Relationship between clinical-epidemiological parameters and outcomes of patients with COVID-19 admitted to the intensive care unit: a report from a Brazilian hospital.

Background: People in low-income countries, especially those with low socio-economic conditions, are likelier to test positive for SARS-CoV-2. The unequal conditions of public health systems also increase the infection rate and make early identification and treatment of at-risk patients difficult. Here, we aimed to characterize the epidemiological profile of COVID-19 patients in intensive care and identify laboratory and clinical markers associated with death. Materials and methods: We conducted an observational, descriptive, and cross-sectional study in a reference hospital for COVID-19 treatment in the Southern Region of Bahia State, in Brazil, to evaluate the epidemiological, clinical, and laboratory characteristics of COVID-19 patients admitted to the intensive care unit (ICU). Additionally, we used the area under the curve (AUC) to classify survivors and non-survivors and a multivariate logistic regression analysis to assess factors associated with death. Data was collected from the hospital databases between April 2020 and July 2021. Results: The use of bladder catheters (OR 79.30; p < 0.0001) and central venous catheters (OR, 45.12; p < 0.0001) were the main factors associated with death in ICU COVID-19 patients. Additionally, the number of non-survivors increased with age (p < 0.0001) and prolonged ICU stay (p < 0.0001). Besides, SAPS3 presents a higher sensibility (77.9%) and specificity (63.1%) to discriminate between survivors and non-survivor with an AUC of 0.79 (p < 0.0001). Conclusion: We suggest that multi-laboratory parameters can predict patient prognosis and guide healthcare teams toward more assertive clinical management, better resource allocation, and improved survival of COVID-19 patients admitted to the ICU.

Searching for bacteria able to metabolize polycyclic aromatic sulfur compounds in 12-years periodically fed bioreactor.

Biodesulfurization is a promising alternative for removing sulfur molecules from the polycyclic aromatic sulfur compounds (PASC) found in petroleum. PASC consists of recalcitrant molecules that can degrade fuel quality and cause a range of health and environmental problems. Therefore, identifying bacteria capable of degrading PASC is essential for handling these recalcitrant molecules. Microorganisms in environments exposed to petroleum derivatives have evolved specific enzymatic machinery, such as the 4S pathway associated with the dszABC genes, which are directly linked to sulfur removal and utilization as nutrient sources in the biodesulfurization process. In this study, bacteria were isolated from a bioreactor containing landfarm soil that had been periodically fed with petroleum for 12 years, using a medium containing dibenzothiophene (DBT), 4.6-dimethylbenzothiophene, 4-methylbenzothiophene, or benzothiophene. This study aimed to identify microorganisms capable of degrading PASC in such environments. Among the 20 colonies isolated from an inoculum containing DBT as the sole sulfur source, only four isolates exhibited amplification of the dszA gene in the dszABC operon. The production of 2-hydroxybiphenyl (HPB) and a decrease in DBT were detected during the growth curve and resting cell assays. The isolates were identified using 16S rRNA sequencing belonging to the genera Stutzerimonas and Pseudomonas. These isolates demonstrated significant potential for biodesulfurization and/or degradation of PASC. All isolates possessed the potential to be utilized in the biotechnological processes of biodesulfurization and degradation of recalcitrant PASC molecules.

BASIDIN as a New Protein Effector of the Phytopathogen Causing Witche's Broom Disease in Cocoa.

The fungus Moniliophthora perniciosa secretes protein effectors that manipulate the physiology of the host plant, but few effectors of this fungus have had their functions confirmed. We performed functional characterization of a promising candidate effector of M. perniciosa. The inoculation of rBASIDIN at 4 µmol L-1 in the mesophyll of leaflets of Solanum lycopersicum caused symptoms of shriveling within 6 h without the presence of necrosis. However, when sprayed on the plant at a concentration of 11 µmol L-1, it caused wilting symptoms only 2 h after application, followed by necrosis and cell death at 48 h. rBASIDIN applied to Theobroma cacao leaves at the same concentration caused milder symptoms. rBASIDIN caused hydrogen peroxide production in leaf tissue, damaging the leaf membrane and negatively affecting the photosynthetic rate of Solanum lycopersicum plants. Phylogenetic analysis indicated that BASIDIN has orthologs in other phytopathogenic basidiomycetes. Analysis of the transcripts revealed that BASIDIN and its orthologs are expressed in different fungal species, suggesting that this protein is differentially regulated in these basidiomycetes. Therefore, the results of applying BASIDIN allow the inference that it is an effector of the fungus M. perniciosa, with a strong potential to interfere in the defense system of the host plant.

