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%.

A community-based study of acute respiratory tract infection in Thai children.

A 2-year longitudinal study was conducted among the population of a socioeconomically depressed urban community in Bangkok, Thailand, from January 1986 through December 1987 to determine the incidence, etiologic agents, and risk factors associated with acute respiratory tract infection (ARI) in children less than 5 years of age. Data were obtained for a total of 674 children, who were visited twice weekly for detection of signs and symptoms of ARI. During the first year of the study, throat-swab specimens were obtained for bacterial culture from both ill and healthy children and a nasal wash was performed on mildly ill children for detection of virus. During both years of the study, nasopharyngeal aspiration for identification of virus was performed for children with more severe infection. The overall incidence of ARI was 11.2 episodes per child-year. The highest (14.9) and lowest (8.8) rates per child-year occurred in age groups 6-11 months and 48-59 months, respectively. Respiratory syncytial virus, parainfluenza virus, adenovirus, Streptococcus pneumoniae, and Haemophilus influenzae were the prevalent pathogenic agents identified. Factors associated with higher risk of ARI were low family income, working mothers, mothers with allergies, chronic malnutrition, and crowding in the home.

Complement activity in children with protein-calorie malnutrition.

Using the hemolytic complement (CH50) assay, we evaluated the complement system of 28 children with severe protein calorie malnutrition (PCM) during their hospital admission and recovery. The mean CH50 activity in children with kwashiorkor was significantly less on hospital days 1 and 4 than in 17 healthy control subjects. On day 8 it rose to normal, and by day 50 it was significantly higher than the controls. The mean CH50 titer of 16 well-nourished febrile children was, in contrast to that of untreated PCM, significantly greater than the healthy controls. Of the children with PCM, 11 (40%) evidenced anticomplementary (AC) activity in their serum on either day or 1 or 4, but only two (7%) had detectable serum AC activity during later convalescence. Significantly, the CH50 titer in a PCM serum correlated inversely to the amount of AC activity in that serum (P less than 0.01). These results indicate that, in children with PCM, the complement system is compromised functionally, and that its repair coincides with intake of adequate diet. The presence of AC activity provides a possible mechanism for depressed complement activity in some untreated PCM children.