bluuebunny/crossref_metadata_2025 huggingface dataset:
The neural regulation of the kidney in hypertension and renal failure
Authors: Johns Edward J.
Published: August 2013

New Findings

What is the topic of this review?

Reports that bilateral renal denervation in resistant hypertensive patients results in a long‐lasting reduction in blood pressure raise the question of the underlying mechanisms involved and how they may be deranged in pathophysiological states of hypertension and renal failure.

What advances does it highlight?

The renal sensory afferent nerves and efferent sympathetic nerves work together to exert an important control over extracellular fluid volume, hence the level at which blood pressure is set. This article emphasizes that both the afferent and the efferent renal innervation may contribute to the neural dysregulation of the kidney that occurs in chronic renal disease and resistant hypertension.

Autonomic control is central to cardiovascular homeostasis, and this is exerted not only at the level of the heart and blood vessels but also at the kidney. At the kidney, the sympathetic neural regulation of renin release and fluid reabsorption may influence fluid balance and, in the longer term, the level at which blood pressure is set. The role of the renal innervation in the regulation of blood pressure has received renewed attention over the past few years, following the reports that bilateral renal denervation of resistant hypertensive patients resulted in a marked reduction in blood pressure, which has been maintained for several years. Such has been the interest that this approach of renal denervation is being applied in other patient groups with diabetes, obesity and renal failure, with the hope that there may be a sustained reduction in blood pressure as well as the amelioration of some aspects of the metabolic syndrome. However, the factors that come into play to cause the rise in blood pressure in these patient groups, particularly the resistant hypertensive patients, are far from clear. Moreover, the mechanisms leading to the fall in blood pressure following renal denervation of resistant hypertensive patients currently elude our understanding and is therefore an area that requires much more investigation to enhance our insight.
Effect of vasopressin on the expression of genes for key enzymes of hyaluronan turnover in Wistar Albino Glaxo and Brattleboro rat kidneys
Authors: Ivanova Lyudmila N. , Babina Alina V. , Baturina Galina S. , Katkova Lyubov E.
Published: August 2013

New findings

What is the central question of this study?

The interstitium of the renal inner medulla contains abundant linear negatively charged glycosaminoglycan hyaluronan (HA), which affects the water flow depending on their polymeric state. It remains an open question whether HA is involved in the long‐term effect of vasopressin on water reabsorption via the regulation of the expression of genes for key enzymes of HA turnover.

What is the main finding and its importance?

Using real‐time RT‐PCR, we show that in rats vasopressin inhibits the synthesis of HA and concomitantly promotes its degradation in the renal interstitium through its effect on genes for key enzymes of HA turnover (hyaluronan synthase‐2 and hyaluronidase‐2). Analysis of the long‐term effect of vasopressin on the transcription of these genes demonstrated a new mechanism in hormonal regulation of the renal concentrating function.

Hyaluronan (HA), the major glycosaminoglycan of the interstitial matrix, is heterogeneously distributed within the kidney. Using real‐time RT‐PCR, we tested the assumption that renal HA may be involved in the long‐term effect of vasopressin on water reabsorption. The expression of the genes encoding hyaluronan synthase‐2 (Has2), hyaluronidase‐1 and hyaluronidase‐2 (Hyal1 and Hyal2) was studied in the kidneys of Wistar Albino Glaxo (WAG) and homozygous vasopressin‐deficient Brattleboro rats treated with the V2 receptor‐selective vasopressin analogue dDAVP (100 μg (kg body wt)−1, i.p., twice a day for 2 days). The Has2 mRNA content was the highest in the kidney papilla of the hydrated WAG and control Brattleboro rats, devoid of vasopressin. In WAG rats, dDAVP induced a considerable decrease in Has2 mRNA content in the papilla, with less pronounced changes in the cortex. The changes elicited by dDAVP in Brattleboro rats tended to be the same as in WAG rats, but weaker. In contrast to Has2, dDAVP treatment caused a significant increase in the Hyal1 and Hyal2 mRNA content in the renal papilla of WAG and Brattleboro rats. In rats of both strains, there was a good fit between Hyal1 and Hyal2 transcriptional levels and changes in hyaluronidase activity in the renal tissue. It is suggested that vasopressin is able to inhibit the synthesis of HA and concomitantly promote its degradation in the interstitium of the renal papilla, thereby facilitating water flow between elements of the renal countercurrent system. The implications for this effect are discussed in the context of the data in the literature.
Admittance‐based pressure–volume loop measurements in a porcine model of chronic myocardial infarction
Authors: van Hout G. P. J. , de Jong R. , Vrijenhoek J. E. P. , Timmers L. , Duckers H. J. , Hoefer I. E.
Published: August 2013

