bluuebunny/crossref_metadata_2025 huggingface dataset:
Alcohol Abuse Enhances Pulmonary Edema in Acute Respiratory Distress Syndrome
Authors: Berkowitz David M. , Danai Pajman A. , Eaton Stephanie , Moss Marc , Martin Greg S.
Published: July 2009

Background:  Pulmonary edema is a cardinal feature of the life‐threatening condition known as acute respiratory distress syndrome (ARDS). Patients with chronic alcohol abuse are known to be at increased risk of developing and dying from ARDS. Based upon preclinical data, we hypothesized that a history of chronic alcohol abuse in ARDS patients is associated with greater quantities and slower resolution of pulmonary edema compared with ARDS patients without a history of alcohol abuse.

Methods:  A PiCCO™ transpulmonary thermodilution catheter was inserted into 35 patients within 72 hours of meeting American European Consensus Criteria definition of ARDS. Pulmonary edema was quantified as extravascular lung water (EVLW) and measured for up to 7 days in 13 patients with a history of chronic alcohol abuse and 22 patients without a history of chronic alcohol abuse.

Results:  Mean EVLW was higher in patients with a history of chronic alcohol abuse (16.6 vs. 10.5 ml/kg, p < 0.0001). Patients with alcohol abuse had significantly greater EVLW over the duration of the study (RM‐ANOVA p = 0.003). There was a trend towards slower resolution of EVLW in patients with a history of alcohol abuse (a decrease of 0.5 ml/kg vs. 2.4 ml/kg, p = 0.17) over the study period. A history of alcohol abuse conferred a greater than 3‐fold increased risk of elevated EVLW [OR 3.16, (1.26 to 7.93)] using multivariate logistic regression analysis.

Conclusions:  In patients who develop ARDS, alcohol abuse is associated with greater levels EVLW and a trend towards slower resolution of EVLW. Combined with mechanistic and preclinical evidence linking chronic alcohol consumption and ARDS, targeted therapies should be developed for these patients.
Upregulation of Serotonin Transporter by Alcohol in Human Dendritic Cells: Possible Implication in Neuroimmune Deregulation
Authors: Babu Dakshayani Kadiyala , Diaz Alain , Samikkannu Thangavel , Rao Kurapati V. K. , Saiyed Zainulabedin M. , Rodriguez Jose W. , Nair Madhavan P.N.
Published: July 2009

Background:  Alcohol is the most widely abused substance and its chronic consumption causes neurobehavioral disorders. It has been shown that alcohol affects the function of immune cells. Dendritic cells (DC) serve as the first line of defense against infections and are known to accumulate neurotransmitters such as 5‐hydroxytryptamine (5‐HT). The enzyme monoamine oxidase‐A (MAO‐A) degrades 5‐HT that is associated with clinical depression and other neurological disorders. 5‐HT is selectively transported into neurons through the serotonin transporter (SERT), which is a member of the sodium‐ and chloride‐dependent neurotransmitter transporter (SLC6) family. SERT also serves as a receptor for psychostimulant recreational drugs. It has been demonstrated that several drugs of abuse such as amphetamine and cocaine inhibit the SERT expression; however, the role of alcohol is yet to be elucidated. We hypothesize that alcohol can modulate SERT and MAO‐A expression in DC, leading to reciprocal downregulation of 5‐HT in extracellular medium.

Methods:  Dendritic cells were treated with different concentrations (0.05% to 0.2%v/v) of alcohol for 24–72 hours and processed for SERT and MAO‐A expression using Q‐PCR and Western blots analysis. In addition, SERT function in DC treated with alcohol both in the presence and absence of imipramine, a SERT inhibitor was measured using 4‐[4‐(dimethylamino)styryl]‐1‐methylpyridinium iodide uptake assay. 5‐HT levels in culture supernatant and intracellular 5‐hydroxy indole acetic acid (5‐HIAA) and cyclic AMP were also quantitated using ELISA.

Results:  Dendritic cells treated with 0.1% alcohol for 24 hours showed significant upregulation of SERT and MAO‐A expression compared with untreated DC. We also observed that 0.1% alcohol enhanced the function of SERT and decreased extracellular 5‐HT levels compared with untreated DC cultures, and this was associated with the elevation of intracellular 5‐HIAA and cyclic AMP levels.

Conclusions:  Our study suggests that alcohol upregulates SERT and MAO‐A by elevating cyclic AMP, which may lead to decreased concentration of 5‐HT in the extracellular medium. As 5‐HT is a major neurotransmitter and an inflammatory mediator, its alcohol‐mediated depletion may cause both neurological and immunological deregulation.
The Canadian Prehospital Evidence‐based Protocols Project: Knowledge Translation in Emergency Medical Services Care
Authors: Jensen Jan L. , Petrie David A. , Travers Andrew H. ,   for the PEP Project Team
Published: July 2009

Objectives:  The principles of evidence‐based medicine are applicable to all areas and professionals in health care. The care provided by paramedics in the prehospital setting is no exception. The Prehospital Evidence‐based Protocols Project Online (PEP) is a repository of appraised research evidence that is applicable to interventions performed in the prehospital setting and is openly available online. This article describes the history, current status, and potential future of the project.

Methods:  The primary objective of the PEP is to catalog and grade emergency medical services (EMS) studies with a level of evidence (LOE). Subsequently, each prehospital intervention is assigned a class of recommendation (COR) based on all the appraised articles on that intervention, in an effort to organize the evidence so it may be put into practice efficiently. An LOE is assigned to each article by the section editor, based on the study rigor and applicability to EMS. The section editor committee consists of EMS physicians and paramedics from across Canada, and two from Ireland and a paramedic coordinator. The evidence evaluation cycle is continuous; as the section editors send back appraisals, the coordinator updates the database and sends out another article for review.

