scholarly journals Characterization of FliL Proteins in Bradyrhizobium diazoefficiens: Lateral FliL Supports Swimming Motility, and Subpolar FliL Modulates the Lateral Flagellar System

2019 ◽  
Vol 202 (5) ◽  
Author(s):  
Florencia Mengucci ◽  
Carolina Dardis ◽  
Elías J. Mongiardini ◽  
María J. Althabegoiti ◽  
Jonathan D. Partridge ◽  
...  
Keyword(s):  
Collective Motion ◽  
Bacterial Species ◽  
Bacterial Motility ◽  
Deletion Mutants ◽  
Semisolid Medium ◽  
Content Type ◽  
Lateral Flagellum ◽  
Important Trait ◽  
Flagellar Proteins

ABSTRACT Bradyrhizobium diazoefficiens is a soil alphaproteobacterium that possesses two evolutionarily distinct flagellar systems, a constitutive subpolar flagellum and inducible lateral flagella that, depending on the carbon source, may be expressed simultaneously in liquid medium and used interactively for swimming. In each system, more than 30 genes encode the flagellar proteins, most of which are well characterized. Among the exceptions is FliL, which has been scarcely studied in alphaproteobacteria and whose function in other bacterial classes is somewhat controversial. Because each B. diazoefficiens flagellar system contains its own fliL paralog, we obtained the respective deletions ΔfliLS (subpolar) and ΔfliLL (lateral) to study their functions in swimming. We determined that FliLL was essential for lateral flagellum-driven motility. FliLS was dispensable for swimming in either liquid or semisolid medium; however, it was found to play a crucial role in upregulation of the lateral flagellum regulon under conditions of increased viscosity/flagellar load. Therefore, although FliLS seems to be not essential for swimming, it may participate in a mechanosensor complex that controls lateral flagellum induction. IMPORTANCE Bacterial motility propelled by flagella is an important trait in most environments, where microorganisms must explore the habitat toward beneficial resources and evade toxins. Most bacterial species have a unique flagellar system, but a few species possess two different flagellar systems in the same cell. An example is Bradyrhizobium diazoefficiens, the N2-fixing symbiont of soybean, which uses both systems for swimming. Among the less-characterized flagellar proteins is FliL, a protein typically associated with a flagellum-driven surface-based collective motion called swarming. By using deletion mutants in each flagellar system’s fliL, we observed that one of them (lateral) was required for swimming, while the other (subpolar) took part in the control of lateral flagellum synthesis. Hence, this protein seems to participate in the coordination of activity and production of both flagellar systems.

scholarly journals Characterization of a Feruloyl Esterase from Lactobacillus plantarum

2013 ◽  
Vol 79 (17) ◽  
pp. 5130-5136 ◽  
Author(s):  
María Esteban-Torres ◽  
Inés Reverón ◽  
José Miguel Mancheño ◽  
Blanca de las Rivas ◽  
Rosario Muñoz
Keyword(s):  
Lactobacillus Plantarum ◽  
Bacterial Species ◽  
Hydrolytic Activity ◽  
Feruloyl Esterase ◽  
Gene Encoding ◽  
Content Type ◽  
Cell Extracts ◽  
Methyl Ferulate ◽  
Methyl Caffeate

ABSTRACTLactobacillus plantarumis frequently found in the fermentation of plant-derived food products, where hydroxycinnamoyl esters are abundant.L. plantarumWCFS1 cultures were unable to hydrolyze hydroxycinnamoyl esters; however, cell extracts from the strain partially hydrolyze methyl ferulate and methylp-coumarate. In order to discover whether the protein Lp_0796 is the enzyme responsible for this hydrolytic activity, it was recombinantly overproduced and enzymatically characterized. Lp_0796 is an esterase that, among other substrates, is able to efficiently hydrolyze the four model substrates for feruloyl esterases (methyl ferulate, methyl caffeate, methylp-coumarate, and methyl sinapinate). A screening test for the detection of the gene encoding feruloyl esterase Lp_0796 revealed that it is generally present amongL. plantarumstrains. The present study constitutes the description of feruloyl esterase activity inL. plantarumand provides new insights into the metabolism of hydroxycinnamic compounds in this bacterial species.

scholarly journals Genomic Characterization of VIM Metallo-β-Lactamase-Producing Alcaligenes faecalis from Gaza, Palestine

