Manganese uptake by MtsABC contributes to the pathogenesis of human pathogen group A streptococcus by resisting host nutritional immune defenses.

The interplay between host nutritional immune mechanisms and bacterial nutrient uptake systems has a major impact on the disease outcome. The host immune factor calprotectin (CP) limits the availability of essential transition metals, such as manganese (Mn) and zinc (Zn), to control the growth of invading pathogens. We previously demonstrated that the competition between CP and the human pathogen group A streptococcus (GAS) for Zn impacts GAS pathogenesis. However, the contribution of Mn sequestration by CP in GAS infection control and the role of GAS Mn acquisition systems in overcoming host-imposed Mn limitation remain unknown. Using a combination of in vitro and in vivo studies, we show that GAS-encoded mtsABC is a Mn uptake system that aids bacterial evasion of CP-imposed Mn scarcity and promotes GAS virulence. Mn deficiency caused by either the inactivation of mtsC or CP also impaired the protective function of GAS-encoded Mn-dependent superoxide dismutase. Our ex vivo studies using human saliva show that saliva is a Mn-scant body fluid, and Mn acquisition by MtsABC is critical for GAS survival in human saliva. Finally, animal infection studies using wild-type (WT) and CP-/- mice showed that MtsABC is critical for GAS virulence in WT mice but dispensable in mice lacking CP, indicating the direct interplay between MtsABC and CP in vivo. Together, our studies elucidate the role of the Mn import system in GAS evasion of host-imposed metal sequestration and underscore the translational potential of MtsABC as a therapeutic or prophylactic target.

ELONGATED HYPOCOTYL 5 regulates BRUTUS and affects iron acquisition and homeostasis in Arabidopsis thaliana.

Iron (Fe) is an essential micronutrient for both plants and animals. Fe-limitation significantly reduces crop yield and adversely impacts on human nutrition. Owing to limited bioavailability of Fe in soil, plants have adapted different strategies that not only regulate Fe-uptake and homeostasis but also bring modifications in root system architecture to enhance survival. Understanding the molecular mechanism underlying the root growth responses will have critical implications for plant breeding. Fe-uptake is regulated by a cascade of basic helix-loop-helix (bHLH) transcription factors (TFs) in plants. In this study, we report that HY5 (Elongated Hypocotyl 5), a member of the basic leucine zipper (bZIP) family of TFs, plays an important role in the Fe-deficiency signaling pathway in Arabidopsis thaliana. The hy5 mutant failed to mount optimum Fe-deficiency responses, and displayed root growth defects under Fe-limitation. Our analysis revealed that the induction of the genes involved in Fe-uptake pathway (FIT-FER-LIKE IRON DEFICIENCY-INDUCED TRANSCRIPTION FACTOR, FRO2-FERRIC REDUCTION OXIDASE 2 and IRT1-IRON-REGULATED TRANSPORTER1) is reduced in the hy5 mutant as compared with the wild-type plants under Fe-deficiency. Moreover, we also found that the expression of coumarin biosynthesis genes is affected in the hy5 mutant under Fe-deficiency. Our results also showed that HY5 negatively regulates BRUTUS (BTS) and POPEYE (PYE). Chromatin immunoprecipitation followed by quantitative polymerase chain reaction revealed direct binding of HY5 to the promoters of BTS, FRO2 and PYE. Altogether, our results showed that HY5 plays an important role in the regulation of Fe-deficiency responses in Arabidopsis.

Erythrina variegata L. bark: an untapped bioactive source harbouring therapeutic properties for the treatment of Alzheimer's disease.

A critical approach for target identification to detect the significant molecular mechanism of lead molecules via computational methods combined with in vitro procedures defines the modern strategy to combat untreatable diseases. Hence, the present investigation dealt to determine the effect of Erythrina variegata L. bark extract/fraction(s) over acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) inhibitory activity followed by target identification and docking analysis of prime phytoconstituents. The in vitro AChE and BChE enzyme inhibitory assay were performed. Phytoconstituents from E. variegata were screened for carcinogenicity and mutagenicity and predicted for their possible targets leading to the identification of two known targets, i.e. AChE and BChE. The alkaloids with non-carcinogenic and non-mutagenic properties were studied for their main moiety responsible for the inhibitory activity. The protein models were checked in ERRAT for their quality and the homology model was created using Modeller9.10v to fill missing amino acid residues. The docking study predicted the binding affinity of bioactive molecules with identified targets using AutoDock 4.2. Molecular dynamics (MD) simulations for top hits were performed by Schrodinger Desmond 6.1v software. Chloroform fraction showed potent inhibition of AChE and BChE with IC50 value of 38.03 ± 1.987 µg/mL and 20.67 ± 2.794 µg/mL, respectively. Among all the six major bioactive compounds, Erysotine and Erythraline scored the highest binding affinity with AChE and Erysodine with BChE. MD simulation for 20 ns production run demonstrated Erysotine and Erysodine stable interaction with Arg49 of AChE and Lys427 of BChE, respectively. The current data provide enough shreds of evidence supporting the utilization of indolo [7a,1-a] isoquinoline derivatives for the identification of a new drug molecule in the management of Alzheimer's disease. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s40203-021-00110-0.

Attenuation of quorum sensing mediated virulence factors production and biofilm formation in Pseudomonas aeruginosa PAO1 by Colletotrichum gloeosporioides HM3.

