Molecular characterization of porcine Pasteurella multocida by REA and RAPD-PCR

Abstract

To determine the genetic relatedness between Pasteurella multocida, twenty one porcine isolates were characterized by using Restriction Endonuclease Analysis (REA) and Randomly Amplified Polymorphic DNA-PCR (RAPD-PCR). The REA using the enzymes, Hha I and Hpa II revealed different fingerprinting patterns. Irrespective of serotypes of Pasteurella multocida isolates, a total of 10 different RE banding patterns within the range of 20292 bp and 977 bp were obtained following treatment with Hpa II enzyme and 10 different RE banding patterns with Hha I enzyme treatment with banding patterns ranging of 18720 bp and 727 bp. Pasteurella multocida serotype D isolates, showed four different banding patterns with Hpa II, ranging from 20292-783 base pair (bp) and five different RE patterns with Hha I, within a range of 18720-2021 bp. However, Pasteurella multocida serotype A isolates exhibited eight different patterns (18382- 478 bp) with Hpa II enzyme and seven patterns (18720-1650 bp) with Hha I enzyme. RAPD-PCR assay of all the Pasteurella multocida isolates using random primer 1 revealed a high degree of discrimination among the isolates, with a banding pattern ranging from 0.180 to 1.344 bp. A common band of size approximately 647 bp was found to be shared by all the 21 isolates. P. multocida serotype D isolates showed a wide range of variation with 5 different banding patterns and serotype A isolates with 10 different banding patterns. The banding pattern of the amplified products with random primer 2 was within a range from 0.246 to 1.754 bp. Both serotype A and D isolates showed six different banding patterns respectively. RAPD-PCR gave clear visible banding patterns, when compared to REA.