1,25 dihydroxyvitamin D3-mediated effects on bovine innate immunity and on biofilm-forming Staphylococcus spp. isolated from cattle with mastitis.

Mastitis is one the most widespread and serious diseases in dairy cattle. Recurrent and chronic infections are often attributable to certain pathogenicity mechanisms in mastitis-causing pathogens such as Staphylococcus spp. These include growing in biofilm and invading cells, both of which make it possible to resist or evade antimicrobial therapies and the host's immune system. This study tested the effects of active vitamin D3 (i.e., calcitriol or 1,25-dihydroxyvitamin D3) on the internalization and phagocytosis of biofilm-forming Staphylococcus spp. isolated from animals with mastitis. Two established bovine cell lines were used: MAC-T (mammary epithelial cells) and BoMac (macrophages). Calcitriol (0-200 nM) did not affect the viability of MAC-T cells nor that of BoMac cells after 24 and 72 h. Concentrations of 0-100 mM for 24 h upregulated the expression of 24-hydroxylase in MAC-T cells, but did not alter that of VDR. Pre-treatment of the cells with calcitriol for 24 h decreased the internalization of S. aureus V329 into MAC-T cells (0-100 nM), and stimulated the phagocytosis of the same strain and of S. xylosus 4913 (0-10 nM). Calcitriol and two conditioned media, obtained by treating the cells with 25-200 nM of the metabolite for 24 h, were also assessed in terms of their antimicrobial and antibiofilm activity. Neither calcitriol by itself nor the conditioned media affected staphylococcal growth or biofilm formation (0-200 nM for 12 and 24 h, respectively). In contrast, the conditioned media (0-100 nM for 24 h) decreased the biomass of preformed non-aureus staphylococcal biofilms and killed the bacteria within them, without affecting metabolic activity. These effects may be mediated by reactive oxygen species and proteins with antimicrobial and/or antibiofilm activity. In short, calcitriol could make pathogens more accessible to antimicrobial therapies and enhance bacterial clearance by professional phagocytes. Moreover, it may modulate the host's endogenous defenses in the bovine udder and help combat preformed non-aureus staphylococcal biofilms (S. chromogenes 40, S. xylosus 4913, and/or S. haemolyticus 6). The findings confirm calcitriol's potential as an adjuvant to prevent and/or treat intramammary infections caused by Staphylococcus spp., which would in turn contribute to reducing antibiotic use on dairy farms.

Alhagi maurorum extract modulates quorum sensing genes and biofilm formation in Proteus mirabilis.

Proteus mirabilis (P. mirabilis) is a frequent cause of catheter-associated urinary tract infections. This study aims to investigate the anti-infective effect of Alhagi maurorum extract (AME), the traditional medicinal plant in the middle east, on the biofilm-forming P. mirabilis isolates. Hydroalcoholic extract and oil of A. maurorum were characterized by HPLC and GC-MS. The antiproliferative, anti-biofilm, and bactericidal activity of AME at various concentrations were assessed by turbidity, crystal violet binding, and agar well diffusion assays, respectively. The AME's effect on adhesion and quorum sensing (QS) were investigated by in vitro adhesion assay on cell culture and agar overlay assay using Janthinobacterium lividum (ATCC 12472) as a biosensor strain. In addition, the expression level of selected genes involved in QS and biofilm regulation were determined by quantitative Real-Time PCR. Furthermore, the bladder phantom model was created to evaluate the assays and investigate the catheter's calcium deposition. The most effective chemical compounds found in AME were tamarixetin, quercetin, and trans-anethole. Although AME did not inhibit swarming motility, it reduced biofilm production and exerted a concentration-dependent anti-adhesive and anti-QS activity against P. mirabilis. AME also downregulated the expression level of selected genes involved in biofilm formation and QS. This study showed that AME as a natural compound reduced biofilm formation of P. mirabilis by targeting virulence factor genes, quorum sensing, and other strategies that include preventing the adhesion of P. mirabilis to the cells. The results suggest that A. maurorum extract might have the potential to be considered for preventing UTIs caused by P. mirabilis.

