A systematic study on the characteristics and applications of laccases produced by fungi: insights on their potential for biotechnologies.

Laccases are polyphenol oxidase enzymes and form the enzyme complex known for their role in wood decomposition and lignin degradation. The present study aimed to systematically review the state-of-the-art trends in scientific publications on laccase enzymes of the last 10 years. The main aspects checked included the laccase-producing fungal genera, the conditions of fungal growth and laccase production, the methods of immobilization, and potential applications of laccase. After applying the systematic search method 177 articles were selected to compound the final database. Although various fungi produce laccase, most studies were Trametes and Pleurotus genera. The submerged fermentation (SmF) has been the most used, however, the use of solid-state fermentation (SSF) appeared as a promising technique to produce laccase when using agro-industrial residues as substrates. Studies on laccase immobilization showed the covalent bonding and entrapment methods were the most used, showing greater efficiency of immobilization and a high number of enzyme reuses. The main use of the laccase was in bioremediation, especially in the discoloration of dyes from the textile industry and the degradation of pharmaceutical waste. Implications and consequences of all these findings in biotechnology and environment, as well as the trends and gaps of laccase research were discussed.

Study of co-occurrence of mycotoxins in cocoa beans in Brazil by liquid chromatography-tandem mass spectrometry (LC-MS/MS).

In this study, 135 samples of cocoa beans collected in the Amazon and Atlantic Forest regions of Brazil were analysed to evaluate the possible co-occurrence of 34 mycotoxins. The results indicate that 42% of the cocoa samples exhibited quantifiable levels for 11 mycotoxins: aflatoxins (AFs) B1, B2 and G1; ochratoxin A; citrinin; cyclopiazonic acid; tenuazonic acid; paxilline; sterigmatocystin; zearalenone and fumonisin B2. Of the samples, 18% exhibited the co-occurrence of up to six mycotoxins. No toxins belonging to the groups of trichothecenes or ergot alkaloids were detected. Contingency analysis of the incidence of mycotoxins did not show significant differences between the two regions evaluated. Seven samples were contaminated with AFs, while only one contained ochratoxin A above 10 µg kg-1. The accuracy of the method was evaluated by proficiency testing for ochratoxin A, where satisfactory Z-scores were obtained.

Unraveling potential enzymes and their functional role in fine cocoa beans fermentation using temporal shotgun metagenomics.

Cocoa beans fermentation is a spontaneous process, essential for the generation of quality starting material for fine chocolate production. The understanding of this process has been studied by the application of high-throughput sequencing technologies, which grants a better assessment of the different microbial taxa and their genes involved in this microbial succession. The present study used shotgun metagenomics to determine the enzyme-coding genes of the microbiota found in two different groups of cocoa beans varieties during the fermentation process. The statistical evaluation of the most abundant genes in each group and time studied allowed us to identify the potential metabolic pathways involved in the success of the different microorganisms. The results showed that, albeit the distinction between the initial (0 h) microbiota of each varietal group was clear, throughout fermentation (24-144 h) this difference disappeared, indicating the existence of selection pressures. Changes in the microbiota enzyme-coding genes over time pointed to the distinct ordering of fermentation at 24-48 h (T1), 72-96 h (T2), and 120-144 h (T3). At T1, the significantly more abundant enzyme-coding genes were related to threonine metabolism and those genes related to the glycolytic pathway, explained by the abundance of sugars in the medium. At T2, the genes linked to the metabolism of ceramides and hopanoids lipids were clearly dominant, which are associated with the resistance of microbial species to extreme temperatures and pH values. In T3, genes linked to trehalose metabolism, related to the response to heat stress, dominated. The results obtained in this study provided insights into the potential functionality of microbial community succession correlated to gene function, which could improve cocoa processing practices to ensure the production of more stable quality end products.

Pectinases Secretion by Saccharomyces cerevisiae: Optimization in Solid-State Fermentation and Identification by a Shotgun Proteomics Approach.

