The essential oil of Melaleuca alternifolia incorporated into hydrogel induces antimicrobial and anti-inflammatory effects on infected wounds by Staphylococcus aureus.

Staphylococcus aureus is one of the most common bacterial isolates found in wounds. Thus, innovative dressings, such as hydrogels, are interesting vehicles for incorporating bioactive compounds like those from Melaleuca alternifolia essential oil (MaEO). In this study, we evaluated the antimicrobial and anti-inflammatory potential of MaEO incorporated into an alginate and chitosan hydrogel for treating wounds infected by S. aureus. The hydrogel incorporated with MaEO 1% (HMa 1%) was homogeneous with a bright pale-yellow color and the characteristic smell of Melaleuca. The incorporation of MaEO 1% does not affect the stability of the hydrogel, which was stable up to 90 days of storage. The Scanning electron microscopy analysis revealed that hydrogels showed irregular surfaces and interconnected porous structures with accumulations of oil crystals distributed throughout the formulation. HMa 1% has a high moisture content (95.1%) and can absorb simulated wound fluid. Regarding the antimicrobial effects, HMa 1% reduced the growth of S. aureus ATCC 6538 in both in vitro conditions and in an ex vivo model of wounds using porcine skin. In addition, the dairy topical treatment of murine skin lesions with HMa 1% induced a significant reduction of the wound area, inflammation score, and bacterial load, as well as tissue re-epithelialization and modulation of inflammatory mediators. Therefore, hydrogel incorporated with MaEO 1% has excellent potential to be used in the pharmacotherapy of infected wounds.

Biocontrol Potential of Serratia Marcescens (B8) and Bacillus sp. (B13) Isolated from Urban Mangroves in Raposa, Brazil.

This study analyzed the antifungal potential of 16 bacterial strains isolated from mangrove sediment. Bacterial selection was conducted in a solid medium. This was followed by the production and extraction of metabolites using ethyl acetate to evaluate chitinase production, antifungal activity, and toxicity toward Allium cepa and Tenebrio molitor. Bacterial strains B8, B11, and B13 produced the largest inhibition halos (>30 mm) toward Fusarium solani, Fusarium oxysporum, and Rhizoctonia solani fungi. Strains B1, B3, B6, B8, B11, B13, B14, and B16 produced chitinases. In assays using liquid media, B8 and B13 produced the largest inhibition halos. Exposing the fungal inocula to metabolic extracts of strains B6, B8, B11, B13, B14, B15, and B16 caused micromorphological alterations in the inocula, culminating in the inhibition of R. solani sporulation and spore germination. Toxicity tests using Allium cepa and Tenebrio molitor revealed that the metabolites showed low toxicity. Six of the bacterial strains were molecularly identified to species levels, and a further two to genus level. These included Serratia marcescens (B8), which exhibited activity in all tests. Mangroves provide a useful resource for the isolation of microorganisms for biocontrol. Among the isolates, Serratia marcescens and Bacillus spp. showed the greatest potential to produce metabolites for use as biocontrol agents in agriculture.

Mannose-Binding Lectins as Potent Antivirals against SARS-CoV-2.

The SARS-CoV-2 entry into host cells is mainly mediated by the interactions between the viral spike protein (S) and the ACE-2 cell receptor, which are highly glycosylated. Therefore, carbohydrate binding agents may represent potential candidates to abrogate virus infection. Here, we evaluated the in vitro anti-SARS-CoV-2 activity of two mannose-binding lectins isolated from the Brazilian plants Canavalia brasiliensis and Dioclea violacea (ConBR and DVL). These lectins inhibited SARS-CoV-2 Wuhan-Hu-1 strain and variants Gamma and Omicron infections, with selectivity indexes (SI) of 7, 1.7, and 6.5, respectively for ConBR; and 25, 16.8, and 22.3, for DVL. ConBR and DVL inhibited over 95% of the early stages of the viral infection, with strong virucidal effect, and also protected cells from infection and presented post-entry inhibition. The presence of mannose resulted in the complete lack of anti-SARS-CoV-2 activity by ConBR and DVL, recovering virus titers. ATR-FTIR, molecular docking, and dynamic simulation between SARS-CoV-2 S and either lectins indicated molecular interactions with predicted binding energies of -85.4 and -72.0 Kcal/Mol, respectively. Our findings show that ConBR and DVL lectins possess strong activities against SARS-CoV-2, potentially by interacting with glycans and blocking virus entry into cells, representing potential candidates for the development of novel antiviral drugs.