Synergistic Inhibitory Effect of Polymyxin B in Combination with Ceftazidime against Robust Biofilm Formed by Acinetobacter baumannii with Genetic Deficiency in AbaI/AbaR Quorum Sensing.

Carbapenem resistance of Acinetobacter baumannii poses challenges to public health. Biofilm contributes to the persistence of A. baumannii cells. This study was designed to investigate the genetic relationships among carbapenem resistance, polymyxin resistance, multidrug resistance, biofilm formation, and surface-associated motility and evaluate the antibiofilm effect of polymyxin in combination with other antibiotics. A total of 103 clinical A. baumannii strains were used to determine antibiotic susceptibility, biofilm formation capacity, and motility. Enterobacterial repetitive intergenic consensus (ERIC)-PCR fingerprinting was used to determine the genetic variation among strains. The distribution of 17 genes related to the resistance-nodulation-cell division (RND)-type efflux, autoinducer-receptor (AbaI/AbaR) quorum sensing, oxacillinases (OXA)-23, and insertion sequence of ISAba1 element was investigated. The representative strains were chosen to evaluate the gene transcription and the antibiofilm activity by polymyxin B (PB) in combination with merapenem, levofloxacin, and ceftazidime, respectively. ERIC-PCR-dependent fingerprints were found to be associated with carbapenem resistance and multidrug resistance. The presence of blaOXA-23 was found to correlate with genes involved in ISAba1 insertion, AbaI/AbaR quorum sensing, and AdeABC efflux. Carbapenem resistance was observed to be negatively correlated with biofilm formation and positively correlated with motility. PB in combination with ceftazidime displayed a synergistic antibiofilm effect against robust biofilm formed by an A. baumannii strain with deficiency in AbaI/AbaR quorum sensing. Our results not only clarify the genetic correlation among carbapenem resistance, biofilm formation, and pathogenicity in a certain level but also provide a theoretical basis for clinical applications of polymyxin-based combination of antibiotics in antibiofilm therapy. IMPORTANCE Deeper explorations of molecular correlation among antibiotic resistance, biofilm formation, and pathogenicity could provide novel insights that would facilitate the development of therapeutics and prevention against A. baumannii biofilm-related infections. The major finding that polymyxin B in combination with ceftazidime displayed a synergistic antibiofilm effect against robust biofilm formed by an A. baumannii strain with genetic deficiency in AbaI/AbaR quorum sensing further provides a theoretical basis for clinical applications of antibiotics in combination with quorum quenching in antibiofilm therapy.

Sustainable phyto-fabrication of silver nanoparticles using Gmelina arborea exhibit antimicrobial and biofilm inhibition activity.

Increase in bacterial resistance to commonly used antibiotics is a major public health concern generating interest in novel antibacterial treatments. Aim of this scientific endeavor was to find an alternative efficient antibacterial agent from non-conventional plant source for human health applications. We used an eco-friendly approach for phyto-fabrication of silver nanoparticles (AgNPs) by utilizing logging residue from timber trees Gmelina arborea (GA). GC-MS analysis of leaves, barks, flowers, fruits, and roots was conducted to determine the bioactive compounds. Biosynthesis, morphological and structural characterization of GA-AgNPs were undertaken by UV-Vis spectroscopy, scanning electron microscopy (SEM), energy-dispersive spectroscopy (EDX), transmission electron microscopy (TEM), Fourier transform infrared spectroscopy (FTIR) and X-ray diffractometer (XRD). GA-AgNPs were evaluated for antibacterial, antibiofilm, antioxidant, wound healing properties and their toxicity studies were carried out. Results identified the presence of terpenoids, sterols, aliphatic alcohols, aldehydes, and flavonoids in leaves, making leaf extract the ideal choice for phyto-fabrication of silver nanoparticles. The synthesis of GA-AgNPs was confirmed by dark brown colored colloidal solution and spectral absorption peak at 420 nm. Spherical, uniformly dispersed, crystalline GA-AgNPs were 34-40 nm in diameter and stable in solutions at room temperature. Functional groups attributed to the presence of flavonoids, terpenoids, and phenols that acted as reducing and capping agents. Antibacterial potency was confirmed against pathogenic bacteria Bacillus cereus, Escherichia coli, Pseudomonas aeruginosa, and Staphylococcus aureus by disc diffusion assay, MIC and MBC assay, biofilm inhibition assay, electron-microscopy, cell staining and colony counting techniques. The results from zone of inhibition, number of ruptured cells and dead-cell-count analysis confirmed that GA-AgNPs were more effective than GA-extract and their bacteria inhibition activity level increased further when loaded on hydrogel as GA-AgNPs-PF127, making it a novel distinguishing feature. Antioxidant activity was confirmed by the free radical scavenging assays (DPPH and ABTS). Wound healing potential was confirmed by cell scratch assay in human dermal fibroblast cell lines. Cell-proliferation study in human chang liver cell lines and optical microscopic observations confirmed non-toxicity of GA-AgNPs at low doses. Our study concluded that biosynthesized GA-AgNPs had enhanced antibacterial, antibiofilm, antioxidant, and wound healing properties.

