bluuebunny/crossref_metadata_2025_split huggingface dataset:
Biochemical Diversity of Carboxyl Esterases and Lipases from Lake Arreo (Spain): a Metagenomic Approach
Authors: Mart�nez-Mart�nez M�nica , Alcaide Mar�a , Tchigvintsev Anatoli , Reva Oleg , Polaina Julio , Bargiela Rafael , Guazzaroni Mar�a-Eugenia , Chicote �lvaro , Canet Albert , Valero Francisco , Rico Eguizabal Eugenio , Guerrero Mar�a del Carmen , Yakunin Alexander F. , Ferrer Manuel 
Published: March 2013

ABSTRACT
The esterases and lipases from the α/β hydrolase superfamily exhibit an enormous sequence diversity, fold plasticity, and activities. Here, we present the comprehensive sequence and biochemical analyses of seven distinct esterases and lipases from the metagenome of Lake Arreo, an evaporite karstic lake in Spain (42�46′N, 2�59′W; altitude, 655 m). Together with oligonucleotide usage patterns and BLASTP analysis, our study of esterases/lipases mined from Lake Arreo suggests that its sediment contains moderately halophilic and cold-adapted proteobacteria containing DNA fragments of distantly related plasmids or chromosomal genomic islands of plasmid and phage origins. This metagenome encodes esterases/lipases with broad substrate profiles (tested over a set of 101 structurally diverse esters) and habitat-specific characteristics, as they exhibit maximal activity at alkaline pH (8.0 to 8.5) and temperature of 16 to 40�C, and they are stimulated (1.5 to 2.2 times) by chloride ions (0.1 to 1.2 M), reflecting an adaptation to environmental conditions. Our work provides further insights into the potential significance of the Lake Arreo esterases/lipases for biotechnology processes (i.e., production of enantiomers and sugar esters), because these enzymes are salt tolerant and are active at low temperatures and against a broad range of substrates. As an example, the ability of a single protein to hydrolyze triacylglycerols, (non)halogenated alkyl and aryl esters, cinnamoyl and carbohydrate esters, lactones, and chiral epoxides to a similar extent was demonstrated.
Detection of Escherichia coli O104 in the Feces of Feedlot Cattle by a Multiplex PCR Assay Designed To Target Major Genetic Traits of the Virulent Hybrid Strain Responsible for the 2011 German Outbreak
Authors: Paddock Z. D. , Bai J. , Shi X. , Renter D. G. , Nagaraja T. G. 
Published: March 2013

ABSTRACT

A multiplex PCR was designed to detect
Escherichia coli
O104:H4, a hybrid pathotype of Shiga toxigenic and enteroaggregative
E. coli
, in cattle feces. A total of 248 fecal samples were tested, and 20.6% were positive for serogroup O104. The O104 isolates did not carry genes characteristic of the virulent hybrid strain.
Culture-Independent Analysis of Aerosol Microbiology in a Metropolitan Subway System
Authors: Robertson Charles E. , Baumgartner Laura K. , Harris J. Kirk , Peterson Kristen L. , Stevens Mark J. , Frank Daniel N. , Pace Norman R. 
Published: March 2013

ABSTRACT

The goal of this study was to determine the composition and diversity of microorganisms associated with bioaerosols in a heavily trafficked metropolitan subway environment. We collected bioaerosols by fluid impingement on several New York City subway platforms and associated sites in three sampling sessions over a 1.5-year period. The types and quantities of aerosolized microorganisms were determined by culture-independent phylogenetic analysis of small-subunit rRNA gene sequences by using both Sanger (universal) and pyrosequencing (bacterial) technologies. Overall, the subway bacterial composition was relatively simple; only 26 taxonomic families made up ∼75% of the sequences determined. The microbiology was more or less similar throughout the system and with time and was most similar to outdoor air, consistent with highly efficient air mixing in the system. Identifiable bacterial sequences indicated that the subway aerosol assemblage was composed of a mixture of genera and species characteristic of soil, environmental water, and human skin commensal bacteria. Eukaryotic diversity was mainly fungal, dominated by organisms of types associated with wood rot. Human skin bacterial species (at 99% rRNA sequence identity) included the
Staphylococcus
spp.
Staphylococcus epidermidis
(the most abundant and prevalent commensal of the human integument),
S. hominis
,
S. cohnii
,
S. caprae
, and
S. haemolyticus
, all well-documented human commensal bacteria. We encountered no organisms of public health concern. This study is the most extensive culture-independent survey of subway microbiota so far and puts in place pre-event information required for any bioterrorism surveillance activities or monitoring of the microbiological impact of recent subway flooding events.
Repetitive, Marker-Free, Site-Specific Integration as a Novel Tool for Multiple Chromosomal Integration of DNA
Authors: Petersen Kia Vest , Martinussen Jan , Jensen Peter Ruhdal , Solem Christian 
Published: March 2013

