bluuebunny/crossref_metadata_2025 huggingface dataset:
Systematic review on the use of omalizumab for the treatment of asthmatic children and adolescents
Authors: Rodrigo Gustavo J. , Neffen Hugo
Published: May 2015

Background

There are less data on omalizumab treatment in pediatric asthma than in adult population. Thus, to establish the efficacy and safety of subcutaneous omalizumab as an add‐on therapy, a systematic review of placebo‐controlled studies was performed.

Methods

Primary outcome was the frequency of asthma exacerbations. Secondary outcomes included spirometric measures, rescue medication use, asthma symptoms, health‐related quality of life, and adverse events.

Results

Three randomized controlled trials (1381 participants) fulfilled the selection criteria. During the stable phase, omalizumab decreased the number of patients with at least one significant asthma exacerbation (26.7% vs. 40.6%, NNTB = 7, 95% CI, 5, 11). The predefined post hoc subgroup analysis showed that duration of treatment did not influence this result. During the steroid reduction phase, omalizumab reduced the number of patients with at least one exacerbation (RR = 0.48, 95% CI, 0.38, 0.61; NNTB = 6, 95% CI, 4, 8) and also the mean number of asthma exacerbations per patient (MD = −0.44, 95% CI, −0.72, −0.17) when compared to placebo. The frequency of serious adverse events was similar between omalizumab (5.2%) and placebo (5.6%), and there were no evidence of increased risk of hypersensitivity reactions, nor malignant neoplasms.

Conclusions

Data indicate that the efficacy of an add‐on omalizumab in patients with moderate‐to‐severe allergic asthma uncontrolled with recommended inhaled steroid treatment is accompanied by an acceptable safety profile.
Intranasal dexmedetomidine for sedation in children undergoing transthoracic echocardiography study—a prospective observational study
Authors: Li Bi L. , Ni Jin , Huang Jun X. , Zhang Na , Song Xing R. , Yuen Vivian M.
Published: May 2015

Background

Intranasal dexmedetomidine has been used for sedation in children undergoing nonpainful procedures.

Objective

The aim of this study was to determine the success rate of intranasal dexmedetomidine sedation for children undergoing transthoracic echocardiography examination.

Methods

This was a prospective observational study of 115 children under the age of 3 years undergoing echocardiography examination under sedation with intranasal dexmedetomidine at 3 mcg·kg−1.

Results

Of the 115 children, 100 (87%) had satisfactory sedation with intranasal dexmedetomidine. The mean onset time was 16.7 ± 7 min (range 5–50 min). The mean wake up time was 44.3 ± 15.1 min (range 12–123 min). The wake up time was significantly correlated with duration of procedure with R = 0.540 (P < 0.001). Aside from one patient who required oxygen supplementation, all children in this investigation had an acceptable heart rate and blood pressure and required no medical intervention.

Conclusion

Sedation by intranasal dexmedetomidine at 3 mcg·kg−1 is associated with acceptable success rate in children undergoing echocardiography with no adverse events in this cohort.
A physiological and biophysical model of coppice willow (Salix spp.) production yields for the contiguous USA in current and future climate scenarios
Authors: WANG DAN , JAISWAL DEEPAK , LEBAUER DAVID S. , WERTIN TIMOTHY M. , BOLLERO GERMÁN A. , LEAKEY ANDREW D. B. , LONG STEPHEN P.
Published: May 2015

High‐performance computing has facilitated development of biomass production models that capture the key mechanisms underlying production at high spatial and temporal resolution. Direct responses to increasing [CO2] and temperature are important to long‐lived emerging woody bioenergy crops. Fast‐growing willow (Salix spp.) within short rotation coppice (SRC) has considerable potential as a renewable biomass source, but performance over wider environmental conditions and under climate change is uncertain. We extended the bioenergy crop modeling platform, BioCro, to SRC willow by adding coppicing and C3 photosynthesis subroutines, and modifying subroutines for perennation, allocation, morphology, phenology and development. Parameterization with measurements of leaf photosynthesis, allocation and phenology gave agreement of modeled with measured yield across 23 sites in Europe and North America. Predictions for the continental USA suggest yields of ≥17 Mg ha−1 year−1 in a 4 year rotation. Rising temperature decreased predicted yields, an effect partially ameliorated by rising [CO2]. This model, based on over 100 equations describing the physiological and biophysical mechanisms underlying production, provides a new framework for utilizing mechanism of plant responses to the environment, including future climates. As an open‐source tool, it is made available here as a community resource for further application, improvement and adaptation.
A systematic review of acute kidney injury in pediatric allogeneic hematopoietic stem cell recipients
Authors: Didsbury Madeleine S. , Mackie Fiona E. , Kennedy Sean E.
Published: May 2015

The process of allogeneic HSCT in children is associated with frequent AKI and mortality, but the epidemiology is not widely reported. The aim of this review was to summarize the available evidence on incidence, risk factors, timing, and prognosis of AKI in children following HSCT. We systematically reviewed all observational studies reporting incidence and outcomes of AKI in pediatric allogenic HSCT recipients. The minimum criteria for AKI were defined as an increase in sCr  ≥ x1.5 or urine output ≤0.5 mL/kg/min over six h. Medline and Embase were searched until March 2014. From 993 electronic records, five were eligible for inclusion (n = 571 patients). The average incidence of AKI within the first 100 days following HSCT was 21.7% (range 11–42%), and the average time of onset was 4–6 wk post‐transplant. Risk factors for AKI included cyclosporine toxicity, amphotericin B and foscarnet, SOS, and having a mismatched donor. There were conflicting reports on whether AKI was associated with the development of CKD. AKI is a common and potentially life‐threatening complication following HSCT in children. Further quality observational studies are needed to accurately determine the epidemiology and prognosis of AKI in this population.
Sequential Quasi Monte Carlo
Authors: Gerber Mathieu , Chopin Nicolas
Published: May 2015

