bluuebunny/crossref_metadata_2025 huggingface dataset:
Food as a Source for Quorum Sensing Inhibitors: Iberin from Horseradish Revealed as a Quorum Sensing Inhibitor of Pseudomonas aeruginosa
Authors: Jakobsen Tim Holm , Bragason Steinn Kristinn , Phipps Richard Kerry , Christensen Louise Dahl , van Gennip Maria , Alhede Morten , Skindersoe Mette , Larsen Thomas Ostenfeld , Høiby Niels , Bjarnsholt Thomas , Givskov Michael
Published: January 2012

Foods with health-promoting effects beyond nutritional values have been gaining increasing research focus in recent years, although not much has been published on this subject in relation to bacterial infections. With respect to treatment, a novel antimicrobial strategy, which is expected to transcend problems with selective pressures for antibiotic resistance, is to interrupt bacterial communication, also known as quorum sensing (QS), by means of signal antagonists, the so-called QS inhibitors (QSIs). Furthermore, QSI agents offer a potential solution to the deficiencies associated with use of traditional antibiotics to treat infections caused by bacterial biofilms and multidrug-resistant bacteria. Several QSIs of natural origin have been identified, and in this study, several common food products and plants were extracted and screened for QSI activity in an attempt to isolate and characterize previously unknown QSI compounds active against the common opportunistic pathogen
Pseudomonas aeruginosa
. Several extracts displayed activity, but horseradish exhibited the highest activity. Chromatographic separation led to the isolation of a potent QSI compound that was identified by liquid chromatography-diode array detector-mass spectrometry (LC-DAD-MS) and nuclear magnetic resonance (NMR) spectroscopy as iberin—an isothiocyanate produced by many members of the
Brassicaceae
family. Real-time PCR (RT-PCR) and DNA microarray studies showed that iberin specifically blocks expression of QS-regulated genes in
P. aeruginosa
.
Phylogeny and Population Structure of Brown Rot- and Moko Disease-Causing Strains of Ralstonia solanacearum Phylotype II
Authors: Cellier G. , Remenant B. , Chiroleu F. , Lefeuvre P. , Prior P.
Published: January 2012

The ancient soilborne plant vascular pathogenRalstonia solanacearumhas evolved and adapted to cause severe damage in an unusually wide range of plants. In order to better describe and understand these adaptations, strains with very similar lifestyles and host specializations are grouped into ecotypes. We used comparative genomic hybridization (CGH) to investigate three particular ecotypes in the American phylotype II group: (i) brown rot strains from phylotypes IIB-1 and IIB-2, historically known as race 3 biovar 2 and clonal; (ii) new pathogenic variants from phylotype IIB-4NPB that lack pathogenicity for banana but can infect many other plant species; and (iii) Moko disease-causing strains from phylotypes IIB-3, IIB-4, and IIA-6, historically known as race 2, that cause wilt on banana, plantain, andHeliconiaspp. We compared the genomes of 72R. solanacearumstrains, mainly from the three major ecotypes of phylotype II, using a newly developed pangenomic microarray to decipher their population structure and gain clues about the epidemiology of these ecotypes. Strain phylogeny and population structure were reconstructed. The results revealed a phylogeographic structure within brown rot strains, allowing us to distinguish European outbreak strains of Andean and African origins. The pangenomic CGH data also demonstrated that Moko ecotype IIB-4 is phylogenetically distinct from the emerging IIB-4NPB strains. These findings improved our understanding of the epidemiology of important ecotypes in phylotype II and will be useful for evolutionary analyses and the development of new DNA-based diagnostic tools.
Nanoparticles Functionalized with Ampicillin Destroy Multiple-Antibiotic-Resistant Isolates of Pseudomonas aeruginosa and Enterobacter aerogenes and Methicillin-Resistant Staphylococcus aureus
Authors: Brown Ashley N. , Smith Kathryn , Samuels Tova A. , Lu Jiangrui , Obare Sherine O. , Scott Maria E.
Published: January 2012

We show here that silver nanoparticles (AgNP) were intrinsically antibacterial, whereas gold nanoparticles (AuNP) were antimicrobial only when ampicillin was bound to their surfaces. Both AuNP and AgNP functionalized with ampicillin were effective broad-spectrum bactericides against Gram-negative and Gram-positive bacteria. Most importantly, when AuNP and AgNP were functionalized with ampicillin they became potent bactericidal agents with unique properties that subverted antibiotic resistance mechanisms of multiple-drug-resistant bacteria.
Factors Driving the Abundance of Ixodes ricinus Ticks and the Prevalence of Zoonotic I. ricinus-Borne Pathogens in Natural Foci
Authors: Ruiz-Fons Francisco , Fernández-de-Mera Isabel G. , Acevedo Pelayo , Gortázar Christian , de la Fuente José
Published: January 2012

