bluuebunny/crossref_metadata_2025_split huggingface dataset:
Bacillus thuringiensis Metalloproteinase Bmp1 Functions as a Nematicidal Virulence Factor
Authors: Luo Xiaoxia , Chen Ling , Huang Qiong , Zheng Jinshui , Zhou Wei , Peng Donghai , Ruan Lifang , Sun Ming 
Published: November 2012

ABSTRACT

Some
Bacillus thuringiensis
strains have high toxicity to nematodes. Nematicidal activity has been found in several families of crystal proteins, such as Cry5, Cry6, and Cry55. The
B. thuringiensis
strain YBT-1518 has three
cry
genes that have high nematicidal activity. The whole genome sequence of this strain contains multiple potential virulence factors. To evaluate the pathogenic potential of virulence factors, we focused on a metalloproteinase called Bmp1. It encompasses a consecutive N-terminal signal peptide, an FTP superfamily domain, an M4 neutral protease GluZincin superfamily, two Big-3 superfamily motifs, and a Gram-positive anchor superfamily motif as a C-terminal domain. Here, we showed that purified Bmp1 protein showed metalloproteinase activity and toxicity against
Caenorhabditis elegans
(the 50% lethal concentration is 610 ± 9.37 μg/ml). In addition, mixing Cry5Ba with Bmp1 protein enhanced the toxicity 7.9-fold (the expected toxicity of the two proteins calculated from their separate toxicities) against
C. elegans
. Confocal microscopic observation revealed that Bmp1 protein was detected from around the mouth and esophagus to the intestine. Striking microscopic images revealed that Bmp1 degrades intestine tissues, and the Cry5Ba causes intestinal shrinkage from the body wall. Thus, the
B. thuringiensis
Bmp1 metalloproteinase is a nematicidal virulence factor. These findings give a new insight into the relationship between
B. thuringiensis
and its host nematodes.
Engineering Signal Peptides for Enhanced Protein Secretion from Lactococcus lactis
Authors: Ng Daphne T. W. , Sarkar Casim A. 
Published: November 2012

ABSTRACT

Lactococcus lactis
is an attractive vehicle for biotechnological production of proteins and clinical delivery of therapeutics. In many such applications using this host, it is desirable to maximize secretion of recombinant proteins into the extracellular space, which is typically achieved by using the native signal peptide from a major secreted lactococcal protein, Usp45. In order to further increase protein secretion from
L. lactis
, inherent limitations of the Usp45 signal peptide (Usp45sp) must be elucidated. Here, we performed extensive mutagenesis on Usp45sp to probe the effects of both the mRNA sequence (silent mutations) and the peptide sequence (amino acid substitutions) on secretion. We screened signal peptides based on their resulting secretion levels of
Staphylococcus aureus
nuclease and further evaluated them for secretion of
Bacillus subtilis
α-amylase. Silent mutations alone gave an increase of up to 16% in the secretion of α-amylase through a mechanism consistent with relaxed mRNA folding around the ribosome binding site and enhanced translation. Targeted amino acid mutagenesis in Usp45sp, combined with additional silent mutations from the best clone in the initial screen, yielded an increase of up to 51% in maximum secretion of α-amylase while maintaining secretion at lower induction levels. The best sequence from our screen preserves the tripartite structure of the native signal peptide but increases the positive charge of the n-region. Our study presents the first example of an engineered
L. lactis
signal peptide with a higher secretion yield than Usp45sp and, more generally, provides strategies for further enhancing protein secretion in bacterial hosts.
Molecular Characterization of Human-Pathogenic Microsporidia and Cyclospora cayetanensis Isolated from Various Water Sources in Spain: a Year-Long Longitudinal Study
Authors: Galván Ana Luz , Magnet Angela , Izquierdo Fernando , Fenoy Soledad , Rueda Cristina , Fernández Vadillo Carmen , Henriques-Gil Nuno , del Aguila Carmen 
Published: November 2012

