bluuebunny/crossref_metadata_2025_split huggingface dataset:
Inactivation of Human Norovirus in Contaminated Oysters and Clams by High Hydrostatic Pressure
Authors: Ye Mu , Li Xinhui , Kingsley David H. , Jiang Xi , Chen Haiqiang 
Published: February 2014

ABSTRACT

Human norovirus (NoV) is the most frequent causative agent of food-borne disease associated with shellfish consumption. In this study, the effect of high hydrostatic pressure (HHP) on inactivation of NoV was determined. Genogroup I.1 (GI.1) or genogroup II.4 (GII.4) NoV was inoculated into oyster homogenates and treated at 300 to 600 MPa at 25, 6, and 1°C for 5 min. After HHP, samples were treated with RNase and viral particles were extracted with porcine gastric mucin (PGM)-conjugated magnetic beads (PGM-MBs). Viral RNA was then quantified by real-time reverse transcription (RT)-PCR. Since PGM contains histo-blood group-like antigens, which can act as receptors for NoV, deficiency for binding to PGM is an indication of loss of infectivity of NoV. After binding to PGM-MBs, RT-PCR-detectable NoV RNA in oysters was reduced by 0.4 to >4 log
10
by HHP at 300 to 600 MPa. The GI.1 NoV was more resistant to HHP than the GII.4 NoV (
P
< 0.05). HHP at lower temperatures significantly enhanced the inactivation of NoV in oysters (
P
< 0.05). Pressure treatment was also conducted for clam homogenates. Treatment at 450 MPa at 1°C achieved a >4 log
10
reduction of GI.1 NoV in both oyster and clam homogenates. It is therefore concluded that HHP could be applied as a potential intervention for inactivating NoV in raw shellfish. The method of pretreatment of samples with RNase, extraction of viral particles using PGM-MB binding, and quantification of viral RNA using RT-PCR can be explored as a practical means of distinguishing between infectious and noninfectious NoV.
H
2
O
2
Production in Species of the Lactobacillus acidophilus Group: a Central Role for a Novel NADH-Dependent Flavin Reductase
Authors: Hertzberger Rosanne , Arents Jos , Dekker Henk L. , Pridmore R. David , Gysler Christof , Kleerebezem Michiel , de Mattos M. Joost Teixeira 
Published: February 2014

ABSTRACT

Hydrogen peroxide production is a well-known trait of many bacterial species associated with the human body. In the presence of oxygen, the probiotic lactic acid bacterium
Lactobacillus johnsonii
NCC 533 excretes up to 1 mM H
2
O
2
, inducing growth stagnation and cell death. Disruption of genes commonly assumed to be involved in H
2
O
2
production (e.g., pyruvate oxidase, NADH oxidase, and lactate oxidase) did not affect this. Here we describe the purification of a novel NADH-dependent flavin reductase encoded by two highly similar genes (
LJ
_
0548
and
LJ
_
0549
) that are conserved in lactobacilli belonging to the
Lactobacillus acidophilus
group. The genes are predicted to encode two 20-kDa proteins containing flavin mononucleotide (FMN) reductase conserved domains. Reductase activity requires FMN, flavin adenine dinucleotide (FAD), or riboflavin and is specific for NADH and not NADPH. The

K
m

for FMN is 30 ± 8 μM, in accordance with its proposed
in vivo
role in H
2
O
2
production. Deletion of the encoding genes in
L. johnsonii
led to a 40-fold reduction of hydrogen peroxide formation. H
2
O
2
production in this mutant could only be restored by in
trans
complementation of both genes. Our work identifies a novel, conserved NADH-dependent flavin reductase that is prominently involved in H
2
O
2
production in
L. johnsonii
.
In-Feed Use of Heavy Metal Micronutrients in U.S. Swine Production Systems and Its Role in Persistence of Multidrug-Resistant Salmonellae
Authors: Medardus Julius J. , Molla Bayleyegn Z. , Nicol Matthew , Morrow W. Morgan , Rajala-Schultz Paivi J. , Kazwala Rudovick , Gebreyes Wondwossen A. 
Published: February 2014