Environmental factors may drive tick ecology and therefore tick-borne pathogen (TBP) epidemiology, which determines the risk to animals and humans of becoming infected by TBPs. For this reason, the aim of this study was to analyze the influence of environmental factors on the abundance of immature-stageIxodes ricinusticks and on the prevalence of two zoonoticI. ricinus-borne pathogens in natural foci of endemicity.I. ricinusabundance was measured at nine sites in the northern Iberian Peninsula by dragging the vegetation with a cotton flannelette, and ungulate abundance was measured by means of dung counts. In addition to ungulate abundance, data on variables related to spatial location, climate, and soil were gathered from the study sites.I. ricinusadults, nymphs, and larvae were collected from the vegetation, and a representative subsample ofI. ricinusnymphs from each study site was analyzed by PCR for the detection ofBorrelia burgdorferisensu lato andAnaplasma phagocytophilumDNA. Mean prevalences of these pathogens were 4.0% ± 1.8% and 20.5% ± 3.7%, respectively. Statistical analyses confirmed the influence of spatial factors, climate, and ungulate abundance onI. ricinuslarva abundance, while nymph abundance was related only to climate. Interestingly, cattle abundance rather than deer abundance was the main driver ofB. burgdorferisensu lato andA. phagocytophilumprevalence inI. ricinusnymphs in the study sites, where both domestic and wild ungulates coexist. The increasing abundance of cattle seems to increase the risk of other hosts becoming infected byA. phagocytophilum, while reducing the risk of being infected byB. burgdorferisensu lato. Controlling ticks in cattle in areas where they coexist with wild ungulates would be more effective for TBP control than reducing ungulate abundance.
Molecular Characterization of the EhaG and UpaG Trimeric Autotransporter Proteins from Pathogenic Escherichia coli
Authors: Totsika Makrina , Wells Timothy J. , Beloin Christophe , Valle Jaione , Allsopp Luke P. , King Nathan P. , Ghigo Jean-Marc , Schembri Mark A.
Published: January 2012

Trimeric autotransporter proteins (TAAs) are important virulence factors of many Gram-negative bacterial pathogens. A common feature of most TAAs is the ability to mediate adherence to eukaryotic cells or extracellular matrix (ECM) proteins via a cell surface-exposed passenger domain. Here we describe the characterization of EhaG, a TAA identified from enterohemorrhagic
Escherichia coli
(EHEC) O157:H7. EhaG is a positional orthologue of the recently characterized UpaG TAA from uropathogenic
E. coli
(UPEC). Similarly to UpaG, EhaG localized at the bacterial cell surface and promoted cell aggregation, biofilm formation, and adherence to a range of ECM proteins. However, the two orthologues display differential cellular binding: EhaG mediates specific adhesion to colorectal epithelial cells while UpaG promotes specific binding to bladder epithelial cells. The EhaG and UpaG TAAs contain extensive sequence divergence in their respective passenger domains that could account for these differences. Indeed, sequence analyses of UpaG and EhaG homologues from several
E. coli
genomes revealed grouping of the proteins in clades almost exclusively represented by distinct
E. coli
pathotypes. The expression of EhaG (in EHEC) and UpaG (in UPEC) was also investigated and shown to be significantly enhanced in an
hns
isogenic mutant, suggesting that H-NS acts as a negative regulator of both TAAs. Thus, while the EhaG and UpaG TAAs contain some conserved binding and regulatory features, they also possess important differences that correlate with the distinct pathogenic lifestyles of EHEC and UPEC.
Supplementation of Intracellular XylR Leads to Coutilization of Hemicellulose Sugars
Authors: Groff Dan , Benke Peter I. , Batth Tanveer S. , Bokinsky Gregory , Petzold Christopher J. , Adams Paul D. , Keasling Jay D.
Published: January 2012

Escherichia coli
has the potential to be a powerful biocatalyst for the conversion of lignocellulosic biomass into useful materials such as biofuels and polymers. One important challenge in using
E. coli
for the transformation of biomass sugars is diauxie, or sequential utilization of different types of sugars. We demonstrate that, by increasing the intracellular levels of the transcription factor XylR, the preferential consumption of arabinose before xylose can be eliminated. In addition, XylR augmentation must be finely tuned for robust coutilization of these two hemicellulosic sugars. Using a novel technique for scarless gene insertion, an additional copy of
xylR
was inserted into the
araBAD
operon. The resulting strain was superior at cometabolizing mixtures of arabinose and xylose and was able to produce at least 36% more ethanol than wild-type strains. This strain is a useful starting point for the development of an
E. coli
biocatalyst that can simultaneously convert all biomass sugars.
Engineered Escherichia coli Silver-Binding Periplasmic Protein That Promotes Silver Tolerance
Authors: Hall Sedlak Ruth , Hnilova Marketa , Grosh Carolynn , Fong Hanson , Baneyx Francois , Schwartz Dan , Sarikaya Mehmet , Tamerler Candan , Traxler Beth
Published: January 2012

Silver toxicity is a problem that microorganisms face in medical and environmental settings. Through exposure to silver compounds, some bacteria have adapted to growth in high concentrations of silver ions. Such adapted microbes may be dangerous as pathogens but, alternatively, could be potentially useful in nanomaterial-manufacturing applications. While naturally adapted isolates typically utilize efflux pumps to achieve metal resistance, we have engineered a silver-tolerant
Escherichia coli
strain by the use of a simple silver-binding peptide motif. A silver-binding peptide, AgBP2, was identified from a combinatorial display library and fused to the C terminus of the
E. coli
maltose-binding protein (MBP) to yield a silver-binding protein exhibiting nanomolar affinity for the metal. Growth experiments performed in the presence of silver nitrate showed that cells secreting MBP-AgBP2 into the periplasm exhibited silver tolerance in a batch culture, while those expressing a cytoplasmic version of the fusion protein or MBP alone did not. Transmission electron microscopy analysis of silver-tolerant cells revealed the presence of electron-dense silver nanoparticles. This is the first report of a specifically engineered metal-binding peptide exhibiting a strong
in vivo
phenotype, pointing toward a novel ability to manipulate bacterial interactions with heavy metals by the use of short and simple peptide motifs. Engineered metal-ion-tolerant microorganisms such as this
E. coli
strain could potentially be used in applications ranging from remediation to interrogation of biomolecule-metal interactions
in vivo
.
Significant Role for Microbial Autotrophy in the Sequestration of Soil Carbon
Authors: Yuan Hongzhao , Ge Tida , Chen Caiyan , O'Donnell Anthony G. , Wu Jinshui
Published: January 2012

