bluuebunny/crossref_metadata_2025 huggingface dataset:
Experimental Transmission of Infectious Pancreatic Necrosis Virus from the Blue Mussel, Mytilus edulis, to Cohabitating Atlantic Salmon (Salmo salar) Smolts
Authors: Molloy Sally D. , Pietrak Michael R. , Bricknell Ian , Bouchard Deborah A.
Published: July 2013

Integrated multitrophic aquaculture (IMTA) reduces the environmental impacts of commercial aquaculture systems by combining the cultivation of fed species with extractive species. Shellfish play a critical role in IMTA systems by filter-feeding particulate-bound organic nutrients. As bioaccumulating organisms, shellfish may also increase disease risk on farms by serving as reservoirs for important finfish pathogens such as infectious pancreatic necrosis virus (IPNV). The ability of the blue mussel (
Mytilus edulis
) to bioaccumulate and transmit IPNV to naive Atlantic salmon (
Salmo salar
) smolts was investigated. To determine the ability of mussels to filter and accumulate viable IPNV, mussels were held in water containing log 4.6 50% tissue culture infective dose(s) (TCID
50
) of the West Buxton strain of IPNV ml
−1
. Viable IPNV was detected in the digestive glands (DGs) of IPNV-exposed mussels as early as 2 h postexposure. The viral load in mussel DG tissue significantly increased with time and reached log 5.35 ± 0.25 TCID
50
g of DG tissue
−1
after 120 h of exposure. IPNV titers never reached levels that were significantly greater than that in the water. Viable IPNV was detected in mussel feces out to 7 days postdepuration, and the virus persisted in DG tissues for at least 18 days of depuration. To determine whether IPNV can be transmitted from mussels to Atlantic salmon, IPNV-exposed mussels were cohabitated with naive Atlantic salmon smolts. Transmission of IPNV did occur from mussels to smolts at a low frequency. The results demonstrate that a nonenveloped virus, such as IPNV, can accumulate in mussels and be transferred to naive fish.
Diet Complexity and Estrogen Receptor β Status Affect the Composition of the Murine Intestinal Microbiota
Authors: Menon Rani , Watson Sara E. , Thomas Laura N. , Allred Clinton D. , Dabney Alan , Azcarate-Peril M. Andrea , Sturino Joseph M.
Published: July 2013

Intestinal microbial dysbiosis contributes to the dysmetabolism of luminal factors, including steroid hormones (sterones) that affect the development of chronic gastrointestinal inflammation and the incidence of sterone-responsive cancers of the breast, prostate, and colon. Little is known, however, about the role of specific host sterone nucleoreceptors, including estrogen receptor β (ERβ), in microbiota maintenance. Herein, we test the hypothesis that ERβ status affects microbiota composition and determine if such compositionally distinct microbiota respond differently to changes in diet complexity that favorProteobacteriaenrichment. To this end, conventionally raised female ERβ+/+and ERβ−/−C57BL/6J mice (mean age of 27 weeks) were initially reared on 8604, a complex diet containing estrogenic isoflavones, and then fed AIN-76, an isoflavone-free semisynthetic diet, for 2 weeks. 16S rRNA gene surveys revealed that the fecal microbiota of 8604-fed mice and AIN-76-fed mice differed, as expected. The relative diversity ofProteobacteria, especially theAlphaproteobacteriaandGammaproteobacteria, increased significantly following the transition to AIN-76. Distinct patterns for beneficialLactobacillaleswere exclusive to and highly abundant among 8604-fed mice, whereas severalProteobacteriawere exclusive to AIN-76-fed mice. Interestingly, representative orders of the phylaProteobacteria,Bacteroidetes, andFirmicutes, including theLactobacillales, also differed as a function of murine ERβ status. Overall, these interactions suggest that sterone nucleoreceptor status and diet complexity may play important roles in microbiota maintenance. Furthermore, we envision that this model for gastrointestinal dysbiosis may be used to identify novel probiotics, prebiotics, nutritional strategies, and pharmaceuticals for the prevention and resolution ofProteobacteria-rich dysbiosis.
Intragenomic Heterogeneity of 16S rRNA Genes Causes Overestimation of Prokaryotic Diversity
Authors: Sun Dong-Lei , Jiang Xuan , Wu Qinglong L. , Zhou Ning-Yi
Published: July 2013

Ever since Carl Woese introduced the use of 16S rRNA genes for determining the phylogenetic relationships of prokaryotes, this method has been regarded as the “gold standard” in both microbial phylogeny and ecology studies. However, intragenomic heterogeneity within 16S rRNA genes has been reported in many investigations and is believed to bias the estimation of prokaryotic diversity. In the current study, 2,013 completely sequenced genomes of bacteria and archaea were analyzed and intragenomic heterogeneity was found in 952 genomes (585 species), with 87.5% of the divergence detected being below the 1% level. In particular, some extremophiles (thermophiles and halophiles) were found to harbor highly divergent 16S rRNA genes. Overestimation caused by 16S rRNA gene intragenomic heterogeneity was evaluated at different levels using the full-length and partial 16S rRNA genes usually chosen as targets for pyrosequencing. The result indicates that, at the unique level, full-length 16S rRNA genes can produce an overestimation of as much as 123.7%, while at the 3% level, an overestimation of 12.9% for the V6 region may be introduced. Further analysis showed that intragenomic heterogeneity tends to concentrate in specific positions, with the V1 and V6 regions suffering the most intragenomic heterogeneity and the V4 and V5 regions suffering the least intragenomic heterogeneity in bacteria. This is the most up-to-date overview of the diversity of 16S rRNA genes within prokaryotic genomes. It not only provides general guidance on how much overestimation can be introduced when applying 16S rRNA gene-based methods, due to its intragenomic heterogeneity, but also recommends that, for bacteria, this overestimation be minimized using primers targeting the V4 and V5 regions.
Combining Individual-Based Modeling and Food Microenvironment Descriptions To Predict the Growth of Listeria monocytogenes on Smear Soft Cheese
Authors: Ferrier Rachel , Hezard Bernard , Lintz Adrienne , Stahl Valérie , Augustin Jean-Christophe
Published: July 2013

