bluuebunny/crossref_metadata_2025_split huggingface dataset:
A Lactobacillus plantarum Esterase Active on a Broad Range of Phenolic Esters
Authors: Esteban-Torres María , Landete José María , Reverón Inés , Santamaría Laura , de las Rivas Blanca , Muñoz Rosario 
Published: March 2015

ABSTRACT

Lactobacillus plantarum
is the lactic acid bacterial species most frequently found in the fermentation of food products of plant origin on which phenolic compounds are abundant.
L. plantarum
strains showed great flexibility in their ability to adapt to different environments and growth substrates. Of 28
L. plantarum
strains analyzed, only cultures from 7 strains were able to hydrolyze hydroxycinnamic esters, such as methyl ferulate or methyl caffeate. As revealed by PCR, only these seven strains possessed the
est_1092
gene. When the
est_1092
gene was introduced into
L. plantarum
WCFS1 or
L. lactis
MG1363, their cultures acquired the ability to degrade hydroxycinnamic esters. These results support the suggestion that Est_1092 is the enzyme responsible for the degradation of hydroxycinnamic esters on the
L. plantarum
strains analyzed. The Est_1092 protein was recombinantly produced and biochemically characterized. Surprisingly, Est_1092 was able to hydrolyze not only hydroxycinnamic esters, since all the phenolic esters assayed were hydrolyzed. Quantitative PCR experiments revealed that the expression of
est_1092
was induced in the presence of methyl ferulate, an hydroxycinnamic ester, but was inhibited on methyl gallate, an hydroxybenzoic ester. As Est_1092 is an enzyme active on a broad range of phenolic esters, simultaneously possessing feruloyl esterase and tannase activities, its presence on some
L. plantarum
strains provides them with additional advantages to survive and grow on plant environments.
Two Phages, phiIPLA-RODI and phiIPLA-C1C, Lyse Mono- and Dual-Species Staphylococcal Biofilms
Authors: Gutiérrez Diana , Vandenheuvel Dieter , Martínez Beatriz , Rodríguez Ana , Lavigne Rob , García Pilar 
Published: March 2015

ABSTRACT

Phage therapy is a promising option for fighting against staphylococcal infections. Two lytic phages, vB_SauM_phiIPLA-RODI (phiIPLA-RODI) and vB_SepM_phiIPLA-C1C (phiIPLA-C1C), belonging to the
Myoviridae
family and exhibiting wide host ranges, were characterized in this study. The complete genome sequences comprised 142,348 bp and 140,961 bp and contained 213 and 203 open reading frames, respectively. The gene organization was typical of
Spounavirinae
members, with long direct terminal repeats (LTRs), genes grouped into modules not clearly separated from each other, and several group I introns. In addition, four genes encoding tRNAs were identified in phiIPLA-RODI. Comparative DNA sequence analysis showed high similarities with two phages, GH15 and 676Z, belonging to the
Twort-like virus
genus (nucleotide identities of >84%); for phiIPLA-C1C, a high similarity with phage phiIBB-SEP1 was observed (identity of 80%). Challenge assays of phages phiIPLA-RODI and phiIPLA-C1C against planktonic staphylococcal cells confirmed their lytic ability, as they were able to remove 5 log units in 8 h. Exposure of biofilms to phages phiIPLA-RODI and phiIPLA-C1C reduced the amount of adhered bacteria to about 2 log units in both monospecies and dual-species biofilms, but phiIPLA-RODI turned out to be as effective as the mixture of both phages. Moreover, the frequencies of bacteriophage-insensitive mutants (BIMs) of
Staphylococcus aureus
and
S. epidermidis
with resistance to phiIPLA-RODI and phiIPLA-C1C were low, at 4.05 × 10
−7
± 2.34 × 10
−9
and 1.1 × 10
−7
± 2.08 × 10
−9
, respectively. Overall, a generally reduced fitness in the absence of phages was observed for BIMs, which showed a restored phage-sensitive phenotype in a few generations. These results confirm that lytic bacteriophages can be efficient biofilm-disrupting agents, supporting their potential as antimicrobials against staphylococcal infections.
Mutational Analyses of Glucose Dehydrogenase and Glucose-6-Phosphate Dehydrogenase Genes in Pseudomonas fluorescens Reveal Their Effects on Growth and Alginate Production
Authors: Maleki Susan , Mærk Mali , Valla Svein , Ertesvåg Helga 
Published: March 2015

