ABSTRACT
Modern broiler breeds allow for high feed efficiency and rapid growth, which come at a cost of increased susceptibility to pathogens and disease. Broiler growth rate, feed efficiency, and health are affected by the composition of the gut microbiota, which in turn is influenced by diet. In this study, we therefore assessed how diet composition can affect the broiler jejunal gut microbiota. A total of 96 broiler chickens were divided into four diet groups: control, coated butyrate supplementation, medium-chain fatty acid supplementation, or a high-fibre low-protein content. Diet groups were sub-divided into age groups (4, 12 and 33 days of age) resulting in groups of 8 broilers per diet per age. The jejunum content was used for metagenomic shotgun sequencing to determine the microbiota taxonomic composition at species level. The composed diets resulted in a total of 104 differentially abundant bacterial species. Most notably were the butyrate-induced changes in the jejunal microbiota of broilers 4 days post-hatch, resulting in the reduced relative abundance of mainly Enterococcus faecium (-1.8 l2fc, Padj = 9.9E-05) and the opportunistic pathogen Enterococcus hirae (-2.9 l2fc, Padj = 2.7E-08), when compared to the control diet. This effect takes place during early broiler development, which is critical for broiler health, thus exemplifying the importance of how diet can influence the microbiota composition in relation to broiler health. Future studies should therefore elucidate how diet can be used to promote a beneficial microbiota in the early stages of broiler development.
Subject(s)
Animal Feed , Chickens , Enterococcus faecium , Enterococcus hirae , Gastrointestinal Microbiome , Jejunum , Animals , Chickens/microbiology , Chickens/growth & development , Enterococcus faecium/genetics , Gastrointestinal Microbiome/drug effects , Jejunum/microbiology , Diet/veterinary , Metagenomics/methods , Dietary SupplementsABSTRACT
Microbial exopolysaccharides (EPS) are high molecular weight polymeric substances with great diversity and variety of applications in the food and pharma industry. In this study, we report the extraction of an EPS from Enterococcus hirae OL616073 strain originally isolated from Indian fermented food and its purification by ion exchange and size exclusion chromatography for physical-functional analyses. The EPS showed two prominent fractions (EPS F1 and EPS F2) with molecular mass 7.7 × 104 and 6.5 × 104 Da respectively by gel permeation chromatography. These fractions were further characterized by FTIR, HPTLC, GC-MS, and NMR as a homopolysaccharide of glucose linked with α-(1 â 6) and α-(1 â 3) glycosidic linkages. The porous, spongy, granular morphology of EPS was observed under scanning electron microscopy. EPS has revealed strong physico-functional properties like water solubility index (76.75 %), water contact angle (65.74°), water activity (0.35), hygroscopicity (3.05 %), water holding capacity (296.19 %), oil holding capacity (379.91 %), foaming capacity (19.58 %), and emulsifying activity (EA1-72.22 %). Rheological analysis showed that aqueous solution of EPS exhibited a non-Newtonian fluid behavior and shear-thinning characteristics. Overall, EPS exhibits techno functional properties with potential applications as a functional biopolymer in food and pharma industry.
Subject(s)
Enterococcus hirae , Glucans , Glucans/chemistry , Solubility , Molecular Weight , Water/chemistry , Polysaccharides, Bacterial/chemistryABSTRACT
Soil salinization has become a prominent obstacle in diverse arid and semi-arid region damaging agricultural productivity globally. From this perspective, present investigation was aimed to compare the potential compatible consortium of bio-inoculants for improving Plant Growth Promoting (PGP) attributes, anti-oxidative enzymes, grain yield and profitability of Vigna radiata in saline soil conditions. A total of 101 rhizobacterium isolated from salt affected regions of Punjab, India were screened for their ability to induce salt tolerance, multifunctional PGP traits and antagonistic activities. The 16S rRNA sequencing identified the strains LSMR-29 and LSMRS-7 as Pseudomonas flourescens and Enterococcus hirae, respectively. In-vitro compatible halo-tolerant dual inoculant (LSMR-29 + LSMRS-7) as bio-inoculants mitigated salt stress in Vigna radiata (spring mungbean) seedling with improved seed germination, biomass and salt tolerance index together with the presence of nifH, acds, pqq and ipdc gene under salinity stress as compared to single inoculants. Further, the potential of single and dual bio-inoculants were also exploited for PGP attributes in pot and field experiments. Results indicated that a significant improvement in chlorophyll content (2.03 fold), nodulation (1.24 fold), nodule biomass (1.23 fold) and leghemoglobin content (1.13 fold) with dual inoculant of LSMR-29 + LSMRS-7 over the LSMR-29 alone. The concentrations of macro & micronutrients, proline, soil enzyme activities i.e. soil dehydrogenase, acid & alkaline phosphatases and antioxidant enzymes such as superoxide dismutase, catalase and peroxidase also found to be high for LSMR-29 + LSMRS-7 as compared to un-inoculated control. The high grain yield thereby leading to Benefit: Cost (B: C) ratio at field scale was indicative of the commercial use bio-inoculants under salt affected Vigna radiata (spring mungbean) to improvement of productivity and soil health. The current finding reveals a co-inoculation of halo-tolerating Pseudomonas fluorescens and Enterococcus hirae containing ACC deaminase could prove to be novel approach for inducing salt tolerance and improving productivity of Vigna radiata (spring mungbean).
