A COMPARISON OF 2 SETS OF PRIMERS FOR THE RT-PCR DETECTION OF ASTROVIRUSES IN ENVIRONMENTAL-SAMPLES

Human astroviruses (HAstV) are associated with sporadic cases and outb reaks of diarrhoe. The faecal-oral route is the predominant mode of tr ansmission and contaminated drinking water and shellfish have been imp licated as vehicles of transmission. Conventional diagnostic technique s have limited sensitivity and in this study two primer pairs, designa ted Jon and Mon, were compared for the detection of HAstV in environme ntal specimens by the reverse transcriptase-polymerase chain reaction (RT-PCR). Both primer pairs yielded positive RT-PCR products for the c ell culture adapted HAstV-1 positive control. The Jon primers, however , also yielded positive results for other viruses as well as for a num ber of water samples. These data suggest that the regions amplified by the Jon primers are not unique to HAstV. The Mon primer pair yielded positive RT-PCR results only for HAstV serotypes 1 to 4 and some envir onmental samples. The results obtained using the Mon primer pair could be confirmed by either hybridisation with an oligonucleotide probe sp ecific for HAstV or a HAstV specific enzyme immunoassay (EIA). RT-PCR, using the Mon primers, proved more sensitive than electron microscopy (EM), immune electron microscopy (IEM) and EIA for the direct detecti on of HAstV in river water. Cell culture amplification using the PLC/P RF/5 human primary liver carcinoma cell line improved the sensitivity of HAstV detection by EIA and RT-PCR, but not EM and IEM. The sensitiv ity of the RT-PCR assay system was enhanced by prior viral recovery by a glass wool adsorption-elution technique.