Staphylococcus aureus Staphyl: grape; coccus: round-shape aureus: golden color Presence of Staphylococcus aureus and/or its enterotoxins in processed foods or on food processing equipment is generally an indication of poor sanitation. Human is the most common carrier.
金黃色葡萄球菌 (Staphylococcus aureus) G (+) 兼氣性 不產孢桿菌 毒素耐熱 A、B、C 、D 、 E、 F、G 、 H A 80℃ 30 min 100 ℃ 1 min B 210 ℃ 30 min
環境適應強 普遍存在人體上 傷口、 鼻腔 膿 食品條理人員 神經性毒 噁心、反覆嘔吐 (特徵)、腹痛、腹瀉 、 發燒 (少數)
Typical characteristics Catalase Coagulase NaCl tolerance
Materials and methods Food: Food samples (sandwich) Sterile cotton Bacteria: Staphylococcus aureus Human Medium Phosphate buffer saline (PBS) TSB with 10% NaCl Baird-Park agar 3% H2O2 Petri-film staphyl
Enumeration 25 g food sample (sandwich) is added into 225 mL of phosphate buffer saline. (x2) Stomach for 2 min. Inoculate S. aureus culture 1-6 groups, 1 mL of 10-4 overnight culture 7-11 groups, 1 mL of 10-5 overnight culture MPN calculation Incubate TSB-10% NaCl 24 hour at 37°C
1 mL TSB-10%NaCl PBS 1 mL 1 mL PBS 1 mL -3 -4 -1 -2
Barid-Parker agar Typical colonies of S. aureus are black, shiny, convex and surrounded by clear zones (egg yolk reaction) of approximately 2-5 mm. Pick up well-isolated colony for catalase test.
Biochemistry tests Catalase test: Place a drop of 3% H2O2 on a glass slide Positive reaction: gas generation S. aureus: positive
Human test Wet a cotton stick in a phosphate buffer saline. Use this cotton stick to swab both nose nasals. Streak the cotton stick onto Baird-Parker plates. Incubate the plates at 35-37℃ for 24 hour. One plate/group
Difco™ Baird-Parker Agar Base Approximate Formula* Per 950 mL Pancreatic Digest of Casein ...................................... 10.0 g Beef Extract ................................................................ 5.0 g Yeast Extract .............................................................. 1.0 g Glycine ..................................................................... 12.0 g Sodium Pyruvate ...................................................... 10.0 g Lithium Chloride ........................................................ 5.0 g Agar ......................................................................... 20.0 g Difco™ EY Tellurite Enrichment Egg yolk emulsion containing potassium tellurite consists of 30% egg yolk suspension with 0.15% potassium tellurite.
Baird-Parker Agar Base contains peptone, beef extract and yeast extract as sources of nutrition. Sodium pyruvate is incorporated to stimulate the growth of S. aureus without destroying the selectivity. Tellurite is toxic to egg yolk clearing strains other than S. aureus and imparts a black color to the colonies. Egg yolk additive, in addition to being an enrichment, aids in the identification process by demonstrating lecithinase activity (egg yolk reaction). Glycine and lithium chloride have inhibitory action for organisms other than S. aureus.
Typical colonies of S. aureus are black, shiny, convex and surrounded by clear zones (egg yolk reaction) of approximately 2-5 mm. Coagulase-negative staphylococci generally do not grow well; if some growth occurs, the typical clear zones are absent.
Pick up a well-isolated colony with typical characteristics with a loop Suspend this colony in a 9 mL PBS by shaking the loop Vortex the tube Take 0.1 mL from the tube, then transfer into another 9 mL PBS tube, vortex Drop one mL onto Petri-film Incubate at 35-37℃ for 24 hour
Typical S. aureus colonies on petrifilm
schedule 12/21 (Mon): prepare sample, TSB-10% NaCl incubation, human test 12/22 (Tue): streak onto Barid-Parker agar plates, (human) observe Barid-Parker plates and mark the typical colonies, store at refrigerator 12/23 (Wed) (food) Observe Barid-Parker plates and mark the typical colonies, store at refrigerator 12/28 (Mon): H2O2, petri-film 12/29 (Tue): observe results