We derive and study sequential quasi Monte Carlo (SQMC), a class of algorithms obtained by introducing QMC point sets in particle filtering. SQMC is related to, and may be seen as an extension of, the array-RQMC algorithm of L'Ecuyer and his colleagues. The complexity of SQMC is O{Nlog(N)}, where N is the number of simulations at each iteration, and its error rate is smaller than the Monte Carlo rate OP(N−1/2). The only requirement to implement SQMC algorithms is the ability to write the simulation of particle xtn given xt−1n as a deterministic function of xt−1n and a fixed number of uniform variates. We show that SQMC is amenable to the same extensions as standard SMC, such as forward smoothing, backward smoothing and unbiased likelihood evaluation. In particular, SQMC may replace SMC within a particle Markov chain Monte Carlo algorithm. We establish several convergence results. We provide numerical evidence that SQMC may significantly outperform SMC in practical scenarios.
Tuberculosis infection control measures in diabetes clinics in China: a rapid assessment of 10 hospitals
Authors: Lin Yan , Harries Anthony D.
Published: May 2015

Objective

To assess tuberculosis infection control measures in diabetes mellitus (DM) clinics in China.

Method

An evaluation questionnaire was developed based on the measures outlined in the WHO policy on TB infection control in healthcare facilities and congregate settings. Ten selected hospitals were assessed between September 2014 and February 2015.

Results

All hospitals had an infection control committee, an action plan and training, but there was no focus on activities to prevent airborne transmission of infection, especially from tuberculosis. All DM clinics had natural or artificial ventilation, but at the time of the evaluation half had all windows closed. While all the hospitals provided surgical masks for hospital staff, none had N95‐specific respiratory masks and only three provided masks for patients with a cough. There were no policies on identifying DM patients with TB symptoms, minimising the time spent by these patients in the clinics or developing health educational material on diabetes and tuberculosis.

Conclusion

Infection control measures to prevent airborne transmission in DM clinics are inadequate. More work is needed to better understand and determine the risk of TB infection in DM clinics and to improve their TB infection control practices.
Medication Adherence Clubs: a potential solution to managing large numbers of stable patients with multiple chronic diseases in informal settlements
Authors: Khabala Kelly B. , Edwards Jeffrey K. , Baruani Bienvenu , Sirengo Martin , Musembi Phylles , Kosgei Rose J. , Walter Kizito , Kibachio Joseph M. , Tondoi Monique , Ritter Helga , Wilkinson Ewan , Reid Tony
Published: May 2015

Objectives

To assess the care of hypertension, diabetes mellitus and/or HIV patients enrolled into Medication Adherence Clubs (MACs).

Methods

Retrospective descriptive study was carried out using routinely collected programme data from a primary healthcare clinic at informal settlement in Nairobi, Kenya. All patients enrolled into MACs were selected for the study. MACs are nurse‐facilitated mixed groups of 25–35 stable hypertension, diabetes mellitus and/or HIV patients who met quarterly to confirm their clinical stability, have brief health discussions and receive medication. Clinical officer reviewed MACs yearly, when a patient developed complications or no longer met stable criteria.

Results

A total of 1432 patients were enrolled into 47 clubs with 109 sessions conducted between August 2013 and August 2014. There were 1020 (71%) HIV and 412 (29%) non‐communicable disease patients. Among those with NCD, 352 (85%) had hypertension and 60 (15%) had DM, while 12 had HIV concurrent with hypertension. A total of 2208 consultations were offloaded from regular clinic. During MAC attendance, blood pressure, weight and laboratory testing were completed correctly in 98–99% of consultations. Only 43 (2%) consultations required referral for clinical officer review before their routine yearly appointment. Loss to follow‐up from the MACs was 3.5%.

Conclusions

This study demonstrates the feasibility and early efficacy of MACs for mixed chronic disease in a resource‐limited setting. It supports burden reduction and flexibility of regular clinical review for stable patients. Further assessment regarding long‐term outcomes of this model should be completed to increase confidence for deployment in similar contexts.
Traditional vs. non‐traditional healing for minor and major morbidities in India: uses, cost and quality comparisons
Authors: Singh Ashish , Madhavan Harilal
Published: May 2015

Objectives

To examine the uses, cost and quality of care of traditional healing for short‐term morbidities and major morbidities in India and to compare them with the non‐traditional healing.

Methods

We used data from a nationally representative survey, the India Human Development Survey (2004‐2005) and descriptive as well as bivariate analyses for the examination.

Results

Use of traditional healing is much less common than use of non‐traditional healing in both rural and urban areas and across all socio‐economic and demographic characteristics; it is slightly more common in rural than urban areas for short‐term morbidities. Use of traditional healing is relatively more frequent for cataract (especially in rural areas), leprosy, asthma, polio, paralysis, epilepsy and mental illnesses; its total cost of care and mean waiting time (in the health facility) are substantially lower than for non‐traditional healing. Among patients who use both traditional and non‐traditional healing, a relatively higher proportion use traditional healing complemented by non‐traditional healing for short‐term illnesses, but vice versa for major morbidities.