Results:  The database currently has 182 individual interventions organized under 103 protocols, with 933 citations.

Conclusions:  This project directly meets recent recommendations to improve EMS by using evidence to support interventions and incorporating it into protocols. Organizing and grading the evidence allows medical directors and paramedics to incorporate research findings into their daily practice. As such, this project demonstrates how knowledge translation can be conducted in EMS.
Altered body composition in preterm infants at hospital discharge
Authors: Cooke Richard J , Griffin Ian
Published: July 2009

Aim:  To test the hypotheses that body size is reduced and body composition altered in preterm infants at hospital discharge.

Methods:  Preterm infants (≤34 weeks gestation, ≤1750 g at birth) were enrolled. Body weight, length and head circumference were converted to standard deviation or z‐scores. Body composition was measured using dual emission X‐ray absorptiometry. The results were analysed using standard statistics.

Results:  One hundred and forty‐nine infants (birth weight = 1406 ± 248 g, gestation = 31 ± 1.7 weeks) were studied. Postmenstrual age at discharge was 37 ± 1.2 weeks. Z‐scores for head circumference, weight and length differed (−0.1 ± 0.6 > −1.4 ± 0.6 > −1.9 ± 0.6; p < 0.0001). Global fat‐free mass was less in study infants than the reference infant at the same weight (2062 < 2252 g; p < 0.0001) or gestation (2062 < 2667 g; p < 0.0001). Global fat mass was greater in study infants than the reference infant at the same weight (307 > 198 g, 13 > 8%) or gestation (307 > 273 g; 13 > 9%; p < 0.0001). Changes in central fat mass closely paralleled those in global fat mass (r2 = 0.76, p < 0.0001).

Conclusion:  Reduced linear growth and a reduced fat‐free mass suggest that dietary protein needs were not met before discharge. A reduced fat‐free mass coupled with an increased global and central fat mass echoes concerns about the development of insulin resistance and metabolic syndrome X in these high‐risk infants.
Nephrocalcinosis in children: a retrospective multi‐centre study
Authors: Ammenti Anita , Pelizzoni Alessandra , Cecconi Milvia , Molinari Pier Paolo , Montini Giovanni
Published: July 2009

Aim:  To review the data of children with NC and to analyse aetiology, clinical manifestations, growth and renal function at presentation; to relate growth and renal function to changes in NC in patients with a follow‐up of at least 12 months.

Methods:  Data of 41 children from four institutions were gathered retrospectively.

Results:  Presenting symptoms were failure to thrive in the first year of life (41%), urinary tract infections, bladder voiding dysfunction or abdominal pain (17%) and psychomotor delay (10%). In 24% of cases NC was detected incidentally. Glomerular function at diagnosis was normal in 83% of children. During a median follow‐up of 4 yrs and 5 months in 28 patients, growth standard deviation score improved from a median of −2.2 to −1.0 and glomerular function remained stable in 89% of patients, in spite of worsening of the degree of NC in 62% of cases. The most frequent causes of NC were hereditary tubulopathies and vitamin D intoxication.

Conclusion:  Our results show that the treatment of the underlying conditions is associated with catch‐up growth and stabilization of glomerular function in many children, but not with the reduction in the degree of NC in the majority of cases. We believe that early recognition of conditions leading to NC is clinically useful and suggest a diagnostic flowchart, which may be helpful in the approach to NC.
Orexins/hypocretins and aminergic systems
Authors: Eriksson K. S. , Sergeeva O. A. , Haas H. L. , Selbach O.
Published: June 2009

Orexin/hypocretin neurones in the posterior hypothalamus are mutually connected with noradrenergic, serotonergic, dopaminergic, histaminergic, and cholinergic neurone systems. They activate these targets by direct post‐synaptic and indirect pre‐synaptic mechanisms and in turn receive inhibitory feedback and excitatory feed forward control. With respect to behavioural state control, orexin/hypocretin neurones are conducting the orchestra of biogenic amines. This review highlights the role of these players in the control of energy administration, sleep–wake architecture, cortical activation, plasticity, and memory functions in health and disease.
Need for uniqueness in older women: an exploratory look
Authors: Armstrong Myrna L. , Saunders Jana C. , Owen Donna C. , Roberts Alden E. , Koch Jerome R.
Published: July 2009

Problem.  Information about older women dwells on age‐related illness and disabilities yet, other empowering traits could be promoting positive forces.

Aim.  While everyone desires some distinctiveness, this study explores higher levels of the motivational construct need for uniqueness (NU) in older women. NU concepts are: 1) perceived feelings of being or thinking different, 2) outspoken social behaviour, and 3) creative consumer consumption.

Design.  An exploratory, descriptive cross‐sectional study provided quantitative data and focus groups generated qualitative data.

Major results.  Significant differences in all three NU concepts were evident between 87 (42%) with no/weak NU and 122 (58%) who ‘preferred to be different from other people’. Focus groups provided examples over their lifetime within the context of their societal expectations.

