GENERAL AND FOOD MICROBIOLOGY PART I - L. V. Krasnikova - 2016

14. CHARACTERISTICS OF FOOD-SPOILAGE BACTERIA

Objective: to study the morphological and PHYSIOLOGICAL CHARACTERISTICS OF Bacteria that most commonly multiply on Food Products and cause their spoilage.

14.1. Genus Micrococcus

Members of the genus Micrococcus are spherical Cells with a diameter of 0.5-2.0 µm, occurring singly or in irregular clusters. Some micrococcal species show a tendency to form tetrads. Micrococci are Gram-positive, non-motile, and do not form spores.

Obligate aerobes. Catalase-positive.

Many representatives of this genus produce pigments, resulting in colonies colored white, yellow, orange, pink, or golden.

Micrococcus luteus (Fig. 14.1). The cells are large, up to 1.5 µm in diameter, and occur singly. On solid media, it forms regular, round, convex colonies with a shiny yellow surface.

Class="center">Figure 14.1. Micrococcus luteus

Micrococcus roseus. Small cells with a diameter of about 0.6 µm. On solid media, it forms small, round, shiny colonies with smooth edges and a pink color.

The optimal growth Temperature for micrococci ranges from 25 to 30 °C. Certain species can withstand heating up to 63-65 °C for 30 minutes and short-term pasteurization at 72-75 °C.

Micrococci are frequently found in the air, from which they enter food products. Intensive multiplication of micrococci in milk leads to a bitter taste and coagulation at low acidity (40-60 °T), which is due to their ability to secrete a rennet-like enzyme. In sweetened condensed milk, along with a bitter taste, flakes appear or thickening occurs. When multiplying On the surface of food products, micrococci form pigmented spots of various colors.

14.2. Genus Enterococcus

According to modern Taxonomy, the genus Enterococcus belongs to the family Enterococcaceae, order Lactobacillales. Enterococci are streptococci of intestinal origin and are part of the normal intestinal microflora of humans and animals. Currently, enterococci, much like coliforms, are considered sanitary-indicator microorganisms.

Enterococcal cells are spherical or oval, ranging from 0.6 to 2.5 µm in diameter, and occur in pairs or chains (Fig. 14.2). They are Gram-positive, non-motile, and do not form spores or capsules.

Figure 14.2. Enterococcus faecalis

With respect to atmospheric oxygen, enterococci are facultative anaerobes. On The surface of solid nutrient media, they form small, convex, colorless colonies, while subsurface colonies resemble the shape of a little boat. Typical Representatives of the genus are Enterococcus faecalis and E. faecium.

Enterococci carry out fermentative METABOLISM, fermenting CARBOHYDRATES primarily to produce lactic acid. The optimal growth temperature for enterococci is 30-37 °C, with a growth temperature range from 10 to 45 °C. Enterococci are quite resistant to environmental factors, withstand short-term heating up to 75-80 °C, and can grow in media containing 6.5% sodium chloride and in alkaline media at pH 9.6. Enterococci hydrolyze Proteins with The formation of bitter Peptides, leading to bitterness in food products.

14.3. Genus Sarcina

The genus Sarcina is included in the family Peptococcaceae. The cells are nearly spherical, 1.5 — 3.0 µm in diameter, arranged in packets (cubes) of eight or more cocci because Cell Division occurs in three mutually perpendicular planes (Fig. 14.3).

Gram-positive, non-motile, and do not form endospores. An exception is the species Sporosarcina ureae, which possesses flagella and forms spores.

Their nutritional type is chemoorganoheterotrophic. The type species is Sarcina ventriculi.

Figure 14.3. Sarcina ventriculi

On the surface of solid media in Petri dishes, sarcinae form medium-sized, round, convex colonies with smooth edges, either smooth or rough/buberculate in texture, granular, and yellow, lemon-yellow, sometimes pink or white in color. They cause food pigmentation.

14.4. Genus Staphylococcus

Staphylococci are predominantly found on the Skin and mucous membranes of warm-blooded animals, but can frequently be isolated from air and food products. Certain species are capable of causing opportunistic infections in humans and animals.

Bacterial Cells of the genus Staphylococcus are spherical, with a diameter of 0.5 — 1.5 µm. They occur singly or in pairs, and also form characteristic irregular clusters resembling grape bunches, which results from cell division in three mutually perpendicular planes (Fig. 14.4).

Staphylococci are Gram-positive, non-motile, and do not form spores or capsules.

Figure 14.4. Staphylococcus aureus

Facultative anaerobes. They are chemoorganotrophs possessing both respiratory and fermentative Types of Metabolism.

On solid media, staphylococci form round, convex, glistening colonies of white, yellow, or golden color. The principal representatives of the genus are S. epidermidis, S. saprophyticus, and S. aureus (golden staphylococcus).

The optimal growth temperature for staphylococci is 30–37 °C. Representatives of this genus are resistant to elevated temperatures applied during thermal food Processing. The multiplication of Staphylococcus aureus in food products leads to the accumulation of exotoxin, which causes food poisoning.

