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1.
Gene ; 807: 145961, 2022 Jan 10.
Artigo em Inglês | MEDLINE | ID: mdl-34530088

RESUMO

Vibrio parahaemolyticus produces two types of IV pili: mannose-sensitive haemagglutinin type IV pili (MSHA) and chitin-regulated pili (ChiRP). Both of them are required for biofilm formation and the pathogen persistence in hosts. However, there are few reports on the regulation of their expression. In the present study, we showed that the master quorum sensing (QS) regulators AphA and OpaR oppositely regulated the transcription of mshA1 encoding the pilin of MSHA pilus in V. parahaemolyticus. At low cell density (LCD), AphA indirectly repressed mshA1 transcription. In contrast, at high cell density (HCD), OpaR bound to the regulatory DNA region of mshA1 to activate its transcription. Oppositely regulation of mshA1 by AphA and OpaR led to a gradual increase in the expression level of mshA1 from LCD to HCD. Thus, regulation of type IV pili production was one of the mechanisms that V. parahaemolyticus adopted to control biofilm formation.


Assuntos
Proteínas de Fímbrias/genética , Percepção de Quorum/genética , Vibrio/genética , Proteínas de Bactérias/genética , Biofilmes/crescimento & desenvolvimento , Contagem de Células , Proteínas de Fímbrias/metabolismo , Expressão Gênica/genética , Regulação Bacteriana da Expressão Gênica/genética , Lectina de Ligação a Manose/genética , Lectina de Ligação a Manose/metabolismo , Transativadores/genética , Transativadores/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Vibrio/metabolismo , Vibrio cholerae/genética , Vibrio cholerae/metabolismo , Vibrio parahaemolyticus/genética , Vibrio parahaemolyticus/metabolismo
2.
J Biol Chem ; 297(3): 101071, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34400168

RESUMO

VhCBP is a periplasmic chitooligosaccharide-binding protein mainly responsible for translocation of the chitooligosaccharide (GlcNAc)2 across the double membranes of marine bacteria. However, structural and thermodynamic understanding of the sugar-binding/-release processes of VhCBP is relatively less. VhCBP displayed the greatest affinity toward (GlcNAc)2, with lower affinity for longer-chain chitooligosaccharides [(GlcNAc)3-4]. (GlcNAc)4 partially occupied the closed sugar-binding groove, with two reducing-end GlcNAc units extending beyond the sugar-binding groove and barely characterized by weak electron density. Mutation of three conserved residues (Trp363, Asp365, and Trp513) to Ala resulted in drastic decreases in the binding affinity toward the preferred substrate (GlcNAc)2, indicating their significant contributions to sugar binding. The structure of the W513A-(GlcNAc)2 complex in a 'half-open' conformation unveiled the intermediary step of the (GlcNAc)2 translocation from the soluble CBP in the periplasm to the inner membrane-transporting components. Isothermal calorimetry data suggested that VhCBP adopts the high-affinity conformation to bind (GlcNAc)2, while its low-affinity conformation facilitated sugar release. Thus, chitooligosaccharide translocation, conferred by periplasmic VhCBP, is a crucial step in the chitin catabolic pathway, allowing Vibrio bacteria to thrive in oceans where chitin is their major source of nutrients.


Assuntos
Quitina/metabolismo , Dissacarídeos/metabolismo , Vibrio/metabolismo , Carboidratos , Quitinases/metabolismo , Quitosana/metabolismo , Cristalografia por Raios X/métodos , Dissacarídeos/fisiologia , Modelos Estruturais , Oligossacarídeos/metabolismo , Periplasma/metabolismo , Proteínas Periplásmicas de Ligação/metabolismo , Relação Estrutura-Atividade
3.
Int J Mol Sci ; 22(16)2021 Aug 05.
Artigo em Inglês | MEDLINE | ID: mdl-34445107

RESUMO

Brown algae is a kind of renewable resource for biofuels production. As the major component of carbohydrate in the cell walls of brown algae, alginate can be degraded into unsaturated monosaccharide by exo-type alginate lyases, then converted into 4-deoxy-L-erythro-5-hexoseulose uronate (DEH) by a non-enzyme reaction, which is an important raw material for the preparation of bioethanol. In our research, a novel exo-type alginate lyase, VsAly7D, belonging to the PL7 family was isolated from marine bacterium Vibrio sp. QY108 and recombinantly expressed in Escherichia coli. The purified VsAly7D demonstrated the highest activity at 35 °C, whereas it still maintained 46.5% and 83.1% of its initial activity at 20 °C and 30 °C, respectively. In addition, VsAly7D exhibited the maximum activity under alkaline conditions (pH 8.0), with the simultaneously remaining stability between pH 8.0 and 10.0. Compared with other reported exo-type enzymes, VsAly7D could efficiently degrade alginate, poly-ß-D-mannuronate (polyM) and poly-α-L-guluronate (polyG) with highest specific activities (663.0 U/mg, 913.6 U/mg and 894.4 U/mg, respectively). These results showed that recombinant VsAly7D is a suitable tool enzyme for unsaturated alginate monosaccharide preparation and holds great promise for producing bioethanol from brown algae.


