bluuebunny/crossref_metadata_2025_split huggingface dataset:
Metatranscriptome of an Anaerobic Benzene-Degrading, Nitrate-Reducing Enrichment Culture Reveals Involvement of Carboxylation in Benzene Ring Activation
Authors: Luo Fei , Gitiafroz Roya , Devine Cheryl E. , Gong Yunchen , Hug Laura A. , Raskin Lutgarde , Edwards Elizabeth A. 
Published: May 2014

ABSTRACT

The enzymes involved in the initial steps of anaerobic benzene catabolism are not known. To try to elucidate this critical step, a metatranscriptomic analysis was conducted to compare the genes transcribed during the metabolism of benzene and benzoate by an anaerobic benzene-degrading, nitrate-reducing enrichment culture. RNA was extracted from the mixed culture and sequenced without prior mRNA enrichment, allowing simultaneous examination of the active community composition and the differential gene expression between the two treatments. Ribosomal and mRNA sequences attributed to a member of the family
Peptococcaceae
from the order
Clostridiales
were essentially only detected in the benzene-amended culture samples, implicating this group in the initial catabolism of benzene. Genes similar to each of two subunits of a proposed benzene-carboxylating enzyme were transcribed when the culture was amended with benzene. Anaerobic benzoate degradation genes from strict anaerobes were transcribed only when the culture was amended with benzene. Genes for other benzoate catabolic enzymes and for nitrate respiration were transcribed in both samples, with those attributed to an
Azoarcus
species being most abundant. These findings indicate that the mineralization of benzene starts with its activation by a strict anaerobe belonging to the
Peptococcaceae
, involving a carboxylation step to form benzoate. These data confirm the previously hypothesized syntrophic association between a benzene-degrading
Peptococcaceae
strain and a benzoate-degrading denitrifying
Azoarcus
strain for the complete catabolism of benzene with nitrate as the terminal electron acceptor.
Beyond the Matrix-Assisted Laser Desorption Ionization (MALDI) Biotyping Workflow: in Search of Microorganism-Specific Tryptic Peptides Enabling Discrimination of Subspecies
Authors: Gekenidis Maria-Theresia , Studer Patrick , Wüthrich Simone , Brunisholz René , Drissner David 
Published: May 2014

ABSTRACT

A well-accepted method for identification of microorganisms uses matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS) coupled to analysis software which identifies and classifies the organism according to its ribosomal protein spectral profile. The method, called MALDI biotyping, is widely used in clinical diagnostics and has partly replaced conventional microbiological techniques such as biochemical identification due to its shorter time to result (minutes for MALDI biotyping versus hours or days for classical phenotypic or genotypic identification). Besides its utility for identifying bacteria, MS-based identification has been shown to be applicable also to yeasts and molds. A limitation to this method, however, is that accurate identification is most reliably achieved on the species level on the basis of reference mass spectra, making further phylogenetic classification unreliable. Here, it is shown that combining tryptic digestion of the acid/organic solvent extracted (classical biotyping preparation) and resolubilized proteins, nano-liquid chromatography (nano-LC), and subsequent identification of the peptides by MALDI-tandem TOF (MALDI-TOF/TOF) mass spectrometry increases the discrimination power to the level of subspecies. As a proof of concept, using this targeted proteomics workflow, we have identified subspecies-specific biomarker peptides for three
Salmonella
subspecies, resulting in an extension of the mass range and type of proteins investigated compared to classical MALDI biotyping. This method therefore offers rapid and cost-effective identification and classification of microorganisms at a deeper taxonomic level.
Comparison of Five Bacteriophages as Models for Viral Aerosol Studies
Authors: Turgeon Nathalie , Toulouse Marie-Josée , Martel Bruno , Moineau Sylvain , Duchaine Caroline 
Published: May 2014