An individual-based modeling (IBM) approach was developed to describe the behavior of a few
Listeria monocytogenes
cells contaminating smear soft cheese surface. The IBM approach consisted of assessing the stochastic individual behaviors of cells on cheese surfaces and knowing the characteristics of their surrounding microenvironments. We used a microelectrode for pH measurements and micro-osmolality to assess the water activity of cheese microsamples. These measurements revealed a high variability of microscale pH compared to that of macroscale pH. A model describing the increase in pH from approximately 5.0 to more than 7.0 during ripening was developed. The spatial variability of the cheese surface characterized by an increasing pH with radius and higher pH on crests compared to that of hollows on cheese rind was also modeled. The microscale water activity ranged from approximately 0.96 to 0.98 and was stable during ripening. The spatial variability on cheese surfaces was low compared to between-cheese variability. Models describing the microscale variability of cheese characteristics were combined with the IBM approach to simulate the stochastic growth of
L. monocytogenes
on cheese, and these simulations were compared to bacterial counts obtained from irradiated cheeses artificially contaminated at different ripening stages. The simulated variability of
L. monocytogenes
counts with the IBM/microenvironmental approach was consistent with the observed one. Contrasting situations corresponding to no growth or highly contaminated foods could be deduced from these models. Moreover, the IBM approach was more effective than the traditional population/macroenvironmental approach to describe the actual bacterial behavior variability.
Adhesion of Human and Animal Escherichia coli Strains in Association with Their Virulence-Associated Genes and Phylogenetic Origins
Authors: Frömmel Ulrike , Lehmann Werner , Rödiger Stefan , Böhm Alexander , Nitschke Jörg , Weinreich Jörg , Groß Julia , Roggenbuck Dirk , Zinke Olaf , Ansorge Hermann , Vogel Steffen , Klemm Per , Wex Thomas , Schröder Christian , Wieler Lothar H. , Schierack Peter
Published: July 2013

Intestinal colonization is influenced by the ability of the bacterium to inhabit a niche, which is based on the expression of colonization factors.
Escherichia coli
carries a broad range of virulence-associated genes (VAGs) which contribute to intestinal (inVAGs) and extraintestinal (exVAGs) infection. Moreover, initial evidence indicates that inVAGs and exVAGs support intestinal colonization. We developed new screening tools to genotypically and phenotypically characterize
E. coli
isolates originating in humans, domestic pigs, and 17 wild mammal and avian species. We analyzed 317 isolates for the occurrence of 44 VAGs using a novel multiplex PCR microbead assay (MPMA) and for adhesion to four epithelial cell lines using a new adhesion assay. We correlated data for the definition of new adhesion genes. inVAGs were identified only sporadically, particularly in roe deer (
Capreolus capreolus
) and the European hedgehog (
Erinaceus europaeus
). The prevalence of exVAGs depended on isolation from a specific host. Human uropathogenic
E. coli
isolates carried exVAGs with the highest prevalence, followed by badger (
Meles meles
) and roe deer isolates. Adhesion was found to be very diverse. Adhesion was specific to cells, host, and tissue, though it was also unspecific. Occurrence of the following VAGs was associated with a higher rate of adhesion to one or more cell lines:
afa-dra
,
daaD
,
tsh
,
vat
,
ibeA
,
fyuA
,
mat
,
sfa-foc
,
malX
,
pic
,
irp2
, and
papC
. In summary, we established new screening methods which enabled us to characterize large numbers of
E. coli
isolates. We defined reservoirs for potential pathogenic
E. coli
. We also identified a very broad range of colonization strategies and defined potential new adhesion genes.
Unique Contribution of the Cell Wall-Binding Endoglucanase G to the Cellulolytic Complex in Clostridium cellulovorans
Authors: Jeon Sang Duck , Lee Ji Eun , Kim Su Jung , Park Sung Hyun , Choi Gi-Wook , Han Sung Ok
Published: July 2013

The cellulosomes produced by
Clostridium cellulovorans
are organized by the specific interactions between the cohesins in the scaffolding proteins and the dockerins of the catalytic components. Using a cohesin biomarker, we identified a cellulosomal enzyme which belongs to the glycosyl hydrolase family 5 and has a domain of unknown function 291 (DUF291) with functions similar to those of the surface layer homology domain in
C. cellulovorans
. The purified endoglucanase G (EngG) had the highest synergistic degree with exoglucanase (ExgS) in the hydrolysis of crystalline cellulose (EngG/ExgS ratio = 3:1; 1.71-fold). To measure the binding affinity of the dockerins in EngG for the cohesins of the main scaffolding protein, a competitive enzyme-linked interaction assay was performed. Competitors, such as ExgS, reduced the percentage of EngG that were bound to the cohesins to less than 20%; the results demonstrated that the cohesins prefer to bind to the common cellulosomal enzymes rather than to EngG. Additionally, in surface plasmon resonance analysis, the dockerin in EngG had a relatively weak affinity (30- to 123-fold) for cohesins compared with the other cellulosomal enzymes. In the cell wall affinity assay, EngG anchored to the cell surfaces of
C. cellulovorans
using its DUF291 domain. Immunofluorescence microscopy confirmed the cell surface display of the EngG complex. These results indicated that in
C. cellulovorans
, EngG assemble into both the cellulolytic complex and the cell wall complex to aid in the hydrolysis of cellulose substrates.
The Carbohydrate Metabolism Signature of Lactococcus lactis Strain A12 Reveals Its Sourdough Ecosystem Origin
Authors: Passerini Delphine , Coddeville Michèle , Le Bourgeois Pascal , Loubière Pascal , Ritzenthaler Paul , Fontagné-Faucher Catherine , Daveran-Mingot Marie-Line , Cocaign-Bousquet Muriel
Published: July 2013