Assuntos
Alginatos/metabolismo , Polissacarídeo-Liases/metabolismo , Vibrio/metabolismo , Sequência de Aminoácidos , Proteínas de Bactérias/metabolismo , Escherichia coli/metabolismo , Ácido Glucurônico/metabolismo , Concentração de Íons de Hidrogênio , Monossacarídeos/metabolismo , Feófitas/microbiologia
4.
Biochem Biophys Res Commun ; 568: 136-142, 2021 09 03.
Artigo em Inglês | MEDLINE | ID: mdl-34214877

RESUMO

Vibrio species are prevalent in the aquatic environments and can infect humans and aquatic organisms. Vibrio parahaemolyticus counteracts ß-lactam antibiotics and enhances virulence using a regulation mechanism mediated by a two-component regulatory system (TCS) consisting of the VbrK histidine kinase and the VbrR response regulator. The periplasmic sensor domain of VbrK (VbrKSD) detects ß-lactam antibiotics or undergoes S-nitrosylation in response to host nitrites. Although V. parahaemolyticus VbrKSD (vpVbrKSD) has recently been characterized through structural studies, it is unclear whether its structural features that are indispensable for biological functions are conserved in other VbrK orthologs. To structurally define the functionally critical regions of VbrK and address the structural dynamics of VbrK, we determined the crystal structures of Vibrio rotiferianus VbrKSD (vrVbrKSD) in two crystal forms and performed a comparative analysis of diverse VbrK structures. vrVbrKSD folds into a curved rod-shaped two-domain structure as observed in vpVbrKSD. The membrane-distal end of the vrVbrKSD structure, including the α3 helix and its neighboring loops, harbors both S-nitrosylation and antibiotic-sensing sites and displays high structural flexibility and diversity. Noticeably, the distal end is partially stabilized by a disulfide bond, which is formed by the cysteine residue that is S-nitrosylated in response to nitrite. Therefore, the distal end of VbrKSD plays a key role in initiating the VbrK-VbrR TCS pathway activation, and it is involved in the nitrosylation-mediated regulation of the structural dynamics of VbrK.


Assuntos
Proteínas de Bactérias/química , Histidina Quinase/química , Vibrio/química , Antibacterianos/metabolismo , Proteínas de Bactérias/metabolismo , Cristalografia por Raios X , Histidina Quinase/metabolismo , Modelos Moleculares , Nitritos/metabolismo , Domínios Proteicos , Vibrio/metabolismo
5.
World J Microbiol Biotechnol ; 37(7): 124, 2021 Jun 25.
Artigo em Inglês | MEDLINE | ID: mdl-34170406

RESUMO

Sulfate-reducing bacteria (SRB) are culprits for microbiologically influenced corrosion, and biofilms are believed to play essential roles in the corrosion induced by SRB. However, little is known about the regulation of SRB biofilms. Quorum sensing signal molecules acyl-homoserine lactones (AHLs) and autoinducer-2 (AI-2) regulate biofilm formation of many bacteria. In this study, the production of AHLs and AI-2 by one SRB strain, Desulfovibrio sp. Huiquan2017, was detected, and the effect of exogenous AI-2 on bacterial biofilm formation was discussed. It was found that the cell-free supernatants of Desulfovibrio sp. Huiquan2017 induced luminescence in a ∆luxS mutant strain Vibrio harveyi BB170, indicating the production of functional AI-2 by the bacterium. In the presence of exogenous AI-2, the growth of Desulfovibrio sp. Huiquan2017 and early biofilm formation were not affected, but the later stage of biofilm development was inhibited significantly. The biofilms became looser, smaller, and thinner, and contained less bacteria and extracellular polymeric substances (EPS). The inhibition effect of AI-2 on the biofilm development of Desulfovibrio sp. Huiquan2017 was mainly achieved through reducing the amount of EPS in biofilms. These findings shed light on the biofilm regulation of SRB.