ABSTRACT

Bacteriophages are perceived to be good models for the study of airborne viruses because they are safe to use, some of them display structural features similar to those of human and animal viruses, and they are relatively easy to produce in large quantities. Yet, only a few studies have investigated them as models. It has previously been demonstrated that aerosolization, environmental conditions, and sampling conditions affect viral infectivity, but viral infectivity is virus dependent. Thus, several virus models are likely needed to study their general behavior in aerosols. The aim of this study was to compare the effects of aerosolization and sampling on the infectivity of five tail-less bacteriophages and two pathogenic viruses: MS2 (a single-stranded RNA [ssRNA] phage of the
Leviviridae
family), Φ6 (a segmented double-stranded RNA [dsRNA] phage of the
Cystoviridae
family), ΦX174 (a single-stranded DNA [ssDNA] phage of the
Microviridae
family), PM2 (a double-stranded DNA [dsDNA] phage of the
Corticoviridae
family), PR772 (a dsDNA phage of the
Tectiviridae
family), human influenza A virus H1N1 (an ssRNA virus of the
Orthomyxoviridae
family), and the poultry virus Newcastle disease virus (NDV; an ssRNA virus of the
Paramyxoviridae
family). Three nebulizers and two nebulization salt buffers (with or without organic fluid) were tested, as were two aerosol sampling devices, a liquid cyclone (SKC BioSampler) and a dry cyclone (National Institute for Occupational Safety and Health two-stage cyclone bioaerosol sampler). The presence of viruses in collected air samples was detected by culture and quantitative PCR (qPCR). Our results showed that these selected five phages behave differently when aerosolized and sampled. RNA phage MS2 and ssDNA phage ΦX174 were the most resistant to aerosolization and sampling. The presence of organic fluid in the nebulization buffer protected phages PR772 and Φ6 throughout the aerosolization and sampling with dry cyclones. In this experimental setup, the behavior of the influenza virus resembled that of phages PR772 and Φ6, while the behavior of NDV was closer to that of phages MS2 and ΦX174. These results provide critical information for the selection of appropriate phage models to mimic the behavior of specific human and animal viruses in aerosols.
Use of Cellulolytic Marine Bacteria for Enzymatic Pretreatment in Microalgal Biogas Production
Authors: Muñoz Camilo , Hidalgo Catalina , Zapata Manuel , Jeison David , Riquelme Carlos , Rivas Mariella 
Published: May 2014

ABSTRACT

In this study, we designed and evaluated a microalgal pretreatment method using cellulolytic bacteria that naturally degrades microalgae in their native habitat. Bacterial strains were isolated from each of two mollusk species in a medium containing 1% carboxymethyl cellulose agar. We selected nine bacterial strains that had endoglucanase activity: five strains from
Mytilus chilensis
, a Chilean mussel, and four strains from
Mesodesma donacium
, a clam found in the Southern Pacific. These strains were identified phylogenetically as belonging to the genera
Aeromonas
,
Pseudomonas
,
Chryseobacterium
, and
Raoultella
. The cellulase-producing capacities of these strains were characterized, and the degradation of cell walls in
Botryococcus braunii
and
Nannochloropsis gaditana
was tested with “whole-cell” cellulolytic experiments.
Aeromonas bivalvium
MA2,
Raoultella ornithinolytica
MA5, and
Aeromonas salmonicida
MC25 degraded
B. braunii
, and
R. ornithinolytica
MC3 and MA5 degraded
N. gaditana
. In addition,
N. gaditana
was pretreated with
R. ornithinolytica
strains MC3 and MA5 and was then subjected to an anaerobic digestion process, which increased the yield of methane by 140.32% and 158.68%, respectively, over that from nonpretreated microalgae. Therefore, a “whole-cell” cellulolytic pretreatment can increase the performance and efficiency of biogas production.
Prevalence, Genetic Diversity, and Host Range of Tectiviruses among Members of the Bacillus cereus Group
Authors: Gillis Annika , Mahillon Jacques 
Published: May 2014

ABSTRACT

GIL01, Bam35, GIL16, AP50, and Wip1 are tectiviruses preying on the
Bacillus cereus
group. Despite the significant contributions of phages in different biological processes, little is known about the dealings taking place between tectiviruses and their Gram-positive bacterial hosts. Therefore, this work focuses on characterizing the interactions between tectiviruses and the
B. cereus
group by assessing their occurrence and genetic diversity and evaluating their host range. To study the occurrence of tectiviruses in the
B. cereus
group, 2,000 isolates were evaluated using primers designed to be specific to two variable regions detected in previously described elements. PCR and propagation tests revealed that tectivirus-like elements occurred in less than 3% of the isolates. Regardless of this limited distribution, several novel tectiviruses were found, and partial DNA sequencing indicated that a greater diversity exists within the family
Tectiviridae
. Analyses of the selected variable regions, along with their host range, showed that tectiviruses in the
B. cereus
group can be clustered mainly into two different groups: the ones infecting
B. anthracis
and those isolated from other
B. cereus
group members. In order to address the host range of some novel tectiviruses, 120 strains were tested for sensitivity. The results showed that all the tested tectiviruses produced lysis in at least one
B. cereus sensu lato
strain. Moreover, no simple relationship between the infection patterns of the tectiviruses and their diversity was found.
Development of a Functional Antibody by Using a Green Fluorescent Protein Frame as the Template
Authors: Wang Rongzhi , Xiang Shuangshuang , Zhang Yonghui , Chen Qiuyu , Zhong Yanfang , Wang Shihua 
Published: May 2014