ABSTRACT

Recent studies suggest the involvement of water in the epidemiology of
Cyclospora cayetanensis
and some microsporidia. A total of 223 samples from four drinking water treatment plants (DWTPs), seven wastewater treatment plants (WWTPs), and six locations of influence (LI) on four river basins from Madrid, Spain, were analyzed from spring 2008 to winter 2009. Microsporidia were detected in 49% of samples (109/223),
Cyclospora
spp. were detected in 9% (20/223), and both parasites were found in 5.4% (12/223) of samples. Human-pathogenic microsporidia were detected, including
Enterocytozoon bieneusi
(C, D, and D-like genotypes),
Encephalitozoon intestinalis
,
Encephalitozoon cuniculi
(genotypes I and III), and
Anncaliia algerae
.
C. cayetanensis
was identified in 17 of 20 samples. To our knowledge, this is the first study that shows a year-long longitudinal study of
C. cayetanensis
in drinking water treatment plants. Additionally, data about the presence and molecular characterization of the human-pathogenic microsporidia in drinking water, wastewater, and locations of influence during 1 year in Spain are shown. It is noteworthy that although the DWTPs and WWTPs studied meet European and national regulations on water sanitary quality, both parasites were found in water samples from these plants, supporting the idea that new and appropriate controls and regulations for drinking water, wastewater, and recreational waters should be proposed to avoid health risks from these pathogens.
Regulons of Three Pseudomonas syringae pv. tomato DC3000 Iron Starvation Sigma Factors
Authors: Markel Eric , Butcher Bronwyn G. , Myers Christopher R. , Stodghill Paul , Cartinhour Sam , Swingle Bryan 
Published: November 2012

ABSTRACT

Pseudomonas syringae
pv. tomato DC3000 contains genes for 15 sigma factors. The majority are members of the extracytoplasmic function class of sigma factors, including five that belong to the iron starvation subgroup. In this study, we identified the genes controlled by three iron starvation sigma factors. Their regulons are composed of a small number of genes likely to be involved in iron uptake.
Construction of a Broad-Host-Range Tn
7
-Based Vector for Single-Copy P
BAD
-Controlled Gene Expression in Gram-Negative Bacteria
Authors: Damron F. Heath , McKenney Elizabeth S. , Schweizer Herbert P. , Goldberg Joanna B. 
Published: November 2012

ABSTRACT

We describe a mini-Tn
7
-based broad-host-range expression cassette for arabinose-inducible gene expression from the P
BAD
promoter. This delivery vector, pTJ1, can integrate a single copy of a gene into the chromosome of Gram-negative bacteria for diverse genetic applications, of which several are discussed, using
Pseudomonas aeruginosa
as the model host.
Diet-Induced Alterations of Host Cholesterol Metabolism Are Likely To Affect the Gut Microbiota Composition in Hamsters
Authors: Martínez Inés , Perdicaro Diahann J. , Brown Andrew W. , Hammons Susan , Carden Trevor J. , Carr Timothy P. , Eskridge Kent M. , Walter Jens 
Published: November 2012

