scholarly journals Expression of Genes Involved in Bacteriocin Production and Self-Resistance inLactobacillus brevis174A Is Mediated by Two Regulatory Proteins

2018 ◽  
Vol 84 (7) ◽  
Author(s):  
Masafumi Noda ◽  
Rumi Miyauchi ◽  
Narandalai Danshiitsoodol ◽  
Yasuyuki Matoba ◽  
Takanori Kumagai ◽  
...  
Keyword(s):  
Lactic Acid ◽  
Gene Cluster ◽  
Cellular Response ◽  
Pathogenic Bacteria ◽  
Response Regulator ◽  
Lactobacillus Brevis ◽  
Regulatory Proteins ◽  
Bacterial Strains ◽  
Content Type ◽  
Transcriptional Regulatory

ABSTRACTWe have previously shown that the lactic acid bacteriumLactobacillus brevis174A, isolated fromCitrus iyofruit, produces a bacteriocin designated brevicin 174A, which is comprised of two antibacterial polypeptides (designated brevicins 174A-β and 174A-γ). We have also found a gene cluster, composed of eight open reading frames (ORFs), that contains genes for the biosynthesis of brevicin 174A, self-resistance to its own bacteriocin, and two transcriptional regulatory proteins. Some lactic acid bacterial strains have a system to start the production of bacteriocin at an adequate stage of growth. Generally, the system consists of a membrane-bound histidine protein kinase (HPK) that senses a specific environmental stimulus and a corresponding response regulator (RR) that mediates the cellular response. We have previously shown that although the HPK- and RR-encoding genes are not found on the brevicin 174A biosynthetic gene cluster in the 174A strain, two putative regulatory genes, designatedbreDandbreG, are in the gene cluster. In the present study, we demonstrate that the expression of brevicin 174A production and self-resistance is positively controlled by two transcriptional regulatory proteins, designated BreD and BreG. BreD is expressed together with BreE as the self-resistance determinant ofL. brevis174A. DNase I footprinting analysis and a promoter assay demonstrated that BreD binds to thebreEDpromoter as a positive autoregulator. The present study also demonstrates that BreG, carrying a transmembrane domain, binds to the common promoter ofbreBandbreC, encoding brevicins 174A-β and 174A-γ, respectively, for positive regulation.IMPORTANCEThe problem of the appearance of bacteria that are resistant to practical antibiotics and the increasing demand for safe foods have increased interest in replacing conventional antibiotics with bacteriocin produced by the lactic acid bacteria. This antibacterial substance can inhibit the growth of pathogenic bacteria without side effects on the human body. The bacteriocin that is produced by aCitrus iyo-derivedLactobacillus brevisstrain inhibits the growth of pathogenic bacteria such asListeria monocytogenes,Staphylococcus aureus, andStreptococcus mutans. In general, lactic acid bacterial strains have a system to start the production of bacteriocin at an adequate stage of growth, which is called a quorum-sensing system. The system consists of a membrane-bound histidine protein kinase that senses a specific environmental stimulus and a corresponding response regulator that mediates the cellular response. The present study demonstrates that the expression of the genes encoding bacteriocin biosynthesis and the self-resistance determinant is positively controlled by two transcriptional regulatory proteins.

scholarly journals ISOLATION AND CHARACTERIZATION OF UNIQUE FRUCTOPHILIC LACTIC ACID BACTERIA FROM DIFFERENT FLOWER SOURCES.

2020 ◽  
Vol 51 (2) ◽  
pp. 508-518
Author(s):  
G. M. Saleh
Keyword(s):  
Lactic Acid ◽  
Lactic Acid Bacteria ◽  
Enterococcus Faecium ◽  
Pathogenic Bacteria ◽  
Lactobacillus Brevis ◽  
Nerium Oleander ◽  
Dual Culture ◽  
Bacterial Strains ◽  
Convolvulus Arvensis ◽  
Isolation And Characterization

