Hesperidin methyl chalcone, a citrus flavonoid, inhibits Aeromonas hydrophila infection mediated by quorum sensing.

Plant-derived phytocompounds are effective in treating a variety of ailments and disorders, the most common of which are bacterial infections in humans, which are a major public health concern. Flavonoids, one of the groups of phytocompounds, are known to have significant antimicrobial and anti-infective properties. Hence, the current study investigates the efficacy of the citrus flavonoid hesperidin methylchalcone (HMC) in addressing this major issue. The results of this study indicate that the anti-quorum sensing (anti-QS) action against Aeromonas hydrophila infections is exhibited with a decrease in biofilm development and virulence factors production through in vitro and in silico analyses. In addition, the qPCR findings indicate that HMC has antivirulence action on A. hydrophila by reducing the expression of QS-related virulence genes, including ahyR, ahyB, ahh1, aerA, and lip. Interestingly, HMC significantly rescued the A. hydrophila-infected zebrafish by reducing the internal colonization, demonstrating the in vivo anti-infective potential of HMC against A. hydrophila infection. Based on these results, this study recommends that HMC could be employed as a possible therapeutic agent to treat A. hydrophila-related infections in humans.

Tocopherol-assisted magnetic Ag-Fe3O4-TiO2 nanocomposite for photocatalytic bacterial-inactivation with elucidation of mechanism and its hazardous level assessment with zebrafish model.

In recent years, many endeavours have been prompted with photocatalytic nanomaterials by the need to eradicate pathogenic microorganisms from water bodies. Herein, a tocopherol-assisted Ag-Fe3O4-TiO2 nanocomposite (TAFTN) was synthesized for photocatalytic bacterial inactivation. The prepared TAFTN became active under sunlight due to its narrowed bandgap, inactivating the bacterial contaminants via photo-induced ROS stress. The ROS radicals destroy bacteria by creating oxidative stress, which damages the cell membrane and cellular components such as nucleic acids and proteins. For the first time, the nano-LC-MS/MS-based quantitative proteomics reveals that the disrupted proteins are involved in a variety of cellular functions; the most of these are involved in the metabolic pathway, eventually leading to bacterial death during TAFTN-photocatalysis under sunlight. Furthermore, the toxicity analysis confirmed that the inactivated bacteria seemed to have no detrimental impact on zebrafish model, showing that the disinfected water via TAFTN-photocatalysis is enormously safe. Furthermore, the TAFTN-photocatalysis successfully killed the bacterial cells in natural seawater, indicating the consistent photocatalytic efficacy when recycled repeatedly. The results of this work demonstrate that the produced nanocomposite might be a powerful recyclable and sunlight-active photocatalyst for environmental water treatment.

Pyrogallol downregulates the expression of virulence-associated proteins in Acinetobacter baumannii and showing anti-infection activity by improving non-specific immune response in zebrafish model.

Acinetobacter baumannii, a virulent uropathogen with widespread antibiotic resistance, has arisen as a critical scientific challenge, necessitating the development of innovative therapeutic agents. This is the first study reveal the proteomic changes in A. baumannii upon pyrogallol treatment for understanding the mechanisms using nano-LC-MS/MS-based quantitative proteomics and qPCR analysis. The obtained results found that pyrogallol treatment dramatically downregulated the expression level of several key proteins such as GroEL, DnaK, ClpB, SodB, KatE, Bap, CsuA/B, PgaA, PgaC, BfmR, OmpA, and SecA in A. baumannii, which are involved in chaperone-mediated oxidative stress responses, antioxidant defence system, biofilm formation, virulence enzyme production, bacterial adhesion, capsule formation, and antibiotic resistance. Accordingly, the pyrogallol dramatically enhanced the lifespan of A. baumannii-infected zebrafish by inhibiting bacterial colonization, demonstrating the anti-infective potential of pyrogallol against A. baumannii. Further, the histopathological results also demonstrated the disease protection efficacy of pyrogallol against the pathognomonic sign of A. baumannii infection. In addition, the pyrogallol treatment effectively improved the immune parameters such as serum myeloperoxidase activity, leukocyte respiratory burst activity, and serum lysozyme activity in zebrafish against A. baumannii infection. Based on the results, the present study strongly proposes pyrogallol as a promising therapeutic agent for treating A. baumannii infection.

Phytosterols in Seaweeds: An Overview on Biosynthesis to Biomedical Applications.