Lactococcus lactis
subsp.
lactis
strain A12 was isolated from sourdough. Combined genomic, transcriptomic, and phenotypic analyses were performed to understand its survival capacity in the complex sourdough ecosystem and its role in the microbial community. The genome sequence comparison of strain A12 with strain IL1403 (a derivative of an industrial dairy strain) revealed 78 strain-specific regions representing 23% of the total genome size. Most of the strain-specific genes were involved in carbohydrate metabolism and are potentially required for its persistence in sourdough. Phenotype microarray, growth tests, and analysis of glycoside hydrolase content showed that strain A12 fermented plant-derived carbohydrates, such as arabinose and α-galactosides. Strain A12 exhibited specific growth rates on raffinose that were as high as they were on glucose and was able to release sucrose and galactose outside the cell, providing soluble carbohydrates for sourdough microflora. Transcriptomic analysis identified genes specifically induced during growth on raffinose and arabinose and reveals an alternative pathway for raffinose assimilation to that used by other lactococci.
Utilization of Heme as an Iron Source by Marine Alphaproteobacteria in the Roseobacter Clade
Authors: Roe Kelly L. , Hogle Shane L. , Barbeau Katherine A.
Published: July 2013

The bioavailability and utilization of porphyrin-bound iron, specifically heme, by marine microorganisms have rarely been examined. This study used
Ruegeria
sp. strain TrichCH4B as a model organism to study heme acquisition by a member of the
Roseobacter
clade. Analogs of known heme transporter proteins were found within the
Ruegeria
sp. TrichCH4B genome. The identified heme uptake and utilization system appears to be functional, as the heme genes were upregulated under iron stress, the bacterium could grow on ferric-porphyrin complexes as the sole iron source, and internalization of
55
Fe from ferric protoporphyrin IX was observed. The potential ability to utilize heme in the
Roseobacter
clade appears to be common, as half of the isolates in the RoseoBase database were found to have a complete heme uptake system. A degenerate primer set was designed and successfully used to identify the putative heme oxygenase gene (
hmus
) in the roseobacter heme uptake system from diverse nonenriched marine environments. This study found that members of the
Roseobacter
clade are capable of utilizing heme as an iron source and that this capability may be present in all types of marine environments. The results of this study add a new perspective to the current picture of iron cycling in marine systems, whereby relatively refractory intracellular pools of heme-bound iron may be taken up quickly and directly reincorporated into living bacteria without previous degradation or the necessity of a siderophore intermediate.
Detoxification of Mercury by Methanobactin from Methylosinus trichosporium OB3b
Authors: Vorobev Alexey , Jagadevan Sheeja , Baral Bipin S. , DiSpirito Alan A. , Freemeier Brittani C. , Bergman Brandt H. , Bandow Nathan L. , Semrau Jeremy D.
Published: July 2013

Many methanotrophs have been shown to synthesize methanobactin, a novel biogenic copper-chelating agent or chalkophore. Methanobactin binds copper via two heterocyclic rings with associated enethiol groups. The structure of methanobactin suggests that it can bind other metals, including mercury. Here we report that methanobactin from
Methylosinus trichosporium
OB3b does indeed bind mercury when added as HgCl
2
and, in doing so, reduced toxicity associated with Hg(II) for both
Alphaproteobacteria
methanotrophs, including
M. trichosporium
OB3b,
M. trichosporium
OB3b Δ
mbnA
(a mutant defective in methanobactin production), and
Methylocystis
sp. strain SB2, and a
Gammaproteobacteria
methanotroph,
Methylomicrobium album
BG8. Mercury binding by methanobactin was evident in both the presence and absence of copper, despite the fact that methanobactin had a much higher affinity for copper due to the rapid and irreversible binding of mercury by methanobactin. The formation of a gray precipitate suggested that Hg(II), after being bound by methanobactin, was reduced to Hg(0) but was not volatilized. Rather, mercury remained associated with methanobactin and was also found associated with methanotrophic biomass. It thus appears that although the mercury-methanobactin complex was cell associated, mercury was not removed from methanobactin. The amount of biomass-associated mercury in the presence of methanobactin from
M. trichosporium
OB3b was greatest for
M. trichosporium
wild-type strain OB3b and the Δ
mbnA
mutant and least for
M. album
BG8, suggesting that methanotrophs may have selective methanobactin uptake systems that may be based on TonB-dependent transporters but that such uptake systems exhibit a degree of infidelity.
Abundant DNase I-Sensitive Bacterial DNA in Healthy Porcine Lungs and Its Implications for the Lung Microbiome
Authors: Pezzulo Alejandro A. , Kelly Patrick H. , Nassar Boulos S. , Rutland Cedric J. , Gansemer Nicholas D. , Dohrn Cassie L. , Costello Andrew J. , Stoltz David A. , Zabner Joseph
Published: July 2013