This study was aimed to investigate Lactic acid bacteria (LAB) that contain an important group of bacterial strains such as fructophilic lactic acid bacteria that usually isolated from fructose rich niches. These groups of bacteria have been considered as probiotics, especially in animal applications. In this study these bacteria were isolated from 6  Plant sources (flowers) such as: Convolvulus arvensis, Hibiscus rosa-sinensis , Nerium oleander, Rosa rugosa, Tagetes erecta and Zinnia elegans. The bacterial isolates were identified by morphological and molecular methods through the detection of 16SrRNA gene. Carbohydrate fermentation profile as well as their antibacterial activity using the dual culture overlay assay was also detected. The results revealed that 5 types of different lactic acid bacteria were isolated included: Fructobacillus fructosus, Lactobacillus kunkeei, Enterococcus durans, Enterococcus faecium , and  Lactobacillus brevis. These isolates showed variety in fermentation of carbohydrates. Also, all isolates exhibit inhibition effect against pathogenic bacteria (Staphylococcus auresو Pseudomonas aeruginosa) except Enterococcus faecium which showed no effect it.

scholarly journals A Novel CO-Responsive Transcriptional Regulator and Enhanced H2Production by an Engineered Thermococcus onnurineus NA1 Strain

2014 ◽  
Vol 81 (5) ◽  
pp. 1708-1714 ◽  
Author(s):  
Min-Sik Kim ◽  
Ae Ran Choi ◽  
Seong Hyuk Lee ◽  
Hae-Chang Jung ◽  
Seung Seob Bae ◽  
...  
Keyword(s):  
Production Rate ◽  
Gene Cluster ◽  
Type Strain ◽  
Wild Type Strain ◽  
Regulatory System ◽  
Transcriptional Regulator ◽  
Bioreactor Culture ◽  
Wild Type ◽  
Content Type ◽  
Transcriptional Regulatory

ABSTRACTGenome analysis revealed the existence of a putative transcriptional regulatory system governing CO metabolism inThermococcus onnurineusNA1, a carboxydotrophic hydrogenogenic archaeon. The regulatory system is composed of CorQ with a 4-vinyl reductase domain and CorR with a DNA-binding domain of the LysR-type transcriptional regulator family in close proximity to the CO dehydrogenase (CODH) gene cluster. Homologous genes of the CorQR pair were also found in the genomes ofThermococcusspecies and “CandidatusKorarchaeum cryptofilum” OPF8. In-frame deletion of eithercorQorcorRcaused a severe impairment in CO-dependent growth and H2production. WhencorQandcorRdeletion mutants were complemented by introducing thecorQRgenes under the control of a strong promoter, the mRNA and protein levels of the CODH gene were significantly increased in a ΔCorR strain complemented with integratedcorQR(ΔCorR/corQR↑) compared with those in the wild-type strain. In addition, the ΔCorR/corQR↑strain exhibited a much higher H2production rate (5.8-fold) than the wild-type strain in a bioreactor culture. The H2production rate (191.9 mmol liter−1h−1) and the specific H2production rate (249.6 mmol g−1h−1) of this strain were extremely high compared with those of CO-dependent H2-producing prokaryotes reported so far. These results suggest that thecorQRgenes encode a positive regulatory protein pair for the expression of a CODH gene cluster. The study also illustrates that manipulation of the transcriptional regulatory system can improve biological H2production.

scholarly journals Trk2 Potassium Transport System in Streptococcus mutans and Its Role in Potassium Homeostasis, Biofilm Formation, and Stress Tolerance

2016 ◽  
Vol 198 (7) ◽  
pp. 1087-1100 ◽  
Author(s):  
Gursonika Binepal ◽  
Kamal Gill ◽  
Paula Crowley ◽  
Martha Cordova ◽  
L. Jeannine Brady ◽  
...  
Keyword(s):  
Stress Tolerance ◽  
Streptococcus Mutans ◽  
Transport System ◽  
Biofilm Formation ◽  
Cellular Response ◽  
Pathogenic Bacteria ◽  
Transport Systems ◽  
Content Type ◽  
Growth And Survival ◽  
Dental Biofilm