Draft genome sequence of Staphylococcus aureus sequence type 5 SA01 isolated from bloodstream infection and comparative analysis with reference strains.

A Staphylococcus aureus isolate (SA01) obtained from bloodstream infection exhibited a remarkable drug resistance profile. In this study, we report the draft genome sequence of S. aureus ST 5 SA01, a multidrug-resistant isolate, and analyzed the genes associated with drug resistance and virulence. The genome sketch of S. aureus ST5 SA01 was sequenced with Illumina and annotated using the Prokka software. Rapid Annotation Subsystem Technology (RAST) was used to verify the gene functions in the genome subsystems. The Comprehensive Antibiotic Resistance Database (CARD) and Virulence Factor Database (VFDB) were used in the analysis. The RAST indicated a contribution of 25 proteins to host adenine, fibronectin-binding protein A (FnbA), and biofilm formation as an intercellular polysaccharide adhesive system (PIA). The MLST indicated that S. aureus ST 5 SA01 belongs to ST5 (CC5). In silico analyses also showed an extensive repertoire of genes associated with toxins, such as LukGH leukocidin, enterotoxins, and superantigen staphylococcal classes (SSL). The 11 genes for antimicrobial resistance in S. aureus ST 5 SA01 showed similarity and identity above ≥ 99% with nucleotide sequences deposited in GenBank. Although studies on ST5 clones in Brazil are scarce, monitoring the clone of S. aureus ST 5 SA01 is essential, as it has become a problem in pediatrics in several countries.

Development and Characterization of Hydroxyethyl Cellulose-Based Gels Containing Lactobacilli Strains: Evaluation of Antimicrobial Effects in In Vitro and Ex Vivo Models.

This study aimed to develop a hydroxyethyl cellulose-based topical formulation containing probiotics and to evaluate its antimicrobial action using in vivo and ex vivo models. Initially, the antagonistic effects of Lacticaseibacillus rhamnosus ATCC 10863, Limosilactobacillus fermentum ATCC 23271, Lactiplantibacillus plantarum ATCC 8014 and Lactiplantibacillus plantarum LP-G18-A11 were analyzed against Enterococcus faecalis ATCC 29212, Klebsiella pneumoniae ATCC 700603, Staphylococcus aureus ATCC 27853 and Pseudomonas aeruginosa ATCC 2785. The best action was seen for L. plantarum LP-G18-A11, which presented high inhibition against S. aureus and P. aeruginosa. Then, lactobacilli strains were incorporated into hydroxyethyl cellulose-based gels (natrosol); however, only the LP-G18-A11-incorporated gels (5% and 3%) showed antimicrobial effects. The LP-G18-A11 gel (5%) maintained its antimicrobial effects and viability up to 14 and 90 days at 25 °C and 4 °C, respectively. In the ex vivo assay using porcine skin, the LP-G18-A11 gel (5%) significantly reduced the skin loads of S. aureus and P. aeruginosa after 24 h, while only P. aeruginosa was reduced after 72 h. Moreover, the LP-G18-A11 gel (5%) showed stability in the preliminary and accelerated assays. Taken together, the results show the antimicrobial potential of L. plantarum LP-G18-A11, which may be applied in the development of new dressings for the treatment of infected wounds.

Cinnamaldehyde for the Treatment of Microbial Infections: Evidence Obtained from Experimental Models.