Cupuassu juice fermentation by Lactiplantibacillus plantarum Lp62 produces an antioxidant enriched probiotic beverage.

The present work aimed to produce a cupuassu juice (Theobroma grandiflorum) fermented by the probiotic bacterium Lactiplantibacillus plantarum Lp62 and to analyze its antioxidant potential, antimicrobial activity, and resistance to biological barriers. The fermented beverage showed an increase in the content of phenolics, flavonoids, and antioxidant potential. The culture showed antagonistic activity against pathogens, but this result was not observed when the juice was tested. The probiotic strain remained viable under refrigeration, even in an acidified environment, and survived simulated gastrointestinal transit in vitro. L. plantarum Lp62 showed 30% adherence to HT-29 intestinal cells and proved to be safe in terms of antibiotic resistance and production of virulence factors. Fermentation increased the functional characteristics of cupuassu juice. This drink proved to be a good vehicle for the delivery of the probiotic bacteria L. plantarum Lp62.

Low Occurrence of Salmonella spp. in Wild Animals in Bahia, Brazil-Population Assessment and Characterization in the Caatinga and Atlantic Forest Biomes.

Salmonella spp. are known to persist in the environment. Wild animals are believed to act as important reservoirs, with antimicrobial resistance frequently occurring in the environment. However, little is known about the role of the wildlife in Bahia as a reservoir for Salmonella in Brazil. This study aimed to isolate and characterize Salmonella spp. from wildlife in the Atlantic Forest and Caatinga biomes considering indicators such as the animal species, degree of anthropization, sampling area, and feeding habits. Convenience wildlife sampling and characterization were conducted, followed by microbiological and molecular identification of Salmonella isolates, serotyping, and antimicrobial susceptibility testing. A total of 674 fecal samples were collected from 12 municipalities during 2015-2021, and 4 were positive for the following Salmonella species: Salmonella enterica subspecies enterica serovar Agona (n = 1), Salmonella enterica subsp. enterica serogroup O:16 (n = 2), and Salmonella enterica subsp. enterica serovar Muenchen (n = 1). Antimicrobial susceptibility analysis revealed that one isolate was resistant to six antibiotics, including extended-spectrum penicillins and beta-lactamase inhibitors. These results indicated a low frequency of Salmonella spp. in the sampled forest fragments. The presence of Salmonella in wild animals increases the risk to public health and biodiversity and indicates that they can act as sentinels of environmental contamination or indicators of preservation.

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.

Bioprotective potential of lactic acid bacteria and their metabolites against enterotoxigenic Escherichia coli.

Escherichia coli is one of the main pathogens that impacts swine production. Given the need for methods for its control, the in vitro effect of lactic acid bacteria (LAB) and their metabolites against E. coli F4 was evaluated through cell culture and microbiological analysis. The strains Limosilactobacillus fermentum 5.2, Lactiplantibacillus plantarum 6.2, and L. plantarum 7.1 were selected. To evaluate the action of their metabolites, lyophilized cell-free supernatants (CFS) were used. The effect of CFS was evaluated in HT-29 intestinal lineage cells; in inhibiting the growth of the pathogen in agar; and in inhibiting the formation of biofilms. The bioprotective activity of LAB was evaluated via their potential for autoaggregation and coaggregation with E. coli. The CFS did not show cytotoxicity at lower concentrations, except for L. fermentum 5.2 CFS, which is responsible for cell proliferation at doses lower than 10 mg ml-1. The CFS were also not able to inhibit the growth of E. coli F4 in agar; however, the CFS of L. plantarum 7.1 resulted in a significant decrease in biofilm formation at a dose of 40 mg ml-1. Regarding LAB, their direct use showed great potential for autoaggregation and coaggregation in vitro, thus suggesting possible effectiveness in animal organisms, preventing E. coli fixation and proliferation. New in vitro tests are needed to evaluate lower doses of CFS to control biofilms and confirm the bioprotective potential of LAB, and in vivo tests to assess the effect of LAB and their metabolites interacting with animal physiology.