2017 ◽  
Vol 61 (11) ◽  
Author(s):  
Nahed Al Laham ◽  
Kalyan D. Chavda ◽  
Astrid V. Cienfuegos-Gallet ◽  
Barry N. Kreiswirth ◽  
Liang Chen
Keyword(s):  
Bacterial Species ◽  
Alcaligenes Faecalis ◽  
Gram Negative Bacteria ◽  
Sequencing Analysis ◽  
Large Hospital ◽  
Content Type ◽  
Next Generation Sequencing Analysis ◽  
Genomic Characterization ◽  
Generation Sequencing

ABSTRACT Carbapenemase-producing Gram-negative bacteria (CP-GNB) have increasingly spread worldwide, and different families of carbapenemases have been identified in various bacterial species. Here, we report the identification of five VIM metallo-β-lactamase-producing Alcaligenes faecalis isolates associated with a small outbreak in a large hospital in Gaza, Palestine. Next-generation sequencing analysis showed bla VIM-2 is harbored by a chromosomal genomic island among three strains, while bla VIM-4 is carried by a novel plasmid in two strains.

scholarly journals Characterization of Phascolarctobacterium succinatutens sp. nov., an Asaccharolytic, Succinate-Utilizing Bacterium Isolated from Human Feces

2011 ◽  
Vol 78 (2) ◽  
pp. 511-518 ◽  
Author(s):  
Yohei Watanabe ◽  
Fumiko Nagai ◽  
Masami Morotomi
Keyword(s):  
Bacterial Species ◽  
Short Chain Fatty Acids ◽  
Pcr Analysis ◽  
Rrna Gene ◽  
Gi Tract ◽  
Bacterial Strains ◽  
High Sequence Identity ◽  
Healthy Human ◽  
Content Type

ABSTRACTIsolation, cultivation, and characterization of the intestinal microorganisms are important for understanding the comprehensive physiology of the human gastrointestinal (GI) tract microbiota. Here, we isolated two novel bacterial strains, YIT 12067Tand YIT 12068, from the feces of healthy human adults. Phylogenetic analysis indicated that they belonged to the same species and were most closely related toPhascolarctobacterium faeciumACM 3679T, with 91.4% to 91.5% 16S rRNA gene sequence similarities, respectively. Substrate availability tests revealed that the isolates used only succinate; they did not ferment any other short-chain fatty acids or carbohydrates tested. When these strains were cocultured with the xylan-utilizing and succinate-producing bacteriumParaprevotella xylaniphilaYIT 11841T, in medium supplemented with xylan but not succinate, their cell numbers became 2 to 3 orders of magnitude higher than those of the monoculture; succinate became undetectable, and propionate was formed. Database analysis revealed that over 200 uncultured bacterial clones from the feces of humans and other mammals showed high sequence identity (>98.7%) to YIT 12067T. Real-time PCR analysis also revealed that YIT 12067T-like bacteria were present in 21% of human fecal samples, at an average level of 3.34 × 108cells/g feces. These results indicate that YIT 12067T-like bacteria are distributed broadly in the GI tract as subdominant members that may adapt to the intestinal environment by specializing to utilize the succinate generated by other bacterial species. The phylogenetic and physiological properties of YIT 12067Tand YIT 12068 suggest that these strains represent a novel species, which we have designatedPhascolarctobacterium succinatutenssp. nov.

scholarly journals Characterization of a Diffusible Signaling Factor from Xylella fastidiosa

mBio ◽  
2013 ◽  
Vol 4 (1) ◽  
Author(s):  
Ellen D. Beaulieu ◽  
Michael Ionescu ◽  
Subhadeep Chatterjee ◽  
Kenji Yokota ◽  
Dirk Trauner ◽  
...  
Keyword(s):  
Cell Signaling ◽  
Unsaturated Fatty Acids ◽  
Xylella Fastidiosa ◽  
Bacterial Species ◽  
Content Type ◽  
Isolation And Characterization ◽  
High Level ◽  
Diffusible Signaling Factor ◽  
Cell Cell