A sequential design strategy was applied to optimize the secretion of pectinases by a Saccharomyces cerevisiae strain, from Brazilian sugarcane liquor vat, on passion fruit residue flour (PFRF), through solid-state fermentation (SSF). A factorial design was performed to determine the influence variables and two rotational central composite designs were executed. The validated experimental result was of 7.1 U mL-1 using 50% PFRF (w/w), pH 5, 30 °C for 24 h, under static SSF. Polygalacturonase, pectin methyl esterase, pectin-lyase and pectate-lyase activities were 3.5; 0.08; 3.1 and 0.8 U mL-1, respectively. Shotgun proteomics analysis of the crude extract enabled the identification of two pectin-lyases, one pectate-lyase and a glucosidase. The crude enzymatic extract maintained at least 80% of its original activity at pH values and temperatures ranging from 2 to 8 and 30 to 80 °C, respectively, over 60 min incubation. Results revealed that PFRF might be a cost-effective and eco-friendly substrate to produce pectinases. Statistical optimization led to fermentation conditions wherein pectin active proteins predominated. To the extent of our knowledge, this is the first study reporting the synthesis of pectate lyase by S. cerevisiae.

Unraveling the Genomic Potential of the Thermophilic Bacterium Anoxybacillus flavithermus from an Antarctic Geothermal Environment.

Antarctica is a mosaic of extremes. It harbors active polar volcanoes, such as Deception Island, a marine stratovolcano having notable temperature gradients over very short distances, with the temperature reaching up to 100 °C near the fumaroles and subzero temperatures being noted in the glaciers. From the sediments of Deception Island, we isolated representatives of the genus Anoxybacillus, a widely spread genus that is mainly encountered in thermophilic environments. However, the phylogeny of this genus and its adaptive mechanisms in the geothermal sites of cold environments remain unknown. To the best of our knowledge, this is the first study to unravel the genomic features and provide insights into the phylogenomics and metabolic potential of members of the genus Anoxybacillus inhabiting the Antarctic thermophilic ecosystem. Here, we report the genome sequencing data of seven A. flavithermus strains isolated from two geothermal sites on Deception Island, Antarctic Peninsula. Their genomes were approximately 3.0 Mb in size, had a G + C ratio of 42%, and were predicted to encode 3500 proteins on average. We observed that the strains were phylogenomically closest to each other (Average Nucleotide Identity (ANI) > 98%) and to A. flavithermus (ANI 95%). In silico genomic analysis revealed 15 resistance and metabolic islands, as well as genes related to genome stabilization, DNA repair systems against UV radiation threats, temperature adaptation, heat- and cold-shock proteins (Csps), and resistance to alkaline conditions. Remarkably, glycosyl hydrolase enzyme-encoding genes, secondary metabolites, and prophage sequences were predicted, revealing metabolic and cellular capabilities for potential biotechnological applications.

Metagenome-Assembled Genomes of Komagataeibacter from Kombucha Exposed to Mars-Like Conditions Reveal the Secrets in Tolerating Extraterrestrial Stresses.

Kombucha mutualistic community (KMC) is composed by acetic acid bacteria and yeasts, producing fermented tea with health benefits. As part of the BIOlogy and Mars EXperiment (BIOMEX) project, the effect of Mars-like conditions on the KMC was analyzed. Here, we analyzed metagenome-assembled genomes (MAGs) of the Komagataeibacter, which is a predominant genus in KMC, to understand their roles in the KMC after exposure to Mars-like conditions (outside the International Space Station) based on functional genetic elements. We constructed three MAGs: K. hansenii, K. rhaeticus, and K. oboediens. Our results showed that (i) K. oboediens MAG functionally more complex than K. hansenii, (ii) K. hansenii is a keystone in KMCs with specific functional features to tolerate extreme stress, and (iii) genes related to the PPDK, betaine biosynthesis, polyamines biosynthesis, sulfate-sulfur assimilation pathway as well as type II toxin-antitoxin (TA) system, quorum sensing (QS) system, and cellulose production could play important roles in the resilience of KMC after exposure to Mars-like stress. Our findings show the potential mechanisms through which Komagataeibacter tolerates the extraterrestrial stress and will help to understand minimal microbial composition of KMC for space travelers.