Ex-vivo mucolytic and anti-inflammatory activity of BromAc in tracheal aspirates from COVID-19.

COVID-19 is a lethal disease caused by the pandemic SARS-CoV-2, which continues to be a public health threat. COVID-19 is principally a respiratory disease and is often associated with sputum retention and cytokine storm, for which there are limited therapeutic options. In this regard, we evaluated the use of BromAc®, a combination of Bromelain and Acetylcysteine (NAC). Both drugs present mucolytic effect and have been studied to treat COVID-19. Therefore, we sought to examine the mucolytic and anti-inflammatory effect of BromAc® in tracheal aspirate samples from critically ill COVID-19 patients requiring mechanical ventilation. METHOD: Tracheal aspirate samples from COVID-19 patients were collected following next of kin consent and mucolysis, rheometry and cytokine analysis using Luminex kit was performed. RESULTS: BromAc® displayed a robust mucolytic effect in a dose dependent manner on COVID-19 sputum ex vivo. BromAc® showed anti-inflammatory activity, reducing the action of cytokine storm, chemokines including MIP-1alpha, CXCL8, MIP-1b, MCP-1 and IP-10, and regulatory cytokines IL-5, IL-10, IL-13 IL-1Ra and total reduction for IL-9 compared to NAC alone and control. BromAc® acted on IL-6, demonstrating a reduction in G-CSF and VEGF-D at concentrations of 125 and 250 µg. CONCLUSION: These results indicate robust mucolytic and anti-inflammatory effect of BromAc® ex vivo in tracheal aspirates from critically ill COVID-19 patients, indicating its potential to be further assessed as pharmacological treatment for COVID-19.

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.

Immobilization of PR4A3 enzyme in pluronic F127 polymeric micelles against colorectal adenocarcinoma cells and increase of in vitro bioavailability.

Traditional therapy for malignant neoplasms involving surgical procedures, radiotherapy and chemotherapy aims to kill neoplastic cells, but also affects normal cells. Therefore, exogenous proteases are the target of studies in cancer therapy, as they have been shown to be effective in suppressing tumors and reducing metastases. Pluronic F127 (F127) is a copolymer of amphiphilic blocks that has shown significant potential for drug administration, as it is capable of incorporating hydrophobic drugs and self-assembling in micrometers of nanometric size. This study investigated the effects of immobilization of the alkaline protease PR4A3 with pluronic F127 micelles on the enzyme-induced cytotoxicity. Protease immobilization was demonstrated through UV-visible and circular dichroism (CD) spectroscopies, as the enzyme interacts with the polymeric micelle of Pluronic F127 without changing its secondary structure. In addition, the immobilized form of the enzyme showed greater bioavailability after passing through the simulated gastrointestinal transit. Cell viability was assessed using the tetrazoic methylthiazole (MTT) assay. The results open perspectives for new research and development for PR4A3 in the treatment of colorectal carcinoma.

Fermentation in fine cocoa type Scavina: Change in standard quality as the effect of use of starters yeast in fermentation.

In the present work we aimed to demonstrate the influence of inoculum starter in support high quality fermentation. Cocoa fermentations were performed in wooden boxes and eight yeasts strains were used in separated fermentations of fine cocoa, type Scavina, as starter inoculum. Temperature, pH, titirable acidity, reducing sugar and free amino acids were evaluated during or after fermentation. The influence of starters yeasts on the decrease of acidity, sugar concentration and free amino acids was significant. The strains Candida parapsilosis, Torulaspora delbrueckii and Pichia kluyveri showed greater changes in the reducing sugar and free amino acids in fermented cocoa beans. These results indicate the ability of yeast used as inoculum starter to modify the end condition and further enhance the quality of fine cocoa beans.

Antimicrobial activity of Lactobacillus fermentum TcUESC01 against Streptococcus mutans UA159.

