Biotechnological Properties Research Articles

Phenotypic and Genomic Insights Into Bacillus spp. and Peribacillus spp. of Spanish Olive Groves With Biotechnological Potential.

Spore-forming organisms are an integral component of the rhizosphere, providing plants with significant advantages. Previous studies determined the antimicrobial activity of the olive sporobiota, identifying five candidates of particular relevance, belonging to the Bacillus subtilis, Peribacillus simplex and Bacillus cereus clade. This study aimed to determine their biotechnological properties, safety aspects, spore structure and resistance, and interaction with the environment through a combined microbiological and genomic approach. We report on the ability of these strains to produce hydrolytic and surface-active enzymes and provide evidence for differences in population behaviour through the formation of strong sessile or floating biofilms. Electron microscopic analysis revealed for the first time the presence of an exosporium layer in olive sporobiota isolates belonging to the P. simplex and B. cereus clade, including numerous pili-like structures on the latter. Spores showed significant differences in their resistance to wet heat, oxidising agents, and UV exposure. Whole genome sequencing of isolate Peribacillus frigoritolerans yielded information on its antimicrobial compound biosynthesis and environmental safety. Overall, our findings provide insights into the phenotypic, morphological and genetic variations of spore-formers from Spanish olive groves, which can be useful for the development of bioactive compounds in sustainable agriculture.

SELECTION OF LACTIC ACID BACTERIA FOR RYE BREAD SOURDOUGH

Lactic acid bacteria are widely used to produce diverse type of fermented products. One of conventional applications for lactic acid bacteria is inclusion into sourdough for bread baking. These microorganisms play a key role in dough fermentation of rye flour or wheat and rye mixture since the yield of stable and well-leavened rye dough is possible only in acidic environment. Lately, despite the development of accelerated baking technologies and technological improvements, which in some cases eliminate the need for sourdough, interest in the tradition of making bread using sourdough as a source of flavoring, nutrient and health-enhancing compounds is constantly growing across the globe. It is natural therefore that focus of the researchers is centered on seeking new strains of lactic acid bacteria attractive as potential constituents of rye bread sourdough. Screening of strains-candidates to be included in the consortia for rye flour fermentation was conducted among collection cultures (23 strains), isolates from rye flour samples (154 strains) and among craft rye dough cultures for spontaneous fermentation kindly provided by private households of Minsk, Turov, Zhitkovichi (21 strains). Identification of lactic acid bacteria following examination of morphological, cultural, physiological and biochemical characteristics of the cultures and guided by the data of MALDI TOF mass spectrometry indicated that the compared microorganisms represented genera Lactiplantibacillus, Limosilactobacillus, Pediococcus, Weissella, Lactococcus. The majority of lactic acid bacteria isolated from rye flour samples belonged to genera Lactiplantibacillus and Pediococcus. Representatives of genera Lactiplantibacillus and Levilactobacillus prevailed among domestic variants of rye bread sourdoughs. Evaluation of acidogenic capacity of analyzed cultures upon 16 h fermentation in water-flour suspension by Neumann technique and organoleptic assessment of the obtained solutions sorted out for further studies 10 strains of lactic acid bacteria referred to species Levilactobacillus brevis (4 strains) Lactiplantibacillus plantarum (3 strains) and singular strains of species Lactiplantibacillus paraplantarum, Limosilactobacillus fermentum, Weissella cibaria. Glycosyl-hydrolases produced by lactic acid bacteria constituting starter cultures largely determine biochemical processes during fermentation of flour mixtures. Chromogenic substrates o-nitrophenyl-β-D-galactopyranoside, o-nitrophenyl-α-D-galactopyranoside, 4-nitrophenyl-β-D-glucopyranoside were used in 1 mmol concentration to assay activities of β-galactosidase, α-galactosidase, β-glucosidase, respectively. It was found that all tested bacteria of species Levilactobacillus brevis and Limosilactobacillus fermentum were characterized by synthesis of proteins with α-galactosidase and β-galactosidase activities in the course of growth on media with glucose and maltose. β-glucosidase activity was revealed in all representatives of species Lactiplantibacillus plantarum, Levilactobacillus brevis and Lactiplantibacillus paraplantarum. Amylolytic activity was detected in two Lactiplantibacillus plantarum strains, whereas bacteria of species Weissella cibaria displayed a weak β-glucosidase activity. An important criterion to select microbial constituents for dough fermentation is antagonistic activity toward contaminating microbiota. Antagonistic response of lactic acid bacterial cultures to causal agents of bread spoilage was demonstrated, including epidemiologically significant fungi of genera Fusarium, Aspergillus, Penicillium, etc. Analysis of obtained data has enabled to compose microbial consortia promising for further elaboration of technologies for producing active bread-making starter formulas with upgraded biotechnological properties and enhanced alimentary, nutritional and biological value.

