The work presented in this thesis focused on two diverse aspects of rhinoviruses, which represent a frequent human pathogen. The first part addressed a basic science theme of virology, namely recombination. Experimental intra- and interspecies HRV recombination using artificially engineered chimeras and non replicative recombination was studied. The small number of recombinants comprised in our results and the usage of a growth deficient parental HRV-C render it difficult to draw general conclusions regarding HRV recombination patterns. Nevertheless, the two different and somehow complementary experimental approaches used in this study provided some valuable information, which may be summarized as follows. First, the 5‟UTR of HRVs, and even HEVs, seem to be largely interchangeable.
Second, recombination in the coding region generated viable intraspecies but not interspecies recombinants. Third, non replicative intraspecies RNA recombination occurs in cell culture upon co-transfection of two complementary deleted HRV genomes and this method could be used to map theoretical HRV recombination sites.
The application of these approaches to an extended number of combinations could expand our understanding of HRV genomic organization and perhaps of genetic determinants of specific phenotypes.
HRV chimeras in which the P2 and P3 regions, coding for non structural proteins, originate from two different HRV genomes could be engineered and tested. Intraspecies chimeras representative of each of the three HRV species may be studied in order to obtain a comprehensive understanding of the experimental HRV recombination potential. Interspecies recombinants in the same genomic regions could also be designed to test the HRV interspecies genetic barrier. Additional non replicative HRV RNA recombination experiments including co-transfection of partially deleted but complementary HRV RNA pairs of each
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HRV species would certainly add supplementary knowledge. Of course, experiments involving HRV-C recombinants will require appropriate culture systems, as members of this species are not cultivable in classic cell lines.
The second part of this thesis concerned a more clinical aspect of rhinoviruses, namely the feasibility of HRV RNA quantification in respiratory specimens using real-time RT-PCR.
Although the Panenterhino/Ge/08 real-time RT-PCR assay was accurate for HRV RNA quantification, the results obtained should be interpreted carefully, according to the issues discussed above. However, this tool might be of use in specific clinical situations, provided that sampling methods are rigorously standardized. For instance, subactute or chronic HRV infections in immunosupressed patients could be monitored, and correlations between viral load and symptom severity could be assessed with this assay. Clinical studies addressing these issues are currently underway. In the future, measuring HRV RNA loads could assess the efficacy of anti HRV drugs.
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