ABSTRACT

We present a tool for repetitive, marker-free, site-specific integration in
Lactococcus lactis
, in which a nonreplicating plasmid vector (pKV6) carrying a phage attachment site (
attP
) can be integrated into a bacterial attachment site (
attB
). The novelty of the tool described here is the inclusion of a minimal bacterial attachment site (

attB
min

), two mutated
loxP
sequences (
lox66
and
lox71
) allowing for removal of undesirable vector elements (antibiotic resistance marker), and a counterselection marker (
oroP
) for selection of
loxP
recombination on the pKV6 vector. When transformed into
L. lactis
expressing the phage TP901-1 integrase, pKV6 integrates with high frequency into the chromosome, where it is flanked by
attL
and
attR
hybrid attachment sites. After expression of Cre recombinase from a plasmid that is not able to replicate in
L. lactis
,
loxP
recombinants can be selected for by using 5-fluoroorotic acid. The introduced

attB
min

site can subsequently be used for a second round of integration. To examine if
attP
recombination was specific to the
attB
site, integration was performed in strains containing the
attB
,
attL
, and
attR
sites or the
attL
and
attR
sites only. Only
attP-attB
recombination was observed when all three sites were present. In the absence of the
attB
site, a low frequency of
attP-attL
recombination was observed. To demonstrate the functionality of the system, the xylose utilization genes (
xylABR
and
xylT
) from
L. lactis
strain KF147 were integrated into the chromosome of
L. lactis
strain MG1363 in two steps.
Enzymatic Synthesis of Novel Phloretin Glucosides
Authors: Pandey Ramesh Prasad , Li Tai Feng , Kim Eun-Hee , Yamaguchi Tokutaro , Park Yong Il , Kim Joong Su , Sohng Jae Kyung 
Published: March 2013

ABSTRACT

A UDP-glycosyltransferase from
Bacillus licheniformis
was exploited for the glycosylation of phloretin. The
in vitro
glycosylation reaction confirmed the production of five phloretin glucosides, including three novel glucosides. Consequently, we demonstrated the application of the same glycosyltransferase for the efficient whole-cell biocatalysis of phloretin in engineered
Escherichia coli
.
Prevalence and Genetic Characterization of Toxoplasma gondii in Bats in Myanmar
Authors: Sun Hongchao , Wang Yiyin , Zhang Yingguang , Ge Wei , Zhang Fuqiang , He Biao , Li Zuosheng , Fan Quanshui , Wang Wei , Tu Changchun , Li Jiping , Liu Quan 
Published: March 2013

ABSTRACT

We detected
Toxoplasma gondii
in 29.3% (95% confidence interval [CI], 25.5% to 33.1%) of 550 insectivorous bats collected in Myanmar. The genotyping of these positive samples revealed they were closely related to or belong to clonal type I, which is highly virulent in mice, showing that these bats are potential reservoirs for
T. gondii
transmission.
Effects of Bartonella spp. on Flea Feeding and Reproductive Performance
Authors: Morick Danny , Krasnov Boris R. , Khokhlova Irina S. , Gutiérrez Ricardo , Fielden Laura J. , Gottlieb Yuval , Harrus Shimon 
Published: March 2013

ABSTRACT

Numerous pathogens are transmitted from one host to another by hematophagous insect vectors. The interactions between a vector-borne organism and its vector vary in many ways, most of which are yet to be explored and identified. These interactions may play a role in the dynamics of the infection cycle. One way to evaluate these interactions is by studying the effects of the tested organism on the vector. In this study, we tested the effects of infection with
Bartonella
species on fitness-related variables of fleas by using
Bartonella
sp. strain OE 1-1,
Xenopsylla ramesis
fleas, and
Meriones crassus
jirds as a model system. Feeding parameters, including blood meal size and metabolic rate during digestion, as well as reproductive parameters, including fecundity, fertility, and life span, were compared between fleas experimentally infected with
Bartonella
and uninfected fleas. In addition, the developmental time, sex ratio, and body size of F
1
offspring fleas were compared between the two groups. Most tested parameters did not differ between infected and uninfected fleas. However, F
1
males produced by
Bartonella-
positive females were significantly smaller than F
1
males produced by
Bartonella
-negative female fleas. The findings in this study suggest that bartonellae are well adapted to their flea vectors, and by minimally affecting their fitness they have evolved to better spread themselves in the natural environment.
Effect of Soil Slope on the Appearance of Mycobacterium avium subsp. paratuberculosis in Water Running off Grassland Soil after Application of Contaminated Slurry
Authors: Salgado M. , Alfaro M. , Salazar F. , Troncoso E. , Mitchell R. M. , Ramirez L. , Naguil A. , Zamorano P. , Collins M. T. 
Published: March 2013