Improving wine fermentation efficiency of Torulaspora delbrueckii by increasing the ploidy of yeast inocula

The life cycle of most non-conventional yeasts, such as Torulaspora delbrueckii (Td), is not as well-understood as that of Saccharomyces cerevisiae (Sc). Td is generally assumed to be haploid, which detracts from some biotechnological properties compared to diploid Sc strains. We analyzed the life cycle of several Td wine strains and found that they were mainly diploid during exponential growth in rich medium. However, most cells became haploid in stationary phase, as observed for Sc haploid heterothallic strains. When transferred and incubated in nutrient-deficient media, these haploid cells became polymorphic, enlarged, and transitioned to diploid or polyploid states. The increased ploidy, that mainly results from supernumerary mitosis without cytokinesis, was followed by sporulation. A similar response was observed in yeasts that remained alive during the second fermentation of base wine for sparkling wine making, or during growth in ethanol-supplemented medium. This response was not observed in the Sc yeast populations under any of the experimental conditions assayed, which suggests that it is a specific adaptation of Td to the stressful fermentation conditions. This response allows Td yeasts to remain alive and metabolically active longer during wine fermentation. Consequently, we designed procedures to increase the cell size and ploidy of haploid Td strains. Td inocula with increased ploidy showed enhanced fermentation efficiency compared to haploid inocula of the same strains.

Raffinocyclicin is a novel plasmid-encoded circular bacteriocin produced by Lactococcus raffinolactis with broad-spectrum activity against many gram-positive food pathogens.

This study describes the discovery and characterization of raffinocyclicin, a novel plasmid-encoded circular bacteriocin, produced by the raw milk isolate Lactococcus raffinolactis APC 3967. This bacteriocin has a molecular mass of 6,092 Da and contains 61 amino acids with a three-amino acid leader peptide. It shows the highest identity to the circular bacteriocins bacicyclicin XIN-1 (42.62%), aureocyclicin 4185 (42.62%), and garvicin ML (41.53%). A broad inhibitory spectrum includes strains from Staphylococcus, Enterococcus, Streptococcus, Micrococcus, Lactobacillus, Leuconostoc, and Listeria, in addition to a pronounced inhibitory effect against Lactococcus and Clostridium. It displays low sensitivity to trypsin, most likely as a result of its circular nature. The raffinocyclicin gene cluster is composed of 10 genes: 6 core genes, genes encoding an accessory three-component ABC transporter (rafCDE), and a putative transcriptional regulator related to the MutR family. A lack of inhibitory activity in the cell-free supernatant combined with the pronounced activity of cell extracts suggests that the majority of raffinocyclicin is associated with the cell rather than being released to the extracellular environment. This is the first report of a bacteriocin produced by the L. raffinolactis species.IMPORTANCEThe present study aimed to characterize raffinocyclicin, a novel circular bacteriocin produced by the lactic acid bacteria Lactococcus raffinolactis APC 3967. Bacteriocins are generally cationic and hydrophobic peptides with antimicrobial activity, which present diverse biotechnological properties of interest for the food industry. Raffinocyclicin inhibits a wide range of bacteria, including foodborne pathogens, and is stable against different treatments which suggest its potential as a natural biopreservative. Whole-genome sequencing and the genetic analysis of the raffinocyclicin gene cluster showed that it is encoded by plasmid that could be used in the future to transfer the ability to produce the bacteriocin to other lactic acid bacteria for industrial applications. These results together highlight the potential of this novel antimicrobial as a biopreservative to be used by the food industry.