Seaweed extracts are considered effective therapeutic alternatives to synthetic anticancer, antioxidant, and antimicrobial agents, owing to their availability, low cost, greater efficacy, eco-friendliness, and non-toxic nature. Since the bioactive constituents of seaweed, in particular, phytosterols, possess plenty of medicinal benefits over other conventional pharmaceutical agents, they have been extensively evaluated for many years. Fortunately, recent advances in phytosterol-based research have begun to unravel the evidence concerning these important processes and to endow the field with the understanding and identification of the potential contributions of seaweed-steroidal molecules that can be used as chemotherapeutic drugs. Despite the myriad of research interests in phytosterols, there is an immense need to fill the void with an up-to-date literature survey elucidating their biosynthesis, pharmacological effects, and other biomedical applications. Hence, in the present review, we summarize studies dealing with several types of seaweed to provide a comprehensive overview of the structural determination of several phytosterol molecules, their properties, biosynthetic pathways, and mechanisms of action, along with their health benefits, which could significantly contribute to the development of novel drugs and functional foods.

Inhibition of biofilm formation and quorum sensing mediated virulence in Pseudomonas aeruginosa by marine sponge symbiont Brevibacterium casei strain Alu 1.

The alternative antimicrobial strategies that mitigate the threat of antibiotic resistance is the quorum-sensing inhibition (QSI) mechanism, which targets autoinducer dependent virulence gene expression in bacterial pathogens. N-acyl homoserine lactone (AHL) acts as a key regulator in the production of virulence factors and biofilm formation in Pseudomonas aeruginosa PAO1 and violacein pigment production in Chromobacterium violaceum ATCC 12472. In the present study, the marine sponge Haliclona fibulata symbiont Brevibacterium casei strain Alu 1 showed potential QSI activity in a concentration-dependent manner (0.5-2% v/v) against the N-acyl homoserine lactone (AHL)-mediated violacein production in C. violaceum (75-95%), and biofilm formation (53-96%), protease (27-82%), pyocyanin (82-95%) and pyoverdin (29-38%) productions in P. aeruginosa. Further, the microscopic analyses validated the antibiofilm activity of the cell-free culture supernatant (CFCS) of B. casei against P. aeruginosa. Subsequently, the biofilm and pyoverdin inhibitory efficacy of the ethyl acetate extract of B. casei CFCS was assessed against P. aeruginosa. Further, the gas chromatography-mass spectrometry (GC-MS) analysis revealed the presence of variety of components in which diethyl phthalate was found to be a major active component. This phthalate ester, known as diethyl ester of phthalic acid, could act as a potential therapeutic agent for preventing bacterial biofilm and virulence associated infectious diseases.

Gene expressing analysis indicates the role of Pyrogallol as a novel antibiofilm and antivirulence agent against Acinetobacter baumannii.

Acinetobacter baumannii has emerged worldwide as a leading cause of hospital-acquired infections. Although A. baumannii was initially regarded to as a low-grade pathogen, evidence has been accumulated suggesting that A. baumannii infections are associated with increased mortality in critically ill patients. Here, we describe the efficacy of pyrogallol, a polyphenolic organic compound found in the galls and barks of various trees, which shows anti-biofilm and anti-virulence potential against A. baumannii. Pyrogallol shows concentration-based biofilm inhibition, as evidenced through light and confocal laser scanning microscopic analysis. The other virulence factors are protease, swarming motility, and extracellular polymeric substances that are also inhibited by pyrogallol. Through real-time PCR, it was found that pyrogallol downregulates expression of the biofilm and virulence-related ompA, bap, csuA/B, katE, pgaA, and pgaC genes. Furthermore, pyrogallol moderately inhibited the mature biofilms of A. baumannii in a concentration-dependent manner (5, 10, and 20 µg/ml). The present study reports that the anti-biofilm and anti-virulence potential of pyrogallol disrupts the biofilm formation, adherence of cells, and cell-to-cell signaling mechanism of A. baumannii. Thus, pyrogallol is a promising therapeutic agent for A. baumannii-related infections.

Inhibitory Effect of Morin Against Candida albicans Pathogenicity and Virulence Factor Production: An in vitro and in vivo Approaches.