New Findings

What is the central question of this study?

Are admittance‐based pressure–volume measurements useful for the assessment of cardiac function in a chronic myocardial infarction model compared with three‐dimensional echocardiography and with classical conductance measurements?

What is the main finding and its importance?

Baseline admittance‐based measurements show good agreement with echocardiography and classical conductance in human‐sized porcine hearts. Myocardial infarction significantly alters ventricular dimensions and pressure–volume loop measurements compared with three‐dimensional echocardiography. The novel admittance system reliably monitors changes in cardiac function after myocardial infarction.

The aim of this study was to validate admittance‐based pressure–volume (PV) loop measurements for the assessment of cardiac function in a porcine model of chronic myocardial infarction. The traditional PV loop measurement technique requires hypertonic saline injections for parallel conductance correction prior to signal conversion into volume. Furthermore, it assumes a linear relationship between conductance and volume. More recently, an admittance‐based technique has been developed, which continuously measures parallel conductance and uses a non‐linear equation for volume calculation. This technique has not yet been evaluated in a large‐animal model of myocardial ischaemia. Eleven pigs underwent invasive PV measurements with the admittance system (AS) and the traditional conductance system followed by three‐dimensional echocardiography (3DE). After baseline measurements, pigs were subjected to 90 min left anterior descending coronary artery occlusion, followed by the same measurements at 8 weeks follow‐up. In the healthy heart, the AS showed good agreement with 3DE for left ventricular volumes and a reasonable correlation for ejection fraction (r= 0.756, P= 0.007). At follow‐up, an increase in end‐systolic volume was observed with 3DE (+15.4 ± 14.4 ml, P= 0.005) and the AS (+34.6 ± 36.1 ml, P= 0.010). The ejection fraction measured with 3DE (−13.2 ± 5.2%, P < 0.001) and the AS (−20.3 ± 11.2%, P < 0.001) significantly decreased. We conclude that the AS can be used for quantitative monitoring of changes in cardiac function induced by myocardial infarction and provides comparable results to 3DE, rendering it a useful tool for functional testing in large‐animal cardiac models.
Modulation of food intake by mTOR signalling in the dorsal motor nucleus of the vagus in male rats: focus on ghrelin and nesfatin‐1
Authors: Zhang Weizhen , Zhang Chao , Fritze Danielle , Chai Biaoxin , Li Jiyao , Mulholland Michael W.
Published: August 2013

New findings

What is the central question of this study?

Does mammalian target of rapamycin (mTOR) signalling in the dorsal vagal complex contribute to the modulation of energy homeostasis?

What is the main finding and its importance?

Our study shows, for the first time, that mTOR signalling in neurons of the dorsal motor nucleus of the vagus regulates both the nutrient and the hormonal signals for the modulation of food intake. These results imply that mTOR signalling in the hindbrain may serve as a potential target for treatment of obesity and appetite‐related disorders.