Characterization and Genome Analysis of the Delftia lacustris Strain LzhVag01 Isolated from Vaginal Discharge

Delftia has been separated from freshwater, sludge, and soil and has emerged as a novel opportunistic pathogen in the female vagina. However, the genomic characteristics, pathogenicity, and biotechnological properties still need to be comprehensively investigated. In this study, a Delftia strain was isolated from the vaginal discharge of a 43-year-old female with histologically confirmed cervical intraepithelial neoplasm (CIN III), followed by whole-genome sequencing. Phylogenetic analysis and average nucleotide identity (ANI) analysis demonstrated that it belongs to Delftia lacustris, named D. lacustris strain LzhVag01. LzhVag01 was sensitive to β-lactams, macrolides, and tetracyclines but exhibited resistance to lincoamines, nitroimidazoles, aminoglycosides, and fluoroquinolones. Its genome is a single, circular chromosome of 6,740,460 bp with an average GC content of 66.59%. Whole-genome analysis identified 16 antibiotic resistance-related genes, which match the antimicrobial susceptibility profile of this strain, and 11 potential virulence genes. These pathogenic factors may contribute to its colonization in the vaginal environment and its adaptation and accelerate the progression of cervical cancer. This study sequenced and characterized the whole-genome of Delftia lacustris isolated from vaginal discharge, which provides investigators and clinicians with valuable insights into this uncommon species.

Self-Assembled Recombinant Elastin and Globular Protein Vesicles with Tunable Properties for Diverse Applications.

Vesicles are self-assembled structures comprised of a membrane-like exterior surrounding a hollow lumen with applications in drug delivery, artificial cells, and micro-bioreactors. Lipid or polymer vesicles are the most common and are made of lipids or polymers, respectively. They are highly useful structures for many applications but it can be challenging to decorate them with proteins or encapsulate proteins in them, owing to the use of organic solvent in their formation and the large size of proteins relative to lipid or polymer molecules. By utilization of recombinant fusion proteins to make vesicles, specific protein domains can be directly incorporated while also imparting tunability and stability. Protein vesicle assembly relies on the design and use of self-assembling amphiphilic proteins. A specific protein vesicle platform made in purely aqueous conditions of a globular, functional protein fused to a glutamate-rich leucine zipper (ZE) and a thermoresponsive elastin-like polypeptide (ELP) fused to an arginine-rich leucine zipper (ZR) is discussed here. The hydrophobic conformational change of the ELP above its transition temperature drives assembly, and strong ZE/ZR binding enables incorporation of the desired functional protein. Mixing the soluble proteins on ice induces zipper binding, and then warming above the ELP transition temperature (Tt) triggers the transition to and growth of protein-rich coacervates and, finally, reorganization of proteins into vesicles. Vesicle size is tunable based on salt concentration, rate of heating, protein concentration, size of the globular protein, molar ratio of the proteins, and the ELP sequence. Increasing the salt concentration decreases vesicle size by decreasing the Tt, resulting in a shorter coacervation transition stage. Likewise, directly changing the heating rate also changes this time and increasing protein concentration increases coalescence. Increasing globular protein size decreases the size of the vesicle due to steric hindrance. By changing the ELP sequence, which consists of (VPGXG)n, through the guest residue (X) or number of repeats (n), Tt is changed, affecting size. Additionally, the chemical nature of X variation has endowed vesicles with stimuli responsiveness and stability at physiological conditions.Protein vesicles have been used for biocatalysis, biomacromolecular drug delivery, and vaccine applications. Photo-cross-linkable vesicles were used to deliver small molecule cargo to cancer cells in vitro and antigen to immune cells in vivo. pH-responsive vesicles effectively delivered functional protein cargo, including cytochrome C, to the cytosol of cancer cells in vitro, using hydrophobic ion pairing to improve cargo distribution in the vesicles and release. The globular protein used to make the vesicles can be varied to achieve different functions. For example, enzyme vesicles exhibit biocatalysis, and antigen vesicles induce antibody and cellular immune responses after vaccination in mice. Collectively, the development and engineering of the protein vesicle platform has employed amphiphilic self-assembly strategies and rational protein engineering to control physical, chemical, and biological properties for biotechnology and nanomedicine applications.

