Food Microbiology II- NFSC 278 Industrial Microbiology

1. Food Microbiology II- NFSC 278 Industrial Microbiology Potential Infective and Toxic Microbiological Hazards Associated with the Consumption of Fermented Foods

2. Fermented Foods and Opportunist Microbial Pathogens • A number of processes exist by which solid-state fermented foods may be rendered safe from opportunist microbial pathogens • The lactic acid bacteria play a dominant role in solid-state fermented foods • LAB produce a number of antimicrobial agents including metabolic products such as short chain organic acids, carbon dioxide, hydrogen peroxide, lactoperoxidase, diacetyl and other more specifically inhibitory substances such as antibiotics and bacteriocins

3. Fermented Foods and Opportunist Microbial Pathogens • It is the cumulative effect of the antimicrobial action which dictates whether competing microorganisms are inhibited or survive and proliferate in foods of concern • Apart from their antagonistic behavior towards unwanted spoilage and opportunist pathogenic organisms, LAB produce lactic acid which imparts the unique tangy flavor to fermented products, lowers the pH, denatures the protein and releases the moisture uniformly

4. Fermented Foods and Opportunist Microbial Pathogens • Low pH and Organic Acids • Acidity may be the primary factor in the preservation and sanitation of fermented foods • There are many bacteria which can grow in acidic conditions, but the majority are neutrophilic (growth pH range 5-9) • The homeostasis of internal pH is essential for neutrophils (7.5-8)

5. Fermented Foods and Opportunist Microbial Pathogens • During food fermentations, the acidity is great, this overcomes the homeostatic pH control mechanism of neutrophilic bacteria and growth ceases completely when the pH drops to 6.6-6.8 • The external acidity (pH) effect is magnified in the presence of organic acids which can cross the cell membrane

6. Fermented Foods and Opportunist Microbial Pathogens • The antimicrobial effects of organic acid production which occur during food fermentations is due: • A- To the depression of pH below the growth range of certain spoilage and pathogenic bacteria • B- To metabolic inhibition by the undissociated acid molecules

7. Fermented Foods and Opportunist Microbial Pathogens • For acetic acid the growth of most food-poisoning organisms and spore-forming bacteria is inhibited by 0.1% and mycotoxigenic molds by 0.3% of the undossociated acid • Apart from their inhibitory effects on food-poisoning bacteria, the LAB also have antifungal properties

8. Fermented Foods and Opportunist Microbial Pathogens • It has been reported that LAB can inhibit aflatoxin biosynthesis, though the extent to which the inhibition is due to lactic acid and/or other LAB metabolites is unclear • It has been suggested that the antimycotic and antimycotoxigenic potential of the LAB might eventually find commercial application as a biopreservative

9. Fermented Foods and Opportunist Microbial Pathogens • Bacteriocins • The role of bacteriocins produced during lactic fermentations in the inhibition of foodborne pathogens has been overlooked • Bacteriocins are defined as protein antibiotics • Many of these proteinaceous antimicrobial cmpds are inhibitory to food poisoning, spoilage and pathogenic bacteria

10. Fermented Foods and Opportunist Microbial Pathogens LAB are known to produce two classes of bacteriocins: a- those that are cidal to a narrow range of target organisms usually closely related to the producer strain B- those that inhibit a broad spectrum of Gram-positive bacteria

11. Fermented Foods and Opportunist Microbial Pathogens • Two major classes of killing activities are displayed by bacteriocins: • 1- Some form ion channels in the cytoplasmic membranes • 2- Others exhibit nuclease activity upon gaining entry to a sensitive cell

12. Fermented Foods and Opportunist Microbial Pathogens • The lantibiotic (e.g. nisin Z produced by Lactococcus lactis) and non-lanthionine-containing bacteriocins (e.g pediocin produced by Pediococcus acidilactis) produced by some LAB appear to affect the cell membrane permeability barrier by forming water-filled membrane channels or pores

13. Fermented Foods and Opportunist Microbial Pathogens • Other Factors • With some fermented foods/beverages, ethanol is a major product of the fermentation • Food spoilage and disease-producing microorganisms cannot develop in either an acidic or an alcoholic fermentation • When both are produced as in some fermented foods, the combination of acid and alcohol gives double protection