Conclusion

This is the first study which has investigated at the national level the uses, complementarities, cost and quality aspects of traditional and non‐traditional healing in India. Traditional healing is more affordable and pro‐poor. Relatively higher use of traditional healing in patients from poorly educated as well as poor households and suffering from diseases, such as, epilepsy and mental illnesses; and higher demand for traditional healing for the above diseases highlight the need for research/policy reorientation in India.
Deep pyoderma caused by Burkholderia cepacia complex associated with ciclosporin administration in dogs: a case series
Authors: Banovic Frane , Koch Sandra , Robson David , Jacob Megan , Olivry Thierry
Published: May 2015

Background

Bacteria of the Burkholderia cepacia complex (Bcc) are ubiquitous Gram‐negative bacilli associated with fatal nosocomial infections in humans; multi‐antibiotic resistance makes this organism a serious threat in hospital settings.

Objective

To describe the historical, clinicopathological and treatment characteristics of Bcc‐associated deep skin infections in dogs.

Animals

Six dogs with skin infections in which skin bacterial cultures resulted in pure growth of Bcc.

Methods

Retrospective study with review of medical records and skin biopsies.

Results

All dogs were receiving oral ciclosporin at the time of skin infection development. All dogs were castrated males and four of six were West Highland white terriers. Cutaneous lesions consistent with deep pyoderma were confined mainly to the trunk. In all dogs skin cytology revealed a strong inflammatory response, with moderate to abundant numbers of intracellular (neutrophils and macrophages) and extracellular bacilli. In three dogs histopathology showed a multifocal, nodular to coalescing pyogranulomatous dermatitis associated with multifocal folliculitis and furunculosis. Tissue Giemsa and Gram stains identified numerous Gram‐negative rods within macrophages. Antimicrobial susceptibility testing revealed multidrug‐resistant Bcc strains with sensitivity to trimethoprim/sulfonamides in all dogs and to marbofloxacin, piperacillin and ceftazidime in three dogs. Successful treatment was achieved in all dogs using trimethoprim/sulfonamides or quinolones (marbofloxacin, ciprofloxacin) or doxycycline in conjunction with ciclosporin withdrawal.

Conclusions and clinical importance

Clinicians should be aware of the rare potential for Bcc‐associated deep skin infections in dogs receiving oral ciclosporin. Owners should be made conscious of the potential transmission risk to humans or other animals.
Modelling the Phenological Relationships of Questing Immature Ixodes Ricinus (Ixodidae) Using Temperature and NDVI Data
Authors: Alonso‐Carné J. , García‐Martín A. , Estrada‐Peña A.
Published: May 2015

All active stages of the tick Ixodes ricinus were collected monthly at two sites in northern Spain between the years 2000 and 2007. We used percentile accumulation of the active stage in the environment to evaluate simple and coherent correlations between accumulation of the active stages of larvae and nymphs and medium‐resolution MODIS satellite‐derived information on the climate, including monthly and accumulated temperature and the Normalized Difference Vegetation Index (NDVI). This framework is not intended to predict the actual abundance of ticks in the field as a measure of the hazard to humans, but to provide a basic structure for addressing the phenology of the tick in its geographic range. We demonstrated that the accumulation of larval ticks in the active stage is a sigmoid function of the accumulated temperature from the beginning of the calendar year. We also demonstrated that the accumulated temperature necessary to recruit nymphs from the questing larval stage is a function of the changes in accumulated larvae and nymphs and the accumulated temperature and NDVI recorded by the Aqua sensor. The low p‐values obtained in the regressions confirmed that such recruitment can be calculated using time intervals to estimate, for example, the beginning of the questing period or the time of the year when a population peak can be expected. The comparison among predicted and actual accumulated temperatures between larvae and nymph recruitment had an averaged error of ±20 days in one complete year. The use of accumulated temperature and NDVI proposed in this study opens up the re‐evaluation of reports on the phenology of the tick in Europe. This framework is intended to evaluate the same correlations along the tick's range and predict its phenological patterns in areas of pathogen transmission risk for humans.
Magnitude of muscle wasting early after on‐pump coronary artery bypass graft surgery and exploration of aetiology
Authors: Hansen Dominique , Linsen Loes , Verboven Kenneth , Hendrikx Marc , Rummens Jean‐Luc , van Erum Monique , Eijnde Bert O. , Dendale Paul
Published: May 2015

New Findings

What is the central question of this study?

It remains uncertain whether significant fat‐free mass wasting occurs early after coronary artery bypass graft surgery, and the aetiology of this wasting in these particular conditions is unexplored.

What is the main finding and its importance?

Significant fat‐free mass wasting is present after coronary artery bypass graft surgery, and this wasting effect is greater in younger patients and in patients with greater increments in blood cortisol‐to‐testosterone ratios after surgery.