ABSTRACTPotassium (K+) is the most abundant cation in the fluids of dental biofilm. The biochemical and biophysical functions of K+and a variety of K+transport systems have been studied for most pathogenic bacteria but not for oral pathogens. In this study, we establish the modes of K+acquisition inStreptococcus mutansand the importance of K+homeostasis for its virulence attributes. TheS. mutansgenome harbors four putative K+transport systems that included two Trk-like transporters (designated Trk1 and Trk2), one glutamate/K+cotransporter (GlnQHMP), and a channel-like K+transport system (Kch). Mutants lacking Trk2 had significantly impaired growth, acidogenicity, aciduricity, and biofilm formation. [K+] less than 5 mM eliminated biofilm formation inS. mutans. The functionality of the Trk2 system was confirmed by complementing anEscherichia coliTK2420 mutant strain, which resulted in significant K+accumulation, improved growth, and survival under stress. Taken together, these results suggest that Trk2 is the main facet of the K+-dependent cellular response ofS. mutansto environment stresses.IMPORTANCEBiofilm formation and stress tolerance are important virulence properties of caries-causingStreptococcus mutans. To limit these properties of this bacterium, it is imperative to understand its survival mechanisms. Potassium is the most abundant cation in dental plaque, the natural environment ofS. mutans. K+is known to function in stress tolerance, and bacteria have specialized mechanisms for its uptake. However, there are no reports to identify or characterize specific K+transporters inS. mutans. We identified the most important system for K+homeostasis and its role in the biofilm formation, stress tolerance, and growth. We also show the requirement of environmental K+for the activity of biofilm-forming enzymes, which explains why such high levels of K+would favor biofilm formation.

scholarly journals Overproduction of Ristomycin A by Activation of a Silent Gene Cluster in Amycolatopsis japonicum MG417-CF17

2014 ◽  
Vol 58 (10) ◽  
pp. 6185-6196 ◽  
Author(s):  
Marius Spohn ◽  
Norbert Kirchner ◽  
Andreas Kulik ◽  
Angelika Jochim ◽  
Felix Wolf ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Gene Cluster ◽  
High Performance ◽  
Pathogenic Bacteria ◽  
Genome Mining ◽  
Gene Clusters ◽  
Von Willebrand Disease ◽  
Biosynthesis Gene Cluster ◽  
Content Type ◽  
Bioinformatic Tools

ABSTRACTThe emergence of antibiotic-resistant pathogenic bacteria within the last decades is one reason for the urgent need for new antibacterial agents. A strategy to discover new anti-infective compounds is the evaluation of the genetic capacity of secondary metabolite producers and the activation of cryptic gene clusters (genome mining). One genus known for its potential to synthesize medically important products isAmycolatopsis. However,Amycolatopsis japonicumdoes not produce an antibiotic under standard laboratory conditions. In contrast to mostAmycolatopsisstrains,A. japonicumis genetically tractable with different methods. In order to activate a possible silent glycopeptide cluster, we introduced a gene encoding the transcriptional activator of balhimycin biosynthesis, thebbrgene fromAmycolatopsis balhimycina(bbrAba), intoA. japonicum. This resulted in the production of an antibiotically active compound. Following whole-genome sequencing ofA. japonicum, 29 cryptic gene clusters were identified by genome mining. One of these gene clusters is a putative glycopeptide biosynthesis gene cluster. Using bioinformatic tools, ristomycin (syn. ristocetin), a type III glycopeptide, which has antibacterial activity and which is used for the diagnosis of von Willebrand disease and Bernard-Soulier syndrome, was deduced as a possible product of the gene cluster. Chemical analyses by high-performance liquid chromatography and mass spectrometry (HPLC-MS), tandem mass spectrometry (MS/MS), and nuclear magnetic resonance (NMR) spectroscopy confirmed thein silicoprediction that the recombinantA. japonicum/pRM4-bbrAbasynthesizes ristomycin A.

scholarly journals The effect of bacterial inoculant supplementation on the fermentation process of crushed maize ears in laboratory silos

2005 ◽  
Vol 53 (4) ◽  
pp. 33-42
Author(s):  
Petr Doležal ◽  
Dušan Kořínek ◽  
Jan Doležal ◽  
Václav Pyrochta
Keyword(s):  
Lactic Acid ◽  
Ph Value ◽  
Lactobacillus Rhamnosus ◽  
Lactobacillus Brevis ◽  
Water Soluble ◽  
Bacterial Strains ◽  
Total Acid ◽  
Ethanol Content ◽  
Fermentation Quality ◽  
Biological Additive

