bluuebunny/crossref_metadata_2025_split huggingface dataset:
Laboratory Evidence of Norwalk Virus Contamination on the Hands of Infected Individuals
Authors: Liu Pengbo , Escudero Blanca , Jaykus Lee-Ann , Montes Julia , Goulter Rebecca M. , Lichtenstein Meredith , Fernandez Marina , Lee Joong-Chul , De Nardo Elizabeth , Kirby Amy , Arbogast James W. , Moe Christine L. 
Published: October 2013

ABSTRACT

Human norovirus (NoV) outbreak investigations suggest that the hands of infected individuals play an important role in NoV transmission. However, there is no experimental evidence documenting the likelihood and degree of NoV contamination on hands. As part of a clinical trial designed to evaluate the efficacy of high-pressure processing for Norwalk virus (NV) inactivation in oysters, 159 hand rinse samples were collected from 6 infected and 6 uninfected subjects. NV was concentrated from the samples by polyethylene glycol precipitation, followed by RNA extraction using an automated guanidinium isothiocyanate-silica method. NV RNA was detected and quantified using multiple NV-specific reverse transcription-quantitative PCR (RT-qPCR) assays. A total of 25.4% (18/71) of the hand rinse samples collected from 6 infected volunteers were presumptively positive for NV, with an average of 3.86 log
10
genomic equivalent copies (GEC) per hand. Dot blot hybridization of PCR products obtained using a different primer set, and DNA sequencing of selected amplicons, provided further confirmation of the presence of NV in the hand rinses. NV contamination was also detected in two hand rinse samples obtained from one uninfected subject. These findings provide definitive evidence of NV contamination on the hands of infected subjects observed under controlled clinical research conditions. Such data support the need for better hand hygiene strategies to prevent NoV transmission.
Aspergillus Collagen-Like Genes (
acl
): Identification, Sequence Polymorphism, and Assessment for PCR-Based Pathogen Detection
Authors: Tuntevski Kiril , Durney Brandon C. , Snyder Anna K. , LaSala P. Rocco , Nayak Ajay P. , Green Brett J. , Beezhold Donald H. , Rio Rita V. M. , Holland Lisa A. , Lukomski Slawomir 
Published: October 2013

ABSTRACT

The genus
Aspergillus
is a burden to public health due to its ubiquitous presence in the environment, its production of allergens, and wide demographic susceptibility among cystic fibrosis, asthmatic, and immunosuppressed patients. Current methods of detection of
Aspergillus
colonization and infection rely on lengthy morphological characterization or nonstandardized serological assays that are restricted to identifying a fungal etiology. Collagen-like genes have been shown to exhibit species-specific conservation across the noncollagenous regions as well as strain-specific polymorphism in the collagen-like regions. Here we assess the conserved region of the
Aspergillus
collagen-like (
acl
) genes and explore the application of PCR amplicon size-based discrimination among the five most common etiologic species of the
Aspergillus
genus, including
Aspergillus fumigatus
,
A. flavus
,
A. nidulans
,
A. niger
, and
A. terreus
. Genetic polymorphism and phylogenetic analysis of the
aclF1
gene were additionally examined among the available strains. Furthermore, the applicability of the PCR-based assay to identification of these five species in cultures derived from sputum and bronchoalveolar fluid from 19 clinical samples was explored. Application of capillary electrophoresis on nanogels was additionally demonstrated to improve the discrimination between
Aspergillus
species. Overall, this study demonstrated that
Aspergillus
acl
genes could be used as PCR targets to discriminate between clinically relevant
Aspergillus
species. Future studies aim to utilize the detection of
Aspergillus
acl
genes in PCR and microfluidic applications to determine the sensitivity and specificity for the identification of
Aspergillus
colonization and invasive aspergillosis in immunocompromised subjects.
SdrA, a New DeoR Family Regulator Involved in Streptomyces avermitilis Morphological Development and Antibiotic Production
Authors: Ulanova Dana , Kitani Shigeru , Fukusaki Eiichiro , Nihira Takuya 
Published: October 2013

ABSTRACT

The SAV3339 (SdrA) protein of
Streptomyces avermitilis
, a member of the DeoR family of regulators, was assessed to determine its
in vivo
function by gene knockdown through the use of
cis
-encoded noncoding RNA and knockout of the
sdrA
gene. These analyses revealed that SdrA represents another class of
Streptomyces
regulator that controls morphological development and antibiotic production.
Dimethyl Adenosine Transferase (KsgA) Deficiency in Salmonella enterica Serovar Enteritidis Confers Susceptibility to High Osmolarity and Virulence Attenuation in Chickens
Authors: Chiok Kim Lam , Addwebi Tarek , Guard Jean , Shah Devendra H. 
Published: October 2013