ABSTRACTCell-cell signaling inXylella fastidiosahas been implicated in the coordination of traits enabling colonization in plant hosts as well as insect vectors. This cell density-dependent signaling has been attributed to a diffusible signaling factor (DSF) produced by the DSF synthase RpfF. DSF produced by related bacterial species are unsaturated fatty acids, but that ofX. fastidiosawas thought to be different from those of other taxa. We describe here the isolation and characterization of anX. fastidiosaDSF (XfDSF) as 2(Z)-tetradecenoic acid. This compound was isolated both from recombinantErwinia herbicolaexpressingX. fastidiosa rpfFand from anX. fastidiosa rpfCdeletion mutant that overproduces DSF. Since anrpfFmutant is impaired in biofilm formation and underexpresses the hemagglutinin-like protein-encoding geneshxfAandhxfB, we demonstrate that these traits can be restored by ca. 0.5 µMXfDSF but not by myristic acid, the fully saturated tetradecenoic acid. AphoA-basedX. fastidiosabiosensor that assesses DSF-dependent expression ofhxfAorhxfBrevealed a high level of molecular specificity of DSF signaling.IMPORTANCEX. fastidiosacauses diseases in many important plants, including grape, where it incites Pierce’s disease. Virulence ofX. fastidiosafor grape is coordinated by cell-cell signaling molecules, designated DSF (Diffusible Signaling Factor). Mutants blocked in DSF production are hypervirulent for grape, suggesting that virulence is suppressed upon DSF accumulation and that disease could be controlled by artificial elevation of the DSF level in plants. In this work, we describe the isolation of the DSF produced byX. fastidiosaand the verification of its biological activity as an antivirulence factor. We also have developedX. fastidiosaDSF biosensors to evaluate the specificity of cell-cell signaling to be investigated.

scholarly journals Characterization of Copy Number Control of Two Haloferax volcanii Replication Origins Using Deletion Mutants and Haloarchaeal Artificial Chromosomes

2017 ◽  
Vol 200 (1) ◽  
Author(s):  
Sandy Maurer ◽  
Katharina Ludt ◽  
Jörg Soppa
Keyword(s):  
Copy Number ◽  
Haloferax Volcanii ◽  
Deletion Mutants ◽  
Genome Copy ◽  
Content Type ◽  
Artificial Chromosomes ◽  
Chromosome Copy Number ◽  
The Core ◽  
Genome Copy Number

ABSTRACT Haloferax volcanii is polyploid and contains about 20 genome copies under optimal conditions. However, the chromosome copy number is highly regulated and ranges from two during phosphate starvation to more than 40 under conditions of phosphate surplus. The aim of this study was the characterization of the influence of two replication origins on the genome copy number. The origin repeats and the genes encoding origin recognition complex (ORC) proteins were deleted. The core origin oriC1-orc1 (ori1) deletion mutant had a lower genome copy number and a higher level of fitness than the wild type, in stark contrast to the oriC2-orc5 (ori2) deletion mutant. The genes adjacent to ori1 could not be deleted, and thus, at least two of them are probably essential, while deletion of the genes adjacent to ori2 was possible. Various fragments of and around the origins were cloned into a suicide plasmid to generate haloarchaeal artificial chromosomes (HACs). The copy number of the oriC1-orc1 HAC was much higher than that of the oriC2-orc5 HAC. The addition of adjacent genes influenced both the HAC copy number and the chromosome copy number. The results indicate that the origins of H. volcanii are not independent but that the copy number is regulated via a network of genes around the origins. IMPORTANCE Several species of archaea have more than one origin of replication on their major chromosome and are thus the only known prokaryotic species that allow the analysis of the evolution of multiorigin replication. The widely studied Haloferax volcanii H26 strain has a major chromosome with four origins of replication. Two origins, ori1 and ori2, were chosen for an in-depth analysis using deletion mutants and haloarchaeal artificial chromosomes. The analysis was not restricted to the core origin regions; origin-adjacent genes were also included. Because H. volcanii is polyploid, the effects on the chromosome copy number were of specific importance. The results revealed extreme differences between the two origins.

scholarly journals Complete Genome Sequences of Two Mycobacterium smegmatis Bacteriophages

2021 ◽  
Vol 10 (1) ◽  
Author(s):  
Darlenys Sanchez ◽  
Anna Acosta ◽  
Nicole Skoumpourdis ◽  
Regina Alvarez ◽  
Bernadette J. Connors
Keyword(s):  
Mycobacterium Smegmatis ◽  
Complete Genome ◽  
Bacterial Species ◽  
Genome Sequences ◽  
Content Type ◽  
Isolation And Characterization

