Indicator Strains Research Articles

4CMenB Breadth of Immune Response, Immunogenicity, and Safety: Results from a Phase 3 Randomized, Controlled, Observer Blind Study in Adolescents and Young Adults

Abstract Background Meningococcal serogroup B (MenB) strains are highly diverse. Breadth of immune response for the MenB vaccine, 4CMenB, administered at 0-2, 0-6, or 0-2-6 months, was demonstrated by endogenous complement-human serum bactericidal antibody (enc-hSBA) assay against an epidemiologically-relevant panel of 110 MenB strains. Methods In a phase 3 trial, 3651 healthy 10–25 year-old participants were randomized 5:5:9:1 to receive 4CMenB (0-6 schedule), 4CMenB (0-2-6 schedule), investigational MenABCWY vaccine, or control MenACWY-CRM vaccine. The primary objectives were to evaluate safety and demonstrate breadth of immune response by enc-hSBA assay against the MenB strain panel using test-based (percentage of samples without bactericidal activity against strains after 4CMenB versus control vaccination) and responder-based (percentage of participants whose post-vaccination sera kill ≥70% strains) approaches. Success was demonstrated with two-sided 97.5% confidence interval (CI) lower limit >65%. Immunogenicity was assessed by traditional hSBA assay against four indicator strains. Results Breadth of immune response (test-based) was 78.7% (97.5% CI 77.2%–80.1%), 81.8% (80.4%–83.1%), 83.2% (81.9%–84.4%) for the 0-2, 0-6, and 0-2-6 schedules, respectively, and (responder-based) 84.8% (81.8%–87.5%), 89.8% (87.2%–92.0%), and 93.4% (91.2%–95.2%), respectively. No clinically relevant differences in immunogenicity were observed across schedules. 4CMenB was well tolerated. Conclusions The two-dose (0-2, 0-6) 4CMenB schedules met pre-defined criteria for success for both breadth of immune response endpoints against a diverse MenB strain panel, had comparable immunogenicity, and safety in line with the established 4CMenB safety profile. The three-dose schedule provided no additional immunological benefit, supporting use of the 4CMenB 0-2 schedule. Clinical Trial Registration NCT04502693.

Synthetic plantaricins show significantly enhanced antibacterial activity against Flavobacterium sp.

The culture supernatant of Lactiplantibacillus plantarum KM2 exhibited antimicrobial activity against seven foodborne pathogens. Genomic analysis revealed the presence of an operon capable of producing plantaricins, specifically bacteriocins known as plantaricins A, E, F, J, and K. In this study, genomic comparisons confirmed that this plantaricin operon was strain-specific to L. plantarum rather than universally present in all strains of the species. Genes corresponding to plantaricins A, E, F, J, and K were identified and their syntheses confirmed, leading to the verification of their antimicrobial activities. Synthesized plantaricins were prepared individually and in combinations of two or three. Their antimicrobial activities were tested against nine indicator strains. The results showed variations in the magnitude and efficacy of their antimicrobial activity, depending on the combination used. Among them, the combination of synthesized plantaricins spPlnAEJ and spPlnAEK showed the greatest effect against Flavobacterium sp. KCCM 11374. To elucidate the mechanism of antimicrobial activity, Raman spectra were examined. Notably, the Raman spectra of Flavobacterium sp. KCCM 11374 treated with synthesized plantaricins differed quite a bit from the untreated spectra, suggesting a distinct effect. Additionally, transmission electron microscopy results revealed that these synthesized plantaricins could induce the cell lysis of Flavobacterium sp. KCCM 11374.

