DETECTION OF RHINOVIRUS INFECTION OF THE NASAL-MUCOSA BY OLIGONUCLEOTIDE IN-SITU HYBRIDIZATION

Human rhinoviruses (HRVs) cause the common cold and often induce lower airway symptoms such as cough and wheezing. Although HRV infection is presumed to involve primarily ciliated epithelial cells, this has not been confirmed in vivo, and the cellular distribution and spread of i nfection as well as the pathogenesis of cold related nasal and chest s ymptoms remain speculative. We have developed in situ hybridization (I SH) to explore localization of the virus to airway tissues, employing HRV 16-derived oligonucleotide probes after sequencing part of the gen ome of this serotype. A reverse transcription-polymerase chain reactio n was used to generate DNA from HRV 16 for sequencing; this yielded 30 5 nucleotide bases that showed considerable homology to other HRVs. Th e HRV 16 sequence was used to design oligonucleotides functioning as a ntisense and sense probes. These probes as well as random sequence and pathogen control oligonucleotides were applied to HRV-infected cell-c lot complexes and finally to sections from six paired nasal biopsies o btained before, during, or after HRV-proven colds. Specificity of hybr ids was established by the absence of signal in uninfected tissue, in cells infected with other viruses, after RNase pretreatment, and with application of control probes. Hybridization signals were observed in epithelial cells in three of six biopsies obtained during a cold, usin g probes to viral (+) strand; intermediate (-) strand, implying viral replication, was present in one biopsy. Evidence for infection of none pithelial cells was inconclusive. HRVs cause productive infection of n asal epithelium during a cold and their intracellular localization may produce perturbation of inflammatory mediators and cytokine profiles. Use of ISH will permit studies exploring the pathogenesis of HRV-rela ted symptoms and clarification of the mechanisms of lower airway invol vement.