APPLICATION OF ?BT & ?BC BACTERIOPHAGES IN INHIBITION AND ERADICATION OF ENTEROBACTER SP. U7 BIOFILM AS BIOCONTROL AGENT FOR INFECTION IN LITOPENAEUS VANNAMEI

• Main Author: Valeria, Bella
• Format: Final Project
• Language: Indonesia
• Online Access: https://digilib.itb.ac.id/gdl/view/65549
• Institution: Institut Teknologi Bandung

The decrease production of white leg shrimp (Litopenaeus vannamei) can be caused by opportunistic pathogenic bacteria such as Vibrio parahaemolyticus which can cause AHPND (Acute Hepatopancreatic Necrosis Disease). AHPND causes shrimp mortality and dysbiosis which characterized by an increase in abundance of Enterobacter sp. that capable to form biofilms. The biofilm can protect other pathogens such as V. parahaemolyticus. Bacteriophages have the potential to be biocontrol agent against biofilm of Enterobacter sp. because of their specificity and safe for the environment, but it is necessary to optimize the Multiplicity of Infection (MOI) so that the infection could be effective. In previous studies, inhibition and eradication test against Enterobacter sp. U7 (106 CFU/mL) with ?Bt and ?Bc still not optimum. The aims for this study were: (1) to identify U7 isolates; (2) to determine the ability of Enterobacter sp. U7 in forming biofilms based on the presence of csgA and csgD genes; (3) to determine the optimum concentraction Enterobacter sp. U7 in forming biofilms; (4) to determine the virulence ability of Enterobacter sp. U7 against L. vannamei based on the presence of pirA gene; (5) to determine the efficiency of inhibition and eradication of biofilm Enterobacter sp. U7 with ?Bt and ?Bc; and (6) to determine the burst size of ?Bt and ?Bc and viability of Enterobacter sp. U7 after treatment with optimum ?Bt and ?Bc. Idenfitication of U7 isolates was carried out by 16s rRNA sequencing. Determination of the efficiency of inhibition and eradication Enterobacter sp. U7 biofilm with ?Bt and ?Bc based on MBIC50 and MBEC50 values were carried out by biofilm assay. The medium used in this study was Luria Bertani, pH 7, salinity 1%, temperature 37oC. The result of this study showed that U7 isolates were Enterobacter cloacae, has csgA and csgD genes to form biofilms and pirA gene for virulence against L. vannamei. The optimum concentration of biofilm formation by Enterobacter sp. U7 was 103 CFU/mL. Phage therapy treatment with ?Bc had the highest MBIC50 at MOI 0.1 with burst size 3.17 x 1010 PFU/cell and the inhibition of viability of planktonic and biofilm bacteria after 24 hours was 65.28% and 76.91%, respectively. While the phage therapy treatment with ?Bt had the highest MBEC50 at MOI 100 with burst size 2.81 x 106 PFU/cell and the inhibition of viability of planktonic and biofilm bacteria after 72 hours was 31.12% and 50%, respectively. Based on this study, ?Bc and ?Bt were confirmed to be able to inhibit and eradicate biofilms of E. U7. The two bacteriophages have different receptors on E. U7., namely ?Bc on the cell surface LPS component, while ?Bt on the biofilm EPS component.