The magnitude and aetiology of muscle wasting early after coronary artery bypass graft (CABG) surgery remains unknown. In the present study, we assessed changes in fat‐free mass early after CABG surgery and explored the possible aetiology (relationships with postsurgical changes in blood hormones, insulin resistance, subject characteristics and inflammation) for these changes. Fat‐free mass was assessed before and 23 (range: 25) days after CABG surgery in 25 subjects. Blood testosterone, cortisol, insulin‐like growth factor‐1, growth hormone, sex hormone‐binding globulin, glucose, insulin, C‐peptide and C‐reactive protein concentrations were determined, and free androgen index, cortisol‐to‐testosterone ratio and HOMA‐IR index were all calculated before surgery, during the first 3 days after surgery and at reassessment of body composition. Relationships between changes in fat‐free mass and changes in blood parameters after surgery or subject characteristics were studied. After surgery, free androgen index and blood sex hormone‐binding globulin, testosterone and insulin‐like growth factor‐1 concentrations decreased significantly, while HOMA‐IR index, cortisol‐to‐testosterone ratio, blood growth hormone, insulin and C‐reactive protein concentrations increased significantly (P < 0.0025, observed α > 0.80). Whole‐body fat‐free mass decreased significantly [by −1.9 (range: 9.1) kg, P < 0.0025, observed α = 0.99] after surgery. According to regression analysis, greater absolute loss of fat‐free mass was observed after CABG surgery in subjects who were younger, who experienced a greater increase in blood cortisol‐to‐testosterone ratio after surgery and/or who underwent earlier reassessment of body composition (P < 0.05). Significant decrements in fat‐free mass were observed early after CABG surgery, especially in younger subjects and/or subjects with elevated blood cortisol‐to‐testosterone ratios after surgery. Interventions to preserve fat‐free mass soon after CABG surgery are thus warranted.
Short‐latency crossed responses in the human biceps femoris muscle
Authors: Stevenson Andrew J. T. , Kamavuako Ernest N. , Geertsen Svend S. , Farina Dario , Mrachacz‐Kersting Natalie
Published: May 2015

Key points

The present study is the first to show short‐latency crossed‐spinal reflexes in the human upper leg muscles following mechanical rotations to the ipsilateral knee (iKnee) joint.

The short‐latency reflex in the contralateral biceps femoris (cBF) was inhibitory following iKnee extension perturbations, and facilitatory following iKnee flexion perturbations. The onset latency was 44 ms, indicating that purely spinal pathways mediate the cBF reflexes.

The short‐latency cBF inhibitory and facilitatory reflexes followed the automatic gain control principle, becoming larger as the level of background contraction in the cBF increased.

The short‐latency cBF reflexes were observed at the motor unit level using i.m. electromyography recordings, and the same population of cBF motor units that was inhibited following iKnee extensions was facilitated following iKnee flexions.

Parallel interneuronal pathways from ipsilateral afferents to common motoneurons in the contralateral leg can therefore probably explain the perturbation direction‐dependent reversal in the sign of the short‐latency cBF reflex.

Interlimb reflexes contribute to the central neural co‐ordination between different limbs in both humans and animals. Although commissural interneurons have only been directly identified in animals, spinally‐mediated interlimb reflexes have been discovered in a number of human lower limb muscles, indicating their existence in humans. The present study aimed to investigate whether short‐latency crossed‐spinal reflexes are present in the contralateral biceps femoris (cBF) muscle following ipsilateral knee (iKnee) joint rotations during a sitting task, where participants maintained a slight pre‐contraction in the cBF. Following iKnee extension joint rotations, an inhibitory reflex was observed in the surface electromyographic (EMG) activity of the cBF, whereas a facilitatory reflex was observed in the cBF following iKnee flexion joint rotations. The onset latency of both cBF reflexes was 44 ms, which is too fast for a transcortical pathway to contribute. The cBF inhibitory and facilitatory reflexes followed the automatic gain control principle, with the size of the response increasing as the level of background pre‐contraction in the cBF muscle increased. In addition to the surface EMG, both short‐latency inhibitory and facilitatory cBF reflexes were recorded directly at the motor unit level by i.m. EMG, and the same population of cBF motor units that were inhibited following iKnee extension joint rotations were facilitated following iKnee flexion joint rotations. Therefore, parallel interneuronal pathways (probably involving commissural interneurons) from ipsilateral afferents to common motoneurons in the contralateral leg can probably explain the perturbation direction‐dependent reversal in the sign of the short‐latency cBF reflex.
A Bio-Inspired Adaptive Control Compensation System for an Aircraft Outside Bounds of Nominal Design
Authors: Perez Andres E. , Moncayo Hever , Perhinschi Mario , Al Azzawi Dia , Togayev Adil
Published: May 2015

This paper presents a novel bio-inspired adaptive control technique that has been designed to maintain the performance of an aircraft under upset conditions. The proposed control approach is inspired by biological principles that govern the humoral response of the immune system of living organisms and is intended to reduce pilot effort while maintaining adequate aircraft operation outside bounds of nominal design. The immunity-based control parameters are optimized offline for multiple sets of failures using a genetic algorithm approach. The performance of the immunity-based augmentation is compared with a neural network (NN)-based augmentation. Different piloted tests were performed on a six degrees-of-freedom (6DOF) motion-based simulator for different types of maneuvers under several flight conditions. The results show that the artificial immune system (AIS) proposed scheme improves the aircraft handling qualities by reducing the tracking errors (TEs) and improving the pilot response required to maintain control of the aircraft under upset conditions.
Barriers to Physical Activity in a Mass Transit Population: A Qualitative Study
Authors: Das Bhibha M. , Petruzzello Steven J.
Published: May 2015

Background:

The physical inactivity epidemic continues be one of the greatest public health challenges in contemporary society in the United States. The transportation industry is at greater risk of physical inactivity, compared with individuals in other sectors of the workforce. The aim of this study was to use the Nominal Group Technique, a focus group technique, to examine mass transit employees’ perceptions of the barriers to physical activity at their worksite.

Methods:

Three focus groups (n = 31) were conducted to examine mass transit employees’ perceptions of barriers to physical activity at the worksite.

Results:

Salient barriers included (1) changing work schedules, (2) poor weather conditions, and (3) lack of scheduled and timely breaks.