Previous studies have demonstrated that mammalian target of rapamycin (mTOR) signalling in the hypothalamus is involved in the control of energy homeostasis. The aim of this study was to characterize the effect of mTOR signalling in the dorsal motor nucleus of the vagus (DMNV) on energy intake. Phospho‐mTOR was detected in the DMNV neurons, and its levels were increased by energy deprivation. Rapamycin significantly inhibited mTOR activity and reduced food intake when administrated into the fourth ventricle. Exposure of DMNV neurons to ghrelin increased the phosphorylation of mTOR. Injection of ghrelin into the fourth ventricle significantly increased food intake relative to the control vehicle. Pretreatment with rapamycin for 15 min attenuated the orexigenic effect of ghrelin. A reduction in the phosphorylation of mTOR was observed following injection of nesfatin‐1 into the fourth ventricle. When administrated by injection into the fourth ventricle, nesfatin‐1 suppressed food intake in comparison with the control vehicle. The anorexigenic effect of nesfatin‐1 was significantly attenuated by pretreatment with leucine for 15 min. All these findings suggest that mTOR signalling in the DMNV neurons regulates both the nutrient and the hormonal signals for the modulation of food intake.
Phosphatidylethanolamine binding protein 1 in vacular endothelial cell autophagy and atherosclerosis
Authors: Wang Li , Li HaiYing , Zhang JinFeng , Lu Wei , Zhao Jing , Su Le , Zhao BaoXiang , Zhang Yun , Zhang ShangLi , Miao JunYing
Published: August 2013

Key points

The enzyme phosphatidylcholine‐specific phospholipase C (PC‐PLC) participates in atherosclerosis development and may negatively regulate autophagy.

Phosphatidylethanolamine binding protein 1 (PEBP1) represents a novel effector of signal transduction pathways that control cellular growth, motility, apoptosis, genomic integrity, and therapeutic resistance.

The disruption of PEBP1 was reported to associate with a wide range of diseases, such as cancer, pancreatitis, and Alzheimer's disease, making it a potential target for disease therapy. However, the roles of PEBP1 in vascular endothelial cell (VEC) autophagy and arteriosclerosis are not clear.

Here we report that PEBP1 interacts with PC‐PLC and positively regulates PC‐PLC activity, while both PEBP1 and PC‐PLC negatively regulate VEC autophagy.

The PEBP1 level is elevated during the development of atherosclerosis, and the PC‐PLC inhibitor D609 significantly decreases the upregulated PEBP1 level in apolipoprotein E−/− mice, suggesting that PEBP1 may be a potential diagnostic indicator for atherosclerosis.

Abstract  We previously found that phosphatidylcholine‐specific phospholipase C (PC‐PLC) was a key inducing element of atherosclerosis, and might negatively regulate human umbilical vein endothelial cell (HUVEC) autophagy. To further investigate the mechanism of PC‐PLC action, we initially identified phosphatidylethanolamine binding protein 1 (PEBP1) as a binding partner of PC‐PLC by using mass spectrometry (MS, MALDI‐TOF/TOF). We found that PEBP1 positively regulated PC‐PLC activity in HUVECs, and inhibition of PC‐PLC by its inhibitor D609 suppressed PEBP1 expression dramatically. Moreover, both PC‐PLC and PEBP1 negatively regulated HUVEC autophagy independently of mammalian target of rapamycin (mTOR). Furthermore, the PEBP1 level was elevated during the development of atherosclerosis, while D609 significantly decreased the upregulated PEBP1 level in apoE−/− mice.
Direct Geometry Processing for Telefabrication
Authors: Chen Yong , Li Kang , Qian Xiaoping
Published: August 2013

This paper presents a new approach for telefabrication where a physical object is scanned in one location and fabricated in another location. This approach integrates three-dimensional (3D) scanning, geometric processing of scanned data, and additive manufacturing (AM) technologies. In this paper, we focus on a set of direct geometric processing techniques that enable the telefabrication. In this approach, 3D scan data are directly sliced into layer-wise contours. Sacrificial supports are generated directly from the contours and digital mask images of the objects and the supports for stereolithography apparatus (SLA) processes are then automatically generated. The salient feature of this approach is that it does not involve any intermediate geometric models such as STL, polygons, or nonuniform rational B-splines (NURBS) that are otherwise commonly used in prevalent approaches. The experimental results on a set of objects fabricated on several SLA machines confirm the effectiveness of the approach in faithfully telefabricating physical objects.
Exploring the Effectiveness of Using Graveyard Data When Generating Design Alternatives
Authors: Foster Garrett , Ferguson Scott
Published: August 2013

