Alkaline peptone water apw

A total of 1900 fecal specimens were collected by Shanghai hospital from patients who presented with acute diarrhea to gastroenteritis outpatient clinics during the period from January 2014 through to December 2015. These fecal samples were placed in sterile sealed plastic bags and stored at 4°C prior to further analysis. Confirmation of V. parahaemolyticus samples were performed using standard culture methods (ISO, 2007 ).
Briefly, 25 g of each fecal specimen was homogenized for 2 min in a stomacher 400 with 225 mL of alkaline peptone water (APW; Beijing Land Bridge Technology Company Ltd., Beijing, China) containing 3% NaCl, and incubated at 37°C for 16-18 h. After incubation, a loop from the top 1 cm was streaked onto thiosulfate-citrate-bile salts-sucrose (TCBS; Beijing Land Bridge Technology Company Ltd., Beijing, China) agar plates and incubated at 37°C for 18–24 h. Presumptive individual bacterial colony (green or blue green colony, 2–3 mm in diameter) were grown in 10 ml tryptic soy broth (TSB; Beijing Land Bridge Technology Company Ltd., Beijing, China) supplemented with 3.0% NaCl and incubated at 37°C for 18–24 h. After cultivation, the bacterial liquid and 50% glycerol in the proportion of 1:1 were placed in a glycerol tube and stored at –80°C for further analysis.

Three Chinese traditional vinegars, white vinegar, aromatic vinegar, and mature vinegar, were purchased from a local supermarket in Shanghai. The major components were listed in Table 1. Each group of five inoculated shrimps was immersed in 500 mL of each condiment for 5, 10, 15, 30, 60, 90 and 120 s, respectively. Inoculated samples treated with sterile 0.85% saline solution were used as control.
After vinegar treatment, shrimp samples were placed in a sterile 400 mL filter stomacher bag (Beijing Land Bridge Technology Company Ltd., Beijing, PR China) with 100 mL of sterile alkaline peptone water (APW, Beijing Land Bridge Technology Company Ltd., Beijing, PR China) with 3% NaCl (pH 8.0), and then homogenized (BagMixer400, Interscience, France) for 2 min. Subsequently, the homogenate was serially diluted with APW and plated onto thiosulfate–citrate–bile salts–sucrose (TCBS,) to enumerate V. parahaemolyticus O3:K6 after incubation at 37 °C for 18–24 h. Three replicates at each sampling time were performed.