ABSTRACT
Single-chain variable fragment (scFv) antibodies are widely used as diagnostic and therapeutic agents or biosensors for a majority of human disease. However, the limitations of the present scFv antibody in terms of stability, solubility, and affinity are challenging to produce by traditional antibody screening and expression formats. We describe here a feasible strategy for creating the green fluorescent protein (GFP)-based antibody. Complementarity-determining region 3 (CDR3), which retains the antigen binding activity, was introduced into the structural loops of superfolder GFP, and the result showed that CDR3-inserted GFP displayed almost the same fluorescence intensity as wild-type GFP, and the purified proteins of CDR3 insertion showed the similar binding activity to antigen as the corresponding scFv. Among of all of the CDRs, CDR3s are responsible for antigen recognition, and only the CDR3a insertion is the best format for producing GFP-based antibody binding to specific antigen. The wide versatility of this system was further verified by introducing CDR3 from other scFvs into loop 9 of GFP. We developed a feasible method for rapidly and effectively producing a high-affinity GFP-based antibody by inserting CDR3s into GFP loops. Further, the affinity can be enhanced by specific amino acids scanning and site-directed mutagenesis. Notably, this method had better versatility for creating antibodies to various antigens using GFP as the scaffold, suggesting that a GFP-based antibody with high affinity and specificity may be useful for disease diagnosis and therapy.
Eutrophication and Bacterial Pathogens as Risk Factors for Avian Botulism Outbreaks in Wetlands Receiving Effluents from Urban Wastewater Treatment Plants
Authors: Anza Ibone , Vidal Dolors , Laguna Celia , Díaz-Sánchez Sandra , Sánchez Sergio , Chicote Álvaro , Florín Máximo , Mateo Rafael 
Published: May 2014

ABSTRACT

Due to the scarcity of water resources in the “Mancha Húmeda” Biosphere Reserve, the use of treated wastewater has been proposed as a solution for the conservation of natural threatened floodplain wetlands. In addition, wastewater treatment plants of many villages pour their effluent into nearby natural lakes. We hypothesized that certain avian pathogens present in wastewater may cause avian mortalities which would trigger avian botulism outbreaks. With the aim of testing our hypothesis, 24 locations distributed in three wetlands, two that receive wastewater effluents and one serving as a control, were monitored during a year. Sediment, water, water bird feces, and invertebrates were collected for the detection of putative avian pathogenic
Escherichia coli
(APEC),
Salmonella
spp.,
Clostridium perfringens
type A, and
Clostridium botulinum
type C/D. Also, water and sediment physicochemical properties were determined. Overall, APEC,
C. perfringens
, and
C. botulinum
were significantly more prevalent in samples belonging to the wetlands which receive wastewater. The occurrence of a botulism outbreak in one of the studied wetlands coincided with high water temperatures and sediment 5-day biochemical oxygen demand (BOD
5
), a decrease in water redox potential, chlorophyll
a
, and sulfate levels, and an increase in water inorganic carbon levels. The presence of
C. botulinum
in bird feces before the onset of the outbreak indicates that carrier birds exist and highlights the risk of botulinum toxin production in their carcasses if they die by other causes such as bacterial diseases, which are more probable in wastewater wetlands.
Pyruvate-Associated Acid Resistance in Bacteria
Authors: Wu Jianting , Li Yannan , Cai Zhiming , Jin Ye 
Published: May 2014

ABSTRACT
Glucose confers acid resistance on exponentially growing bacteria by repressing formation of the cyclic AMP (cAMP)-cAMP receptor protein (CRP) complex and consequently activating acid resistance genes. Therefore, in a glucose-rich growth environment, bacteria are capable of resisting acidic stresses due to low levels of cAMP-CRP. Here we reveal a second mechanism for glucose-conferred acid resistance. We show that glucose induces acid resistance in exponentially growing bacteria through pyruvate, the glycolysis product. Pyruvate and/or the downstream metabolites induce expression of the small noncoding RNA (sncRNA) Spot42, and the sncRNA, in turn, activates expression of the master regulator of acid resistance, RpoS. In contrast to glucose, pyruvate has little effect on levels of the cAMP-CRP complex and does not require the complex for its effects on acid resistance. Another important difference between glucose and pyruvate is that pyruvate can be produced by bacteria. This means that bacteria have the potential to protect themselves from acidic stresses by controlling glucose-derived generation of pyruvate, pyruvate-acetate efflux, or reversion from acetate to pyruvate. We tested this possibility by shutting down pyruvate-acetate efflux and found that the resulting accumulation of pyruvate elevated acid resistance. Many sugars can be broken into glucose, and the subsequent glycolysis generates pyruvate. Therefore, pyruvate-associated acid resistance is not confined to glucose-grown bacteria but is functional in bacteria grown on various sugars.
Comparative Biochemical Characterization of Three Exolytic Oligoalginate Lyases from Vibrio splendidus Reveals Complementary Substrate Scope, Temperature, and pH Adaptations
Authors: Jagtap Sujit Sadashiv , Hehemann Jan-Hendrik , Polz Martin F. , Lee Jung-Kul , Zhao Huimin 
Published: May 2014

