Påvisning af virus i vand og toskallede bløddyr

1.  Summary

The Danish routine microbiological quality assessment of water and shellfish detects the presence of coliform bacteria. Detection of human enteric viruses is not carried out as a routine procedure. The standard method of detection of infectious viruses involves cell cultures which is expensive and time-consuming. Rapid and reliable methods have to be developed for assessment of the health and environmental aspects of viruses in water and shellfish.

The development of molecular biological techniques, e.g. Polymerase Chain Reaction (PCR) for the detection of viral nucleic acid, has resulted in the fact that analysis results can be obtained within a very short time. PCR has a high sensitivity and specificity, and it can detect the Norwalk viruses which cannot grow in cell cultures as well as other viruses which are difficult to cultivate such as hepatitis A. The sensitivity and the specificity of PCR can be increased by nested PCR which is a two-step PCR reaction. The difficulties tied to PCR are mainly contributed to chemical components in the samples inhibiting the enzyme reactions and thereby reducing the sensitivity. There is still a need for developing new methods which efficiently remove PCR-inhibiting substances.

Viruses which generally are found in very small numbers in water must be concentrated in order to be detected. According to the American Public Health Association the filter method is the preferred method of concentration of virus in water: the virus is absorbed by and eluted from filter membranes. This method can concentrate viruses from water with high or low content of organic material. Subsequently the eluates are re-concentrated by organic flocculation or polyethylene glycol (PEG) precipitation.

The PCR technique detects viral nucleic acid and therefore not necessarily infectious viruses. The genome of the enteroviruses is protected by a capsid against the external environment, and detected nucleic acid is probably isolated from intact virus particles. In cases where it is desirable to detect infectious viruses the cell cultivation technique can be combined with PCR under the condition that the viruses in question are cultivable.The obtained results recently indicate that PCR has some substantial advantages, and that the method can be of importance to the future progress of techniques for the detection of human enteric viruses in water and shellfish.