ABSTRACTThe study aimed to characterize the role of heavy metal micronutrients in swine feed in emergence of heavy-metal-tolerant and multidrug-resistantSalmonellaorganisms. We conducted a longitudinal study in 36 swine barns over a 2-year period. The feed and fecal levels of Cu2+and Zn2+were measured.Salmonellawas isolated at early and late finishing. MICs of copper sulfate and zinc chloride were measured using agar dilution. Antimicrobial susceptibility was tested using the Kirby-Bauer method, and 283 isolates were serotyped. We amplifiedpcoAandczcDgenes that encode Cu2+and Zn2+tolerance, respectively. Of the 283 isolates, 113 (48%) showed Cu2+tolerance at 24 mM and 164 (58%) showed Zn2+tolerance at 8 mM. In multivariate analysis, serotype and source of isolates were significantly associated with Cu2+tolerance (P< 0.001). Fecal isolates were more likely to be Cu2+tolerant than those of feed origin (odds ratio [OR], 27.0; 95% confidence interval [CI], 2.8 to 250;P= 0.0042) or environmental origin (OR, 5.8), implying the significance of gastrointestinal selective pressure.Salmonella entericaserotypes Typhimurium and Heidelberg, highly significant for public health, had higher odds of having >20 mM MICs of Cu2+than did “other” serotypes. More than 60% ofSalmonellaisolates with resistance type (R-type) AmStTeKm (32 of 53) carriedpcoA; only 5% with R-type AmClStSuTe carried this gene.czcDgene carriage was significantly associated with a higher Zn2+MIC (P< 0.05). The odds of having a high Zn2+MIC (≥8 mM) were 14.66 times higher in isolates with R-type AmClStSuTe than in those with R-type AmStTeKm (P< 0.05). The findings demonstrate strong association between heavy metal tolerance and antimicrobial resistance, particularly amongSalmonellaserotypes important in public health.
Two-Component Histidine Phosphotransfer Protein Ypd1 Is Not Essential for Viability in Candida albicans
Authors: Mavrianos John , Desai Chirayu , Chauhan Neeraj 
Published: February 2014

ABSTRACT

Prokaryotes and lower eukaryotes, such as yeasts, utilize two-component signal transduction pathways to adapt cells to environmental stress and to regulate the expression of genes associated with virulence. One of the central proteins in this type of signaling mechanism is the phosphohistidine intermediate protein Ypd1. Ypd1 is reported to be essential for viability in the model yeast
Saccharomyces cerevisiae
. We present data here showing that this is not the case for
Candida albicans
. Disruption of
YPD1
causes cells to flocculate and filament constitutively under conditions that favor growth in yeast form. To determine the function of Ypd1 in the Hog1 mitogen-activated protein kinase (MAPK) pathway, we measured phosphorylation of Hog1 MAPK in
ypd1
Δ/Δ and wild-type strains of
C. albicans
. Constitutive phosphorylation of Hog1 was observed in the
ypd1
Δ/Δ strain compared to the wild-type strain. Furthermore, fluorescence microscopy revealed that green fluorescent protein (GFP)-tagged Ypd1 is localized to both the nucleus and the cytoplasm. The subcellular segregation of GFP-tagged Ypd1 hints at an important role(s) of Ypd1 in regulation of Ssk1 (cytosolic) and Skn7 (nuclear) response regulator proteins via phosphorylation in
C. albicans
. Overall, our findings have profound implications for a mechanistic understanding of two-component signaling pathways in
C. albicans
, and perhaps in other pathogenic fungi.
The Acetylation Motif in AMP-Forming Acyl Coenzyme A Synthetases Contains Residues Critical for Acetylation and Recognition by the Protein Acetyltransferase Pat of Rhodopseudomonas palustris
Authors: Crosby Heidi A. , Escalante-Semerena Jorge C. 
Published: February 2014

ABSTRACT

The AMP-forming acyl coenzyme A (acyl-CoA) synthetases are a large class of enzymes found in both anabolic and catabolic pathways that activate fatty acids to acyl-CoA molecules. The protein acetyltransferase (Pat) from
Rhodopseudomonas palustris
(
Rp
Pat) inactivates AMP-forming acyl-CoA synthetases by acetylating the ε-amino group of a conserved, catalytic lysine residue. In all of the previously described
Rp
Pat substrates, this lysine residue is located within a PX
4
GK motif that has been proposed to be a recognition motif for
Rp
Pat. Here, we report five new substrates for
Rp
Pat, all of which are also AMP-forming acyl-CoA synthetases. This finding supports the idea that Pat enzymes may have evolved to control the activity of this family of enzymes. Notably,
Rp
Pat did not acetylate the wild-type long-chain acyl-CoA synthetase B (
Rp
LcsB; formerly Rpa2714) enzyme of this bacterium. However, a single amino acid change two residues upstream of the acetylation site was sufficient to convert
Rp
LcsB into an
Rp
Pat substrate. The results of mutational and functional analyses of
Rp
LcsB and
Rp
PimA variants led us to propose PK/R
T
X
S/T/V/N
GKX
2
K/R as a substrate recognition motif. The underlined positions within this motif are particularly important for acetylation by
Rp
Pat. The first residue, threonine, is located 4 amino acids upstream of the acetylation site. The second residue can be S/T/V/N and is located two positions upstream of the acetylation site. Analysis of published crystal structures suggests that the side chains of these two residues are very close to the acetylated lysine residue, indicating that they may directly interact with
Rp
Pat.
Roles of Cyclic Di-GMP and the Gac System in Transcriptional Control of the Genes Coding for the Pseudomonas putida Adhesins LapA and LapF
Authors: Martínez-Gil Marta , Ramos-González María Isabel , Espinosa-Urgel Manuel 
Published: February 2014