ABSTRACT Mycobacterium smegmatis, an acid-fast bacterial species in the phylum Actinobacteria, has often been used as a substitute for pathogenic mycobacteria in research. Here, we describe the isolation and characterization of two M. smegmatis bacteriophages, Penelope2018 and Miniwave.

scholarly journals Characterization of the Novel DNA Gyrase Inhibitor AZD0914: Low Resistance Potential and Lack of Cross-Resistance in Neisseria gonorrhoeae

2014 ◽  
Vol 59 (3) ◽  
pp. 1478-1486 ◽  
Author(s):  
Richard A. Alm ◽  
Sushmita D. Lahiri ◽  
Amy Kutschke ◽  
Linda G. Otterson ◽  
Robert E. McLaughlin ◽  
...  
Keyword(s):  
Neisseria Gonorrhoeae ◽  
Oral Treatment ◽  
Bacterial Species ◽  
Treatment Options ◽  
Dna Gyrase ◽  
Cross Resistance ◽  
Topoisomerase Iv ◽  
Content Type

ABSTRACTThe unmet medical need for novel intervention strategies to treatNeisseria gonorrhoeaeinfections is significant and increasing, as rapidly emerging resistance in this pathogen is threatening to eliminate the currently available treatment options. AZD0914 is a novel bacterial gyrase inhibitor that possesses potentin vitroactivities against isolates with high-level resistance to ciprofloxacin and extended-spectrum cephalosporins, and it is currently in clinical development for the treatment ofN. gonorrhoeaeinfections. The propensity to develop resistance against AZD0914 was examined inN. gonorrhoeaeand found to be extremely low, a finding supported by similar studies withStaphylococcus aureus. The genetic characterization of both first-step and second-step mutants that exhibited decreased susceptibilities to AZD0914 identified substitutions in the conserved GyrB TOPRIM domain, confirming DNA gyrase as the primary target of AZD0914 and providing differentiation from fluoroquinolones. The analysis of available bacterial gyrase and topoisomerase IV structures, including those bound to fluoroquinolone and nonfluoroquinolone inhibitors, has allowed the rationalization of the lack of cross-resistance that AZD0914 shares with fluoroquinolones. Microbiological susceptibility data also indicate that the topoisomerase inhibition mechanisms are subtly different betweenN. gonorrhoeaeand other bacterial species. Taken together, these data support the progression of AZD0914 as a novel treatment option for the oral treatment ofN. gonorrhoeaeinfections.

scholarly journals Secretion of Flagellar Proteins by the Pseudomonas aeruginosa Type III Secretion-Injectisome System

2015 ◽  
Vol 197 (12) ◽  
pp. 2003-2011 ◽  
Author(s):  
Dilek Ince ◽  
Fayyaz S. Sutterwala ◽  
Timothy L. Yahr
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Type Iii Secretion ◽  
Bacterial Species ◽  
Opportunistic Pathogen ◽  
Host Cells ◽  
Inflammasome Activation ◽  
Content Type ◽  
Type Iii ◽  
Amino Terminal ◽  
Flagellar Proteins

ABSTRACTThe opportunistic pathogenPseudomonas aeruginosautilizes an injectisome-type III secretion system (injectisome-T3SS) to elicit cytotoxicity toward epithelial cells and macrophages. Macrophage killing results from the cytotoxic properties of the translocated effector proteins (ExoS, ExoT, ExoU, and ExoY) and inflammasome-mediated induction of pyroptosis. Inflammasome activation can occur following Nlrc4-mediated recognition of cytosolic translocated flagellin (FliC). In the present study, we demonstrate that FliC is a secretion substrate of both the injectisome- and flagellum-associated T3SSs. Molecular analyses indicate that the first 20 amino-terminal residues of FliC are sufficient for secretion by the injectisome-T3SS and that the first 100 residues are sufficient for translocation of FliC into host cells. Although maximal inflammasome activation requires FliC, activation can also occur in the absence of FliC. This prompted us to examine whether other flagellar components might also be translocated into cells to elicit inflammasome activation. Indeed, we find that the flagellar cap (FliD), hook-associated (FlgK and FlgL), hook (FlgE), and rod (FlgE) proteins are secretion substrates of the injectisome-T3SS. None of these proteins, however, result in increased inflammasome activation when they are overexpressed in afliCmutant and appear to be translocated into host cells. While a role in inflammasome activation has been excluded, these data raise the possibility that flagellar components, which are highly conserved between different bacterial species, trigger other specific host responses from the extracellular milieu or contribute to the pathogenesis ofP. aeruginosa.IMPORTANCEThe inflammasome is a host defense mechanism that recognizes invading bacteria and triggers an inflammatory immune response. The opportunistic pathogenP. aeruginosaproduces both inflammasome agonists and antagonists. In this study, we demonstrate that overexpression of an agonist suppresses the activity of an antagonist, thereby resulting in inflammasome activation. Since the relative expression levels of agonists and antagonists likely vary between strains, these differences could be important predictors of whether a particularP. aeruginosastrain elicits inflammasome activation.

