Screening and isolation of microbes from a Mud Community of Ischia Island Thermal Springs: preliminary analysis of a bioactive compound

Introduction: Balneotherapy centers of Ischia island (Italy) offer treatments for different dermatological diseases (psoriasis, acne, atopic dermatitis) and upper respiratory tract infections. In this study, we integrated morphological and molecular approaches to give bacterial diversity of microbial mat samples collected from the thermae of Ischia. Methods: Samples were collected during 2019 at four sites. Some bacterial strains ATCC for antibacterial and antibiofilm Activity were tested. After morphological characterization, screening for antagonistic isolates was made. The colonies isolated from thermal mud samples were submitted to molecular characterization. Susceptibility testing by dilution spotting was carried out and antibacterial efficacies of most active isolate were evaluated with a Minimal inhibition concentration assay. Biofilm formation, inhibition, eradication were examined. Statistical analyses were carried out utilizing Microsoft® Excel 2016/XLSTAT©-Pro. Results: We isolated a natural compound with antimicrobial and antibiofilm activities. Conclusions: The results obtained in this study are discussed in the context of how hydrothermal systems are important environmental source of uncharted antimicrobial and antibiofilm compounds. This is, to our knowledge, the first view of a spring water microbiome analysis of Ischia

A preliminary study on a novel bioaugmentation technique enhancing lactic acid production by mixed cultures fermentation

The paper is a preliminary study on the selection of lactic acid producing microorganisms from a mixed microbial population via bioaugmentation. The bioaugmentation technique is based on pH sudden variations occurring in sequential batch steps of a dark fermentation process applied to simple substrates. Different conditions are tested and compared. The structure of microbial communities and concentrations of metabolic intermediates are analyzed to study the possible substrate conversion routes. Obtained results indicate that the initial mixed culture produced a lactic acid percentage of 5% in terms of CODLA/CODPRODUCTS. In the most favourable conditions, the selected culture produced a lactic acid percentage of 59%. The analysis of the composition of microbial communities before and after the bioaugmentation processes, indicates that lactic acid production mainly results from the population change to bacteria belonging to the genus Bacillus. Indeed, the relative abundance of Bacilli increased from 0.67%, to 8.40% during the bioaugmentation cycle

An Ecotoxicological Evaluation of Four Fungal Metabolites with Potential Application as Biocides for the Conservation of Cultural Heritage

Biocides based on chemical synthetic compounds have been commonly used to counteract damages caused by microorganisms on stone cultural heritage. However, in the last few years, the use of commercial and traditional biocides has been banned and/or limited due to their dangerous profile for the environment, as well as human and animal health. Natural products could be used as suitable alternatives for cultural heritage purposes, as they have low toxicity and stability compared with synthetic pesticides. Even if most of the investigated solutions have already shown promising results, their efficiency, ecotoxicological, and chemical features are poorly investigated. In this manuscript, we aimed to evaluate the ecotoxicological profile of four fungal metabolites—namely, cavoxin, epiepoformin, seiridin, and sphaeropsidone—with potential antimicrobial properties for monumental artworks. A battery of ecotoxicological tests using Aliivibrio fischeri (bacterium), Raphidocelis subcapitata (alga), Raphanus sativus L. (macrophyte), Daphnia magna (crustacean), and Caenorhabditis elegans (nematode) revealed a relative lower toxicity of these compounds, especially when compared with Preventol® and Rocima®, commercial biocides mainly used for the conservation of cultural heritage

Allium ursinum and Allium oschaninii against Klebsiella pneumoniae and Candida albicans Mono- and Polymicrobic Biofilms in In Vitro Static and Dynamic Models

The present study assesses the in vitro antibiofilm potential activity of extracts of wild Allium ursinum and Allium oschaninii. The active ingredients of the extracts were obtained with a technique named Naviglio (rapid solid–liquid dynamic extraction, RSLDE) which is based on an innovative and green solid–liquid extraction methodology. The extracts were tested against models of mono‐ and polymicrobial biofilm structures of clinically antibiotic‐resistant pathogens, Klebsiella pneumoniae ATCC 10031 and Candida albicans ATCC 90028. Biofilms were studied using a static and a dynamic model (microtiter plates and a CDC reactor) on three different surfaces reproducing what happens on implantable medical devices. Antimicrobic activities were determined through minimum inhibitory concentration (MIC), while antibiofilm activity was assessed by minimum biofilm eradication concentration (MBEC) using a crystal violet (CV) biofilm assay and colony forming unit (CFU) counts. Results showed that both Allium extracts eradicated biofilms of the tested microorganisms well; biofilms on Teflon were more susceptible to extracts than those on polypropylene and polycarbonate, suggesting that when grown on a complex substrate, biofilms may be more tolerant to antibiotics. Our data provide significant advances on antibiotic susceptibility testing of biofilms grown on biologically relevant materials for future in vitro and in vivo applications