ABSTRACT

LapA and LapF are large extracellular proteins that play a relevant role in biofilm formation by
Pseudomonas putida
. Current evidence favors a sequential model in which LapA is first required for the initial adhesion of individual bacteria to a surface, while LapF participates in later stages of biofilm development. In agreement with this model,
lapF
transcription was previously shown to take place at late times of growth and to respond to the stationary-phase sigma factor RpoS. We have now analyzed the transcription pattern of
lapA
and other regulatory elements that influence expression of both genes. The
lapA
promoter shows a transient peak of activation early during growth, with a second increase in stationary phase that is independent of RpoS. The same pattern is observed in biofilms although expression is not uniform in the population. Both
lapA
and
lapF
are under the control of the two-component regulatory system GacS/GacA, and their transcription also responds to the intracellular levels of the second messenger cyclic diguanylate (c-di-GMP), although in surprisingly reverse ways. Whereas expression from the
lapA
promoter increases with high levels of c-di-GMP, the opposite is true for
lapF
. The transcriptional regulator FleQ is required for the modulation of
lapA
expression by c-di-GMP but has a minor influence on
lapF
. This work represents a further step in our understanding of the regulatory interactions controlling biofilm formation in
P. putida
.
Natural Competence and the Evolution of DNA Uptake Specificity
Authors: Mell Joshua Chang , Redfield Rosemary J. 
Published: February 2014

ABSTRACT

Many bacteria are naturally competent, able to actively transport environmental DNA fragments across their cell envelope and into their cytoplasm. Because incoming DNA fragments can recombine with and replace homologous segments of the chromosome, competence provides cells with a potent mechanism of horizontal gene transfer as well as access to the nutrients in extracellular DNA. This review starts with an introductory overview of competence and continues with a detailed consideration of the DNA uptake specificity of competent proteobacteria in the
Pasteurellaceae
and
Neisseriaceae
. Species in these distantly related families exhibit strong preferences for genomic DNA from close relatives, a self-specificity arising from the combined effects of biases in the uptake machinery and genomic overrepresentation of the sequences this machinery prefers. Other competent species tested lack obvious uptake bias or uptake sequences, suggesting that strong convergent evolutionary forces have acted on these two families. Recent results show that uptake sequences have multiple “dialects,” with clades within each family preferring distinct sequence variants and having corresponding variants enriched in their genomes. Although the genomic consensus uptake sequences are 12 and 29 to 34 bp, uptake assays have found that only central cores of 3 to 4 bp, conserved across dialects, are crucial for uptake. The other bases, which differ between dialects, make weaker individual contributions but have important cooperative interactions. Together, these results make predictions about the mechanism of DNA uptake across the outer membrane, supporting a model for the evolutionary accumulation and stability of uptake sequences and suggesting that uptake biases may be more widespread than currently thought.
Functional Analysis of the Accessory Protein TapA in Bacillus subtilis Amyloid Fiber Assembly
Authors: Romero Diego , Vlamakis Hera , Losick Richard , Kolter Roberto 
Published: February 2014

ABSTRACT

Bacillus subtilis
biofilm formation relies on the assembly of a fibrous scaffold formed by the protein TasA. TasA polymerizes into highly stable fibers with biochemical and morphological features of functional amyloids. Previously, we showed that assembly of TasA fibers requires the auxiliary protein TapA. In this study, we investigated the roles of TapA sequences from the C-terminal and N-terminal ends and TapA cysteine residues in its ability to promote the assembly of TasA amyloid-like fibers. We found that the cysteine residues are not essential for the formation of TasA fibers, as their replacement by alanine residues resulted in only minor defects in biofilm formation. Mutating sequences in the C-terminal half had no effect on biofilm formation. However, we identified a sequence of 8 amino acids in the N terminus that is key for TasA fiber formation. Strains expressing TapA lacking these 8 residues were completely defective in biofilm formation. In addition, this TapA mutant protein exhibited a dominant negative effect on TasA fiber formation. Even in the presence of wild-type TapA, the mutant protein inhibited fiber assembly
in vitro
and delayed biofilm formation
in vivo
. We propose that this 8-residue sequence is crucial for the formation of amyloid-like fibers on the cell surface, perhaps by mediating the interaction between TapA or TapA and TasA molecules.
Comparative Functional Genomics of Lactobacillus spp. Reveals Possible Mechanisms for Specialization of Vaginal Lactobacilli to Their Environment
Authors: Mendes-Soares Helena , Suzuki Haruo , Hickey Roxana J. , Forney Larry J. 
Published: February 2014