ABSTRACT

Dimethyl adenosine transferase (KsgA) performs diverse roles in bacteria, including ribosomal maturation and DNA mismatch repair, and synthesis of KsgA is responsive to antibiotics and cold temperature. We previously showed that a
ksgA
mutation in
Salmonella enterica
serovar Enteritidis results in impaired invasiveness in human and avian epithelial cells. In this study, we tested the virulence of a
ksgA
mutant (the
ksgA
::Tn
5
mutant) of
S
. Enteritidis in orally challenged 1-day-old chickens. The
ksgA
::Tn
5
mutant showed significantly reduced intestinal colonization and organ invasiveness in chickens compared to those of the wild-type (WT) parent. Phenotype microarray (PM) was employed to compare the
ksgA
::Tn
5
mutant and its isogenic wild-type strain for 920 phenotypes at 28°C, 37°C, and 42°C. At chicken body temperature (42°C), the
ksgA
::Tn
5
mutant showed significantly reduced respiratory activity with respect to a number of carbon, nitrogen, phosphate, sulfur, and peptide nitrogen nutrients. The greatest differences were observed in the osmolyte panel at concentrations of ≥6% NaCl at 37°C and 42°C. In contrast, no major differences were observed at 28°C. In independent growth assays, the
ksgA
::Tn
5
mutant displayed a severe growth defect in high-osmolarity (6.5% NaCl) conditions in nutrient-rich (LB) and nutrient-limiting (M9 minimum salts) media at 42°C. Moreover, the
ksgA
::Tn
5
mutant showed significantly reduced tolerance to oxidative stress, but its survival within macrophages was not impaired. Unlike
Escherichia coli
, the
ksgA
::Tn
5
mutant did not display a cold-sensitivity phenotype; however, it showed resistance to kasugamycin and increased susceptibility to chloramphenicol. To the best of our knowledge, this is the first report showing the role of
ksgA
in
S
. Enteritidis virulence in chickens, tolerance to high osmolarity, and altered susceptibility to kasugamycin and chloramphenicol.
Resistance of Bacillus subtilis Spore DNA to Lethal Ionizing Radiation Damage Relies Primarily on Spore Core Components and DNA Repair, with Minor Effects of Oxygen Radical Detoxification
Authors: Moeller Ralf , Raguse Marina , Reitz Günther , Okayasu Ryuichi , Li Zuofeng , Klein Stuart , Setlow Peter , Nicholson Wayne L. 
Published: October 2013

ABSTRACT

The roles of various core components, including α/β/γ-type small acid-soluble spore proteins (SASP), dipicolinic acid (DPA), core water content, and DNA repair by apurinic/apyrimidinic (AP) endonucleases or nonhomologous end joining (NHEJ), in
Bacillus subtilis
spore resistance to different types of ionizing radiation including X rays, protons, and high-energy charged iron ions have been studied. Spores deficient in DNA repair by NHEJ or AP endonucleases, the oxidative stress response, or protection by major α/β-type SASP, DPA, and decreased core water content were significantly more sensitive to ionizing radiation than wild-type spores, with highest sensitivity to high-energy-charged iron ions. DNA repair via NHEJ and AP endonucleases appears to be the most important mechanism for spore resistance to ionizing radiation, whereas oxygen radical detoxification via the MrgA-mediated oxidative stress response or KatX catalase activity plays only a very minor role. Synergistic radioprotective effects of α/β-type but not γ-type SASP were also identified, indicating that α/β-type SASP's binding to spore DNA is important in preventing DNA damage due to reactive oxygen species generated by ionizing radiation.
Establishing the Yeast Kluyveromyces lactis as an Expression Host for Production of the Saposin-Like Domain of the Aspartic Protease Cirsin
Authors: Curto Pedro , Lufrano Daniela , Pinto Cátia , Custódio Valéria , Gomes Ana Catarina , Trejo Sebastián A. , Bakás Laura , Vairo-Cavalli Sandra , Faro Carlos , Simões Isaura 
Published: October 2013