Discovery of <i>Brevibacterium</i> predominating in fecal samples from three children with persistent diarrhea negative for common pathogens

Acute diarrhea is a common disease in children under 5 years old and can develop into persistent diarrhea, greatly affecting the children's health. Although advanced techniques had been used to diagnose and detect common pathogens in hospitals, however, 40% of cases are negative for the pathogens. In this study, to investigate dominant bacteria in stool samples of three persistent-diarrheal children with unidentified pathogenic agents, the V3 and V68 regions of the 16S rRNA gene were amplified from fecal bacterial metagenomic DNA, separated on DGGE gel, and the dominant DNA bands were sequenced. As a result, the V3 and V68 regions of bacteria in persistent diarrheal children were less diverse and different from the corresponding DNA bands of the indicator strains. Sequence analysis and similarity comparison of six DNA bands of V3 regions and seven DNA bands of V68 regions showed that two V3 sequences (of 160 bp) derived from two samples were novel and did not match any genes from the non-redundant database, but they shared 93.75% similarity to each other. The four V3 sequences left derived from all three samples were the most similar (94.53-100%) with the corresponding genes of Brevibacterium. Six of the seven V68 sequences derived from dominant DNA bands of all three samples were the most similar (from 99.4% to 100%) to the corresponding genes of referent strains belonging to the genus Enterococcus. In sample D3, a sequence of the V68 region possessed 100% identity to the E. faecalis ATCC 19433 strain. This is the first study report that Brevibacterium was the dominant bacteria in the gastrointestinal microflora of children with persistent diarrhea although the bacterial genus has been reported to cause dangerous diseases in humans with immunodeficiency.

Calcium Sulfate Disks for Sustained-Release of Amoxicillin and Moxifloxacin for the Treatment of Osteomyelitis.

The purpose of this in vitro study was to develop calcium sulfate (CS)-based disks infused with an antimicrobial drug, which can be used as a post-surgical treatment modality for osteomyelitis. CS powder was embedded with 10% antibiotic, amoxicillin (AMX) or moxifloxacin (MFX), to form composite disks 11 mm in diameter that were tested for their degradation and antibiotic release profiles. For the disk degradation study portion, the single drug-loaded disks were placed in individual meshes, subsequently submerged in phosphate-buffered saline (PBS), and incubated at 37 °C. The disks were weighed once every seven days and analyzed via Fourier-transform infrared spectroscopy, X-ray diffraction, energy dispersive X-ray spectroscopy, and scanning electron microscopy. During the antibiotic release analysis, composite disks were placed in PBS solution, which was changed every 3 days, and analyzed for antibiotic activity and efficacy. The antibacterial effects of these sustained-release composites were tested by agar diffusion assay using Streptococcus mutans (S. mutans) UA 159 as an indicator strain. The degradation data showed significant increases in the degradation of all disks with the addition of antibiotics. Following PBS incubation, there were significant increases in the amount of phosphate and decreases in the amount of sulfate. The agar diffusion assay demonstrated that the released concentrations of the respective antibiotics from the disks were significantly higher than the minimum inhibitory concentration exhibited against S. mutans over a 2-3-week period. In conclusion, CS-antibiotic composite disks can potentially serve as a resorbable, osteoconductive, and antibacterial therapy in the treatment of bone defects and osteomyelitis.

Pseudocin 196, a novel lantibiotic produced by Bifidobacterium pseudocatenulatum elicits antimicrobial activity against clinically relevant pathogens

ABSTRACT Bacteriocins are broad or narrow-spectrum antimicrobial compounds that have received significant scientific attention due to their potential to treat infections caused by antibiotic-resistant pathogenic bacteria. The genome of Bifidobacterium pseudocatenulatum MM0196, an antimicrobial-producing, fecal isolate from a healthy pregnant woman, was shown to contain a gene cluster predicted to encode Pseudocin 196, a novel lantibiotic, in addition to proteins involved in its processing, transport and immunity. Following antimicrobial assessment against various indicator strains, protease-sensitive Pseudocin 196 was purified to homogeneity from cell-free supernatant. MALDI TOF mass spectrometry confirmed that the purified antimicrobial compound corresponds to a molecular mass of 2679 Da, which is consistent with that deduced from its genetic origin. Pseudocin 196 is classified as a lantibiotic based on its similarity to lacticin 481, a lanthionine ring-containing lantibiotic produced by Lactococcus lactis. Pseudocin 196, the first reported bacteriocin produced by a B. pseudocatenulatum species of human origin, was shown to inhibit clinically relevant pathogens, such as Clostridium spp. and Streptococcus spp. thereby highlighting the potential application of this strain as a probiotic to treat and prevent bacterial infections.

Biosynthesis of bacteriocin BacZY05-silver nanoconjugates and evaluation of their antibacterial properties.