In the experiment was the effect of biological additive on the fermentation quality of crushed maize ears of two hybrids by comparing with the untreated control. The bacterial inoculant „A“ contained selected bacterial strains of Lactobacillus rhamnosus (NCIMB 30121) and Enterococcus faecium (NCIMB 30122). As effective substances of bacterial water–soluble inoculant „B“ were selected bacterial strains of Lactobacillus rhamnosus (NCIMB 30121), Lactobacillus plantarum (DSM 12836), Lactobacillus brevis (DSM 12835), Lactobacillus buchneri (DSM 12856), Pediococcus acidialactici (P. pentosaceus) (DSM 12834). The addition of inoculant „A“ in our experiment conditions increased statistically significantly (P<0.01) the pH value (4.09±0.01), resp. 4.02±0.02 in second trial with Pedro hybrid. The bacterial inoculant „B“ increased significantly (P<0.01) the contents of lactic acid (50.95±0.1.87 g/kg DM), acetic acid (18.61±0.34 g/kg DM), sum of acids (69.55±1.62 g/kg DM) and decreased (P<0.01) in the first trial the ethanol content (5.41±0.45 g/kg DM). The highest DM content (P<0.01) was in all experimental inoculated silages with additive „A“ (54.26±0.86%, and 53.56±0.54%, resp.). The bacterial inoculant „A“ increased significantly (P<0.01) in comparison with control silage in the second trial the content of lactic acid (34.66Ī2.81 g/kg DM), sum of acids (44.68±3.54 g/kg DM), the total acids content (32.87±2.88 g/kg DM), and ethanol content (17.33±0.79 g/kg DM). The inoculation positive effect was demonstrated in reduction of ethanol amount and of total acid production. The pH value of inoculated silages was not significantly lower than that in the control silage.

scholarly journals Gene Cluster Responsible for Secretion of and Immunity to Multiple Bacteriocins, the NKR-5-3 Enterocins

2014 ◽  
Vol 80 (21) ◽  
pp. 6647-6655 ◽  
Author(s):  
Naoki Ishibashi ◽  
Kohei Himeno ◽  
Yoshimitsu Masuda ◽  
Rodney Honrada Perez ◽  
Shun Iwatani ◽  
...  
Keyword(s):  
Gene Cluster ◽  
Abc Transporter ◽  
Response Regulator ◽  
Regulatory System ◽  
Content Type ◽  
Expression Studies ◽  
A Genome ◽  
Wide Range ◽  
Close Proximity ◽  
Immunity Genes

ABSTRACTEnterococcus faeciumNKR-5-3, isolated from Thai fermented fish, is characterized by the unique ability to produce five bacteriocins, namely, enterocins NKR-5-3A, -B, -C, -D, and -Z (Ent53A, Ent53B, Ent53C, Ent53D, and Ent53Z). Genetic analysis with a genome library revealed that the bacteriocin structural genes (enkA[ent53A],enkC[ent53C],enkD[ent53D], andenkZ[ent53Z]) that encode these peptides (except for Ent53B) are located in close proximity to each other. This NKR-5-3ACDZ (Ent53ACDZ) enterocin gene cluster (approximately 13 kb long) includes certain bacteriocin biosynthetic genes such as an ABC transporter gene (enkT), two immunity genes (enkIazandenkIc), a response regulator (enkR), and a histidine protein kinase (enkK). Heterologous-expression studies ofenkTand ΔenkTmutant strains showed thatenkTis responsible for the secretion of Ent53A, Ent53C, Ent53D, and Ent53Z, suggesting that EnkT is a wide-range ABC transporter that contributes to the effective production of these bacteriocins. In addition, EnkIaz and EnkIc were found to confer self-immunity to the respective bacteriocins. Furthermore, bacteriocin induction assays performed with the ΔenkRKmutant strain showed that EnkR and EnkK are regulatory proteins responsible for bacteriocin production and that, together with Ent53D, they constitute a three-component regulatory system. Thus, the Ent53ACDZ gene cluster is essential for the biosynthesis and regulation of NKR-5-3 enterocins, and this is, to our knowledge, the first report that demonstrates the secretion of multiple bacteriocins by an ABC transporter.

scholarly journals Inulin Fermentation by Lactobacilli and Bifidobacteria from Dairy Calves