Candida albicans is considered an exclusive etiologic agent of candidiasis, a very common fungal infection in human. The expression of virulence factors contributes highly to the pathogenicity of C. albicans. These factors include biofilm formation, yeast-to-hyphal transition, adhesins, aspartyl proteases, and phospholipases secretion. Moreover, resistance development is a critical issue for the therapeutic failure of antifungal agents against systemic candidiasis. To circumvent resistance development, the present study investigated the virulence targeted therapeutic activity of the phyto-bioactive compound morin against C. albicans. Morin is a natural compound commonly found in medicinal plants and widely used in the pharmaceutical and cosmetic products/industries. The present study explicated the significant inhibitory potential of morin against biofilm formation and other virulence factors' production, such as yeast-hyphal formation, phospholipase, and exopolymeric substances, in C. albicans. Further, qPCR analysis confirmed the downregulation of biofilm and virlence-related genes in C. albicans upon morin treatment, which is in correspondence with the in vitro bioassays. Further, the docking analysis revealed that morin shows strong affinity with Hwp-1 protein, which regulates the expression of biofilm and hyphal formation in C. albicans and, thereby, abolishes fungal pathogenicity. Moreover, the anti-infective potential of morin against C. albicans-associated systemic candidiasis is confirmed through an in vivo approach using biomedical model organism zebrafish (Danio rerio). The outcomes of the in vivo study demonstrate that the morin treatment effectively rescues animals from C. albicans infections and extends their survival rate by inhibiting the internal colonization of C. albicans. Histopathology analysis revealed extensive candidiasis-related pathognomonic changes in the gills, intestine, and kidney of animals infected with C. albicans, while no extensive abnormalities were observed in morin-treated animals. The results evidenced that morin has the ability to protect against the pathognomonic effect and histopathological lesions caused by C. albicans infection in zebrafish. Thus, the present study suggests that the utilization of morin could act as a potent therapeutic medication for C. albicans instigated candidiasis.

Proteomic analysis deciphers the multi-targeting antivirulence activity of tannic acid in modulating the expression of MrpA, FlhD, UreR, HpmA and Nrp system in Proteus mirabilis.

In the present study, the multi-targeting antivirulence activity of tannic acid (TA) was explored against Proteus mirabilis through MS-based proteomic approach. The in vitro biofilm biomass quantification assay and microscopic analysis demonstrated the antibiofilm activity of TA against P. mirabilis in which, minimum biofilm inhibitory concentration (MBIC) of TA was found to be 200 µg/mL concentration. Moreover, the nanoscale liquid chromatography coupled to tandem mass spectrometry (nano LC-MS/MS) analysis revealed that TA (at MBIC) differentially regulated the proteins involved in fimbrial adhesion, flagellar motility, iron acquisition, Fe-S cluster assembly, heat shock response, virulence enzymes, and toxin secretion. Further, the transcriptomic analysis validated the outcomes of proteomic analysis in which, the expression level of virulence genes responsible for MR/P fimbrial adhesion (mrpA), flagellar transcriptional activation (flhD), biosynthesis of urease (ureR), hemolysin (hpmA), non-ribosomal peptide siderophore system (Nrp), oxidative stress responsible enzymes and fitness factors proteins were down-regulated in TA exposed P. mirabilis. These observations were also in correspondence with the in vitro bioassays. Thus, this study reports the feasibility of TA to act as a promising therapeutic agent against multifactorial P. mirabilis infections.

Metal sensing-carbon dots loaded TiO2-nanocomposite for photocatalytic bacterial deactivation and application in aquaculture.

Nowadays, bioactive nanomaterials have been attracted the researcher's enthusiasm in various fields. Herein, Diplocyclos palmatus leaf extract-derived green-fluorescence carbon dots (DP-CDs) were prepared using the hydrothermal method. Due to the strong fluorescence stability, the prepared DP-CDs were coated on filter-paper to make a fluorometric sensor-strip for Fe3+ detection. After, a bandgap-narrowed DP-CDs/TiO2 nanocomposite (DCTN) was prepared using the methanolic extract of D. palmatus. The prepared DCTN exhibited improved photocatalytic bacterial deactivation under sunlight irradiation. The DCTN-photocatalysis slaughtered V. harveyi cells by the production of reactive oxygen species, which prompting oxidative stress, damaging the cell membrane and cellular constituents. These results suggest the plausible mode of bactericidal action of DCTN-photocatalysis under sunlight. Further, the DCTN has shown potent anti-biofilm activity against V. harveyi, and thereby, DCTN extended the survival of V. harveyi-infected shrimps during the in vivo trial with Litopenaeus vannamei. Notably, this is the first report for the disinfection of V. harveyi-mediated acute-hepatopancreatic necrosis disease (AHPND) using nanocomposite. The reduced internal-colonization of V. harveyi on the hepatopancreas as well as the rescue action of the pathognomonic effect in the experimental animals demonstrated the anti-infection potential of DCTN against V. harveyi-mediated AHPND in aquaculture.