Soils were incubated for 80 days in a continuously labeled
14
CO
2
atmosphere to measure the amount of labeled C incorporated into the microbial biomass. Microbial assimilation of
14
C differed between soils and accounted for 0.12% to 0.59% of soil organic carbon (SOC). Assuming a terrestrial area of 1.4 × 10
8
km
2
, this represents a potential global sequestration of 0.6 to 4.9 Pg C year
−1
. Estimated global C sequestration rates suggest a “missing sink” for carbon of between 2 and 3 Pg C year
−1
. To determine whether
14
CO
2
incorporation was mediated by autotrophic microorganisms, the diversity and abundance of CO
2
-fixing bacteria and algae were investigated using clone library sequencing, terminal restriction fragment length polymorphism (T-RFLP), and quantitative PCR (qPCR) of the ribulose 1,5-bisphosphate carboxylase/oxygenase (RubisCO) gene (
cbbL
). Phylogenetic analysis showed that the dominant
cbbL
-containing bacteria were
Azospirillum lipoferum
,
Rhodopseudomonas palustris
,
Bradyrhizobium japonicum
,
Ralstonia eutropha
, and
cbbL
-containing chromophytic algae of the genera
Xanthophyta
and
Bacillariophyta
. Multivariate analyses of T-RFLP profiles revealed significant differences in
cbbL
-containing microbial communities between soils. Differences in
cbbL
gene diversity were shown to be correlated with differences in SOC content. Bacterial and algal
cbbL
gene abundances were between 10
6
and 10
8
and 10
3
to 10
5
copies g
−1
soil, respectively. Bacterial
cbbL
abundance was shown to be positively correlated with RubisCO activity (
r
= 0.853;
P
< 0.05), and both
cbbL
abundance and RubisCO activity were significantly related to the synthesis rates of [
14
C]SOC (
r
= 0.967 and 0.946, respectively;
P
< 0.01). These data offer new insights into the importance of microbial autotrophy in terrestrial C cycling.
Functional Cloning and Characterization of Antibiotic Resistance Genes from the Chicken Gut Microbiome
Authors: Zhou Wei , Wang Ying , Lin Jun
Published: January 2012

Culture-independent sampling in conjunction with a functional cloning approach identified diverse antibiotic resistance genes for different classes of antibiotics in gut microbiomes from both conventionally raised and free-range chickens. Many of the genes are phylogenetically distant from known resistance genes. Two unique genes that conferred ampicillin and spectinomycin resistance were also functional in
Campylobacter
, a distant relative of the
Escherichia coli
host used to generate the genomic libraries.
Disulfide Bond Formation and Activation of Escherichia coli β-Galactosidase under Oxidizing Conditions
Authors: Seras-Franzoso Joaquin , Affentranger Roman , Ferrer-Navarro Mario , Daura Xavier , Villaverde Antonio , García-Fruitós Elena
Published: January 2012

Escherichia coli
β-galactosidase is probably the most widely used reporter enzyme in molecular biology, cell biology, and biotechnology because of the easy detection of its activity. Its large size and tetrameric structure make this bacterial protein an interesting model for crystallographic studies and atomic mapping. In the present study, we investigate a version of
Escherichia coli
β-galactosidase produced under oxidizing conditions, in the cytoplasm of an Origami strain. Our data prove the activation of this microbial enzyme under oxidizing conditions and clearly show the occurrence of a disulfide bond in the β-galactosidase structure. Additionally, the formation of this disulfide bond is supported by the analysis of a homology model of the protein that indicates that two cysteines located in the vicinity of the catalytic center are sufficiently close for disulfide bond formation.
Influence of Cations on Growth of Thermophilic Geobacillus spp. and Anoxybacillus flavithermus in Planktonic Culture
Authors: Somerton Ben , Palmer Jon , Brooks John , Smolinski Edward , Lindsay Denise , Flint Steve
Published: January 2012

Free ions of Na
+
, K
+
, Ca
2+
, and Mg
2+
influenced the optical density of planktonic cultures of thermophilic bacilli.
Anoxybacillus flavithermus
E16 and
Geobacillus
sp. strain F75 (milk powder manufacturing plant isolates) and
A. flavithermus
DSM 2641 and
G. thermoleovorans
DSM 5366 were studied. Ca
2+
and Mg
2+
were associated with increases in optical density more so than Na
+
and K
+
. Overall, it appeared that Ca
2+
and/or Mg
2+
was required for the production of protein in thermophilic bacilli, as shown by results obtained with
A. flavithermus
E16, which was selected for further study.
Roles of Protein Kinase A and Adenylate Cyclase in Light-Modulated Cellulase Regulation in Trichoderma reesei
Authors: Schuster André , Tisch Doris , Seidl-Seiboth Verena , Kubicek Christian P. , Schmoll Monika
Published: January 2012

