Isolation of bacteriophage from shrimp ponds in Soc Trang Province for inhibiting biofilm-forming <i>Micrococcus luteus</i>

The bacterial biofilm can support several pathogens to be resistant to antibiotics, fungicides, and parasiticides. Slime from shrimp ponds is the common habitat of some biofilm-forming bacteria and their natural enemies, especially bacteriophage. Therefore, this study aimed to isolate these biofilm-forming bacteria from the bottom slime of shrimp ponds and subsequently isolate several bacteriophage strains that were capable of infecting these bacteria. For the methods, the disadvantageous bacteria were collected from the samples and isolated through morphology, biochemical reactions, and molecular biological identification. The bacteriophage was simultaneously collected from the same slime samples and isolated through spot test and double agar overlay assay. Moreover, the bacterial biofilm detachment ability of the bacteriophage was evaluated through the optical density measurement of the bacterial biofilm with and without bacteriophage. As a result, the isolated biofilm-forming bacteria STM1 was both diplococci and tetrad, Gram-positive, non-motile, and yellow colony on the TSA medium. The catalase and oxidase activities of the STM1 were positive, and this strain did not ferment glucose and mannitol. Through morphological observations, biochemical reactions, and 16S rRNA gene analysis, STM1 was similar to Micrococcus luteus (PP495084.1) at 99.5 %. For the bacteriophage selection, four strains detaching bacterial biofilm were isolated at salinities between 5 and 20 ppt, wherein the phage pST9Δ2 strongly detached the STM1 biofilm (peaking at 82 % at 15ppt salinity).