Conclusions:

Findings were consistent with previous research demonstrating shift work, poor weather, and lack of breaks can negatively impact mass transit employees’ ability to be physically active. Although physical activity barriers for this population have been consistent for the last 20 years, public health practice and policy have not changed to address these barriers. Future studies should include conducing focus groups stratified by job classification (eg, operators, maintenance, and clerical) along with implementing and evaluating worksite-based physical activity interventions and policy changes.
EGFR-activated Src family kinases maintain GAB1-SHP2 complexes distal from EGFR
Authors: Furcht Christopher M. , Buonato Janine M. , Lazzara Matthew J.
Published: May 2015

Activation of Src family kinases enables ligand-initiated signaling to persist in the cytoplasm after the receptor is turned off.
Discrete PTEN signaling
Authors: Prabakaran Sudhakaran
Published: May 2015

The phosphatase PTEN must be localized to cytoplasmic vesicles to oppose PI3K signaling.
Epigenetically programming gender identity
Authors: Ferrarelli Leslie K.
Published: May 2015

Gender identity in the brain is regulated by DNA methylation during a short time around birth.
Optimization of Methanotrophic Growth and Production of Poly(3-Hydroxybutyrate) in a High-Throughput Microbioreactor System
Authors: Sundstrom Eric R. , Criddle Craig S.
Published: May 2015

Production of poly(3-hydroxybutyrate) (P3HB) from methane has economic and environmental advantages over production by agricultural feedstock. Identification of high-productivity strains and optimal growth conditions is critical to efficient conversion of methane to polymer. Current culture conditions, including serum bottles, shake flasks, and agar plates, are labor-intensive and therefore insufficient for systematic screening and isolation. Gas chromatography, the standard method for analysis of P3HB content in bacterial biomass, is also incompatible with high-throughput screening. Growth in aerated microtiter plates coupled with a 96-well Nile red flow-cytometric assay creates an integrated microbioreactor system for high-throughput growth and analysis of P3HB-producing methanotrophic cultures, eliminating the need for individual manipulation of experimental replicates. This system was tested in practice to conduct medium optimization for P3HB production in pure cultures of
Methylocystis parvus
OBBP. Optimization gave insight into unexpected interactions: for example, low calcium concentrations significantly enhanced P3HB production under nitrogen-limited conditions. Optimization of calcium and copper concentrations in the growth medium increased final P3HB content from 18.1% to 49.4% and P3HB concentration from 0.69 g/liter to 3.43 g/liter while reducing doubling time from 10.6 h to 8.6 h. The ability to culture and analyze thousands of replicates with high mass transfer in completely mixed culture promises to streamline medium optimization and allow the detection and isolation of highly productive strains. Applications for this system are numerous, encompassing analysis of biofuels and other lipid inclusions, as well as analysis of heterotrophic and photosynthetic systems.
Yersinia enterocolitica Isolates from Wild Boars Hunted in Lower Saxony, Germany
Authors: von Altrock Alexandra , Seinige Diana , Kehrenberg Corinna
Published: May 2015

Yersiniosis is strongly associated with the consumption of pork contaminated with enteropathogenic
Yersinia enterocolitica
, which is harbored by domestic pigs without showing clinical signs of disease. In contrast to data on
Y. enterocolitica
isolated from conventionally reared swine, investigations into the occurrence of
Y. enterocolitica
in wild boars in Germany are rare. The objectives of the study were to get knowledge about these bacteria and their occurrence in wild boars hunted in northern Germany by isolation of the bacteria from the tonsils, identification of the bioserotypes, determination of selected virulence factors, macrorestriction analysis, multilocus sequence typing (MLST), and testing of antimicrobial susceptibility. Altogether, tonsils from 17.1% of 111 tested wild boars were positive for
Y. enterocolitica
by culture methods. All but two isolates belonged to biotype (BT) 1A, with the majority of isolates bearing a
ystB
nucleotide sequence which was revealed to have 85% identity to internal regions of
Y. enterocolitica
heat-stable enterotoxin type B genes. The remaining
Y. enterocolitica
isolates were identified to be BT 1B and did not carry the virulence plasmid. However, two BT 1A isolates carried the
ail
gene. Macrorestriction analysis and results from MLST showed a high degree of genetic diversity of the isolates, although the region where the samples were taken was restricted to Lower Saxony, Germany, and wild boars were shot during one hunting season. In conclusion, most
Y. enterocolitica
isolates from wild boars investigated in this study belonged to biotype 1A. Enteropathogenic
Y. enterocolitica
bioserotypes 4/O:3 and 2/O:9, usually harbored by commercially raised pigs in Europe, could not be identified.
Depth-Dependent Survival of Escherichia coli and Enterococci in Soil after Manure Application and Simulated Rainfall
Authors: Stocker M. D. , Pachepsky Y. A. , Hill R. L. , Shelton D. R.
Published: May 2015

Once released, manure-borne bacteria can enter runoff via interaction with the thin mixing layer near the soil surface. The objectives of this work were to document temporal changes in profile distributions of manure-borne
Escherichia coli
and enterococci in the near-surface soil layers after simulated rainfalls and to examine differences in survival of the two fecal indicator bacteria. Rainfall simulations were performed in triplicate on soil-filled boxes with grass cover and solid manure application for 1 h with rainfall depths of 30, 60, and 90 mm. Soil samples were collected weekly from depth ranges of 0 to 1, 1 to 2, 2 to 5, and 5 to 10 cm for 1 month. Rainfall intensity was found to have a significant impact on the initial concentrations of fecal indicator bacteria in the soil. While total numbers of enterococci rapidly declined over time,
E. coli
populations experienced initial growth with concentration increases of 4, 10, and 25 times the initial levels at rainfall treatment depths of 30, 60, and 90 mm, respectively.
E. coli
populations grew to the approximately the same level in all treatments. The 0- to 1-cm layer contained more indicator bacteria than the layers beneath it, and survival of indicator bacteria was better in this layer, with decimation times between 12 and 18 days after the first week of growth. The proportion of bacteria in the 0- to 1-cm layer grew with time as the total number of bacteria in the 0- to 10-cm layer declined. The results of this work indicate the need to revisit the bacterial survival patterns that are assumed in water quality models.
Stoichiometric Assembly of the Cellulosome Generates Maximum Synergy for the Degradation of Crystalline Cellulose, as Revealed by
In Vitro
Reconstitution of the Clostridium thermocellum Cellulosome
Authors: Hirano Katsuaki , Nihei Satoshi , Hasegawa Hiroki , Haruki Mitsuru , Hirano Nobutaka
Published: May 2015