The objective of this paper is to demonstrate that unique alternative designs can be efficiently found by searching the discarded data (or graveyard) from a multiobjective genetic algorithm (MOGA). Motivation for using graveyard data to generate design alternatives arises from the computational cost associated with real-time design space exploration of multiobjective optimization problems. The effectiveness of this approach is explored by comparing (1) the uniqueness of alternatives found using graveyard data and those generated using an optimization-based search, and (2) how alternative generation near the Pareto frontier is impacted. Two multiobjective case study problems are introduced—a two bar truss and an I-beam design optimization. Results from these studies indicate that using graveyard data allows for the discovery of alternative designs that are at least 70% as unique as alternatives found using an optimization-based alternative identification approach, while saving a significant number of functional evaluations. Additionally, graveyard data are shown to be better suited for alternative generation near the Pareto frontier than standard sampling techniques. Finally, areas of future work are also discussed.
Batch Process and Sensitivity Analysis of Collision Detection of Planar Convex Polygons in Motion
Authors: Chen Cheng-fu
Published: August 2013

A new method for formulation, solution, and sensitivity analysis of collision detection of convex objects in motion is presented. The collision detection problem is formulated as a parametric programming problem governed by the changes in the relative translation and relative rotation between the two objects considered. The two parameters together determine all the possible relative configurations between two moving convex objects. Therefore, solving this parametric problem allows for knowing the proximity information for all the possible configurations of the objects. We develop a two-step decomposition procedure to solve this parametric programming problem, and show that the solution is a convex function of the two parameters. This convexity feature enables an archive of the proximity information and sensitivity analysis for the collision detection problem.
Multistack Close Range Photogrammetry for Low Cost Submillimeter Metrology
Authors: Galantucci L. M. , Lavecchia F. , Percoco G.
Published: August 2013

Considerable research effort has been focused on evaluating the accuracy of meso- and macroscale digital close range photogrammetry. However, evaluations of accuracy and applications in the submillimeter scale are rare. In this paper the authors propose the development of a three-dimensional (3D) photogrammetric scanner, based on macrolens cameras, able to reconstruct the three-dimensional surface topography of objects with submillimeter features. The system exploits multifocal image composition and has been designed for installation on all types of Numerical Controlled or Robotic systems. The approach is exploitable for digitizing submillimeter features at mesoscale as well as macroscale objects.
Whole-Genome Sequencing of Gentamicin-Resistant Campylobacter coli Isolated from U.S. Retail Meats Reveals Novel Plasmid-Mediated Aminoglycoside Resistance Genes
Authors: Chen Yuansha , Mukherjee Sampa , Hoffmann Maria , Kotewicz Michael L , Young Shenia , Abbott Jason , Luo Yan , Davidson Maureen K. , Allard Marc , McDermott Patrick , Zhao Shaohua
Published: August 2013