Assuntos
Biofilmes/efeitos dos fármacos , Biofilmes/crescimento & desenvolvimento , Desulfovibrio/efeitos dos fármacos , Desulfovibrio/crescimento & desenvolvimento , Desulfovibrio/metabolismo , Homosserina/análogos & derivados , Lactonas/metabolismo , Lactonas/farmacologia , Agrobacterium tumefaciens/metabolismo , Proteínas de Bactérias/metabolismo , Proteínas de Bactérias/farmacologia , Corrosão , Matriz Extracelular de Substâncias Poliméricas/metabolismo , Homosserina/metabolismo , Homosserina/farmacologia , Percepção de Quorum , Vibrio/metabolismo
6.
Fish Shellfish Immunol ; 114: 253-262, 2021 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-33979691

RESUMO

Vibriosis, an illness caused by the Vibrio bacteria species, results in significant economic loss in olive flounder farms. Here we present a novel anti-Vibrio feed vaccine protecting multiple strains of Vibrio pathogens, a universal vaccine effect. The vaccine was generated by engineering Lactococcus lactis BFE920 to express the fusion antigens of Vibrio outer membrane protein K (OmpK) and flagellin B subunit (FlaB). These antigen genes are highly conserved among Vibrio species. Olive flounder (7.1 ± 0.8 g and 140 ± 10 g) were fed the vaccine adsorbed to a regular feed (1 × 107 CFU/g) for one week with a 1-week interval, repeating three times (a triple boost). The vaccinated fish increased the significant levels of antigen-specific antibodies, T cell numbers (CD4-1, CD4-2, and CD8α), cytokine production (T-bet and IFN-γ), and innate immune responses (TLR5M, IL-1ß, and IL-12p40). Also, the survival rates of adult and juvenile fish fed the vaccine were significantly elevated when challenged with V. anguillarum, V. alginolyticus, and V. harveyi. In addition, weight gain rate and feed conversion ratio were improved in vaccinated fish. The feed vaccine protected multiple Vibrio pathogens, a universal vaccine effect, by activating innate and adaptive immune responses. This oral vaccine may be developed as an anti-Vibrio vaccine to protect against a broad spectrum of Vibrio pathogens.


Assuntos
Proteínas de Bactérias/imunologia , Vacinas Bacterianas/imunologia , Linguado , Lactococcus lactis/metabolismo , Vibrioses/veterinária , Vibrio/metabolismo , Imunidade Adaptativa , Animais , Vacinas Bacterianas/administração & dosagem , Doenças dos Peixes/microbiologia , Doenças dos Peixes/prevenção & controle , Imunidade Inata , Probióticos , Vibrio/imunologia , Vibrioses/prevenção & controle
7.
J Bacteriol ; 203(15): e0017221, 2021 07 08.
Artigo em Inglês | MEDLINE | ID: mdl-34031037

RESUMO

Vitamin B12 belongs to a family of structurally diverse cofactors with over a dozen natural analogs, collectively referred to as cobamides. Most bacteria encode cobamide-dependent enzymes, many of which can only utilize a subset of cobamide analogs. Some bacteria employ a mechanism called cobamide remodeling, a process in which cobamides are converted into other analogs to ensure that compatible cobamides are available in the cell. Here, we characterize an additional pathway for cobamide remodeling that is distinct from the previously characterized ones. Cobamide synthase (CobS) is an enzyme required for cobamide biosynthesis that attaches the lower ligand moiety in which the base varies between analogs. In a heterologous model system, we previously showed that Vibrio cholerae CobS (VcCobS) unexpectedly conferred remodeling activity in addition to performing the known cobamide biosynthesis reaction. Here, we show that additional Vibrio species perform the same remodeling reaction, and we further characterize VcCobS-mediated remodeling using bacterial genetics and in vitro assays. We demonstrate that VcCobS acts upon the cobamide pseudocobalamin directly to remodel it, a mechanism which differs from the known remodeling pathways in which cobamides are first cleaved into biosynthetic intermediates. This suggests that some CobS homologs have the additional function of cobamide remodeling, and we propose the term "direct remodeling" for this process. This characterization of yet another pathway for remodeling suggests that cobamide profiles are highly dynamic in polymicrobial environments, with remodeling pathways conferring a competitive advantage. IMPORTANCE Cobamides are widespread cofactors that mediate metabolic interactions in complex microbial communities. Few studies directly examine cobamide profiles, but several have shown that mammalian gastrointestinal tracts are rich in cobamide analogs. Studies of intestinal bacteria, including beneficial commensals and pathogens, show variation in the ability to produce and utilize different cobamides. Some bacteria can convert imported cobamides into compatible analogs in a process called remodeling. Recent discoveries of additional cobamide remodeling pathways, including this work, suggest that remodeling is an important factor in cobamide dynamics. Characterization of such pathways is critical in understanding cobamide flux and nutrient cross-feeding in polymicrobial communities, and it facilitates the establishment of microbiome manipulation strategies via modulation of cobamide profiles.


