Mechanisms of Inactivation of Hepatitis A Virus by Chlorine

Abstract

The study was intended to investigate the feasibility of reverse transcription-PCR (RT-PCR) for evaluation of the efficacy of inactivation of viruses in water and to elucidate the mechanisms of inactivation of hepatitis A virus (HAV) by chlorine. Cell culture, enzyme-linked immunosorbent assay, and long-overlap RT-PCR were used to detect the infectivity, antigenicity, and entire genome of HAV inactivated or destroyed by chlorine. The cell culture results revealed the complete inactivation of infectivity after 30 min of exposure to 10 or 20 mg of chlorine per liter and the highest level of sensitivity in the 5 nontranslated regions (5NTR), inactivation of which took as much time as the inactivation of infectivity of HAV by chlorine. However, antigenicity was not completely destroyed under these conditions. Some fractions in the coding region were resistant to chlorine. To determine the specific region of the 5NTR lost, three segments of primers were redesigned to monitor the region from bp 1 to 1023 across the entire genome. It was shown that the sequence from bp 1 to 671 was the region most sensitive to chlorine. The results suggested that the inactivation of HAV by chlorine was due to the loss of the 5NTR. It is believed that PCR can be used to assess the efficacy of disinfection of HAV by chlorine as well as to research the mechanisms of inactivation of viruses by disinfectants.Hepatitis A virus (HAV) is a positive-strand RNA virus that belongs to the Hepatovirus group of the family Picornaviridae, which can bring about infectious hepatitis (hepatitis A). The infection rate of HAV is universally very high, being about 70% in China (18), and person-to-person spread of HAV is also very common. Disinfection with chlorine has been adopted worldwide to ensure the safety of drinking water, since HAV spreads mainly through drinking water and food. However, research on the mechanisms of chlorine inactivation of HAV has been inadequate. It is generally believed that chlorine inactivates HAV by damaging the nucleic acid of the virus (2,3,12). With the development of molecular biology methods, nucleic acid probes and PCR have been used to evaluate the effects of virus disinfection and to research disinfection mechanisms. However, the research problem most frequently encountered and difficult to solve is that, after disinfection, the virus nucleic acid can still be detected by PCR, although the infectivity of the virus actually no longer exists. Therefore, it is generally believed that molecular biology methods cannot be used to evaluate either the effects or the mechanisms of disinfection (2,8,9,16).The assumption that chlorine acts on the virus nucleic acid is likely to be measured by PCR. The main reason why no ideal result has ever been obtained in previous studies is that the length of the virus nucleic acid segment detected is limited and fails to reflect the overall status of viral nucleic acid. So in this study, in addition to using such techniques as cell culture and enzyme-linked immunosorbent assay (ELISA) to d...

Internalization of Sapovirus, a Surrogate for Norovirus, in Romaine Lettuce and the Effect of Lettuce Latex on Virus Infectivity

“…4), suggesting that the latex sap affected the viral capsids but not the genomes. Ag-ELISA has been used previously to show the negative effects of disinfectants on virus antigenicity (33,34). Using Ag-ELISA, we showed that the latex sap affected the antigenicity of SaV (Fig.…”

Application of PCR-Based Methods To Assess the Infectivity of Enteric Viruses in Environmental Samples

“…Two different RT-PCR approaches have been used for determining viral infectivity. One approach involves determining the presence of an intact genome or amplifiable undamaged genome by direct RT-PCR (48,51,74,75). The second uses coupling of the RT-PCR with a pre-PCR sample treatment that can determine the integrity of the viral capsid prior to extraction and purification of nucleic acid and subsequent enzymatic amplification (56,57).…”

Effect of Exposure to UV-C Irradiation and Monochloramine on Adenovirus Serotype 2 Early Protein Expression and DNA Replication

“…Monochloramine disinfection also decreased the numbers of infectious viruses, as measured by plaque assays. Chemical disinfectants such as monochloramine have been reported to damage both the viral genome (22,23,25,27) and capsid proteins (22,28,42). To assess which mechanism was more likely for monochloramine inactivation, monochloramine-disinfected virus samples were further analyzed for E1A protein synthesis and DNA replication, as a method to assess the ability of the disinfected viral capsid to enter the host cell and to deliver genetic material to the host cell nucleus and as a measure of DNA damage by monochloramine disinfection, respectively.…”