Desenvolvimento de ferramentas moleculares para identificação de variantes genotípicas do vírus causador da Síndrome da Mancha Branca em camarões

Abstract

The northeastern region is responsible for more than 90% of shrimp produced in Brazil, including the two largest producers, the states of Ceará and Rio Grande do Norte. The virus that causes white spot syndrome (WSSV) has been considered a major challenge faced by global shrimp farming in recent decades. Despite the fact that it is the biggest problem facing Brazilian shrimp farming today, the epidemiological status and the genetic variability of the WSSV have been little studied in Brazil to date. The genetic variability of WSSV has been investigated in different regions of the world, especially in studies of viral dispersion and in attempts to correlate genotype and virulence. This variability is accessed primarily through the variable regions present at the insertions between ORFs 23/24 and 14/15, and variable number of tandem repeats (VNTRs) occurring in ORFs 75, 94 and 125. The primers used to genotype the WSSV based on these regions have been described for at least 10 years, and some do not work efficiently to genotype new WSSV variants including those occurring in Brazil. In the present work, a new set of primers was developed to amplify the variable regions in the WSSV genome by PCR, based on alignments of all the sequences available so far in the public databases. The new primers were synthesized, and new PCR reactions were standardized, having their efficiency compared to previously described protocols and primers. The new primers were used in a pilot epidemiological study in which shrimp samples were collected on farms located along the coast of Rio Grande do Norte. The prevalence of WSSV was 58%, 100% and 60% in the farms investigated in the southern, central and northern regions respectively. The samples with the highest viral load were genotyped. The new set of primers proved to be more efficient in comparison with the primers already described, allowing the genotyping of more samples. The results indicate that WSSV prevalence in the state of Rio Grande do Norte was high at the time the samples were collected and that the genetic variability of WSSV in the state is low and was observed mainly in the ORF94 marker. The results allowed to observe some evidence that events of genetic recombination may have originated the Brazilian isolates of WSSV.