Assuntos
Proteínas de Bactérias/metabolismo , Cobamidas/biossíntese , Vibrio cholerae/metabolismo , Proteínas de Bactérias/genética , Cobamidas/química , Estrutura Molecular , Vibrio/genética , Vibrio/metabolismo , Vibrio cholerae/química , Vibrio cholerae/genética
8.
Biophys J ; 120(11): 2124-2137, 2021 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-33812846

RESUMO

VhChiP, a sugar-specific porin found on the outer membrane of Vibrio campbellii, is responsible for the transport of chitooligosaccharides, allowing the bacterium to thrive in aquatic environments using chitin as a nutrient. We previously showed that VhChiP is composed of three identical subunits, each containing a 16-stranded ß-barrel connected by eight extracellular loops and eight short periplasmic turns. This study is focused on the specific roles of three prominent extracellular loops of VhChiP-L2, L3, and L8. The deletion of L2 completely disrupted the L2-L2 interactions, thus destabilizing the protein trimers as well as the integrity of the secondary structure. The deletion of L3 caused a drastic loss in the binding affinity for sugar substrates because of the absence of a cluster of key amino acid residues that form the affinity sites. The removal of L8 induced pronounced gating, which is highly responsive to elevated potentials. Our data provide further information on the important roles of the three prominent loops of VhChiP: loop L2 maintains the trimeric structure and the integrity of secondary structure, loop L3 controls the binding affinity for sugar substrates, and loop L8 retains the stably open state of the channel.


Assuntos
Proteínas da Membrana Bacteriana Externa , Vibrio , Proteínas da Membrana Bacteriana Externa/genética , Proteínas da Membrana Bacteriana Externa/metabolismo , Porinas/genética , Porinas/metabolismo , Estrutura Secundária de Proteína , Vibrio/metabolismo
9.
Front Immunol ; 12: 634497, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33868255

RESUMO

Programmed cell death (PCD) is an essential process for the immune system's development and homeostasis, enabling the remotion of infected or unnecessary cells. There are several PCD's types, depending on the molecular mechanisms, such as non-inflammatory or pro-inflammatory. Hemocytes are the main component of cellular immunity in bivalve mollusks. Numerous infectious microorganisms produce toxins that impair hemocytes functions, but there is little knowledge on the role of PCD in these cells. This study aims to evaluate in vitro whether marine toxins induce a particular type of PCD in hemocytes of the bivalve mollusk Crassostrea gigas during 4 h at 25°C. Hemocytes were incubated with two types of marine toxins: non-proteinaceous toxins from microalgae (saxitoxin, STX; gonyautoxins 2 and 3, GTX2/3; okadaic acid/dynophysistoxin-1, OA/DTX-1; brevetoxins 2 and 3, PbTx-2,-3; brevetoxin 2, PbTx-2), and proteinaceous extracts from bacteria (Vibrio parahaemolyticus, Vp; V. campbellii, Vc). Also, we used the apoptosis inducers, staurosporine (STP), and camptothecin (CPT). STP, CPT, STX, and GTX 2/3, provoked high hemocyte mortality characterized by apoptosis hallmarks such as phosphatidylserine translocation into the outer leaflet of the cell membrane, exacerbated chromatin condensation, DNA oligonucleosomal fragments, and variation in gene expression levels of apoptotic caspases 2, 3, 7, and 8. The mixture of PbTx-2,-3 also showed many apoptosis features; however, they did not show apoptotic DNA oligonucleosomal fragments. Likewise, PbTx-2, OA/DTX-1, and proteinaceous extracts from bacteria Vp, and Vc, induced a minor degree of cell death with high gene expression of the pro-inflammatory initiator caspase-1, which could indicate a process of pyroptosis-like PCD. Hemocytes could carry out both PCD types simultaneously. Therefore, marine toxins trigger PCD's signaling pathways in C. gigas hemocytes, depending on the toxin's nature, which appears to be highly conserved both structurally and functionally.


Assuntos
Apoptose/efeitos dos fármacos , Toxinas Bacterianas/toxicidade , Crassostrea/efeitos dos fármacos , Hemócitos/efeitos dos fármacos , Toxinas Marinhas/toxicidade , Animais , Toxinas Bacterianas/isolamento & purificação , Caspases/metabolismo , Montagem e Desmontagem da Cromatina/efeitos dos fármacos , Crassostrea/imunologia , Crassostrea/metabolismo , Quebras de DNA de Cadeia Dupla , Hemócitos/imunologia , Hemócitos/metabolismo , Hemócitos/patologia , Fosfatidilserinas/metabolismo , Vibrio/metabolismo , Vibrio parahaemolyticus/metabolismo
10.
mBio ; 12(2)2021 04 27.
Artigo em Inglês | MEDLINE | ID: mdl-33906925