Bacteriocins are antimicrobial peptides produced by bacteria to prevent the growth of pathogens. Combining bacteriocins with metal nanoparticles, like silver nanoparticles (AgNPs), has developed into a viable strategy to get over bacteriocin limitations. In this study, bacteriocin BacZY05 was extracted from Bacillus subtilis ZY05 and purified using various techniques. The resulting purified bacteriocin was then combined with silver nanoparticles to form bacteriocin silver nanoconjugates (BacZY05-AgNPs). The physicochemical properties of the BacZY05-AgNPs were characterized using various analytical techniques. The mean diameter of the synthesized AgNPs was approximately 20-60nm with an oval or spherical shape. The antimicrobial activity of the BacZY05-AgNPs was evaluated against several indicator strains by their zone of inhibition (ZOI), using the agar well diffusion method. Compared to bacteriocin (ZOI- 13 to 20mm) and AgNPs (ZOI- 10-22mm) alone, the antibacterial activity data demonstrated a 1.3-1.5-fold increase in the activity of bacteriocin-nanoconjugates (ZOI- 22 to 26mm). For Staphylococcus aureus MTCC3103 and Klebsiella pneumoniae MTCC109, BacZY05-capped AgNPs exhibited the lowest minimum inhibitory concentration (MIC), measuring 10.93µg/mL. For Salmonella typhi NCIM2501, the MIC was 28.75µg/mL. The highest MIC value was 57.5µg/mL for Escherichia coli DH5α and Vibrio cholerae MTCC3909. With BacZY05-capped AgNPs, the lowest minimum bactericidal concentration (MBC) of 28.75µg/mL was observed for Staphylococcus aureus MTCC31003. In the cases of Salmonella typhi NCIM2501 and Klebsiella pneumoniae MTCC109 concentration was 57.5µg/mL. Vibrio cholerae MTCC3909 and Escherichia coli DH5α had the highest MBC values at 115µg/mL.

Characterization of carotenoids extracted from Haloferax larsenii NCIM 5678 isolated from Pachpadra salt lake, Rajasthan.

Carotenoids are a diverse group of pigments known for their broad range of biological functions and applications. This study delves into multifaceted potential of carotenoids extracted from Haloferax larsenii NCIM 5678 previously isolated from Pachpadra Salt Lake in Rajasthan, India. H. larsenii NCIM 5678 was able to grow up to OD600 1.77 ± 0.03 with carotenoid concentration, 3.3 ± 0.03µg/ml. The spectrophotometric analysis of carotenoid extract indicated the presence of three-fingered peak (460, 490 and 520nm) which is a characteristic feature of bacterioruberin and its derivatives. The bacterioruberin was purified using silica gel column chromatography and thin layer chromatography. The carotenoid extract showed 12.3 ± 0.09mm zone of growth inhibition with a minimum inhibitory concentration 546ng/ml against indicator strain, H. larsenii HA4. The percentage antioxidant activity of carotenoid was found to be 84% which was higher as compared to commercially available ascorbic acid (56.74%). Thus, carotenoid extract from H. larsenii NCIM 5678 possesses unique attributes with compelling evidence of antimicrobial and antioxidant potential for the development of novel pharmaceuticals and nutraceuticals.

Fermentation Performance Evaluation of Lactic Acid Bacteria Strains for Sichuan Radish Paocai Production.

Fermented vegetable products play a significant role in various cuisines, and understanding the fermentation dynamics of lactic acid bacteria (LAB) strains is essential for optimizing their production and quality. Here, we sought to investigate the fermentation performance of five LAB strains isolated from Sichuan paocai as starters for paocai. Sensory evaluation revealed that the inoculation of radish paocai samples with LAB strains effectively improved the overall liking and sensory satisfaction of participants, increasing the scores to varying degrees in terms of taste, flavor, texture, and coloration. Lactiplantibacillus plantarum and Lacticaseibacillus rhamnosus exhibited a good salt resistance in radish juice and could grow in a medium containing 10% NaCl. Four indicator strains commonly found in contaminated paocai were effectively inhibited by fermented LAB broths, which improved the edibility and safe production of paocai. Compared to spontaneous fermentation (CK), radish paocai inoculated with LAB showed a significantly accelerated acid production rate, shortening the fermentation period by approximately two days. The contents of titratable total acids, organic acids, and free amino acids were higher in the inoculated samples and were enriched in the taste of radish paocai. The content of volatile organic compounds in the inoculated samples was higher than that in CK. Based on OPLS-DA analysis, 31 key indicators of paocai quality were screened and used to rank the fermentation performances of the five strains using the TOPSIS method; here, Lpb. plantarum and Lcb. rhamnosus achieved the highest scores. This study provides a reference for selecting LAB strains as efficient and secure fermentation starters to optimize paocai quality.