Aminoglycoside resistance in
Campylobacter
has been routinely monitored in the United States in clinical isolates since 1996 and in retail meats since 2002. Gentamicin resistance first appeared in a single human isolate of
Campylobacter coli
in 2000 and in a single chicken meat isolate in 2007, after which it increased rapidly to account for 11.3% of human isolates and 12.5% of retail isolates in 2010. Pulsed-field gel electrophoresis analysis indicated that gentamicin-resistant
C. coli
isolates from retail meat were clonal. We sequenced the genomes of two strains of this clone using a next-generation sequencing technique in order to investigate the genetic basis for the resistance. The gaps of one strain were closed using optical mapping and Sanger sequencing, and this is the first completed genome of
C. coli
. The two genomes are highly similar to each other. A self-transmissible plasmid carrying multiple antibiotic resistance genes was revealed within both genomes, carrying genes encoding resistance to gentamicin, kanamycin, streptomycin, streptothricin, and tetracycline. Bioinformatics analysis and experimental results showed that gentamicin resistance was due to a phosphotransferase gene,
aph(2″)-Ig
, not described previously. The phylogenetic relationship of this newly emerged clone to other
Campylobacter
spp. was determined by whole-genome single nucleotide polymorphisms (SNPs), which showed that it clustered with the other poultry isolates and was separated from isolates from livestock.
The Mevalonate Auxotrophic Mutant of Staphylococcus aureus Can Adapt to Mevalonate Depletion
Authors: Yu Wenqi , Leibig Martina , Schäfer Tina , Bertram Ralph , Ohlsen Knut , Götz Friedrich
Published: August 2013

In this study, we attempted to adopt the auxotrophic mevalonate synthase mutant (Δ
mvaS
mutant) of
Staphylococcus aureus
to study whether a nongrowing but viable cell population is tolerant to bactericidal antibiotics. The mevalonate-depleted nongrowing Δ
mvaS
mutant was found tolerant to antibiotics. Surprisingly, after prolonged cultivation, we obtained stable Δ
mvaS
variants that were able to grow without mevalonate, which suggested unknown mechanisms for compensating undecaprenyl pyrophosphate production without mevalonate in
S. aureus
.
Regional Resistance Surveillance Program Results for 12 Asia-Pacific Nations (2011)
Authors: Mendes Rodrigo E. , Mendoza Myrna , Banga Singh Kirnpal K. , Castanheira Mariana , Bell Jan M. , Turnidge John D. , Lin Stephen S. F. , Jones Ronald N.
Published: August 2013

The Regional Resistance Surveillance program monitored susceptibility rates and developing resistance by geographic region, including 12 Asia-Pacific (APAC) countries. Reference broth microdilution methods for susceptibility/interpretations were applied, processing 5,053 strains. Among
Staphylococcus aureus
isolates (37% methicillin-resistant
S. aureus
[MRSA], highest in South Korea [73%]), linezolid (LZD), tigecycline (TIG), and vancomycin were 100% active, but 33 and 34% of strains were levofloxacin (LEV) or macrolide resistant, respectively.
Streptococcus pneumoniae
was most resistant to β-lactams and macrolides (45%) but was LZD, LEV, and TIG susceptible (>98%). Extended-spectrum β-lactamase (ESBL) phenotype rates in
Escherichia coli
and
Klebsiella
spp. were 48 and 47%, respectively, and were highest in Taiwan, at 75 to 91%. The best anti-ESBL-phenotype agents were amikacin (81 to 96% susceptible), colistin (COL; >98%), TIG (>98%), and carbapenems (81 to 97%).
Pseudomonas aeruginosa
showed ≥20% resistance to all drugs except COL (99% susceptible). In conclusion, endemic evolving antimicrobial resistances in APAC nations show compromised roles for many commonly used antimicrobials.
Subtyping of a Large Collection of Historical Listeria monocytogenes Strains from Ontario, Canada, by an Improved Multilocus Variable-Number Tandem-Repeat Analysis (MLVA)
Authors: Saleh-Lakha S. , Allen V. G. , Li J. , Pagotto F. , Odumeru J. , Taboada E. , Lombos M. , Tabing K. C. , Blais B. , Ogunremi D. , Downing G. , Lee S. , Gao A. , Nadon C. , Chen S.
Published: August 2013