The cellulosome is a supramolecular multienzyme complex formed by species-specific interactions between the cohesin modules of scaffoldin proteins and the dockerin modules of a wide variety of polysaccharide-degrading enzymes. Cellulosomal enzymes bound to the scaffoldin protein act synergistically to degrade crystalline cellulose. However, there have been few attempts to reconstitute intact cellulosomes due to the difficulty of heterologously expressing full-length scaffoldin proteins. We describe the synthesis of a full-length scaffoldin protein containing nine cohesin modules, CipA; its deletion derivative containing two cohesin modules, ΔCipA; and three major cellulosomal cellulases, Cel48S, Cel8A, and Cel9K, of the
Clostridium thermocellum
cellulosome. The proteins were synthesized using a wheat germ cell-free protein synthesis system, and the purified proteins were used to reconstitute cellulosomes. Analysis of the cellulosome assembly using size exclusion chromatography suggested that the dockerin module of the enzymes stoichiometrically bound to the cohesin modules of the scaffoldin protein. The activity profile of the reconstituted cellulosomes indicated that cellulosomes assembled at a CipA/enzyme molar ratio of 1/9 (cohesin/dockerin = 1/1) and showed maximum synergy (4-fold synergy) for the degradation of crystalline substrate and ∼2.4-fold-higher synergy for its degradation than minicellulosomes assembled at a ΔCipA/enzyme molar ratio of 1/2 (cohesin/dockerin = 1/1). These results suggest that the binding of more enzyme molecules on a single scaffoldin protein results in higher synergy for the degradation of crystalline cellulose and that the stoichiometric assembly of the cellulosome, without excess or insufficient enzyme, is crucial for generating maximum synergy for the degradation of crystalline cellulose.
Metabolism of Doubly
para
-Substituted Hydroxychlorobiphenyls by Bacterial Biphenyl Dioxygenases
Authors: Pham Thi Thanh My , Sondossi Mohammad , Sylvestre Michel
Published: May 2015

In this work, we examined the profile of metabolites produced from the doubly
para
-substituted biphenyl analogs 4,4′-dihydroxybiphenyl, 4-hydroxy-4′-chlorobiphenyl, 3-hydroxy-4,4′-dichlorobiphenyl, and 3,3′-dihydroxy-4,4′-chlorobiphenyl by biphenyl-induced
Pandoraea pnomenusa
B356 and by its biphenyl dioxygenase (BPDO). 4-Hydroxy-4′-chlorobiphenyl was hydroxylated principally through a 2,3-dioxygenation of the hydroxylated ring to generate 2,3-dihydro-2,3,4-trihydroxy-4′-chlorobiphenyl and 3,4-dihydroxy-4′-chlorobiphenyl after the removal of water. The former was further oxidized by the biphenyl dioxygenase to produce ultimately 3,4,5-trihydroxy-4′-chlorobiphenyl, a dead-end metabolite. 3-Hydroxy-4,4′-dichlorobiphenyl was oxygenated on both rings. Hydroxylation of the nonhydroxylated ring generated 2,3,3′-trihydroxy-4′-chlorobiphenyl with concomitant dechlorination, and 2,3,3′-trihydroxy-4′-chlorobiphenyl was ultimately metabolized to 2-hydroxy-4-chlorobenzoate, but hydroxylation of the hydroxylated ring generated dead-end metabolites. 3,3′-Dihydroxy-4,4′-dichlorobiphenyl was principally metabolized through a 2,3-dioxygenation to generate 2,3-dihydro-2,3,3′-trihydroxy-4,4′-dichlorobiphenyl, which was ultimately converted to 3-hydroxy-4-chlorobenzoate. Similar metabolites were produced when the biphenyl dioxygenase of
Burkholderia xenovorans
LB400 was used to catalyze the reactions, except that for the three substrates used, the BPDO of LB400 was less efficient than that of B356, and unlike that of B356, it was unable to further oxidize the initial reaction products. Together the data show that BPDO oxidation of doubly
para
-substituted hydroxychlorobiphenyls may generate nonnegligible amounts of dead-end metabolites. Therefore, biphenyl dioxygenase could produce metabolites other than those expected, corresponding to dihydrodihydroxy metabolites from initial doubly
para
-substituted substrates. This finding shows that a clear picture of the fate of polychlorinated biphenyls in contaminated sites will require more insights into the bacterial metabolism of hydroxychlorobiphenyls and the chemistry of the dihydrodihydroxylated metabolites derived from them.
Liposome-Encapsulated Bacteriophages for Enhanced Oral Phage Therapy against Salmonella spp
Authors: Colom Joan , Cano-Sarabia Mary , Otero Jennifer , Cortés Pilar , Maspoch Daniel , Llagostera Montserrat
Published: May 2015