Histological assessment, anti-quorum sensing, and anti-biofilm activities of Dioon spinulosum extract: in vitro and in vivo approach.

Pseudomonas aeruginosa is an opportunistic bacterium causing several health problems and having many virulence factors like biofilm formation on different surfaces. There is a significant need to develop new antimicrobials due to the spreading resistance to the commonly used antibiotics, partly attributed to biofilm formation. Consequently, this study aimed to investigate the anti-biofilm and anti-quorum sensing activities of Dioon spinulosum, Dyer Ex Eichler extract (DSE), against Pseudomonas aeruginosa clinical isolates. DSE exhibited a reduction in the biofilm formation by P. aeruginosa isolates both in vitro and in vivo rat models. It also resulted in a decrease in cell surface hydrophobicity and exopolysaccharide quantity of P. aeruginosa isolates. Both bright field and scanning electron microscopes provided evidence for the inhibiting ability of DSE on biofilm formation. Moreover, it reduced violacein production by Chromobacterium violaceum (ATCC 12,472). It decreased the relative expression of 4 quorum sensing genes (lasI, lasR, rhlI, rhlR) and the biofilm gene (ndvB) using qRT-PCR. Furthermore, DSE presented a cytotoxic activity with IC50 of 4.36 ± 0.52 µg/ml against human skin fibroblast cell lines. For the first time, this study reports that DSE is a promising resource of anti-biofilm and anti-quorum sensing agents.

Regulatory network controls microbial biofilm development, with Candida albicans as a representative: from adhesion to dispersal.

Microorganisms mainly exist in the form of biofilm in nature. Biofilm can contaminate food and drinking water system, as well as cause chronic wound infections, thereby posing a potential threat to public health safety. In the last two decades, researchers have made efforts to investigate the genetic contributors control different stages of biofilm development (adherence, initiation, maturation, and dispersal). As an opportunistic pathogen, C. albicans causes severe superficial or systemic infections with high morbidity and mortality under conditions of immune dysfunction. It has been reported that 80% of C. albicans infections are directly or indirectly associated with biofilm formation on host or abiotic surfaces including indwelling medical devices, resulting in high morbidity and mortality. Significantly, the outcome of C. albicans biofilm development includes enhanced invasion, exacerbated inflammatory responses and intrinsic resistance to antimicrobial chemotherapy. Thus, this review aimed at providing a comprehensive overview of the regulatory network controls microbial biofilm development, with C. albicans as a representative, served as reference for therapeutic targets.

Evaluating Metabolic Pathways and Biofilm Formation in Stenotrophomonas maltophilia.