Relevance to clinical practice.  The nurse’s strategy to partner with older women to individualize their care and teaching will be important in maintaining their well‐being. Examining their decision‐making with how they respond to perceived feelings of being or thinking different (such as taking risks), how outspoken are they, and their patterns of consumer consumption are a few questions that could be asked to examine an older woman’s NU. Further research examining specific health care decisions in those with NU will also be needed to note their responses toward future self‐care activities.
Intraoperative use of a reflective blanket (Sirius®rescue sheet) for temperature management in dogs less than 10 kg
Authors: Tünsmeyer J. , Bojarski I. , Nolte I. , Kramer S.
Published: July 2009

Objectives:To compare the effects of the Sirius®rescue sheet with gel padsversusgel pads alone on intraoperative body temperature in dogs less than 10 kg.

Methods:Forty small breed dogs undergoing elective surgical procedures were randomly assigned to two groups. One group was intraoperatively laid on warmed gel pads, and the other group was additionally wrapped in a Sirius®rescue sheet. Oesophageal body temperature was determined every 10 minutes and compared between groups. Temperature of gel pads was measured preoperatively and postoperatively to compare heat loss of the gel pads between groups.

Results:The body temperature of dogs wrapped with the Sirius®rescue sheet increased intraoperatively. In dogs just lying on warmed gel pads, a decrease in mean body temperature was revealed and mean body temperatures differed between groups after 40 minutes. Extent of heat loss from the gel pads did not differ between the groups.

ClinicalSignificance:The Sirius®rescue sheet, used in addition to warmed gel pads, led to higher intraoperative body temperatures in small breed dogs undergoing surgical procedures to the extremities and the head. The cost‐effectiveness and ease of handling make this a useful addition to clinical practice.
Diagnostic accuracy of positive contrast computed tomography arthrography for the detection of injuries to the medial meniscus in dogs with naturally occurring cranial cruciate ligament insufficiency
Authors: Tivers M. S. , Mahoney P. N. , Baines E. A. , Corr S. A.
Published: July 2009

Objective: To assess the usefulness of computed tomography arthrography of the stifle in diagnosing meniscal tears in dogs with cranial cruciate ligament insufficiency.

Methods: A prospective clinical study was performed. Dogs were included if they had evidence of cranial cruciate ligament insufficiency or persistent or recurrent lameness following surgery for cranial cruciate ligament insufficiency. Dogs were sedated for a computed tomography scan of the affected stifle, orientated in the dorsal plane. A survey computed tomography scan was followed by a computed tomography arthrogram. A stifle arthrotomy was performed, and the surgical findings were recorded. The computed tomography scans were reviewed by three blinded reviewers, and the results were compared to the surgical findings.

Results: Twenty‐one computed tomography arthrograms from 20 dogs were included. At surgery, damage to the medial meniscus was identified in 14 stifles. Initial interpretation of computed tomography arthrography images was 57 to 64 per cent sensitive and 71 to 100 per cent specific for diagnosing medial meniscal injuries. Interpretation of the images on retrospective analysis was 71 per cent sensitive and 100 per cent specific, with an accuracy of 0·857.

Clinical Significance: The accuracy of stifle computed tomography arthrography for the diagnosis of tears to the medial meniscus was found to be good. It is a minimally invasive and repeatable technique, which does not require general anaesthesia or specialist training to obtain the images. The ability to reliably diagnose meniscal injury without the need for surgery may be advantageous, particularly in dogs which had previously had surgery for cranial cruciate ligament insufficiency.
Introduction to the Special Issue on Marketing and Operations Management Interfaces and Coordination
Authors: Ho Teck H. , Tang Christopher S.
Published: July 2009

In this special issue, the contributing authors address several emerging marketing and operations interface problems and develop innovative approaches for solving them. Specifically, by explicitly modeling active consumer behavior under different pricing schemes, the papers in this special issue examine how firms can coordinate their marketing and operations to improve their competitiveness and profit. The papers also provide insights on how to develop and operate new and innovative market mechanisms.
Effects of pre‐ and periconceptional undernutrition on arterial function in adult female sheep are vascular bed dependent
Authors: Torrens Christopher , Snelling Tristram H. , Chau Ryan , Shanmuganathan Meena , Cleal Jane K. , Poore Kirsten R. , Noakes David E. , Poston Lucilla , Hanson Mark A. , Green Lucy R.
Published: June 2009

The nutritional environment during development and even prior to conception may contribute to cardiovascular risk. In mature adult female sheep, we investigated the effect of preconceptional and periconceptional maternal nutritional restriction on the vascular reactivity of arteries from four vascular beds supplying the heart, thorax, kidney and hindlimb. Welsh Mountain ewes received 100% of nutrient requirements throughout gestation (control group, C, n= 18), or 50% of nutrient requirements for 30 days prior to conception (preconceptional group, PRE, n= 20) or for 15 days either side of conception (periconceptional group, PERI, n= 31) and 100% thereafter. In 3.5‐year‐old female offspring, the left anterior descending coronary (LAD), left internal thoracic (LITA), right renal and second and third order femoral arteries were dissected and their reactivity was assessed by organ bath or wire myography. Vasoconstrictor responses were greater in both LAD and LITA from PERI offspring compared with C (P < 0.01), while vasoconstriction was unaffected by maternal diet in arteries from the renal and femoral circulations (P= n.s.). Endothelium‐dependent and ‐independent vasodilatation was attenuated in third order femoral arteries of PRE and PERI groups compared with C (P < 0.05). Endothelium‐independent vasodilatation was attenuated in both the LAD and renal arteries in the PERI group compared with C (P < 0.05). These data show that moderate maternal undernutrition either prior to or around conception affects vascular function of adult offspring. The effect depends on the timing of the insult, but also on the vascular bed studied and vessel hierarchy in the vascular tree.
Voluntary activation of human knee extensors measured using transcranial magnetic stimulation
Authors: Goodall S. , Romer L. M. , Ross E. Z.
Published: June 2009