ABSTRACT

The biosynthesis of alginate has been studied extensively due to the importance of this polymer in medicine and industry. Alginate is synthesized from fructose-6-phosphate and thus competes with the central carbon metabolism for this metabolite. The alginate-producing bacterium
Pseudomonas fluorescens
relies on the Entner-Doudoroff and pentose phosphate pathways for glucose metabolism, and these pathways are also important for the metabolism of fructose and glycerol. In the present study, the impact of key carbohydrate metabolism enzymes on growth and alginate synthesis was investigated in
P. fluorescens
. Mutants defective in glucose-6-phosphate dehydrogenase isoenzymes (Zwf-1 and Zwf-2) or glucose dehydrogenase (Gcd) were evaluated using media containing glucose, fructose, or glycerol. Zwf-1 was shown to be the most important glucose-6-phosphate dehydrogenase for catabolism. Both Zwf enzymes preferred NADP as a coenzyme, although NAD was also accepted. Only Zwf-2 was active in the presence of 3 mM ATP, and then only with NADP as a coenzyme, indicating an anabolic role for this isoenzyme. Disruption of
zwf-1
resulted in increased alginate production when glycerol was used as the carbon source, possibly due to decreased flux through the Entner-Doudoroff pathway rendering more fructose-6-phosphate available for alginate biosynthesis. In alginate-producing cells grown on glucose, disruption of
gcd
increased both cell numbers and alginate production levels, while this mutation had no positive effect on growth in a non-alginate-producing strain. A possible explanation is that alginate synthesis might function as a sink for surplus hexose phosphates that could otherwise be detrimental to the cell.
New Shuttle Vector-Based Expression System To Generate Polyhistidine-Tagged Fusion Proteins in Staphylococcus aureus and Escherichia coli
Authors: Schwendener Sybille , Perreten Vincent 
Published: March 2015

ABSTRACT

Four
Staphylococcus aureus
-
Escherichia coli
shuttle vectors were constructed for gene expression and production of tagged fusion proteins. Vectors pBUS1-HC and pTSSCm have no promoter upstream of the multiple cloning site (MCS), and this allows study of genes under the control of their native promoters, and pBUS1-P

cap

-HC and pTSSCm-P

cap

contain the strong constitutive promoter of
S. aureus
type 1 capsule gene 1A (P

cap

) upstream of a novel MCS harboring codons for the peptide tag Arg-Gly-Ser-hexa-His (
rgs
-
his
6
). All plasmids contained the backbone derived from pBUS1, including the
E. coli
origin ColE1, five copies of terminator
rrnB
T
1
, and tetracycline resistance marker
tet
(L) for
S. aureus
and
E. coli
. The minimum pAMα1 replicon from pBUS1 was improved through either complementation with the single-strand origin
oriL
from pUB110 (pBUS1-HC and pBUS1-P

cap

-HC) or substitution with a pT181-family replicon (pTSSCm and pTSSCm-P

cap

). The new constructs displayed increased plasmid yield and segregational stability in
S. aureus
. Furthermore, pBUS1-P

cap

-HC and pTSSCm-P

cap

offer the potential to generate C-terminal RGS-His
6
translational fusions of cloned genes using simple molecular manipulation. BcgI-induced DNA excision followed by religation converts the TGA stop codon of the MCS into a TGC codon and links the
rgs
-
his
6
codons to the 3′ end of the target gene. The generation of the
rgs
-
his
6
codon-fusion, gene expression, and protein purification were demonstrated in both
S. aureus
and
E. coli
using the macrolide-lincosamide-streptogramin B resistance gene
erm
(44) inserted downstream of P

cap

. The new His tag expression system represents a helpful tool for the direct analysis of target gene function in staphylococcal cells.
Novel
bla
ROB-1
-Bearing Plasmid Conferring Resistance to β-Lactams in Haemophilus parasuis Isolates from Healthy Weaning Pigs
Authors: Moleres Javier , Santos-López Alfonso , Lázaro Isidro , Labairu Javier , Prat Cristina , Ardanuy Carmen , González-Zorn Bruno , Aragon Virginia , Garmendia Junkal 
Published: March 2015

