Saturday, July 31, 2010


Jana Moerbe Rocker and Jeffrey M. Lotz*
Department of Coastal Sciences
University of Southern Mississippi
Gulf Coast Research Laboratory
Ocean Springs, MS 39564 USA Geographic genetic variability has been noted in Taura Syndrome Virus (TSV) and this variability has been used in molecular epidemiological studies. However, little is known of the genetic variability that exists within a single host during an infection. Quasispecies is a term originally applied to a mathematical model describing the genetic variability of RNA viral particles within a host. Quasi-species result from a balance between a high mutation rate and selection against the assumed less fit mutations. Due to the lack of a proofreading mechanism in RNA-dependent RNA polymerase (RdRp), the replication of RNA is an inherently error-prone process Therefore, the RNA virus exists within its host as a heterogeneous mixture of sequences: the master sequence (the most common and fittest sequence) and various non-lethal but less fit mutants.

Our objective was to determine whether TSV exists as a quasi-species by characterizing the genetic variability within individual hosts with TSV infections. Further we compared the genetic variation between acutely infected shrimp and chronically infected shrimp. We approached characterizing the genetic variability by cloning and sequencing numerous TSV samples from one infected host. The assumption is that the relative abundance of sequences among the clones reflects the relative abundances of sequences within the host. In order to reduce any sequence errors during the PCR process we used high fidelity polymerases. We employed the CP2 gene, which is used for most geographical genetic characterization, for our quasi-species analysis. Five shrimp with acute infections and five shrimp with chronic infections were used in the analysis. We attempted to clone 25 hemolymph samples from each shrimp for a total of 250 clones. Each clone was sequenced and the sequences were analyzed.

We found that individual shrimp were carrying TSV with considerable sequence variation in the CP2 gene. The mean mutation frequency was 1.7 x 10-4. The mutation frequency in chronic infections (2.18 x 10-4) was higher than in acute infections (1.2 x 10-4).

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