Module 2. Classification of dairy microorganisms

Lesson 9

CHARACTERISTICS OF SPOILAGE AND PATHOGENIC MICROORGANISMS – III

9.1 Introduction

As discussed in earlier lesson different microorganisms possess different characteristics such as pigments, uptake and utilization of different sugars, selective medium for their growth and different colony pattern on media. All microorganisms have different biochemical and molecular characteristics which distinguish them from each other. Here a number of characteristics of spoilage and pathogenic microorganisms are discussed and presented.

9.2 Bacillus

Bacillus is a genus of Gram-positive and rod-shaped bacteria of the division Firmicutes. These can be obligate aerobes or facultative anaerobes, and test positive for catalase. These are ubiquitous in nature, Bacillus includes both free-living and pathogens. Under stressful conditions, the cells produce oval endospores that can stay dormant longer.

The ‘Bacillus’ means a little stick. These are Gram-positive large rods (ranging from 3 to 9 um in length), aerobic, saprophytic soil bacteria, and these may occur either singly or in pairs or chains. These are aerobic spore formers. The main sources of Bacillus are air, water, soil, feed, fodder etc. These can be isolated by heating raw milk to 80°C/ 10 minutes prior to plating. On incubation the large and rough colonies are formed. These are acid sensitive, potent spoilage microorganisms. These may be psychrotrophic, mesophilic, and thermophilic and are catalase positive. The Bacillus spores are ellipsoidal to cylindrical in shape.

Bacillus cereus is a Gram-positive, rod-shaped and β-hemolytic bacterium. Some strains are harmful and cause foodborne illness, while others can be beneficial. It causes ‘Fried Rice Syndrome’. B. cereus is an aerobe, and like other members, it can produce protective endospores. Its virulence factors include cereolysin and phospholipase C. It cannot grow at 5°C but grows well in the range of 20-35°C. It causes food poisoning by producing enterotoxins (if number exceeds 1x10⁶/g). This produces ‘bitty cream’ defect that means broken cream by the action of lecithinase that is an extra-cellular phospholipase. It produces ‘sweet curdling’ by coagulating milk at lower acidity by producing rennet like enzyme.

Bacillus subtilis, known also as hay or grass bacillus, is a Gram-positive, catalase-positive bacterium commonly found in soil. It is a member of the genus Bacillus, B. subtilis is rod-shaped, and has the ability to form a tough, protective endospore, allowing the micro-organism to tolerate extreme environments. Unlike several other well-known species, B. subtilis has been classified as an aerobe. It grows well in the range of 20-45°C. It causes ropiness or sliminess in raw or pasteurized milk. It causes spoilage of UHT milk and concentrated or canned milk products. It causes ‘sweet curdling’ by coagulating milk at lower acidity. It produces levan extracellular from sucrose. It degrades pectin, casein, polysaccharides of plant tissue and also produces polypeptide antibiotic ‘subtilin’.

Bacillus licheniformis is commonly found in soil and bird feathers. It is a Gram-positive, thermophilic bacterium. Its optimal growth temperature is around 30°C, though it can survive at much higher temperatures. The optimal temperature for enzyme secretion is 37°C. It can exist in spore form to resist harsh environment, or in a vegetative state when conditions are good. This is an important micro-organism because it:

· Causes spoilage of UHT milk and concentrated or canned milk products similar to B. subtilis

· Produces levan extracellular from sucrose similar to B. subtilis

· Causes ropiness or sliminess in raw, pasteurized milk

Bacillus coagulans is a lactic acid-forming species within the genus it was first isolated and described in 1932 and initially considered to be a spore-forming Lactobacillus. Since, Bacillus coagulans exhibits characteristics of both genera Lactobacillus and Bacillus, its taxonomic position between the families Lactobacillaceae and Bacillaceae is often debated, where it was finally placed in the genus Bacillus. DNA-based technology was used in distinguishing between the two genera that are morphologically similar and possess similar physiological and biochemical characteristics.  B. coagulans is a Gram-positive rod (0.9μm by 3.0μm to 5.0μm in size); catalase positive, spore-forming, motile, a facultative anaerobe. B. coagulans may appear Gram-negative when entering the stationary phase of growth. The optimum temperature for growth is 50°C (the range of temperature tolerated is 30°C - 55°C). It mainly causes spoilage of UHT milk and concentrated or canned milk products.

Bacillus stearothermophilus is a rod-shaped, Gram-positive bacterium. These are thermophillic and are widely distributed in soil, hot springs, and are a cause of spoilage in food products. It can grow within a temperature range of 30-75°C. It is commonly used as a challenge micro-organism for sterilization validation. The biological indicator contains spores of the micro-organism on filter paper inside a vial. After sterilizing, the cap is closed, an ampoule of growth medium inside of the vial is crushed and the whole vial is incubated. A colour and/or turbidity change indicates the results of the sterilization process; no change indicates that the sterilization conditions have been achieved; otherwise the growth of the spores indicates that the sterilization process has not been met.

It was first described in 1920 as Bacillus stearothermophilus, but following a reclassification it is officially a member of the genus Geobacillus. It is an obligate thermophilic, cause flat sour spoilage of canned and UHT milk products and used in the detection of antibiotics in milk.

9.3 Clostridium

Clostridium is a genus of Gram-positive bacteria. These are obligate anaerobes capable of producing endospores. Individual cells are rod-shaped, that gives them their name, from the Greek kloster or spindle. These characteristics traditionally defined the genus; however, many species originally classified as Clostridium have been reclassified in other genera. Clostridium consists of around 100 species that include common free-living bacteria as well as important pathogens.

C. botulinum, an organism that produces botulinum toxin in food/wound and can cause botulism. Honey sometimes contains spores of Clostridium botulinum, which may cause infant botulism in humans one year old and younger. The toxin eventually paralyzes the infant's breathing muscles. Adults and older children can eat honey safely, because Clostridium does not compete well with the other rapidly growing bacteria present in the gastrointestinal tract. This same toxin is known as ‘Botox’ and is used cosmetically to paralyze facial muscles to reduce the signs of aging; it also has numerous therapeutic uses.

C. difficile, which can flourish when other bacteria in the gut are killed during antibiotic therapy, leading to pseudomembranous colitis (a cause of antibiotic-associated diarrhea).

C. perfringens, formerly called C. welchii, causes a wide range of symptoms, from food poisoning to gas gangrene. It is also responsible for enterotoxemia (also called ‘overeating’ or ‘pulpy kidney’ disease) in sheep and goats. C. perfringens also takes the place of yeast in the making of salt rising bread. The name perfringens means 'breaking through' or 'breaking in pieces'.

C. tetani is the causative organism of tetanus. The name derives from ‘of a tension’, referring to the tension (caused by tetanus) in the muscles.