Dental caries is a multifactorial chronic-infection disease, which starts with a bacterial biofilm formation caused mainly by Streptococcus mutans. The use of probiotics has shown numerous health benefits, including in the fight against oral diseases. Strains of Lactobacillus fermentum have already shown probiotic potential against S. mutans, but there are still few studies. Thus, the aim of our study was to evaluate the antimicrobial activity of the inoculum and metabolites produced by L. fermentum TcUESC01 against S. mutans UA159. For this, a growth curve of L. fermentum was performed and both the inoculum and the metabolites formed in the fermentation were tested against the growth of S. mutans UA159 in agar diffusion tests, and only its metabolites were tested to determine the minimum inhibitory concentration, minimal bactericidal concentration and inhibition of cell adhesion. Inhibition of biofilm formation, pH drop and proton permeability were also tested with the metabolites. The zone of inhibition began to be formed at 14 h and continued until 16 h. The inoculum containing L. fermentum also showed zone of inhibition. The MIC for the metabolites was 1280 mg/mL and the MBC was obtained with a concentration higher than the MIC equal to 5120 mg/mL. Half of the MIC concentration (640 mg/mL) was required to inhibit S. mutans adhesion to the surface of the microplates. In the biofilm analyzes, the treatment with the metabolites in the tested concentration was not able to reduce biomass, insoluble glucans and alkali soluble compared to the control biofilm (p > 0.05). The metabolites also did not affect acid production and acid tolerance of S. mutans cells in biofilms compared to saline group (p > 0.05). Lactic acid (50.38%) was the most abundant organic acid produced by L. fermentum. This is the first report showing that the metabolites produced by the Lactobacillus fermentum TcUESC01 have a potential to be used as an antimicrobial agent against S. mutans, showing anti-adherence and bactericidal activity against planktonic cells of S. mutans. Thus, further studies should be carried out in order to better understand the antimicrobial activity of metabolites of L. fermentum TCUESC01.

Biotechnological potential of mangrove sediments: Identification and functional attributes of thermostable and salinity-tolerant ß-glucanase.

Microorganisms native to mangroves are expected to contain enzymes capable of hydrolyzing different carbon sources. However, most of these microorganisms aren't cultivable; hence, alternative techniques as metagenomics are tools for studying and obtaining some of the natural genomes, genes and enzymes of biotechnological interest. The ß-glucanase was produced using a metagenomic clone of mangrove sediments and detected by functional screening on carboxymethylcellulose substrate. The enzyme was purified by cation exchange chromatography. The peptides detected by mass spectrometry showed 20% identity with the polypeptide deduced from the genomic fragment sequenced. The ORF identified as BglfosD9 possessed 729 bp and the encoded protein showed predicted MW and pI of 28kD and 6.8, respectively. The enzyme was active in a wide range of pH (5-10) with optimum pH at 8, had relative activity greater than 50% at all temperatures tested (5-90 °C), was stable at temperatures of 5, 50 and 90 °C and showed excellent relative activity at high NaCl concentrations. This ß-glucanase also showed high relative activity in the presence of SDS and it could hydrolyze ß-glucan, CMC and Avicel as substrates. These findings support the idea of a new thermostable and active enzyme at basic pH from metagenomic library of mangrove sediment.

Aflatoxins and ochratoxin A: occurrence and contamination levels in cocoa beans from Brazil.

In the present study, the occurrence of aflatoxins (AFs) and ochratoxin A (OTA) was evaluated in 123 samples of cocoa beans produced in five Brazilian states. The presence of these mycotoxins was determined by high-performance liquid chromatography with fluorescence detection (HPLC-FLD) after immunoaffinity column clean-up. The mean level of total AFs in cocoa beans samples was 5.7 µg.kg-1. Four (3.3%) samples exceeded the maximum limit of 10 µg.kg-1 established by the Brazilian legislation for total AFs. The mean level of OTA contamination was 1.2 µg.kg-1, and none of the samples exceeded the maximum limit established by the Brazilian legislation. The co-occurrence of AFs and OTA was observed in 4.9% of the samples. The results of the present study demonstrated that, in relation to the levels of AFs and OTA established by the Brazilian legislation, most samples of cocoa beans analyzed are safe for consumption. This is the first report on the occurrence and levels of AFs and OTA in cocoa beans from the five main Brazilian states producing cocoa. The data in this study provide important information for farmers, traders, industry, consumers and law enforcement agencies.