Conservation techniques for promising bacteria in the oil industry to preserve biological properties

Background: The conservation of valuable bacterial strains is crucial for various scientific, industrial, and environmental applications. Microorganisms with oil-emulsifying and oil-displacing properties are potentially significant for biotechnologies applied in the oil industry, particularly in such areas as bioremediation and tertiary enhanced oil recovery. To supply enterprises with pure cultures of microorganisms, they should be constantly maintained in the collection conditions in an active state while monitoring the preservation of their biotechnological properties. Therefore, keeping microorganism strains in working conditions and preserving their valuable properties are important for almost any work with microorganisms, ranging from primary research to their use in the production of various biopreparations.
 Aim: The article focuses on studying a method for preserving bacteria that are useful in the oil industry. This method involves modifying the technique of microencapsulating microorganism cells in alginate gel by adding glycerin, which is used as an agent with biostatic action.
 Materials and methods: The subject of research are eight hydrocarbon-oxidizing cultures of microorganisms that were sourced from the Department of Biotechnology of the al-Farabi Kazakh National University. Of these, four cultures were spore-bearing, while the other four were non-spore-bearing. The research employed microbiological methods of cultivation and storage of microorganisms in both solid and liquid media under aerobic conditions. In addition, Cooper's method was used to determine oil emulsification index and statistical methods for data analysis.
 Results: It has been found that adding glycerin (15% vol.) as a biostatic to the gel-forming matrix of sodium alginate can ensure long-term (up to 6 months) cell viability for the studied bacteria in the range of 88-96% while maintaining functionality of immobilized cells. The values of the bacteria’s oil emulsification remained at the pre-conservation levels, whereas traditional storage methods result in a lower number of viable cells after six months. It should be noted that after six months of being stored in encapsulated form with glycerin, the viability of non-sporе-forming Pseudomonas cultures is lower (88-91%) than spore-forming Bacillus (95-98%). This correlation is also is observed for traditional methods.
 Conclusion: The modern method of preserving bacteria allows for their long-term storage while maintaining functionality and viability.

Diversity and enzymatic, biosurfactant and phytotoxic activities of culturable Ascomycota fungi present in marine sediments obtained near the South Shetland Islands, maritime Antarctica.

We studied the culturable fungal community recovered from deep marine sediments in themaritime Antarctic, and assessed their capabilities to produce exoenzymes, emulsifiers and metabolites with phytotoxic activity. Sixty-eight Ascomycota fungal isolates were recovered and identified. The most abundant taxon recovered was the yeast Meyerozyma guilliermondii, followed by the filamentous fungi Penicillium chrysogenum, P. cf. palitans, Pseudeurotium cf. bakeri, Thelebolus balaustiformis, Antarctomyces psychrotrophicus and Cladosporium sp. Diversity indices displayed low values overall, with the highest values obtained at shallow depth, decreasing to the deepest location sampled. Only M. guilliermondii and P. cf. palitans were detected in the sediments at all depths sampled, and were the most abundanttaxa at all sample sites. The most abundant enzymes detected were proteases, followed by invertases, cellulases, lipases, carrageenases, agarases, pectinases and esterases. Four isolates showed good biosurfactant activity, particularly the endemic species A. psychrotrophicus. Twenty-four isolates of P. cf. palitans displayed strong phytotoxic activities against the models Lactuca sativa and Allium schoenoprasum. The cultivable fungi recovered demonstrated good biosynthetic activity in the production of hydrolytic exoenzymes, biosurfactant molecules and metabolites with phytotoxic activity, reinforcing the importance of documenting the taxonomic, ecological and biotechnological properties of fungi present in deep oceanic sediments of the Southern Ocean.

A novel thermostable YtnP lactonase from Stenotrophomonas maltophilia inhibits Pseudomonas aeruginosa virulence in vitro and in vivo

Infections caused by multidrug-resistant pathogens are one of the biggest challenges facing the healthcare system today. Quorum quenching (QQ) enzymes have the potential to be used as innovative enzyme-based antivirulence therapeutics to combat infections caused by multidrug-resistant pathogens. The main objective of this research was to describe the novel YtnP lactonase derived from the clinical isolate Stenotrophomonas maltophilia and to investigate its antivirulence potential against multidrug-resistant Pseudomonas aeruginosa MMA83. YtnP lactonase, the QQ enzyme, belongs to the family of metallo-β-lactamases. The recombinant enzyme has several advantageous biotechnological properties, such as high thermostability, activity in a wide pH range, and no cytotoxic effect. High-performance liquid chromatography analysis revealed the activity of recombinant YtnP lactonase toward a wide range of N-acyl-homoserine lactones (AHLs), quorum sensing signaling molecules, with a higher preference for long-chain AHLs. Recombinant YtnP lactonase was shown to inhibit P. aeruginosa MMA83 biofilm formation, induce biofilm decomposition, and reduce extracellular virulence factors production. Moreover, the lifespan of MMA83-infected Caenorhabditis elegans was prolonged with YtnP lactonase treatment. YtnP lactonase showed synergistic inhibitory activity in combination with gentamicin and acted additively with meropenem against MMA83. The described properties make YtnP lactonase a promising therapeutic candidate for the development of next-generation antivirulence agents.

