The role of serological diagnosis of viral infections in the 21st century

Discuss the role of serology in virus diagnosis in the early 21st century

Serology remains the mainstay for the diagnosis of virus infections in a routine diagnostic laboratory, especially for the diagnosis of virus infections. However, it plays a much lesser or absent role. in other areas of virological testing, such as epidemiological research, monitoring the response to anti-viral therapy, and antiviral resistance testing. There are 3 basic categories of methods used in the initial diagnosis of virus infections: 1. Direct examination, 2. Indirect examination, and 3. Serology

Direct examination

In direct examination, the clinical specimen is examined directly for the presence of virus particles, virus antigens, viral nucleic acids and virus-induced histological changes. The main advantage of direct examination methods is the short length of time required for a result, often within the same day. With the advent of specific antiviral chemotherapy, this is becoming an increasingly important consideration. On the debit side, these tests are usually labor intensive and require experienced personnel. However, this is likely to change with the arrival to simple automated molecular biology techniques. Current commercial examples include the PCR-based Roche Amplicor system, Abbott LCR system, and the Chiron branched DNA system. Indeed the Roche Amplicor and the Chiron branched DNA system are now widely used for the determination of HIV viral load. The major drawback of these tests is that they are very expensive. However, when the costs come down, they should gain widespread acceptance in the virology laboratory. The importance of molecular techniques became evident with the SARS crisis in 2003 when PCR assays were rapidly developed and were the mainstay of the diagnosis. It took several weeks before antibody were detectable and serology was mainly used to confirm infection and for epidemiological research.

Indirect examination

The virus in the clinical specimen is amplified by growing in tissue culture, eggs or animals. Cell culture is by far the most commonly used method. The presence of growing virus may be revealed by changes such as cytopathic effect (CPE) or the ability to haemadsorb. The identity of the isolated virus can be further confirmed by various tests eg. virus neutralization, immunofluorescence, complement-fixation, electron microscopy etc. The obvious disadvantage of virus isolation is the length of time required for the CPE or the ability to haemadsorb to become apparent, which may take from a few days to a few weeks. Longer, if confirmatory tests such as virus neutralization are to be carried out. However, rapid culture methods such as the DEAFF tests for CMV are becoming available whereby the cell culture is examined for the presence of CMV early antigens by fluorescent antibody technique. Another problem with virus culture is that the sensitivity is often low and depends on a large extent the quality of the clinical specimen received. Also, virus isolation would not be applicable to viruses which are difficult or cannot be cultivated such as hepatitis B and parvovirus. Morover, there is considerable expense and expertise involved in setting up a cell culture facility, particular those that meet biosafety level II and level III standards.

Serology

Serology remains the bulk of the work carried out by a routine diagnostic virus laboratory. A large variety of serological tests are available eg. complement-fixation (CFT) , haeagglutination-inhibition (HAI), enzyme-linked immunoassay (EIA), radioimmunoassay (RIA), particle agglutination, immunofluorescence, single radial haemolysis, western blot etc. The sensitivity and specificity varies greatly between different techniques. Most techniques will detect all classes of antibody whereas some assays eg. RIA, EIA and IF can be made to detect one specific class only ie. IgM, IgG or IgA. The following virus infections are usually diagnosed by serology and are likely to remain so in the foreseeable future:-

Hepatitis Viruses - hepatitis A, B and C infections are usually diagnosed by serology as these viruses cannot be routinely cultured. Current serological tests including the test for HBsAg are well established and despite the availablity of molecular biological techniques for the detection of viral nucleic acid, serology is unlikely to be challenged as the main means of diagnosis.
HIV - HIV infection is normally diagnosed by serology The only instance when serology cannot be relied on is in diagnosing HIV infection in the newborn.
Rubella and parvovirus - rubella and parvovirus infections are usually diagnosed by serology. Rubella virus is difficult to isolate and parvovirus cannot be isolated by routine cell culture. The onset of clinical symptoms for these infections coincide with the appearance of antibodies and thus there is little need for other means of diagnosis.
EBV - although EBV serology is reliable, the heterophile antibody test is usually used for diagnosing cases of infectious mononucleosis

Some virus infections may be diagnosed by serology but it not the method of choice. This includes herpesviruses that reactivate from time to time to cause diseases, and respiratory and enteroviruses where the illness has passed by the time antibody is detectable. Direct and indirect detection methods are commonly used for these viruses and molecular biology techniques are likely to play an increasing role in the diagnosis of these viral infections.

HSV - although CFT and other serological tests are available for HSV, HSV infections are usually diagnosed by cell culture. Electron microscopy, immunofluorescence and PCR are available as rapid diagnostic methods. Serology is not that reliable in the case of HSV infections, in particular reactivations.
CMV - although serology is available for diagnosing CMV infections, it is not reliable as most cases of CMV infections are a result of reactivation/reinfection. Cell culture (including the DEAFF test) and rapid methods such as the CMV antigenaemia test and PCR are preferred means of diagnosis
VZV - serology can be used to diagnose acute infection and is also used for immunity screening. Cell culture is difficult but a rapid diagnosis may be reached in more severe cases by electron microscopy and immunofluorescence of the vesicle fluid.
Respiratory viruses - diagnosis of respiratory virus infections is more commonly made by cell culture or more rapidly by immunofluorescence of the clinical material. CFT and HAI techniques are usually used for serology and any diagnosis is going to be retrospective.
Enteroviruses - enterovirus infections are usually diagnosed by cell culture. Serology has a very limited role to play as available tests such as neutralization, are cumbersome to perform and in any case, the diagnosis would be retrospective. Serology is of some value in diagnosing cases of coxsackie B myopericarditis as it would be impractical to obtain biopsies of heart tissue.
Rabies - serology is used along with other direct detection methods in diagnosing rabies and it may be used to check for immunity after vaccination
Arboviruses - arbovirus infections may be diagnosed by serology or virus isolation. Arboviruses will not usually grow in routine cell cultures and may require mosquito cell lines or animal inoculation.

The following virus infections are not normally diagnosed by serology, and the role of molecular biology techniques is likely to increase in the near future;-

Diarrhoeal viruses - serology is of virtually no value in the diagnosis of diarrhoeal virus infections as the diagnosis is going to be retrospective. Diarrhoeal virus infections are normally diagnosed by electron microscopy and the detection of viral antigens by ELISA or particle agglutination
Papovavirus - serology is of virtually no value in diagnosing papovavirus infections.
Poxviruses - serology is of little value in diagnosing poxvirus infections.

Epidemiological research, monitoring the response to anti-viral therapy, and antiviral resistance testing are playing an increasingly greater role in the work of a virus laboratory, especially reference laboratories. These techniques depend mainly on molecular biology methods. Therefore, it is envisaged that molecular methods will become increasingly used in the 21^st century. However, serology is likely remain the mainstay of diagnosis for many viral infections and it will continue to play an important role in the 21^st century.