Varicella-zoster virus (VZV) viremia at different stages of infection was characterized. Different approaches were used, polymerase chain reaction (PCR), isothermal transcription based nucleic acid amplification (NASBA), and immunofluorescence to describe and quantitate viral infection of peripheral blood mononuclear cells (PBMC). In patients with acute varicella 200 to 5,000 copies of the viral genome in every 150,000 PBMC were found with quantitative competitive PCR (QCPCR). With NASBA, viral transcriptional activity was detected in these cells. RNA transcribed from the immediate early gene IE 63 as well as from the late gene 68 were found, indicating a productive infection. Glycoprotein gE specific immunofluorescence visualized by confocal laser scanning microscopy revealed that only 1 in 10,000 to 100,000 PBMC was infected. T and B lymphocytes as well as monocytes expressed viral protein on their surface. Similar results were obtained with PBMC from immunocompetent zoster patients. In some cases a transient viremia was found shortly after the onset of rash, although the viral load seemed to be lower than in patients with varicella. Examination of blood samples from 16 persons with postherpetic neuralgia (PHN) signs of viral replication in PBMC were not detected. In conclusion, the data suggest that VZV viremia is a frequent event in patients with varicella and zoster, but not in those with postherpetic neuralgia. Moreover, the results indicated that subclinical reactivations occur both in immunocompromised and immunocompetent individuals.

The occurrence of subclinical reactivation of varicella-zoster virus (VZV) in peripheral blood mononuclear cells (PBMC) from immunocompetent subjects >60 years old without any signs of VZV-caused illnesses, and from immunocompromised patients was investigated. Altogether, 223 samples were tested by nested ORF 63 PCR assay. In addition, all positive samples were tested by ORF 14, ORF 29 and ORF 63 PCR assays, as well as by ORF 63 and ORF 68 nucleic acid sequence-based amplification assays. In 5 samples, VZV-specific DNA, but no transcripts, could be detected. Three of them belonged to the group of >60-year-olds, 1 was HIV positive, the other was being treated with chemotherapy. The results confirm the observation of other authors that subclinical reactivation occurs in both immunocompromised and healthy individuals. The failure to detect DNA in samples taken from 2 individuals several weeks later excludes a long-lasting infection of VZV in PBMC.

Abstracts molecules/ml. Most of them were detected by the long probe but not by the short probe. When PCR for the Sor PreS-gene was included it was possible to detect all 24 HBV-positive sera (not shown) by ELOSA. The reliable lower quantification limit for the long probe is 250 molecules/ml and for the short probe 2500 molecules/ml. Surprisingly, chemiluminescence did not produce better qualitative or quantitative results. The data suggest that the usage of several replicates allows relative quantification ~ in most cases. One possible drawback we see is the hybridization efficiency. Six of our positive samples showed great differences between the number of target molecules suggested by agarose gel electrophoreses or by hybridization (Southern blot or ELOSA). All of them contained more than::,! 06 molecules/ml. For these cases and for the samples where the short probe and the long probe gave discordant result (2 cases) we think that competitive PCR will be the method of choice, but in most cases ELOSA with the long probe gives reliable results and is highly sensitive.sponding to expected target copy numbers (5000, 500 and 50 copies/reaction) was employed. The NASBA showed a high reproducibility; the intra-and inter-assay coefficients of variation were < 10%. The results obtained by the amplification of the dilution panel corresponded to the quantification of NASBA QT using the undiluted (5000 copies) HIV-1 RNA standard. So far, the NASBA HIV-1 RNA QT is the only commercially available amplification assay which permits a quantitative detection of HIV-1 RNA. Our results, although preliminary, show that the NASBA allows an accurate and reproducible quantitation of HIV RNA combined with a simple nucleic acid isolation procedure and non-radioactive detection of the amplificates.

No abstract

No abstract