Cinnamaldehyde (CNM) is a cyclic terpene alcohol found as the major compound of essential oils from some plants of the genus Cinnamomum (Lauraceae). CNM has several reported pharmacological activities, including antimicrobial, antivirulence, antioxidant, and immunomodulatory effects. These properties make CNM an attractive lead molecule for the development of anti-infective agents. In this descriptive review, we discuss the application of CNM in experimental models of microbial infection using invertebrate and vertebrate organisms. CNM (pure or in formulations) has been successfully applied in the treatment of infections caused by a range of bacterial (such as Cronobacter sakazakii, Escherichia coli, Listeria monocytogenes, Mycobacterium tuberculosis, Pseudomonas aeruginosa, Salmonella enterica, Staphylococcus aureus, Streptococcus agalactiae, Vibrio cholerae) and fungal (such as Aspergillus fumigatus, Candida albicans and Cryptococcus neoformans) pathogens. All these experimental evidence-based findings have promoted the use of cinnamaldehyde as the leading molecule for developing new anti- infective drugs.

Recent advances to combat ESKAPE pathogens with special reference to essential oils.

Biofilm-associated bacteria, especially ESKAPE pathogens (Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter spp.), are a serious challenge worldwide. Due to the lack of discovery of novel antibiotics, in the past two decades, it has become necessary to search for new antibiotics or to study synergy with the existing antibiotics so as to counter life-threatening infections. Nature-derived compounds/based products are more efficient than the chemically synthesized ones with less resistance and lower side effects. In this descriptive review, we discuss the most promising therapeutics for the treatment of ESKAPE-related biofilms. The first aspect includes different types of natural agents [botanical drugs, essential oils (EOs), antimicrobial peptides, bacteriophages, and endolysins] effective against ESKAPE pathogens. The second part of the review deals with special references to EOs/essential oil components (EOCs) (with some exclusive examples), mode of action (via interfering in the quorum-sensing pathways, disruption of biofilm and their inhibitory concentrations, expression of genes that are involved, other virulence factors), existing in literature so far. Moreover, different essential oils and their major constituents were critically discussed using in vivo models to target ESKAPE pathogens along with the studies involving existing antibiotics.

Ethyl Acetate Fraction of Punica granatum and Its Galloyl-HHDP-Glucose Compound, Alone or in Combination with Fluconazole, Have Antifungal and Antivirulence Properties against Candida spp.

Candidiasis is the most common fungal infection among immunocompromised patients. Its treatment includes the use of antifungals, which poses limitations such as toxicity and fungal resistance. Plant-derived extracts, such as Punica granatum, have been reported to have antimicrobial activity, but their antifungal effects are still unknown. We aimed to evaluate the antifungal and antiviral potential of the ethyl acetate fraction of P. granatum (PgEA) and its isolated compound galloyl-hexahydroxydiphenoyl-glucose (G-HHDP-G) against Candida spp. In silico analyses predicted the biological activity of G-HHDP-G. The minimum inhibitory concentrations (MIC) of PgEA and G-HHDP-G, and their effects on biofilm formation, preformed biofilms, and phospholipase production were determined. In silico analysis showed that G-HHDP-G has antifungal and hepatoprotective effects. An in vitro assay confirmed the antifungal effects of PgEA and G-HHDP-G, with MIC in the ranges of 31.25-250 µg/mL and 31.25 ≥ 500 µg/mL, respectively. G-HHDP-G and PgEA synergistically worked with fluconazole against planktonic cells. The substances showed antibiofilm action, alone or in combination with fluconazole, and interfered with phospholipase production. The antifungal and antibiofilm actions of PgEA and G-HHDP-G, alone or in combination with fluconazole, in addition to their effects on reducing Candida phospholipase production, identify them as promising candidates for therapeutics.

Molecular Detection of Drug-Resistance Genes of blaOXA-23-blaOXA-51 and mcr-1 in Clinical Isolates of Pseudomonas aeruginosa.