RESUMO

Unlike nucleobase modifications in canonical restriction-modification systems, DNA phosphorothioate (PT) epigenetic modification occurs in the DNA sugar-phosphate backbone when the nonbridging oxygen is replaced by sulfur in a double-stranded (ds) or single-stranded (ss) manner governed by DndABCDE or SspABCD, respectively. SspABCD coupled with SspE constitutes a defense barrier in which SspE depends on sequence-specific PT modifications to exert its antiphage activity. Here, we identified a new type of ssDNA PT-based SspABCD-SspFGH defense system capable of providing protection against phages through a mode of action different from that of SspABCD-SspE. We provide further evidence that SspFGH damages non-PT-modified DNA and exerts antiphage activity by suppressing phage DNA replication. Despite their different defense mechanisms, SspFGH and SspE are compatible and pair simultaneously with one SspABCD module, greatly enhancing the protection against phages. Together with the observation that the sspBCD-sspFGH cassette is widely distributed in bacterial genomes, this study highlights the diversity of PT-based defense barriers and expands our knowledge of the arsenal of phage defense mechanisms.IMPORTANCE We recently found that SspABCD, catalyzing single-stranded (ss) DNA phosphorothioate (PT) modification, coupled with SspE provides protection against phage infection. SspE performs both PT-simulated NTPase and DNA-nicking nuclease activities to damage phage DNA, rendering SspA-E a PT-sensing defense system. To our surprise, ssDNA PT modification can also pair with a newly identified 3-gene sspFGH cassette to fend off phage infection with a different mode of action from that of SspE. Interestingly, both SspFGH and SspE can pair with the same SspABCD module for antiphage defense, and their combination provides Escherichia coli JM109 with additive phage resistance up to 105-fold compared to that for either barrier alone. This agrees with our observation that SspFGH and SspE coexist in 36 bacterial genomes, highlighting the diversity of the gene contents and molecular mechanisms of PT-based defense systems.


Assuntos
Subfamília D de Transportador de Cassetes de Ligação de ATP/genética , Proteínas de Bactérias/genética , DNA Bacteriano/genética , Fosfatos , Vibrio/genética , Subfamília D de Transportador de Cassetes de Ligação de ATP/classificação , Subfamília D de Transportador de Cassetes de Ligação de ATP/metabolismo , Proteínas de Bactérias/metabolismo , Bacteriófagos/genética , Bacteriófagos/patogenicidade , Escherichia coli/genética , Genoma Bacteriano , Vibrio/metabolismo
11.
J Agric Food Chem ; 69(11): 3380-3389, 2021 Mar 24.
Artigo em Inglês | MEDLINE | ID: mdl-33705122

RESUMO

Carbohydrates are structurally and functionally diverse materials including polysaccharides, and marine organisms are known to have many enzymes for the breakdown of complex polysaccharides. Here, we identified an α-l-fucosidase enzyme from the marine bacterium Vibrio sp. strain EJY3 (VejFCD) that has dual α-1,4-glucosidic and ß-1,4-galactosidic specificities. We determined the crystal structure of VejFCD and provided the structural basis underlying the dual α- and ß-glycosidase activities of the enzyme. Unlike other three-domain FCDs, in VejFCD, carbohydrate-binding module-B (CBM-B) with a novel ß-sandwich fold tightly contacts with the CatD/CBM-B main body and provides key residues for the ß-1,4-glycosidase activity of the enzyme. The phylogenetic tree analysis suggests that only a few FCDs from marine microorganisms have the key structural features for dual α-1,4- and ß-1,4-glycosidase activities. This study provides the structural insights into the mechanism underlying the novel glycoside hydrolase activities and could be applied for more efficient utilization in the hydrolysis of complex carbohydrates in biotechnological applications.


Assuntos
Vibrio , alfa-L-Fucosidase , Carboidratos , Glicosídeo Hidrolases/genética , Glicosídeo Hidrolases/metabolismo , Hidrólise , Filogenia , Especificidade por Substrato , Vibrio/metabolismo , alfa-L-Fucosidase/genética , alfa-L-Fucosidase/metabolismo
13.
Int J Biol Macromol ; 174: 457-465, 2021 Mar 31.
Artigo em Inglês | MEDLINE | ID: mdl-33493561

RESUMO

Vibrio anguillarum is a globally distributed aquatic pathogen, and its flagellin B (FlaB) protein can evoke innate immune responses in hosts. In order to explore the role of FlaB in V. anguillarum infection, we constructed a FlaB-deficient mutant using overlapping PCR and two-step homologous recombination, and gene sequencing confirmed successful knockout of the FlaB gene. Scanning electron microscopy showed no significant differences in the morphological structure of the flagellum between wild-type and FlaB-deficient strains. The mutant was subsequently injected into the freshwater prawn (Macrobrachium rosenbergii) to explore its pathogenicity in the host, and expression of myeloid differentiation factor 88, prophenoloxidase, catalase, superoxide dismutase and glutathione peroxidase was investigated by real-time PCR. The results showed that deletion of FlaB had little effect on V. anguillarum-induced expression of these immune-related genes (p > 0.05). In general, the FlaB mutant displayed similar flagella morphology and immune characteristics to the wild-type strain, hence we speculated that knockout of FlaB might promote the expression and function of other flagellin proteins. Furthermore, this study provides a rapid and simple method for obtaining stable mutants of V. anguillarum free from foreign plasmid DNA.