Methicillin-resistant Staphylococcus aureus and coagulase-negative Staphylococcus produce antimicrobial substances against members of the skin microbiota in children with atopic dermatitis.

Coagulase-negative Staphylococcus (CoNS) species inhibiting Staphylococcus aureus has been described in the skin of atopic dermatitis (AD) patients. This study evaluated whether Staphylococcus spp. from the skin and nares of AD and non-AD children produced antimicrobial substances (AMS). AMS production was screened by an overlay method and tested against NaOH, proteases and 30 indicator strains. Clonality was assessed by pulsed-field gel electrophoresis. Proteinaceous AMS-producers were investigated for autoimmunity by the overlay method and presence of bacteriocin genes by polymerase chain reaction. Two AMS-producers had their genome screened for AMS genes. A methicillin-resistant S. aureus (MRSA) produced proteinaceous AMS that inhibited 51.7% of the staphylococcal indicator strains, and it was active against 60% of the colonies selected from the AD child where it was isolated. On the other hand, 57 (8.8%) CoNS from the nares and skin of AD and non-AD children, most of them S. epidermidis (45.6%), reduced the growth of S. aureus and other CoNS species. Bacteriocin-related genes were detected in the genomes of AMS-producers. AMS production by CoNS inhibited S. aureus and other skin microbiota species from children with AD. Furthermore, an MRSA colonizing a child with AD produced AMS, reinforcing its contribution to dysbiosis and disease severity.

Newly Isolated Limosilactobacillus reuteri B1/1 Modulates the Expression of Cytokines and Antimicrobial Proteins in a Porcine ex Vivo Model.

The epithelia of the intestine perform various functions, playing a crucial role in providing a physical barrier and an innate immune defense against infections. By generating a "three-dimensional" (3D) model of cell co-cultures using the IPEC-J2 cell line and porcine blood monocyte-derived macrophages (MDMs), we are getting closer to mimicking the porcine intestine ex vivo.Methods: The effect of Limosilactobacillus reuteri B1/1 and Limosilactobacillus fermentum CCM 7158 (indicator strain) on the relative gene expression of interleukins (IL-1β, IL-6, IL-8, IL-18 and IL-10), genes encoding receptors for TLR4 and TLR2, tight junction proteins such as claudin-1 (CLDN1), occludin (OCLN) and important antimicrobial proteins such as lumican (LUM) and olfactomedin-4 (OLMF-4) was monitored in this model. The results obtained from this pilot study point to the immunomodulatory potential of newly isolated L. reuteri B1/1, as it was able to suppress the enhanced pro-inflammatory response to lipopolysaccharide (LPS) challenge in both cell types. L. reuteri B1/1 was even able to up-regulate the mRNA levels of genes encoding antimicrobial proteins LUM and OLFM-4 and to increase tight junction (TJ)-related genes CLDN1 and OCLN, which were significantly down-regulated in LPS-induced IPEC-J2 cells. Conversely, L. fermentum CCM 7158, chosen as an indicator lactic acid bacteria (LAB) strain, increased the mRNA levels of the investigated pro-inflammatory cytokines (IL-18, IL-6, and IL-1β) in MDMs when LPS was simultaneously applied to basally deposited macrophages. Although L. fermentum CCM 7158 induced the production of pro-inflammatory cytokines, synchronous up-regulation of the anti-inflammatory cytokine IL-10 was detected in both LAB strains used in both cell cultures. The obtained results suggest that the recently isolated LAB strain L. reuteri B1/1 has the potential to alleviate epithelial disruption caused by LPS and to influence the production of antimicrobial molecules by enterocytes.