Pseudomonas aeruginosa has caused high rates of mortality due to the appearance of strains with multidrug resistance (MDR) profiles. This study aimed to characterize the molecular profile of virulence and resistance genes in 99 isolates of P. aeruginosa recovered from different clinical specimens. The isolates were identified by the automated method Vitek2, and the antibiotic susceptibility profile was determined using different classes of antimicrobials. The genomic DNA was extracted and amplified by multiplex polymerase chain reaction (mPCR) to detect different virulence and antimicrobial resistance genes. Molecular typing was performed using the enterobacterial repetitive intergenic consensus (ERIC-PCR) technique to determine the clonal relationship among P. aeruginosa isolates. The drug susceptibility profiles of P. aeruginosa for all strains showed high levels of drug resistance, particularly, 27 (27.3%) isolates that exhibited extensively drug-resistant (XDR) profiles, and the other isolates showed MDR profiles. We detected the polymyxin E (mcr-1) gene in one strain that showed resistance against colistin. The genes that confer resistance to oxacillin (blaOXA-23 and blaOXA-51) were present in three isolates. One of these isolates carried both genes. As far as we know from the literature, this is the first report of the presence of blaOXA-23 and blaOXA-51 genes in P. aeruginosa.

Antimicrobial Potential of Streptomyces ansochromogenes (PB3) Isolated From a Plant Native to the Amazon Against Pseudomonas aeruginosa.

The objective of this study was to evaluate the antibacterial action of filamentous bacteria isolated from the Byrsonima crassifolia leaf. An endophytic bacterium has been identified by classical and molecular techniques as Streptomyces ansochromogene. Screening for antibacterial action against pathogens with medical relevance (Klebsiella pneumoniae ATCC 700603, Pseudomonas aeruginosa ATCC 15692, Staphylococcus aureus ATCC 6538, Corynebacterium diphtheriae ATCC 27012, Mycobacterium abscessus, Cryptococcus gattii ATCC 24065, and Cryptococcus neoformans ATCC 24067) demonstrated activity against the bacterium P. aeruginosa ATCC 0030 with inhibition diameter zones (IDZ) of 17.6 ± 0.25 mm in the preliminary screening in solid medium. After fermentation in liquid medium, an IDZ of 19.6 ± 0.46 mm and a minimum inhibitory concentration (MIC) of 0.5 mg/mL were detected. The antibiofilm action was observed with 100% inhibition of biofilm formation at a concentration of 0.250 mg/mL. When the infection curve was prepared, it was observed that the metabolite was effective in protecting the larvae of Tenebrio molitor. The metabolite does not show toxicity for eukaryotic cells. The leishmanicidal activity demonstrated that the metabolite presented a dose-dependent effect on the promastigotes forms of Leishmania amazonensis growth and the estimated IC50/72 h was 71.65 ± 7.4 µg/mL. Therefore, it can be concluded that the metabolite produced by the endophytic bacterium Streptomyces sp. is promising for future use as an alternative strategy against bacterial resistance.

Antioxidant Action and In Vivo Anti-Inflammatory and Antinociceptive Activities of Myrciaria floribunda Fruit Peels: Possible Involvement of Opioidergic System.

This work evaluated the antioxidant properties and in vivo antinociceptive and anti-inflammatory effects of extracts obtained from fruit peels of Myrciaria floribunda (H. West ex Willd.) O. Berg (Myrtaceae). This plant is popularly known in Brazil as Cambuí or camboim. Different extracts were submitted to comparative analysis to determine the content of selected phytochemical classes (levels of total phenols, flavonoids, and monomeric anthocyanins) and the in vitro antioxidant potentials. The extract with higher potential was selected for in vivo evaluation of its antinociceptive and anti-inflammatory action. Finally, the chemical characterization of this extract was performed by high-performance liquid chromatography (HPLC). MfAE (extract obtained using acetone as solvent) showed the higher levels of phenols (296 mg GAE/g) and anthocyanins contents (35.65 mg Cy-3-glcE/g) that were associated with higher antioxidant activity. MfAE also exhibited in vivo anti-inflammatory and analgesic propertiers. This fraction inhibited the inflammatory and neurogenic phases of pain, and this effect was reversed by naloxone (suggesting the involvement of opioidergic system). MfAE reduced the abdominal contortions induced by acetic acid. The HPLC analysis revealed the presence of gallic acid (and its derivatives) and ellagic acid. Taken together, these data support the use of M. floribunda fruit peels for development of functional foods and nutraceutics.