14.5. Genus Escherichia

The genus Escherichia belongs to the family Enterobacteriaceae. Typical representatives of this genus are small, Gram-negative rods with rounded ends, measuring (0.5–0.7) × (2.0–3.0) µm. In smears, they are arranged randomly. They are motile in liquid media due to peritrichously arranged flagella. Most cells possess pili. They do not form spores, cysts, or capsules. The type species is Escherichia coli.

Escherichia coli — the colon bacillus (Fig. 14.5) — is a facultative anaerobe that grows well on simple nutrient media at an optimal temperature of 37 °C. Colonies on meat-peptone Agar are smooth, glistening, with smooth edges, slightly convex, and grayish.

Figure 14.5. Escherichia coli

The proliferation of coliform bacteria in food products leads to spoilage defects such as off-flavors and gassiness (bloating).

The presence of Escherichia coli in food products serves as an indicator of sanitary regulation violations during production.

14.6. Genus Proteus

The genus Proteus belongs to the family Enterobacteriaceae. Representatives of this genus are frequently found in the intestines of humans and animals, in manure, soil, and polluted waters. Proteus species are easily identified by their ability to swarm. Swarming results in the formation of a veil-like film (swarming growth) on the surface of solid nutrient media. The type species is Proteus vulgaris.

Proteus vulgaris (Proteus bacillus). Proteus cells are small, Gram-negative rods measuring (0.4–0.6) × (1.0–3.0) µm. They do not form endospores or capsules. They are highly motile peritrichs (Fig. 14.6).

Fig. 14.6. Proteus vulgaris

Facultative anaerobes. Chemoorganotrophs with respiratory and fermentative types of metabolism. The optimal growth temperature is 37 °C.

Proteus bacillus exhibits active Proteolytic Enzymes and causes putrefactive spoilage in meat and meat products, fish, and dairy products. During protein decomposition, it produces hydrogen sulfide, ammonia, and indole. The multiplication of Proteus species in food can lead to food poisoning.

14.7. Genus Pseudomonas

The genus Pseudomonas belongs to the family Pseudomonadaceae. Pseudomonads are widely distributed in nature and are commonly found in soil, Water, and food products. Pseudomonas cells are small, Gram-negative rods measuring (0.5–1.0) × (1.5–5.0) µm. They do not form spores or capsules and are motile by means of one or more polar flagella.

Strict aerobes. On solid media, they form circular or irregular, flat to convex, mucoid, and often pigmented colonies. Their metabolism is strictly respiratory, utilizing oxygen as the terminal electron acceptor. They are capable of growing within a temperature range of 4 to 43 °C.

Pseudomonas fluorescens (fluorescent bacterium) forms large, mucoid colonies on nutrient agar that are yellow or yellow-green in color and often fluorescent (Fig. 14.7).

Fig. 14.7. Pseudomonas fluorescens

Pseudomonas aeruginosa (pyocyac bacillus) secretes the blue pigment pyocyanin into the medium, giving its colonies a blue coloration. It is an opportunistic pathogen.

Most pseudomonads are psychrophiles capable of multiplying at low food storage temperatures (from 2 to 4 °C).

Pseudomonads possess active proteolytic and lipolytic enzymes that break down proteins and fats, leading to the rapid spoilage of meat, dairy, and fish products.

14.8. Genus Bacillus

The genus Bacillus belongs to the family Bacillaceae. Bacteria of this genus are Gram-positive rods of varying length and thickness, mostly motile, and capable of forming endospores.

Aerobes or facultative anaerobes. Chemoorganotrophs with a respiratory or fermentative type of metabolism.

Bacillus subtilis (hay bacillus) consists of slender, short, Gram-positive rods measuring (0.5–2.5) × (3.0–5.0) µm, occurring singly or in chains. The cells are motile via peritrichous flagella. Spores are oval, 0.6 to 0.9 µm in size, and centrally located within The Cell (Fig. 14.8, a).

On meat-peptone agar, B. subtilis forms dry, wrinkled, or velvety flat colonies with wavy edges, which are colorless or pinkish.

The spores of the hay bacillus are highly heat-resistant and can withstand brief sterilization. The optimum growth temperature is 25–37 °C. The hay bacillus is classified among putrefactive bacteria; it produces active proteases that break down milk and meat proteins, as well as amylases responsible for the "rope" spoilage (potato disease) of bread.

Bacillus megaterium (from Lat. terra — earth) consists of large, granular rods with rounded ends measuring (5–7) × (1.2–1.5) µm, arranged in long chains. Spores are elliptical and centrally located in the cell (Fig. 14.8, b).

Fig. 14.8. Genus Bacillus: a — Bacillus subtilis; b — Bacillus megaterium; c — Bacillus mycoides

On solid nutrient media, it forms large, smooth, matte or glossy, grayish-white colonies with wavy-fringed edges. It possesses active proteases and cleaves carbohydrates with acid production.

