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Certain strains of V. parahaemolyticus carry a gene that encodes a toxin that causes Acute hepatopancreatic necrosis disease (AHPND) in P. vannamei. AHPND attacks shrimp post larvae within 20-30 days after stocking causing up to 100% mortality. The expression of these virulent genes is controlled by the quorum sensing system. This system is inhibited by an anti-quorum sensing (AQS) mechanism. Several Bacillus strains have AQS mechanism by producing AHLLactonase enzyme. Therefore, this study aimed to obtain Bacillus spp. having AQS activity for controlling AHPND. The study was conducted from isolation and selection of Bacillus isolates, as well as determination of AQS activity. From 22 samples consisting of shrimp intestines, water and pond sediment samples, a total of 151 isolates of Bacillus spp. were isolated. The screening test for AQS activity obtained 11 isolates that showed AQS activity on Cromobacterium violaceum. Determination of violacein pigment in liquid cultures of C. violaceum showed the index value of the pigment formation was between 0.025-0.166 and 0.026-0.567 at 24-hour and between 48-hour incubations, respectively. The quantitative analysis of violacein production showed that there were six isolates of Bacillus could inhibit the pigment production more than 75%. The isolates were identified as Bacillus cereus (four isolates), Bacillus thuringiensis (one isolate), and Bacillus velezensis (one isolate), respectively. The molecular analysis had confirmed that the isolates have aiiA genes encoding AHL-lactonase enzyme. These Bacillus isolates have potential application for controlling AHNPD disease. [ABSTRACT FROM AUTHOR]

Pathogenic bacteria use quorum sensing (QS) mechanism to express virulence gene. QS mechanism is induced by acyl homoserine lactone (AHL) as universal QS autoinducer in Gram negative bacteria. Inhibition of virulence genes expression can be done by degrading the AHL compounds using AHL-lactonase encoding aiiA gene. The study aimed to clone and express aiiA gene of Bacillus cereus B10 into E.coli BL21 (DE3) pLysS. The aiiA gene was cloned into DH5α through pGEMT-easy vector and expressed using pETt15b vector. The aiiA sequences and 3D structure of AiiA protein were analyzed with ORF finder, swiss model program and PyMOL software. Overexpression of aiiA gene in E.coli BL21 (DE3) pLysS was induced by 0.1 mM, 0.5 mM, and 1 mM IPTG through SDS PAGE method. The result showed that aiiA gene consisted of 250 amino acids and had high similarity with AiiA proteins in Bacillus cereus AAL98271. AHL lactonase of B.cereus B10 showed high homology with zinc metallo-beta-lactamase family with HXHXDH motif. The SDS- PAGE analysis showed that AiiA proteins of Bacillus cereus B10 had a molecular weight of 28,18 Kda. The aiiA gene from B.cereus B10 was successfully expressed into E.coli BL21(DE3). [ABSTRACT FROM AUTHOR]

The virulence gene expression of phytopathogenic bacteria, such as Dickeya dadantii is regulated by quorum sensing (QS) mechanism. The QS mechanism is induced by acyl homoserine lactone (AHL) that is universal autoinducer of Gram-negative bacteria. Degradation of AHL compound can be used to control the virulence of phytopathogenic bacteria. The enzyme of AHL lactonase is encoded by the aiiA gene and can degrade the lactone ring of AHL. This study was aimed to characterize aiiA gene encoding AHL lactonase from 20 isolates of shrimp pond and soil forest, Indonesia and examine the isolates ability to reduce violacein pigment of Chromobacterium violaceum and soft rot disease symptoms caused by D. dadantii. The results showed nine bacterial isolates (B4, B10, B11, B13, B14, B16, B17, B18 and B19) containing the aiiA gene encoding AHL lactonase. Based on 16S rRNA gene identification, 9 isolates had the highest similarity with Bacillus toyonensis, B. cereus, B. thuringiensis, B. mycoides and B. subtilis. These isolates had the capability to reduce the formation of violacein pigment and the symptoms of soft rot disease in potatoes. These results show that the AHL lactonase produced by these isolates can be used as Quorum Quenching (QQ) agent to inhibit the QS mechanism [ABSTRACT FROM AUTHOR]

Bekasam is an Indonesian fermented fish product that tastes sour and mostly contains lactic acid bacteria (LAB). The aim of this study is to obtain and characterize LAB isolates from bekasam and to study their potency in inhibiting the growth of pathogenic bacteria, i.e. Escherichia coli, Salmonella typhimurium ATCC 14028, Bacillus cereus, Staphylococcus aureus and Listeria monocytogenes. LAB were isolated from bekasam using media of MRSA supplemented with CaCO3 0.5%. Incubation was at 37°C for 48 hours. The pure cultures were verified as LAB based on morphological and biochemical characteristics. Eight bekasam samples showed that the total average of LAB were 1.4 x 108 to 9.0 x 108 CFU/mL. Seventy four isolates were successfully isolated. It was found that 62 isolates (84%) belonged to LAB, and 23 isolates of them could inhibit the growth of the five pathogenic bacteria. The highest inhibition zone was accounted for S. aureus. However, neutralized supernatant of the LAB culture did not inhibit the growth of the pathogenic bacteria. These results indicate that the LAB inhibition to the pathogenic bacteria was due to the organic acid, and that perhaps the main factor in the bekasam preservation by LAB. [ABSTRACT FROM AUTHOR]