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Characterization of an anti‐pathogenic lactonase from Palk Bay metagenome
Author(s) -
PANDIAN SHUNMUGIAH THEVAR KARUTHA,
NITHYA CHARI,
VISZWAPRIYA DHARMAPRAKASH
Publication year - 2012
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.26.1_supplement.1002.2
Subject(s) - chromobacterium violaceum , microbiology and biotechnology , quorum sensing , chemistry , biology , fosmid , bacteria , biofilm , metagenomics , biochemistry , gene , genetics
Objectives Screening of metaclones for anti‐pathogenic activity and identification of anti‐pathogenic agents. Methods Screening for anti‐pathogenic activity using Chromobacterium violaceum (ATCC 12472) and CV026 as indicators. Mode of action by TLC (Thin Layer Chromatography), HPLC (High Performance Liquid Chromatography) and PCR (Polymerase Chain Reaction). Results Recombinant fosmid clones (N=10200) were propagated in E. coli DH10B. When individually 1056 clones were used to challenge C. violaceum CV026, eight clones could able to inactivate the perception of QS signal HHL (C6‐HSL) by the Chromobacterium biosensor. Additional assays revealed that the metagenomic clone M8‐70 disrupted AHL signals, including C6‐HSL, heptanoyl‐ (C7‐ ), octanoyl‐ (C8‐) and decanoyl‐homoserine lactone (C10‐HSL), as well as the hydroxy‐ or oxo‐substituted derivatives at carbon 3, such as OC6‐HSL, OC8‐HSL and OHC8‐HSL. Culture supernatants of M8‐70 was examined for its effect on P. aeruginosa PAO1 (LasA, LasB, Pyocyanin, Pyoverdin and and Total protease), S. marcescens (prodigiosin production, swarming, secreted caseinase and lipase) QS system and distinct profiles were obtained for las and rhl QS genes. Furthermore, the growth curve analysis and cell count method made it clear that the supernatant of M8‐70 had no antibacterial activity toward P. aeruginosa PAO1 and S. marcescens . The results of the ring closure assay and the sequence analysis of the PCR products from M8‐70 (GenBank accession number HQ876560 ) revealed that its activity as lactonase activity. Conclusion In this study a lactonase is reported through metagenomics approach from the sediment of Palk Bay which was found responsible for the inhibition of QS mediated virulence factors of Gram negative bacterial pathogens without inhibiting their growth.