Microbial quality of wastewater: detection of hepatitis A virus by reverse transcriptase-polymerase chain reaction.
OBJECTIVE
The persistent circulation of hepatitis A virus (HAV) in the Mediterranean area suggests the need for monitoring its presence in the environment. A reverse transcriptase-polymerase chain reaction (RT-PCR) was used to detect the presence of HAV in several consecutive raw sewage and final effluent samples, collected over an 8-month period from an activated sludge treatment plant in southern Italy.


RESULTS
Two distinct purification protocols, either based on antigen-capture with monoclonal antibody (AC) or RNA extraction, were compared. The possible influence of the antibody used in the AC phase was evaluated in preliminary experiments on HAV-spiked samples, using two different monoclonal antibodies. Hepatitis A virus RNA was detected in all but one sewage environmental sample examined. The contemporary presence of enteroviruses, reoviruses and phages was observed, while HAV growth in cell culture was hampered.


CONCLUSIONS
The RT-PCR technique was confirmed to be a valuable tool for the rapid monitoring of HAV in sewage samples. In addition, this study demonstrated that application of different sample purification methods can result in different levels of sensitivity of the assay and that, in the antigen-capture method, the choice of antibody can have a crucial role.


CONCLUSIONS
This work underlines the need for technical uniformity in environmental studies from different laboratories for a correct and useful comparison of the results.
Morace
G
G
Laboratorio di Virologia, Istituto Superiore di Sanità, Rome, Italy.
Aulicino
F A
FA
Angelozzi
C
C
Costanzo
L
L
Donadio
F
F
Rapicetta
M
M
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D000956:Antigens, Viral / Q000737:chemistry / Q000276:immunology / Q000302:isolation & purification; D016679:Genome, Viral; D030041:Hepatitis A virus / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification*; D006801:Humans; D008969:Molecular Sequence Data; D012367:RNA, Viral / Q000032:analysis / Q000302:isolation & purification; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012680:Sensitivity and Specificity; D017422:Sequence Analysis, DNA; D012722:Sewage / Q000382:microbiology*; D014764:Viral Proteins / Q000737:chemistry; D014871:Water Microbiology
D000911:Antibodies, Monoclonal; D000956:Antigens, Viral; D012367:RNA, Viral; D012722:Sewage; D014764:Viral Proteins