ABSTRACT

Lactobacilli are found in a wide variety of habitats. Four species,
Lactobacillus crispatus
,
L. gasseri
,
L. iners
, and
L. jensenii
, are common and abundant in the human vagina and absent from other habitats. These may be adapted to the vagina and possess characteristics enabling them to thrive in that environment. Furthermore, stable codominance of multiple
Lactobacillus
species in a single community is infrequently observed. Thus, it is possible that individual vaginal
Lactobacillus
species possess unique characteristics that confer to them host-specific competitive advantages. We performed comparative functional genomic analyses of representatives of 25 species of
Lactobacillus
, searching for habitat-specific traits in the genomes of the vaginal lactobacilli. We found that the genomes of the vaginal species were significantly smaller and had significantly lower GC content than those of the nonvaginal species. No protein families were found to be specific to the vaginal species analyzed, but some were either over- or underrepresented relative to nonvaginal species. We also found that within the vaginal species, each genome coded for species-specific protein families. Our results suggest that even though the vaginal species show no general signatures of adaptation to the vaginal environment, each species has specific and perhaps unique ways of interacting with its environment, be it the host or other microbes in the community. These findings will serve as a foundation for further exploring the role of lactobacilli in the ecological dynamics of vaginal microbial communities and their ultimate impact on host health.
Germination of Spores of Bacillus Species: What We Know and Do Not Know
Authors: Setlow Peter 
Published: February 2014

ABSTRACT

Spores of
Bacillus
species can remain in their dormant and resistant states for years, but exposure to agents such as specific nutrients can cause spores' return to life within minutes in the process of germination. This process requires a number of spore-specific proteins, most of which are in or associated with the inner spore membrane (IM). These proteins include the (i) germinant receptors (GRs) that respond to nutrient germinants, (ii) GerD protein, which is essential for GR-dependent germination, (iii) SpoVA proteins that form a channel in spores' IM through which the spore core's huge depot of dipicolinic acid is released during germination, and (iv) cortex-lytic enzymes (CLEs) that degrade the large peptidoglycan cortex layer, allowing the spore core to take up much water and swell, thus completing spore germination. While much has been learned about nutrient germination, major questions remain unanswered, including the following. (i) How do nutrient germinants penetrate through spores' outer layers to access GRs in the IM? (ii) What happens during the highly variable and often long lag period between the exposure of spores to nutrient germinants and the commitment of spores to germinate? (iii) What do GRs and GerD do, and how do these proteins interact? (iv) What is the structure of the SpoVA channel in spores' IM, and how is this channel gated? (v) What is the precise state of the spore IM, which has a number of novel properties even though its lipid composition is very similar to that of growing cells? (vi) How is CLE activity regulated such that these enzymes act only when germination has been initiated? (vii) And finally, how does the germination of spores of clostridia compare with that of spores of bacilli?
RNase E Affects the Expression of the Acyl-Homoserine Lactone Synthase Gene
sinI
in Sinorhizobium meliloti
Authors: Baumgardt Kathrin , Charoenpanich Pornsri , McIntosh Matthew , Schikora Adam , Stein Elke , Thalmann Sebastian , Kogel Karl-Heinz , Klug Gabriele , Becker Anke , Evguenieva-Hackenberg Elena 
Published: February 2014