Assuntos
Proteínas de Artrópodes/genética , Flagelina/administração & dosagem , Mutação , Palaemonidae/imunologia , Vibrio/metabolismo , Animais , Proteínas de Bactérias/administração & dosagem , Proteínas de Bactérias/genética , Proteínas de Bactérias/imunologia , Catalase/genética , Catecol Oxidase/genética , Clonagem Molecular , Precursores Enzimáticos/genética , Flagelina/genética , Flagelina/imunologia , Regulação da Expressão Gênica , Técnicas de Inativação de Genes , Glutationa Peroxidase/genética , Imunidade Inata , Microscopia Eletrônica de Varredura , Fator 88 de Diferenciação Mieloide/genética , Palaemonidae/genética , Superóxido Dismutase/genética , Vibrio/imunologia
14.
Electron J Biotechnol ; 49: 22-28, Jan. 2021. ilus, graf, tab
Artigo em Inglês | LILACS | ID: biblio-1291938

RESUMO

BACKGROUND: Vibrio species display variable and plastic fitness strategies to survive and interact with multiple hosts, including marine aquaculture species that are severely affected by pathogenic Vibrios. The culturable Vibrio sp. strain ArtGut-C1, the focus of this study, provides new evidence of such phenotypic plasticity as it accumulates polyhydroxybutyrate (PHB), a biodegradable polymer with anti-pathogen activity, particularly in the marine larviculture phase. The strain was isolated from the gut of laboratory-reared Artemia individuals, the live diet and PHB carrier used in larviculture. Its main phenotypic properties, taxonomic status and genomic properties are reported based on the whole-genome sequencing. RESULTS: Vibrio sp. ArtGut-C1 yielded 72.6% PHB of cells' dry weight at 25 C. The genomic average nucleotide identity (ANI) shows it is closely related to V. diabolicus (ANI: 88.6%). Its genome contains 5,236,997- bp with 44.8% GC content, 3,710 protein-coding sequences, 96 RNA, 9 PHB genes functionally related to PHB metabolic pathways, and several genes linked to competing and colonizing abilities. CONCLUSIONS: This culturable PHB-accumulating Vibrio strain shows high genomic and phenotypic variability. It may be used as a natural pathogen biocontrol in the marine hatchery and as a potential cell factory for PHB production.


Assuntos
Animais , Artemia/microbiologia , Vibrio/metabolismo , Poli-Hidroxialcanoatos/metabolismo , Hidroxibutiratos/metabolismo , Variação Genética , Vibrio/isolamento & purificação , Vibrio/classificação , Aquicultura , Probióticos , Crustáceos/microbiologia , Microbioma Gastrointestinal , Variação Biológica da População
15.
J Fish Dis ; 44(5): 591-599, 2021 May.
Artigo em Inglês | MEDLINE | ID: mdl-33210340

RESUMO

Vibriosis caused by luminous Vibrio species is one of the biggest challenges to shrimp industry in Bangladesh. This study aimed to characterize whole microbial communities from Vibrio-infected black tiger shrimp (Penaeus monodon) using 16S rRNA-based amplicon sequencing. A total of 36 disease-free and infected shrimp were collected from six different hatcheries in Bagerhat, Bangladesh. A final pool of 12 samples (n = 6) was created by homogenization of the hepatopancreas samples from three shrimps collected from each hatchery for the same group. The amplicon sequencing data revealed significant (p < .05) decrease of alpha diversity measurements and subsequent effects (p < .05) on the hepatopancreas microbiota in the infected group, compared to control shrimp. Proteobateria and Aeromonas were the most dominant bacteria at phylum and genus level in both groups and identified as core microbiota in the community. Two bacterial groups at phyla level and eight at genus level were found associated with the alteration of hepatopancreas microbial communities and associated gene functions in vibriosis-infected shrimp, revealed by differential abundance and KEGG pathway analysis. The overwhelming abundance of Citroibacter, Shewanella and Candidatus lineages in vibriosis-infected shrimp needs further investigations.