Effect of UV pretreatment on the source control of floR during subsequent biotreatment of florfenicol wastewater.

UV photolysis has been recommended as an alternative pretreatment method for the elimination of antibacterial activity of antibiotics against the indicator strain, but the pretreated antibiotic intermediates might not lose their potential to induce antibiotic resistance genes (ARGs) proliferation during subsequent biotreatment processes. The presence of florfenicol (FLO) in wastewater seriously inhibits the metabolic performance of anaerobic sludge microorganisms, especially the positive correlation between UV irradiation doses and ATP content, while it did not significantly affect the organics utilization ability and protein biosynthetic process of aerobic microorganisms. After sufficient UV pretreatment, the relative abundances of floR from genomic or plasmid DNA in subsequent aerobic and anaerobic biotreatment processes both decreased by two orders of magnitude, maintained at the level of the groups without FLO selective pressure. Meanwhile, the abundances of floR under anaerobic condition were always lower than that under aerobic condition, suggesting that anaerobic biotreatment systems might be more suitable for the effective control of target ARGs. The higher abundance of floR in plasmid DNA than in genome also indicated that the potential transmission risk of mobile ARGs should not be ignored. In addition, the relative abundance of intI1 was positively correlated with floR in its corresponding genomic or plasmid DNA (p < 0.05), which also increased the potential horizontal transfer risk of target ARGs. This study provides new insights into the effect of preferential UV photolysis as a pretreatment method for the enhancement of metabolic performance and source control of target ARGs in subsequent biotreatment processes. KEY POINTS: • Sufficient UV photolytic pretreatment efficiently controlled the abundance of floR • A synchronous decrease in abundance of intI1 reduced the risk of horizontal transfer • An appreciable abundance of floR in plasmid DNA was a potential source of total ARGs.

Comparative Study of Bioactive Compounds and Biological Activities of Five Rose Hip Species Grown in Sicily.

Nowadays, research on plant extracts has attracted increasing interest. The aim of this study was to compare phenolic profile, vitamin C, and carotenoid content, as well as the biological activities of five different rose species, including Rosa canina, R. corymbifera, R. micrantha, R. rubiginosa, and R. rugosa. These species had different morphological characteristics, with R. rugosa showing higher size of flower petals and higher weight of hips. The highest vitamin C content was found in hip extracts of R. rubiginosa and R. rugosa, which also showed the highest carotenoid amount. R. corymbifera showed the highest phenolic content. No significant antimicrobial activity of extracts containing phenolic compounds against different indicator strains could be detected. Cell monolayer integrity was not affected by treatments with the above-mentioned extracts of R. canina, R. micrantha, and R. rugosa at different concentrations for up to 24 h, while those of R. rubiginosa and R. corymbifera affected intestinal permeability at the highest concentration tested. The partial least squares regression analysis generated a predictive model correlating phenolic compounds with cell monolayer integrity, suggesting a relevant role for catechin, quercitrin, and p-coumaric acid. In conclusion, this study highlights how rose hips belonging to different species can have a diverse phenolic profile, differently influencing intestinal monolayer integrity.

Development of a fumonisin-sensitive Saccharomyces cerevisiae indicator strain and utilization for activity testing of candidate detoxification genes.

Fumonisins can cause diseases in animals and humans consuming Fusarium-contaminated food or feed. The search for microbes capable of fumonisin degradation, or for enzymes that can detoxify fumonisins, currently relies primarily on chemical detection methods. Our constructed fumonisin B1-sensitive yeast strain can be used to phenotypically detect detoxification activity and should be useful in screening for novel fumonisin resistance genes and to elucidate fumonisin metabolism and resistance mechanisms in fungi and plants, and thereby, in the long term, help to mitigate the threat of fumonisins in feed and food.