Human lungs are constantly exposed to bacteria in the environment, yet the prevailing dogma is that healthy lungs are sterile. DNA sequencing-based studies of pulmonary bacterial diversity challenge this notion. However, DNA-based microbial analysis currently fails to distinguish between DNA from live bacteria and that from bacteria that have been killed by lung immune mechanisms, potentially causing overestimation of bacterial abundance and diversity. We investigated whether bacterial DNA recovered from lungs represents live or dead bacteria in bronchoalveolar lavage (BAL) fluid and lung samples in young healthy pigs. Live bacterial DNA was DNase I resistant and became DNase I sensitive upon human antimicrobial-mediated killingin vitro. We determined live and total bacterial DNA loads in porcine BAL fluid and lung tissue by comparing DNase I-treated versus untreated samples. In contrast to the case for BAL fluid, we were unable to culture bacteria from most lung homogenates. Surprisingly, total bacterial DNA was abundant in both BAL fluid and lung homogenates. In BAL fluid, 63% was DNase I sensitive. In 6 out of 11 lung homogenates, all bacterial DNA was DNase I sensitive, suggesting a predominance of dead bacteria; in the remaining homogenates, 94% was DNase I sensitive, and bacterial diversity determined by 16S rRNA gene sequencing was similar in DNase I-treated and untreated samples. Healthy pig lungs are mostly sterile yet contain abundant DNase I-sensitive DNA from inhaled and aspirated bacteria killed by pulmonary host defense mechanisms. This approach and conceptual framework will improve analysis of the lung microbiome in disease.
Enantioselective Dehydrochlorination of δ-Hexachlorocyclohexane and δ-Pentachlorocyclohexene by LinA1 and LinA2 from Sphingobium indicum B90A
Authors: Geueke Birgit , Miska Milena E. , Poiger Thomas , Rentsch Daniel , Lal Rup , Holliger Christof , Kohler Hans-Peter E.
Published: July 2013

δ-Hexachlorocyclohexane (δ-HCH), one of the prevalent isomers of technical HCH, was enantioselectively dehydrochlorinated by the dehydrochlorinases LinA1 and LinA2 from
Sphingobium indicum
B90A to the very same δ-pentachlorocyclohexene enantiomer. Racemic δ-pentachlorocyclohexene, however, was transformed with opposite enantioselectivities by the two enzymes. A transformation pathway based on an
anti
-1,2-elimination, followed by a
syn
-1,4-elimination and a subsequent
syn
-1,2-elimination is postulated.
Genetic Characterization of Atypical Enteropathogenic Escherichia coli Isolates from Ewes' Milk, Sheep Farm Environments, and Humans by Multilocus Sequence Typing and Pulsed-Field Gel Electrophoresis
Authors: Otero Verónica , Rodríguez-Calleja José-María , Otero Andrés , García-López María-Luisa , Santos Jesús A.
Published: July 2013

A collection of 81 isolates of enteropathogenic
Escherichia coli
(EPEC) was obtained from samples of bulk tank sheep milk (62 isolates), ovine feces (4 isolates), sheep farm environment (water, 4 isolates; air, 1 isolate), and human stool samples (9 isolates). The strains were considered atypical EPEC organisms, carrying the
eae
gene without harboring the pEAF plasmid. Multilocus sequence typing (MLST) was carried out with seven housekeeping genes and 19 sequence types (ST) were detected, with none of them having been previously reported for atypical EPEC. The most frequent ST included 41 strains isolated from milk and human stool samples. Genetic typing by pulsed-field gel electrophoresis (PFGE) resulted in 57 patterns which grouped in 24 clusters. Comparison of strains isolated from the different samples showed phylogenetic relationships between milk and human isolates and also between milk and water isolates. The results obtained show a possible risk for humans due to the presence of atypical EPEC in ewes' milk and suggest a transmission route for this emerging pathogen through contaminated water.
Alternative Sigma Factor σ
E
Has an Important Role in Stress Tolerance of Yersinia pseudotuberculosis IP32953
Authors: Palonen Eveliina , Lindström Miia , Somervuo Panu , Korkeala Hannu
Published: July 2013

Yersinia pseudotuberculosis
is an important pathogen that probably survives well in the modern food chain. However, little is known about the mechanisms that allow the growth of this pathogen in foods under stress conditions. The expression of
rpoE
encoding σ
E
was defined by quantitative real-time reverse transcription-PCR. Expression of
rpoE
was induced at 3°C, 37°C, and 42°C, under exposure to 3% NaCl, 3% ethanol, or high and low pH, in relation to its expression at the optimum growth temperature of 28°C of
Y. pseudotuberculosis
. Mutation of
rpoE
either impaired or abolished growth under stresses caused by low or high temperature, low pH, and ethanol. In addition, the growth temperature range of the mutant was significantly diminished compared to that of the wild-type strain IP32953. The results were confirmed with complementation of the mutant. Thus, σ
E
plays a significant role in the stress tolerance of
Y. pseudotuberculosis
IP32953 and probably contributes to the survival of this pathogen in the food chain.
Extending the Cellulosome Paradigm: the Modular Clostridium thermocellum Cellulosomal Serpin PinA Is a Broad-Spectrum Inhibitor of Subtilisin-Like Proteases
Authors: Ó Cuív Páraic , Gupta Rajesh , Goswami Hareshwar P. , Morrison Mark
Published: July 2013