The aim of this study was to determine the applicability and reliability of a transcranial magnetic stimulation twitch interpolation technique for measuring voluntary activation of a lower limb muscle group. Cortical voluntary activation of the knee extensors was determined in nine healthy men on two separate visits by measuring superimposed twitch torques evoked by transcranial magnetic stimulation during isometric knee extensions of varying intensity. Superimposed twitch amplitude decreased linearly with increasing voluntary torque between 50 and 100% of mean maximal torque, allowing estimation of resting twitch amplitude and subsequent calculation of voluntary activation. There were no systematic differences for maximal voluntary activation within day (mean ±s.d. 90.9 ± 6.2 versus 90.7 ± 5.9%; P= 0.98) or between days (90.8 ± 6.0 versus 91.2 ± 5.7%; P= 0.92). Systematic bias and random error components of the 95% limits of agreement were 0.23 and 9.3% within day versus−0.38 and 7.5% between days. Voluntary activation was also determined immediately after a 2 min maximal voluntary isometric contraction; in four of these subjects, voluntary activation was determined 30 min after the sustained contraction. Immediately after the sustained isometric contraction, maximal voluntary activation was reduced from 91.2 ± 5.7 to 74.2 ± 12.0% (P < 0.001), indicating supraspinal fatigue. After 30 min, voluntary activation had recovered to 85.4 ± 8.8% (P= 0.39 versus baseline). These results demonstrate that transcranial magnetic stimulation enables reliable measurement of maximal voluntary activation and assessment of supraspinal fatigue of the knee extensors.
Metabolic Capacity of
Sinorhizobium
(
Ensifer
)
meliloti
Strains as Determined by Phenotype MicroArray Analysis
Authors: Biondi Emanuele G. , Tatti Enrico , Comparini Diego , Giuntini Elisa , Mocali Stefano , Giovannetti Luciana , Bazzicalupo Marco , Mengoni Alessio , Viti Carlo
Published: June 2009

Sinorhizobium meliloti
is a soil bacterium that fixes atmospheric nitrogen in plant roots. The high genetic diversity of its natural populations has been the subject of extensive analysis. Recent genomic studies of several isolates revealed a high content of variable genes, suggesting a correspondingly large phenotypic differentiation among strains of
S. meliloti
. Here, using the Phenotype MicroArray (PM) system, hundreds of different growth conditions were tested in order to compare the metabolic capabilities of the laboratory reference strain Rm1021 with those of four natural
S. meliloti
isolates previously analyzed by comparative genomic hybridization (CGH). The results of PM analysis showed that most phenotypic differences involved carbon source utilization and tolerance to osmolytes and pH, while fewer differences were scored for nitrogen, phosphorus, and sulfur source utilization. Only the variability of the tested strain in tolerance to sodium nitrite and ammonium sulfate of pH 8 was hypothesized to be associated with the genetic polymorphisms detected by CGH analysis. Colony and cell morphologies and the ability to nodulate
Medicago truncatula
plants were also compared, revealing further phenotypic diversity. Overall, our results suggest that the study of functional (phenotypic) variability of
S. meliloti
populations is an important and complementary step in the investigation of genetic polymorphism of rhizobia and may help to elucidate rhizobial evolutionary dynamics, including adaptation to diverse environments.
Evaluation of Different Methods for Extracting Extracellular DNA from the Biofilm Matrix
Authors: Wu Jianfeng , Xi Chuanwu
Published: June 2009

The occurrence of high concentrations of extracellular DNA (eDNA) in the extracellular matrices of biofilms plays an important role in biofilm formation and development and possibly in horizontal gene transfer through natural transformation. Studies have been conducted to characterize the nature of eDNA and its potential function in biofilm development, but it is difficult to extract eDNA from the extracellular matrices of biofilms without any contamination from genomic DNA released by cell lysis during the extraction process. In this report, we compared several different extraction methods in order to obtain highly pure eDNA from different biofilm samples. After different extraction methods were explored, it was concluded that using chemical treatment or enzymatic treatment of biofilm samples may obtain larger amounts of eDNA than using the simple filtration method. There was no detectable cell lysis when the enzymatic treatment methods were used, but substantial cell lysis was observed when the chemical treatment methods were used. These data suggest that eDNA may bind to other extracellular polymers in the biofilm matrix and that enzymatic treatment methods are effective and favorable for extracting eDNA from biofilm samples. Moreover, randomly amplified polymorphic DNA analysis of eDNA in
Acinetobacter
sp. biofilms and
Acinetobacter
sp. genomic DNA and DNA sequencing analysis revealed that eDNA originated from genomic DNA but was not structurally identical to the genomic DNA.
Monochloramine Disinfection Kinetics of
Nitrosomonas europaea
by Propidium Monoazide Quantitative PCR and Live/Dead BacLight Methods
Authors: Wahman David G. , Wulfeck-Kleier Karen A. , Pressman Jonathan G.
Published: June 2009

