Diagnosis of HCV infections is not routinely done in most diagnostic virus laboratories.
This is mainly due to their fastidious growth requirements in cell cultures and to their inadequate biochemical and immunological characterisation, which makes serological diagnosis difficult.
The infections caused by HCVs have so far proved to be of minor clinical significance and therefore have not merited extensive study.
The routine diagnistic procedures used at present comprise:
1. Virus isolation in cell and/or organ cultures and
2. serological diagnosis on paired sera from infected individuals.
Direct detection of virus Antigen from nasopharyngeal aspirates and nasal or throat swabs has been shown to be successful. However, these procedures have not yet been used for routine diagnosis.
VIRUS ISOLATION IN CELL AND ORGAN CULTURES:
Virus isolation can be done from nasal and throat swabs, and nasopharyngeal aspirates taken from infected individuals.
TISSUE CULTURE ISOLATION:
229E and related strains can be isolated in roller culture monolayers of human Embryonic Lung Fibroblasts, such as WI-38 and MRC-5 cells.
In virus-positive cultures, a gradual cytopathic effect consisting of small, granular, round cells appear throughout the monolayers.
Isolates are generally confirmed as being 229E- related strains by standard serum neutralization tests.
ORGAN CULTURE ISOLATION:
OC43- related strains usually cannot be grown in cell cultures and hence isolation is performed in organ cultures of Human Embryonic Tissues.
Small pieces of tracheal or nasal epithelium with the ciliated surface uppermost are inoculated with the respiratory specimens and the culture examined for ciliary activity daily for upto two weeks.
The medium from these preparations is then harvested and examined after negative staining by direct electron microscopy or by immuno electron microscopy using convalescent-phase serum from the infected patient.
Immobilization or reduction in ciliary activity of infected organ cultures in comparison with controls, has also been used to identify HCV infection, but this is not a reliable measure of infection.
SEROLOGICAL DIAGNOSIS:
Identification of 229E and OC43 type infections can be performed using paired serum samples taken from acute and convalescent stages of infection.
The most frequently used test is the Neutralisation test.
Serum dilutions are incubated with aliquotes of virus. After incubation the mixture is inoculated onto cell culture tubes and the cultures are examined microscopically at three-day intervals for upto two weeks.
A four-fold or greater rise in neutralising antibody titre is diagnostic of a recent infection.
Other serological tests include Complement Fixation (CF) test and Haemagglutination Inhibition (HI) tests using chicken red blood cells for OC43 and related strains.
The antigens used for these tests are crude preparations of 229E (for CF) and OC43 (for CF and HI) and group-specific antibodies are detected.
The most sensitive method so far reported for detecting significant changes in HCV antibody in paired human sera is ELISA.
DIRECT DETECTION OF VIRUS:
Recent studies have shown that HCV particles can be detected in epithelial cells shed from the nasopharyx of individuals with Corona virus infections.
Indirect Immunofluorescence assay have shown that Corona virus fluorescence can be visualised in nasopharyngeal cells from volunteers inoculated with HCVs.
In another study ELISA has been used to detect HCV Antigens in nasal and throat swabs and nasopharyngeal aspirates taken from children suffering fron acute respiratory infections.
Nasal swabs were the best specimens for obtaining suitable quantities of antigen for the assay.