Antimicrobial and Antivirulence Action of Eugenia brejoensis Essential Oil in vitro and in vivo Invertebrate Models.

Eugenia brejoensis L. (Myrtaceae) is an endemic plant from caatinga ecosystem (brazilian semi-arid) which have an E. brejoensis essential oil (EbEO) with reported antimicrobial activity. In this work, in vitro and in vivo models were used to characterize the inhibitory effects of EbEO in relation to Staphylococcus aureus. EbEO inhibited the growth of all tested S. aureus strains (including multidrug resistance isolates) with values ranging from 8 to 516 µg/mL. EbEO also synergistically increased the action of ampicillim, chloramphenicol, and kanamycin. The treatment with subinhibitory concentrations (Sub-MIC) of EbEO decreased S. aureus hemolytic activity and its ability to survive in human blood. EbEO strongly reduced the levels of staphyloxanthin (STX), an effect related to increased susceptibility of S. aureus to hydrogen peroxide. The efficacy of EbEO against S. aureus was further demonstrated using Caenorhabditis elegans and Galleria mellonella. EbEO increased the lifespan of both organisms infected by S. aureus, reducing the bacterial load. In addition, EbEO reduced the severity of S. aureus infection in G. mellonella, as shown by lower levels of melanin production in those larvae. In summary, our data suggest that EbEO is a potential source of lead molecules for development of new therapeutic alternatives against S. aureus.

Oxidative and nitrosative stresses in cerebral malaria: can we target them to avoid a bad prognosis?

There is currently a global effort to reduce malaria morbidity and mortality. However, malaria still results in the deaths of thousands of people every year. Malaria is caused by Plasmodium spp., parasites transmitted through the bite of an infected female Anopheles mosquito. Treatment timing plays a decisive role in reducing mortality and sequelae associated with the severe forms of the disease such as cerebral malaria (CM). The available antimalarial therapy is considered effective but parasite resistance to these drugs has been observed in some countries. Antimalarial drugs act by increasing parasite lysis, especially through targeting oxidative stress pathways. Here we discuss the roles of reactive oxygen species and reactive nitrogen intermediates in CM as a result of host-parasite interactions. We also present evidence of the potential contribution of oxidative and nitrosative stress-based antimalarial drugs to disease treatment and control.

Antifungal and Antivirulence Activities of Hydroalcoholic Extract and Fractions of Platonia insignis Leaves against Vaginal Isolates of Candida Species.

Vulvovaginal candidiasis is a common fungal infection in women. In this study, Platonia insignis hydroalcoholic extract (PiHE) and its fractions were evaluated for antifungal and antivirulence activities against vaginal Candida species. Dichloromethane (DCMF) and ethyl acetate fractions (EAF) obtained from PiHE effectively inhibited the pathogen. Electrospray ionization mass spectrometry was used for identifying the main compounds in extracts. Minimal inhibitory and fungicidal concentrations (MIC and MFC, respectively) were determined by a broth microdilution assay. Furthermore, we evaluated the effect of the extract and fractions on the virulence properties of Candida albicans, and their cytotoxicity effect was determined on RAW 264.7 cells. Compounds found in extracts were flavonoid glycosides, mainly derivatives of quercetin and myricetin. Extracts showed antifungal potential, with the lowest MIC value for EAF (1.3 mg/mL) and inhibited Candida adherence and biofilm formation. EAF disrupted 48 h biofilms with an inhibition rate of more than 90%. The extract and its fractions exhibited no cytotoxicity. The antifungal effects were attributed to the ability of these extracts to alter the mitochondrial membrane potential for the release of pro-apoptotic factors in the cytosol. In conclusion, our data suggest that PiHE and EAF could act as novel candidates for the development of new therapeutic treatments against fungal infections.