2020 ◽  
Vol 87 (1) ◽  
Author(s):  
Yuanting Zhu ◽  
Jinxin Liu ◽  
Julian M. Lopez ◽  
David A. Mills
Keyword(s):  
Lactic Acid ◽  
Lactic Acid Bacteria ◽  
Gut Microbiome ◽  
Animal Health ◽  
Dairy Calves ◽  
Milk Replacer ◽  
Comparative Genomic ◽  
Bacterial Strains ◽  
Content Type ◽  
Long Chain

ABSTRACT Prebiotics are increasingly examined for their ability to modulate the neonate gut microbiota of livestock, and products such as inulin are commonly added to milk replacer used in calving. However, the ability of specific members of the bovine neonate microbiota to respond to inulin remains to be determined, particularly among indigenous lactobacilli and bifidobacteria, beneficial genera commonly enriched by inulin. Screening of Bifidobacterium and Lactobacillus isolates obtained from fresh feces of dairy calves revealed that lactobacilli had a higher prevalence of inulin fermentation capacity (58%) than bifidobacteria (17%). Several Ligilactobacillus agilis (synonym Lactobacillus agilis) isolates exhibited vigorous growth on, and complete degradation of, inulin; however, the phenotype was strain specific. The most vigorous inulin-fermenting strain, L. agilis YZ050, readily degraded long-chain inulin not consumed by bifidobacterial isolates. Comparative genomic analysis of both L. agilis fermenter and nonfermenter strains indicated that strain YZ050 encodes an inulinase homolog, previously linked to extracellular degradation of long-chain inulin in Lacticaseibacillus paracasei, that was strongly induced during growth on inulin. Inulin catabolism by YZ050 also generates extracellular fructose, which can cross-feed other non-inulin-fermenting lactic acid bacteria isolated from the same bovine feces. The presence of specific inulin-responsive bacterial strains within calf gut microbiome provides a mechanistic rationale for enrichment of specific lactobacilli and creates a foundation for future synbiotic applications in dairy calves aimed at improving health in early life. IMPORTANCE The gut microbiome plays an important role in animal health and is increasingly recognized as a target for diet-based manipulation. Inulin is a common prebiotic routinely added to animal feeds; however, the mechanism of inulin consumption by specific beneficial taxa in livestock is ill defined. In this study, we examined Lactobacillus and Bifidobacterium isolates from calves fed inulin-containing milk replacer and characterized specific strains that robustly consume long-chain inulin. In particular, novel Ligilactobacillus agilis strain YZ050 consumed inulin via an extracellular fructosidase, resulting in complete consumption of all long-chain inulin. Inulin catabolism resulted in temporal release of extracellular fructose, which can promote growth of other non-inulin-consuming strains of lactic acid bacteria. This work provides the mechanistic insight needed to purposely modulate the calf gut microbiome via the establishment of networks of beneficial microbes linked to specific prebiotics.

scholarly journals GlnR Negatively Regulates Glutamate-Dependent Acid Resistance in Lactobacillus brevis

2020 ◽  
Vol 86 (7) ◽  
Author(s):  
Luchan Gong ◽  
Cong Ren ◽  
Yan Xu
Keyword(s):  
Lactic Acid ◽  
Lactic Acid Bacteria ◽  
Glutamate Decarboxylase ◽  
Wild Type Strain ◽  
Acid Resistance ◽  
Lactobacillus Brevis ◽  
Deletion Strain ◽  
Content Type ◽  
Gaba Production ◽  
Encoding Gene