Clostridium thermocellum
encodes a cellulosomal, modular, and thermostable serine protease inhibitor (serpin), PinA. PinA stability but not inhibitory activity is affected by the Fn(III) and Doc(I) domains, and PinA is a broad inhibitor of subtilisin-like proteases and may play a key role in protecting the cellulosome from protease attack.
Clostridium acidurici Electron-Bifurcating Formate Dehydrogenase
Authors: Wang Shuning , Huang Haiyan , Kahnt Jörg , Thauer Rudolf K.
Published: July 2013

Cell extracts of uric acid-grown
Clostridium acidurici
catalyzed the coupled reduction of NAD
+
and ferredoxin with formate at a specific activity of 1.3 U/mg. The enzyme complex catalyzing the electron-bifurcating reaction was purified 130-fold and found to be composed of four subunits encoded by the gene cluster
hylCBA-fdhF2
.
Characterization of AmiBA2446, a Novel Bacteriolytic Enzyme Active against Bacillus Species
Authors: Mehta Krunal K. , Paskaleva Elena E. , Azizi-Ghannad Saba , Ley Daniel J. , Page Martin A. , Dordick Jonathan S. , Kane Ravi S.
Published: July 2013

There continues to be a need for developing efficient and environmentally friendly treatments for
Bacillus anthracis
, the causative agent of anthrax. One emerging approach for inactivation of vegetative
B. anthracis
is the use of bacteriophage endolysins or lytic enzymes encoded by bacterial genomes (autolysins) with highly evolved specificity toward bacterium-specific peptidoglycan cell walls. In this work, we performed
in silico
analysis of the genome of
Bacillus anthracis
strain Ames, using a consensus binding domain amino acid sequence as a probe, and identified a novel lytic enzyme that we termed AmiBA2446. This enzyme exists as a homodimer, as determined by size exclusion studies. It possesses
N
-acetylmuramoyl-
l
-alanine amidase activity, as determined from liquid chromatography-mass spectrometry (LC-MS) analysis of muropeptides released due to the enzymatic digestion of peptidoglycan. Phylogenetic analysis suggested that AmiBA2446 was an autolysin of bacterial origin. We characterized the effects of enzyme concentration and phase of bacterial growth on bactericidal activity and observed close to a 5-log reduction in the viability of cells of
Bacillus cereus
4342, a surrogate for
B. anthracis
. We further tested the bactericidal activity of AmiBA2446 against various
Bacillus
species and demonstrated significant activity against
B. anthracis
and
B. cereus
strains. We also demonstrated activity against
B. anthracis
spores after pretreatment with germinants. AmiBA2446 enzyme was also stable in solution, retaining its activity after 4 months of storage at room temperature.
Bacterial Community Response to Petroleum Hydrocarbon Amendments in Freshwater, Marine, and Hypersaline Water-Containing Microcosms
Authors: Jurelevicius Diogo , Alvarez Vanessa Marques , Marques Joana Montezano , de Sousa Lima Laryssa Ribeiro Fonseca , Dias Felipe de Almeida , Seldin Lucy
Published: July 2013

Hydrocarbon-degrading bacterial communities from freshwater, marine, and hypersaline Brazilian aquatic ecosystems (with water salinities corresponding to 0.2%, 4%, and 5%, respectively) were enriched with different hydrocarbons (heptadecane, naphthalene, or crude oil). Changes within the different microcosms of bacterial communities were analyzed using cultivation approaches and molecular methods (DNA and RNA extraction, followed by genetic fingerprinting and analyses of clone libraries based on the 16S rRNA-coding gene). A redundancy analysis (RDA) of the genetic fingerprint data and a principal component analysis (PCA) of the clone libraries revealed hydrocarbon-enriched bacterial communities specific for each ecosystem studied. However, within the same ecosystem, different bacterial communities were selected according to the petroleum hydrocarbon used. In general, the results demonstrated that
Acinetobacter
and
Cloacibacterium
were the dominant genera in freshwater microcosms; the
Oceanospirillales
order and the
Marinobacter
,
Pseudomonas
, and
Cycloclasticus
genera predominated in marine microcosms; and the
Oceanospirillales
order and the
Marinobacter
genus were selected in the different hydrocarbon-containing microcosms in hypersaline water. Determination of total petroleum hydrocarbons (TPHs) in all microcosms after 32 days of incubation showed a decrease in the hydrocarbon concentration compared to that for the controls. A total of 50 (41.3%) isolates from the different hydrocarbon-contaminated microcosms were associated with the dominant operational taxonomic units (OTUs) obtained from the clone libraries, and their growth in the hydrocarbon contaminating the microcosm from which they were isolated as the sole carbon source was observed. These data provide insight into the general response of bacterial communities from freshwater, marine, and hypersaline aquatic ecosystems to petroleum hydrocarbon contamination.
Comparative Proteomic Analysis of Streptomyces lividans Wild-Type andppkMutant Strains Reveals the Importance of Storage Lipids for Antibiotic Biosynthesis
Authors: Le Maréchal Pierre , Decottignies Paulette , Marchand Christophe H. , Degrouard Jeril , Jaillard Danièle , Dulermo Thierry , Froissard Marine , Smirnov Aleksey , Chapuis Violaine , Virolle Marie-Joelle
Published: July 2013