The latex of Euphorbia tirucalli inhibits staphyloxanthin production and protects Tenebrio molitor larvae against Staphylococcus aureus infection.

The latex of Euphorbia tirucalli L. (LET) has great etnopharmacological relevance for several traditional communities. In this study, the in vitro and in vivo (using Tenebrio molitor larvae) antimicrobial effects of LET were evaluated. LET did not inhibit the growth of S. aureus, however, a reduction on staphyloxanthin production (an important virulence factor of S. aureus) was observed. LET (at 10 µL/kg) was also able to enhance the survival of larvae infected with a lethal dose of S. aureus, an effect associated with reduction in the numbers of haemocytes. Furthermore, haemocytes from LET-treated larvae exhibited dysfunctional lysosome activity. These results indicate the effectiveness of LET as an anti-infective agent which could be useful as source of lead molecules for the development of new therapies against S. aureus-induced infections.

Asteraceae Plants as Sources of Compounds Against Leishmaniasis and Chagas Disease.

Leishmaniasis and Chagas disease cause great impact on social and economic aspects of people living in developing countries. The treatments for these diseases are based on the same regimen for over 40 years, thus, there is an urgent need for the development of new drugs. In this scenario, Asteraceae plants (a family widely used in folk medicine worldwide) are emerging as an interesting source for new trypanocidal and leishmanicidal compounds. Herein, we provide a non-exhaustive review about the activity of plant-derived products from Asteraceae with inhibitory action toward Leishmania spp. and T. cruzi. Special attention was given to those studies aiming the isolation (or identification) of the bioactive compounds. Ferulic acid, rosmarinic acid, and ursolic acid (Baccharis uncinella DC.) were efficient to treat experimental leishmaniasis; while deoxymikanolide (Mikania micrantha) and (+)-15-hydroxy-labd-7-en-17-al (Aristeguietia glutinosa Lam.) showed in vivo anti-T. cruzi action. It is also important to highlight that several plant-derived products (compounds, essential oils) from Artemisia plants have shown high inhibitory potential against Leishmania spp., such as artemisinin and its derivatives. In summary, these compounds may help the development of new effective agents against these neglected diseases.

Evaluation of in vitro Antifungal Activity of Xylosma prockia (Turcz.) Turcz. (Salicaceae) Leaves Against Cryptococcus spp.

Cryptococcus species are responsible for important systemic mycosis and are estimated to cause millions of new cases annually. The available therapy is limited due to the high toxicity and the increasing rates of yeast resistance to antifungal drugs. Popularly known as "sucará," Xylosma prockia (Turcz.) Turcz. (Salicaceae) is a native plant from Brazil with little information on its pharmacological potential. In this work, we evaluated in vitro anticryptococcal effects of the leaf ethanolic extract of X. prockia and its fractions against Cryptococcus gattii and Cryptococcus neoformans. We also evaluated phenotypic alterations caused by ethyl acetate fraction (EAF) (chosen according to its biological results). The liquid chromatography-mass spectrometry (LC-MS) analysis of EAF demonstrated the presence of phenolic metabolites that belong to three structurally related groups as majority compounds: caffeoylquinic acid, coumaroyl-glucoside, and caffeoyl-glucoside/deoxyhexosyl-caffeoyl glucoside derivatives. The minimum inhibitory concentration (MIC) values against C. gattii and C. neoformans ranged from 8 to 64 mg/L and from 0.5 to 8 mg/L, for ethanolic extract and EAF, respectively. The EAF triggered an oxidative burst and promoted lipid peroxidation. EAF also induced a reduction of ergosterol content in the pathogen cell membrane. These effects were not associated with alterations in the cell surface charge or in the thermodynamic fingerprint of the molecular interaction between EAF and the yeasts evaluated. Cytotoxic experiments with peripheral blood mononuclear cells (PBMCs) demonstrated that EAF was more selective for yeasts than was PBMCs. The results may provide evidence that X. prockia leaf extract might indeed be a potential source of antifungal agents.