ABSTRACT

Marine microbes use alginate lyases to degrade and catabolize alginate, a major cell wall matrix polysaccharide of brown seaweeds. Microbes frequently contain multiple, apparently redundant alginate lyases, raising the question of whether these enzymes have complementary functions. We report here on the molecular cloning and functional characterization of three exo-type oligoalginate lyases (OalA, OalB, and OalC) from
Vibrio splendidus
12B01 (12B01), a marine bacterioplankton species. OalA was most active at 16°C, had a pH optimum of 6.5, and displayed activities toward poly-β-
d
-mannuronate [poly(M)] and poly-α-
l
-guluronate [poly(G)], indicating that it is a bifunctional enzyme. OalB and OalC were most active at 30 and 35°C, had pH optima of 7.0 and 7.5, and degraded poly(M·G) and poly(M), respectively. Detailed kinetic analyses of oligoalginate lyases with poly(G), poly(M), and poly(M·G) and sodium alginate as substrates demonstrated that OalA and OalC preferred poly(M), whereas OalB preferred poly(M·G). The catalytic efficiency (
k
cat
/

K
m

) of OalA against poly(M) increased with decreasing size of the substrate. OalA showed
k
cat
/

K
m

from 2,130 mg
−1
ml s
−1
for the trisaccharide to 224 mg
−1
ml s
−1
for larger oligomers of ∼50 residues, and 50.5 mg
−1
ml s
−1
for high-molecular-weight alginate. Although OalA was most active on the trisaccharide, OalB and OalC preferred dimers. Taken together, our results indicate that these three Oals have complementary substrate scopes and temperature and pH adaptations.
Coordinated and Distinct Functions of Velvet Proteins in Fusarium verticillioides
Authors: Lan Nan , Zhang Hanxing , Hu Chengcheng , Wang Wenzhao , Calvo Ana M. , Harris Steven D. , Chen She , Li Shaojie 
Published: May 2014

ABSTRACT

Velvet-domain-containing proteins are broadly distributed within the fungal kingdom. In the corn pathogen
Fusarium verticillioides
, previous studies showed that the velvet protein
F. verticillioides
VE1 (FvVE1) is critical for morphological development, colony hydrophobicity, toxin production, and pathogenicity. In this study, tandem affinity purification of FvVE1 revealed that FvVE1 can form a complex with the velvet proteins
F. verticillioides
VelB (FvVelB) and FvVelC. Phenotypic characterization of gene knockout mutants showed that, as in the case of FvVE1, FvVelB regulated conidial size, hyphal hydrophobicity, fumonisin production, and oxidant resistance, while FvVelC was dispensable for these biological processes. Comparative transcriptional analysis of eight genes involved in the ROS (reactive oxygen species) removal system revealed that both FvVE1 and FvVelB positively regulated the transcription of a catalase-encoding gene,
F. verticillioides
CAT2
(
FvCAT2
). Deletion of
FvCAT2
resulted in reduced oxidant resistance, providing further explanation of the regulation of oxidant resistance by velvet proteins in the fungal kingdom.
Metal-Mediated Modulation of Streptococcal Cysteine Protease Activity and Its Biological Implications
Authors: Chella Krishnan Karthickeyan , Mukundan Santhosh , Landero Figueroa Julio A. , Caruso Joseph A. , Kotb Malak 
Published: May 2014