Streptomyces lividansTK24 is a strain that naturally produces antibiotics at low levels, but dramatic overproduction of antibiotics occurs upon interruption of theppkgene. However, the role of the Ppk enzyme in relation to the regulation of antibiotic biosynthesis remains poorly understood. In order to gain a better understanding of the phenotype of theppkmutant, the proteomes of the wild-type (wt) andppkmutant strains, grown for 96 h on R2YE medium limited in phosphate, were analyzed. Intracellular proteins were separated on two-dimensional (2D) gels, spots were quantified, and those showing a 3-fold variation or more were identified by mass spectrometry. The expression of 12 proteins increased and that of 29 decreased in theppkmutant strain. Our results suggested that storage lipid degradation rather than hexose catabolism was taking place in the mutant. In order to validate this hypothesis, the triacylglycerol contents of the wt andppkmutant strains ofS. lividansas well as that ofStreptomyces coelicolorM145, a strain that produces antibiotics at high levels and is closely related toS. lividans, were assessed using electron microscopy and thin-layer chromatography. These studies highlighted the large difference in triacylglycerol contents of the three strains and confirmed the hypothetical link between storage lipid metabolism and antibiotic biosynthesis inStreptomyces.
Physiological Role of Acyl Coenzyme A Synthetase Homologs in Lipid Metabolism in Neurospora crassa
Authors: Roche Christine M. , Blanch Harvey W. , Clark Douglas S. , Glass N. Louise
Published: July 2013

Acyl coenzyme A (CoA) synthetase (ACS) enzymes catalyze the activation of free fatty acids (FAs) to CoA esters by a two-step thioesterification reaction. Activated FAs participate in a variety of anabolic and catabolic lipid metabolic pathways, including
de novo
complex lipid biosynthesis, FA β-oxidation, and lipid membrane remodeling. Analysis of the genome sequence of the filamentous fungus
Neurospora crassa
identified seven putative fatty ACSs (ACS-1 through ACS-7). ACS-3 was found to be the major activator for exogenous FAs for anabolic lipid metabolic pathways, and consistent with this finding, ACS-3 localized to the endoplasmic reticulum, plasma membrane, and septa. Double-mutant analyses confirmed partial functional redundancy of ACS-2 and ACS-3. ACS-5 was determined to function in siderophore biosynthesis, indicating alternative functions for ACS enzymes in addition to fatty acid metabolism. The
N. crassa
ACSs involved in activation of FAs for catabolism were not specifically defined, presumably due to functional redundancy of several of ACSs for catabolism of exogenous FAs.
Novel Thioredoxin-Like Proteins Are Components of a Protein Complex Coating the Cortical Microtubules of Toxoplasma gondii
Authors: Liu Jun , Wetzel Laura , Zhang Ying , Nagayasu Eiji , Ems-McClung Stephanie , Florens Laurence , Hu Ke
Published: July 2013

Microtubules are versatile biopolymers that support numerous vital cellular functions in eukaryotes. The specific properties of microtubules are dependent on distinct microtubule-associated proteins, as the tubulin subunits and microtubule structure are exceptionally conserved. Highly specialized microtubule-containing assemblies are often found in protists, which are rich sources for novel microtubule-associated proteins. A protozoan parasite,Toxoplasma gondii, possesses several distinct tubulin-containing structures, including 22 microtubules closely associated with the cortical membrane. Early ultrastructural studies have shown that the cortical microtubules are heavily decorated with associating proteins. However, little is known about the identities of these proteins. Here, we report the discovery of a novel protein, TrxL1 (forThioredoxin-Like protein 1), and an associating complex that coats the cortical microtubules. TrxL1 contains a thioredoxin-like fold. To visualize its localization in live parasites by fluorescence, we replaced the endogenous TrxL1 gene with an mEmeraldFP-TrxL1 fusion gene. Structured illumination-based superresolution imaging of this parasite line produced a detailed view of the microtubule cytoskeleton. Despite its stable association with the cortical microtubules in the parasite, TrxL1 does not seem to bind to microtubules directly. Coimmunoprecipitation experiments showed that TrxL1 associates with a protein complex containing SPM1, a previously reported microtubule-associated protein inT. gondii. We also found that SPM1 recruits TrxL1 to the cortical microtubules. Besides SPM1, several other novel proteins are found in the TrxL1-containing complex, including TrxL2, a close homolog of TrxL1. Thus, our results reveal for the first time a microtubule-associated complex inT. gondii.
Synchronous Activation of Cell Division by Light or Temperature Stimuli in the Dimorphic Yeast Schizosaccharomyces japonicus
Authors: Okamoto Sho , Furuya Kanji , Nozaki Shingo , Aoki Keita , Niki Hironori
Published: July 2013

Many fungi respond to light and regulate fungal development and behavior. A blue light-activated complex has been identified in
Neurospora crassa
as the product of the
wc-1
and
wc-2
genes. Orthologs of WC-1 and WC-2 have hitherto been found only in filamentous fungi and not in yeast, with the exception of the basidiomycete pathogenic yeast
Cryptococcus
. Here, we report that the fission yeast
Schizosaccharomyces japonicus
responds to blue light depending on Wcs1 and Wcs2, orthologs of components of the WC complex. Surprisingly, those of ascomycete
S. japonicus
are more closely related to those of the basidiomycete.
S. japonicus
reversibly changes from yeast to hyphae in response to environmental stresses. After incubation at 30°C, a colony of yeast was formed, and then hyphal cells extended from the periphery of the colony. When light cycles were applied, distinct dark- and bright-colored hyphal cell stripes were formed because the growing hyphal cells had synchronously activated cytokinesis. In addition, temperature cycles of 30°C for 12 h and 35°C for 12 h or of 25°C for 12 h and 30°C for 12 h during incubation in the dark induced a response in the hyphal cells similar to that of light. The stripe formation of the temperature cycles was independent of the
wcs
genes. Both light and temperature, which are daily external cues, have the same effect on growing hyphal cells. A dual sensing mechanism of external cues allows organisms to adapt to daily changes of environmental alteration.
Independent Localization of Plasma Membrane and Chloroplast Components during Eyespot Assembly
Authors: Mittelmeier Telsa M. , Thompson Mark D. , Öztürk Esra , Dieckmann Carol L.
Published: July 2013