Stenotrophomonas maltophilia has recently arisen as a prominent nosocomial pathogen because of its high antimicrobial resistance and ability to cause chronic respiratory infections. Often the infections are worsened by biofilm formation which enhances antibiotic tolerance. We have previously found that mutation of the gpmA gene, encoding the glycolytic enzyme phosphoglycerate mutase, impacts the formation of this biofilm on biotic and abiotic surfaces at early time points. This finding, indicating an association between carbon source and biofilm formation, led us to hypothesize that metabolism would influence S. maltophilia biofilm formation and planktonic growth. In the present study, we tested the impact of various growth substrates on biofilm levels and growth kinetics to determine metabolic requirements for these processes. We found that S. maltophilia wild type preferred amino acids versus glucose for planktonic and biofilm growth and that gpmA deletion inhibited growth in amino acids. Furthermore, supplementation of the ΔgpmA strain by glucose or ribose phenotypically complemented growth defects. These results suggest that S. maltophilia shuttles amino acid carbon through gluconeogenesis to an undefined metabolic pathway supporting planktonic and biofilm growth. Further evaluation of these metabolic pathways might reveal novel metabolic activities of this pathogen. IMPORTANCE Stenotrophomonas maltophilia is a prominent opportunistic pathogen that often forms biofilms during infection. However, the molecular mechanisms of virulence and biofilm formation are poorly understood. The glycolytic enzyme phosphoglycerate mutase appears to play a role in biofilm formation, and we used a mutant in its gene (gpmA) to probe the metabolic circuitry potentially involved in biofilm development. The results of our study indicate that S. maltophilia displays unique metabolic activities, which could be exploited for inhibiting growth and biofilm formation of this pathogen.

UPLC-ESI-MS/MS profiling of the underground parts of common Iris species in relation to their anti-virulence activities against Staphylococcusaureus.

ETHNOPHARMACOLOGICAL RELEVANCE: The use of plant extracts and their phytochemicals as candidates for targeting the microbial resistance inhibition is increasingly focused in last decades. In Mongolian traditional medicine, Irises were long used for the treatment of bacterial infections. Irises have been used since the Ancient Egyptians. AIM OF THE STUDY: Chemical composition and virulence inhibition potential of both polar (PF) and non-polar fractions (NPF) of three common Iris species (I. confusa, I. pseudacorus and I. germanica) were explored. MATERIAL AND METHODS: Secondary metabolites profiling was characterized by the UPLC-HRMS/MS technique. Multi-variate data analysis was performed using Metaboanalyst 3.0. Anti-virulence inhibitory activity was evaluated via anti-haemolytic assay and Quantitative biofilm inhibition assay. RESULTS: I. pseudacorus PF exhibited the most potent effect against S. aureus haemolytic activity. All the tested fractions from all species, except I. pseudacorus NPF, have no significant inhibition on the biofilm formation of methicillin resistant and sensitive (MRSA and MSSA) S. aureus. I. pseudacorus NPF showed potent biofilm inhibitory potential of 71.4 and 85.8% against biofilm formation of MRSA and MSSA, respectively. Metabolite profiling of the investigated species revealed ninety and forty-five metabolites detected in the PFs and NPFs, respectively. Nigricin-type, tectorigenin-type isoflavonids and xanthones allowed the discrimination of I. pseudacorus PF from the other species, highlighting the importance of those metabolites in exerting its promising activity. On the other hand, triterpene acids, iridals, triacylglycerols and ceramides represented the metabolites detected in highest abundance in I. pseudacorus NPF. CONCLUSIONS: This is the sole map represents the secondary metabolites profiling of the PFs and NPFs of common Iris species correlating them with the potent explored Staphylococcus aureus anti-virulence activity.

Bioactive triterpenoids from Solanum torvum fruits with antifungal, resistance modulatory and anti-biofilm formation activities against fluconazole-resistant candida albicans strains.

Vulvovaginal candidiasis (VVC) is the second most common vaginal infection that affects women of reproductive age. Its increased occurrence and associated treatment cost coupled to the rise in resistance of the causative pathogen to current antifungal therapies has necessitated the need for the discovery and development of novel effective antifungal agents for the treatment of the disease. We report in this study the anti-Candida albicans activity of Solanum torvum 70% ethanol fruit extract (STF), fractions and some isolated compounds against four (4) fluconazole-resistant strains of C. albicans. We further report on the effect of the isolated compounds on the antifungal activity of fluconazole and voriconazole in the resistant isolates as well as their inhibitory effect on C. albicans biofilm formation. STF was fractionated using n-hexane, chloroform (CHCl3) and ethyl acetate (EtOAc) to obtain four respective major fractions, which were then evaluated for anti-C. albicans activity using the microbroth dilution method. The whole extract and fractions recorded MICs that ranged from 0.25 to 16.00 mg/mL. From the most active fraction, STF- CHCl3 (MIC = 0.25-1.00 mg/mL), four (4) known compounds were isolated as Betulinic acid, 3-oxo-friedelan-20α-oic acid, Sitosterol-3-ß-D-glucopyranoside and Oleanolic acid. The compounds demonstrated considerably higher antifungal activity (0.016 to 0.512 mg/mL) than the extract and fractions and caused a concentration-dependent anti-biofilm formation activity. They also increased the sensitivity of the C. albicans isolates to fluconazole. This is the first report of 3-oxo-friedelan-20α-oic acid in the plant as well as the first report of betulinic acid, sitosterol-3-ß-D-glucopyranoside and oleanolic acid from the fruits of S. torvum. The present study has demonstrated the anti-C. albicans activity of the constituents of S. torvum ethanol fruit extract and also shown that the constituents possess anti-biofilm formation and resistance modulatory activities against fluconazole-resistant clinical C. albicans isolates.