Subject(s)
Pseudomonas fluorescens , Vigna , Enterococcus hirae/genetics , RNA, Ribosomal, 16S/genetics , Salt Stress , SoilABSTRACT
The vacuolar-type ATPase from Enterococcus hirae (EhV-ATPase) is a thus-far unique adaptation of V-ATPases, as it performs Na+ transport and demonstrates an off-axis rotor assembly. Recent single molecule studies of the isolated V1 domain have indicated that there are subpauses within the three major states of the pseudo three-fold symmetric rotary enzyme. However, there was no structural evidence for these. Herein we activate the EhV-ATPase complex with ATP and identified multiple structures consisting of a total of six states of this complex by using cryo-electron microscopy. The orientations of the rotor complex during turnover, especially in the intermediates, are not as perfectly uniform as expected. The densities in the nucleotide binding pockets in the V1 domain indicate the different catalytic conditions for the six conformations. The off-axis rotor and its' interactions with the stator a-subunit during rotation suggests that this non-uniform rotor rotation is performed through the entire complex.
Subject(s)
Vacuolar Proton-Translocating ATPases , Vacuolar Proton-Translocating ATPases/metabolism , Enterococcus hirae/metabolism , Cryoelectron Microscopy , Rotation , CatalysisABSTRACT
Animal farming copiously generates indoles, which contribute to odor and pose a challenge for deodorization. While biodegradation is widely accepted, there is a lack of suitable indole-degrading bacteria for animal husbandry. In this study, we aimed to construct genetically engineered strains with indole-degrading abilities. Enterococcus hirae GDIAS-5 is a highly efficient indole-degrading bacterium, which functions via a monooxygenase YcnE presumably contributes to indole oxidation. However, the efficiency of engineered Escherichia coli expressing YcnE for indole degradation is lower than that of GDIAS-5. To improve its efficacy, the underlying indole-degradation mechanisms in GDIAS-5 were analyzed. An ido operon that responds to a two-component indole oxygenase system was identified. In vitro experiments showed that the reductase component of YcnE, YdgI, can improve the catalytic efficiency. The reconstruction of the two-component system in E. coli exhibited higher indole removal efficiency than GDIAS-5. Furthermore, isatin, the key intermediate metabolite in indole degradation, might be degraded via a novel isatin-acetaminophen-aminophenol pathway involving an amidase whose coding gene is located near the ido operon. The two-component anaerobic oxidation system, upstream degradation pathway, and engineering strains investigated in this study provide important insights into indole degradation metabolism and offer efficient resources for achieving bacterial odor elimination.