14. Fermented Foods and Opportunist Microbial Pathogens • Acid interferes with cell pH homeostasis, alcohol is considered to act by denaturing proteins and possible interaction with membrane lipid components • Ethanol produces a rapid release of intercellular constituents from exposed bacteria and disorganization of the membrane probably results from penetration of the solvents into the hydrocarbon interior of the cytoplasmic membrane

15. Fermented Foods and Opportunist Microbial Pathogens • Acid fermentation combined with salting remains one of the most practical methods of preserving and often enhancing the organoleptic and nutritional qualities of fresh vegetables, cereal gruels and milk-cereal mixtures • Salting increases the osmotic pressure and together with acidity and some chemical components is considered to be responsible for bacteriocidal properties of soy sauce

16. Acid Resistance of Some Foodborne Pathogens • An emerging pathogen which has caused concern with some fermented food products is E. coli 0157:H7 • Some outbreaks have involved acidic foods: apple cider, dry fermented sausage, yoghurt and mayonnaise • Acid tolerance may play a role in E.coli 0157:H7 pathogenesis by promoting survival during passage through the stomach

17. Acid Resistance of Some Foodborne Pathogens • It has been found that E.coli 0157:H7 growth occur at pH levels ranging from 4.0-9.0 • It is also known that the acid tolerance in E.coli 0157:H7 is not dependent upon adaptation but is induced by stationary phase or starvation • E.coli 0157:H7 poses a new threat to fermented foods should these become contaminated at any stage of preparation

18. Acid Resistance of Some Foodborne Pathogens • With other pathogenic bacteria, it has been established that exposure to certain conditions can result in a marked increase in acid tolerance • The so-called acid tolerance response (ATR) was shown to occur with Listeria monocytogenes which after 1 hour exposure to mild acid (pH 5.5) were then capable of withstanding severe acid stress (pH 3.5)

19. Acid Resistance of Some Foodborne Pathogens • Salmonella typhimurium has also been shown to exhibit an ATR • In S. typhimurium, the ATR has been characterized as a complex defense system that can minimize the lethal effects of extremely low pH (pH 3) • ATR can also defend against weak acids such as butyric, acetic and propionic acids

20. Factors increasing susceptibility of the consumers of fermented foods to microbial hazards • The study and control of established and new emerging pathogens in solid-state fermented foods must always be examined under the conditions which exist in the environment where these products are prepared, fermented and consumed

21. Factors increasing susceptibility of the consumers of fermented foods to microbial hazards • Inter-related factors are: • Preparation of foods under unhygienic conditions • Poor personal hygiene • Contaminated water supply • Presence of enteropathogenic in raw materials or fermented foods as contaminants • Type and virulence of enteropathogens present

22. Factors increasing susceptibility of the consumers of fermented foods to microbial hazards • Immune status of the individual being compromised or underdeveloped • Consumption of inadequately fermented foods (possible an indigenous fermentation process without the involvement of a LAB starter culture) • Worldwide in 1990 over 3 million children died as a result of diarrhea

23. Factors increasing susceptibility of the consumers of fermented foods to microbial hazards • Food borne disease can cause severe and/or long term damage to health, including acute, watery and bloody diarrhea, meningitis as well as chronic diseases affecting the renal, articular, cardiovascular, respiratory and immune system • The most serious implications of foodborne infections are their effects on nutritional status • A poor food intake, aggravated by loss of nutrients from vomiting, diarrhea, malabsorption and fever over an extended period leads to nutritional deficiencies

24. Factors increasing susceptibility of the consumers of fermented foods to microbial hazards • A child whose resistance is suppressed becomes vulnerable to other diseases and is subsequently caught in a vicious circle of infection and malnutrition • Annually, about 13 million children under the age of 5 yrs die in developing countries; in the majority of cases, these deaths are due to infections and associated malnutrition

25. Malnutrition and Food borne Infection Cycle Fermented Food Contamination Disease –Age, immunity status, incidence, severity, duration Weight Loss, Growth Faltering, Lowered Immunity, Mucosal Damage Appetite loss, Nutrient loss, Malabsorption, Altered metabolism Inadequate Dietary Intake

26. Factors increasing susceptibility of the consumers of fermented foods to microbial hazards • Overall, it is of paramount importance that fermented products be under good sanitary and hygiene conditions • Sources of food contamination are numerous: soil, polluted water, flies, pests, domestic animals, unclean utensils and pots, dirty hands, and a polluted environment caused by lack of sanitation, domestic animal droppings, dust and dirt, etc.