Monochloramine disinfection kinetics were determined for the pure-culture ammonia-oxidizing bacterium
Nitrosomonas europaea
(ATCC 19718) by two culture-independent methods, namely, Live/Dead BacLight (LD) and propidium monoazide quantitative PCR (PMA-qPCR). Both methods were first verified with mixtures of heat-killed (nonviable) and non-heat-killed (viable) cells before a series of batch disinfection experiments with stationary-phase cultures (batch grown for 7 days) at pH 8.0, 25°C, and 5, 10, and 20 mg Cl
2
/liter monochloramine. Two data sets were generated based on the viability method used, either (i) LD or (ii) PMA-qPCR. These two data sets were used to estimate kinetic parameters for the delayed Chick-Watson disinfection model through a Bayesian analysis implemented in WinBUGS. This analysis provided parameter estimates of 490 mg Cl
2
-min/liter for the lag coefficient (
b
) and 1.6 × 10
−3
to 4.0 × 10
−3
liter/mg Cl
2
-min for the Chick-Watson disinfection rate constant (
k
). While estimates of
b
were similar for both data sets, the LD data set resulted in a greater
k
estimate than that obtained with the PMA-qPCR data set, implying that the PMA-qPCR viability measure was more conservative than LD. For
N. europaea
, the lag phase was not previously reported for culture-independent methods and may have implications for nitrification in drinking water distribution systems. This is the first published application of a PMA-qPCR method for disinfection kinetic model parameter estimation as well as its application to
N. europaea
or monochloramine. Ultimately, this PMA-qPCR method will allow evaluation of monochloramine disinfection kinetics for mixed-culture bacteria in drinking water distribution systems.
Dead-End Hollow-Fiber Ultrafiltration for Recovery of Diverse Microbes from Water
Authors: Smith Carmela M. , Hill Vincent R.
Published: June 2009

Dead-end ultrafiltration (DEUF) is an alternative approach to tangential-flow hollow-fiber ultrafiltration that can be readily employed under field conditions to recover microbes from water. The hydraulics of DEUF and microbe recovery for a new DEUF method were investigated using 100-liter tap water samples. Pressure, flow rate, and temperature were investigated using four hollow-fiber ultrafilter types. Based on hydraulic performance, the Asahi Kasei REXEED 25S ultrafilter was selected for microbe recovery experiments. Microbe recovery experiments were performed using MS2 bacteriophage,
Enterococcus faecalis
,
Clostridium perfringens
spores, and
Cryptosporidium parvum
oocysts. Microbes were recovered from ultrafilters by backflushing using a surfactant solution. Average flow rates were 2.1 liters/min for 100-liter water samples having turbidities of 0.28 to 4.3 nephelometric turbidity units (NTU), and no evidence of appreciable filter clogging was observed. The DEUF average recovery efficiencies for each study analyte in tap water were as follows: for
E. faecalis
, 93% ± 16%; for MS2, 57% ± 7.7%; for
C. perfringens
spores, 94% ± 22%; and for
C. parvum
, 87% ± 18%. Average microbe recoveries for tap water amended with surface water (average turbidity = 4.3 NTU) were as follows: for
E. faecalis
, 78% ± 12%; for MS2, 73% ± 13%; for
C. perfringens
, 57% ± 21%; and for
C. parvum
, 83% ± 21%. These data demonstrate that DEUF is an effective method for recovering diverse microbes from water and should be a useful tool for field-based environmental investigations.
Adsorption ofMycobacterium aviumsubsp.paratuberculosisto Soil Particles
Authors: Dhand Navneet K. , Toribio Jenny-Ann L. M. L. , Whittington Richard J.
Published: June 2009

Attachment ofMycobacterium aviumsubsp.paratuberculosisto soil particles could increase their availability to farm animals, as well as influence the transportation ofM. aviumsubsp.paratuberculosisto water sources. To investigate the possibility of such attachment, we passed a known quantity ofM. aviumsubsp.paratuberculosisthrough chromatography columns packed with clay soil, sandy soil, pure silica, clay-silica mixture, or clay-silica complexes and measured the organisms recovered in the eluent using culture or quantitative PCR. Experiments were repeated using buffer at a range of pH levels with pure silica to investigate the effect of pH onM. aviumsubsp.paratuberculosisattachment. Linear mixed-model analyses were conducted to compare the proportional recovery ofM. aviumsubsp.paratuberculosisin the eluent between different substrates and pH levels. Of the organisms added to the columns, 83 to 100% were estimated to be retained in the columns after adjustment for those retained in empty control columns. The proportions recovered were significantly different across different substrates, with the retention being significantly greater (P< 0.05) in pure substrates (silica and clay-silica complexes) than in soil substrates (clay soil and sandy soil). However, there were no significant differences in the retention ofM. aviumsubsp.paratuberculosisbetween silica and clay-silica complexes or between clay soil and sandy soil. The proportion retained decreased with increasing pH in one of the experiments, indicating greater adsorption ofM. aviumsubsp.paratuberculosisto soil particles at an acidic pH (P< 0.05). The results suggest that under experimental conditionsM. aviumsubsp.paratuberculosisadsorbs to a range of soil particles, and this attachment is influenced by soil pH.
Comparison of Species Richness Estimates Obtained Using Nearly Complete Fragments and Simulated Pyrosequencing-Generated Fragments in 16S rRNA Gene-Based Environmental Surveys
Authors: Youssef Noha , Sheik Cody S. , Krumholz Lee R. , Najar Fares Z. , Roe Bruce A. , Elshahed Mostafa S.
Published: June 2009