Pestalotiopsis mangiferae isolated from cocoa leaves and concomitant tannase and gallic acid production.

The enzyme tannase is of great industrial and biotechnological importance for the hydrolysis of vegetable tannins, reducing their undesirable effects and generating products for a wide range of processes. Thus, the search for new microorganisms that permit more stable tannase production is of considerable importance. A strain of P. mangiferae isolated from cocoa leaves was selected and investigated for its capacity to produce tannase enzymes and gallic acid through submerged fermentation. The assessment of the variables affecting tannase production by P. mangiferae showed that tannic acid, ammonium nitrate and temperature were the most significant (8.4 U/mL). The variables were analyzed using Response Surface Methodology - RSM (Box-Behnken design), with the best conditions for tannase production being: 1.9% carbon source, 1% nitrogen source and temperature of 23 °C. Tannase activity doubled (16.9 U/mL) after the optimization process when compared to the initial fermentation. A pH of 7.0 was optimal for the tannase and it presented stability above 80% with pH between 4.0 and 7.0 after 2h of incubation. The optimal temperature was 30 °C and activity remained at above 80% at 40-60 °C after 1 h. Production of gallic acid was achieved with 1% tannic acid (0.9 mg/mL) and P. mangiferae had not used up the gallic acid produced by tannic acid hydrolysis after 144 h of fermentation. A 5% tannic acid concentration was the best for gallic acid production (1.6 mg/mL). These results demonstrate P. mangiferae's potential for tannase and gallic acid production for biotechnological applications.

Flavanone Glycosides, Triterpenes, Volatile Compounds and Antimicrobial Activity of Miconia minutiflora (Bonpl.) DC. (Melastomataceae).

Chemical composition of the essential oils and extracts and the antimicrobial activity of Miconia minutiflora were investigated. The flavanone glycosides, pinocembroside and pinocembrin-7-O-[4″,6″-HHDP]-ß-D-glucose, were identified, along with other compounds that belong mainly to the triterpene class, besides the phenolics, gallic acid and methyl gallate. Sesquiterpenes and monoterpenes were the major compounds identified from the essential oils. Screening for antimicrobial activity from the methanolic extract of the leaves showed that the MIC and MMC values against the tested microorganisms ranged from 0.625 to 5 mg·mL−1 and that the extract was active against microorganisms, Staphyloccocus aureus, Escherichia coli, and Bacillus cereus.

High-throughput screening for distinguishing nitrilases from nitrile hydratases in Aspergillus and application of a Box-Behnken design for the optimization of nitrilase.

Nitrilases and nitrile hydratases/amidases hydrolyze nitriles into carboxylic acids and/or amides, which are used in industrial chemical processes. In the present study, 26 microorganisms, including yeasts and filamentous fungi, in a minimum solid mineral medium supplemented with glucose and phenylacetonitrile were screened to evaluate their biocatalytic potential. Of these microorganisms, five fungi of the genus Aspergillus were selected and subjected to colorimetry studies to evaluate the production and distinction of nitrilase and nitrile hydratase/amidase enzymes. Aspergillus parasiticus Speare 7967 and A. niger Tiegh. 8285 produced nitrilases and nitrile hydratase, respectively. Nitrilase optimization was performed using a Box-Behnken design (BBD) and fungus A. parasiticus Speare 7967 with phenylacetonitrile volume (µl), pH, and carbohydrate source (starch:glucose; g/g) as independent variables and nitrilase activity (U ml-1 ) as dependent variable. Maximum activity (2.97 × 10-3  U ml-1 ) was obtained at pH 5.5, 80 µl of phenylacetonitrile, and 15 g of glucose. A. parasiticus Speare 7967 showed promise in the biotransformation of nitriles to carboxylic acids.

Citral modulates human monocyte responses to Staphylococcus aureus infection.