ABSTRACT

Typical plant aspartic protease zymogens comprise a characteristic and plant-specific insert (PSI). PSI domains can interact with membranes, and a role as a defensive weapon against pathogens has been proposed. However, the potential of PSIs as antimicrobial agents has not been fully investigated and explored yet due to problems in producing sufficient amounts of these domains in bacteria. Here, we report the development of an expression platform for the production of the PSI domain of cirsin in the generally regarded as safe (GRAS) yeast
Kluyveromyces lactis
. We successfully generated
K. lactis
transformants expressing and secreting significant amounts of correctly processed and glycosylated PSI, as well as its nonglycosylated mutant. A purification protocol with protein yields of ∼4.0 mg/liter was established for both wild-type and nonglycosylated PSIs, which represents the highest reported yield for a nontagged PSI domain. Subsequent bioactivity assays targeting phytopathogenic fungi indicated that the PSI of cirsin is produced in a biologically active form in
K. lactis
and provided clear evidence for its antifungal activity. This yeast expression system thereby emerges as a promising production platform for further exploring the biotechnological potential of these plant saposin-like proteins.
Expression of Functional Plasmodium falciparum Enzymes Using a Wheat Germ Cell-Free System
Authors: Mudeppa Devaraja G. , Rathod Pradipsinh K. 
Published: October 2013

ABSTRACT

One decade after the sequencing of the
Plasmodium falciparum
genome, 95% of malaria proteins in the genome cannot be expressed in traditional cell-based expression systems, and the targets of the best new leads for antimalarial drug discovery are either not known or not available in functional form. For a disease that kills up to 1 million people per year, routine expression of recombinant malaria proteins in functional form is needed both for the discovery of new therapeutics and for identification of targets of new drugs. We tested the general utility of cell-free systems for expressing malaria enzymes. Thirteen test enzyme sequences were reverse amplified from total RNA, cloned into a plant-like expression vector, and subjected to cell-free expression in a wheat germ system. Protein electrophoresis and autoradiography confirmed the synthesis of products of expected molecular masses. In rare problematic cases, truncated products were avoided by using synthetic genes carrying wheat codons. Scaled-up production generated 39 to 354 μg of soluble protein per 10 mg of translation lysate. Compared to rare proteins where cell-based systems do produce functional proteins, the cell-free yields are comparable or better. All 13 test products were enzymatically active, without failure. This general path to produce functional malaria proteins should now allow the community to access new tools, such as biologically active protein arrays, and lead to the discovery of new chemical functions, structures, and inhibitors of previously inaccessible malaria gene products.
Essential Roles for Mycobacterium tuberculosis Rel beyond the Production of (p)ppGpp
Authors: Weiss Leslie A. , Stallings Christina L. 
Published: October 2013

ABSTRACT

In
Mycobacterium tuberculosis
, the stringent response to amino acid starvation is mediated by the
M. tuberculosis
Rel (Rel
Mtb
) enzyme, which transfers a pyrophosphate from ATP to GDP or GTP to synthesize ppGpp and pppGpp, respectively. (p)ppGpp then influences numerous metabolic processes. Rel
Mtb
also encodes a second, distinct catalytic domain that hydrolyzes (p)ppGpp into pyrophosphate and GDP or GTP. Rel
Mtb
is required for chronic
M. tuberculosis
infection in mice; however, it is unknown which catalytic activity of Rel
Mtb
mediates pathogenesis and whether (p)ppGpp itself is necessary. In order to individually investigate the roles of (p)ppGpp synthesis and hydrolysis during
M. tuberculosis
pathogenesis, we generated Rel
Mtb
point mutants that were either synthetase dead (Rel
Mtb
H344Y
) or hydrolase dead (Rel
Mtb
H80A
).
M. tuberculosis
strains expressing the synthetase-dead Rel
Mtb
H344Y
mutant did not persist in mice, demonstrating that the Rel
Mtb
(p)ppGpp synthetase activity is required for maintaining bacterial titers during chronic infection. Deletion of a second predicted (p)ppGpp synthetase had no effect on pathogenesis, demonstrating that Rel
Mtb
was the major contributor to (p)ppGpp production during infection. Interestingly, expression of an allele encoding the hydrolase-dead Rel
Mtb
mutant, Rel
Mtb
H80A
, that is incapable of hydrolyzing (p)ppGpp but still able to synthesize (p)ppGpp decreased the growth rate of
M. tuberculosis
and changed the colony morphology of the bacteria. In addition, Rel
Mtb
H80A
expression during acute or chronic
M. tuberculosis
infection in mice was lethal to the infecting bacteria. These findings highlight a distinct role for Rel
Mtb
-mediated (p)ppGpp hydrolysis that is essential for
M. tuberculosis
pathogenesis.
Identification of a DNA-Damage-Inducible Regulon in Acinetobacter baumannii
Authors: Aranda Jesús , Poza Margarita , Shingu-Vázquez Miguel , Cortés Pilar , Boyce John D. , Adler Ben , Barbé Jordi , Bou Germán 
Published: October 2013

