M. du Preez

A sensitive seminested PCR method for the detection of Shigella in spiked environmental water samples

A rapid seminested polymerase chain reaction (PCR) method for the specific, sensitive detection of virulent Shigella spp. in spiked environmental water samples was developed. A set of primers specific for the invasion plasmid antigen gene (ipaH) of virulent Shigella spp. and enteroinvasive Escherichia coli produced a 620-bp fragment that was used as template for the seminested primer pair delineating a 401-bp fragment. By using agarose gel electrophoresis for detection of the seminested PCR-amplified products, a detection limit of 1.6 x 10(3) cfu S. flexneri was obtained with amplification reactions from crude bacterial lysates. The PCR procedure coupled with an enrichment culture incubated for 6 h detected as few as 1.6 S. flexneri organisms in pure culture. Treated sewage, ground, surface and drinking water samples collected from various sources were seeded with S. flexneri and incubated in GN broth for 6 h before detection by seminested PCR. A detection limit lower than 14 cfu/ml was achieved in some water samples. The results indicate that the described seminested PCR has the advantage of a rapid turnaround time and it fulfills the requirements of sensitivity and specificity for use in an environmental laboratory.

Detection of toxigenic Vibrio cholerae from environmental water samples by an enrichment broth cultivation-pit-stop semi-nested PCR procedure.

A pit‐stop semi‐nested PCR assay for the detection of toxigenic Vibrio cholerae in environmental water samples was developed and its performance evaluated. The PCR technique amplifies sequences within the cholera toxin operon specific for toxigenic V. cholerae. The PCR procedure coupled with an enrichment culture detected as few as four V. cholerae organisms in pure culture. Treated sewage, surface, ground and drinking water samples were seeded with V. cholerae and following enrichment, a detection limit of as few as 1 V. cholerae cfu ml−1 was obtained with amplification reactions from crude bacterial lysates. The proposed method, which includes a combination of enrichment, rapid sample preparation and a pit‐stop semi‐nested PCR, could be applicable in the rapid detection of toxigenic V. cholerae in environmental water samples.

Use of Handheld Computers for Collecting Water Quality Data in Developing Countries

Abstract A novel system is described for recording and storing details of water samples collected from rural African households and the water sources that they use. The system makes use of handheld computers for data entry instead of paper-based records. Bar codes are used to track water samples from the field to the laboratory, where total coliforms and Escherichia coli counts are enumerated for each sample. Participating household members and their water sources are documented using instamatic photography, enabling more rapid identification on subsequent visits. The system significantly reduces the lag-time between data collection and analysis. The use of bar codes and direct data entry reduces transcription errors that occur with paper-based recording.