Staphylococcus aureus is a Gram-positive bacterium that is considered an important human pathogen. Due to its virulence and ability to acquire mechanisms of resistance to antibiotics, the clinical severity of S. aureus infection is driven by inflammatory responses to the bacteria. Thus, the present study aimed to investigate the modulating role of citral in inflammation caused by S. aureus infection. For this, we used an isolate obtained from a nasal swab sample of a healthy child attending a day-care centre in Vitória da Conquista, Bahia, Brazil. The role of citral in modulating immunological factors against S. aureus infection was evaluated by isolating and cultivating human peripheral blood mononuclear cells. The monocytes were treated with 4%, 2%, and 1% citral before and after inoculation with S. aureus. The cells were analysed by immunophenotyping of monocyte cell surface molecules (CD54, CD282, CD80, HLA-DR, and CD86) and cytokine dosage (IL-1ß, IL-6, IL-10, IL-12p70, IL-23, IFN-γ, TGF-ß, and TNF-α), and evaluated for the expression of 84 genes related to innate and adaptive immune system responses. GraphPad Prism software and variables with P values < 0.05, were used for statistical analysis. Our data demonstrated citral's action on the expression of surface markers involved in recognition, presentation, and migration, such as CD14, CD54, and CD80, in global negative regulation of inflammation with inhibitory effects on NF-κB, JNK/p38, and IFN pathways. Consequently, IL-1ß, IL-6, IL-12p70, IL-23, IFN-γ, and TNF-α cytokine expression was reduced in groups treated with citral and groups treated with citral at 4%, 2%, and 1% and infected, and levels of anti-inflammatory cytokines such as IL-10 were increased. Furthermore, citral could be used as a supporting anti-inflammatory agent against infections caused by S. aureus. There are no data correlating citral, S. aureus, and the markers analysed here; thus, our study addresses this gap in the literature.

Lactiplantibacillus plantarum strains isolated from spontaneously fermented cocoa exhibit potential probiotic properties against Gardnerella vaginalis and Neisseria gonorrhoeae.

BACKGROUND: Probiotics are important tools in therapies against vaginal infections and can assist traditional antibiotic therapies in restoring healthy microbiota. Recent research has shown that microorganisms belonging to the genus Lactobacillus have probiotic potential. Thus, this study evaluated the potential in vitro probiotic properties of three strains of Lactiplantibacillus plantarum, isolated during the fermentation of high-quality cocoa, against Gardnerella vaginalis and Neisseria gonorrhoeae. Strains were evaluated for their physiological, safety, and antimicrobial characteristics. RESULTS: The hydrophobicity of L. plantarum strains varied from 26.67 to 91.67%, and their autoaggregation varied from 18.10 to 30.64%. The co-aggregation of L. plantarum strains with G. vaginalis ranged from 14.73 to 16.31%, and from 29.14 to 45.76% with N. gonorrhoeae. All L. plantarum strains could moderately or strongly produce biofilms. L. plantarum strains did not show haemolytic activity and were generally sensitive to the tested antimicrobials. All lactobacillus strains were tolerant to heat and pH resistance tests. All three strains of L. plantarum showed antimicrobial activity against the tested pathogens. The coincubation of L. plantarum strains with pathogens showed that the culture pH remained below 4.5 after 24 h. All cell-free culture supernatants (CFCS) demonstrated activity against the two pathogens tested, and all L. plantarum strains produced hydrogen peroxide. CFCS characterisation in conjunction with gas chromatography revealed that organic acids, especially lactic acid, were responsible for the antimicrobial activity against the pathogens evaluated. CONCLUSION: The three strains of L. plantarum presented significant probiotic characteristics against the two pathogens of clinical importance. In vitro screening identified strong probiotic candidates for in vivo studies for the treatment of vaginal infections.

Gardnerella vaginalis and Neisseria gonorrhoeae Are Effectively Inhibited by Lactobacilli with Probiotic Properties Isolated from Brazilian Cupuaçu (Theobroma grandiflorum) Fruit.