Partial genomic characterization of Chromobacterium piscinae from India reveals multi drug resistance.

Species of genus Chromobacterium have been isolated from diverse geographical settings, which exhibits significant metabolic flexibility as well as biotechnological and pathogenic properties. This study describes the isolation, characterization, draft assembly, and detailed sequence analysis of Chromobacterium piscinae strain W1B-CG-NIBSM isolated from water samples from multi use community pond. The organism was characterized by biochemical tests, Matrix Assisted Laser Desorption/Ionization Time of Flight Mass Spectrometry (MALDI TOF-MS) and partial genome sequencing. The partial genomic data ofChromobacterium pisciane isolate W1B NIBSM strainwas submitted to GenBank with Bio project number PRJNA803347 and accession no CP092474. An integrated genome analysis of Chromobacterium piscinae has been accomplished with PATRIC which indicates good quality genome. DNA sequencing using the illumina HiSeq 4000 system generated total length of 4,155,481bp with 63 contig with G + C content is 62.69%. This partial genome contains 4,126 protein-coding sequences (CDS), 27 repeats region and 78 transfer RNA (tRNA) genes as well as 3 ribosomal RNA (rRNA) genes. The genomic annotation of Chromobacterium W1B depicts 2,925 proteins with functional assignments and 1201 hypothetical proteins. A repertoire of specialty genes implicated in antibiotic resistance (45 genes), drug target (6 genes), Transporter (3 genes) and virulence factor (10 genes). The genomic analysis reveals the adaptability, displays metabolic varied pathways and shows specific structural complex and various virulence factors which makes this strain multi drug resistant. The isolate was found to be highly resistant to β-lactam antibiotics whereas it showed sensitivity towards aminoglycosides and fluoroquinolone antibiotics. Hence, the recovery of Chromobacterium piscinae from community pond evidenced for uncertain hidden source of public health hazard. To the best of authors knowledge this is first report of isolation and genomic description of C. piscinae from India.

Assessment of Potential Probiotic Properties and Biotechnological Activities of Lactobacillus Strains Isolated from Traditional Algerian Fermented Wheat ELHAMOUM

Background: The objective of the current study was to explore the in vitro probiotic and technological characteristics of lactic acid bacteria (LAB) isolated from Algerian fermented wheat. Methods: Three LAB strains were selected and identified using 16S rRNA gene sequencing, in vitro assessments of probiotic properties included resistance to simulated gastric and intestinal conditions, auto-aggregation and antimicrobial susceptibility against common pathogens. Furthermore, the strains were evaluated for their biotechnological properties. Results: The results of 16S rRNA gene sequencing showed that two of them were Levilactobacillus brevis (LAn91 and LAn110) and LAn12a was Lactiplantibacillus pentosus. The tolerance of all strains to gastric acid and bile salts was more than 90%; they showed a high auto-aggregation capacity and a variable antagonistic property against pathogenic strains. Strain LAn12a presented the best biotechnological characteristics (proteolytic, acidifying and lactose hydrolysis), while the LAn91 and LAn110 strains were not able to acidify milk. All the strains exhibited a survival count in fermented milk exceeding 7 log CFU/ml during storage at 4°C on day 21, which surpasses the requirement of 6 log CFU/ml to provide health benefits. These bacteria can also be employed for the creation of functional foods and to validate their suitability as probiotic starter cultures.

Lipopeptides from Bacillus: unveiling biotechnological prospects-sources, properties, and diverse applications.