Anti-Biofilm Activity of Phenyllactic Acid against Clinical Isolates of Fluconazole-Resistant Candida albicans

: Commonly found colonizing the human microbiota, Candida albicans is a microorganism known for its ability to cause infections, mainly in the vulvovaginal region, and is responsible for 85% to 90% of vulvovaginal candidiasis (VVC) cases. The development of drug resistance in C. albicans isolates after long-term therapy with fluconazole is an important complication to solve and new therapeutic strategies are required to target this organism and its pathogenicity. In the present study, phenyllactic acid (PLA) an important broad-spectrum antimicrobial compound was investigated for its antifungal and antivirulence activities against clinical isolates of C. albicans. Previously characterized strains of C. albicans isolates from women with VVC and C. albicans ATCC90028 were used to evaluate the antimicrobial and time dependent killing assay activity of PLA showing a MIC 7.5 mg mL-1 and a complete reduction of viable Candida cells detected by killing kinetics after 4 h of treatment with PLA. Additionally, PLA significantly reduced the biomass and the metabolic activity of C. albicans biofilms and impaired biofilm formation also with changes in ERG11, ALS3, and HWP1 genes expression as detected by qPCR. PLA eradicated pre-formed biofilms as showed also with confocal laser scanning microscopy (CLSM) observations. Furthermore, the compound prolonged the survival rate of Galleria mellonella infected by C. albicans isolates. These results indicate that PLA is a promising candidate as novel and safe antifungal agents for the treatment of vulvovaginal candidiasis

Evaluation of Microbial Communities of Bottled Mineral Waters and Preliminary Traceability Analysis Using NGS Microbial Fingerprints

The microbiological monitoring of mineral bottled waters results is crucial for the prevention of outbreaks in consumers. European and International regulations establish the quality of water intended for human consumption in order to preserve human health from the negative effects deriving from water contamination. Advanced methods targeting the faster detection of potential pathogens in drinking water may consent to the creation of an early warning system, enhancing water quality management. This study aimed to suggest the implementation of standard water quality evaluations, based on the characterization of the microbial composition of mineral bottled water brands, contributing to the periodic control of the water’s microbiological stability along with the shelf life, and, consequently, the stability of the supplying sources. Bottled water microbiota analysis was combined with the qualitative and quantitative evaluation of microbial loads in time, and the monitoring was performed in two seasons and two different storage conditions for a total of sixty days. The employment of molecular microbiology techniques (NGS and Sanger sequencing), compared to standardized cultural methods and integrated with metagenomic analysis, combining chemical and physical indicators for each sample, allowing for the generation of specific fingerprints for mineral bottled waters, pointing at simplifying and improving the foreseen risk assessment strategies to ensure the adequate traceability, quality and safety management of drinking water

Chemical Composition and Biological Activities of Oregano and Lavender Essential Oils

Folk medicine uses wild herbs, especially from the Lamiaceae family, such as oregano and lavender, in the treatment of many diseases. In the present study, we investigated the antibacterial activity of the essential oils of Origanum glandulosum Desf. and Lavandula dentata L. against multidrug- resistant Klebsiella pneumoniae strains. The chemical composition of essential oils and their effect on the ultrastructure of the tested bacteria and on the release of cellular components that absorb at 260 nm were studied. Furthermore, the cytotoxicity and the production of reactive oxygen species in human lymphocytes treated with essential oils were evaluated. Thymol (33.2%) was the major constituent in O. glandulosum, and β-pinene (17.3%) was the major constituent in L. dentata. We observed ultrastructural damage in bacteria and increased release of cellular material. Furthermore, ROS production in human lymphocytes treated with essential oils was lower than in untreated lymphocytes and no cytotoxicity was observed. Therefore, the essential oils of lavender and oregano could be used as a source of natural antibacterial and antioxidant agents with potential pharmacological applications