In recent years, certain Lactobacillus sp. have emerged in health care as an alternative therapy for various diseases. Based on this, this study is aimed at evaluating in vitro the potential probiotics of five lactobacilli strains isolated from pulp of cupuaçu fruit fermentation against Gardnerella vaginalis and Neisseria gonorrhoeae. Our lactobacilli strains were classified as safe for use in humans, and they were tolerant to heat and pH. Our strains were biofilm producers, while hydrophobicity and autoaggregation varied from 13% to 86% and 13% to 25%, respectively. The coaggregation of lactobacilli used in this study with G. vaginalis and N. gonorrhoeae ranged from 15% to 36% and 32% to 52%, respectively. Antimicrobial activity was present in all tested Lactobacillus strains against both pathogens, and the growth of pathogens in coculture was reduced by the presence of our lactobacilli. Also, all tested lactobacilli reduced the pH of the culture, even in incubation with pathogens after 24 hours. The cell-free culture supernatants (CFCS) of all five lactobacilli demonstrated activity against the two pathogens with a halo presence and CFCS characterization assay together with gas chromatography revealed that lactic acid was the most abundant organic acid in the samples (50% to 62%). Our results demonstrated that the organic acid production profile is strain-specific. This study revealed that cupuaçu is a promising source of microorganisms with probiotic properties against genital pathogens. We demonstrated by in vitro tests that our Lactobacillus strains have probiotic properties. However, the absence of in vivo tests is a limitation of our work due to the need to evaluate the interaction of our lactobacilli with pathogens in the vaginal mucosa. We believe that these findings may be useful in developing a product containing our lactobacilli and their supernatants in order to support with vaginal health.

Anticariogenic activities of Libidibia ferrea, gallic acid and ethyl gallate against Streptococcus mutans in biofilm model.

ETHNOPHARMACOLOGICAL RELEVANCE: In Brazil, ethnopharmacological studies show that Libidibia ferrea (Mart. ex Tul.) L. P. Queiroz is commonly used in folk medicine as an antifungal, antimicrobial and anti-inflammatory. In the Amazon region, the dried fruit powder of L. ferrea are widely used empirically by the population in an alcoholic tincture as an antimicrobial mouthwash in oral infections and the infusion is also recommended for healing oral wounds. However, there are few articles that have evaluated the antimicrobial activity against oral pathogens in a biofilm model, identifying active compounds and mechanisms of action. AIM OF THE STUDY: The aim of this study was to evaluate the antimicrobial and anti-adherence activities of the ethanolic extract, fractions and isolated compounds (gallic acid and ethyl gallate) of the fruit and seed of L. ferrea against Streptococcus mutans. The inhibition of acidicity/acidogenicity and the expression of the S. mutans GTF genes in biofilms were also evaluated. MATERIALS AND METHODS: Minimal Inhibitory Concentration (MIC), Minimum Bactericidal Concentration (MBC) and Minimum Inhibitory Concentration of Cell Adhesion (MICA) were evaluated with ethanolic extract (EELF), fractions, gallic acid (GA) and ethyl gallate (EG) against S. mutans. Inhibition of biofilm formation, pH drop and proton permeability tests were conducted with EELF, GA and EG, and also evaluated the expression of the GTF genes in biofilms. The compounds of dichloromethane fraction were identified by GC-MS. RESULTS: This is the first report of shikimic, pyroglutamic, malic and protocatechuic acids identified in L. ferrea. EELF, GA and EG showed MIC at 250 µg/mL, and MBC at 1000 µg/mL by EELF. EELF biofilms showed reduced dry weight and acidogenicity of S. mutans in biofilms. GA and EG reduced viable cells, glucans soluble in alkali, acidogenicity, aciduricity and downregulated expression of gtfB, gtfC and gtfD genes in biofilms. SEM images of GA and EG biofilms showed a reduction of biomass, exopolysaccharide and microcolonies of S. mutans. CONCLUSIONS: The ethanolic extract of fruit and seed of L. ferrea, gallic acid and ethyl gallate showed great antimicrobial activity and inhibition of adhesion, reduction of acidogenicity and aciduricity in S. mutans biofilms. The results obtained in vitro validate the use of this plant in ethnopharmacology, and open opportunities for the development of new oral anticariogenic agents, originated by plants that can inhibit pathogenic biofilm that leads to the development of caries.

