This method is suitable for the detection of small amounts of Staphylococcus aureus in foods and their raw materials considered to contain large amounts of competing bacteria and in untreated foods.
1. Sample preparation:
Solid or semi-solid food: 25g sample was weighed aseptically, placed in a sterilized homogenized cup containing 225mL of sterile physiological saline, homogenized at 8000r/min for 1-2 minutes, and made into 1:10 sample aliquot .
Liquid food: Pipette 25mL sample into a sterilized glass bottle containing 225mL sterile saline (pre-positioned glass beads in the bottle), and shake it to make 1:10 sample aliquot .
For the counting test, the sample homogenate can be appropriately diluted according to the decimal increment dilution method.
2. Select three consecutive dilutions, take 1 mL of diluted sample solution from each dilution, and inoculate 3 tubes containing 10% sodium chloride why? Remember? Staphylococcus aureus has a high salt tolerance and can be grown in 10-15% NaCl broth. Therefore, this is in fact a selective enrichment. Tryptone soy broth. The high dilution of the sample must be such that a negative endpoint can be obtained and cultured at 36 ± 1 °C for 48 h.
3. Using a 3 mm inoculating loop, remove 1 loop from each tube with bacterial growth, streak it onto a dry-surfaced Baird-Parker agar plate, and incubate at 36 ± 1 °C for 45-48 h.
4. Pick at least one suspected Staphylococcus aureus colony from each plate with bacterial growth, transfer it to the broth culture medium, and incubate at 36±1°C for 20-24 hours.
5. Take broth culture 0.3mL and 0.5mL coagulase test rabbit plasma in 8mm × 100mm test tube, mix well, set at 36 ± 1 °C culture, regularly observe whether there is clot formation, at least 6 h, to complete the contents Coagulation, the tube is not positive when the tube is inverted or tilted. Both positive and negative controls should be performed simultaneously in the test. Suspicious results should be confirmed by Gram staining, microscopy and other ancillary tests {such as thermonuclease test).
6. Report results: According to the results of the coagulase test, the zui approximation (MPN) table was used to report the MPN/g (mL) of S. aureus.
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