Demonstration of Allium sativum Extract Inhibitory Effect on Biodeteriogenic Microbial Strain Growth, Biofilm Development, and Enzymatic and Organic Acid Production.

To the best of our knowledge, this is the first study demonstrating the efficiency of Allium sativum hydro-alcoholic extract (ASE) againstFigure growth, biofilm development, and soluble factor production of more than 200 biodeteriogenic microbial strains isolated from cultural heritage objects and buildings. The plant extract composition and antioxidant activities were determined spectrophotometrically and by HPLC-MS. The bioevaluation consisted of the qualitative (adapted diffusion method) and the quantitative evaluation of the inhibitory effect on planktonic growth (microdilution method), biofilm formation (violet crystal microtiter method), and production of microbial enzymes and organic acids. The garlic extract efficiency was correlated with microbial strain taxonomy and isolation source (the fungal strains isolated from paintings and paper and bacteria from wood, paper, and textiles were the most susceptible). The garlic extract contained thiosulfinate (307.66 ± 0.043 µM/g), flavonoids (64.33 ± 7.69 µg QE/g), and polyphenols (0.95 ± 0.011 mg GAE/g) as major compounds and demonstrated the highest efficiency against the Aspergillus versicolor (MIC 3.12-6.25 mg/mL), A. ochraceus (MIC: 3.12 mg/mL), Penicillium expansum (MIC 6.25-12.5 mg/mL), and A. niger (MIC 3.12-50 mg/mL) strains. The extract inhibited the adherence capacity (IIBG% 95.08-44.62%) and the production of cellulase, organic acids, and esterase. This eco-friendly solution shows promising potential for the conservation and safeguarding of tangible cultural heritage, successfully combating the biodeteriogenic microorganisms without undesirable side effects for the natural ecosystems.

Composition, Anti-MRSA Activity and Toxicity of Essential Oils from Cymbopogon Species.

Many of the essential oils obtained from medicinal plants possess proven antimicrobial activity and are suitable for medicinal purposes and applications in the food industry. The aim of the present work was the chemical analysis of 19 essential oils (EOs) from seven different Cymbopogon species (C. nardus, C. citratus, C winterianus, C. flexuosus, C. schoenanthus, C. martinii, C. giganteus). Five different chemotypes were established by GC/MS and TLC assay. The EOs, as well as some reference compounds, i.e., citronellol, geraniol and citral (neral + geranial), were also tested for their antimicrobial and antibiofilm activity against methicillin-resistant Staphylococcus aureus (MRSA) by the microdilution method and direct bioautography. The toxicity of EOs was evaluated by Danio rerio 'Zebrafish' model assay. All examined EOs showed moderate to high activity against MRSA, with the highest activity noted for C. flexuosus-lemongrass essential oil, both in microdilution and direct autobiography method. Significant difference in the toxicity of the examined EOs was also detected.

Serum levels of immunoglobulin and complement in UTI of patients caused by Proteus mirabilis and using AgNPs as antiswarming.