Bacteriophages UAB_Phi20, UAB_Phi78, and UAB_Phi87 were encapsulated in liposomes, and their efficacy in reducing
Salmonella
in poultry was then studied. The encapsulated phages had a mean diameter of 309 to 326 nm and a positive charge between +31.6 and +35.1 mV (pH 6.1). In simulated gastric fluid (pH 2.8), the titer of nonencapsulated phages decreased by 5.7 to 7.8 log units, whereas encapsulated phages were significantly more stable, with losses of 3.7 to 5.4 log units. The liposome coating also improved the retention of bacteriophages in the chicken intestinal tract. When cocktails of the encapsulated and nonencapsulated phages were administered to broilers, after 72 h the encapsulated phages were detected in 38.1% of the animals, whereas the nonencapsulated phages were present in only 9.5%. The difference was significant. In addition, in an
in vitro
experiment, the cecal contents of broilers promoted the release of the phages from the liposomes. In broilers experimentally infected with
Salmonella
, the daily administration of the two cocktails for 6 days postinfection conferred similar levels of protection against
Salmonella
colonization. However, once treatment was stopped, protection by the nonencapsulated phages disappeared, whereas that provided by the encapsulated phages persisted for at least 1 week, showing the enhanced efficacy of the encapsulated phages in protecting poultry against
Salmonella
over time. The methodology described here allows the liposome encapsulation of phages of different morphologies. The preparations can be stored for at least 3 months at 4°C and could be added to the drinking water and feed of animals.
Generation of a Novel Staphylococcus aureus Ghost Vaccine and Examination of Its Immunogenicity against Virulent Challenge in Rats
Authors: Vinod Nagarajan , Oh Sung , Park Hyun Jung , Koo Jung Mo , Choi Chang Won , Kim Sei Chang
Published: May 2015

Staphylococcus aureusis a Gram-positive pathogen that causes a wide range of infections in humans and animals. Bacterial ghosts are nonliving, empty cell envelopes and are well represented as novel vaccine candidates. In this study, we examined the immunogenicity and protective efficacy ofS. aureusghosts (SAGs) against a virulent challenge in rats. Nonliving SAGs were generated by using the MIC of sodium hydroxide. The formation of a transmembrane lysis tunnel structure in SAGs was visualized by scanning electron microscopy. To investigate these SAGs as a vaccine candidate, rats were divided into four groups, A (nonimmunized control), B (orally immunized), C (subcutaneously immunized), and D (intravenously immunized). The IgG antibody responses were significantly stronger in the SAG-immunized groups than in the nonimmunized control group (P< 0.05). Moreover, a significant increase in the populations of CD4+and CD8+T cells was observed in all three immunized groups (P< 0.05). We also found that serum bactericidal antibodies were significantly elicited in the SAG-immunized groups (P< 0.05). Most importantly, the bacterial loads in the immunized groups were significantly lower than those in the nonimmunized control group (P< 0.01). These results suggest that immunization with SAGs induces immune responses and provides protection against a virulentS. aureuschallenge.
Comparative Roles of the Two Helicobacter pylori Thioredoxins in Preventing Macromolecule Damage
Authors: Kuhns Lisa G. , Wang Ge , Maier Robert J.
Published: May 2015

Thioredoxins are highly conserved throughout a wide range of organisms, and they are essential for the isurvival of oxygen-sensitive cells. The gastric pathogen
Helicobacter pylori
uses the thioredoxin system to maintain its thiol/disulfide balance. There are two thioredoxins present in
H. pylori
, Trx1 and Trx2 (herein referred to as TrxA and TrxC). TrxA has been shown to be important as an electron donor for some antioxidant enzymes, but the function of TrxC remains unknown (L. M. Baker, A. Raudonikiene, P. S. Hoffman, and L. B. Poole, J Bacteriol 183:1961–1973, 2001; P. Alamuri and R. J. Maier, J Bacteriol 188:5839–5850, 2006). We demonstrate that both TrxA and TrxC are important in protecting
H. pylori
from oxidative stress. Individual Δ
trxA
and Δ
trxC
deletion mutant strains each show a greater abundance of lipid peroxides and suffer more DNA damage and more protein carbonylation than the parent. Both deletion mutants were much more sensitive to O
2
-mediated viability loss than the parent. Unexpectedly, the oxidative DNA damage and protein carbonylation was more severe in the Δ
trxC
mutant than in the Δ
trxA
mutant; it had 20-fold- and 4-fold-more carbonylated protein content than the wild type and the Δ
trxA
strain, respectively, after 4 h of atmospheric O
2
stress.
trx
transcript abundance was altered by the deletion of the heterologous
trx
gene. The Δ
trxC
mutant lacked mouse colonization ability, while the ability to colonize mouse stomachs was significantly reduced in the Δ
trxA
mutant.
The PduL Phosphotransacylase Is Used To Recycle Coenzyme A within the Pdu Microcompartment
Authors: Liu Yu , Jorda Julien , Yeates Todd O. , Bobik Thomas A.
Published: May 2015

In
Salmonella enterica
, 1,2-propanediol (1,2-PD) utilization (Pdu) is mediated by a bacterial microcompartment (MCP). The Pdu MCP consists of a multiprotein shell that encapsulates enzymes and cofactors for 1,2-PD catabolism, and its role is to sequester a reactive intermediate (propionaldehyde) to minimize cellular toxicity and DNA damage. For the Pdu MCP to function, the enzymes encapsulated within must be provided with a steady supply of substrates and cofactors. In the present study, Western blotting assays were used to demonstrate that the PduL phosphotransacylase is a component of the Pdu MCP. We also show that the N-terminal 20-residue-long peptide of PduL is necessary and sufficient for targeting PduL and enhanced green fluorescent protein (eGFP) to the lumen of the Pdu MCP. We present the results of genetic tests that indicate that PduL plays a role in the recycling of coenzyme A internally within the Pdu MCP. However, the results indicate that some coenzyme A recycling occurs externally to the Pdu MCP. Hence, our results support a model in which a steady supply of coenzyme A is provided to MCP lumen enzymes by internal recycling by PduL as well as by the movement of coenzyme A across the shell by an unknown mechanism. These studies expand our understanding of the Pdu MCP, which has been linked to enteric pathogenesis and which provides a possible basis for the development of intracellular bioreactors for use in biotechnology.