Subject(s)
Isatin , Enterococcus hirae/metabolism , Escherichia coli/genetics , Escherichia coli/metabolism , Indoles/metabolismABSTRACT
BACKGROUND: Bloodstream infections are a matter of concern in small animal veterinary practice. Few reports are avaiable, especially regarding the role of opportunistic bacteria in becoming infectious. This report aims to add to the current veterinary literature on two opportunistic bacterial species (Enterococcus hirae and Enterobacter xiangfangensis) associated with bloodstream infections in small animals admitted to the Bologna University Veterinary Hospital. CASE PRESENTATION: In the first case, a 15-year-old, immunocompromised, cardiopathic dog was admitted to the hospital for anorexia and diarrhea. The patient had a history of previous surgery and hospitalization. After three days, hyperthermia, leukopenia and hyperlactatemia were recorded, and blood culture revealed positivity for Enterococcus hirae, identified using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). The patient's general conditions progressively worsened, and the patient was euthanized. In the second case, a 2-year-old cat with chronic ocular herpesvirus infection and hypertrophic cardiomyopathy was admitted to the hospital for anorexia and hyperthermia. The cat was hospitalized one week before and received antimicrobial treatment for urinary tract infection by Staphylococcus felis. Hypokalemia and lymphopenia were also diagnosed. The patient progressively improved and was discharged after three days. On the same day, blood culture taken at admission revealed positivity for Enterobacter xiangfangensis, identified using MALDI-TOF MS. After five days, the patient returned with neurological symptoms, hypothermia and bradycardia, and was euthanized. CONCLUSIONS: In small animal veterinary practice, the impact of opportunistic bacterial agents (such as E.hirae and E.xiangfangensis) on bloodstream infections remains unclear. As in human medicine, they can be contracted in every healthcare setting and considered hospital-acquired infections. In this report, we highlighted the threat they pose especially in patients with multiple risk factors. Rapid and accurate diagnostic tools (such as MALDI-TOF MS) could be particularly important for reducing the severity of the infections.
Subject(s)
Dog Diseases , Sepsis , Humans , Animals , Dogs , Enterococcus hirae , Pets , Anorexia/veterinary , Sepsis/diagnosis , Sepsis/veterinary , Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization/veterinary , Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization/methodsABSTRACT
l-Lactate oxidase (LOx) is a flavin mononucleotide (FMN)-dependent triose phosphate isomerase (TIM) barrel fold enzyme that catalyzes the oxidation of l-lactate using oxygen as a primary electron acceptor. Although reductive half-reaction mechanism of LOx has been studied by structure-based kinetic studies, oxidative half-reaction and substrate/product-inhibition mechanisms were yet to be elucidated. In this study, the structure and enzymatic properties of wild-type and mutant LOxs from Enterococcus hirae (EhLOx) were investigated. EhLOx structure showed the common TIM-barrel fold with flexible loop region. Noteworthy observations were that the EhLOx crystal structures prepared by co-crystallization with product, pyruvate, revealed the complex structures with "d-lactate form ligand," which was covalently bonded with a Tyr211 side chain. This observation provided direct evidence to suggest the product-inhibition mode of EhLOx. Moreover, this structure also revealed a flip motion of Met207 side chain, which is located on the flexible loop region as well as Tyr211. Through a saturation mutagenesis study of Met207, one of the mutants Met207Leu showed the drastically decreased oxidase activity but maintained dye-mediated dehydrogenase activity. The structure analysis of EhLOx Met207Leu revealed the absence of flipping in the vicinity of FMN, unlike the wild-type Met207 side chain. Together with the simulation of the oxygen-accessible channel prediction, Met207 may play as an oxygen gatekeeper residue, which contributes oxygen uptake from external enzyme to FMN. Three clades of LOxs are proposed based on the difference of the Met207 position and they have different oxygen migration pathway from external enzyme to active center FMN.
Subject(s)
Enterococcus hirae , Flavin Mononucleotide , Catalytic Domain , Enterococcus hirae/metabolism , Flavin Mononucleotide/chemistry , Kinetics , Lactates , Ligands , Mixed Function Oxygenases/chemistry , Oxygen , Pyruvic Acid , Triose-Phosphate Isomerase/metabolismABSTRACT
Probiotic attributes of lactic acid bacteria isolated from goat and sheep milk samples were analysed by culturing them on an MRS agar medium. The most potential isolates, GMB24 and SMB16, were identified by biochemical tests which had ability to tolerate different concentrations of acid and bile and phenol resistance. They were further identified as Enterococcus faecium GMB24 and Enterococcus hirae SMB16 by 16S rRNA gene sequencing approach. The probiotic potential of the isolates GMB24 and SMB16 were recorded including antimicrobial activity against pathogenic bacteria viz., Escherichia coli (MTCC118), Staphylococcus aureus (MTCC7443), Pseudomonas aeruginosa (MTCC424), Listeria monocytogens (MTCC657) and Salmonella typhimurium (MTCC733), and antibiotic susceptibility test. The isolates SMB16 and GMB24 exhibited a higher zone of inhibition against P. aeruginosa (19.00 ± 0.57 mm) and S. aureus (25.66 ± 0.88 mm), respectively. The data from these experiments were used for the principal component analysis (PCA) to assess the survivability of the isolates under different factors. The heatmap generated in this study clustered the bacterial isolates based on their phenotype properties. Further, immunomodulating activities of these probiotic bacteria were tested on neutrophil adhesion test, haemagglutinating antibody titer and delayed-type hypersensitivity. Probiotic E. faecium GMB24 and E. hirae SMB16, at 109 cells/mL doses per day, increased the neutrophil adhesion, haemagglutinating antibody titer and DTH in comparison to the untreated control group. The isolates showed negative test for haemolytic and gelatinase activities and hence were considered safe. E. faecium GMB24 and E. hirae SMB16 were shown to have high probiotic potential and immune-stimulant action.