Metabolomic profiling and molecular characterization of food matrices: identification of potential markers of microbial contamination

The research aims to generate an early warning system able to highlight, in real time, bacterial contamination of meat matrices and to provide information which could support companies in accepting or rejecting batches. Current microorganisms detection methods rely on techniques (plate counting), which provide retrospective values for microbial contamination. The disposal of fast headspace air measurement, using gas chromatography-mass spectrometry analysis, able to accurately and rapidly (30 min per sample) detect microbial spoilage in raw meat, could result a valid replacement to traditional and time-consuming (3 to 4 days) standardized microbiological analysis required by regulations. The experiments focused on the qualitative and quantitative evaluation of volatile organic compounds (VOCs) produced by Salmonella Typhimurium, Campylobacter jejuni and Staphylococcus aureus in different types of raw meat (beef, pork, chicken). The reduction of analysis times represents the strength of the alternative method to ISO protocols, although these are currently the official procedures provided by International regulations. The applied method requires smaller sample aliquots and does not need any sample processing, thus consenting its application on different food matrices (not only meat) and for the detection of a wide variety of pathogens. The validation of the suggested analytical approach would therefore result innovative, by evaluating further samples in order to demonstrate the benefits of the technique in terms of times, costs and preservation of consumers health. Data analysis allowed the characterization of unique VOC profiles and possible marker compounds of meat contamination due to certain pathogens. The identification of marker volatile compounds resulted essential to outline specific metabolic profiles for each microorganism responsible of meat spoilage

Prevalence, Distribution, and Diversity of Salmonella spp. in Meat Samples Collected from Italian Slaughterhouses

Recently worldwide food safety authorities indicated the rise of foodborne outbreaks linked to Salmonella: this highlighted the need to intensify monitoring and apply more targeted controls to help manage the spread of the disease. The aim of this study was to assess the prevalence and distribution of Salmonella serotypes in 7 slaughterhouses, located in different areas of Naples province (Regione Campania, Italy). Meat samples collected from the slaughterhouses were submitted for standardized microbiological analysis in 2015. Results of routine testing for Salmonella spp. were analyzed and then compared to biochemical and molecular evaluations. Salmonella spp. were detected in 12% of 320 samples examined (39/320) and the isolation rates ranged from 87% (32 samples) for raw poultry meat to 13% (7 samples) for pork meat. Biochemical serotyping showed that approximately 50% of the isolates belonged to Salmonella enterica serotype Choleraesuis. Rapid detection methods, such as molecular analysis (polymerase chain reaction and gel electrophoresis), able to confirm food matrices contamination, represent a valid support to the fast identification of Salmonella species. A further aspect of the study consisted, indeed, on analyzing isolated strains through molecular evaluations. By amplifying bacterial DNA-using invA primers, selective for Salmonella-it was possible, in less than 3 h, to classify the isolates as Salmonella spp., confirming the results of microbiological outcomes. Results of distribution analysis, supported by rapid molecular approaches, showed the difficulty of reducing Salmonella risk on food chain. This emphasized the importance of periodic surveillance to prevent outbreaks

Rapid and Positive Effect of Bicarbonate Addition on Growth and Photosynthetic Efficiency of The Green Microalgae Chlorella Sorokiniana (Chlorophyta, Trebouxiophyceae)

Bicarbonate ions are the primary source of inorganic carbon for autotrophic organisms living in aquatic environments. In the present study, we evaluated the short‐term (hours) effects of sodium bicarbonate (NaHCO3) addition on the growth and photosynthetic efficiency of the green algae Chlorella sorokiniana (211/8k). Bicarbonate was added to nonaxenic cultures at concentrations of 1, 2, and 3 g L‐1 leading to a significant increase in biomass especially at the highest salt concentration (3 g L‐1) and also showing a bactericidal and bacteriostatic effect that helped to keep a reduced microbial load in the algal culture. Furthermore, bicarbonate stimulated the increase in cellular content of chlorophyll a, improving the photosynthetic performance of cells. Since microalgae of genus Chlorella spp. show great industrial potential for the production of biofuels, nutraceuticals, cosmetics, health, and dietary supplements and the use of bicarbonate as a source of inorganic carbon led to short‐term responses in Chlorella sorokiniana, this method represents a valid alternative not only to the insufflation of carbon dioxide for the intensive cultures but also for the production of potentially bioactive compounds in a short period