ABSTRACT

The study assessed the effect of soil slope on
Mycobacterium avium
subsp.
paratuberculosis
transport into rainwater runoff from agricultural soil after application of
M. avium
subsp.
paratuberculosis
-contaminated slurry. Under field conditions, 24 plots of undisturbed loamy soil 1 by 2 m
2
were placed on platforms. Twelve plots were used for water runoff: 6 plots at a 3% slope and 6 plots at a 15% slope. Half of the plots of each slope were treated with
M. avium
subsp.
paratuberculosis
-contaminated slurry, and half were not treated. Using the same experimental design, 12 plots were established for soil sampling on a monthly basis using the same spiked slurry application and soil slopes. Runoff following natural rainfall was collected and analyzed for
M. avium
subsp.
paratuberculosis
, coliforms, and turbidity.
M. avium
subsp.
paratuberculosis
was detected in runoff from all plots treated with contaminated slurry and one control plot. A higher slope (15%) increased the likelihood of
M. avium
subsp.
paratuberculosis
detection but did not affect the likelihood of finding coliforms. Daily rainfall increased the likelihood that runoff would have coliforms and the coliform concentration, but it decreased the
M. avium
subsp.
paratuberculosis
concentration in the runoff. When there was no runoff, rain was associated with increased
M. avium
subsp.
paratuberculosis
concentrations. Coliform counts in runoff were related to runoff turbidity.
M. avium
subsp.
paratuberculosis
presence/absence, however, was related to turbidity. Study duration decreased bacterial detection and concentration. These findings demonstrate the high likelihood that
M. avium
subsp.
paratuberculosis
in slurry spread on pastures will contaminate water runoff, particularly during seasons with high rainfall.
M. avium
subsp.
paratuberculosis
contamination of water has potential consequences for both animal and human health.
Subtle Differences in Virus Composition Affect Disinfection Kinetics and Mechanisms
Authors: Sigstam Thérèse , Gannon Greg , Cascella Michele , Pecson Brian M. , Wigginton Krista Rule , Kohn Tamar 
Published: March 2013

ABSTRACT

Viral disinfection kinetics have been studied in depth, but the molecular-level inactivation mechanisms are not understood. Consequently, it is difficult to predict the disinfection behavior of nonculturable viruses, even when related, culturable viruses are available. The objective of this work was to determine how small differences in the composition of the viral genome and proteins impact disinfection. To this end, we investigated the inactivation of three related bacteriophages (MS2, fr, and GA) by UV
254
, singlet oxygen (
1
O
2
), free chlorine (FC), and chlorine dioxide (ClO
2
). Genome damage was quantified by PCR, and protein damage was assessed by quantitative matrix-assisted laser desorption ionization (MALDI) mass spectrometry. ClO
2
caused great variability in the inactivation kinetics between viruses and was the only treatment that did not induce genome damage. The inactivation kinetics were similar for all viruses when treated with disinfectants possessing a genome-damaging component (FC,
1
O
2
, and UV
254
). On the protein level, UV
254
subtly damaged MS2 and fr capsid proteins, whereas GA's capsid remained intact.
1
O
2
oxidized a methionine residue in MS2 but did not affect the other two viruses. In contrast, FC and ClO
2
rapidly degraded the capsid proteins of all three viruses. Protein composition alone could not explain the observed degradation trends; instead, molecular dynamics simulations indicated that degradation is dictated by the solvent-accessible surface area of individual amino acids. Finally, despite the similarities of the three viruses investigated, their mode of inactivation by a single disinfectant varied. This explains why closely related viruses can exhibit drastically different inactivation kinetics.
Phenotype Overlap in Xylella fastidiosa Is Controlled by the Cyclic Di-GMP Phosphodiesterase Eal in Response to Antibiotic Exposure and Diffusible Signal Factor-Mediated Cell-Cell Signaling
Authors: de Souza Alessandra A. , Ionescu Michael , Baccari Clelia , da Silva Aline M. , Lindow Steven E. 
Published: March 2013

