Inhibition of biofilm formation, quorum sensing and other virulence factors in Pseudomonas aeruginosa by polyphenols of Gynura procumbens leaves.

Quorum sensing (QS) enables virulence factors in bacteria for biofilm formation and pathogenic invasion. Therefore, quorum quenching (QQ), disruption of QS circuit, becomes an alternative antimicrobial therapy. In this study, leaf extract of Gynura procumbens (GP) was used to inhibit biofilm and virulent factors in Pseudomonas aeruginosa. The extract inhibited the biofilm production (p ≤ 0.05) in P. aeruginosa strains MZ2F and MZ4A. The minimum biofilm eradication concentration (MBEC) was recorded at 250 and 500 µg/ml while total activity was found at 288 and 144 ml/g, respectively. Moreover, a significant reduction of virulence factors (p ≤ 0.05) at sub-MBEC without affecting the growth implies the QQ action of the extract. The bioactive fractions were rich in polyphenols and tentatively identified as quercetin and myricetin (Rf=0.53-0.60). Furthermore, we employed computational methods to validate our findings and their interactions with QS receptors (LasR and RhlR). Interestingly, docking studies have also shown that quercetin and myricetin are the promising anti-QS agents out of 31 GP compounds. Notably, their binding affinity ranged between -9.77 and -10.52 kcal/mol for both QS receptors, with controls ranging from -5.40 to -8.97 kcal/mol. Besides, ΔG of quercetin and myricetin with LasR was -71.56 and -74.88 kcal/mol, respectively. Moreover, compounds were suitable drug candidates with stable binding interactions. Therefore, the anti-QS activity of GP leaves and the identified polyphenols can be used in developing QQ-based therapeutics. Communicated by Ramaswamy H. Sarma.

Computational prediction of active sites and ligands in different AHL quorum quenching lactonases and acylases.

With the emergence of multidrug-resistant 'superbug', conventional treatments become obsolete. Quorum quenching (QQ), enzyme-dependent alteration of quorum sensing (QS), is now considered as a promising antimicrobial therapy because of its potentiality to impede virulence gene expression without resulting in growth inhibition and antibiotic resistance. In our study, we intended to compare between two major QQ enzyme groups (i.e., AHL lactonases and AHL acylases) in terms of their structural and functional aspects. The amino acid composition-based principal component analysis (PCA) suggested that probably there is no structural and functional overlapping between the two groups of enzymes as well as within the lactonase enzymes but the acylases may functionally be affected by one another. In subcellular localization analysis, we also found that most lactonases are cytoplasmic while acylases are periplasmic. Investigation on the secondary structural features showed random coil dominates over alpha-helix and beta-sheet in all evaluated enzymes. For structural comparison, the tertiary structures of the selected proteins were modelled and submitted to the PMDB database (Accession ID: PM0081007 to PM0081018). Interestingly, sequence alignment revealed the presence of several conserved domains important for functions in both protein groups. In addition, three amino acid residues, namely aspartic acid, histidine, and isoleucine, were common in the active sites of all protein models while most frequent ligands were found to be 3C7, FEO, and PAC. Importantly, binding interactions of predicted ligands were similar to that of native QS signal molecules. Furthermore, hydrogen bonds analysis suggested six proteins are more stable than others. We believe that the knowledge of this comparative study could be useful for further research in the development of QSbased universal antibacterial strategies.

The periodontopathogenic bacterium Eikenella corrodens produces an autoinducer-2-inactivating enzyme.

Eikenella corrodens produces autoinducer-2 (AI-2) in the mid log phase, and AI-2 activity decreases dramatically during the stationary phase. We investigated the mechanism underlying this decrease in AI-2 activity. To analyze the mechanism, we extracted and purified AI-2 from the supernatant of mid-log-phase culture. Simultaneously, the stationary-phase culture supernatant was fractionated by ammonium sulfate precipitation. On incubating purified AI-2 and 4-hydroxy-5-methyl-3(2H)-furanone (MHF) with each fraction, the 30% fraction decreased both AI-2 and MHF activities. The data suggest that AI-2 and MHF were rendered inactive in the same manner. Heat and/or trypsin treatment of the 30% fraction did not completely arrest AI-2 inactivation, suggesting that partially heat-stable proteins are involved in AI-2 inactivation. We observed that an enzyme converted MHF to another form. This suggests that E. corrodens produces an AI-2 inactivating enzyme, and that AI-2 can be degraded or modified by it.