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. 2017 Apr 24;8:727.
doi: 10.3389/fmicb.2017.00727. eCollection 2017.

Leaf Extracts of Mangifera indica L. Inhibit Quorum Sensing - Regulated Production of Virulence Factors and Biofilm in Test Bacteria

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Free PMC article

Leaf Extracts of Mangifera indica L. Inhibit Quorum Sensing - Regulated Production of Virulence Factors and Biofilm in Test Bacteria

Fohad M Husain et al. Front Microbiol. .
Free PMC article

Abstract

Quorum sensing (QS) is a global gene regulatory mechanism in bacteria for various traits including virulence factors. Disabling QS system with anti-infective agent is considered as a potential strategy to prevent bacterial infection. Mangifera indica L. (mango) has been shown to possess various biological activities including anti-QS. This study investigates the efficacy of leaf extracts on QS-regulated virulence factors and biofilm formation in Gram negative pathogens. Mango leaf (ML) extract was tested for QS inhibition and QS-regulated virulence factors using various indicator strains. It was further correlated with the biofilm inhibition and confirmed by electron microscopy. Phytochemical analysis was carried out using ultra performance liquid chromatography (UPLC) and gas chromatography-mass spectrometry (GC-MS) analysis. In vitro evaluation of anti-QS activity of ML extracts against Chromobacterium violaceum revealed promising dose-dependent interference in violacein production, by methanol extract. QS inhibitory activity is also demonstrated by reduction in elastase (76%), total protease (56%), pyocyanin (89%), chitinase (55%), exopolysaccharide production (58%) and swarming motility (74%) in Pseudomonas aeruginosa PAO1 at 800 μg/ml concentration. Biofilm formation by P. aeruginosa PAO1 and Aeromonas hydrophila WAF38 was reduced considerably (36-82%) over control. The inhibition of biofilm was also observed by scanning electron microscopy. Moreover, ML extracts significantly reduced mortality of Caenorhabditis elegans pre-infected with PAO1 at the tested concentration. Phytochemical analysis of active extracts revealed very high content of phenolics in methanol extract and a total of 14 compounds were detected by GC-MS and UPLC. These findings suggest that phytochemicals from the ML could provide bioactive anti-infective and needs further investigation to isolate and uncover their therapeutic efficacy.

Keywords: C. elegans; GC-MS; Mangifera indica; biofilm inhibition; leaf extract; quorum sensing; virulence factors.

Figures

FIGURE 1
FIGURE 1
(A) Inhibition of violacein by methanol extract of Mangifera indica (leaf) in C. violaceum at the concentration of 200 (1), 400 (2), 800 (3), and 1600 (4) μg/ml. (B) Quantitative assessment of violacein inhibition in CVO26 at sub-inhibitory concentrations of M. indica (leaf) extract. All of the data are presented as mean ± standard deviation. p ≤ 0.05, ∗∗∗p ≤ 0.001.
FIGURE 2
FIGURE 2
Scanning electron microscopic (SEM) images for inhibition of biofilm of Pseudomonas aeruginosa PA01 at Sub minimum inhibitory concentrations. Images shown the effect on biofilm formation at 400 μg/ml (B) and 800 μg/ml (C) concentration of Mangifera indica (methanol) extract compared to control (A).
FIGURE 3
FIGURE 3
Inhibition of swarming motility in P. aeruginosa PAO1 by sub-MICs of methanol extract of M. indica (leaf), (A) Untreated control; (B) 200 μg/ml; (C) 400 μg/ml; (D) 800 μg/ml.
FIGURE 4
FIGURE 4
Effect of M. indica (methanol) extract on β-galactosidase activity in E. coli MG4/pKDT17. All of the data are presented as mean ± SD. ∗∗p ≤ 0.01, ∗∗∗p ≤ 0.001.
FIGURE 5
FIGURE 5
Anti-infection potential of M. indica (methanol) extract on P. aeruginosa infected C. elegans. Figure represents effect of mango extract (800 μg/ml) on the survival of C. elegans. Nematodes were pre-infected with P. aeruginosa and treated in the presence and absence of mango extract for 4 days. Graph represents the average of three independent experiments and SD.
FIGURE 6
FIGURE 6
HPLC profile of Mangifera indica (methanol) leaf extract. Twenty microliter of extracts (1 mg/ml) were injected and analyzed using photo diode array detector on a C18 (250 mm × 4.6 mm ID × 5 μm) reverse phase column in a gradient of 3.5% aqueous phosphoric acid and acetonitrile. Chromatograms in (A) represent various polyphenolics detected at 280 nm. Chromatograms in the (B) represent analysis of different peaks as indicated in (A) in the range of 200–600 nm. mAu, milli absorbance units. For clarity of data, y-axis scales are different.

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