Respiratory disease caused by a species B2 adenovirus in a military camp in Turkey.

In April 2004, two patients were admitted to hospital in Berlin, Germany, with clinical signs of acute respiratory infection after returning from a military exercise in their home country of Turkey. They were admitted to a high security infectious disease unit as epidemiological data pointed to an outbreak of unknown etiology. Samples taken at the time of admission proved to be strongly positive for Adenovirus by PCR, but negative for Influenza A/H1N1 virus, Influenza A/H3N2 virus, Influenza B virus, Respiratory syncytial virus, and SARS coronavirus. No evidence for bacterial infection was obtained by serological tests and blood cultures. The adenovirus detected was characterized further by genotyping and was identified as a species B2 virus with the highest similarity to adenovirus type 11a.

Development of a PCR-based assay for detection, quantification, and genotyping of human adenoviruses.

BACKGROUND: Adenoviruses (AdVs) can cause serious disease in immunosuppressed patients, particularly those undergoing allogeneic stem cell transplantation. A method for virus quantification in clinical specimens is essential for monitoring patient adenoviral loads and evaluating new therapeutic approaches. METHODS: We developed a PCR-based assay that combines detection and genotyping of human AdVs, targeting a highly conserved region of the adenoviral genome coding for the DNA polymerase (AdV DPol PCR). We tested the diagnostic applicability of this PCR-based assay by analyzing 159 clinical specimens from children with respiratory disease and comparing the results with those obtained by nested PCR analysis. RESULTS: The PCR assay detected all currently known AdV serotypes, with a detection limit of approximately 10 genome equivalents per reaction for 49 of 51 serotypes. No cross-reactivity to human DNA or other DNA viruses was observed. In addition, genotyping of PCR-positive samples was achieved within minutes by fluorescence curve melting analysis in a LightCycler instrument using 6 pairs of hybridization probes, each specific for a single AdV species. Results for clinical specimens were in good concordance with those obtained by nested PCR. CONCLUSION: The presented assay is a suitable tool for the detection and genotyping of human AdVs in clinical samples.

Approaching virus safety in xenotransplantation: a search for unrecognized herpesviruses in pigs.

The identification of porcine viruses so far unrecognized is required to minimize virus-related risks associated with xenotransplantation. We used a pan-herpes consensus polymerase chain reaction assay to search for unrecognized porcine species of the Herpesviridae. The assay targets conserved regions of the herpesvirus DNA polymerase (DPOL) gene, using primers that were modified to diminish the assay's recognition capacity for the highly prevalent porcine lymphotropic herpesviruses 1, 2 and 3 (PLHV-1, -2, -3), without substantially lowering the universal detection capacity of the assay. Analysis of 495 porcine blood and tissue samples from 294 animals, including 35 samples from 20 immunosuppressed pigs, resulted in the amplification of 128 herpesviral DPOL sequences. Sequence analysis attributed 127 of the amplimers to the known porcine herpesviruses (PLHV-1, -2, -3; porcine cytomegalovirus; pseudorabiesvirus). In none of the pig samples analyzed here, evidence was obtained for the presence of additional novel porcine herpesvirus species. Therefore we conclude that pigs bred for the purpose of xenotransplantation pose a negligible risk of transmitting presently unrecognized herpesviruses to organ recipients.

A novel porcine gammaherpesvirus.

A novel porcine gammaherpesvirus was detected in the blood of domestic pigs by PCR. With degenerate-primer PCR and subsequent long-distance PCR approaches a 60-kbp genome stretch was amplified. Sequence analysis revealed the presence of the gammaherpesvirus ORFs 03 to 46 as well as a putative chemokine receptor and a v-bcl-2 gene. The 60-kbp sequence was compared with the corresponding sequence of the porcine lymphotropic herpesvirus 1 (PLHV-1) published recently and the sequence of PLHV-2, which was amplified from porcine tonsil. Considerable sequence differences (amino acid identities: 49-89%) were found between the novel virus and PLHV-1 as well as PLHV-2, which were very closely related to each other (amino acid identities: 85-98%). The novel virus had essentially the same genome organization as PLHV-1 and -2 and was therefore designated PLHV-3. Like PLHV-1 and -2, PLHV-3 was frequently found in the blood and in lymphoid organs of domestic and feral pigs from different geographic locations. In the blood, the PLHVs were detected predominantly in B-cells. Indication for latent as well as productive PLHV-3 infection was found in the porcine B-cell line L23. It can be concluded that the PLHVs are widespread and are likely to cause a persistent B-lymphotropic infection. Since PLHV-1 has been implicated in the development of porcine posttransplantation lymphoproliferative disease, all porcine lymphotropic gammaherpesviruses are of concern when pigs are used as donors in xenotransplantation.