ABSTRACT

The gastrointestinal microbiota affects the metabolism of the mammalian host and has consequences for health. However, the complexity of gut microbial communities and host metabolic pathways make functional connections difficult to unravel, especially in terms of causation. In this study, we have characterized the fecal microbiota of hamsters whose cholesterol metabolism was extensively modulated by the dietary addition of plant sterol esters (PSE). PSE intake induced dramatic shifts in the fecal microbiota, reducing several bacterial taxa within the families
Coriobacteriaceae
and
Erysipelotrichaceae
. The abundance of these taxa displayed remarkably high correlations with host cholesterol metabolites. Most importantly, the associations between several bacterial taxa with fecal and biliary cholesterol excretion showed an almost perfect fit to a sigmoidal nonlinear model of bacterial inhibition, suggesting that host cholesterol excretion can shape microbiota structure through the antibacterial action of cholesterol.
In vitro
experiments suggested a modest antibacterial effect of cholesterol, and especially of cholesteryl-linoleate, but not plant sterols when included in model bile micelles. The findings obtained in this study are relevant to our understanding of gut microbiota-host lipid metabolism interactions, as they provide the first evidence for a role of cholesterol excreted with the bile as a relevant host factor that modulates the gut microbiota. The findings further suggest that the connections between
Coriobacteriaceae
and
Erysipelotrichaceae
and host lipid metabolism, which have been observed in several studies, could be caused by a metabolic phenotype of the host (cholesterol excretion) affecting the gut microbiota.
Surveying the Microbiome of Ants: Comparing 454 Pyrosequencing with Traditional Methods To Uncover Bacterial Diversity
Authors: Kautz Stefanie , Rubin Benjamin E. R. , Russell Jacob A. , Moreau Corrie S. 
Published: November 2012

ABSTRACT

We are only beginning to understand the depth and breadth of microbial associations across the eukaryotic tree of life. Reliably assessing bacterial diversity is a key challenge, and next-generation sequencing approaches are facilitating this endeavor. In this study, we used 16S rRNA amplicon pyrosequencing to survey microbial diversity in ants. We compared 454 libraries with Sanger-sequenced clone libraries as well as cultivation of live bacteria. Pyrosequencing yielded 95,656 bacterial 16S rRNA reads from 19 samples derived from four colonies of one ant species. The most dominant bacterial orders in the microbiome of the turtle ant
Cephalotes varians
were
Rhizobiales
,
Burkholderiales
,
Opitutales
,
Xanthomonadales
, and
Campylobacterales
, as revealed through both 454 sequencing and cloning. Even after stringent quality filtering, pyrosequencing recovered 445 microbe operational taxonomic units (OTUs) not detected with traditional techniques. In comparing bacterial communities associated with specific tissues, we found that gut tissues had significantly higher diversity than nongut tissues, and many of the OTUs identified from these groups clustered within ant-specific lineages, indicating a deep coevolutionary history of
Cephalotes
ants and their associated microbes. These lineages likely function as nutritional symbionts. One of four ant colonies investigated was infected with a
Spiroplasma
sp. (order
Entomoplasmatales
), a potential ant pathogen. Our work shows that the microbiome associated with
Cephalotes varians
is dominated by a few dozen bacterial lineages and that 454 sequencing is a cost-efficient tool to screen ant symbiont diversity.
Circadian Activation of the Mitogen-Activated Protein Kinase MAK-1 Facilitates Rhythms in Clock-Controlled Genes in Neurospora crassa
Authors: Bennett Lindsay D. , Beremand Phillip , Thomas Terry L. , Bell-Pedersen Deborah 
Published: November 2012

ABSTRACT

The circadian clock regulates the expression of many genes involved in a wide range of biological functions through output pathways such as mitogen-activated protein kinase (MAPK) pathways. We demonstrate here that the clock regulates the phosphorylation, and thus activation, of the MAPKs MAK-1 and MAK-2 in the filamentous fungus
Neurospora crassa
. In this study, we identified genetic targets of the MAK-1 pathway, which is homologous to the cell wall integrity pathway in
Saccharomyces cerevisiae
and the extracellular signal-regulated kinase 1/2 (ERK1/2) pathway in mammals. When MAK-1 was deleted from
Neurospora
cells, vegetative growth was reduced and the transcript levels for over 500 genes were affected, with significant enrichment for genes involved in protein synthesis, biogenesis of cellular components, metabolism, energy production, and transcription. Additionally, of the ∼500 genes affected by the disruption of MAK-1, more than 25% were previously identified as putative clock-controlled genes. We show that MAK-1 is necessary for robust rhythms of two morning-specific genes, i.e.,
ccg-1
and the mitochondrial phosphate carrier protein gene NCU07465. Additionally, we show clock regulation of a predicted chitin synthase gene, NCU04352, whose rhythmic accumulation is also dependent upon MAK-1. Together, these data establish a role for the MAK-1 pathway as an output pathway of the circadian clock and suggest a link between rhythmic MAK-1 activity and circadian control of cellular growth.
Normal Adaptation of Candida albicans to the Murine Gastrointestinal Tract Requires Efg1p-Dependent Regulation of Metabolic and Host Defense Genes
Authors: Pierce Jessica V. , Dignard Daniel , Whiteway Malcolm , Kumamoto Carol A. 
Published: November 2012