ABSTRACT

Quorum sensing of
Sinorhizobium meliloti
relies on
N
-acyl-homoserine lactones (AHLs) as autoinducers. AHL production increases at high population density, and this depends on the AHL synthase SinI and two transcriptional regulators, SinR and ExpR. Our study demonstrates that ectopic expression of the gene
rne
, coding for RNase E, an endoribonuclease that is probably essential for growth, prevents the accumulation of AHLs at detectable levels. The ectopic
rne
expression led to a higher level of
rne
mRNA and a lower level of
sinI
mRNA independently of the presence of ExpR, the AHL receptor, and AHLs. In line with this, IPTG (isopropyl-β-
d
-thiogalactopyranoside)-induced overexpression of
rne
resulted in a shorter half-life of
sinI
mRNA and a strong reduction of AHL accumulation. Moreover, using translational
sinI-egfp
fusions, we found that
sinI
expression is specifically decreased upon induced overexpression of
rne
, independently of the presence of the global posttranscriptional regulator Hfq. The 28-nucleotide 5′ untranslated region (UTR) of
sinI
mRNA was sufficient for this effect. Random amplification of 5′ cDNA ends (5′-RACE) analyses revealed a potential RNase E cleavage site at position +24 between the Shine-Dalgarno site and the translation start site. We postulate therefore that RNase E-dependent degradation
of sinI
mRNA from the 5′ end is one of the steps mediating a high turnover of
sinI
mRNA, which allows the Sin quorum-sensing system to respond rapidly to changes in transcriptional control of AHL production.
Mechanics of turning and jumping and skier speed are associated with injury risk in men's World Cup alpine skiing: a comparison between the competition disciplines
Authors: Gilgien Matthias , Spörri Jörg , Kröll Josef , Crivelli Philip , Müller Erich 
Published: February 2014

Background/aimIn alpine ski racing, there is limited information about skiers’ mechanical characteristics and their relation to injury risk, in particular for World Cup (WC) competitions. Hence, current findings from epidemiological and qualitative research cannot be linked to skiers’ mechanics. This study was undertaken to investigate whether recently reported differences in numbers of injuries per 1000 runs for competition disciplines can be explained by differences in the skiers’ mechanics.MethodsDuring seven giant slalom, four super-G and five downhill WC competitions, mechanical characteristics of a forerunner were captured using differential global navigation satellite technology and a precise terrain surface model. Finally, the discipline-specific skiers’ mechanics were compared with the respective number of injuries per hour skiing.ResultsWhile the number of injuries per hour skiing was approximately equal for all disciplines, kinetic energy, impulse, run time, turn radius and turn speed were significantly different and increased from giant slalom to super-G and downhill. Turn ground reaction forces were largest for giant slalom, followed by super-G and downhill. The number of jumps was doubled from super-G to downhill.ConclusionsAssociating the number of injuries per hour in WC skiing with skiers’ mechanical characteristics, injuries in super-G and downhill seem to be related to increased speed and jumps, while injuries in giant slalom may be related to high loads in turning. The reported differences in the number of injuries per 1000 runs might be explained by a bias in total exposure time per run and thus potentially by emerged fatigue.
Hospital-based surveillance of rotavirus gastroenteritis among children under 5 years of age in the Republic of Ivory Coast: a cross-sectional study
Authors: Akoua-Koffi Chantal , Asse Kouadio Vincent , Yao Atteby Jean Jacques 
Published: February 2014

ObjectivesTo estimate the proportion of rotavirus gastroenteritis (RVGE) among children aged less than 5 years who had been diagnosed with acute gastroenteritis (AGE) and admitted to hospitals and emergency rooms (ERs). The seasonal distribution of RVGE and most prevalent rotavirus (RV) strains was also assessed.DesignA cross-sectional hospital-based surveillance study.Setting5 reference paediatric hospitals across Abidjan.ParticipantsChildren aged less than 5 years, who were hospitalised/visiting ERs for WHO-defined AGE, were enrolled. Written informed consent was obtained from parents/guardians before enrolment. Children who acquired nosocomial infection were excluded from the study.Primary and secondary outcome measuresThe proportion of RVGE among AGE hospitalisations and ER visits was expressed with 95% exact CI. Stool samples were collected from all enrolled children and were tested for the presence of RV using an enzyme immunoassay. RV-positive samples were serotyped using reverse transcriptase-PCR.ResultsOf 357 enrolled children (mean age 13.6±11.14 months), 332 were included in the final analyses; 56.3% (187/332) were hospitalised and 43.7% (145/332) were admitted to ERs. The proportion of RVGE hospitalisations and ER visits among all AGE cases was 30.1% (95% CI 23.6% to 37.3%) and 26.9% (95% CI 19.9% to 34.9%), respectively. Ninety-five children (28.6%) were RV positive; the highest number of RVGE cases was observed in children aged 6–11 months. The number of GE cases peaked in July and August 2008; the highest percentage of RV-positive cases was observed in January 2008. G1P[8] wild-type and G8P[6] were the most commonly detected strains.ConclusionsRVGE causes substantial morbidity among children under 5 years of age and remains a health concern in the Republic of Ivory Coast, where implementation of prevention strategies such as vaccination might help to reduce disease burden.