ABSTRACT

The transcriptional response of
Acinetobacter baumannii
, a major cause of nosocomial infections, to the DNA-damaging agent mitomycin C (MMC) was studied using DNA microarray technology. Most of the 39 genes induced by MMC were related to either prophages or encoded proteins involved in DNA repair. Electrophoretic mobility shift assays demonstrated that the product of the
A. baumannii
MMC-inducible
umuD
gene (
umuDAb
) specifically binds to the palindromic sequence TTGAAAATGTAACTTTTTCAA present in its promoter region. Mutations in this palindromic region abolished UmuDAb protein binding. A comparison of the promoter regions of all MMC-induced genes identified four additional transcriptional units with similar palindromic sequences recognized and specifically bound by UmuDAb. Therefore, the UmuDAb regulon consists of at least eight genes encoding seven predicted error-prone DNA polymerase V components and DddR, a protein of unknown function. Expression of these genes was not induced in the MMC-treated
recA
mutant. Furthermore, inactivation of the
umuDAb
gene resulted in the deregulation of all DNA-damage-induced genes containing the described palindromic DNA motif. Together, these findings suggest that UmuDAb is a direct regulator of the DNA damage response in
A. baumannii
.
A Mutation in the N Domain of Escherichia coli Lon Stabilizes Dodecamers and Selectively Alters Degradation of Model Substrates
Authors: Wohlever Matthew L. , Baker Tania A. , Sauer Robert T. 
Published: October 2013

ABSTRACT

Escherichia coli
Lon, an ATP-dependent AAA
+
protease, recognizes and degrades many different substrates, including the RcsA and SulA regulatory proteins. More than a decade ago, the E240K mutation in the N domain of Lon was shown to prevent degradation of RcsA but not SulA
in vivo
. Here, we characterize the biochemical properties of the E240K mutant
in vitro
and present evidence that the effects of this mutation are complex. For example, Lon
E240K
exists almost exclusively as a dodecamer, whereas wild-type Lon equilibrates between hexamers and dodecamers. Moreover, Lon
E240K
displays degradation defects
in vitro
that do not correlate in any simple fashion with degron identity, substrate stability, or dodecamer formation. The Lon sequence segment near residue 240 is known to undergo nucleotide-dependent conformational changes, and our results suggest that this region may be important for coupling substrate binding with allosteric activation of Lon protease and ATPase activity.
Structural and Functional Characterization of an Orphan ATP-Binding Cassette ATPase Involved in Manganese Utilization and Tolerance in Leptospira spp
Authors: Benaroudj Nadia , Saul Frederick , Bellalou Jacques , Miras Isabelle , Weber Patrick , Bondet Vincent , Murray Gerald L. , Adler Ben , Ristow Paula , Louvel Hélène , Haouz Ahmed , Picardeau Mathieu 
Published: October 2013

ABSTRACT

Pathogenic
Leptospira
species are the etiological agents of the widespread zoonotic disease leptospirosis. Most organisms, including
Leptospira
, require divalent cations for proper growth, but because of their high reactivity, these metals are toxic at high concentrations. Therefore, bacteria have acquired strategies to maintain metal homeostasis, such as metal import and efflux. By screening
Leptospira biflexa
transposon mutants for their ability to use Mn
2+
, we have identified a gene encoding a putative orphan ATP-binding cassette (ABC) ATPase of unknown function. Inactivation of this gene in both
L. biflexa
and
L. interrogans
strains led to mutants unable to grow in medium in which iron was replaced by Mn
2+
, suggesting an involvement of this ABC ATPase in divalent cation uptake. A mutation in this ATPase-coding gene increased susceptibility to Mn
2+
toxicity. Recombinant ABC ATPase of the pathogen
L. interrogans
exhibited Mg
2+
-dependent ATPase activity involving a P-loop motif. The structure of this ATPase was solved from a crystal containing two monomers in the asymmetric unit. Each monomer adopted a canonical two-subdomain organization of the ABC ATPase fold with an α/β subdomain containing the Walker motifs and an α subdomain containing the ABC signature motif (LSSGE). The two monomers were arranged in a head-to-tail orientation, forming a V-shaped particle with all the conserved ABC motifs at the dimer interface, similar to functional ABC ATPases. These results provide the first structural and functional characterization of a leptospiral ABC ATPase.
SmbFT, a Putative ABC Transporter Complex, Confers Protection against the Lantibiotic Smb in Streptococci
Authors: Biswas Saswati , Biswas Indranil 
Published: October 2013