The use of plant extracts represents a promising approach for the synthesis of silver nanoparticles (AgNPs). This study reports the low-cost, green synthesis of AgNPs using the extract of clove and black seeds. The biosynthesized AgNPs were confirmed and characterized by analysis of the spectroscopy profile of the UV-visible spectrophotometer. The purpose of the present study is to evaluate the inhibitory effect concentration (MIC) of AgNPs, clove, and black cumin seed extracts on the growth and swarming of P. mirabilis. Clinical isolates of P. mirabilis were isolated from patients suffering from urinary tract infections. Thirteen types of antibiotics were used in the present study to detect their ability to inhibit P. mirabilis's resistance. Immunological findings included the determination of serum levels of IgG, IgM, IgA and complement protein C3 and C4. Results showed that IgG and IgA concentrations significantly increased (1311.13 ± 72.54 and 279 ± 21.31) respectively in UTI patients in comparison to the healthy control group which was 1089.88 ± 37.33 and 117.611 ± 4.19 respectively, While IgM concentrations were increased non significantly in UTI patients (153.331 ± 6.45) in comparison to healthy control (145.2 ± 13.49). Complement components C3 showed a significant increase in UTI patients with mean values of 125.95 ± 6.22 compared to the control group with mean values of 55.191 ± 9.64, while C4 showed statically non-significant among UTI patients in comparison with the control group (35.195 ± 2.34 and 34.371 ± 1.22) respectively.

Bio-assisted synthesis of bimetallic nanoparticles featuring antibacterial and photothermal properties for the removal of biofilms.

Biofilms are responsible for about considerable amounts of cases of bacterial infections in humans. They are considered a major threat to transplant and chronic wounds patients due to their highly resistant nature against antibacterial materials and due to the limited types of techniques that can be applied to remove them. Here we demonstrate a successful in-situ bio-assisted synthesis of dual functionality nanoparticles composed of Silver and Gold. This is done using a jellyfish-based scaffold, an antibacterial material as the templating host in the synthesis. We further explore the scaffold's antibacterial and photothermal properties against various gram-negative and positive model bacteria with and without photo-induced heating at the Near-IR regime. We show that when the scaffold is loaded with these bimetallic nanoparticles, it exhibits dual functionality: Its photothermal capabilities help to disrupt and remove bacterial colonies and mature biofilms, and its antibacterial properties prevent the regrowth of new biofilms.

Evaluation of antimicrobial peptide LL-37 for treatment of Staphylococcus aureus biofilm on titanium plate.

ABSTRACT: The antimicrobial peptide LL-37 belongs to the cathelicidin family and is one of the few human bactericidal peptides with potent antistaphylococcal activity. Staphylococcus aureus is one of the main infection bacteria in orthopedic implant therapy. Biofilm formation after bacterial infection brings more and more severe test for clinical antiinfection treatment.However, there are few studies on LL-37 in S. aureus infection of prosthesis. In this work, addition to research the antibacterial activity and the inhibitory effect on bacterial adhesion of LL-37, an in vitro model of S. aureus biofilm formation on titanium alloy surface was established to observe the inhibitory effect of LL-37.The results showed that LL-37 has a strong antibacterial effect on S. aureus in vitro, and the minimum inhibitory concentration (MIC) is about 0.62 µΜ. Moreover, LL-37 has significant impact on the adhesion of S. aureus when the concentration ≥0.16 µM and significant anti-staphylococcal biofilm effects on static biofilm models at the concentration of 0.31 to 10 µM. Additionally, LL-37 at 5 µM had a significant destructive effect on S. aureus biofilm (P < .05) that formed on the titanium alloy surface.This study further confirmed the role of LL-37 in the process of S. aureus infection, including antimicrobial activities, inhibition of bacterial adhesion, and inhibition of mature biofilm. LL-37 can significantly destroy the stable biofilm structure on the titanium alloy surface in vitro, which may provide a new way for refractory infection caused by S. aureus in titanium alloy prosthesis infection.

Therapeutic Effects of Inhibitor of ompA Expression against Carbapenem-Resistant Acinetobacter baumannii Strains.