ABSTRACT

Eal is an EAL domain protein in
Xylella fastidiosa
homologous to one involved in resistance to tobramycin in
Pseudomonas aeruginosa
. EAL and HD-GYP domain proteins are implicated in the hydrolysis of the secondary messenger bis-(3′-5′)-cyclic dimeric GMP (cyclic di-GMP). Cell density-dependent communication mediated by a Diffusible Signal Factor (DSF) also modulates cyclic di-GMP levels in
X. fastidiosa
, thereby controlling the expression of virulence genes and genes involved in insect transmission. The possible linkage of Eal to both extrinsic factors such as antibiotics and intrinsic factors such as quorum sensing, and whether both affect virulence, was thus addressed. Expression of
eal
was induced by subinhibitory concentrations of tobramycin, and an
eal
deletion mutant was more susceptible to this antibiotic than the wild-type strain and exhibited phenotypes similar to those of an
rpfF
deletion mutant blocked in DSF production, such as hypermotility, reduced biofilm formation, and hypervirulence to grape. Consistent with that, the
rpfF
mutant was more susceptible than the wild-type strain to tobramycin. Therefore, we propose that cell-cell communication and antibiotic stress can apparently lead to similar modulations of cyclic di-GMP in
X. fastidiosa
, resulting in similar phenotypes. However, the effect of cell density is dominant compared to that of antibiotic stress, since
eal
is suppressed by RpfF, which may prevent inappropriate behavioral changes in response to antibiotic stress when DSF accumulates.
An Expanded Genetic Code in Candida albicans To Study Protein-Protein Interactions
In Vivo
Authors: Palzer Silke , Bantel Yannick , Kazenwadel Franziska , Berg Michael , Rupp Steffen , Sohn Kai 
Published: March 2013

ABSTRACT

For novel insights into the pathogenicity of
Candida albicans
, studies on molecular interactions of central virulence factors are crucial. Since methods for the analysis of direct molecular interactions of proteins
in vivo
are scarce, we expanded the genetic code of
C. albicans
with the synthetic photo-cross-linking amino acid
p
-azido-
l
-phenylalanine (AzF). Interacting molecules in close proximity of this unnatural amino acid can be covalently linked by UV-induced photo-cross-link, which makes unknown interacting molecules available for downstream identification. Therefore, we applied an aminoacyl-tRNA synthetase and a suppressor tRNA pair (
Ec
TyrtRNA
CUA
) derived from
Escherichia coli
, which was previously reported to be orthogonal in
Saccharomyces cerevisiae
. We further optimized the aminoacyl-tRNA synthetase for AzF (AzF-RS) and
Ec
TyrtRNA
CUA
for
C. albicans
and identified one AzF-RS with highest charging efficiency. Accordingly, incorporation of AzF into selected model proteins such as Tsa1p or Tup1p could be considerably enhanced. Immunologic detection of C-terminally tagged Tsa1p and Tup1p upon UV irradiation in a strain background containing suppressor tRNA and optimized AzF-RS revealed not only the mutant monomeric forms of these proteins but also higher-molecular-weight complexes, strictly depending on the specific position of incorporated AzF and UV excitation. By Western blotting and tandem mass spectrometry, we could identify these higher-molecular-weight complexes as homodimers consisting of one mutant monomer and a differently tagged, wild-type version of Tsa1p or Tup1p, respectively, demonstrating that expanding the genetic code of
C. albicans
with the unnatural photo-cross-linker amino acid AzF and applying it for
in vivo
binary protein interaction analyses is feasible.
Central Carbon Metabolism and Electron Transport in Chlamydomonas reinhardtii: Metabolic Constraints for Carbon Partitioning between Oil and Starch
Authors: Johnson Xenie , Alric Jean 
Published: March 2013

ABSTRACT

The metabolism of microalgae is so flexible that it is not an easy task to give a comprehensive description of the interplay between the various metabolic pathways. There are, however, constraints that govern central carbon metabolism in
Chlamydomonas reinhardtii
that are revealed by the compartmentalization and regulation of the pathways and their relation to key cellular processes such as cell motility, division, carbon uptake and partitioning, external and internal rhythms, and nutrient stress. Both photosynthetic and mitochondrial electron transfer provide energy for metabolic processes and how energy transfer impacts metabolism and vice versa is a means of exploring the regulation and function of these pathways. A key example is the specific chloroplast localization of glycolysis/gluconeogenesis and how it impacts the redox poise and ATP budget of the plastid in the dark. To compare starch and lipids as carbon reserves, their value can be calculated in terms of NAD(P)H and ATP. As microalgae are now considered a potential renewable feedstock, we examine current work on the subject and also explore the possibility of rerouting metabolism toward lipid production.
Posttranslational Modification of Flagellin FlaB in Shewanella oneidensis
Authors: Sun Linlin , Jin Miao , Ding Wen , Yuan Jie , Kelly John , Gao Haichun 
Published: March 2013