Subject(s)
Enterococcus faecium , Probiotics , Animals , Enterococcus faecium/genetics , Enterococcus hirae/genetics , Goats , Milk/microbiology , Probiotics/pharmacology , RNA, Ribosomal, 16S/genetics , Sheep , Staphylococcus aureus/geneticsABSTRACT
Young rabbits are susceptible to gastrointestinal diseases caused by bacteria. Enterococcus hirae can be associated with diseases. But enterocins produced by some enterococcal species can prevent/reduce this problem. Therefore, the interaction of enterocin M with a biofilm-forming, autochthonous E. hirae Kr8+ strain was tested in rabbits to assess enterocin potential in vivo. Rabbits (96), aged 35 days, both sexes, meat line M91 breed were divided into four groups, control C and three experimental groups. The rabbits in C received the standard diet, rabbits in experimental group 1 (E1) received 108 CFU/mL of Kr8+, a dose 500 µL/animal/day, E2 received Ent M (50 µL/animal/day), and E3 received both Kr8+ and Ent M in their drinking water over 21 days. The experiment lasted 42 days. Feces and blood were sampled at day 0/1 (at the start of the experiment, fecal mixture of 96 animals, n = 10), at day 21 (five fecal mixtures per group, n = 5), and at day 42 (21 days after additives cessation, the same). At days 21 and 42, four rabbits from each group were slaughtered, and cecum and appendix were sampled for standard microbial analysis. Ent M showed decreased tendency of Kr8+. Using next-generation sequencing, the phyla detected with the highest abundance were Firmicutes, Verrucomicrobia, Bacteroidetes, Tenericutes, Proteobacteria, Cyanobacteria, Saccharibacteria, and Actinobacteria. Interaction of Ent M with some phyla resulted in reduced abundance percentage. At day 21, significantly increased phagocytic activity (PA) was found in E1 and E2 (p < 0.001). Kr8+ did not attack PA and did not stimulate oxidative stress. But Ent M supported PA. The prospective importance of this study lies in beneficial interaction of enterocin in host body.
Subject(s)
Bacteriocins , Enterococcus hirae , Rabbits , Animals , Biofilms , Female , Male , Prospective StudiesABSTRACT
Indole is an inter-species and inter-kingdom signaling molecule widespread in the natural world. A large amount of indole in livestock wastes makes it difficult to be degraded, which causes serious malodor. Identifying efficient and eco-friendly ways to eliminate it is an urgent task for the sustainable development of husbandry. While bioconversion is a widely accepted means, the mechanism of indole microbial degradation is little understood, especially under anaerobic conditions. Herein, a new Enterococcus hirae isolate GDIAS-5, effectively degraded 100 mg/L indole within 28 h aerobically or 5 days anaerobically. Three intermediates (oxindole, isatin, and catechol) were identified in indole degradation, and catechol was further degraded by a meta-cleavage catabolic pathway. Two important processes for GDIAS-5 indole utilization were discovered. One is Fe(III) uptake and reduction, which may be a critical process that is coupled with indole oxidation, and the other is the entire pathway directly involved in indole oxidation and metabolism. Furthermore, monooxygenase ycnE responsible for indole oxidation via the indole-oxindole-isatin pathway was identified for the first time. Bioinformatic analyses showed that ycnE from E. hirae formed a phylogenetically separate branch from monooxygenases of other species. These findings provide new targets and strategies for synthetic biological reconstruction of indole-degrading bacteria.