The widespread of carbapenem-resistant Acinetobacter baumannii (CRAB) is of great concern in clinical settings worldwide. It is urgent to develop new therapeutic agents against this pathogen. This study aimed to evaluate the therapeutic potentials of compound 62520, which has been previously identified as an inhibitor of the ompA promoter activity of A. baumannii, against CRAB isolates, both in vitro and in vivo. Compound 62520 was found to inhibit the ompA expression and biofilm formation in A. baumannii ATCC 17978 at sub-inhibitory concentrations in a dose-dependent manner. These inhibitory properties were also observed in clinical CRAB isolates belonging to sequence type (ST) 191. Additionally, compound 62520 exhibited a bacteriostatic activity against clinical clonal complex (CC) 208 CRAB isolates, including ST191, and ESKAPE pathogens. This bacteriostatic activity was not different between STs of CRAB isolates. Bacterial clearance was observed in mice infected with bioimaging A. baumannii strain 24 h after treatment with compound 62520. Compound 62520 was shown to significantly increase the survival rates of both immunocompetent and neutropenic mice infected with A. baumannii ATCC 17978. This compound also increased the survival rates of mice infected with clinical CRAB isolate. These results suggest that compound 62520 is a promising scaffold to develop a novel therapeutic agent against CRAB infections.

Antimicrobial and antibiofilm potential of Curcuma longa Linn. Rhizome extract against biofilm producing Staphylococcus aureus and Pseudomonas aeruginosa isolates.

More than 65% of all human bacterial infection are associated with biofilm. Bacteria in such biofilms are 10 to 1000-fold more resistant to antibiotics than free living bacteria cells. Organisms such as S. aureus and P. aeruginosa are responsible for a significant number of biofilm related infections. In this study, we investigated the antimicrobial and anti-biofilm activity of C. longa L. rhizome extract against biofilm producing S. aureus and P. aeruginosa isolates. The results of MIC and MBC demonstrated promising antibacterial activity of the rhizome extract. TLC and column chromatography detected various curcuminoids while phytochemical analysis also reveals presence of number of bioactive compounds such as alkaloids, flavonoids, phenolics, terpenoids, etc. Micro titer plate assay indicated significant inhibition of biofilm formation in clinical isolates treated with turmeric extract. Thus, on basis of our results turmeric extracts can be considered as natural antibiofilm and antibacterial agent.

Antibacterial, Antibiofilm, and Efflux Pump Inhibitory Properties of the Crude Extract and Fractions from Acacia macrostachya Stem Bark.

Microbial infections remain a public health problem due to the upsurge of bacterial resistance. In this study, the antibacterial, antibiofilm, and efflux pump inhibitory activities of the stem bark of Acacia macrostachya, an indigenous African medicinal plant, were investigated. In traditional medicine, the plant is used in the treatment of microbial infections and inflammatory conditions. A crude methanol extract obtained by Soxhlet extraction was partitioned by column chromatography to obtain the petroleum ether, ethyl acetate, and methanol fractions. Antibacterial, efflux pump inhibition and antibiofilm formation activities were assessed by the high-throughput spot culture growth inhibition (HT-SPOTi), ethidium bromide accumulation, and the crystal violet retention assay, respectively. The minimum inhibitory concentrations (MICs) of the crude extract and major fractions ranged from 250 to ≥500 µg/mL. At a concentration of 3.9-250 µg/mL, all extracts demonstrated >80% inhibition of biofilm formation in S. aureus. In P. aeruginosa, the EtOAc fraction showed the highest antibiofilm activity (59-69%) while the pet-ether fraction was most active against E. coli biofilms (45-67%). Among the test samples, the crude extract, methanol, and ethyl acetate fractions showed remarkable efflux pump inhibition in S. aureus, E. coli, and P. aeruginosa. At ½ MIC, the methanol fraction demonstrated significant accumulation of EtBr in E. coli having superior efflux inhibition over the standard EPIs: chlorpromazine and verapamil. Tannins, flavonoids, triterpenoids, phytosterols, coumarins, and saponins were identified in preliminary phytochemical studies. Stigmasterol was identified in the EtOAc fraction. This study justifies the use of A. macrostachya in the treatment of infections in traditional medicine and highlights its potential as a source of bioactive compounds that could possibly interact with some resistance mechanisms in bacteria to combat antimicrobial resistance.

Antimicrobial and cytotoxic capacity of pyroligneous extracts films of Eucalyptus grandis and chitosan for oral applications.

