Surface films

Saccharomyces cerevisiae Torulopsis holmii Candida krusei Brettanomyces spp. T. holmii Hansenula subpelliculosa Sacch. rosei Sacch. rouxii Debaryomyces spp. Rhodotorula glutinis Rh. minuta Rh. rubra Sacch. oleaginosus Sachh. kluvveri Hansenula anomala Zygosaccharomyces bailii Pichia membranaefaciens

Off odour Gas production Gas production

Mixed salads

vegetable

and

fru Gas production Off odour

Sacch. exiguus Sacch. dairensis Sacch. exiguus Sacch. dairensis Pichia membranaefaciens Saccharomyces spp.

BACTERIA THAT CAUSE SOFT ROT OF VEGETABLES AND FRUIT Temperatures for Growth (C) Bacterium Min 3 Opt 27 Max 35 Produce Affected Most vegetables, particularly potatoes, some fruit Most vegetables and some fruit Pineapple Many vegetables Beans Chicory, endive, cabbage, lettuce

Erwinia carotovora subsp. atroseptica Erwinia carotovora subsp. carotovora Erwinia chrysanthemi Pseudomonas marginalis Pseudomonas viridiflava Psedomonas cichorii

6 6 >0.2 -

28-30 34-37 25-30 c. 30

37-42 >45 >41 >41

Pseudomonas cepacia Pseudomonas gladioli pv. Allicola Bacillus polymyxa Bacillus subtilis Clostridium puniceum Low temperature clostridia

>4 >4 5-10 5-20 7 0-4

30-35 30-35 9-22

40-41 40-41 35-40 45-55 39 17-30

Onion Onion Potato, pepper Potato, tomato Potato Potato

MONITORING AND DETECTION OF MICROBIAL SPOILAGE Direct microbiological methods Chemical/physical/physiochemical methods Acceptability criteria include sensory determinations colour , texture, odour, flavour and general appearance.

MICROBIAL INDICATORS OF PRODUCT QUALITY Microbial product quality or shelf-life indicators are organisms and/or their metabolic products whose presence in given foods at certain levels may be used to assess existing quality or, better, to predict shelf-life. These may be: - the spoilage organisms themselves, or - their metabolic products Microbial Indicators that are Highly Correlated with Food Quality: Organisms Products Acetobacter Fresh cider Bacillus Bread dough Clostridium Hard cheeses Beers, wines Lactic acid bacteria Sugar (during refinery) Leuconostoc mesenteroides Fruit juice concentrates Yeasts Butter Pseudomonas putrefaciens Canned vegetables Flat-sour spores Canned fruits Byssochlamys spp. Mayonnaise, salad dressing Zygosaccharomyces bailii

Microbial Metabolites that Correlate with Food Quality: Metabolite Food Product Vacuum packaged beef Cadaverine and putresaine Frozen juice concentrate Diacetyl Canned tuna Histamine Canned vegetables Lactic acid Fish Trimethylamine (TMA) Seafish Total volatile bases (TVB) Seafish Total volatile nitrogen (TVN) Butter Volatile fatty acids

Use Of Total Viable Count (TVC) Total viable count is of greater value in assessing quality than predicting shelf-life. A Spoilage spectrum has been developed by Jay (1992) Range A (TVC 103-106 cfu/g) o Microbial spoilage is generally not recognised (with the possible exception of milk) Range B (TVC 106-107 cfu/g) o Some food products show incipiency o Vacuum packed meats often display objectionable odours and may be spoiled Range C (TVC 107-108 cfu/g) o Off odours are generally associated with aerobically stored meat and some vegetables Range D (TVC 5x107-108 cfu/g) o Almost all food products display obvious signs of spoilage o Slime will be common on aerobically stored meats Range E (TVC in excess of 5x108 cfu/g) o Definite structural change in product occurs

PREVENTION OF MICROBIAL SPOILAGE Use of quality raw materials Correct storage for the food type Allocation of appropriate shelf-life HACCP and other quality systems Hygiene of processing environment Use of predictive methods Training and education

Note: Above text derived from notes provided by Dr P Voysey, Campden & Chorleywood Food Research Association