ABSTRACT

Streptococcal cysteine protease (SpeB), the major secreted protease produced by group A streptococcus (GAS), cleaves both host and bacterial proteins and contributes importantly to the pathogenesis of invasive GAS infections. Modulation of SpeB expression and/or its activity during invasive GAS infections has been shown to affect bacterial virulence and infection severity. Expression of SpeB is regulated by the GAS CovR-CovS two-component regulatory system, and we demonstrated that bacteria with mutations in the CovR-CovS two-component regulatory system are selected for during localized GAS infections and that these bacteria lack SpeB expression and exhibit a hypervirulent phenotype. Additionally, in a separate study, we showed that expression of SpeB can also be modulated by human transferrin- and/or lactoferrin-mediated iron chelation. Accordingly, the goal of this study was to investigate the possible roles of iron and other metals in modulating SpeB expression and/or activity in a manner that would potentiate bacterial virulence. Here, we report that the divalent metals zinc and copper inhibit SpeB activity at the posttranslational level. Utilizing online metal-binding site prediction servers, we identified two putative metal-binding sites in SpeB, one of which involves the catalytic-dyad residues
47
Cys and
195
His. Based on our findings, we propose that zinc and/or copper availability in the bacterial microenvironment can modulate the proteolytic activity of SpeB in a manner that preserves the integrity of several other virulence factors essential for bacterial survival and dissemination within the host and thereby may exacerbate the severity of invasive GAS infections.
Aggregation Interplay between Variants of the RepA-WH1 Prionoid in Escherichia coli
Authors: Molina-García Laura , Giraldo Rafael 
Published: May 2014

ABSTRACT

The N-terminal domain (winged-helix domain, or WH1) of the
Pseudomonas
pPS10 plasmid DNA replication protein RepA can assemble into amyloid fibers
in vitro
and, when expressed in
Escherichia coli
, leads to a unique intracellular amyloid proteinopathy by hampering bacterial proliferation. RepA-WH1 amyloidosis propagates along generations through the transmission of aggregated particles across the progeny, but it is unable to propagate horizontally as an infectious agent and is thus the first synthetic bacterial prionoid. RepA-WH1 amyloidosis is promoted by binding to double-stranded DNA (dsDNA)
in vitro
, and it is modulated by the Hsp70 chaperone DnaK
in vivo
. Different mutations in the
repA
-WH1 gene result in variants of the protein with distinct amyloidogenic properties. Here, we report that intracellular aggregates of the hyperamyloidogenic RepA with an A31V change in WH1 [RepA-WH1(A31V)] are able to induce and enhance the growth
in vivo
of new amyloid particles from molecules of wild-type RepA-WH1 [RepA-WH1(WT)], which otherwise would remain soluble in the cytoplasm. In contrast, RepA-WH1(ΔN37), a variant lacking a clear amyloidogenic sequence stretch that aggregates as conventional inclusion bodies (IBs), can drive the aggregation of the soluble protein into IBs only if expressed at high molar ratios over RepA-WH1(WT). The cytotoxic bacterial intracellular prionoid RepA-WH1 thus exhibits a hallmark feature of amyloids, as characterized in eukaryotes: cross-aggregation between variants of the same protein.
Effects of (p)ppGpp on the Progression of the Cell Cycle of Caulobacter crescentus
Authors: Gonzalez Diego , Collier Justine 
Published: May 2014

ABSTRACT

Bacteria must control the progression of their cell cycle in response to nutrient availability. This regulation can be mediated by guanosine tetra- or pentaphosphate [(p)ppGpp], which are synthesized by enzymes of the RelA/SpoT homologue (Rsh) family, particularly under starvation conditions. Here, we study the effects of (p)ppGpp on the cell cycle of
Caulobacter crescentus
, an oligotrophic bacterium with a dimorphic life cycle.
C. crescentus
divides asymmetrically, producing a motile swarmer cell that cannot replicate its chromosome and a sessile stalked cell that is replication competent. The swarmer cell rapidly differentiates into a stalked cell in appropriate conditions. An artificial increase in the levels of (p)ppGpp in nonstarved
C. crescentus
cells was achieved by expressing a truncated
relA
gene from
Escherichia coli
, encoding a constitutively active (p)ppGpp synthetase. By combining single-cell microscopy, flow cytometry approaches, and swarming assays, we show that an increase in the intracellular concentration of (p)ppGpp is sufficient to slow down the swarmer-to-stalked cell differentiation process and to delay the initiation of chromosome replication. We also present evidence that the intracellular levels of two master regulators of the cell cycle of
C. crescentus
, DnaA and CtrA, are modulated in response to (p)ppGpp accumulation, even in the absence of actual starvation. CtrA proteolysis and DnaA synthesis seem indirectly inhibited by (p)ppGpp accumulation. By extending the life span of the motile nonreproductive swarmer cell and thus promoting dispersal and foraging functions over multiplication under starvation conditions, (p)ppGpp may play a central role in the ecological adaptation of
C. crescentus
to nutritional stresses.