A polymerase chain reaction (PCR) using primers specific to the internal region of the Salmonella gatD gene encoding galactitol-1-phosphate dehydrogenase was developed for a rapid, specific and sensitive detection of Salmonella spp. Four 20 mer oligonucleotides, named P1, P2, P3 and P4, were used in combinations of Pl-P3, P2-P4 and P3-P4 for amplification of the Salmonella specific region of the gatD gene. Single PCR products with about 780, 140 and 920 by were generated with all DNAs prepared from 24 Salmonella strains and P1-P3, P2-P4 and P3-P4 GatD primer sets, respectively. No PCR products were amplified from the DNA samples of 24 non-Salmonella bacteria using any of the GatD primer sets . These PCR bands were detected by ethidium bromide staining even when genomic DNA prepared from 180 cells of S. Typhimurium was used for the PCR. By using PCR with the primer sets following 6 h preenrichment in Enterobacteriaceae enrichment mannitol broth and 18 h selective enrichment in dulcitol-magnesium chloride-pyridinesulfonic acid-brilliant green-novobiocin medium, 1.7 cfu of S. Typhimurium per 25g of autoclave-sterilized chicken meat were detected even in the presence of E. coli at 2.1×106 cfu/25g.
This study was performed to increase the speed and sensitivity of the detection of E. coli O157: H7 in edible meats. We examined the efficiency and assay time of an improved enrichment culture procedure of the BAXTM screening system developed by Qua1icon Ltd. in the United States. The resu1ts are follows: 1) At 105 cfu/ml, the sensitivity of the PCR kit in detecting E. coli O157: H7 (VT2, VT 1, 2) in broth was less than that obtained by the culture method at 104 cfu/ml. On the other hand, the ability of the PCR kit to detect E. coli O157: H7 inoculated into edible beef, pork, chicken and calf/ox liver was greater than the culture method (1-40cfu/25g compared with 10-2, 100cfu/25g) except in the case of VT1, 2-producing strain in chicken. 2) An assay time of 40 hours was required with the culture method, but with the PCRkit, the assay time was only 25 hours. It is therefore possible to detect the presence of E. coli 0157: H7 in slightly over one day before a product is distributed. 3) Because the main reagents of the PCR kit were in tablet form, the whole PCR kit process was much simpler than that of the culture method. Consequently, the PCR kit is practical and useful. 4) Enrichment by shaking the culture shortened the time required for the PCR kit from 20 to 10 hours.
To apply the Bactometer for the rapid detection of Salmonella from foods, proper enrichment broths and incubation temperatures were examined in this study. The results obtained are shown as follows: 1. Detection of Salmonella was available within 10h by the conductance method with Impedance-Splitting Salmonell broth (ISS) 41°C incubation. 2. Growth of Citrobacter freundii, Escherichia coli, Klebsiella pneumoniae and Proteus mirabilis were inhibited in ISS-41°C incubation. 3. By using Modified Lysine decarboxylase broth (MLD) 41°, C Salmonella was detected, but E. coli was not inhibited.
Fresh meat is a highly perishable product due to its biological composition, which frequently leads to foodborne diseases. This study focuses on aerobic bacteria counts of fresh meat during storage periods, and helped to determine dates of minimum durability. Cultures of 963 samples of fresh meat (beef, pork and chicken) were purchased from a retail shop. The Samples were kept in a refrigerator at 0°C, 4°C and 10°C for 1 to 2 weeks under normal aerobic packing conditions, and the aerobic bacteria countS of coliforms, Salmonella and psychrophilic bacteria were examined. The detected aerobic bacteria were incubated at 30°C and 35°C for 48 hours using standard plate count agar plates. The isolation of aerobic bacteria from samples was better at 30°C than at 35°C, and there were significant differences between the efficiency of the two cultures. As bacteria counts increased, the manifestations of spoilage (slime, odor, color, and flavor changes) were noticeable on meat when bacteria numbers reached about 107cfu/g.
Environmental monitoring for microorganisms concerned with quantitative detection of the contamination on surfaces are not often carried out in the same way, because it is unknow how much to press against a surface with forceps . In this study, we evaluated the recovery of microorganisms with the swab method, using newly developed Torque Forceps with different levels of force. The recovery from a force of 150g/cm2 was relatively low. However, forces of 300g/cm2 and 450g/cm2 yielded almost the same results.