Witches' broom resistant genotype CCN51 shows greater diversity of symbiont bacteria in its phylloplane than susceptible genotype catongo.

BACKGROUND: Theobroma cacao L. (cacao) is a perennial tropical tree, endemic to rainforests of the Amazon Basin. Large populations of bacteria live on leaf surfaces and these phylloplane microorganisms can have important effects on plant health. In recent years, the advent of high-throughput sequencing techniques has greatly facilitated studies of the phylloplane microbiome. In this study, we characterized the bacterial microbiome of the phylloplane of the catongo genotype (susceptible to witch's broom) and CCN51 (resistant). Bacterial microbiome was determined by sequencing the V3-V4 region of the bacterial 16S rRNA gene. RESULTS: After the pre-processing, a total of 1.7 million reads were considered. In total, 106 genera of bacteria were characterized. Proteobacteria was the predominant phylum in both genotypes. The exclusive genera of Catongo showed activity in the protection against UV radiation and in the transport of substrates. CCN51 presented genus that act in the biological control and inhibition in several taxonomic groups. Genotype CCN51 presented greater diversity of microorganisms in comparison to the Catongo genotype and the total community was different between both. Scanning electron microscopy analysis of leaves revealed that on the phylloplane, many bacterial occur in large aggregates in several regions of the surface and isolated nearby to the stomata. CONCLUSIONS: We describe for the first time the phylloplane bacterial communities of T. cacao. The Genotype CCN51, resistant to the witch's broom, has a greater diversity of bacterial microbioma in comparison to Catongo and a greater amount of exclusive microorganisms in the phylloplane with antagonistic action against phytopathogens.

Production of xylitol and bio-detoxification of cocoa pod husk hemicellulose hydrolysate by Candida boidinii XM02G.

The use of cocoa pod husk hemicellulose hydrolysate (CPHHH) was evaluated for the production of xylitol by Candida boidinii XM02G yeast isolated from soil of cocoa-growing areas and decaying bark, as an alternative means of reusing this type of waste. Xylitol was obtained in concentrations of 11.34 g.L-1, corresponding to a yield (Yp/s) of 0.52 g.g-1 with a fermentation efficiency (ε) of 56.6%. The yeast was tolerant to inhibitor compounds present in CPHHH without detoxification in different concentration factors, and was able to tolerate phenolic compounds at approximately 6 g.L-1. The yeast was also able to metabolize more than 99% (p/v) of furfural and hydroxymethylfurfural present in the non-detoxified CPHHH without extension of the cell-growth lag phase, showing the potential of this microorganism for the production of xylitol. The fermentation of cocoa pod husk hydrolysates appears to provide an alternative use which may reduce the impact generated by incorrect disposal of this waste.

In Vitro Activity of Lactobacilli with Probiotic Potential Isolated from Cocoa Fermentation against Gardnerella vaginalis.

Study of the probiotic potential of microorganisms isolated from fermented foods has been increasing, especially studies related to lactobacilli. In intestinal models, lactobacilli have demonstrated beneficial properties, such as anti-inflammatory activity and increased antibody production, but the molecular mechanisms involving probiotic and antagonistic action as well as their effect on human vaginal cells have not yet been fully elucidated. The aim of this study was to evaluate the functional and antagonistic properties of three strains of lactobacilli isolated from cocoa fermentation (Lactobacillus fermentum 5.2, L. plantarum 6.2, and L. plantarum 7.1) against Gardnerella vaginalis. Our results show that the lactobacilli have potential use as probiotics, since they have high hydrophobicity and autoaggregation properties and effectively adhere to vaginal cells. Metabolites secreted into the culture medium and whole cells of the strains under study are capable of interfering with the growth of G. vaginalis to different degrees. The elucidation of the antagonistic mechanisms as well as their effect on human cells may be useful in the development of a product containing such microorganisms or products secreted by them.

Functional Profile Evaluation of Lactobacillus fermentum TCUESC01: A New Potential Probiotic Strain Isolated during Cocoa Fermentation.

