THE IDENTIFICATION OF MYCOPLASMA HAEMOSUIS IN THAI SWINE FARMS WITH CONVENTIONAL AND MOLECULAR TECHNIQUES

Abstract

Swine hemoplasmas caused by Mycoplasma haemosuis (M. haemosuis) or Mycoplasma suis (M. suis) or previously known as Eperythrozoon suis (E. suis) is the infectious disease via insect vectors that the information of its pathogenesis and prevalence had been still unclear. Therefore, this study aimed to diagnose M. suis in swine herds in Thailand by using conventional and molecular techniques. Three hundred suspected pigs tending to M. suis infection were collected for blood samples from open-air swine farms in six provinces of Thailand such as Nakhon Pathom, Chachoengsao, Prachinburi, Ratchaburi, Sisaket, and Chiang Mai. Blood were detected by thin blood smear staining with 10% Giemsa, acridine orange (AO) and conventional PCR using 10 samples that were positive to M. suis by 10% Giemsa and AO and nested PCR based on conserve gene (16srRNA) of 27 strains of M. suis from domestic pigs and wild pigs in GenBank. Outer primers were designed using Bioedit V7.2.5 and Multalin programs and inner primers were designed based on DNA sequence of derived recombinant DNA as positive control from Japan using Primer3 program. Those primers in this study were examined sensitivity and specificity tests with other Mycoplasma in feline and swine together with 10 blood samples that were positive to M. suis based on Giemsa and acridine orange. The result of primers investigation found that designed primers were highly specificity to M. suis at position 603 bp (outer) and 222 bp (inner) or only 222 bp (inner) and highly sensitivity that could scrutinize a quantity of DNA at least 2.351 x 10-6 copies/µl. Based on 300 blood samples, twenty-three samples (7.67%) showed basophilic discoid shape using 10% Giemsa. Forty-five blood samples (15%) were found using acridine orange and 70 samples (23.33%) using nested PCR were positive. Consequently, this study is the first to investigate M. suis within swine farms in Thailand using molecular techniques (PCR and nested PCR). The results of this study could be used as alternative diagnostic assays especially carrier pigs for continuing control of outbreak and treatment planning.