Bacillus sp. has proven to be a goldmine of diverse bioactive lipopeptides, finding wide-range of industrial applications. This review highlights the importance of three major families of lipopeptides (iturin, fengycin, and surfactin) produced by Bacillus sp. and their diverse activities against plant pathogens. This review also emphasizes the role of non-ribosomal peptide synthetases (NRPS) as significant enzymes responsible for synthesizing these lipopeptides, contributing to their peptide diversity. Literature showed that these lipopeptides exhibit potent antifungal activity against various plant pathogens and highlight their specific mechanisms, such as siderophore activity, pore-forming properties, biofilm inhibition, and dislodging activity. The novelty of this review comes from its comprehensive coverage of Bacillus sp. lipopeptides, their production, classification, mechanisms of action, and potential applications in plant protection. It also emphasizes the importance of ongoing research for developing new and enhanced antimicrobial agents. Furthermore, this review article highlights the need for future research to improve the production efficiency of these lipopeptides for commercial applications. It recognizes the potential for these lipopeptides to expand the field of biological pest management for both existing and emerging plant diseases.

Research of biotechnological, physical and mechanical properties and fractional composition of pressed feed bales

To develop and optimize the parameters of a compressed feed chopper device, a comprehensive understanding of the biotechnological, physical, and mechanical properties of the processed compressed feed is essential. An experiment was conducted to analyze the fractional composition of compressed straw, alfalfa, and camel thorn. The compressed feed was divided into fractions based on length categories: up to 30 mm, 30-50 mm, and 50 mm. The fractional composition was determined relative to the total mass, and size-mass indicators were analyzed. Results indicated that fractions longer than 50 mm constituted 10.42 kg or 78.2% of the straw bale, with an average mass of 13.3 kg. Fractions in the 30-50 mm range accounted for 2.34 kg or 17.6%, while fractions shorter than 30 mm made up 0.56 kg or 4.2% of the total mass. The average density of the straw bale was calculated to be 83.1 kg/m3. Subsequent experiments revealed the diameter range of stalks for straw (1.5-5.5 mm), alfalfa hay (1.5-4.0 mm), and camel thorn (2-10 mm). Samples of each feed type, ranging from 50 mm to 100 mm in length, were selected for bending and breaking experiments.

Como a pandemia da covid-19 levantou novas questões éticas e legais em relação à propriedade intelectual

The book “Saúde & propriedade intelectual”, edited by Marcos Wachowicz and Ângela Kretschmann, provides a comprehensive view of the complex ethical and legal issues related to the COVID-19 pandemic and intellectual property in the context of global health. It is a work that explores the critical intersection of intellectual property and public health, with a focus on the COVID-19 pandemic. The book examines ethical issues arising from the pandemic, particularly the conflict between private economic interests and the preservation of people’s lives. It emphasizes the need to rethink intellectual property in biotechnology, considering its social function and pointing to the necessity of balancing public and private interests within the patent system. The work also delves into the Brazilian National Public Health System and international negotiations regarding the intellectual protection of genetic resources, along with the challenges of patenting medicines in resource-constrained countries. Due to its wide coverage, this publication is essential reading for academics, public health experts, legal professionals, and anyone interested in the intersection of intellectual property and public health.

Biotechnological properties and antifungal activity of lactic acid bacteria isolated from two marine algae Ulva lactuca and Sargassum muticum collected from the Moroccan coast of Sidi Bouzid–El Jadida

Biotechnological properties and antifungal activity of lactic acid bacteria isolated from two marine algae Ulva lactuca and Sargassum muticum collected from the Moroccan coast of Sidi Bouzid–El Jadida

Antifungal activity of eukaryotic microalgae in dermatophytes

Introduction: Microalgae are considered a promising source of biologically active secondary metabolites, with several biotechnological properties already recorded, including antifungal activity. Objective: Thus, the properties of ethanol extracts of microalgae Ankistrodesmus falcatus, Chaetoceros neogracilis, Desmodesmus brasiliensis, Dunaliella tertiolecta, Kirchneriella lunaris and Tetraselmis gracilis were investigated in vitro for antifungal activity against dermatophytes Nannizzia gypsea, Nannizzia nana, Trichophyton mentagrophytes and Trichophyton tonsurans. Methods: The extracts were evaluated using broth microdilution methodology, with a test interval of 0.0115 to 6 mg mL-1. The antifungal itraconazole was tested at concentrations 0.0313 to 16 μg mL-1. Results: All microalgae extracts showed antifungal activity, especially extracts from C. neogracilis, D. brasiliensis and K. lunaris that totally inhibited growth of all species of dermatophytes evaluated. The lowest MIC values recorded were for the extracts of D. brasiliensis and K. lunaris (MIC 0.188 mg mL-1), against T. tonsurans e N. nana. Conclusions: This was a pioneering work about the antifungal activity of these microalgae against dermatophytes. It is expected that, from this study, further research will be carried out in order to identify and isolate the active biomolecules responsible for the antifungal activity of these microalgae species.