Signal dependent microbial communication in Pseudomonas aeruginosa PAO1 is a typical phenomenon mediated by acyl homo-serine lactone molecules that helps in developing biofilm and enhance antibiotic resistance. Microbial sources provide insight to the hidden treasure of secondary metabolites, and these structurally diversified chemical motifs can be used as antimicrobial and anti-infective agents. In the present study, endophytic fungus, Colletotrichum gloeosporioides HM3 isolated from Carica papaya leaves was explored for anti-infective potential against P. aeruginosa PAO1. The crude extract of C. gloeosporioides HM3 displayed bacteriostatic effect on P. aeruginosa PAO1 growth at 750 µg/ml concentration. A significant decline was observed in the production of quorum sensing regulated virulence factors, i.e. 56.32%, 62.54%, and 66.67% of pyocyanin, chitinase, and elastase enzyme, respectively. A drastic reduction in pathogenic determinant behaviour after treatment with crude extract of C. gloeosporioides HM3 i.e. EPS, rhamnolipid, and HCN production was noted. Light microscopy and CLSM analysis revealed that fungal extract treatment has reduced bacterial ability to form dense biofilm architecture. In silico analysis demonstrated the binding efficiency of bioactive compound, 4-(2,3-dimethoxybenzylidene)-3-methyl-1-(4-nitrophenyl)-2-pyrazolin-5-one, which is equipotent to the natural ligand and displayed a docking score of -5.436 kcal/mol with QS transcriptional regulator (LasR). Whereas the compound Acetamide, n-[tetrahydro-3-(phenylmethyl) thieno [3,4-d]thiazol-2 (3 h)-ylidene]-, s,s-dioxide exhibits a docking score of -4.088 kcal/mol (LasR) and -1.868 kcal/mol (RhlR) with cognate receptor proteins. Henceforth, the research report suggests C. gloeosporioides HM3 derived metabolites could be considered as a potential inhibitors of QS regulated virulence factors and biofilm production in P. aeruginosa PAO1.

Phomopsis tersa as Inhibitor of Quorum Sensing System and Biofilm Forming Ability of Pseudomonas aeruginosa.

Endophytic fungi provide rich reservoir for novel antimicrobial compounds. An endophytic fungus, from Carica papaya plant identified as Phomopsis tersa, was investigated for attenuating the quorum sensing mediated pathogenicity of Pseudomonas aeruginosa PAO1. Crude extract of P. tersa was found to reduce the production of redox-active pigments-pyocyanin and pyoverdine in P. aeruginosa PAO1 by 92.46% and 71.55%, respectively at sub-MIC concentration of 900 µg/mL. In addition, the crude extract was also able to inhibit the expression of virulence factors involved in biofilm formation: exopolysaccharide (72.21%) and alginate (72.50%). Secretion of cell-lytic enzymes was also found to be reduced: chitinase by 79.73% and elastase by 74.30%. 3-Isobutylhexahydropyrrolo[1,2-a]pyrazine-1,4-dione identified from GC-MS analysis, displayed favorable molecular interactions with P. aeruginosa transcriptional regulators, LasR and RhlR with good docking scores of - 6.873 kJ/mol and - 6.257 kJ/mol, respectively. The study thus reveals the potential use of P. tersa for discovering drugs against infectious pathogens.

Anti-quorum sensing and antibiofilm potential of Alternaria alternata, a foliar endophyte of Carica papaya, evidenced by QS assays and in-silico analysis.

In the present study, secondary metabolites from an endophytic fungus, Alternaria alternata, colonizing Carica papaya, demonstrated antiquorum sensing properties against Pseudomonas aeruginosa. This study reports the antagonistic effects of fungal crude extract of A. alternata against the various quorum sensing (QS) associated virulent factors such as percentage decrease in production of pyocyanin, alginate, chitinase and rhamnolipid; significant decrease in proteases activity such as LasA protease activity, staphylolytic activity, Las B elastase; and a marked decrease in biofilm formation and associated factors such as exopolysaccharide (EPS) production and cell surface hydrophobicity (CSH). Further, motility pattern i.e., swimming and swarming was also found to be inhibited. This down regulation of QS and associated factors are further supported by in-silico analysis of interaction between QS receptor LasR and bioactive molecules viz., sulfurous acid, 2-propyl tridecyl ester and 1,2-benzenedicarboxylic acid, bis(2-methylpropyl) ester present in fungal crude extract, found based on GCMS analysis, sketches the modulating ability of QS expression. This is the first report on an endophytic fungus of C. papaya having a role in QS inhibition against P. aeruginosa and lays a platform to explore further the endophytes for potent therapeutic agents in QS.

Green synthesis of anisotropic gold nanoparticles using hordenine and their antibiofilm efficacy against Pseudomonas aeruginosa.

Pseudomonas aeruginosa is a notorious pathogen that causes biofilm aided infections in patients with cystic fibrosis and burn wounds, resulting in significant mortality in immunocompromised individuals. This study reports a novel one-step biosynthesis of gold nanoparticles using phytocompound, hordenine (HD), as a reducing and capping agent. The synthesis of the anisotropic hordenine-fabricated gold nanoparticles (HD-AuNPs) with an average particle size of 136.87 nm was achieved within 12 h of incubation at room temperature. Both HD and HD-AuNPs exhibited significant antibiofilm activity against P. aeruginosa PAO1, although greater biofilm inhibition was observed for the nanoparticles as compared to hordenine alone. In the microtitre plate assay and tube method, the nanoparticles significantly inhibited the biofilm formation by 73.69 and 78.41%, respectively. The exopolysaccharide production by the test pathogen was arrested by 68.46% on treatment with the nanoparticles. Further, the effect of HD and HD-AuNPs on the biofilm architecture of P. aeruginosa was revealed by light and confocal laser-scanning microscopy micrographs. The overall results of this study suggested the synergistic antibiofilm effect of AuNPs and HD for the treatment of chronic bacterial infections caused by biofilms forming pathogens.