ABSTRACT Lactic acid bacteria often encounter a variety of multiple stresses in their natural and industrial fermentation environments. The glutamate decarboxylase (GAD) system is one of the most important acid resistance systems in lactic acid bacteria. In this study, we demonstrated that GlnR, a nitrogen regulator in Gram-positive bacteria, directly modulated γ-aminobutyric acid (GABA) conversion from glutamate and was involved in glutamate-dependent acid resistance in Lactobacillus brevis. The glnR deletion strain (ΔglnR mutant) achieved a titer of 284.7 g/liter GABA, which is 9.8-fold higher than that of the wild-type strain. The cell survival of the glnR deletion strain was significantly higher than that of the wild-type strain under the condition of acid challenge and was positively correlated with initial glutamate concentration and GABA production. Quantitative reverse transcription-PCR assays demonstrated that GlnR inhibited the transcription of the glutamate decarboxylase-encoding gene (gadB), glutamate/GABA antiporter-encoding gene (gadC), glutamine synthetase-encoding gene (glnA), and specific transcriptional regulator-encoding gene (gadR) involved in gadCB operon regulation. Moreover, GABA production and glutamate-dependent acid resistance were absolutely abolished in the gadR glnR deletion strain. Electrophoretic mobility shift and DNase I footprinting assays revealed that GlnR directly bound to the 5′-untranslated regions of the gadR gene and gadCB operon, thus inhibiting their transcription. These results revealed a novel regulatory mechanism of GlnR on glutamate-dependent acid resistance in Lactobacillus. IMPORTANCE Free-living lactic acid bacteria often encounter acid stresses because of their organic acid-producing features. Several acid resistance mechanisms, such as the glutamate decarboxylase system, F1Fo-ATPase proton pump, and alkali production, are usually employed to relieve growth inhibition caused by acids. The glutamate decarboxylase system is vital for GAD-containing lactic acid bacteria to protect cells from DNA damage, enzyme inactivation, and product yield loss in acidic habitats. In this study, we found that a MerR-type regulator, GlnR, was involved in glutamate-dependent acid resistance by directly regulating the transcription of the gadR gene and gadCB operon, resulting in an inhibition of GABA conversion from glutamate in L. brevis. This study represents a novel mechanism for GlnR's regulation of glutamate-dependent acid resistance and also provides a simple and novel strategy to engineer Lactobacillus strains to elevate their acid resistance as well as GABA conversion from glutamate.

scholarly journals The Two-Component Signal Transduction System VxrAB Positively Regulates Vibrio cholerae Biofilm Formation

2017 ◽  
Vol 199 (18) ◽  
Author(s):  
Jennifer K. Teschler ◽  
Andrew T. Cheng ◽  
Fitnat H. Yildiz
Keyword(s):  
Signal Transduction ◽  
Vibrio Cholerae ◽  
Histidine Kinase ◽  
Biofilm Formation ◽  
Pathogenic Bacteria ◽  
Response Regulator ◽  
Systematic Analysis ◽  
Content Type ◽  
Two Component ◽  
Two Component Signal Transduction

ABSTRACT Two-component signal transduction systems (TCSs), typically composed of a sensor histidine kinase (HK) and a response regulator (RR), are the primary mechanism by which pathogenic bacteria sense and respond to extracellular signals. The pathogenic bacterium Vibrio cholerae is no exception and harbors 52 RR genes. Using in-frame deletion mutants of each RR gene, we performed a systematic analysis of their role in V. cholerae biofilm formation. We determined that 7 RRs impacted the expression of an essential biofilm gene and found that the recently characterized RR, VxrB, regulates the expression of key structural and regulatory biofilm genes in V. cholerae. vxrB is part of a 5-gene operon, which contains the cognate HK vxrA and three genes of unknown function. Strains carrying ΔvxrA and ΔvxrB mutations are deficient in biofilm formation, while the ΔvxrC mutation enhances biofilm formation. The overexpression of VxrB led to a decrease in motility. We also observed a small but reproducible effect of the absence of VxrB on the levels of cyclic di-GMP (c-di-GMP). Our work reveals a new function for the Vxr TCS as a regulator of biofilm formation and suggests that this regulation may act through key biofilm regulators and the modulation of cellular c-di-GMP levels. IMPORTANCE Biofilms play an important role in the Vibrio cholerae life cycle, providing protection from environmental stresses and contributing to the transmission of V. cholerae to the human host. V. cholerae can utilize two-component systems (TCS), composed of a histidine kinase (HK) and a response regulator (RR), to regulate biofilm formation in response to external cues. We performed a systematic analysis of V. cholerae RRs and identified a new regulator of biofilm formation, VxrB. We demonstrated that the VxrAB TCS is essential for robust biofilm formation and that this system may regulate biofilm formation via its regulation of key biofilm regulators and cyclic di-GMP levels. This research furthers our understanding of the role that TCSs play in the regulation of V. cholerae biofilm formation.

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