scholarly journals Characterization of the pJB12 Plasmid from Pseudomonas aeruginosa Reveals Tn6352, a Novel Putative Transposon Associated with Mobilization of the blaVIM-2-Harboring In58 Integron

2017 ◽  
Vol 61 (5) ◽  
Author(s):  
João Botelho ◽  
Filipa Grosso ◽  
Luísa Peixe
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Inverted Repeat ◽  
Chromosomal Location ◽  
Bacterial Species ◽  
Content Type

ABSTRACT The bla VIM-2-carrying In58 integron has been linked to a chromosomal location in different bacterial species, including Pseudomonas aeruginosa. This work reports the first fully sequenced In58-harboring plasmid, which is significantly different from the two previously identified bla VIM-2-carrying plasmids in P. aeruginosa. bla VIM-2 might have been acquired by transposition of Tn6352, a novel transposon composed of the In58 and ISPa17 elements. The recognition of similar inverted repeat (IR) sites by ISPa17 reveals a common mobilization process associated with acquisition of the bla VIM-2 and bla VIM-1 genes.

scholarly journals Characterization of a Type II-A CRISPR-Cas System in Streptococcus mutans

mSphere ◽  
2020 ◽  
Vol 5 (3) ◽  
Author(s):  
Cas Mosterd ◽  
Sylvain Moineau
Keyword(s):  
Streptococcus Mutans ◽  
Defense Mechanisms ◽  
Bacterial Species ◽  
Oral Microbiota ◽  
Type Ii ◽  
Content Type ◽  
C Terminus ◽  
Protospacer Adjacent Motif ◽  
Cas9 Protein

ABSTRACT Streptococcus mutans and its virulent phages are important members of the human oral microbiota. S. mutans is also the primary causal agent of dental caries. To survive in this ecological niche, S. mutans must encode phage defense mechanisms, which include CRISPR-Cas systems. Here, we describe the CRISPR-Cas type II-A system of S. mutans strain P42S, which was found to display natural adaptation and interference activity in response to phage infection and plasmid transformation. Newly acquired spacers were integrated both at the 5′ end of the CRISPR locus and ectopically. In comparisons of the cas genes of P42S to those of other strains of S. mutans, cas1, cas2, and csn2 appear to be highly conserved within the species. However, more diversity was observed with cas9. While the nuclease domains of S. mutans Cas9 (SmCas9) are conserved, the C terminus of the protein, including the protospacer adjacent motif (PAM) recognition domain, is less conserved. In support of these findings, we experimentally demonstrated that the PAMs associated with SmCas9 of strain P42S are NAA and NGAA. These PAMs are different from those previously reported for the CRISPR-Cas system of the model strain S. mutans UA159. This study illustrates the diversity of CRISPR-Cas type II-A systems that can be found within the same bacterial species. IMPORTANCE CRISPR-Cas is one of the mechanisms used by bacteria to defend against viral predation. Increasing our knowledge of the biology and diversity of CRISPR-Cas systems will also improve our understanding of virus-bacterium interactions. As CRISPR-Cas systems acquiring novel immunities under laboratory conditions are rare, Streptococcus mutans strain P42S provides an alternative model to study the adaptation step, which is still the least understood step in CRISPR-Cas biology. Furthermore, the availability of a natural Cas9 protein recognizing an AT-rich PAM opens up new avenues for genome editing purposes.

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