ANTIMYCOTIC ACTIVITY OF THE ISOLATES OF LACTOBACTERIA FROM WA-TER AND MUSSELS OF BLACK SEA

Aim. Determination of the presence of antimycotic activity and its level in thirteen lactobacteria isolates from Black Sea water and mussels. Methods. Antimycotic activity of lactic acid bacteria (LAB) isolates was tested by agar diffusion method in 24-well plates. To determine character of action the plates were incubated for 7 days at 25 °C. Results. For the first time antimycotic activity of marine LAB from Odesa coast was estimated. The majority of the tested isolates of lactobateria exhibited antimycotic activity of high level completely inhibiting (in 100%) as mycelium growth as well as spore formation of Penicillium expansum UKM F-575 and Aspergillus niger UKM F-16706 on second day of the study. Four isolates (M.5.1, M.5.2, M.7.1, M.7.2) showed lower antimycotic activity (from 0 to 75%). Eight LAB isolates from seawater (W.1.1, W.1.2, W.1.3, W.1.4, W.1.5, W.1.дк, W.2.3, W.2.4) and one isolate from mussel liquor (M4.1) completely inhibited mycelial growth and sporulation of P. expansum UКМ F-575 and A. niger UКМ F-16706 even within seven days indicating fungicidal character of action. Conclusions. The most of tested LAB isolates from Black Sea exhibited high antimycotic activity against P. expansum UКМ F-575 and A. niger UКМ F-16706 indicator strains. The most active lactobacteria isolates were W.1.1, W.1.2, W.1.3, W.1.4, W.1.5, W.1.дк, W.2.3, W.2.4 isolated from Black Sea water and M4.1 – from mussel liquor. The character of the revealed antimycotic activity was determined as fungicidal.

Study of a combined biological product with antibacterial and probiotic activity

Scientific relevance. The issue of antimicrobial resistance has acquired global significance, urging the search for and use of alternative antibacterial agents to treat infectious diseases. In particular, this situation prompts wider use of biotechnology-derived medicinal products based on bacteriophages and probiotics, including those of the metabolite type. The bacteriotropic (antimicrobial and probiotic) properties of pharmaceutical compositions based on bacteriophages and metabolites of probiotic bacteria suggest a qualitatively new combined antibacterial effect, as well as effectiveness against microorganisms resistant to antibiotics.Aim. The authors aimed to study the properties of a combined biological product with antibacterial and probiotic effects.Materials and methods. The study focused on a combined biological product consisting of a probiotic agent based on Lactobacillus exometabolites and a complex bacteriophage. The antimicrobial activity evaluation involved the Appelmans method, a disc diffusion method using lawn cultures homologous (Staphylococcus spp., Pseudomonas aeruginosa, Escherichia coli, Proteus spp., Enterococcus spp., Salmonella spp., and Shigella spp.) and non-homologous (Acinetobacter baumannii, Klebsiella pneumoniae, and Yersinia enterocolitica) to the complex bacteriophage, and a bioluminescence inhibition test using the genetically modified indicator strain E. coli lum+. The study used bacterial strains isolated from various clinical samples and biotopes in bacteriological laboratories of healthcare institutions in the Perm Territory in 2019–2022. The probiotic activity was assessed by the stimulating effect on model microorganisms of the normal flora.Results. The Appelmans method showed that the combined biological product had high antimicrobial activity against microorganisms homologous to the complex bacteriophage. The titres calculated for the combined biological product and its complex bacteriophage component were comparable and amounted to 10–6.6±0.01 and 10–6.9±0.01, respectively. The disc diffusion method demonstrated that the combined biological product had a more pronounced antibacterial effect on all tested strains than its individual components. The optical density values obtained with the combined biological product were 1.5 and 2 times higher than the values observed with control samples in Bifidobacterium bifidum 1 and Lactobacillus plantarum 8P-А3 cultures, respectively, which demonstrated the stimulating effect of the product on the normal flora. The study results suggest the compatibility of the phage and probiotic components of the combined biological product, as well as its high antimicrobial activity.Conclusions. The novel combined biological product has high specific and non-specific antimicrobial activity, which consists in the inhibition of pathogenic bacteria growth without affecting the normal flora. The combined biological product broadens the range of medicinal products having probiotic and antibacterial effects, particularly on microorganisms resistant to antibiotics.

Effect of lactic acid bacteria by different concentrations of copper based on non-target metabolomic analysis.