Pyrosequencing-based 16S rRNA gene surveys are increasingly utilized to study highly diverse bacterial communities, with special emphasis on utilizing the large number of sequences obtained (tens to hundreds of thousands) for species richness estimation. However, it is not yet clear how the number of operational taxonomic units (OTUs) and, hence, species richness estimates determined using shorter fragments at different taxonomic cutoffs correlates with the number of OTUs assigned using longer, nearly complete 16S rRNA gene fragments. We constructed a 16S rRNA clone library from an undisturbed tallgrass prairie soil (1,132 clones) and used it to compare species richness estimates obtained using eight pyrosequencing candidate fragments (99 to 361 bp in length) and the nearly full-length fragment. Fragments encompassing the V1 and V2 (V1+V2) region and the V6 region (generated using primer pairs 8F-338R and 967F-1046R) overestimated species richness; fragments encompassing the V3, V7, and V7+V8 hypervariable regions (generated using primer pairs 338F-530R, 1046F-1220R, and 1046F-1392R) underestimated species richness; and fragments encompassing the V4, V5+V6, and V6+V7 regions (generated using primer pairs 530F-805R, 805F-1046R, and 967F-1220R) provided estimates comparable to those obtained with the nearly full-length fragment. These patterns were observed regardless of the alignment method utilized or the parameter used to gauge comparative levels of species richness (number of OTUs observed, slope of scatter plots of pairwise distance values for short and nearly complete fragments, and nonparametric and parametric species richness estimates). Similar results were obtained when analyzing three other datasets derived from soil, adult Zebrafish gut, and basaltic formations in the East Pacific Rise. Regression analysis indicated that these observed discrepancies in species richness estimates within various regions could readily be explained by the proportions of hypervariable, variable, and conserved base pairs within an examined fragment.
Culture-Independent Characterization of Archaeal Biodiversity in Swine Confinement Building Bioaerosols
Authors: Nehmé Benjamin , Gilbert Yan , Létourneau Valérie , Forster Robert J. , Veillette Marc , Villemur Richard , Duchaine Caroline
Published: June 2009

It was previously demonstrated that microbial communities of pig manure were composed of both bacteria and archaea. Recent studies have shown that bacteria are aerosolized from pig manure, but none have ever focused on the airborne archaeal burden. We sought here to develop and apply molecular ecology approaches to thoroughly characterize airborne archaea from swine confinement buildings (SCBs). Eight swine operations were visited, twice in winter and once during summer. Institute of Occupational Medicine cassettes loaded with 25-mm gelatin filters were used to capture the inhalable microbial biomass. The total genomic DNA was extracted and used as a template for PCR amplification of the archaeal 16S rRNA gene. High concentrations of archaea were found in SCB bioaerosols, being as high as 10
8
16S rRNA gene copies per cubic meter of air. Construction and sequencing of 16S rRNA gene libraries revealed that all sequences were closely related to methanogenic archaea, such as
Methanosphaera stadtmanae
(94.7% of the archaeal biodiversity). Archaeal community profiles were compared by 16S rRNA gene denaturing gradient gel electrophoresis. This analysis showed similar fingerprints in each SCB and confirmed the predominance of methanogenic archaea in the bioaerosols. This study sheds new light on the nature of bioaerosols in SCBs and suggests that archaea are also aerosolized from pig manure.
Identification of a Novel Two-Peptide Lantibiotic, Lichenicidin, following Rational Genome Mining for LanM Proteins
Authors: Begley Máire , Cotter Paul D. , Hill Colin , Ross R. Paul
Published: June 2009

Lantibiotics are ribosomally synthesized peptide antimicrobials which contain considerable posttranslational modifications. Given their usually broad host range and their highly stable structures, there have been renewed attempts to identify and characterize novel members of the lantibiotic family in recent years. The increasing availability of bacterial genome sequences means that in addition to traditional microbiological approaches, in silico screening strategies may now be employed to the same end. Taking advantage of the highly conserved nature of lantibiotic biosynthetic enzymes, we screened publicly available microbial genome sequences for genes encoding LanM proteins, which are required for the posttranslational modification of type 2 lantibiotics. By using this approach, 89 LanM homologs, including 61 in strains not known to be lantibiotic producers, were identified. Of these strains, five (
Streptococcus pneumoniae
SP23-BS72,
Bacillus licheniformis
ATCC 14580,
Anabaena variabilis
ATCC 29413,
Geobacillus thermodenitrificans
NG80-2, and
Herpetosiphon aurantiacus
ATCC 23779) were subjected to a more detailed bioinformatic analysis. Four of the strains possessed genes potentially encoding a structural peptide in close proximity to the
lanM
determinants, while two,
S. pneumoniae
SP23-BS72 and
B. licheniformis
ATCC 14580, possess two potential structural genes. The
B. licheniformis
strain was selected for a proof-of-concept exercise, which established that a two-peptide lantibiotic, lichenicidin, which exhibits antimicrobial activity against all
Listeria monocytogenes
, methicillin-resistant
Staphylococcus aureus
, and vancomycin-resistant enterococcus strains tested, was indeed produced, thereby confirming the benefits of such a bioinformatic approach when screening for novel lantibiotic producers.
Metabolic Flux Analysis of
Escherichia coli creB
and
arcA
Mutants Reveals Shared Control of Carbon Catabolism under Microaerobic Growth Conditions
Authors: Nikel Pablo I. , Zhu Jiangfeng , San Ka-Yiu , Méndez Beatriz S. , Bennett George N.
Published: June 2009