ABSTRACT

Although gastrointestinal colonization by the opportunistic fungal pathogen
Candida albicans
is generally benign, severe systemic infections are thought to arise due to escape of commensal
C. albicans
from the gastrointestinal (GI) tract. The
C. albicans
transcription factor Efg1p is a major regulator of GI colonization, hyphal morphogenesis, and virulence. The goals of this study were to identify the Efg1p regulon during GI tract colonization and to compare
C. albicans
gene expression during colonization of different organs of the GI tract. Our results identified significant differences in gene expression between cells colonizing the cecum and ileum. During colonization,
efg1

−

null mutant cells expressed higher levels of genes involved in lipid catabolism, carnitine biosynthesis, and carnitine utilization than did colonizing wild-type (WT) cells. In addition, during laboratory growth,
efg1

−

null mutant cells grew to a higher density than WT cells. The
efg1

−

null mutant grew in depleted medium, while WT cells could grow only if the depleted medium was supplemented with carnitine, a compound that promotes the metabolism of fatty acids. Altered gene expression and altered growth capability support the ability of
efg1
−
cells to hypercolonize naïve mice. Also, Efg1p was shown to be important for transcriptional responses to the stresses present in the cecum environment. For example, during colonization,
SOD5
, encoding a superoxide dismutase, was highly upregulated in an Efg1p-dependent manner. Ectopic expression of
SOD5
in an
efg1

−

null mutant increased the fitness of the
efg1

−

null mutant cells during colonization. These data show that
EFG1
is an important regulator of GI colonization.
Dual Functions of α-Ketoglutarate Dehydrogenase E2 in the Krebs Cycle and Mitochondrial DNA Inheritance in Trypanosoma brucei
Authors: Sykes Steven E. , Hajduk Stephen L. 
Published: November 2012

ABSTRACT
The dihydrolipoyl succinyltransferase (E2) of the multisubunit α-ketoglutarate dehydrogenase complex (α-KD) is an essential Krebs cycle enzyme commonly found in the matrices of mitochondria. African trypanosomes developmentally regulate mitochondrial carbohydrate metabolism and lack a functional Krebs cycle in the bloodstream of mammals. We found that despite the absence of a functional α-KD, bloodstream form (BF) trypanosomes express α-KDE2, which localized to the mitochondrial matrix and inner membrane. Furthermore, α-KDE2 fractionated with the mitochondrial genome, the kinetoplast DNA (kDNA), in a complex with the flagellum. A role for α-KDE2 in kDNA maintenance was revealed in α-KDE2 RNA interference (RNAi) knockdowns. Following RNAi induction, bloodstream trypanosomes showed pronounced growth reduction and often failed to equally distribute kDNA to daughter cells, resulting in accumulation of cells devoid of kDNA (dyskinetoplastic) or containing two kinetoplasts. Dyskinetoplastic trypanosomes lacked mitochondrial membrane potential and contained mitochondria of substantially reduced volume. These results indicate that α-KDE2 is bifunctional, both as a metabolic enzyme and as a mitochondrial inheritance factor necessary for the distribution of kDNA networks to daughter cells at cytokinesis.
Molecular and Morphological Data Support the Existence of a Sexual Cycle in Species of the Genus Paracoccidioides
Authors: Teixeira Marcus de Melo , Theodoro Raquel Cordeiro , Derengowski Lorena da Silveira , Nicola André Moraes , Bagagli Eduardo , Felipe Maria Sueli 
Published: November 2012

