© Borgis - Nowa Medycyna 2/2009, s. 149-150
*Kristian Schřnning
Respiratory Viruses – their detection and significance in respiratory infections
Department of Clinical Microbiology, Hvidovre University Hospital
Streszczenie
Acute respiratory disease causes the majority of morbidities in the community in temperate countries. The contribution of respiratory viruses of to the disease burden in the community as well as in the hospital setting is becoming well recognized (Tsolia et al, 2004; Jennings et al., 2007; Johnstone et al., 2008) as methods for their detection has become increasingly refined. Virus culture and especially the application of molecular techniques to clinical samples in the research setting has led to and the description of novel respiratory virus as possible agents causing human disease. Examples include human coronavirus NL63 (Fouchier et al., 2004) and HKU1 (Woo et al., 2005), human Bocavirus (Allander et al., 2005), polyomavirus Wu (Allander et al., 2007) and Ki (Gaynor et al., 2007). The application of molecular techniques to clinical samples has also led to the description of new syndromes caused by well known agents. For example Pneumocystis jirovecii, now generally accepted as a fungus, is associated with a "viral-like” self-resolving syndrome in immunocompetent infants (Larsen et al., 2007). To further complicate the matter co-infections are frequent and evidence suggest that viral and bacterial coinfection may cause an synergistic increase in virulence (Bakeletz, 1995; Jennings et al., 2008). The time interval between the description of a novel agent and the possible application of a diagnostic test in a clinical setting has considerably shortened with the advent of molecular tests. This advantageous development has in turn yielded less time for clinical evidence to accumulate and this in turn makes clinical interpretation of test results more difficult. In this review, I will discuss recent advances in molecular detection of respiratory viruses, how testing is done typically done in Denmark with special reference to my own laboratory and finally discuss the problems of clinical interpretation of new diagnostic tests of new agents using human bocavirus as an example.
Diagnostic methods
Virus isolation has been a golden standard due to its inherent high specificity, however, in Denmark it has never been established as a routine diagnostic test due to high turn-around time, high cost and labour intensiveness. Serological tests have been valuable in seroepidemiological surveys, but has had limited impact in routine clinical testing as paired sera are seldom obtained. Direct fluorescent antibody (DFA) tests have been largely abandoned and replaced by molecular methods. ELISA and immunochromatographic antigen demonstration of virus-Ag is still important as rapid tests and point of care-tests in Denmark.
PCR and other molecular methods (NATs) for detection of respiratory viruses are generally believed to be highly specific and sensitivity compares favourably with virus isolation and DFA (Kim et al., 2009; Nolte et al., 2007). In particular the detection of virus that are difficult to isolate by culture such as rhinovirus and coronavirus are considerably improved by NAT. At Hvidovre Hospital, Copenhagen we are currently opting for in-house real-time TaqMan-assays and are currently offering individual tests for RSV-A/B, Influenza A and B, Parainfluenza 1-3, human metapneumovirus, adenovirus, rhinovirus and bocavirus. Performing individual assays for an increasing number of possible pathogens is both labour demanding and costly in reagents. A solution to this problem is multiplexing; a multiplex assay analyzes for multiple pathogens in a single reaction. This may be technically demanding to accomplish without loosing sensitivity. Different commercial companies are now providing different solutions to diagnostic testing for respiratory virus. Abbotts xTag respiratory virus panel is a multiplex PCR assay detecting 20 different respiratory virus using a Luminex suspension microarray for detection. This assay has obtained FDA approval as an IVD and has been favourably compared with in-house NATs (Pabbaraju et al., 2008). Qiagens Resplex II is a similar highly multiplexed PCR assay (18 analytes) using the same detection system and has in an older version been compared to testing by individual NAT assays (Li et al., 2007). Other companies offer other solutions with using different detection strategies; e.g. Seeplex RV12 (Kimet al., 2009). The advantage of these highly multiplexed systems is that they allow detection of pathogens not suspected by the clinician and therefore increase the diagnostic yield compared to individual tests specified by the clinician. Companies will be competing for providing the most complete solutions with the most extensive test reports. This will in turn make clinical interpretation more complex and the need for clinical studies to aid this more pertinent. Clinical investigations of pediatric cases of human bocavirus infections may illustrate this point.
Human Bocavirus (HboV) – A commensal or a pathogen?
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Piśmiennictwo
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