Like many algae,Chlamydomonas reinhardtiiis phototactic, using two anterior flagella to swim toward light optimal for photosynthesis. The flagella are responsive to signals initiated at the photosensory eyespot, which comprises photoreceptors in the plasma membrane and layers of pigment granules in the chloroplast. Phototaxis depends on placement of the eyespot at a specific asymmetric location relative to the flagella, basal bodies, and bundles of two or four highly acetylated microtubules, termed rootlets, which extend from the basal bodies toward the posterior of the cell. Previous work has shown that the eyespot is disassembled prior to cell division, and new eyespots are assembled in daughter cells adjacent to the nascent four-membered rootlet associated with the daughter basal body (D4), but the chronology of these assembly events has not been determined. Here we use immunofluorescence microscopy to follow assembly and acetylation of the D4 rootlet, localization of individual eyespot components in the plasma membrane or chloroplast envelope, and flagellar emergence during and immediately following cell division. We find that the D4 rootlet is assembled before the initiation of eyespot assembly, which occurs within the same time frame as rootlet acetylation and flagellar outgrowth. Photoreceptors in the plasma membrane are correctly localized in eyespot mutant cells lacking pigment granule layers, and chloroplast components of the eyespot assemble in mutant cells in which photoreceptor localization is retarded. The data suggest that plasma membrane and chloroplast components of the eyespot are independently responsive to a cytoskeletal positioning cue.
Protein Kinase CK2 Holoenzyme Promotes Start-Specific Transcription in Saccharomyces cerevisiae
Authors: Tripodi Farida , Nicastro Raffaele , Busnelli Sara , Cirulli Claudia , Maffioli Elisa , Tedeschi Gabriella , Alberghina Lilia , Coccetti Paola
Published: July 2013

In
Saccharomyces cerevisiae
, the entrance into S phase requires the activation of a specific burst of transcription, which depends on SBF (SCB binding factor, Swi4/Swi6) and MBF (MCB binding factor, Mbp1/Swi6) complexes. CK2 is a pleiotropic kinase involved in several cellular processes, including the regulation of the cell cycle. CK2 is composed of two catalytic subunits (α and α′) and two regulatory subunits (β and β′), both of which are required to form the active holoenzyme. Here we investigate the function of the CK2 holoenzyme in Start-specific transcription. The
ckb1
Δ
ckb2
Δ mutant strain, bearing deletions of both genes encoding CK2 regulatory subunits, shows a delay of S-phase entrance due to a severe reduction of the expression of SBF- and MBF-dependent genes. This transcriptional defect is caused by an impaired recruitment of Swi6 and Swi4 to G
1
gene promoters. Moreover, CK2 α and β′ subunits interact with RNA polymerase II, whose binding to G
1
promoters is positively regulated by the CK2 holoenzyme. Collectively, these findings suggest a novel role for the CK2 holoenzyme in the activation of G
1
transcription.
Candida albicans Czf1 and Efg1 Coordinate the Response to Farnesol during Quorum Sensing, White-Opaque Thermal Dimorphism, and Cell Death
Authors: Langford Melanie L. , Hargarten Jessica C. , Patefield Krista D. , Marta Elizabeth , Blankenship Jill R. , Fanning Saranna , Nickerson Kenneth W. , Atkin Audrey L.
Published: July 2013

Quorum sensing by farnesol in
Candida albicans
inhibits filamentation and may be directly related to its ability to cause both mucosal and systemic diseases. The Ras1-cyclic AMP signaling pathway is a target for farnesol inhibition. However, a clear understanding of the downstream effectors of the morphological farnesol response has yet to be unraveled. To address this issue, we screened a library for mutants that fail to respond to farnesol. Six mutants were identified, and the
czf1
Δ/
czf1
Δ mutant was selected for further characterization. Czf1 is a transcription factor that regulates filamentation in embedded agar and also white-to-opaque switching. We found that Czf1 is required for filament inhibition by farnesol under at least three distinct environmental conditions: on agar surfaces, in liquid medium, and when embedded in a semisolid agar matrix. Since Efg1 is a transcription factor of the Ras1-cyclic AMP signaling pathway that interacts with and regulates Czf1, an
efg1
Δ/
efg1
Δ
czf1
Δ/
czf1
Δ mutant was tested for filament inhibition by farnesol. It exhibited an opaque-cell-like temperature-dependent morphology, and it was killed by low farnesol levels that are sublethal to wild-type cells and both
efg1
Δ/
efg1
Δ and
czf1
Δ/
czf1
Δ single mutants. These results highlight a new role for Czf1 as a downstream effector of the morphological response to farnesol, and along with Efg1, Czf1 is involved in the control of farnesol-mediated cell death in
C. albicans
.
Physiological Consequences of Multiple-Target Regulation by the Small RNA SgrS in Escherichia coli
Authors: Sun Yan , Vanderpool Carin K.
Published: July 2013