Differential Immune Response of Lactobacillus plantarum 286 Against Salmonella Typhimurium Infection in Conventional and Germ-Free Mice.

We aimed at evaluating in vivo the probiotic potential of Lactobacillus plantarum 286 against Salmonella enterica serov. Typhimurium. Colonization capacity and antagonistic activity were determined in feces of gnotobiotic mice. Survival to infection, translocation, histopathology, IgA and cytokine levels (IL-10, IL-6, IFN-γ, TNF-α, TGF-ß) were determined both in conventional and germ-free mice followed L. plantarum 286 administration and Salmonella infection. L. plantarum 286 colonized the intestine of gnotobiotic mice, where it produced antagonistic substances against S. Typhimurium. In conventional animals, the administration of this strain increased intestinal IgA levels and reduced the inflammatory response and the tissue damage caused by S. Typhimurium. Reduction of tissue damage in the intestine and liver of germ-free animals was also observed, however the immune response elicited was different in either model. L. plantarum 286 showed in vivo probiotic properties in both murine models. Probiotic capacity results may depend on the animal model chosen.

Effects of chronic treatment with new strains of Lactobacillus plantarum on cognitive, anxiety- and depressive-like behaviors in male mice.

Psychobiotics correspond to a class of probiotics, mainly of the genus Lactobacillus and Bifidobacterium, capable of producing neuroactive substances, such as γ-aminobutyric acid (GABA) and serotonin, which exert effects on the brain-gut axis. Evidence suggests that psychobiotics can have a beneficial effect on mood, anxiety and cognition. The present study evaluated the effects of chronic administration of two new strains of Lactobacillus plantarum, L. plantarum 286 (Lp 286) and L. plantarum 81 (Lp 81) isolated from the fermentation of cocoa (Theobroma cacao L.) and cupuaçu (Theobroma grandiflorum), respectively, on cognitive, anxiety- and depressive-like behaviors in male Swiss mice. Different groups of animals were administered (oral gavage) solutions of vehicle (0.85% saline plus 15% skim milk), Lp 286 (109/0.1 ml CFU) or Lp 81 (109/0.1 ml CFU) for 30 days, and animals were tested for general locomotor activity, depressive-like behavior in the forced swim test, and learning/memory and anxiety-like behavior in the plus-maze discriminative avoidance task. Treatment with the strains Lp 286 and Lp 81 did not interfere with locomotor activity or learning and memory. The Lp 286 strain exerted anti-depressant- and anxiolytic-like effects under our experimental conditions. Our findings add to the current body of evidence suggesting that probiotics from the genus Lactobacillus may exert psychobiotic potential and introduce a new strain, Lp 286, as a potential candidate in the prevention or as therapeutic adjuvant in the treatment of mental disorders.

Cocoa pulp in beer production: Applicability and fermentative process performance.

This work evaluated the effect of cocoa pulp as a malt adjunct on the parameters of fermentation for beer production on a pilot scale. For this purpose, yeast isolated from the spontaneous fermentation of cachaça (SC52), belonging to the strain bank of the State University of Feira de Santana-Ba (Brazil), and a commercial strain of ale yeast (Safale S-04 Belgium) were used. The beer produced was subjected to acceptance and purchase intention tests for sensorial analysis. At the beginning of fermentation, 30% cocoa pulp (adjunct) was added to the wort at 12°P concentration. The production of beer on a pilot scale was carried out in a bioreactor with a 100-liter capacity, a usable volume of 60 liters, a temperature of 22°C and a fermentation time of 96 hours. The fermentation parameters evaluated were consumption of fermentable sugars and production of ethanol, glycerol and esters. The beer produced using the adjunct and yeast SC52 showed better fermentation performance and better acceptance according to sensorial analysis.

