RAPD-PCR and Outer Membrane Protein Characterization of Vibrio alginolyticus and Vibrio parahaemolyticus Isolated from Diseased shrimp

Abstract

Vibrio alginolyticus and V. parahaemolyticus were characterized by random amplified polymorphic DNA-polymerase chain reaction (RAPD-PCR) and profiling of outer membrane proteins (OMP) to the genetic level to enable easier identification of these microorganisms from diseased shrimp (Penaeus monodon) and adopt control measures. Of 20 tested Operon primers (OPC), only OPC5 and OPC18 were used for PCR amlification of 70 isolates representing three strains of each species. OPC5 amplified 1-8 bands with sizes ranging 0.42-2.95 Kb in V. alginolyticus, 5-10 fragments with sizes ranging 0.449-3.55 Kb in V. parahaemolyticus, and one unique common band of 1.34 Kb in both species. When OPC18 was used, one dark band (2.4 Kb) was common to all V. alginolyticus strains and another dark band (1.77 Kb) was common to two strains and the V. alginolyticus ATCC reference strain. One protein band (37.63 kDa) was common to both V. alginolyticus and V. parahaemolyticus and could be used as a diagnostic OMP marker for both species. The simpler and more rapid dot-ELISA and indirect ELISA methods confirmed identification of these pathogenic bacteria.