Bacillus mycoides (mycelial bacillus) comprises small rods measuring (5–7) × (0.8–1.2) µm, with vacuoles and reserve nutrient granules present in the Cytoplasm. Spores are oval in shape (Fig. 14.8, c).

On nutrient agar, it forms colonies resembling fungal growth: rhizoid or mycelium-like, spreading across the surface of the agar.

The mycelial bacillus possesses active proteases and degrades proteins with the formation of bitter peptides.

Bacillus cereus (from Lat. cera — wax) features short, thick cells measuring (3–5) × (1.0–1.5) µm, occurring singly or in chains. Spores are oval (1.5 × 0.9 µm) and located subterminally (shifted from the center toward the end of the cell).

Colonies on nutrient agar are waxy, thick, matte, with a folded center and rhizoid, wavy edges, occasionally finely bumpy with fringed borders from which delicate interlacing filaments extend.

B. cereus is a potentially hazardous microorganism, as foodstuffs contaminated with it can cause food poisoning.

14.9. Genus Clostridium

The genus Clostridium belongs to the family Clostridiaceae. These are rod-shaped, Gram-positive bacteria that, when young, are typically motile (peritrichous). They form endospores that usually exceed the diameter of the cell.

Strict anaerobes. Some clostridia are saccharolytic, breaking down sugars via Butyric acid and acetone-butanol Fermentation, while others are proteolytic, causing putrefactive processes. Certain clostridia can cause foodborne intoxications: botulism, caused by Clostridium botulinum, and foodborne toxi-infections, caused by Clostridium perfringens. Among clostridia, There are also species pathogenic to humans that cause gas gangrene (Clostridium septicum, C. perfringens, C. novyi) and tetanus (Clostridium tetani).

Clostridium butyricum represents butyric acid bacteria. These are large rods measuring (0.5-1.5) × (5-12) μm, Gram-positive, and motile until sporulation begins. Spores are located terminally or subterminally. Spore-bearing cells resemble a club, a tennis racket, or a spoon.

Butyric acid bacteria belong to the group of saccharolytic clostridia; they ferment carbohydrates with The production of butyric, acetic, and formic acids, as well as a large amount of gases, thereby causing food defects such as a rancid taste, container bulging (elater/swell), and cheese blowing (Swelling).

Clostridium perfringens is a large rod measuring (1-2) × (5-8) μm; unlike other clostridia, it is non-motile and forms a capsule in the Human and Animal body. Spores are located terminally or subterminally (Fig. 14.9).

Fig. 14.9. Clostridium perfringens

Chemoorganotrophs with a fermentative type of metabolism.

On solid media, the bacteria form small, round colonies with smooth or serrated edges. When grown on Blood agar, C. perfringens colonies are round, smooth, grayish, later turning green and surrounded by a zone of β-hemolysis.

Bacteria of this species synthesize A number of aggressive enzymes: lecithinase, collagenase, hyaluronidase, etc.

Clostridium perfringens belongs to saccharolytic clostridia. This species causes rapid milk coagulation with the formation of a spongy clot. Its spores are heat-resistant and capable of withstanding boiling for 2 hours. It exhibits sulfite-reducing ability. Certain serological variants of this species (A and C) produce endotoxins and are therefore capable of causing foodborne toxi-infections.

Clostridium putrificum is a large Gram-positive rod with oval spores located at the cell pole. The microbe belongs to the group of proteolytic bacteria, breaking down proteins with the production of hydrogen sulfide and ammonia. It causes the defect known as "putrid cheese".

Clostridium sporogenes is a motile rod 3–4 μm long and 0.9 μm in diameter with rounded ends. Large oval spores are located in the center of the cell. A characteristic feature is the rapid formation of spores (within the first day of growth). It ferments carbohydrates with the production of acid and gas. During protein degradation, abundant evolution of hydrogen sulfide is observed. The optimal growth temperature ranges from 35 to 40 °C, with a minimum of 8 °C. It is the CAUSATIVE AGENT OF spoilage in canned meat, fish, and vegetable products.

Clostridium botulinum consists of large rods measuring (3.4-8.6) × (1.0-1.3) μm, Gram-positive, motile until sporulation, and non-capsulated. Spores are located terminally. Spore-bearing cells resemble a tennis racket, a small boat, or a spoon (Fig. 14.10).

Fig. 14.10. Clostridium botulinum

C. botulinum produces two exotoxins: a neurotoxin and a hemolysin. The accumulation of the neurotoxin in food products leads to severe food poisoning, frequently resulting in death.

Review Questions

1. What types of food spoilage are caused by bacteria possessing high proteolytic activity?

2. Which of the described bacteria cause food poisoning?

3. What is the oxygen requirement (attitude to atmospheric oxygen) of bacilli and clostridia?

4. List the genera of bacteria that form and do not form endospores.

5. Which of the specified genera belong to the family Enterobacteriaceae?