2-Hydroxy-4-methoxybenzaldehyde from Hemidesmus indicus is antagonistic to Staphylococcus epidermidis biofilm formation.

Staphylococcus epidermidis (SE) is an opportunistic nosocomial pathogen that accounts for recalcitrant device-related infections worldwide. Owing to the growing interest in plants and their secondary metabolites targeting bacterial adhesion, this study was intended to uncover the anti-biofilm potential of Hemidesmus indicus and its major constituent 2-hydroxy-4-methoxybenzaldehyde (HMB) against SE. The minimum biofilm inhibitory concentration (MBIC) of H. indicus root extract and HMB were found to be 500 and 250 µg ml-1, respectively. The results of time-dependent biofilm inhibition and mature biofilm disruption assays confirmed that HMB targets initial cell adhesion. Furthermore, interference by HMB in the expression of adhesin genes (icaA, aap and bhp) and biofilm components was associated with an increased susceptibility of SE to oxidative stress and antibiotics. To conclude, this study reports for the first time HMB as a potential drug against SE biofilms.

Attenuation of Proteus mirabilis colonization and swarming motility on indwelling urinary catheter by antibiofilm impregnation: An in vitro study.

Proteus mirabilis is one of the important etiologic agents of urinary tract infections (UTI), which complicates the long-term urinary catheterization process in clinical settings. Owing to its crystalline biofilm forming ability and flagellar motility, elimination of P. mirabilis from urinary system becomes very difficult. Thus, the present study is focused to prepare antibiofilm-impregnated Silicone Foley Catheter (SFC) to prevent P. mirabilis instigated UTIs. Through solvent swelling method, the antibiofilm compounds such as linalool (LIN) and 2-hydroxy-4-methoxy benzaldehyde (HMB) were successfully infused into SFCs. Surface topography was studied using AFM analysis, which unveiled the unmodified surface roughness of normal and antibiofilm-impregnated SFCs. In addition, UV-spectrometric and FT-IR analyses revealed good impregnation efficacy and prolonged stability of antibiofilm compounds. Further, in vitro biofilm biomass quantification assay exhibited a maximum of 87 % and 84 % crystalline biofilm inhibition in LIN (350 µg/cm3) and HMB (120 µg/cm3) impregnated SFCs, respectively against P. mirabilis in artificial urine medium. Also, the LIN & HMB-impregnated SFCs demonstrated long-term crystalline biofilm inhibitory activity for more than 30 days, which is ascribed to the sustained release of the compounds. Furthermore, the results of swarming motility analysis revealed the efficacy of antibiofilm-impregnated catheters to mitigate the migration of pathogens over them. Thus, antibiofilm-impregnated catheter is proposed to act as a suitable strategy for reducing P. mirabilis infections and associated complications in long-term urinary catheter users.

Protective effect of neglected plant Diplocyclos palmatus on quorum sensing mediated infection of Serratia marcescens and UV-A induced photoaging in model Caenorhabditis elegans.

Plants are considered to be a leading source for possible human therapeutic agents. This holistic study has investigated the anti-quorum sensing (anti-QS), anti-infection, antioxidant and anti-photoaging properties of neglected plant Diplocyclos palmatus. The results showed that D. palmatus methanolic leaf extract (DPME) effectively inhibited the quorum sensing (QS) regulated virulence factor production as well as biofilm formation in Serratia marcescens. The transcriptomic analysis revealed that DPME significantly downed the expression of QS-regulated genes such as fimA, fimC, flhC, bsmB, pigP and shlA in S. marcescens, which supports the outcome of in vitro bioassays. Further, the docking study revealed that the presence of active compounds, namely tocopherols and phytol, DPME exhibited its anti-QS activity against S. marcescens. In addition, DPME treatment extended the lifespan of S. marcescens infected C. elegans by the action of dropping the internal accumulation. Further, qPCR analysis clearly revealed that DPME treatment significantly up-regulated the expression of the lifespan-related gene (daf-16) and immune-related genes (clec-60, clec-87, lys-7 and bec-1) in S. marcescens infected C.elegans. On the other hand, DPME extensively reduced the UV-A induced ROS stress, thereby, extended the lifespan in UV-A photoaged C. elegans. Further, the qPCR analysis also confirmed the up-regulation of daf-16, clec-60, clec-87 and col-19 genes which advocated the improvement of the lifespan, healthspan and collagen production in UV-A photoaged C. elegans. Further bioassays evidenced that that the lifespan extension of photoaged C. elegans was accomplished by the actions of antioxidants such as tocopherols and phytol in DPME.