Characteristics of the metabolite complex produced L.reuteri LR1

Introduction: Lactic acid microorganisms have valuable biotechnological and probiotic properties. Probiotic properties are often mediated by biologically active metabolites produced by these microorganisms. The ability of lactic acid microorganisms to produce biologically active compounds has received special attention in recent years, as their positive effect on the human body has been determined. However, there is a huge knowledge gap regarding the composition of metabolite complexes that requires study to ensure their safe use.Purpose: The current study of the metabolite complex produced by L. reuteri LR1, obtained by cultivating the strain in the MRS broth nutrient medium at a temperature of (37±1)ºС for 24 hours.Materials and Methods: The object of research was a cell-free supernatant (metabolite complex) obtained by cultivating the Lactobacillus reuteri LR1 strain from the collection of the Federal State Scientific Institution “VNIMI”. The antimicrobial activity of MK L. reuteri LR1 against E. coli ATCC 25922, S. aureus ATCC 6538, S. typhimurium ATCC 14028 was determined by the agar diffusion method. The antioxidant activity of the samples was determined using the ORAC fluorescence method. The content of organic and amino acids in LA was determined by capillary electrophoresis. Identification of secondary metabolites present in MK was carried out using gas chromatography-mass spectrometry (GC-MS).Results: The biological activity of MK produced by L. reuteri LR1 was determined. The composition of the cell-free metabolite complex secreted by L.reuteri LR1 during cultivation in the MRS broth nutrient medium at a temperature of 37°C for 24 hours was characterized. The presence of amino acids and organic acids in LA was confirmed and their content was determined. In addition, secondary metabolites present in MK have been identified, some of them have confirmed biological activity.Conclusion: The results obtained may be useful for predicting the probiotic potential of MK, however, it is necessary to determine the likely correlation between the composition of MK and its beneficial properties, which will allow us to identify new possibilities for the use of MK produced by probiotic microorganisms.

Antilisterial activity of raw sheep milk from two native Epirus breeds: Culture-dependent identification, bacteriocin gene detection and primary safety evaluation of the antagonistic LAB biota

Raw milk from native small ruminant breeds in Epirus, Greece, is a valuable natural source of autochthonous lactic acid bacteria (LAB) strains with superior biotechnological properties. In this study, two bulk milks (RM1, RM2) from two local sheep yards, intended for traditional Kefalotyri cheese production, were preselected for bacteriocin-like antilisterial activity by in vitro tests. Their antagonistic LAB biota was quantified followed by polyphasic (16S rRNA gene sequencing; IGS for Enterococcus; a multiplex-PCR for Leuconostoc) identification of 42 LAB (RM1/18; RM2/24) isolates further evaluated for bacteriocin encoding genes and primary safety traits. Representative isolates of the numerically dominant mesophilic LAB were Leuconostoc mesenteroides (10) in both RMs, Streptococcus parauberis (7) in RM2, and Lactococcus lactis (1) in RM1; the subdominant thermophilic LAB isolates were Enterococcus durans (8), E. faecium (6), E. faecalis (3), E. hirae (1), E. hermanniensis (1), Streptococcus lutetiensis (2), S. equinus (1) and S. gallolyticus (1). Based on their rpoB, araA, dsr and sorA profiles, six Ln. mesenteroides strains (8 isolates) were atypical lying between the subspecies mesenteroides and dextranicum, whereas two strains profiled with Ln. mesenteroides subsp. jonggajibkimchii that is first-time reported in Greek dairy food. Two RM1 E. faecium strain biotypes (3 isolates) showed strong, enterocin-mediated antilisterial activity due to entA/entB/entP possession. One E. durans from RM1 possessed entA and entP, while additional nine RM2 isolates of the E. faecium/durans group processed entA or entP singly. All showed direct (cell-associated) antilisterial activity only, as also both S. lutetiensis strains from RM2 did strongly. Desirably, no LAB isolate was β-hemolyrtic, or cytolysin-positive, or possessed vanA, vanB for vancomycin resistance, or agg, espA, hyl, and IS16 virulence genes. However, all three E. faecalis from RM2 possessed gelE and/or ace virulence genes. In conclusion, all Ln. mesenteroides strains, the two safe, enterocin A-B-P-producing E. faecium strains, and the two antilisterial S. lutetiensis strains should be validated further as potential costarter or adjunct cultures in Kefalotyri cheese. The prevalence of α-hemolytic pyogenic streptococci in raw milk, mainly S. parauberis in RM2, requires consideration in respect to subclinical mastitis in sheep and the farm hygiene overall.