Cells use complex mechanisms to regulate glucose transport and metabolism to achieve optimal energy and biomass production while avoiding accumulation of toxic metabolites. Glucose transport and glycolytic metabolism carry the risk of the buildup of phosphosugars, which can inhibit growth at high concentrations. Many enteric bacteria cope with phosphosugar accumulation and associated stress (i.e., sugar-phosphate stress) by producing a small RNA (sRNA) regulator, SgrS, which decreases phosphosugar accumulation in part by repressing translation of sugar transporter mRNAs (
ptsG
and
manXYZ
) and enhancing translation of a sugar phosphatase mRNA (
yigL
). Despite a molecular understanding of individual target regulation by SgrS, previously little was known about how coordinated regulation of these multiple targets contributes to the rescue of cell growth during sugar-phosphate stress. This study examines how SgrS regulation of different targets impacts growth under different nutritional conditions when sugar-phosphate stress is induced. The severity of stress-associated growth inhibition depended on nutrient availability. Stress in nutrient-rich media necessitated SgrS regulation of only sugar transporter mRNAs (
ptsG
or
manXYZ
). However, repression of transporter mRNAs was insufficient for growth rescue during stress in nutrient-poor media; here SgrS regulation of the phosphatase (
yigL
) and as-yet-undefined targets also contributed to growth rescue. The results of this study imply that regulation of only a subset of an sRNA's targets may be important in a given environment. Further, the results suggest that SgrS and perhaps other sRNAs are flexible regulators that modulate expression of multigene regulons to allow cells to adapt to an array of stress conditions.
A review of causes of Stevens–Johnson syndrome and toxic epidermal necrolysis in children
Authors: Ferrandiz-Pulido Carla , Garcia-Patos Vicente
Published: July 2013

Stevens–Johnson syndrome (SJS) and toxic epidermal necrolysis (TEN) are rare life-threatening conditions almost exclusively attributed to drugs. The incidence in children is lower than in adults and has a better outcome.Mycosplama pneumoniaeinfection may be involved in some cases of paediatric SJS. The main etiologic factors for both SSJ and TEN are sulphonamides and anticonvulsants, followed by penicillins and non-steroidal anti-inflammatory drugs. In rare instances, paracetamol is the only suspected drug. By contrast with adults, allopurinol, oxicams and nevirapine are not identified as causative agents in children, probably due to differences in drug prescriptions. The only aspects of treatment that have been proved to improve survival are the rapid withdrawal of the suspected offending drugs and an optimal supportive therapy with emphasis in nutritional support, accompanied by management of denuded skin areas. The use of specific therapies remains controversial.
Association of work-related stress with mental health problems in a special police force unit
Authors: Garbarino Sergio , Cuomo Giovanni , Chiorri Carlo , Magnavita Nicola
Published: July 2013

Objectives

Law and order enforcement tasks may expose special force police officers to significant psychosocial risk factors. The aim of this work is to investigate the relationship between job stress and the presence of mental health symptoms while controlling sociodemographical, occupational and personality variables in special force police officers.

Method

At different time points, 292 of 294 members of the ‘VI Reparto Mobile’, a special police force engaged exclusively in the enforcement of law and order, responded to our invitation to complete questionnaires for the assessment of personality traits, work-related stress (using the Demand–Control–Support (DCS) and the Effort–Reward–Imbalance (ERI) models) and mental health problems such as depression, anxiety and burnout.

Results

Regression analyses showed that lower levels of support and reward and higher levels of effort and overcommitment were associated with higher levels of mental health symptoms. Psychological screening revealed 21 (7.3%) likely cases of mild depression (Beck Depression Inventory, BDI≥10). Officers who had experienced a discrepancy between work effort and rewards showed a marked increase in the risk of depression (OR 7.89, 95% CI 2.32 to 26.82) when compared with their counterparts who did not perceive themselves to be in a condition of distress.

Conclusions

The findings of this study suggest that work-related stress may play a role in the development of mental health problems in police officers. The prevalence of mental health symptoms in the cohort investigated here was low, but not negligible in the case of depression. Since special forces police officers have to perform sensitive tasks for which a healthy psychological functioning is needed, the results of this study suggest that steps should be taken to prevent distress and improve the mental well-being of these workers.
Belimumab: a technological advance for systemic lupus erythematosus patients? Report of a systematic review and meta-analysis
Authors: Kandala Ngianga-Bakwin , Connock Martin , Grove Amy , Sutcliffe Paul , Mohiuddin Syed , Hartley Louise , Court Rachel , Cummins Ewen , Gordon Caroline , Clarke Aileen
Published: July 2013

Objectives

To undertake a systematic review and meta-analysis to investigate clinical effectiveness of belimumab for patients with systemic lupus erythematosus (SLE) and antinuclear and/or anti-double-stranded DNA (dsDNA) autoantibodies.

Methods

We searched eight electronic databases and reference lists for randomised controlled trials (RCTs) of belimumab against placebo or best supportive care. Quality assessment and random effects meta-analysis were undertaken.

Design

A meta-analysis of RCTs.

Participants

2133 SLE patients.

Primary and secondary outcome measures

SLE Responder Index (SRI) at week 52.

Results

Three double-blind placebo-controlled RCTs (L02, BLISS-52 BLISS-76) investigated 2133 SLE patients. BLISS-52 and BLISS-76 trials recruited patients with antinuclear and/or anti-dsDNA autoantibodies and demonstrated belimumab effectiveness for the SRI at week 52. Ethnicity and geographical location of participants varied considerably between BLISS trials. Although tests for statistical heterogeneity were negative, BLISS-52 results were systematically more favourable for all measured outcomes. Meta-analysis of pooled 52-week SRI BLISS results showed benefit for belimumab (OR 1.63, 95% CI 1.27 to 2.09). By week 76, the primary SRI outcome in BLISS-76 was not statistically significant (OR 1.31, 95% CI 0.919 to 1.855).