Chitosan films containing distilled pyroligneous extracts of Eucalyptus grandis (DPEC), characterized and developed by Brazilian Agricultural Research Corporation-Embrapa Temperate Agriculture (EMBRAPA-CPACT), were evaluated for antimicrobial activity against Candida albicans, Streptococcus mutans, and Lactobacillus acidophilus by direct contact test. Further, their capacity for the prevention of teeth enamel demineralization and cytotoxicity in vitro were also determined. The natural polymers were tested at different concentrations (1500-7500 µg mL-1) and the formulation of an experimental fluoride varnish with antimicrobial activity was evaluated by direct contact test, whereas cytotoxicity was analyzed through the colorimetric MTT assay. Preliminary data showed no statistically significant differences in cytotoxicity to NIH/3T3 cell line when DPEC is compared to the control group. On the other hand, the antimicrobial capacity and demineralization effects were found between the test groups at the different concentrations tested. Chitosan films containing distilled pyroligneous extracts of E. grandis may be an effective control strategy to prevent biofilm formation related to dental caries when applied as a protective varnish. They may inhibit the colonization of oral microorganisms and possibly control dental caries through a decrease in pH and impairment of enamel demineralization.

Feedback between bottom-up and top-down control of stream biofilm mediated through eutrophication effects on grazer growth.

Algal biofilms in streams are simultaneously controlled by light and nutrient availability (bottom-up control) and by grazing activity (top-down control). In addition to promoting algal growth, light and nutrients also determine the nutritional quality of algae for grazers. While short-term experiments have shown that grazers increase consumption rates of nutrient-poor algae due to compensatory feeding, nutrient limitation in the long run can constrain grazer growth and hence limit the strength of grazing activity. In this study, we tested the effects of light and phosphorus availability on grazer growth and thus on the long-term control of algal biomass. At the end of the experiment, algal biomass was significantly affected by light, phosphorus and grazing, but the interactive effects of the three factors significantly changed over time. At both high light and phosphorus supply, grazing did not initially reduce algal biomass, but the effect of grazing became stronger in the final three weeks of the experiment. Snail growth was enhanced by light, rather than phosphorus, suggesting that algal quantity rather than quality was the main limiting factor for grazer growth. Our results highlight the role of feedback effects and the importance of long-term experiments in the study of foodweb interactions.

A new Kunitz trypsin inhibitor from Erythrina poeppigiana exhibits antimicrobial and antibiofilm properties against bacteria.

Erythrina poeppigiana belongs to Fabaceae family (subfamily Papillionoideae) and is commonly found in tropical and subtropical regions in Brazil. Herein, we described the purification and characterization of a new Kunitz-type inhibitor, obtained from E. poeppigiana seeds (EpTI). EpTI is composed by three isoforms of identical amino-terminal sequences with a molecular weight ranging from 17 to 20 kDa. The physicochemical features showed by EpTI are common to Kunitz inhibitors, including the dissociation constant (13.1 nM), stability against thermal (37-100 °C) and pH (2-10) ranging, and the presence of disulfide bonds stabilizing its reactive site. Furthermore, we investigated the antimicrobial, anti-adhesion, and anti-biofilm properties of EpTI against Gram-positive and negative bacteria. The inhibitor showed antimicrobial activity with a minimum inhibitory concentration (MIC, 5-10 µM) and minimum bactericidal concentration (MBC) of 10 µM for Enterobacter aerogenes, Enterobacter cloacae, Klebsiella pneumoniae, Staphylococcus aureus, and Staphylococcus haemolyticus. The combination of EpTI with ciprofloxacin showed a marked synergistic effect, reducing the antibiotic concentration by 150%. The increase in crystal violet uptake for S. aureus and K. pneumoniae strains was approximately 30% and 50%, respectively, suggesting that the bacteria plasma membrane is targeted by EpTI. Treatment with EpTI at 1x and 10 x MIC significantly reduced the biofilm formation and prompted the disruption of a mature biofilm. At MIC/2, EpTI decreased the bacterial adhesion to polystyrene surface within 2 h. Finally, EpTI showed low toxicity in animal model Galleria mellonella. Given its antimicrobial and anti-biofilm properties, the EpTI sequence might be used to design novel drug prototypes.