The cyclic AMP (cAMP) pathway represents a central signaling cascade with crucial functions in all organisms. Previous studies of
Trichoderma reesei
(anamorph of
Hypocrea jecorina
) suggested a function of cAMP signaling in regulation of cellulase gene expression. We were therefore interested in how the crucial components of this pathway, adenylate cyclase (ACY1) and cAMP-dependent protein kinase A (PKA), would affect cellulase gene expression. We found that both ACY1 and PKA catalytic subunit 1 (PKAC1) are involved in regulation of vegetative growth but are not essential for sexual development. Interestingly, our results showed considerably increased transcript abundance of cellulase genes in darkness compared to light (light responsiveness) upon growth on lactose. This effect is strongly enhanced in mutant strains lacking PKAC1 or ACY1. Comparison to the wild type showed that ACY1 has a consistently positive effect on cellulase gene expression in light and darkness, while PKAC1 influences transcript levels of cellulase genes positively in light but negatively in darkness. A function of PKAC1 in light-modulated cellulase gene regulation is also reflected by altered complex formation within the
cel6a/cbh2
promoter in light and darkness and in the absence of
pkac1
. Analysis of transcript levels of cellulase regulator genes indicates that the regulatory output of the cAMP pathway may be established via adjustment of XYR1 abundance. Consequently, both adenylate cyclase and protein kinase A are involved in light-modulated cellulase gene expression in
T. reesei
and have a dampening effect on the light responsiveness of this process.
Population Structure of and Mycotoxin Production by Fusarium graminearum from Maize in South Korea
Authors: Lee Jungkwan , Kim Hun , Jeon Jae-Jin , Kim Hye-Seon , Zeller Kurt A. , Carter Laurel L. A. , Leslie John F. , Lee Yin-Won
Published: January 2012

Fusarium graminearum
(
Gibberella zeae
) is an important pathogen of wheat, maize, barley, and rice in South Korea, and harvested grain often is contaminated with trichothecenes such as deoxynivalenol and nivalenol. In this study, we examined 568 isolates of
F. graminearum
collected from maize at eight locations in South Korea. We used amplified fragment length polymorphisms (AFLPs) to identify four lineages (2, 3, 6, and 7); lineage 7 was the most common (75%), followed by lineage 6 (12%), lineage 3 (12%), and lineage 2 (1%). The genetic identity among populations was high (>0.98), and the effective migration rate between locations was higher than that between lineages. Female fertility varied by lineage: all lineage 7 isolates were fertile, while 70%, 26%, and 14% of the isolates in lineages 6, 3, and 2, respectively, were fertile. All lineage 3 and lineage 7 isolates produced deoxynivalenol, whereas most lineage 2 and 6 isolates produced nivalenol. Genotypic diversity in lineage 3 and lineage 6 populations is similar to that found in previously described Korean rice populations, but genotypic diversity in lineage 7 is much lower, even though similar levels of gene flow occur between lineage 7 populations. We conclude that lineage 7 was relatively recently introduced into South Korea, perhaps accompanying imported maize seeds.
Chimeric Phage Lysins Act Synergistically with Lysostaphin To Kill Mastitis-Causing Staphylococcus aureus in Murine Mammary Glands
Authors: Schmelcher Mathias , Powell Anne M. , Becker Stephen C. , Camp Mary J. , Donovan David M.
Published: January 2012

Staphylococci cause bovine mastitis, with
Staphylococcus aureus
being responsible for the majority of the mastitis-based losses to the dairy industry (up to $2 billion/annum). Treatment is primarily with antibiotics, which are often ineffective and potentially contribute to resistance development. Bacteriophage endolysins (peptidoglycan hydrolases) present a promising source of alternative antimicrobials. Here we evaluated two fusion proteins consisting of the streptococcal λSA2 endolysin endopeptidase domain fused to staphylococcal cell wall binding domains from either lysostaphin (λSA2-E-Lyso-SH3b) or the staphylococcal phage K endolysin, LysK (λSA2-E-LysK-SH3b). We demonstrate killing of 16 different
S. aureus
mastitis isolates, including penicillin-resistant strains, by both constructs. At 100 μg/ml in processed cow milk, λSA2-E-Lyso-SH3b and λSA2-E-LysK-SH3b reduced the
S. aureus
bacterial load by 3 and 1 log units within 3 h, respectively, compared to a buffer control. In contrast to λSA2-E-Lyso-SH3b, however, λSA2-E-LysK-SH3b permitted regrowth of the pathogen after 1 h. In a mouse model of mastitis, infusion of 25 μg of λSA2-E-Lyso-SH3b or λSA2-E-LysK-SH3b into mammary glands reduced
S. aureus
CFU by 0.63 or 0.81 log units, compared to >2 log for lysostaphin. Both chimeras were synergistic with lysostaphin against
S. aureus
in plate lysis checkerboard assays. When tested in combination in mice, λSA2-E-LysK-SH3b and lysostaphin (12.5 μg each/gland) caused a 3.36-log decrease in CFU. Furthermore, most protein treatments reduced gland wet weights and intramammary tumor necrosis factor alpha (TNF-α) concentrations, which serve as indicators of inflammation. Overall, our animal model results demonstrate the potential of fusion peptidoglycan hydrolases as antimicrobials for the treatment of
S. aureus
-induced mastitis.
Abundance and Single-Cell Activity of Heterotrophic Bacterial Groups in the Western Arctic Ocean in Summer and Winter
Authors: Nikrad Mrinalini P. , Cottrell M. T. , Kirchman D. L.
Published: January 2012

Environmental conditions in the western Arctic Ocean range from constant light and nutrient depletion in summer to complete darkness and sea ice cover in winter. This seasonal environmental variation is likely to have an effect on the use of dissolved organic matter (DOM) by heterotrophic bacteria in surface water. However, this effect is not well studied and we know little about the activity of specific bacterial clades in the surface oceans. The use of DOM by three bacterial subgroups in both winter and summer was examined by microautoradiography combined with fluorescencein situhybridization. We found selective use of substrates by these groups, although the abundances of Ant4D3 (AntarcticGammaproteobacteria),Polaribacter(Bacteroidetes), and SAR11 (Alphaproteobacteria) were not different between summer and winter in the Beaufort and Chukchi Seas. The number of cells taking up glucose within all three bacterial groups decreased significantly from summer to winter, while the percentage of cells using leucine did not show a clear pattern between seasons. The uptake of the amino acid mix increased substantially from summer to winter by the Ant4D3 group, although such a large increase in uptake was not seen for the other two groups. Use of glucose by bacteria, but not use of leucine or the amino acid mix, related strongly to inorganic nutrients, chlorophylla, and other environmental factors. Our results suggest a switch in use of dissolved organic substrates from summer to winter and that the three phylogenetic subgroups examined fill different niches in DOM use in the two seasons.
Construction and Characterization of a Gradually Inducible Expression Vector for Halobacterium salinarum, Based on the
kdp
Promoter
Authors: Kixmüller Dorthe , Greie Jörg-Christian
Published: January 2012