ABSTRACT

Haemophilus parasuis
, the causative agent of Glässer's disease, is one of the early colonizers of the nasal mucosa of piglets. It is prevalent in swine herds, and lesions associated with disease are fibrinous polyserositis and bronchopneumonia. Antibiotics are commonly used in disease control, and resistance to several antibiotics has been described in
H. parasuis
. Prediction of
H. parasuis
virulence is currently limited by our scarce understanding of its pathogenicity. Some genes have been associated with
H. parasuis
virulence, such as
lsgB
and group 1
vtaA
, while biofilm growth has been associated with nonvirulent strains. In this study, 86
H. parasuis
nasal isolates from farms that had not had a case of disease for more than 10 years were obtained by sampling piglets at weaning. Isolates were studied by enterobacterial repetitive intergenic consensus PCR and determination of the presence of
lsgB
and group 1
vtaA
, biofilm formation, inflammatory cell response, and resistance to antibiotics. As part of the diversity encountered, a novel 2,661-bp plasmid, named pJMA-1, bearing the
bla
ROB-1
β-lactamase was detected in eight colonizing strains. pJMA-1 was shown to share a backbone with other small plasmids described in the
Pasteurellaceae
, to be 100% stable, and to have a lower biological cost than the previously described plasmid pB1000. pJMA-1 was also found in nine
H. parasuis
nasal strains from a separate collection, but it was not detected in isolates from the lesions of animals with Glässer's disease or in nontypeable
Haemophilus influenzae
isolates. Altogether, we show that commensal
H. parasuis
isolates represent a reservoir of β-lactam resistance genes which can be transferred to pathogens or other bacteria.
Novel Substrate Specificity and Temperature-Sensitive Activity of Mycosphaerella graminicola CYP51 Supported by the Native NADPH Cytochrome P450 Reductase
Authors: Price Claire L. , Warrilow Andrew G. S. , Parker Josie E. , Mullins Jonathan G. L. , Nes W. David , Kelly Diane E. , Kelly Steven L. 
Published: March 2015

ABSTRACT

Mycosphaerella graminicola
(
Zymoseptoria tritici
) is an ascomycete filamentous fungus that causes Septoria leaf blotch in wheat crops. In Europe the most widely used fungicides for this major disease are demethylation inhibitors (DMIs). Their target is the essential sterol 14α-demethylase (CYP51), which requires cytochrome P450 reductase (CPR) as its redox partner for functional activity. The
M. graminicola
CPR (MgCPR) is able to catalyze the sterol 14α-demethylation of eburicol and lanosterol when partnered with
Candida albicans
CYP51 (CaCYP51) and that of eburicol only with
M. graminicola
CYP51 (MgCYP51). The availability of the functional
in vivo
redox partner enabled the
in vitro
catalytic activity of MgCYP51 to be demonstrated for the first time. MgCYP51 50% inhibitory concentration (IC
50
) studies with epoxiconazole, tebuconazole, triadimenol, and prothioconazole-desthio confirmed that MgCYP51 bound these azole inhibitors tightly. The characterization of the MgCPR/MgCYP51 redox pairing has produced a functional method to evaluate the effects of agricultural azole fungicides, has demonstrated eburicol specificity in the activity observed, and supports the conclusion that prothioconazole is a profungicide.
Simple Method for Markerless Gene Deletion in Multidrug-Resistant Acinetobacter baumannii
Authors: Oh Man Hwan , Lee Je Chul , Kim Jungmin , Choi Chul Hee , Han Kyudong 
Published: March 2015

ABSTRACT

The traditional markerless gene deletion technique based on overlap extension PCR has been used for generating gene deletions in multidrug-resistant
Acinetobacter baumannii
. However, the method is time-consuming because it requires restriction digestion of the PCR products in DNA cloning and the construction of new vectors containing a suitable antibiotic resistance cassette for the selection of
A. baumannii
merodiploids. Moreover, the availability of restriction sites and the selection of recombinant bacteria harboring the desired chimeric plasmid are limited, making the construction of a chimeric plasmid more difficult. We describe a rapid and easy cloning method for markerless gene deletion in
A. baumannii
, which has no limitation in the availability of restriction sites and allows for easy selection of the clones carrying the desired chimeric plasmid. Notably, it is not necessary to construct new vectors in our method. This method utilizes direct cloning of blunt-end DNA fragments, in which upstream and downstream regions of the target gene are fused with an antibiotic resistance cassette via overlap extension PCR and are inserted into a blunt-end suicide vector developed for blunt-end cloning. Importantly, the antibiotic resistance cassette is placed outside the downstream region in order to enable easy selection of the recombinants carrying the desired plasmid, to eliminate the antibiotic resistance cassette via homologous recombination, and to avoid the necessity of constructing new vectors. This strategy was successfully applied to functional analysis of the genes associated with iron acquisition by
A. baumannii
ATCC 19606 and to
ompA
gene deletion in other
A. baumannii
strains. Consequently, the proposed method is invaluable for markerless gene deletion in multidrug-resistant
A. baumannii
.
Integrated Kinetic and Probabilistic Modeling of the Growth Potential of Bacterial Populations
Authors: George S. M. , Métris A. , Baranyi J. 
Published: March 2015