IMPORTANCE
Bacterial MCPs are widespread organelles that play important roles in pathogenesis and global carbon fixation. Here we show that the PduL phosphotransacylase is a component of the Pdu MCP. We also show that PduL plays a key role in cofactor homeostasis by recycling coenzyme A internally within the Pdu MCP. Further, we identify a potential N-terminal targeting sequence using a bioinformatic approach and show that this short sequence extension is necessary and sufficient for directing PduL as well as heterologous proteins to the lumen of the Pdu MCP. These findings expand our general understanding of bacterial MCP assembly and cofactor homeostasis.
Bacillus anthracis Overcomes an Amino Acid Auxotrophy by Cleaving Host Serum Proteins
Authors: Terwilliger Austen , Swick Michelle C. , Pflughoeft Kathryn J. , Pomerantsev Andrei , Lyons C. Rick , Koehler Theresa M. , Maresso Anthony
Published: May 2015

Bacteria sustain an infection by acquiring nutrients from the host to support replication. The host sequesters these nutrients as a growth-restricting strategy, a concept termed “nutritional immunity.” Historically, the study of nutritional immunity has centered on iron uptake because many bacteria target hemoglobin, an abundant circulating protein, as an iron source. Left unresolved are the mechanisms that bacteria use to attain other nutrients from host sources, including amino acids. We employed a novel medium designed to mimic the chemical composition of human serum, and we show here that
Bacillus anthracis
, the causative agent of anthrax disease, proteolyzes human hemoglobin to liberate essential amino acids which enhance its growth. This property can be traced to the actions of InhA1, a secreted metalloprotease, and extends to at least three other serum proteins, including serum albumin. The results suggest that we must also consider proteolysis of key host proteins to be a way for bacterial pathogens to attain essential nutrients, and we provide an experimental framework to determine the host and bacterial factors involved in this process.

IMPORTANCE
The mechanisms by which bacterial pathogens acquire nutrients during infection are poorly understood. Here we used a novel defined medium that approximates the chemical composition of human blood serum, blood serum mimic (BSM), to better model the nutritional environment that pathogens encounter during bacteremia. Removing essential amino acids from BSM revealed that two of the most abundant proteins in blood—hemoglobin and serum albumin—can satiate the amino acid requirement for
Bacillus anthracis
, the causative agent of anthrax. We further demonstrate that hemoglobin is proteolyzed by the secreted protease InhA1. These studies highlight that common blood proteins can be a nutrient source for bacteria. They also challenge the historical view that hemoglobin is solely an iron source for bacterial pathogens.
Importance of Real-Time Assays To Distinguish Multidrug Efflux Pump-Inhibiting and Outer Membrane-Destabilizing Activities in Escherichia coli
Authors: Misra Rajeev , Morrison Keith D. , Cho Hyun Jae , Khuu Thanh
Published: May 2015

The constitutively expressed AcrAB multidrug efflux system ofEscherichia colishows a high degree of homology with the normally silent AcrEF system. Exposure of a strain withacrABdeleted to antibiotic selection pressure frequently leads to the insertion sequence-mediated activation of the homologous AcrEF system. In this study, we used strains constitutively expressing either AcrAB or AcrEF from their normal chromosomal locations to resolve a controversy about whether phenylalanylarginine β-naphthylamide (PAβN) inhibits the activities of AcrAB and AcrEF and/or acts synergistically with antibiotics by destabilizing the outer membrane permeability barrier. Real-time efflux assays allowed a clear distinction between the efflux pump-inhibiting activity of PAβN and the outer membrane-destabilizing action of polymyxin B nonapeptide (PMXBN). When added in equal amounts, PAβN, but not PMXBN, strongly inhibited the efflux activities of both AcrAB and AcrEF pumps. In contrast, when outer membrane destabilization was assessed by the nitrocefin hydrolysis assay, PMXBN exerted a much greater damaging effect than PAβN. Strong action of PAβN in inhibiting efflux activity compared to its weak action in destabilizing the outer membrane permeability barrier suggests that PAβN acts mainly by inhibiting efflux pumps. We concluded that at low concentrations, PAβN acts specifically as an inhibitor of both AcrAB and AcrEF efflux pumps; however, at high concentrations, PAβN in the efflux-proficient background not only inhibits efflux pump activity but also destabilizes the membrane. The effects of PAβN on membrane integrity are compounded in cells unable to extrude PAβN.

IMPORTANCEThe increase in multidrug-resistant bacterial pathogens at an alarming rate has accelerated the need for implementation of better antimicrobial stewardship, discovery of new antibiotics, and deeper understanding of the mechanism of drug resistance. The work carried out in this study highlights the importance of employing real-time fluorescence-based assays in differentiating multidrug efflux-inhibitory and outer membrane-destabilizing activities of antibacterial compounds.