Gradually inducible expression vectors which are governed by variations of growth conditions are powerful tools for gene expression of conditionally lethal mutants. Furthermore, controlled expression allows monitoring of overproduction of proteins at various stages in their expressing hosts. For
Halobacterium salinarum
, which is often used as a paradigm for halophilic archaea, such an inducible expression system is not available to date. Here we show that the
kdp
promoter (P
kdp
), which facilitates gene expression upon K
+
limitation, can be used to establish such a system for molecular applications. P
kdp
features a rather high expression rate, with an approximately 50-fold increase that can be easily varied by K
+
concentrations in the growth medium. Besides the construction of an expression vector, our work describes the characterization of expression patterns and, thus, offers a gradually inducible expression system to the scientific community.
Three Types of Taxis Used in the Response of
Acidovorax
sp. Strain JS42 to 2-Nitrotoluene
Authors: Rabinovitch-Deere Christine A. , Parales Rebecca E.
Published: January 2012

Acidovorax
sp. strain JS42 is able to utilize 2-nitrotoluene (2NT) as its sole carbon, nitrogen, and energy source. We report here that strain JS42 is chemotactic to 2NT and that the response is increased when cells are grown on compounds such as 2NT that are known to induce the first step of 2NT degradation. Assays with JS42 mutants unable to oxidize 2NT showed that the first step of 2NT metabolism was required for the induced response, but not for a portion of the constitutive response, indicating that 2NT itself is an attractant. The 2NT metabolite nitrite was shown to be a strong attractant for strain JS42, and sufficient nitrite was produced during the taxis assay to account for a large part of the induced response. A mutant with an inactivated
ntdY
gene, which is located adjacent to the 2NT degradation genes and codes for a putative methyl-accepting chemotaxis protein, showed a defect in taxis toward 2NT that may involve a reduced response to nitrite. Responses of a mutant defective for the energy-taxis receptor, Aer, indicated that a functional
aer
gene is required for a substantial part of the wild-type induced response to 2NT. In summary, strain JS42 utilizes three types of taxis to sense and respond to 2NT: constitutive 2NT-specific chemotaxis to directly sense 2NT, metabolism-dependent nitrite-specific chemotaxis that may be mediated by NtdY, and energy taxis mediated by Aer.
Differential Isotopic Fractionation during Cr(VI) Reduction by an Aquifer-Derived Bacterium under Aerobic versus Denitrifying Conditions
Authors: Han Ruyang , Qin Liping , Brown Shaun T. , Christensen John N. , Beller Harry R.
Published: January 2012

We studied Cr isotopic fractionation during Cr(VI) reduction by
Pseudomonas stutzeri
strain RCH2. Despite the fact that strain RCH2 reduces Cr(VI) cometabolically under both aerobic and denitrifying conditions and at similar specific rates, fractionation was markedly different under these two conditions (ε was ∼2‰ aerobically and ∼0.4‰ under denitrifying conditions).
Geraniol and Geranial Dehydrogenases Induced in Anaerobic Monoterpene Degradation by Castellaniella defragrans
Authors: Lüddeke Frauke , Wülfing Annika , Timke Markus , Germer Frauke , Weber Johanna , Dikfidan Aytac , Rahnfeld Tobias , Linder Dietmar , Meyerdierks Anke , Harder Jens
Published: January 2012

Castellaniella defragrans
is a
Betaproteobacterium
capable of coupling the oxidation of monoterpenes with denitrification. Geraniol dehydrogenase (GeDH) activity was induced during growth with limonene in comparison to growth with acetate. The N-terminal sequence of the purified enzyme directed the cloning of the corresponding open reading frame (ORF), the first bacterial gene for a GeDH (
geoA
, for
ge
raniol
o
xidation pathway). The
C. defragrans
geraniol dehydrogenase is a homodimeric enzyme that affiliates with the zinc-containing benzyl alcohol dehydrogenases in the superfamily of medium-chain-length dehydrogenases/reductases (MDR). The purified enzyme most efficiently catalyzes the oxidation of perillyl alcohol (
k
cat
/
K
m
= 2.02 × 10
6
M
−1
s
−1
), followed by geraniol (
k
cat
/
K
m
= 1.57 × 10
6
M
−1
s
−1
). Apparent
K
m
values of <10 μM are consistent with an
in vivo
toxicity of geraniol above 5 μM. In the genetic vicinity of
geoA
is a putative aldehyde dehydrogenase that was named
geoB
and identified as a highly abundant protein during growth with phellandrene. Extracts of
Escherichia coli
expressing
geoB
demonstrated
in vitro
a geranial dehydrogenase (GaDH) activity. GaDH activity was independent of coenzyme A. The irreversible formation of geranic acid allows for a metabolic flux from β-myrcene via linalool, geraniol, and geranial to geranic acid.
Molecular Basis and Phylogenetic Implications of Deoxycylindrospermopsin Biosynthesis in the Cyanobacterium Raphidiopsis curvata
Authors: Jiang Yongguang , Xiao Peng , Yu Gongliang , Sano Tomoharu , Pan Qianqian , Li Renhui
Published: January 2012

