Detection of tetracycline-resistance determinants by multiplex polymerase chain reaction in Edwardsiella tarda isolated from fish farms in Korea

To determine and discriminate the types of tetracycline (Tc)-resistance determinants (tet), we developed a multiplex polymerase chain reaction (PCR). With minimized numbers of primers derived from the variable and conserved regions of six different types of tet genes, tet(A)–(E) and (G), the multiplex PCR was sensitive and specific enough to discriminate the various types of tet genes, even multiple tet genes in an individual resistant isolate, by the different sizes of the resulting PCR products. Each of 20 Tc-resistant Edwardsiella tarda (Ed. tarda) isolates from diseased fish from aquatic farms in Korea carrying either one or two tet genes of types (A), (D), (B), or (G) gave PCR products of the appropriate lengths. Among the four types of tet genes found in Ed. tarda, two types, tet(A) and (D), were always present on mobile plasmids, and the other two types, tet(B) and (G), were located on the nonmobile nucleic acids. This is the first time that either of these two genes, type (B) or (G), have been found in Ed. tarda isolates. The two most common types of Tc-resistance determinant were, tet(A) and (D), occurring in 55% and 45% of total Tc-resistant Ed. tarda isolates, respectively.