Selection of Lactic Acid Bacteria with Probiotic Potential Isolated from the Fermentation Process of "Cupuaçu" (Theobroma grandiflorum).

In the present study, nine lactic acid bacteria isolated from the fermentation process of "cupuaçu" (Theobroma grandiflorum) were selected for probiotic use. In vitro (resistance to gastrointestinal environment, in vitro antagonism and co-aggregation with pathogens) and in vivo (intestinal colonization and ex vivo antagonism in germ-free mice, cumulative mortality, translocation to liver and spleen, histopathological examination of liver and ileum and mRNA cytokine gene expression during an experimental infection with S. Typhimurium) assays were used. Among the nine Lactobacillus strains isolated from the "cupuaçu" fermentation, L. plantarum 81 and L. plantarum 90 were selected as potential probiotics based on better results obtained in in vitro evaluations (production of diffusible inhibitory compounds and co-aggregation) as well as in vivo experiments (resistance to gastrointestinal environment, ex vivo antagonism, higher survival after enteropathogen challenge, lower hepatic translocation of enteropathogen, lower histopathological lesions in ileum and liver and anti-inflammatory pattern of immunological response). Concluding, L. plantarum 81 and L. plantarum 90 showed in vitro and in vivo capacities for probiotic use through different mechanisms of protection and its origin would allow an easier adaptation in an alimentary matrix for its administration.

Chemical composition and pharmacological properties of the essential oils obtained seasonally from Lippia thymoides.

CONTEXT: Lippia thymoides Mart. & Schauer (Verbenaceae) is used in folk medicine to treat wounds, fever, bronchitis, rheumatism, headaches, and weakness. OBJECTIVE: This study determinates the chemical composition of essential oils from L. thymoides, obtained at during each of the four seasons and correlates with pharmacological properties. MATERIALS AND METHODS: Essential oils were obtained by hydrodistillation and analyzed by gas chromatography coupled to mass spectroscopy (GC-MS). Antioxidant activity was determined by DPPH free radical scavenging and ß-carotene bleaching methods. The antimicrobial assays were performed by minimum inhibitory concentration (MIC) and minimum microbicidal concentration (MMC) methods. Isolated rat aorta and uterus, and guinea-pig trachea were utilized to evaluate relaxant potential in pre-contracted smooth muscle. RESULTS AND DISCUSSION: Essential oils from leaves of L. thymoides had the sesquiterpene ß-caryophyllene (17.22-26.27%) as the major constituent followed by borneol (4.45-7.36%), camphor (3.22-8.61%), camphene (2.64-5.66%), and germacrene D (4.72-6.18%). In vitro assays showed that these essential oils do not have antioxidant activity, have antimicrobial selectivity to Gram-positive bacteria Staphylococcus aureus (MIC = 0.004 mg/mL and MMC = 0.26-10.19 mg/mL) and Micrococcus luteus (MIC = 0.03 mg/mL and MMC = 8.43 mg/mL), relax isolated rat aorta (EC50 = 305-544 µg/mL, with endothelium; and EC50 = 150-283 µg/mL, without endothelium), and uterus (EC50 = 74-257 µg/mL), and minor potency, isolated guinea-pig trachea. CONCLUSIONS: Lippia thymoides is a source of natural products of pharmaceutical interest, being necessary additional studies to determine the substances involved in the biological activities.

Atividade antimicrobiana do óleo essencial de Ocimum americanum e Ocimum basilicum sobre periodontopatógenos / Antimicrobial activity of essential oil of Ocimum americanum and Ocimum basilicum against periodontopathogens

