Introduction Staphylococcus aureus • In 1878, Koch observed staphylococci. • Staphylococcus recognized as a separate genus in 1880 by Pasteur. S. aureus Properties Introduction • 1884 - Rosenback grew • In the Greek language: staphylococci on a solid medium. � staphyle = a bunch of grapes � coccus = round • 1884 - Sternberg associated staphylococci with “ptomaine” formation in cheese that caused human illness. Introduction Introduction • 1894 - Denys associated illness with • 1914 – Barber related eating of meat from a cow sick with staphylococcal food poisoning to a pyogenic staphylococci. toxic substance produced in food. • 1907 - Owen recovered • He isolated staphylococci from staphylococci from dried beef that contaminated milk that came from had caused poisoning characteristic a sick cow with mastitis. of what now is called staphylococcal food poisoning. 1
Introduction Introduction • 1929 - Dack studied an outbreak of food • 1948–1974 studies demonstrated: poisoning caused from eating X-mas cake. � The presence of preformed • Re-discovered the role of staphylococci in enterotoxin in foods that had caused food poisoning. staphylococcal food poisoning. • He showed with human volunteers that the � Antitoxin in the blood of people that isolated staphylococci produced a toxic had suffered from this type of substance in culture, this substance caused poisoning. typical staphylococcal food poisoning. S. aureus in the US S. aureus (estimated) • In 1994, S. aureus was considered to be the cause of one of the most Agent Cases % Deaths % common bacterial food intoxications. S. aureus 185,060 1.3 2 0.1 Total bacterial 4,175,565 30.2 1,297 71.7 • Holt et al. (1994) estimated S. aureus food intoxication to be the second Total foodborne 13,814,924 100 1,809 100 most prevalent disease in the US. Illness & Causative Agent Contemporary Problems • S. aureus causes foodborne • Foods associated with intoxication. staphylococcal food poisoning: � In the US • The thermostable enterotoxins and not the bacterium are responsible – Meat products (e.g., ham) for the foodborne illness. – Desserts � In Japan – Rice balls • Staphylococcus / staphylococcal food poisoning. • Seasonal variations 2
Illness & Causative Agent S. aureus Properties • Only enterotoxin-producing • The pathogen produces the toxins staphylococci cause food poisoning. while growing in the food. • The ability to produce enterotoxin(s) • When the toxins are ingested by a is associated with production of susceptible person they will cause coagulase and heat resistant DNase. the illness. S. aureus Properties S. aureus Properties • It has a coccus shape. • S. aureus is ~0.5-1.5 µm in diameter • Occurs in clusters of irregular arrangement like • Gram positive, non-sporeforming, the bunch of grapes. non-motile, facultative anaerobe • May occur singly, in pairs, • Coagulase and catalase positive or in short chains. Coagulase Test • Suspect colonies are incubated in 2 ml of Brain Heart Infusion (BHI) broth for 18–24 hr at 35–37°C. • 0.5 ml coagulase plasma (with 0.5 ml of EDTA) is added to 0.5 ml of broth culture and mixed. • Tubes are incubated and examined after 4 hr. 3
S. aureus Properties • S. aureus produces a variety of extracellular enzymes and metabolites. • The most important metabolite produced is a group of heat-stable toxins called enterotoxins (staphylococcal enterotoxins). S. aureus Properties S. aureus Properties • Temperature range: • pH range: 4.0 – 9.8 (Optimal 6-7) 7–47.8°C (Opt. 35–37°C) • Salt tolerant (10 – 20% NaCl) • Enterotoxins produced between 10–46°C (Opt. 40–45°C) Environmental Effects S. aureus Properties • >10% NaCl inhibits SEA and SEB • Can grow at a sucrose concentration production. up to 50–60% • Enterotoxins are not formed: • Water activity as low as 0.86 under aerobic conditions, and 0.90 under – Below pH 5.3 at 30°C anaerobic conditions. – Below pH 5.6 at 10°C • Greater toxin production under • Minimal water activity -- 0.86 for aerobic conditions. growth 4