Escherichia coli
has several elaborate sensing mechanisms for response to availability of oxygen and other electron acceptors, as well as the carbon source in the surrounding environment. Among them, the CreBC and ArcAB two-component signal transduction systems are responsible for regulation of carbon source utilization and redox control in response to oxygen availability, respectively. We assessed the role of CreBC and ArcAB in regulating the central carbon metabolism of
E. coli
under microaerobic conditions by means of
13
C-labeling experiments in chemostat cultures of a wild-type strain, Δ
creB
and Δ
arcA
single mutants, and a Δ
creB
Δ
arcA
double mutant. Continuous cultures were conducted at
D
= 0.1 h
−1
under carbon-limited conditions with restricted oxygen supply. Although all experimental strains metabolized glucose mainly through the Embden-Meyerhof-Parnas pathway, mutant strains had significantly lower fluxes in both the oxidative and the nonoxidative pentose phosphate pathways. Significant differences were also found at the pyruvate branching point. Both pyruvate-formate lyase and the pyruvate dehydrogenase complex contributed to acetyl-coenzyme A synthesis from pyruvate, and their activity seemed to be modulated by both ArcAB and CreBC. Strains carrying the
creB
deletion showed a higher biomass yield on glucose compared to the wild-type strain and its Δ
arcA
derivative, which also correlated with higher fluxes from building blocks to biomass. Glyoxylate shunt and lactate dehydrogenase were active mainly in the Δ
arcA
strain. Finally, it was observed that the tricarboxylic acid cycle reactions operated in a rather cyclic fashion under our experimental conditions, with reduced activity in the mutant strains.
In Vivo Domain-Based Functional Analysis of the Major Sporulation Sensor Kinase, KinA, inBacillus subtilis
Authors: Eswaramoorthy Prahathees , Guo Tao , Fujita Masaya
Published: June 2009

Sensor histidine kinases are widely used by bacteria to detect and respond to environmental signals. InBacillus subtilis, KinA is a major kinase providing phosphate input to the phosphorelay that activates the sporulation pathway upon starvation via the phosphorylated Spo0A transcription factor. KinA contains three PAS domains in its amino-terminal sensor domain, which appear to be involved in the sensing of an unidentified sporulation signal(s) produced upon starvation. Prior biochemical studies have suggested that KinA forms a homodimer as a functional enzyme and that the most amino-terminal PAS domain (PAS-A) plays an important role in sensing the signal(s) to activate an ATP-dependent autophosphorylation reaction to a histidine residue. To analyze the structure and function of the kinase in vivo, we have used a strain in which the synthesis of KinA is under the control of an isopropyl-β-d-thiogalactopyranoside (IPTG)-inducible promoter. In vivo functional studies in combination with domain-based deletion analysis show that the cytosolic KinA forms a homo-oligomer as an active form under both nutrient-rich and nutrient-depleted conditions via its amino- and carboxyl-terminal domains independently. Furthermore, we found that a mutant in which the PAS-A domain was deleted was still able to induce sporulation at a wild-type level irrespective of nutrient availability, suggesting that PAS-BC domains are sufficient to maintain the kinase activity. Based on these results, we propose that the primary role of the amino-terminal sensor domain is to form a stable complex as a functional kinase, but possibly not for the binding of an unidentified sporulation signal(s).
Production of 3-Nitrosoindole Derivatives by
Escherichia coli
during Anaerobic Growth
Authors: Kwon Young-Man , Weiss Bernard
Published: June 2009

When
Escherichia coli
K-12 is grown anaerobically in medium containing tryptophan and sodium nitrate, it produces red compounds. The reaction requires functional genes for trytophanase (
tnaA
), a tryptophan permease (
tnaB
), and a nitrate reductase (
narG
), as well as a natural drop in the pH of the culture. Mass spectrometry revealed that the purified chromophores had mass/charge ratios that closely match those for indole red, indoxyl red, and an indole trimer. These compounds are known products of chemical reactions between indole and nitrous acid. They are derived from an initial reaction of 3-nitrosoindole with indole. Apparently, nitrite that is produced from the metabolic reduction of nitrate is converted in the acid medium to nitrous acid, which leads to the nitrosation of the indole that is generated by tryptophanase. An
nfi
(endonuclease V) mutant and a
recA
mutant were selectively killed during the period of chromophore production, and a
uvrA
strain displayed reduced growth. These effects depended on the addition of nitrate to the medium and on tryptophanase activity in the cells. Unexpectedly, the killing of a
tnaA
+
nfi
mutant was not accompanied by marked increases in mutation frequencies for several traits tested. The vulnerability of three DNA repair mutants indicates that a nitrosoindole or a derivative of a nitrosoindole produces lethal DNA damage.
Two-step stimulation of intestinal Ca2+absorption during lactation by long-term prolactin exposure and suckling-induced prolactin surge
Authors: Charoenphandhu Narattaphol , Nakkrasae La-iad , Kraidith Kamonshanok , Teerapornpuntakit Jarinthorn , Thongchote Kanogwun , Thongon Narongrit , Krishnamra Nateetip
Published: July 2009

