Bacterial causes of AIDS-associated diarrhea in Thailand.

The incidence of bacterial diarrhea in AIDS patients has increased steadily and has led to enormous medical and public health problems. In this study, the clinical data together with 350 rectal swab samples each from AIDS patients with diarrhea (APD) and non-AIDS patients with diarrhea (NAPD), were collected and examined for bacterial enteropathogens at the Bamrasnaradura Infectious Diseases Hospital (BIDH), Nonthaburi, Thailand from May to December 1996. Patients were matched by age and sex. The majority of these patients were male (79%, 554/700), aged between 15 and 34 years (70.9%). The study found that the isolation rates of bacterial enteropathogens causing diarrhea in APD (18%, 62/350) were considerably lower than those in NAPD (43%, 152/350) (p<0.05). The infection rate with Salmonella group B (19.7%, 12/61) in APD was found to be significantly higher than that in NAPD (14.3%, 2/14) (p<0.05). Vibrio parahaemolyticus (53.3%, 81/152), Plesiomonas shigelloides (27%, 41/152), Aeromonas spp (19.1%, 29/152) and V. cholerae O1 (15.1%, 23/152), were more frequently detected in NAPD than in APD (p<0.05). Only nine Escherichia coli strains were isolated from APD, of which six were enteroinvasive E. coli, two enterotoxigenic E. coli and one enterohemorrhagic E. coli (non O157) possessing both vtl and vt2. No V. cholerae strains were detected in APD. The least effective antibiotics were ampicillin, tetracycline and cotrimoxazole. Antibiotic resistant patterns of the isolated organisms were similar from both groups. The results from this study might be useful in Thailand in the diagnosis and management of clinical cases of bacterial diarrhea, especially APD.

Rapid detection of polioviruses in environmental water samples by one-step duplex RT-PCR.

This study describes the rapid detection of polioviruses in environmental waters by a simple reverse transcriptase-polymerase chain reaction (RT-PCR) using two primer pairs for differentiation of poliovirus from non-polio enteroviruses in a single reaction by a one-step method, combining RT and PCR in a single tube. The detection by agarose gel electrophoresis yielded 2 bands of 153-bp and 293-bp for poliovirus tested without the need for further hybridization. The detection sensitivity of this one-step duplex RT-PCR, as measured with RNA extracted by heat treatment from supernatant of infected cell extracts, was 10(-1) 50% tissue culture effective doses (TCID50). This assay was used to evaluate the ability of sample concentration by membrane filter-based adsorption and elution, and purification by a simple RNA isolation based on guanidine isothiocyanate-phenol-chloroform extraction; the system yielded a detection limit of 5 x 10(-1) TCID50 seeded in 5 liters of tap water. This protocol was applied to the poliovirus detection in environmental water collected from 2 communities in Bangkok, Thailand during February and May 1998. Of 100 samples tested, 2 water samples collected from the same open sewage pipeline at one location were positive for polioviruses and one sample collected from another sewage pipeline was positive for non-polio enterovirus while a further 97 water samples were negative for both polioviruses and non-polio enteroviruses. With poliovirus detection by cell culture technique, none of the 100 samples tested was positive for poliovirus type 1, 2 or 3. RT-PCR was more sensitive, rapid, simple and cost-effective than the cell culture technique since the two water samples which were positive for polioviruses by RT-PCR failed to be detected by cell culture. Sequence data of 293-bp amplicons from positive samples were compared with those of reference poliovirus strains in the Genbank and the EMBL databases and identity to the sequence of type 1 strain Sabin was found to be 99%.

Detection of poliovirus, hepatitis A virus and rotavirus from sewage and water samples.

A modified adsorption-elution technique for concentration of enteric viruses from sewage and water samples was developed. The viruses in water were concentrated by negatively charged membrane filtration, eluted with 2.9% tryptose phosphate broth containing 6% glycine pH 9.0, and reconcentrated using centrifugation by a speedVac concentrator. The presence of poliovirus, hepatitis A virus (HAV) RNA, and rotavirus antigen was determined by cell culture isolation, nested polymerase chain reaction (nested PCR), and enzyme-linked immunosorbent assay (ELISA), respectively. A total of 100 sewage and water samples were collected from various sources in congested communities in Bangkok, concentrated and examined for those enteric viruses. Of 20 surface water samples from canals which located near sewage drains, 15% were positive for HAV RNA by nested PCR. Of 48 domestic sewage samples from man-holes of underground sewers, 8% were positive for rotavirus antigen by ELISA. Even though the samples were concentrated 256-2,000 fold, poliovirus was not found by isolation in cell culture.