Assuntos
Genes Bacterianos , Penaeidae/microbiologia , RNA Bacteriano/genética , RNA Ribossômico 16S/genética , Vibrio/genética , Animais , RNA Bacteriano/metabolismo , RNA Ribossômico 16S/metabolismo , Análise de Sequência de RNA , Vibrio/metabolismo
16.
Environ Microbiol ; 23(1): 160-173, 2021 01.
Artigo em Inglês | MEDLINE | ID: mdl-33107668

RESUMO

A major conundrum in the isolation of prokaryotes from open environments is stochasticity. It is especially difficult to study low abundance groups where very little biological information exists, although single-cell genomics and metagenomics have alleviated some of this bottleneck. Here, we report an approach to capture lignin-utilizing bacteria by linking a physical model to actual organisms. Extracellular enzymes, lignin degradation and cell growth are crucial phenotypes of lignin-utilizing bacteria, but their interrelationships remain poorly understood. In this study, the phenotypes of bacteria isolated from in situ lignocellulose enrichment samples in coastal waters were traced and statistically analysed. It suggested cell growth, dye-decolorizing peroxidase (DyP) and reactive oxygen species (ROS) were significantly correlated with lignin degradation, exhibiting a genus-specific property. The established models enabled us to efficiently capture lignin-utilizing bacteria and rapidly evaluate lignin degradation for Bacillus and Vibrio strains. Through the model, we identified several previously unrecognized marine bacterial lignin degraders. Moreover, it demonstrated that the isolated marine lignin-utilizing bacteria employ a DyP-based system and ROS for lignin depolymerization, providing insights into the mechanism of marine bacterial lignin degradation. Our findings should have implications beyond the capture of lignin-utilizing bacteria, in the isolation of other microorganisms with as-yet-unknown molecular biomarkers.


Assuntos
Bacillus/metabolismo , Lignina/metabolismo , Peroxidase/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Vibrio/metabolismo , Organismos Aquáticos/metabolismo , Bacillus/isolamento & purificação , Fenômenos Bioquímicos , Oxirredutases/metabolismo , Peroxidases/metabolismo , Metabolismo Secundário/fisiologia , Vibrio/isolamento & purificação
17.
Eur J Med Chem ; 209: 112883, 2021 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-33035924

RESUMO

Methionine aminopeptidases (MetAPs) have been recognized as drug targets and have been extensively studied for discovery of selective inhibitors. MetAPs are essential enzymes in all living cells. While most prokaryotes contain a single gene, some prokaryotes and all eukaryotes including human have redundancy. Due to the similarity in the active sites of the MetAP enzyme between the pathogens and human limited the success of discovering selective inhibitors. We recently have discovered that MetAPs with small inserts within the catalytic domain to have different susceptibilities against some inhibitors compared to those that do not have. Using this clue we used bioinformatic tools to identify new variants of MetAPs with inserts in pathogenic species. Two new isoforms were identified in Vibrio species with two and three inserts in addition to an isoform without any insert. Multiple sequence alignment suggested that inserts are conserved in several of the Vibrio species. Two of the three inserts are common between two and three insert isoforms. One of the inserts is identified to have "NNKNN" motif that is similar to well-characterized quorum sensing peptide, "NNWNN". Another insert is predicted to have a posttranslational modification site. Three Vibrio proteins were cloned, expressed, purified, enzyme kinetics established and inhibitor screening has been performed. Several of the pyridinylpyrimidine derivatives selectively inhibited MetAPs with inserts compared to those that do not have, including the human enzyme. Crystal structure and molecular modeling studies provide the molecular basis for selective inhibition.


Assuntos
Proteínas de Bactérias/antagonistas & inibidores , Inibidores Enzimáticos/química , Inibidores Enzimáticos/farmacologia , Metionil Aminopeptidases/antagonistas & inibidores , Vibrio/enzimologia , Sequência de Aminoácidos , Antibacterianos/química , Antibacterianos/farmacologia , Proteínas de Bactérias/química , Proteínas de Bactérias/metabolismo , Domínio Catalítico/efeitos dos fármacos , Cristalografia por Raios X , Humanos , Metionil Aminopeptidases/química , Metionil Aminopeptidases/metabolismo , Simulação de Acoplamento Molecular , Isoformas de Proteínas/antagonistas & inibidores , Isoformas de Proteínas/química , Isoformas de Proteínas/metabolismo , Pirimidinas/química , Pirimidinas/farmacologia , Vibrio/química , Vibrio/metabolismo
18.
Arch Microbiol ; 203(1): 399-404, 2021 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-32844278

RESUMO

Description of a Gram-negative, motile, circular-shaped bacterial strain, designated A511T obtained from the skin of the pufferfish Sphoeroides spengleri (Family Tetraodontidae), collected in Arraial do Cabo, Brazil. Optimum growth occurs at 20-28 °C in the presence of 3% NaCl. The genome sequence of the novel isolate consisted of 4.36 Mb, 3,976 coding genes and G + C content of 42.5%. Genomic taxonomy analyses based on average amino acid (AAI), genome-to-genome-distance (GGDH) and phylogenetic reconstruction placed A511T (= CBAS 712T = CAIM 1939T) into a new species of the genus Vibrio (Vibrio tetraodonis sp. nov.). The genome of the novel species contains eight genes clusters (~ 183.9 Kbp in total) coding for different types of bioactive compounds that hint to several possible ecological roles in the pufferfish host.