During pregnancy and lactation, the enhanced intestinal Ca2+absorption serves to provide Ca2+for fetal development and lactogenesis; however, the responsible hormone and its mechanisms remain elusive. We elucidated herein that prolactin (PRL) markedly stimulated the transcellular and paracellular Ca2+transport in the duodenum of pregnant and lactating rats as well as in Caco-2 monolayer in a two-step manner. Specifically, a long-term exposure to PRL in pregnancy and lactation induced an adaptation in duodenal cells at genomic levels by upregulating the expression of genes related to transcellular transport, e.g., TRPV5/6 and calbindin-D9k, and the paracellular transport, e.g., claudin-3, thereby raising Ca2+absorption rate to a new “baseline” (Step 1). During suckling, PRL surge further increased Ca2+absorption to a higher level (Step 2) in a nongenomic manner to match Ca2+loss in milk. PRL-enhanced apical Ca2+uptake was responsible for the increased transcellular transport, whereas PRL-enhanced paracellular transport required claudin-15, which regulated epithelial cation selectivity and paracellular Ca2+movement. Such nongenomic PRL actions were mediated by phosphoinositide 3-kinase, protein kinase C, and RhoA-associated coiled-coil-forming kinase pathways. In conclusion, two-step stimulation of intestinal Ca2+absorption resulted from long-term PRL exposure, which upregulated Ca2+transporter genes to elevate the transport baseline, and the suckling-induced transient PRL surge, which further increased Ca2+transport to the maximal capacity. The present findings also suggested that Ca2+supplementation at 15–30 min prior to breastfeeding may best benefit the lactating mother, since more Ca2+could be absorbed as a result of the suckling-induced PRL surge.
CORM-3-derived CO modulates polymorphonuclear leukocyte migration across the vascular endothelium by reducing levels of cell surface-bound elastase
Authors: Mizuguchi Shinjiro , Stephen Jancy , Bihari Relka , Markovic Nevena , Suehiro Shigefumi , Capretta Alfredo , Potter Richard F. , Cepinskas Gediminas
Published: June 2009

Recently, it has been shown that carbon monoxide (CO)-releasing molecule (CORM)-released CO can suppress inflammation. In this study, we assessed the effects and potential mechanisms of a ruthenium-based water-soluble CO carrier [tricarbonylchloroglycinate-ruthenium(II) (CORM-3)] in the modulation of polymorphonuclear leukocyte (PMN) inflammatory responses in an experimental model of sepsis. Sepsis in mice was induced by cecal ligation and puncture. CORM-3 (3 mg/kg iv) was administered 15 min after the induction of cecal ligation and puncture. PMN accumulation in the lung (myeloperoxidase assay), bronchoalveolar lavage (BAL) fluid, and lung vascular permeability (protein content in BAL fluid) were assessed 6 h later. In in vitro experiments, human PMNs were primed with LPS (10 ng/ml) and subsequently stimulated with formyl-methionyl-leucylphenylalanine (fMLP; 100 nM). PMN production of ROS (L-012/dihydrorhodamine-123 oxidation), degranulation (release of elastase), and PMN rolling, adhesion, and migration to/across human umbilical vein endothelial cells (HUVECs) were assessed in the presence or absence of CORM-3 (1–100 μM). The obtained results indicated that systemically administered CORM-3 attenuates PMN accumulation and vascular permeability in the septic lung. Surprisingly, in in vitro experiments, treatment of PMNs with CORM-3 further augmented LPS/fMLP-induced ROS production and the release of elastase. The latter effects, however, were accompanied by an inability of PMNs to mobilize elastase to the cell surface (plasma membrane), an event required for efficient PMN transendothelial migration. The CORM-3-induced decrease in cell surface levels of elastase was followed by decreased PMN rolling/adhesion to HUVECs and complete prevention of PMN migration across HUVECs. In contrast, treatment of HUVECs with CORM-3 had no effect on PMN transendothelial migration. Taken together, these findings indicate that, in sepsis, CORM3-released CO, while further amplifying ROS production and degranulation of PMNs, concurrently reduces the levels of cell surface-bound elastase, which contributes to suppressed PMN transendothelial migration.
Resveratrol blocks interleukin-18-EMMPRIN cross-regulation and smooth muscle cell migration
Authors: Venkatesan Balachandar , Valente Anthony J. , Reddy Venkatapuram Seenu , Siwik Deborah A. , Chandrasekar Bysani
Published: June 2009

Vascular smooth muscle cell (SMC) migration is an important mechanism in atherogenesis and postangioplasty arterial remodeling. Previously, we demonstrated that the proinflammatory cytokine interleukin (IL)-18 is a potent inducer of SMC migration. Since extracellular matrix metalloproteinase inducer (EMMPRIN) stimulates ECM degradation and facilitates cell migration, we investigated whether IL-18 and EMMPRIN regulate each other's expression, whether their cross talk induces SMC migration, and whether the phytoalexin resveratrol inhibits IL-18-EMMPRIN signaling and SMC migration. Our studies demonstrate that 1) IL-18 induces EMMPRIN mRNA and protein expressions and stimulates EMMPRIN secretion from human aortic SMCs; 2) IL-18 stimulates EMMPRIN expression via oxidative stress and phosphatidylinositol 3-kinase (PI3K)-Akt-ERK signaling; 3) IL-18-stimulated SMC migration is significantly blunted by EMMPRIN knockdown, EMMPRIN function-blocking antibodies, or adenoviral transduction of mutant EMMPRIN; 4) conversely, EMMPRIN stimulates IL-18 expression and secretion via PI3K, Akt, and ERK; and 5) resveratrol attenuates IL-18- and EMMPRIN-mediated PI3K, Akt, and ERK activations; blunts IL-18-mediated oxidative stress; blocks IL-18-EMMPRIN cross-regulation; and inhibits SMC migration. Collectively, our results demonstrate that the coexpression and regulation of IL-18 and EMMPRIN in the vessel wall may amplify the inflammatory cascade and promote atherosclerosis and remodeling. Resveratrol, via its antioxidant and anti-inflammatory properties, has the potential to inhibit the progression of atherosclerosis by blocking IL-18 and EMMPRIN cross-regulation and SMC migration.