ABSTRACT

Streptococcus mutans
, a dental pathogen, secretes different kinds of lantibiotic and nonlantibiotic bacteriocins. For self-protection, a bacteriocin producer strain must possess one or more cognate immunity mechanisms. We report here the identification of one such immunity complex in
S. mutans
strain GS-5 that confers protection against Smb, a two-component lantibiotic. The immunity complex that we identified is an ABC transporter composed of two proteins: SmbF (the ATPase component) and SmbT (the permease component). Both of the protein-encoding genes are located within the
smb
locus. We show that GS-5 becomes sensitized to Smb upon deletion of
smbT
, which makes the ABC transporter nonfunctional. To establish the role SmbFT in providing immunity, we heterologously expressed this ABC transporter complex in four different sensitive streptococcal species and demonstrated that it can confer resistance against Smb. To explore the specificity of SmbFT in conferring resistance, we tested mutacin IV (a nonlantibiotic), nisin (a single peptide lantibiotics), and three peptide antibiotics (bacitracin, polymyxin B, and vancomycin). We found that SmbFT does not recognize these structurally different peptides. We then tested whether SmbFT can confer protection against haloduracin, another two-component lantibiotic that is structurally similar to Smb; SmbFT indeed conferred protection against haloduracin. SmbFT can also confer protection against an uncharacterized but structurally similar lantibiotic produced by
Streptococcus gallolyticus
. Our data suggest that SmbFT truly displays immunity function and confer protection against Smb and structurally similar lantibiotics.
Variables associated with return to sport following anterior cruciate ligament reconstruction: a systematic review
Authors: Czuppon Sylvia , Racette Brad A , Klein Sandra E , Harris-Hayes Marcie 
Published: October 2013

BackgroundAs one of the purposes of anterior cruciate ligament reconstruction (ACLR) is to return athletes to their preinjury activity level, it is critical to understand variables influencing return to sport. Associations between return to sport and variables representing knee impairment, function and psychological status have not been well studied in athletes following ACLR.PurposeThe purpose of this review was to summarise the literature reporting on variables proposed to be associated with return to sport following ACLR.Study designSystematic review.MethodsMedline, EMBASE, CINAHL and Cochrane databases were searched for articles published before November 2012. Articles included in this review met these criteria: (1) included patients with primary ACLR, (2) reported at least one knee impairment, function or psychological measure, (3) reported a return to sport measure and (4) analysed the relationship between the measure and return to sport.ResultsWeak evidence existed in 16 articles suggesting variables associated with return to sport included higher quadriceps strength, less effusion, less pain, greater tibial rotation, higher Marx Activity score, higher athletic confidence, higher preoperative knee self-efficacy, lower kinesiophobia and higher preoperative self-motivation.ConclusionsWeak evidence supports an association between knee impairment, functional and psychological variables and return to sport. Current return to sport guidelines should be updated to reflect all variables associated with return to sport. Utilising evidence-based return to sport guidelines following ACLR may ensure that athletes are physically and psychologically capable of sports participation, which may reduce reinjury rates and the need for subsequent surgery.
The financial cost of hamstring strain injuries in the Australian Football League
Authors: Hickey Jack , Shield Anthony J , Williams Morgan D , Opar David A 
Published: October 2013

BackgroundHamstring strain injuries (HSIs) have remained the most prevalent injury in the Australian Football League (AFL) over the past 21 regular seasons. The effect of HSIs in sports is often expressed as regular season games missed due to injury. However, the financial cost of athletes missing games due to injury has not been investigated. The aim of this report is to estimate the financial cost of games missed due to HSIs in the AFL.MethodData were collected using publicly available information from the AFL's injury report and the official AFL annual report for the past 10 competitive AFL seasons. Average athlete salary and injury epidemiology data were used to determine the average yearly financial cost of HSIs for AFL clubs and the average financial cost of a single HSI over this time period.ResultsAcross the observed period, average yearly financial cost of HSIs per club increased by 71% compared with a 43% increase in average yearly athlete salary. Over the same time period the average financial cost of a single HSI increased by 56% from $A25 603 in 2003 to $A40 021 in 2012, despite little change in the HSI rates during the period.ConclusionsThe observed increased financial cost of HSIs was ultimately explained by the failure of teams to decrease HSI rates, but coupled with increases in athlete salaries over the past 10 season. The information presented in this report highlights the financial cost of HSIs and other sporting injuries, raising greater awareness and the need for further funding for research into injury prevention strategies to maximise economical return for investment in athletes.