Water Activity Microbial Ecology • Enterotoxin production occurs at 0.86 – 0.99, Opt. 0.99 • S. aureus does not compete well with the normal flora of most foods. • Reducing a w minimizes production of enterotoxins: –0.90 a w reduces SEB by 90-99% (Maradona, 1998) S. aureus Toxins S. aureus Toxins • S. aureus is the common species • Staphylococcal enterotoxin A (SEA) associated with food intoxication. most common in gastroenteritis. • 12 enterotoxins: A, B, C, D, E, G, H, I, J, K, L, M. • Three variants of SEC – C 1 , C 2 , C 3 (minor antigenic differences) S. aureus Toxins S. aureus Enterotoxins • Enterotoxins are simple proteins. • Low molecular weight (~30 kDa) • Easily soluble in water and salt simple proteins solutions. • Heat resistant simple • Resistant to trypsin, chymotrypsin, • S. aureus itself is not heat resistant. and papain. • Enterotoxins A and D are the most • Pepsin destroys the toxin at pH 2. heat resistant. • Toxin is resistant to radiation (200 • When active, A and D exhibit kGy), and boiling (resists 121.1°C proteolytic enzyme resistance. for 0.5 hr) 5
Pathogenesis – target area Clinical Symptoms • S. aureus enterotoxins cause: • Enterotoxins expected to act on � severe gastroenteritis the receptors in the gut that � nausea, vomiting, retching, transmit impulse to medullary abdominal cramps, sweating, centers. chills, prostration, weak pulse, shock, shallow respiration, subnormal body temperatures. S. aureus Food Poisoning S. aureus Food Poisoning • About $ 10 6 cells/gram of S. aureus • Onset of illness takes <30 min – 8 hr. in food is needed for toxin following ingestion of the toxin production. containing food. • Most illness, however, occurs within 2–4 hr. • About 200 ng of toxin can cause illness in humans. • Recovery is within 24–48 hr. • Illness is rarely fatal. Clinical Symptoms S. aureus Infections • The enterotoxins acts on the • S. aureus is a feared receptors in the gut that transmit hospital pathogen. impulse to medullary centers. • Treatment of patients consists of bed • Sometimes it can be very rest and maintenance of body fluids virulent, and often and electrolytes. resistant to antibiotics. 6
How Is S. aureus Introduced to Food? How Is S. aureus Introduced to Food? • S. aureus is commonly found in: • Any food that requires handling and � Nose preparation is susceptible for � Throat contamination. � Hands � Fingertips • S. aureus is also found on the skin or � Hair and skin hides of animals. • Found in more than 50% of healthy • Cross-contamination may result people. from these animals during • Found on skins or hides of animals. • Found in the environment. slaughtering. Food Sources for Staphylococcal Foods Often Incriminated Outbreaks (1973-1987) • Meats and meat products • Poultry and Fish 120 Pork 96 • Cream-filled baked goods 100 Bakery Products Beef • Baked foods 80 Turkey Chicken • Potato Salad 60 Eggs • Salads containing any of the above items 40 26 22 20 • Any nutrient-rich, moist food that is 20 14 9 temperature abused. 0 (Bean et al., 1990) Contributing Factors Prevention • Adequate storage and refrigeration of foods • Improper storage and holding temperatures • Not preparing foods far in advance • Inadequate cooking/processing temperatures • Adequate cooking and/or heat processing • Contaminated Equipment • Avoiding poor personal hygiene • Unsafe food sources • Not holding foods between 40 – 140°F (4.4 – • Poor personal hygiene 60°C) for prolonged periods � 10 – 50% adults are reservoirs of S. aureus � 40-135°F (4.4-57°C); new numbers 7
Pathogen Detection Indicators for the Presence of S. aureus • Laboratory media: • Coagulase Test � Trypticase soy broth with 10% NaCl • Thermostable Nuclease Test (TNase) � Mannitol salt agar � Baird-Parker agar • Polymerase Chain Reaction (PCR) Polymerase Chain Reaction TNase Testing (PCR) • Culture is boiled for 15 min. • Thermostable DNA polymerase catalyzes the • Toluidine blue agar plates are prepared. gene probe amplification. • 2 mm wells are dug in the plates and filled with the boiled cultures • Amplified DNA is detected by hybridization • Plates incubated for 2 – 4 hr at 37 – 50°C ring using radio- and non-radiolabeled probes. • Pink halos around wells indicates positive reaction. • Can amplify a single DNA molecule to 10 7 molecules. (Maradona, 1998) (Maradona, 1998) Biological Detection Detection Methods of Enterotoxins • Each new toxin type had to be detected biologically • Biological • Biological subjects used are cats, kittens, and monkeys. • Kittens--emetic response • Immunological (many, • Can determine the enterotoxin activity by including kits) observing responses. • Monkeys used to simulate human response. (Maradona, 1998) 8
Recommend
More recommend