ABSTRACT

Shewanella oneidensis
is a highly motile organism by virtue of a polar, glycosylated flagellum composed of flagellins FlaA and FlaB. In this study, the functional flagellin FlaB was isolated and analyzed with nano-liquid chromatography-mass spectrometry (MS) and tandem MS. In combination with the mutational analysis, we propose that the FlaB flagellin protein from
S. oneidensis
is modified at five serine residues with a series of novel
O
-linked posttranslational modifications (PTMs) that differ from each other by 14 Da. These PTMs are composed in part of a 274-Da sugar residue that bears a resemblance to the nonulosonic acids. The remainder appears to be composed of a second residue whose mass varies by 14 Da depending on the PTM. Further investigation revealed that synthesis of the glycans initiates with PseB and PseC, the first two enzymes of the Pse pathway. In addition, a number of lysine residues are found to be methylated by SO4160, an analogue of the lysine methyltransferase of
Salmonella enterica
serovar
Typhimurium
.
The Rcs Stress Response and Accessory Envelope Proteins Are Required for
De Novo
Generation of Cell Shape in Escherichia coli
Authors: Ranjit Dev K. , Young Kevin D. 
Published: March 2013

ABSTRACT

Interactions with immune responses or exposure to certain antibiotics can remove the peptidoglycan wall of many Gram-negative bacteria. Though the spheroplasts thus created usually lyse, some may survive by resynthesizing their walls and shapes. Normally, bacterial morphology is generated by synthetic complexes directed by FtsZ and MreBCD or their homologues, but whether these classic systems can recreate morphology in the absence of a preexisting template is unknown. To address this question, we treated
Escherichia coli
with lysozyme to remove the peptidoglycan wall while leaving intact the inner and outer membranes and periplasm. The resulting lysozyme-induced (LI) spheroplasts recovered a rod shape after four to six generations. Recovery proceeded via a series of cell divisions that produced misshapen and branched intermediates before later progeny assumed a normal rod shape. Importantly, mutants defective in mounting the Rcs stress response and those lacking penicillin binding protein 1B (PBP1B) or LpoB could not divide or recover their cell shape but instead enlarged until they lysed. LI spheroplasts from mutants lacking the Lpp lipoprotein or PBP6 produced spherical daughter cells that did not recover a normal rod shape or that did so only after a significant delay. Thus, to regenerate normal morphology
de novo
,
E. coli
must supplement the classic FtsZ- and MreBCD-directed cell wall systems with activities that are otherwise dispensable for growth under normal laboratory conditions. The existence of these auxiliary mechanisms implies that they may be required for survival in natural environments, where bacterial walls can be damaged extensively or removed altogether.
The Clostridium difficile
cpr
Locus Is Regulated by a Noncontiguous Two-Component System in Response to Type A and B Lantibiotics
Authors: Suárez Jose M. , Edwards Adrianne N. , McBride Shonna M. 
Published: March 2013

ABSTRACT

The intestinal pathogen
Clostridium difficile
is known to grow only within the intestines of mammals, yet little is known about how the bacterium subsists in this environment. In the intestine,
C. difficile
must contend with innate defenses within the host, such as cationic antimicrobial peptides (CAMPs) produced by the host and the indigenous microbiota. In this study, we investigated the mechanism of activation and regulation of the CprABC transporter system, which provides resistance to multiple CAMPs and shows homology to the immunity systems of bacterial antimicrobial peptide producers. The CprABC system proved to be controlled by a noncontiguous two-component system consisting of the CprK sensor kinase and an orphan response regulator (CD3320; CprR). The CprK-CprR regulators were shown to activate
cprABCK
transcription in a manner similar to that by lantibiotic regulatory systems. Unlike lantibiotic producer regulation, regulation by CprK-CprR was activated by multiple lantibiotics produced by diverse Gram-positive bacteria. We identified a motif within these lantibiotics that is likely required for activation of
cpr
. Based on the similarities between the Cpr system and lantibiotic systems, we propose that the CprABC transporter and its regulators are relatives of lantibiotic systems that evolved to recognize multiple substrates to defend against toxins made by the intestinal microbiota.