Assuntos
Genoma Bacteriano/genética , Filogenia , Vibrio/classificação , Vibrio/genética , Composição de Bases , Brasil , RNA Ribossômico 16S/genética , Cloreto de Sódio/metabolismo , Especificidade da Espécie , Vibrio/crescimento & desenvolvimento , Vibrio/metabolismo
19.
mSphere ; 5(6)2020 12 16.
Artigo em Inglês | MEDLINE | ID: mdl-33328352

RESUMO

The type II secretion system (T2SS) is a conserved transport pathway responsible for the secretion of a range of virulence factors by many pathogens, including Vibrio cholerae Disruption of the T2SS genes in V. cholerae results in loss of secretion, changes in cell envelope function, and growth defects. While T2SS mutants are viable, high-throughput genomic analyses have listed these genes among essential genes. To investigate whether secondary mutations arise as a consequence of T2SS inactivation, we sequenced the genomes of six V. cholerae T2SS mutants with deletions or insertions in either the epsG, epsL, or epsM genes and identified secondary mutations in all mutants. Two of the six T2SS mutants contain distinct mutations in the gene encoding the T2SS-secreted protease VesC. Other mutations were found in genes coding for V. cholerae cell envelope proteins. Subsequent sequence analysis of the vesC gene in 92 additional T2SS mutant isolates identified another 19 unique mutations including insertions or deletions, sequence duplications, and single-nucleotide changes resulting in amino acid substitutions in the VesC protein. Analysis of VesC variants and the X-ray crystallographic structure of wild-type VesC suggested that all mutations lead to loss of VesC production and/or function. One possible mechanism by which V. cholerae T2SS mutagenesis can be tolerated is through selection of vesC-inactivating mutations, which may, in part, suppress cell envelope damage, establishing permissive conditions for the disruption of the T2SS. Other mutations may have been acquired in genes encoding essential cell envelope proteins to prevent proteolysis by VesC.IMPORTANCE Genome-wide transposon mutagenesis has identified the genes encoding the T2SS in Vibrio cholerae as essential for viability, but the reason for this is unclear. Mutants with deletions or insertions in these genes can be isolated, suggesting that they have acquired secondary mutations that suppress their growth defect. Through whole-genome sequencing and phenotypic analysis of T2SS mutants, we show that one means by which the growth defect can be suppressed is through mutations in the gene encoding the T2SS substrate VesC. VesC homologues are present in other Vibrio species and close relatives, and this may be why inactivation of the T2SS in species such as Vibrio vulnificus, Vibrio sp. strain 60, and Aeromonas hydrophila also results in a pleiotropic effect on their outer membrane assembly and integrity.


Assuntos
Proteínas de Bactérias/química , Proteínas de Membrana/química , Vibrio cholerae/genética , Vibrio/metabolismo , Sequência de Aminoácidos , Proteínas de Bactérias/genética , Cristalografia por Raios X , Proteínas de Membrana/genética , Mutagênese , Mutação , Peptídeo Hidrolases/metabolismo , Supressão Genética
20.
Environ Microbiol ; 22(12): 5467-5482, 2020 12.
Artigo em Inglês | MEDLINE | ID: mdl-33169914

RESUMO

Amphiphilic siderophores, including amphibactins, are the most abundant siderophores in oceans. Genes putatively encoding the amphibactin system were proposed in some bacteria and homologues of these genes are particularly abundant in multiple bacterial lineages inhabitant of low-iron seawater. However, since no defective mutant strains in any of these genes were studied to date, their role in amphibactin synthesis or uptake was not demonstrated. In this work, an in silico analysis of the genome of the mollusc pathogen Vibrio neptunius leads us to identify a gene cluster (denoted absABDEF) that is predicted to encode an amphibactin-like siderophore and several mutant strains unable to synthesize or use siderophores were constructed. The results showed that genes absABDEF are required for amphibactin synthesis. A comparative chemical analysis of V. neptunius wild type and biosynthesis mutants allowed us to identify a mixture of nine amphibactin forms produced by this bacterium. In addition, the gene abtA is predicted to encode the ferri-amphibactin outer membrane transporter. The prevalence of the amphibactin system in bivalve hemolymph microbiota was also studied. We found that the amphibactin system is widespread in hemolymph microbiota including both commensal and pathogenic bacterial species. Thus, its contribution to bacterial fitness must be more related to environmental persistence than to pathogenicity.


Assuntos
Proteínas de Bactérias/metabolismo , Bivalves/microbiologia , Microbiota , Sideróforos/biossíntese , Vibrio/metabolismo , Animais , Proteínas de Bactérias/genética , Genoma Bacteriano/genética , Microbiota/genética , Família Multigênica , Mutação , Água do Mar/microbiologia , Sideróforos/genética , Vibrio/genética
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