In vitro antimicrobial activity of the crude extract of the endophytic bacterium Pseudomonas aeruginosa (SS93) isolated from Sapindus saponaria L.

Endophytes colonize the interior of plant tissues without causing any damage to their hosts. The plant Sapindus saponaria L., popularly known as ‘sabão-de-soldado’, presents a diversified endophytic microbiota and also medicinal properties. Endophytic microorganisms may produce secondary metabolites with different biotechnological properties. The present study aimed to evaluate the in vitro antibacterial and antifungal capacity of the crude extract of secondary metabolites produced by the endophytic bacteria P. aeruginosa SS93 isolated from S. saponaria leaves. The metabolites extract was obtained using the organic solvent ethyl acetate, and the antimicrobial activities were tested against six pathogenic bacteria (Enterococcus faecalis [ATCC 29212], Pseudomonas aeruginosa [ATCC 27853], Shigella flexneri [ATCC 12022], Salmonella enterica [CCCD a016], Escherichia coli [ATCC 25922], and Staphylococcus aureus [ATCC 25923]), and pathogenic fungi (Fusarium oxysporum, Glomerella sp., Sphaceloma sp., Fusarium solani, Maniliophtora perniciosa, and Sclerotinia sclerotiorum), by agar diffusion method. In the antibacterial assay, the best results were obtained against E. faecalis and S. aureus, where the formation of inhibition halos was observed in all tested concentrations, especially at 500 and 700 µg mL-1. Positive inhibitory activity against phytopathogenic fungi was observed, with the highest inhibition recorded against F. oxysporum (61.1%), followed by Sphaceloma sp. (55.7%), M. perniciosa (35.6%), F. solani (34.4%), and Glomerella sp. (30.4%).

Biodiversity and biotechnological properties of lactic acid bacteria isolated from traditional Moroccan sourdoughs.

The present study aimed at characterizing lactic acid bacteria (LAB) strains isolated from traditional sourdoughs collected in different regions of Morocco. Isolated strains were firstly identified using Gram staining and catalase reaction test. Presumptive LAB strains were then checked for various phenotypical properties including growth at 45°C, resistance to NaCl, enzyme production, acidification capacity, diacetyl and exopolysaccharide (EPS) production, and antifungal activity. Finally, selected LAB strains were identified using 16S rDNA sequencing. Results showed that 32.1% of the isolates were thermophilic (45°C) and 83.9% were resistant to NaCl (6.5%). Moreover, 51.7 and 37.5% were able to produce diacetyl and EPS, respectively. Regarding enzyme production, 55.3 and 7.1% of the isolates showed lipolytic and proteolytic activities, respectively. Low pH values (3.37-3.76) were obtained after 24h of incubation of LAB strains in de Man, Rogosa and Sharpe (MRS) broth. Antifungal activity test against Aspergillus flavus, Aspergillus niger and Penicillium spp. showed an inhibition rate up to 50%. Bacterial DNA sequencing showed that LAB isolates belong to seven species, chiefly Levilactobacillus brevis, Lentilactobacillus parabuchneri, Lactiplantibacillus plantarum, Pediococcus pentosaceus, Enterococcus hirae, Bifidobacterium pseudocatenulatum, and Companilactobacillus paralimentarius. These findings, for the first time in Moroccan sourdoughs, indicate that the isolated LAB strains have good multifunctional properties and could be suitable as good starters for sourdough bread production under controlled conditions.