Virulence targeted inhibitory effect of linalool against the exclusive uropathogen Proteus mirabilis.

Proteus mirabilis is one of the leading causes of catheter-associated UTIs (CAUTI) in individuals with prolonged urinary catheterization. Since, biofilm assisted antibiotic resistance is reported to complicate the treatment strategies of P. mirabilis infections, the present study was aimed to attenuate biofilm and virulence factor production in P. mirabilis. Linalool is a naturally occurring monoterpene alcohol found in a wide range of flowers and spice plants and has many biological applications. In this study, linalool exhibited concentration dependent anti-biofilm activity against crystalline biofilm of P. mirabilis through reduced production of the virulence enzyme urease that raises the urinary pH and drives the formation of crystals (struvite) in the biofilm. The results of q-PCR analysis unveiled the down regulation of biofilm/virulence associated genes upon linalool treatment, which was in correspondence with the in vitro bioassays. Thus, this study reports the feasibility of linalool acting as a promising anti-biofilm agent against P. mirabilis mediated CAUTI.

Explication of the Potential of 2-Hydroxy-4-Methoxybenzaldehyde in Hampering Uropathogenic Proteus mirabilis Crystalline Biofilm and Virulence.

Proteus mirabilis is an important etiological agent of catheter-associated urinary tract infections (CAUTIs) owing to its efficient crystalline biofilm formation and virulence enzyme production. Hence, the present study explicated the antibiofilm and antivirulence efficacies of 2-hydroxy-4-methoxybenzaldehyde (HMB) against P. mirabilis in a non-bactericidal manner. HMB showed concentration-dependent biofilm inhibition, which was also evinced in light, confocal, and scanning electron microscopic (SEM) analyses. The other virulence factors such as urease, hemolysin, siderophores, and extracellular polymeric substances production as well as swimming and swarming motility were also inhibited by HMB treatment. Further, HMB treatment effectively reduced the struvite/apatite production as well as crystalline biofilm formation by P. mirabilis. Furthermore, the results of gene expression analysis unveiled the ability of HMB to impair the expression level of virulence genes such as flhB, flhD, rsbA, speA, ureR, hpmA, and hpmB, which was found to be in correlation with the results of in vitro bioassays. Additionally, the cytotoxicity analysis divulged the innocuous characteristic of HMB against human embryonic kidney cells. Thus, the present study reports the potency of HMB to act as a promising therapeutic remedy for P. mirabilis-instigated CAUTIs.

Biogenic synthesis of silver nanoparticles using Piper betle aqueous extract and evaluation of its anti-quorum sensing and antibiofilm potential against uropathogens with cytotoxic effects: an in vitro and in vivo approach.

Urinary tract infections are the utmost common bacterial infections caused by Proteus mirabilis, Pseudomonas aeruginosa, Escherichia coli, and Serratia marcescens. These uropathogens resist the action of several antibiotics due to their ability to form biofilms. Most of these bacterial pathogens use the quorum sensing (QS) machinery to co-ordinate their cells and regulate several virulence factors and biofilm formation. On the other hand, the anti-quorum sensing (anti-QS) and antibiofilm potential of silver nanoparticles have been well reported against certain bacterial pathogens, but to the best of our knowledge, no report is available against the pathogenicity of uropathogens in particular S. marcescens and P. mirabilis. Therefore, the present study is primarily focused on the anti-QS and antibiofilm potential of Piper betle-based synthesized silver nanoparticles (PbAgNPs) against S. marcescens and P. mirabilis. Initially, the silver nanoparticles were synthesized by the aqueous extract of P. betle and characterized by UV-absorbance spectroscopy, XRD, FT-IR, SEM, TEM, and DLS. The synthesized silver nanoparticles were assessed for their anti-QS activity and the obtained results revealed that the PbAgNPs inhibited the QS-mediated virulence factors such as prodigiosin, protease, biofilm formation, exopolysaccharides and hydrophobicity productions in uropathogens. The gene expression analysis divulged the downregulation of fimA, fimC, flhD, and bsmB genes in S. marcescens and flhB, flhD, and rsbA genes in P. mirabilis, respectively. The in vivo Caenorhabditis elegans assays revealed the non-toxic and anti-adherence efficiency of PbAgNPs. Furthermore, the non-toxic effect of PbAgNPs was also confirmed through peripheral blood mononuclear cells and normal lung epithelial cells. Therefore, the contemporary study demonstrates the use of PbAgNPs as a possible alternative toward conventional antibiotics in controlling QS and biofilm-related uropathogen infections.