ABSTRACT

The genus
Paracoccidioides
includes the thermodimorphic species
Paracoccidioides brasiliensis
and
P. lutzii
, both of which are etiologic agents of paracoccidioidomycosis, a systemic mycosis that affects humans in Latin America. Despite the common occurrence of a sexual stage among closely related fungi, this has not been observed with
Paracoccidioides
species, which have thus been considered asexual. Molecular evolutionary studies revealed recombination events within isolated populations of the genus
Paracoccidioides
, suggesting the possible existence of a sexual cycle. Comparative genomic analysis of all dimorphic fungi and
Saccharomyces cerevisiae
demonstrated the presence of conserved genes involved in sexual reproduction, including those encoding mating regulators such as MAT, pheromone receptors, pheromone-processing enzymes, and mating signaling regulators. The expression of sex-related genes in the yeast and mycelial phases of both
Paracoccidioides
species was also detected by real-time PCR, with nearly all of these genes being expressed preferentially in the filamentous form of the pathogens. In addition, the expression of sex-related genes was responsive to the putative presence of pheromone in the supernatants obtained from previous cocultures of strains of two different mating types.
In vitro
crossing of isolates of different mating types, discriminated by phylogenetic analysis of the α-box (
MAT1-1
) and the high-mobility-group (HMG) domain (
MAT1-2
), led to the identification of the formation of young ascocarps with constricted coiled hyphae related to the initial stage of mating. These genomic and morphological analyses strongly support the existence of a sexual cycle in species of the genus
Paracoccidioides
.
Missense Mutations in Cytochrome
c
Maturation Genes Provide New Insights into Rhodobacter capsulatus cbb
3
-Type Cytochrome
c
Oxidase Biogenesis
Authors: Ekici Seda , Jiang Xinpei , Koch Hans-Georg , Daldal Fevzi 
Published: November 2012

ABSTRACT

The
Rhodobacter capsulatus cbb
3
-type cytochrome
c
oxidase (
cbb
3
-Cox) belongs to the heme-copper oxidase superfamily, and its subunits are encoded by the
ccoNOQP
operon. Biosynthesis of this enzyme is complex and needs dedicated biogenesis genes (
ccoGHIS
). It also relies on the
c
-type cytochrome maturation (Ccm) process, which requires the
ccmABCDEFGHI
genes, because two of the
cbb
3
-Cox subunits (CcoO and CcoP) are
c
-type cytochromes. Recently, we reported that mutants lacking CcoA, a major facilitator superfamily type transporter, produce very small amounts of
cbb
3
-Cox unless the growth medium is supplemented with copper. In this work, we isolated “Cu-unresponsive” derivatives of a
ccoA
deletion strain that exhibited no
cbb
3
-Cox activity even upon Cu supplementation. Molecular characterization of these mutants revealed missense mutations in the
ccmA
or
ccmF
gene, required for the Ccm process. As expected, Cu-unresponsive mutants lacked the CcoO and CcoP subunits due to Ccm defects, but remarkably, they contained the CcoN subunit of
cbb
3
-Cox. Subsequent construction and examination of single
ccm
knockout mutants demonstrated that membrane insertion and stability of CcoN occurred in the absence of the Ccm process. Moreover, while the
ccm
knockout mutants were completely incompetent for photosynthesis, the Cu-unresponsive mutants grew photosynthetically at lower rates and produced smaller amounts of cytochromes
c
1
and
c
2
than did a wild-type strain due to their restricted Ccm capabilities. These findings demonstrate that different levels of Ccm efficiency are required for the production of various
c
-type cytochromes and reveal for the first time that maturation of the heme-Cu-containing subunit CcoN of
R. capsulatus cbb
3
-Cox proceeds independently of that of the
c
-type cytochromes during the biogenesis of this enzyme.
Roles of AtpI and Two YidC-Type Proteins from Alkaliphilic Bacillus pseudofirmus OF4 in ATP Synthase Assembly and Nonfermentative Growth
Authors: Liu Jun , Hicks David B. , Krulwich Terry A. 
Published: November 2012