New insights into the distribution and biochemistry of the cyanotoxin cylindrospermopsin (CYN) have been provided by the recent determination of its biosynthesis gene cluster (
cyr
) in several cyanobacterial species.
Raphidiopsis curvata
CHAB1150 isolated from China was analyzed for CYN analogues. Only 7-deoxy-CYN was detected in the cell extracts. The
cyr
gene cluster of
R. curvata
CHAB1150 was sequenced, and the
cyr
genes of this strain were found to have extremely high similarities (96% to 100%) to those from other nostocalean species. These species include
Cylindrospermopsis raciborskii
AWT205,
Aphanizomenon
sp. strain 10E6, and
Aphanizomenon ovalisporum
ILC-146. Insertion mutation was identified within the
cyrI
gene, and transcripts of
cyrI
and another functional gene
cyrJ
were detected in
R. curvata
CHAB1150. General congruence between the phylogenetic trees based on both
cyr
and 16S
rrn
was displayed. Neutral evolution was found on the whole sequences of the
cyr
genes, and 0 to 89 negative selected codons were detected in each gene. Therefore, the function of CyrI is to catalyze the oxygenation of 7-deoxy-CYN in CYN biosynthesis. The transcripts of the mutated
cyrI
gene may result from polycistronic transcription. The high conservation of the
cyr
genes may be ascribed to purifying selection and horizontal gene transfer.
Toward Biosynthetic Design and Implementation of Escherichia coli-Derived Paclitaxel and Other Heterologous Polyisoprene Compounds
Authors: Jiang Ming , Stephanopoulos Gregory , Pfeifer Blaine A.
Published: January 2012

Escherichia coli
offers unparalleled engineering capacity in the context of heterologous natural product biosynthesis. However, as with other heterologous hosts, cellular metabolism must be designed or redesigned to support final compound formation. This task is at once complicated and aided by the fact that the cell does not natively produce an abundance of natural products. As a result, the metabolic engineer avoids complicated interactions with native pathways closely associated with the outcome of interest, but this convenience is tempered by the need to implement the required metabolism to allow functional biosynthesis. This review focuses on engineering
E. coli
for the purpose of polyisoprene formation, as it is related to isoprenoid compounds currently being pursued through a heterologous approach. In particular, the review features the compound paclitaxel and early efforts to design and overproduce intermediates through
E. coli
.
Cloning and Sequencing of Inulinase and β-Fructofuranosidase Genes of a Deep-Sea Microbulbifer Species and Properties of Recombinant Enzymes
Authors: Kobayashi Tohru , Uchimura Kohsuke , Deguchi Shigeru , Horikoshi Koki
Published: January 2012

An inulinase-producing
Microbulbifer
sp. strain, JAM-3301, was isolated from a deep-sea sediment. An inulin operon that contained three open reading frames was cloned and sequenced. Two of the three genes were expressed. One product was an endo-inulinase, and the other was a β-fructofuranosidase. Both enzymes worked together to effectively degrade inulin.
Caenorhabditis elegans, a Model Organism for Investigating Immunity
Authors: Marsh Elizabeth K. , May Robin C.
Published: January 2012

The nematode
Caenorhabditis elegans
has been a powerful experimental organism for almost half a century. Over the past 10 years, researchers have begun to exploit the power of
C. elegans
to investigate the biology of a number of human pathogens. This work has uncovered mechanisms of host immunity and pathogen virulence that are analogous to those involved during pathogenesis in humans or other animal hosts, as well as novel immunity mechanisms which appear to be unique to the worm. More recently, these investigations have uncovered details of the natural pathogens of
C. elegans
, including the description of a novel intracellular microsporidian parasite as well as new nodaviruses, the first identification of viral infections of this nematode. In this review, we consider the application of
C. elegans
to human infectious disease research, as well as consider the nematode response to these natural pathogens.
Comprehensive Set of Integrative Plasmid Vectors for Copper-Inducible Gene Expression in Myxococcus xanthus
Authors: Gómez-Santos Nuria , Treuner-Lange Anke , Moraleda-Muñoz Aurelio , García-Bravo Elena , García-Hernández Raquel , Martínez-Cayuela Marina , Pérez Juana , Søgaard-Andersen Lotte , Muñoz-Dorado José
Published: January 2012

Myxococcus xanthus
is widely used as a model system for studying gliding motility, multicellular development, and cellular differentiation. Moreover,
M. xanthus
is a rich source of novel secondary metabolites. The analysis of these processes has been hampered by the limited set of tools for inducible gene expression. Here we report the construction of a set of plasmid vectors to allow copper-inducible gene expression in
M. xanthus
. Analysis of the effect of copper on strain DK1622 revealed that copper concentrations of up to 500 μM during growth and 60 μM during development do not affect physiological processes such as cell viability, motility, or aggregation into fruiting bodies. Of the copper-responsive promoters in
M. xanthus
reported so far, the multicopper oxidase
cuoA
promoter was used to construct expression vectors, because no basal expression is observed in the absence of copper and induction linearly depends on the copper concentration in the culture medium. Four different plasmid vectors have been constructed, with different marker selection genes and sites of integration in the
M. xanthus
chromosome. The vectors have been tested and gene expression quantified using the
lacZ
gene. Moreover, we demonstrate the functional complementation of the motility defect caused by lack of PilB by the copper-induced expression of the
pilB
gene. These versatile vectors are likely to deepen our understanding of the biology of
M. xanthus
and may also have biotechnological applications.
Function of the Pyruvate Oxidase-Lactate Oxidase Cascade in Interspecies Competition between Streptococcus oligofermentans and Streptococcus mutans
Authors: Liu Lei , Tong Huichun , Dong Xiuzhu
Published: January 2012