Na doença periodontal, a resposta imune ao desafio microbiano resulta em ativação de osteoclastos e reabsorção do osso alveolar, culminando com a perda do dente. Deste modo, a busca por compostos com atividade antimicrobiana torna-se relevante no controle da formação do biofilme dental. Com o crescente aumento da resistência bacteriana aos antimicrobianos, o descobrimento de novas drogas seria uma ferramenta útil e neste cenário as plantas medicinais são alternativas promissoras. No presente estudo a atividade antimicrobiana de óleos essenciais extraídos de folhas e flores de Ocimum americanum e Ocimum basilicum foi avaliada frente aos periodontopatógenos Aggregatibacter actinomycetemcomitans (ATCC 43717), Fusobacterium nucleatum (ATCC 25586), Porphyromonas gingivalis (ATCC 33277) e frente ao micro-organismo Bacteroides fragilis (ATCC 25285). O método de macrodiluição em tubos foi utilizado para determinação da concentração inibitória mínima (CIM) e concentração bactericida mínima (CBM). Foram registrados menores valores de CIM (0,00625 mg.mL-1 a 0,0125 mg.mL-1) frente a bactéria P. gingivalis, sendo os maiores valores ( <3,2 mg.mL-1) registrados para A. actinomycetemcomitans. Estas concentrações foram consideradas bactericidas quando a CBM foi avaliada. A composição química dos óleos mostrou a presença de linalol e metil cinamato, compostos com reconhecida atividade antimicrobiana, o que poderia explicar os resultados encontrados. Este trabalho mostrou que as plantas medicinais do gênero Ocimum estudadas foram capazes de inibir o crescimento microbiano, especialmente de P. gingivalis, apresentando, portanto, potencial biotecnológico para uso na área de odontologia

In Periodontal disease, the immune response to microbial challenge results in osteoclast activation and alveolar bone resorption, leading to tooth loss, so the search for chemical compounds with antimicrobial activity is relevant to control biofilm formation. With the increasing of bacterial resistance to antibiotics, the discovery of new drugs would be a useful tool, and in this scenario the medicinal plants are promising alternatives. In this study, the antimicrobial activity of essential oils extracted from leaves and flowers of Ocimum americanum and Ocimum basilicum was evaluated against the periodontopathogens Aggregatibacter actinomycetemcomitans (ATCC 43717), Fusobacterium nucleatum (ATCC 25586), Porphyromonas gingivalis (ATCC 33277) and against the bacteria Bacteroides fragilis (ATCC 25285). The macrodilution method was used to determine the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC). There were recorded lower values of MIC (0.00625 mg.mL-1 to 0.0125 mg.mL-1) against P. gingivalis, and the highest values (<3,2 mg.mL-1) were recorded for A. actinomycetemcomitans. These concentrations were considered bactericidal when MBC was assessed. The chemical composition of the oils showed the presence of linalool and methyl cinnamate, compounds with proven antimicrobial activity, which could explain the results. This project showed that medicinal plants studied were able to inhibit microbial growth, especially against P. gingivalis, presenting therefore biotechnological potential for use in dentistry

Actinobacteria from Termite Mounds Show Antiviral Activity against Bovine Viral Diarrhea Virus, a Surrogate Model for Hepatitis C Virus.

Extracts from termite-associated bacteria were evaluated for in vitro antiviral activity against bovine viral diarrhea virus (BVDV). Two bacterial strains were identified as active, with percentages of inhibition (IP) equal to 98%. Both strains were subjected to functional analysis via the addition of virus and extract at different time points in cell culture; the results showed that they were effective as posttreatments. Moreover, we performed MTT colorimetric assays to identify the CC50, IC50, and SI values of these strains, and strain CDPA27 was considered the most promising. In parallel, the isolates were identified as Streptomyces through 16S rRNA gene sequencing analysis. Specifically, CDPA27 was identified as S. chartreusis. The CDPA27 extract was fractionated on a C18-E SPE cartridge, and the fractions were reevaluated. A 100% methanol fraction was identified to contain the compound(s) responsible for antiviral activity, which had an SI of 262.41. GC-MS analysis showed that this activity was likely associated with the compound(s) that had a peak retention time of 5 min. Taken together, the results of the present study provide new information for antiviral research using natural sources, demonstrate the antiviral potential of Streptomyces chartreusis compounds isolated from termite mounds against BVDV, and lay the foundation for further studies on the treatment of HCV infection.