ABSTRACT
When bacteria are exposed to osmotic stress, some cells recover and grow, while others die or are unculturable. This leads to a viable count growth curve where the cell number decreases before the onset of the exponential growth phase. From such curves, it is impossible to estimate what proportion of the initial cells generates the growth because it leads to an ill-conditioned numerical problem. Here, we applied a combination of experimental and statistical methods, based on optical density measurements, to infer both the probability of growth and the maximum specific growth rate of the culture. We quantified the growth potential of a bacterial population as a quantity composed from the probability of growth and the “suitability” of the growing subpopulation to the new environment. We found that, for all three laboratory media studied, the probability of growth decreased while the “work to be done” by the growing subpopulation (defined as the negative logarithm of their suitability parameter) increased with NaCl concentration. The results suggest that the effect of medium on the probability of growth could be described by a simple shift parameter, a differential NaCl concentration that can be accounted for by the change in the medium composition. Finally, we highlighted the need for further understanding of the effect of the osmoprotectant glycine betaine on metabolism.
Accumulation of
d
-Glucose from Pentoses by Metabolically Engineered Escherichia coli
Authors: Xia Tian , Han Qi , Costanzo William V. , Zhu Yixuan , Urbauer Jeffrey L. , Eiteman Mark A. 
Published: March 2015

ABSTRACT

Escherichia coli
that is unable to metabolize
d
-glucose (with knockouts in
ptsG
,
manZ
, and
glk
) accumulates a small amount of
d
-glucose (yield of about 0.01 g/g) during growth on the pentoses
d
-xylose or
l
-arabinose as a sole carbon source. Additional knockouts in the
zwf
and
pfkA
genes, encoding, respectively,
d
-glucose-6-phosphate 1-dehydrogenase and 6-phosphofructokinase I (
E. coli
MEC143), increased accumulation to greater than 1 g/liter
d
-glucose and 100 mg/liter
d
-mannose from 5 g/liter
d
-xylose or
l
-arabinose. Knockouts of other genes associated with interconversions of
d
-glucose-phosphates demonstrate that
d
-glucose is formed primarily by the dephosphorylation of
d
-glucose-6-phosphate. Under controlled batch conditions with 20 g/liter
d
-xylose, MEC143 generated 4.4 g/liter
d
-glucose and 0.6 g/liter
d
-mannose. The results establish a direct link between pentoses and hexoses and provide a novel strategy to increase carbon backbone length from five to six carbons by directing flux through the pentose phosphate pathway.
Evolutionary Engineering Improves Tolerance for Replacement Jet Fuels in Saccharomyces cerevisiae
Authors: Brennan Timothy C. R. , Williams Thomas C. , Schulz Benjamin L. , Palfreyman Robin W. , Krömer Jens O. , Nielsen Lars K. 
Published: March 2015

ABSTRACT

Monoterpenes are liquid hydrocarbons with applications ranging from flavor and fragrance to replacement jet fuel. Their toxicity, however, presents a major challenge for microbial synthesis. Here we evolved limonene-tolerant
Saccharomyces cerevisiae
strains and sequenced six strains across the 200-generation evolutionary time course. Mutations were found in the tricalbin proteins Tcb2p and Tcb3p. Genomic reconstruction in the parent strain showed that truncation of a single protein (tTcb3p
1-989
), but not its complete deletion, was sufficient to recover the evolved phenotype improving limonene fitness 9-fold. tTcb3p
1-989
increased tolerance toward two other monoterpenes (β-pinene and myrcene) 11- and 8-fold, respectively, and tolerance toward the biojet fuel blend AMJ-700t (10% cymene, 50% limonene, 40% farnesene) 4-fold. tTcb3p
1-989
is the first example of successful engineering of phase tolerance and creates opportunities for production of the highly toxic C
10
alkenes in yeast.
Mortality Caused by Bath Exposure of Zebrafish (Danio rerio) Larvae to Nervous Necrosis Virus Is Limited to the Fourth Day Postfertilization
Authors: Morick Danny , Faigenbaum Or , Smirnov Margarita , Fellig Yakov , Inbal Adi , Kotler Moshe 
Published: March 2015