Complex interspecies interactions occur constantly between oral commensals and the opportunistic pathogen
Streptococcus mutans
in dental plaque. Previously, we showed that oral commensal
Streptococcus oligofermentans
possesses multiple enzymes for H
2
O
2
production, especially lactate oxidase (Lox), allowing it to out-compete
S. mutans
. In this study, through extensive biochemical and genetic studies, we identified a pyruvate oxidase (
pox
) gene in
S. oligofermentans
. A
pox
deletion mutant completely lost Pox activity, while ectopically expressed
pox
restored activity. Pox was determined to produce most of the H
2
O
2
in the earlier growth phase and log phase, while Lox mainly contributed to H
2
O
2
production in stationary phase. Both
pox
and
lox
were expressed throughout the growth phase, while expression of the
lox
gene increased by about 2.5-fold when cells entered stationary phase. Since lactate accumulation occurred to a large degree in stationary phase, the differential Pox- and Lox-generated H
2
O
2
can be attributed to differential gene expression and substrate availability. Interestingly, inactivation of
pox
causes a dramatic reduction in H
2
O
2
production from lactate, suggesting a synergistic action of the two oxidases in converting lactate into H
2
O
2
. In an
in vitro
two-species biofilm experiment, the
pox
mutant of
S. oligofermentans
failed to inhibit
S. mutans
even though
lox
was active. In summary,
S. oligofermentans
develops a Pox-Lox synergy strategy to maximize its H
2
O
2
formation so as to win the interspecies competition.
Phylogenetic Clustering of Soil Microbial Communities by 16S rRNA but Not 16S rRNA Genes
Authors: DeAngelis Kristen M. , Firestone Mary K.
Published: January 2012

We evaluated phylogenetic clustering of bacterial and archaeal communities from redox-dynamic subtropical forest soils that were defined by 16S rRNA and rRNA gene sequences. We observed significant clustering for the RNA-based communities but not the DNA-based communities, as well as increasing clustering over time of the highly active taxa detected by only rRNA.
The Histone-Like Nucleoid Structuring Protein (H-NS) Is a Repressor of Vibrio cholerae Exopolysaccharide Biosynthesis (
vps
) Genes
Authors: Wang Hongxia , Ayala Julio C. , Silva Anisia J. , Benitez Jorge A.
Published: January 2012

The capacity of
Vibrio cholerae
to form biofilms has been shown to enhance its survival in the aquatic environment and play important roles in pathogenesis and disease transmission. In this study, we demonstrated that the histone-like nucleoid structuring protein is a repressor of exopolysaccharide (
vps
) biosynthesis genes and biofilm formation.
Ring-Cleaving Dioxygenases with a Cupin Fold
Authors: Fetzner Susanne
Published: January 2012

Ring-cleaving dioxygenases catalyze key reactions in the aerobic microbial degradation of aromatic compounds. Many pathways converge to catecholic intermediates, which are subject toorthoormetacleavage by intradiol or extradiol dioxygenases, respectively. However, a number of degradation pathways proceed via noncatecholic hydroxy-substituted aromatic carboxylic acids like gentisate, salicylate, 1-hydroxy-2-naphthoate, or aminohydroxybenzoates. The ring-cleaving dioxygenases active toward these compounds belong to the cupin superfamily, which is characterized by a six-stranded β-barrel fold and conserved amino acid motifs that provide the 3His or 2- or 3His-1Glu ligand environment of a divalent metal ion. Most cupin-type ring cleavage dioxygenases use an FeIIcenter for catalysis, and the proposed mechanism is very similar to that of the canonical (type I) extradiol dioxygenases. The metal ion is presumed to act as an electron conduit for single electron transfer from the metal-bound substrate anion to O2, resulting in activation of both substrates to radical species. The family of cupin-type dioxygenases also involves quercetinase (flavonol 2,4-dioxygenase), which opens up two C-C bonds of the heterocyclic ring of quercetin, a wide-spread plant flavonol. Remarkably, bacterial quercetinases are capable of using different divalent metal ions for catalysis, suggesting that the redox properties of the metal are relatively unimportant for the catalytic reaction. The major role of the active-site metal ion could be to correctly position the substrate and to stabilize transition states and intermediates rather than to mediate electron transfer. The tentative hypothesis that quercetinase catalysis involves direct electron transfer from metal-bound flavonolate to O2is supported by model chemistry.
Biology and Genome Sequence of Streptococcus mutans Phage M102AD
Authors: Delisle Allan L. , Guo Ming , Chalmers Natalia I. , Barcak Gerard J. , Rousseau Geneviève M. , Moineau Sylvain
Published: January 2012

M102AD is the new designation for a
Streptococcus mutans
phage described in 1993 as phage M102. This change was necessitated by the genome analysis of another
S. mutans
phage named M102, which revealed differences from the genome sequence reported here. Additional host range analyses confirmed that
S. mutans
phage M102AD infects only a few serotype c strains. Phage M102AD adsorbed very slowly to its host, and it cannot adsorb to serotype e and f strains of
S. mutans
. M102AD adsorption was blocked by c-specific antiserum. Phage M102AD also adsorbed equally well to heat-treated and trypsin-treated cells, suggesting carbohydrate receptors. Saliva and polysaccharide production did not inhibit plaque formation. The genome of this siphophage consisted of a linear, double-stranded, 30,664-bp DNA molecule, with a GC content of 39.6%. Analysis of the genome extremities indicated the presence of a 3′-overhang
cos
site that was 11 nucleotides long. Bioinformatic analyses identified 40 open reading frames, all in the same orientation. No lysogeny-related genes were found, indicating that phage M102AD is strictly virulent. No obvious virulence factor gene candidates were found. Twelve proteins were identified in the virion structure by mass spectrometry. Comparative genomic analysis revealed a close relationship between
S. mutans
phages M102AD and M102 as well as with
Streptococcus thermophilus
phages. This study also highlights the importance of conducting research with biological materials obtained from recognized microbial collections.
Different Virucidal Activities of Hyperbranched Quaternary Ammonium Coatings on Poliovirus and Influenza Virus
Authors: Tuladhar Era , de Koning Martijn C. , Fundeanu Irina , Beumer Rijkelt , Duizer Erwin
Published: January 2012