Copper (Cu) is an essential element for mammals, but excess intake can have detrimental health consequences. However, Cu is no longer present in the "Limit of Contaminants in Foods" promulgated in 2022. The potential impact of different Cu (II) concentrations on human health remains unclear. In this study, a strain of lactic acid bacteria (LAB), namely, Lactiplantibacillus plantarum CICC 23121 (L23121), was selected as a prebiotic indicator strain to indirectly assess the effects of food-limited Cu (II) concentrations (issued by Tolerance limit of copper in foods in 1994) on the functions of intestinal microbes. We used non-target metabolomics, automatic growth curve detector, scanning electron microscopy (SEM), and Fourier transform infrared spectroscopy (FTIR) to investigate the effects of Cu (II) on L23121. The study revealed shows that the 50% minimum inhibitory concentration (MIC50) of Cu (II) for most lactic acid bacteria was 4 mg/L. At low Cu (II) concentrations (≤ 4 mg/L), the pentose phosphate pathway and pyrimidine metabolism of the lactic acid bacteria were affected, resulting in a decrease in the content of beneficial secondary metabolites and a significant decrease in the cell activity. As Cu (II) concentrations increase (≥ 6 mg/L), the key amino acid and lipid metabolisms were affected, leading to the inhibition of growth and primary metabolite production of the bacteria. Under high concentration of Cu (II) (6 mg/L), the surface adhesion of the bacteria was distorted and covered with significantly large particles, and the functional groups of the cells were significantly shifted. As a probiotic, the abundance of lactic acid bacteria in the intestine is significantly reduced, which will inevitably seriously damage intestinal homeostasis. Thus, to protect human intestinal microbes' health, it is recommended to limit the concentration of Cu in food to less than 4 mg/L.

A Bacteriophage Cocktail Reduces Five Relevant Salmonella Serotypes at Low Multiplicities of Infection and Low Temperatures.

Salmonella are important pathogenic bacteria and, following Campylobacter, they are the second most common cause of bacterial foodborne infections worldwide. To reduce the presence of bacteria along the food chain, the application of bacteriophages (phages) may be a promising tool. In this study, the lytic properties of six phages against five relevant Salmonella serotypes (S. Enteritidis, S. Typhimurium, S. Infantis, S. Paratyphi B and S. Indiana) were analyzed. Three phages were able to lyse all five serotypes. We determined the lytic potential of each phage on indicator strains in vitro at room temperature (RT) and at 37 °C using low multiplicities of infection (MOIs). Most phages reduced their host more efficiently at RT than at 37 °C, even at the lowest MOI of 0.001. Following this, the lytic activity of a cocktail comprising five phages (MOI = 0.1) was examined with each of the five serotypes and a mix of them at RT, 15, 12, 10, 8 and 6 °C. All cultures of single serotypes as well as the mixture of strains were significantly reduced at temperatures as low as 8 °C. For single serotypes, reductions of up to 5 log10 units and up to 2.3 log10 units were determined after 6 h (RT) and 40 h (8 °C), respectively. The mixture of strains was reduced by 1.7 log10 units at 8 °C. The data clearly suggest that these phages are suitable candidates for biocontrol of various Salmonella serotypes under food manufacturing conditions.

Selection and Characterization of Bacteriocinogenic Lactic Acid Bacteria from the Intestine of Gilthead Seabream (Sparus aurata) and Whiting Fish (Merlangius merlangus): Promising Strains for Aquaculture Probiotic and Food Bio-Preservation.