27. Role of Fungal Toxins (mycotoxins) in Fermented Foods • Mycotoxins are secondary metabolites of certain filamentous fungi that can cause toxicity to humans and animals when low concentrations are ingested or inhaled • In nature, such toxins are derived from agricultural crops like cereal grains and oil seeds and products derived from them and from animal-derived foods such as milk

28. Role of Fungal Toxins (mycotoxins) in Fermented Foods • Mycotoxins can enter the human dietary system by indirect or direct contamination • Indirect contamination can occur when an ingredient of a food or beverage fermentation (e.g cereals, legumes. etc) has been previously contaminated with toxin-producing molds • The mold may be killed or removed during processing, mycotoxins will often remain in the final product

29. Role of Fungal Toxins (mycotoxins) in Fermented Foods • Direct contamination can occur in 2 ways: • 1- the fermentation may involve a fungus essential for the process but also capable of producing mycotoxins • 2- the process or final product can become infected with toxigenic mold and subsequent toxin formation

30. Role of Fungal Toxins (mycotoxins) in Fermented Foods • All fermented foods and beverages have the potential to be affected by toxigenic molds at some stage during their production, processing, transport or storage • Consumption of fermented foods and beverages heavily contaminated with mycotoxins is not likely to occur in most advanced societies because of the existence of strict food regulations

31. Role of Fungal Toxins (mycotoxins) in Fermented Foods • High quality agricultural practices with improved storage and transportation facilities, have reduced toxigenic mold growth in raw agricultural materials destined for the human food chain • Where poor agricultural practices and warm, hot climates prevail, such as in many developing countries, higher endemic levels of mycotoxins must be anticipated in raw food materials like corn, rice and peanuts

32. Role of Fungal Toxins (mycotoxins) in Fermented Foods • In Asia, many molds including Aspergillus, Rhizopus, Mucor are widely used in the manufacture of fermented foods and beverages • Such products are usually derived from raw materials like soya beans, rice, wheat, groundnuts, etc. • It is widely accepted that there have been no examples of overt mycotoxicity related to the fermentations generated by the above molds

33. Role of Fungal Toxins (mycotoxins) in Fermented Foods • With the discovery of Aflatoxins in 1960s from Aspergillus flavus and other Aspergillus spp., the koji molds (A. oryzae and A. sojae) were studied for any mycotoxin production • Aflatoxin production was not recognized however, it is now known that A. oryzae and A. sojae can produce other mycotoxins when exposed to specific stressful environmental conditions

34. Role of Fungal Toxins (mycotoxins) in Fermented Foods • Microbial detoxification of aflatoxin and other mycotoxins is now well documented and is highly relevant to all fermented food and beverage production processes • LAB can affect mold growth and aflatoxin production

35. Role of Fungal Toxins (mycotoxins) in Fermented Foods • In Europe and North America, mold-ripened foods are primarily cheeses and meats ripened by starter cultures of Penicillium spp. • Since Penicillium spp are potentially toxigenic, it has been imperative to pursue selection processes to produce non-toxigenic starter cultures

36. Role of Fungal Toxins (mycotoxins) in Fermented Foods • In establishing suitable non-toxigenic but technically suitable starter cultures, the following criteria must be applied: • 1- Toxicological screening, with chemical and biological methods, of isolates obtained from high-quality products • 2- Sensory evaluation of foods inoculated with potentially suitable strains and ripened under the usual conditions

37. Role of Fungal Toxins (mycotoxins) in Fermented Foods • 3- Extraction of the finished product and investigation of the extracts for mycotoxins • At present, totally non-toxigenic strains of P. roqueforti and P. camemberti are not available • It should be noted that mycotoxins production by these Penicillium starter cultures is usually only obtained under laboratory stress conditions and if mycotoxins are found in the final cheese products, the levels are extremely low