Subject(s)
Enterococcus hirae , Isatin , Bacteria/metabolism , Catechols , Enterococcus hirae/metabolism , Ferric Compounds , Indoles/metabolism , OxindolesABSTRACT
The development of antimicrobial agents against multidrug-resistant bacteria is an important medical challenge. Antimicrobial peptides (AMPs), human cathelicidin LL-37 and its derivative OP-145, possess a potent antimicrobial activity and were under consideration for clinical trials. In order to overcome some of the challenges to their therapeutic potential, a very promising AMP, SAAP-148 was designed. Here, we studied the mode of action of highly cationic SAAP-148 in comparison with OP-145 on membranes of Enterococcus hirae at both cellular and molecular levels using model membranes composed of major constituents of enterococcal membranes, that is, anionic phosphatidylglycerol (PG) and cardiolipin (CL). In all assays used, SAAP-148 was consistently more efficient than OP-145, but both peptides displayed pronounced time and concentration dependences in killing bacteria and performing at the membrane. At cellular level, Nile Red-staining of enterococcal membranes showed abnormalities and cell shrinkage, which is also reflected in depolarization and permeabilization of E. hirae membranes. At the molecular level, both peptides abolished the thermotropic phase transition and induced disruption of PG/CL. Interestingly, the membrane was disrupted before the peptides neutralized the negative surface charge of PG/CL. Our results demonstrate that SAAP-148, which kills bacteria at a significantly lower concentration than OP-145, shows stronger effects on membranes at the cellular and molecular levels.
Subject(s)
Antimicrobial Peptides , Enterococcus hirae , Anti-Bacterial Agents/chemistry , Cell Membrane/metabolism , Drug Resistance, Multiple, Bacterial , Humans , Microbial Sensitivity Tests , PhosphatidylglycerolsABSTRACT
Application of cryo-electron microscopy (cryo-EM) is crucially important for ascertaining the atomic structure of large biomolecules such as ribosomes and protein complexes in membranes. Advances in cryo-EM technology and software have made it possible to obtain data with near-atomic resolution, but the method is still often capable of producing only a density map with up to medium resolution, either partially or entirely. Therefore, bridging the gap separating the density map and the atomic model is necessary. Herein, we propose a methodology for constructing atomic structure models based on cryo-EM maps with low-to-medium resolution. The method is a combination of sensitive and accurate homology modeling using our profile-profile alignment method with a flexible-fitting method using molecular dynamics simulation. As described herein, this study used benchmark applications to evaluate the model constructions of human two-pore channel 2 (one target protein in CASP13 with its structure determined using cryo-EM data) and the overall structure of Enterococcus hirae V-ATPase complex.
Subject(s)
Adenosine Triphosphatases/chemistry , Protein Conformation , Sequence Alignment , Sequence Homology, Amino Acid , Cryoelectron Microscopy/methods , Enterococcus hirae/metabolism , Humans , Models, Chemical , Molecular Dynamics Simulation , SoftwareABSTRACT
Prodigiosenes are a family of red pigments with versatile biological activity. Their tripyrrolic core structure has been modified many times in order to manipulate the spectrum of activity. We have been looking systematically at prodigiosenes substituted at the C ring with alkyl chains of different lengths, in order to assess the relevance of this substituent in a context that has not been investigated before for these derivatives: Cu(II) complexation, DNA binding, self-activated DNA cleavage, photoinduced cytotoxicity and antimicrobial activity. Our results indicate that the hydrophobic substituent has a clear influence on the different aspects of their biological activity. The cytotoxicity study of the Cu(II) complexes of these prodigiosenes shows that they exhibit a strong cytotoxic effect towards the tested tumor cell lines. The Cu(II) complex of a prodigiosene lacking any alkyl chain excelled in its photoinduced anticancer activity, thus demonstrating the potential of prodigiosenes and their metal complexes for an application in photodynamic therapy (PDT). Two derivatives along with their Cu(II) complexes showed also antimicrobial activity against Staphylococcus aureus strains.