Virucidal activity of immobilized quaternary ammonium compounds (IQACs) coated onto glass and plastic surfaces was tested against enveloped influenza A (H1N1) virus and nonenveloped poliovirus Sabin1. The IQACs tested were virucidal against the influenza virus within 2 min, but no virucidal effect against poliovirus was found in 6 h.
Tumor Necrosis Factor Alpha Modulates the Dynamics of the Plasminogen-Mediated Early Interaction between Bifidobacterium animalis subsp.
lactis
and Human Enterocytes
Authors: Centanni Manuela , Bergmann Simone , Turroni Silvia , Hammerschmidt Sven , Chhatwal Gursharan Singh , Brigidi Patrizia , Candela Marco
Published: January 2012

The capacity to intervene with the host plasminogen system has recently been considered an important component in the interaction process between
Bifidobacterium animalis
subsp.
lactis
and the human host. However, its significance in the bifidobacterial microecology within the human gastrointestinal tract is still an open question. Here we demonstrate that human plasminogen favors the
B. animalis
subsp.
lactis
BI07 adhesion to HT29 cells. Prompting the HT29 cell capacity to activate plasminogen, tumor necrosis factor alpha (TNF-α) modulated the plasminogen-mediated bacterium-enterocyte interaction, reducing the bacterial adhesion to the enterocytes and enhancing migration to the luminal compartment.
Analysis of the Functions of Recombination-Related Genes in the Generation of Large Chromosomal Deletions by Loop-Out Recombination in Aspergillus oryzae
Authors: Takahashi Tadashi , Ogawa Masahiro , Koyama Yasuji
Published: January 2012

Loop-out-type recombination is a type of intrachromosomal recombination followed by the excision of a chromosomal region. The detailed mechanism underlying this recombination and the genes involved in loop-out recombination remain unknown. In the present study, we investigated the functions of
ku70
,
ligD
,
rad52
,
rad54
, and
rdh54
in the construction of large chromosomal deletions via loop-out recombination and the effect of the position of the targeted chromosomal region on the efficiency of loop-out recombination in
Aspergillus oryzae
. The efficiency of generation of large chromosomal deletions in the near-telomeric region of chromosome 3, including the aflatoxin gene cluster, was compared with that in the near-centromeric region of chromosome 8, including the tannase gene. In the Δ
ku70
and Δ
ku70-rdh54
strains, only precise loop-out recombination occurred in the near-telomeric region. In contrast, in the Δ
ligD
, Δ
ku70-rad52
, and Δ
ku70-rad54
strains, unintended chromosomal deletions by illegitimate loop-out recombination occurred in the near-telomeric region. In addition, large chromosomal deletions via loop-out recombination were efficiently achieved in the near-telomeric region, but barely achieved in the near-centromeric region, in the Δ
ku70
strain. Induction of DNA double-strand breaks by I-SceI endonuclease facilitated large chromosomal deletions in the near-centromeric region. These results indicate that
ligD
,
rad52
, and
rad54
play a role in the generation of large chromosomal deletions via precise loop-out-type recombination in the near-telomeric region and that loop-out recombination between distant sites is restricted in the near-centromeric region by chromosomal structure.
Mitochondrial Sorting and Assembly Machinery Subunit Sam37 in Candida albicans: Insight into the Roles of Mitochondria in Fitness, Cell Wall Integrity, and Virulence
Authors: Qu Yue , Jelicic Branka , Pettolino Filomena , Perry Andrew , Lo Tricia L. , Hewitt Victoria L. , Bantun Farkad , Beilharz Traude H. , Peleg Anton Y. , Lithgow Trevor , Djordjevic Julianne T. , Traven Ana
Published: January 2012

Recent studies indicate that mitochondrial functions impinge on cell wall integrity, drug tolerance, and virulence of human fungal pathogens. However, the mechanistic aspects of these processes are poorly understood. We focused on the mitochondrial outer membrane SAM (
S
orting and
A
ssembly
M
achinery) complex subunit Sam37 in
Candida albicans
. Inactivation of
SAM37
in
C. albicans
leads to a large reduction in fitness, a phenotype not conserved with the model yeast
Saccharomyces cerevisiae
. Our data indicate that slow growth of the
sam37ΔΔ
mutant results from mitochondrial DNA loss, a new function for Sam37 in
C. albicans
, and from reduced activity of the essential SAM complex subunit Sam35. The
sam37ΔΔ
mutant was hypersensitive to drugs that target the cell wall and displayed altered cell wall structure, supporting a role for Sam37 in cell wall integrity in
C. albicans
. The sensitivity of the mutant to membrane-targeting antifungals was not significantly altered. The
sam37ΔΔ
mutant was avirulent in the mouse model, and bioinformatics showed that the fungal Sam37 proteins are distant from their animal counterparts and could thus represent potential drug targets. Our study provides the first direct evidence for a link between mitochondrial function and cell wall integrity in
C. albicans
and is further relevant for understanding mitochondrial function in fitness, antifungal drug tolerance, and virulence of this major pathogen. Beyond the relevance to fungal pathogenesis, this work also provides new insight into the mitochondrial and cellular roles of the SAM complex in fungi.