This study sought to evaluate the probiotic properties and the food preservation ability of lactic acid bacteria isolates collected from the intestines of wild marine fishes (gilthead seabream (Sparus aurata) (n = 60) and whiting fish (Merlangius merlangus) (n = 40)) from the Mediterranean sea in the area of Mostaganem city, Algeria. Forty-two isolates were identified as: Enterococcus durans (n = 19), Enterococcus faecium (n = 15), Enterococcus faecalis (n = 4), Lactococcus lactis subp. lactis (n = 3), and Lactobacillus plantarum (n = 1). All isolates showed inhibition to at least one indicator strain, especially against Listeria monocytogenes, Staphylococcus aureus, Paenibacillus larvae, Vibrio alginolyticus, Enterococcus faecalis, Bacillus cereus, and Bacillus subtilis. In all collected isolates, PCR analysis of enterocin-encoding genes showed the following genes: entP (n = 21), ent1071A/B (n = 11), entB (n = 8), entL50A/B (n = 7), entAS48 (n = 5), and entX (n = 1). Interestingly, 15 isolates harbored more than one ent gene. Antimicrobial susceptibility, phenotypic virulence, and genes encoding virulence factors were investigated by PCR. Resistance to tetracycline (n = 8: tetL + tetK), erythromycin (n = 7: 5 ermA, 2 msrA, and 1 mef(A/E)), ciprofloxacin (n = 1), gentamicin (n = 1: aac(6')-aph(2″)), and linezolid (n = 1) were observed. Three isolates were gelatinase producers and eight were α-hemolytic. Three E. durans and one E. faecium harbored the hyl gene. Eight isolates showing safety properties (susceptible to clinically relevant antibiotics, free of genes encoding virulence factors) were tested to select probiotic candidates. They showed high tolerance to low pH and bile salt, hydrophobicity power, and co-culture ability. The eight isolates showed important phenotypic and genotypic traits enabling them to be promising probiotic candidates or food bio-conservers and starter cultures.

Identification and characterisation of antimicrobial compound produced by probiotic Alkalihalobacillus clausii 088AE.

In the present study, Alkalihalobacillus clausii 088AE was evaluated for the production of antimicrobial compounds, their characterisation, and identification. The results showed that, 48-h-old A. clausii supernatant is able to inhibit the growth of indicator bacteria Micrococcus luteus MTCC 106T (7.0 ± 0.51 mm). The cultivation of 088AE on solid media along with XAD16N beads for 5 days, and isopropanol extraction of antimicrobial compound thereof showed enhanced antimicrobial activity (19.67 ± 0.58 mm) against indicator strain. Further purification with a reversed-phase C18 cartridge yielded a powder with 389.12 ± 10.08 μg of protein per mg. The UV spectra of the sample revealed the characteristic aromatic amino acid peaks at 262 and 276 nm. The results of triple-quadrupole mass spectrometry showed a peak of 1055 m/z, confirming the existence of 10542+, i.e., 2108 Da, suggestive of class I lantibiotic clausin. Furthermore, in silico analysis confirmed the presence of genes responsible for the production of lanthipeptide clausin. The clausin is stable in the presence of proteases (trypsin and pepsin), high temperature (100 °C), pH up to 11 and showed antimicrobial activity against Listeria monocytogenes ATCC 19115, M. luteus, and Staphylococcus aureus ATCC 6538P. Therefore, it could be useful to control gut infections caused by Gram-positive pathogens.

Metabolomic Profiling of Lactobacillus plantarum Isolated from Ocimum gratissimum Rhizosphere

L. plantarum is a known member of the lactic acid bacteria that possesses antimicrobial activities. Organic acids, hydrogen peroxide and bacteriocins have been majorly identified as the contributors to its antimicrobial activity. The present study sought to employ the GC-MS technique to further classify biological compounds that comprise the antimicrobial metabolite produced by L. plantarum through sub-merged fermentation. The bacterium was isolated from the rhizosphere of O. gratissimum and confirmed using molecular typing. Preliminary antibacterial screening of the organism was done with indicator strains isolated from urinary tract and wound surface, after which sub-merged fermentation was employed for the production of secondary metabolites for a 24 h period. The GC-MS technique was employed to identify the volatile bioactive compounds that comprise the secondary metabolite produced. The organism had significant (p&lt;0.05) inhibition of the indicator strains when compared to the ciprofloxacin standard antibiotic. Metabolomics analyses identified Hydroxylamine, O-decyl-, 2,4-Di-tert-butylphenol, and a wide range of organic compounds mainly from the alkane, amine, carboxylic acids and phenol functional groups, as the components of its antibacterial metabolite. GC-MS based metabolomics analyses is a profitable tool for identifying key components of the antibacterial substance produced by L. plantarum as this will give a room for its bio-prospecting potentials as alternative and sustainable source of novel antimicrobial compounds and other beneficial medications used by humans.