Subject(s)
Anti-Bacterial Agents/pharmacology , Antineoplastic Agents/pharmacology , Coordination Complexes/pharmacology , Copper/pharmacology , DNA/drug effects , Alkylation , Animals , Anti-Bacterial Agents/chemical synthesis , Anti-Bacterial Agents/chemistry , Antineoplastic Agents/chemical synthesis , Antineoplastic Agents/chemistry , Cell Line , Cell Proliferation/drug effects , Cell Survival/drug effects , Coordination Complexes/chemical synthesis , Coordination Complexes/chemistry , Copper/chemistry , DNA Cleavage/drug effects , Dose-Response Relationship, Drug , Drug Screening Assays, Antitumor , Enterococcus hirae/drug effects , Escherichia coli/drug effects , Humans , Mice , Microbial Sensitivity Tests , Molecular Structure , Photochemotherapy , Pseudomonas aeruginosa/drug effects , Rats , Reactive Oxygen Species/metabolism , Staphylococcus aureus/drug effects , Structure-Activity RelationshipABSTRACT
Enterococcus cecorum and Enterococcus hirae can cause locomotor problems, septicaemia, and endocarditis in broiler chickens. Understanding transmission routes and resistance patterns are essential for effective treatment. The aim of this study was to follow the same animals from the breeder flock to the hatchery and up to 14-day-old broiler chickens on the farm to find the source of E. cecorum and E. hirae. During the production cycle, only faeces and organs of broilers were E. hirae positive in all three sampled farms in which recurrent enterococcal infections were previously confirmed. None of the isolates possessed virulence genes. Based on resistance profiles, a variety of different strains were present in faeces and organs of different broilers' ages. Samples from the breeder flock and hatchery were negative. Faecal shedding on the farm and tolerance of enterococci to the environmental conditions enable persistence of pathogenic enterococci in farm dust; therefore, adequate cleaning and disinfection after depopulation of the farms could prevent disease recurrence in the new cycle. Susceptibility testing of E. hirae isolates showed no resistance to the drugs of choice for the treatment of enterococcal infections in poultry.
Subject(s)
Gram-Positive Bacterial Infections , Poultry Diseases , Animals , Anti-Bacterial Agents , Chickens , Drug Resistance, Microbial , Enterococcus , Enterococcus hirae , Poultry , SloveniaABSTRACT
BACKGROUND: Infections caused by Enterococcus hirae are common in animals, with instances of transmission to humans being rare. Further, few cases have been reported in humans because of the difficulty in identifying the bacteria. Herein, we report a case of pyelonephritis caused by E. hirae bacteremia and conduct a literature review on E. hirae bacteremia. CASE PRESENTATION: A 57-year-old male patient with alcoholic cirrhosis and neurogenic bladder presented with fever and chills that had persisted for 3 days. Physical examination revealed tenderness of the right costovertebral angle. Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) of the patient's blood and urine samples revealed the presence of E. hirae, and pyelonephritis was diagnosed. The patient was treated successfully with intravenous ampicillin followed by oral linezolid for a total of three weeks. CONCLUSION: The literature review we conducted revealed that E. hirae bacteremia is frequently reported in urinary tract infections, biliary tract infections, and infective endocarditis and is more likely to occur in patients with diabetes, liver cirrhosis, and chronic kidney disease. However, mortality is not common because of the high antimicrobial susceptibility of E. hirae. With the advancements in MALDI-TOF MS, the number of reports of E. hirae infections has also increased, and clinicians need to consider E. hirae as a possible causative pathogen of urinary tract infections in patients with known risk factors.