ABSTRACT

AtpI, a membrane protein encoded by many bacterial
atp
operons, is reported to be necessary for
c
-ring oligomer formation during assembly of some ATP synthase complexes. We investigated chaperone functions of AtpI and compared them to those of AtpZ, a protein encoded by a gene upstream of
atpI
that has a role in magnesium acquisition at near-neutral pH, and of SpoIIIJ and YqjG, two YidC/OxaI/Alb3 family proteins, in alkaliphilic
Bacillus pseudofirmus
OF4. A strain with a chromosomal deletion of
atpI
grew nonfermentatively, and its purified ATP synthase had a
c
-ring of normal size, indicating that AtpI is not absolutely required for ATP synthase function. However, deletion of
atpI
, but not
atpZ
, led to reduced stability of the ATP synthase rotor, reduced membrane association of the F
1
domain, reduced ATPase activity, and modestly reduced nonfermentative growth on malate at both pH 7.5 and 10.5. Both
spoIIIJ
and
yqjG
, but not
atpI
or
atpZ
, complemented a YidC-depleted
Escherichia coli
strain. Consistent with such overlapping functions, single deletions of
spoIIIJ
or
yqjG
in the alkaliphile did not affect membrane ATP synthase levels or activities, but functional specialization was indicated by YqjG and SpoIIIJ showing respectively greater roles in malate growth at pH 7.5 and 10.5. Expression of
yqjG
was elevated at pH 7.5 relative to that at pH 10.5 and in Δ
spoIIIJ
strains, but it was lower than constitutive
spoIIIJ
expression. Deletion of
atpZ
caused the largest increase among the mutants in magnesium concentrations needed for pH 7.5 growth. The basis for this phenotype is not yet resolved.
Role of Homologous Recombination in Adaptive Diversification of Extraintestinal Escherichia coli
Authors: Paul Sandip , Linardopoulou Elena V. , Billig Mariya , Tchesnokova Veronika , Price Lance B. , Johnson James R. , Chattopadhyay Sujay , Sokurenko Evgeni V. 
Published: November 2012

ABSTRACT

The contribution of homologous exchange (recombination) of core genes in the adaptive evolution of bacterial pathogens is not well understood. To investigate this, we analyzed fully assembled genomes of two
Escherichia coli
strains from sequence type 131 (ST131), a clonal group that is both the leading cause of extraintestinal
E. coli
infections and the main source of fluoroquinolone-resistant
E. coli
. Although the sequences of each of the seven multilocus sequence typing genes were identical in the two ST131 isolates, the strains diverged from one another by homologous recombination that affected at least 9% of core genes. This was on a par with the contribution to genomic diversity of horizontal gene transfer and point gene mutation. The genomic positions of recombinant and mobile genetic regions were partially linked, suggesting their concurrent occurrence. One of the genes affected by homologous recombination was
fimH
, which encodes mannose-specific type 1 fimbrial adhesin, resulting in functionally distinct copies of the gene in ST131 strains. One strain, a uropathogenic isolate, had a pathoadaptive variant of
fimH
that was acquired by homologous replacement into the commensal strain background. Close examination of FimH structure and function in additional ST131 isolates revealed that recombination led to acquisition of several functionally distinct variants that, upon homologous exchange, were targeted by a variety of pathoadaptive mutations under strong positive selection. Different recombinant
fimH
strains also showed a strong clonal association with ST131 isolates that had distinct fluoroquinolone resistance profiles. Thus, homologous recombination of core genes plays a significant role in adaptive diversification of bacterial pathogens, especially at the level of clonally related groups of isolates.