An investigation was carried out to survey the prevalence of Marek’s disease virus serotype 1 (MDV-1) on unvaccinated healthy backyard chicken flocks from four districts in Cantho city and to analyze the genetic characterization of the meq gene. A total of 353 feather follicle samples were collected from 50 unvaccinated chicken flocks for Marek’s disease to investigate prevalent MDV-1 by polymerase chain reaction (PCR). The four represent strains were chosen for meq gene sequencing. The results revealed that 26 out of 353 samples were positive for MDV-1 accounting for 7.37%. There were significant differences in the prevalence of MDV-1 on chickens among districts (P<0.05). The Meq protein of the four strains (VT, CD, PD, and TL) contained 20.14% proline and three regions of proline repeats. Besides, all four strains occurred with amino acid point mutations. These strains had only a substitution of proline at position 217 as well as interruption of consecutive proline at site 2. Phylogenetic analysis indicated that these strains were within a group relating to virulent Italian isolates.