Subject(s)
Bacteremia , Pyelonephritis , Ampicillin , Animals , Bacteremia/complications , Bacteremia/drug therapy , Enterococcus hirae , Humans , Liver Cirrhosis, Alcoholic/complications , Male , Middle Aged , Pyelonephritis/complications , Pyelonephritis/drug therapy , Spectrometry, Mass, Matrix-Assisted Laser Desorption-IonizationABSTRACT
Bacteriocins produced by lactic acid bacteria have potential use as natural food preservatives, which may alleviate current problems associated with the overuse of antibiotics and emerging multi-drug-resistant microbes. In this work, Lactiplantibacillus plantarum RUB1 was found to produce a class IIb bacteriocin with strong antibacterial activity. Except for plnXY encoding putative proteins, L. plantarum RUB1 contains most genes in five operons (plnABCD, plnGHSTUVW, plnMNOP, plnIEF, and plnRLJK) related to bacteriocin synthesis. Adding low (100 and 500 ng/mL) and medium (1 µg/mL) concentrations of PlnA to broth promoted bacteriocin production and upregulated bacteriocin gene plnA, while high concentrations (50 and 200 µg/mL) inhibited expression of these genes. Co-culturing L. plantarum RUB1 with Enterococcus hirae 1003, Enterococcus hirae LWS, Limosilactobacillus fermentum RC4, L. plantarum B6, and even Listeria monocytogenes ATCC 19111 and Staphylococcus aureus ATCC 6538 enhanced bacteriocin activity and expression of bacteriocin-related genes. This study verifies that PlnA can indeed upregulate the expression of bacteriocin genes, and also bacteriocin production can be induced by co-culture with some specific bacteria or their cell-free supernatants. Bacteriocin production by L. plantarum RUB1 is mediated by a quorum sensing mechanism, directly influenced by autoinducing peptide or specific strains. The findings provide new methods and insight into bacteriocin production mechanisms.
Subject(s)
Anti-Bacterial Agents , Bacteriocins , Lactobacillus plantarum , Anti-Bacterial Agents/pharmacology , Bacteriocins/biosynthesis , Bacteriocins/pharmacology , Enterococcus hirae/drug effects , Quorum Sensing , Staphylococcus aureus/drug effectsABSTRACT
In canine nutrition, the use of goat nutraceutical dairy products is an innovative proposal. Therefore, the objective of this study was to prepare fermented goat milk with probiotic potential in dogs in an in vitro model. A total of 40 lactic acid bacteria (LAB) species were grown, of which 30 were CAP isolates originally from goat milk and 10 were CAN isolates originally from fecal material of newborn dogs. The isolates were selected based on resistance to the simulated canine gastrointestinal condition and acidifying ability. After this preliminary screening, the analyses were performed regarding ß-galactosidase and exopolysaccharide formation, diacetyl production, adhesion proteins Mub and mapa, hydrophobicity, DPPH assay, virulence and antibiotic resistance. With these evaluations, four LAB isolates were identified using sequencing of the 16S rRNA gene. These were identified as Enterococcus hirae and were used to produce fermented goat milk. For statistical analysis, the data were analyzed using the Scott-Knott test and also submitted to analysis of variance and the Tukey test (P < 0.05). In the evaluation of goat milk fermented with E. hirae and control, over the 36-day storage period there was a reduction in pH and an increase in acidity, and higher levels of LAB were observed in goat milk fermented with E. hirae. Therefore, both these E. hirae isolates and the fermented goat milk produced showed satisfactory results in vitro, demonstrating probiotic efficiency and food safety for dogs.
Subject(s)
Bacterial Physiological Phenomena , Dogs/microbiology , Enterococcus hirae/chemistry , Gastrointestinal Tract/microbiology , Goats/microbiology , Milk/microbiology , Probiotics/administration & dosage , Animals , Dogs/physiology , Feces/microbiology , Fermentation , Gastrointestinal Tract/physiologyABSTRACT
A deviated repertoire of the gut microbiome predicts resistance to cancer immunotherapy. Enterococcus hirae compensated cancer-associated dysbiosis in various tumor models. However, the mechanisms by which E. hirae restored the efficacy of cyclophosphamide administered with concomitant antibiotics remain ill defined. Here, we analyzed the multifaceted modes of action of this anticancer probiotic. Firstly, E. hirae elicited emigration of thymocytes and triggered systemic and intratumoral IFNγ-producing and CD137-expressing effector memory T cell responses. Secondly, E. hirae activated the autophagy machinery in enterocytes and mediated ATG4B-dependent anticancer effects, likely as a consequence of its ability to increase local delivery of polyamines. Thirdly, E. hirae shifted the host microbiome toward a Bifidobacteria-enriched ecosystem. In contrast to the live bacterium, its pasteurized cells or membrane vesicles were devoid of anticancer properties. These pleiotropic functions allow the design of optimal immunotherapies combining E. hirae with CD137 agonistic antibodies, spermidine, or Bifidobacterium animalis. We surmise that immunological, metabolic, epithelial, and microbial modes of action of the live E. hirae cooperate to circumvent primary resistance to therapy.