ABSTRACT

Nervous necrosis virus (NNV) is a member of the
Betanodavirus
genus that causes fatal diseases in over 40 species of fish worldwide. Mortality among NNV-infected fish larvae is almost 100%. In order to elucidate the mechanisms responsible for the susceptibility of fish larvae to NNV, we exposed zebrafish larvae to NNV by bath immersion at 2, 4, 6, and 8 days postfertilization (dpf). Here, we demonstrate that developing zebrafish embryos are resistant to NNV at 2 dpf due to the protection afforded by the egg chorion and, to a lesser extent, by the perivitelline fluid. The zebrafish larvae succumbed to NNV infection during a narrow time window around the 4th dpf, while 6- and 8-day-old larvae were much less sensitive, with mortalities of 24% and 28%, respectively.
Influence of Adhesion Force on
icaA
and
cidA
Gene Expression and Production of Matrix Components in Staphylococcus aureus Biofilms
Authors: Harapanahalli Akshay K. , Chen Yun , Li Jiuyi , Busscher Henk J. , van der Mei Henny C. 
Published: March 2015

ABSTRACT

The majority of human infections are caused by biofilms. The biofilm mode of growth enhances the pathogenicity of
Staphylococcus
spp. considerably, because once they adhere, staphylococci embed themselves in a protective, self-produced matrix of extracellular polymeric substances (EPSs). The aim of this study was to investigate the influence of forces of staphylococcal adhesion to different biomaterials on
icaA
(which regulates the production of EPS matrix components) and
cidA
(which is associated with cell lysis and extracellular DNA [eDNA] release) gene expression in
Staphylococcus aureus
biofilms. Experiments were performed with
S. aureus
ATCC 12600 and its isogenic mutant,
S. aureus
ATCC 12600 Δ
pbp4
, deficient in peptidoglycan cross-linking. Deletion of
pbp4
was associated with greater cell wall deformability, while it did not affect the planktonic growth rate, biofilm formation, cell surface hydrophobicity, or zeta potential of the strains. The adhesion forces of
S. aureus
ATCC 12600 were the strongest on polyethylene (4.9 ± 0.5 nN), intermediate on polymethylmethacrylate (3.1 ± 0.7 nN), and the weakest on stainless steel (1.3 ± 0.2 nN). The production of poly-
N
-acetylglucosamine, eDNA presence, and expression of
icaA
genes decreased with increasing adhesion forces. However, no relation between adhesion forces and
cidA
expression was observed. The adhesion forces of the isogenic mutant
S. aureus
ATCC 12600 Δ
pbp4
(deficient in peptidoglycan cross-linking) were much weaker than those of the parent strain and did not show any correlation with the production of poly-
N
-acetylglucosamine, eDNA presence, or expression of the
icaA
and
cidA
genes. This suggests that adhesion forces modulate the production of the matrix molecule poly-
N
-acetylglucosamine, eDNA presence, and
icaA
gene expression by inducing nanoscale cell wall deformation, with cross-linked peptidoglycan layers playing a pivotal role in this adhesion force sensing.
Neospora caninum Recruits Host Cell Structures to Its Parasitophorous Vacuole and Salvages Lipids from Organelles
Authors: Nolan Sabrina J. , Romano Julia D. , Luechtefeld Thomas , Coppens Isabelle 
Published: March 2015

ABSTRACT

Toxoplasma gondii
and
Neospora caninum
, which cause the diseases toxoplasmosis and neosporosis, respectively, are two closely related apicomplexan parasites. They have similar heteroxenous life cycles and conserved genomes and share many metabolic features. Despite these similarities,
T. gondii
and
N. caninum
differ in their transmission strategies and zoonotic potential. Comparative analyses of the two parasites are important to identify the unique biological features that underlie the basis of host preference and pathogenicity.
T. gondii
and
N. caninum
are obligate intravacuolar parasites; in contrast to
T. gondii
, events that occur during
N. caninum
infection remain largely uncharacterized. We examined the capability of
N. caninum
(Liverpool isolate) to interact with host organelles and scavenge nutrients in comparison to that of
T. gondii
(RH strain).
N. caninum
reorganizes the host microtubular cytoskeleton and attracts endoplasmic reticulum (ER), mitochondria, lysosomes, multivesicular bodies, and Golgi vesicles to its vacuole though with some notable differences from
T. gondii
. For example, the host ER gathers around the
N. caninum
parasitophorous vacuole (PV) but does not physically associate with the vacuolar membrane; the host Golgi apparatus surrounds the
N. caninum
PV but does not fragment into ministacks.
N. caninum
relies on plasma lipoproteins and scavenges cholesterol from NPC1-containing endocytic organelles. This parasite salvages sphingolipids from host Golgi Rab14 vesicles that it sequesters into its vacuole. Our data highlight a remarkable degree of conservation in the intracellular infection program of
N. caninum
and
T. gondii
. The minor differences between the two parasites related to the recruitment and rearrangement of host organelles around their vacuoles likely reflect divergent evolutionary paths.