The use of intestinal probiotic bacteria is very common in the food industry and has been the focus of the majority of research in this field. Yet in recent years, research on extraintestinal microorganisms has greatly increased due to their well-known potential as probiotics. Thus, we studied a strain of Lactobacillus fermentum (TCUESC01) extracted from fermenting cocoa. First, we examined the impact of pH on the growth of this strain and studied its survival under conditions similar to those of the human gastrointestinal tract. L. fermentum TCUESC01 demonstrated resistance to conditions mimicking the human stomach and intestines and grew well between pH 5 and pH 7. Next, we subjected L. fermentum TCUESC01 to storage at 4°C in a milk solution and found that it survived well for 28 days. Lastly, we measured the susceptibility of this strain to numerous antibiotics and its tendency to autoaggregate. L. fermentum TCUESC01 showed significant autoaggregation, as well as susceptibility to the majority of antibiotics tested. Overall, our findings support the potential use of this extraintestinal bacterium as a dietary probiotic.

Microbiological Quality and Prevalence of ß-Lactam Antibiotic Resistance Genes in Oysters ( Crassostrea rhizophorae ).

The microbiological quality of oysters reflects the microbiological quality of their habitats because they are filter feeders. The objective of this study was to assess the bacterial composition of the edible oyster Crassostrea rhizophorae in urban and preserved estuaries. Particularly, we assessed the presence of pathogenic bacteria, investigated antibiotic susceptibility in bacterial isolates, and quantified ß-lactam antibiotic resistance genes (blaTEM, blaSHV, and blaKPC) via quantitative PCR of oyster DNA. Our results detected total coliforms, Escherichia coli , and enterobacteria in the oysters from urban estuaries, which is indicative of poor water quality. In addition, our detection of the eaeA and stxA2 virulence genes in 16.7% of E. coli isolates from oysters from this region suggests the presence of multiantibiotic-resistant enteropathogenic and enterohemorrhagic E. coli strains. During periods of low precipitation, increased contamination by E. coli (in winter) and Vibrio parahaemolyticus (in autumn) was observed. In contrast, cultivated oysters inhabiting monitored farms in preserved areas had low levels of bacterial contamination, emphasizing that oyster culture monitoring enhances food quality and makes oysters fit for human consumption. Distinct antibiotic resistance profiles were observed in bacteria isolated from oysters collected from different areas, including resistance to ß-lactam antibiotics. The presence of the blaTEM gene in 91.3% of oyster samples indicated that microorganisms in estuarine water conferred the capability to produce ß-lactamase. To our knowledge, this is the first study to directly quantify and detect ß-lactam antibiotic resistance genes in oysters. We believe our study provides baseline data for bacterial dynamics in estuarine oysters; such knowledge contributes to developing risk assessments to determine the associated hazards and consequences of consuming oysters from aquatic environments containing pathogenic bacteria that may possess antibiotic resistance genes.

Inhibition of Staphylococcus aureus biofilm by Lactobacillus isolated from fine cocoa.

BACKGROUND: Biofilm production represents an important virulence and pathogenesis factor for Staphylococcus aureus. The formation of biofilms on medical devices is a major concern in hospital environments, as they can become a constant source of infection. Probiotic bacteria, such as Lactobacillus fermentum and L. plantarum, have been found to inhibit biofilm formation; however little is known about the underlying mechanism. In this study, we tested the activity of supernatants produced by L. fermentum TCUESC01 and L. plantarum TCUESC02, isolated during the fermentation of fine cocoa, against S. aureus CCMB262 biofilm production. We measured inhibition of biofilm formation in vitro and analyzed biofilm structure by confocal and electronic microscopy. Additionally, we quantified the expression of S. aureus genes icaA and icaR involved in the synthesis of the biofilm matrix by real-time PCR. RESULTS: Both Lactobacillus supernatants inhibited S. aureus growth. However, only L. fermentum TCUESC01 significantly reduced the thickness of the biofilm, from 14 µm to 2.83 µm (at 18 mg∙mL-1, 90 % of the minimum inhibitory concentration, MIC), 3.12 µm (at 14 mg∙mL-1, 70 % of the MIC), and 5.21 µm (at 10 mg∙mL-1, 50 % of the MIC). Additionally, L. fermentum TCUESC01 supernatant modulated the expression of icaA and icaR. CONCLUSIONS: L. fermentum TCUESC01 reduces the formation of S. aureus biofilm under subinhibitory conditions. Inhibition of biofilm production probably depends on modulation of the ica operon.