Evaluation of the Roche LightCycler parvovirus B19 quantification kit for the diagnosis of parvovirus B19 infections.
BACKGROUND
The rapid and quantitative detection of viral DNA is important in the diagnosis of parvovirus B19 infection in immunocompromised patients and in congenital infection. It is also valuable for monitoring progress following therapeutic interventions.


OBJECTIVE
To evaluate the diagnostic sensitivity and specificity of the Roche LightCycler (LC) parvovirus B19 quantification kit in comparison with previously described nested PCR and dot blot hybridisation assays.


METHODS
Two hundred and twenty eight clinical samples and two standard B19 DNA sera were tested to assess the diagnositic performance of the Roche LC kit.


RESULTS
Ten clinical samples (4.3%) gave invalid LC results, including three of five bone marrow samples but only two of 165 serum samples. In the remaining 218 samples, the LC assay detected B19 DNA in 97.5% (79/81) samples that were positive by the nested PCR. The two samples (from the same patient) that were LC negative were sequenced in a 511-nucleotide region of the NS gene and 42 nucleotide changes were found. The Roche LC assay detected B19 DNA in 9.5% (13/137) samples that were negative by nested PCR. Analysis of the available clinical and serological data associated with these samples suggested that the LC results in the majority of these cases were true positive. In patients with resolving persistent infection, the LC assay remained positive for longer than nested PCR.


CONCLUSIONS
The Roche LC assay was more sensitive than the nested PCR used in this study. The additional sensitivity and the quantitative DNA measurements were valuable for monitoring patients with persistent B19 infection. Practical advantages of the LC assay include a short running time and the possibility to automate the assay. The LC assay provides a controlled and standardised method for quantitative detection of viral DNA for the diagnosis and monitoring of parvovirus B19 infections but failed to detect a variant strain.
Enteric, Respiratory and Neurological Virus Laboratory, Specialist and Reference Microbiology Division, Health Protection Agency, 61 Colindale Avenue, London NW9 5HT, UK.
Sharleen
S
Braham
Jayshree
J
Gandhi
Stuart
S
Beard
Bernard
B
Cohen
D000328:Adult; D000914:Antibodies, Viral / Q000097:blood; D001483:Base Sequence; D001853:Bone Marrow / Q000821:virology; D002648:Child; D002675:Child, Preschool; D004279:DNA, Viral / Q000032:analysis / Q000737:chemistry; D005189:False Positive Reactions; D005260:Female; D006801:Humans; D007223:Infant; D008297:Male; D008875:Middle Aged; D025202:Molecular Diagnostic Techniques / Q000379:methods* / Q000592:standards; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D010322:Parvoviridae Infections / Q000175:diagnosis* / Q000821:virology; D016732:Parvovirus B19, Human / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification*; D012680:Sensitivity and Specificity; D017422:Sequence Analysis, DNA; D044967:Serum / Q000821:virology; D017361:Viral Nonstructural Proteins / Q000235:genetics
D000914:Antibodies, Viral; D004279:DNA, Viral; D017361:Viral Nonstructural Proteins