Listeria monocytogenes
is responsible for severe and often fatal food-borne infections in humans. A collection of 2,421
L. monocytogenes
isolates originating from Ontario's food chain between 1993 and 2010, along with Ontario clinical isolates collected from 2004 to 2010, was characterized using an improved multilocus variable-number tandem-repeat analysis (MLVA). The MLVA method was established based on eight primer pairs targeting seven variable-number tandem-repeat (VNTR) loci in two 4-plex fluorescent PCRs. Diversity indices and amplification rates of the individual VNTR loci ranged from 0.38 to 0.92 and from 0.64 to 0.99, respectively. MLVA types and pulsed-field gel electrophoresis (PFGE) patterns were compared using Comparative Partitions analysis involving 336 clinical and 99 food and environmental isolates. The analysis yielded Simpson's diversity index values of 0.998 and 0.992 for MLVA and PFGE, respectively, and adjusted Wallace coefficients of 0.318 when MLVA was used as a primary subtyping method and 0.088 when PFGE was a primary typing method. Statistical data analysis using BioNumerics allowed for identification of at least 8 predominant and persistent
L. monocytogenes
MLVA types in Ontario's food chain. The MLVA method correctly clustered epidemiologically related outbreak strains and separated unrelated strains in a subset analysis. An MLVA database was established for the 2,421
L. monocytogenes
isolates, which allows for comparison of data among historical and new isolates of different sources. The subtyping method coupled with the MLVA database will help in effective monitoring/prevention approaches to identify environmental contamination by pathogenic strains of
L. monocytogenes
and investigation of outbreaks.
Molecular Analysis of the Acinetobacter baumannii Biofilm-Associated Protein
Authors: Goh H. M. Sharon , Beatson Scott A. , Totsika Makrina , Moriel Danilo G. , Phan Minh-Duy , Szubert Jan , Runnegar Naomi , Sidjabat Hanna E. , Paterson David L. , Nimmo Graeme R. , Lipman Jeffrey , Schembri Mark A.
Published: August 2013

Acinetobacter baumannii
is a multidrug-resistant pathogen associated with hospital outbreaks of infection across the globe, particularly in the intensive care unit. The ability of
A. baumannii
to survive in the hospital environment for long periods is linked to antibiotic resistance and its capacity to form biofilms. Here we studied the prevalence, expression, and function of the
A. baumannii
biofilm-associated protein (Bap) in 24 carbapenem-resistant
A. baumannii
ST92 strains isolated from a single institution over a 10-year period. The
bap
gene was highly prevalent, with 22/24 strains being positive for
bap
by PCR. Partial sequencing of
bap
was performed on the index case strain MS1968 and revealed it to be a large and highly repetitive gene approximately 16 kb in size. Phylogenetic analysis employing a 1,948-amino-acid region corresponding to the C terminus of Bap showed that Bap
MS1968
clusters with Bap sequences from clonal complex 2 (CC2) strains ACICU, TCDC-AB0715, and 1656-2 and is distinct from Bap in CC1 strains. By using overlapping PCR, the
bap
MS1968
gene was cloned, and its expression in a recombinant
Escherichia coli
strain resulted in increased biofilm formation. A Bap-specific antibody was generated, and Western blot analysis showed that the majority of
A. baumannii
strains expressed an ∼200-kDa Bap protein. Further analysis of three Bap-positive
A. baumannii
strains demonstrated that Bap is expressed at the cell surface and is associated with biofilm formation. Finally, biofilm formation by these Bap-positive strains could be inhibited by affinity-purified Bap antibodies, demonstrating the direct contribution of Bap to biofilm growth by
A. baumannii
clinical isolates.
Temporal Changes and Altitudinal Distribution of Aerobic Anoxygenic Phototrophs in Mountain Lakes
Authors: Čuperová Zuzana , Holzer Evelyn , Salka Ivette , Sommaruga Ruben , Koblížek Michal
Published: August 2013

Aerobic anoxygenic phototrophs (AAPs) are bacteriochlorophylla-containing microorganisms that use organic substrates for growth but can supplement their energy requirements with light. They have been reported from various marine and limnic environments; however, their ecology remains largely unknown. Here infrared epifluorescence microscopy was used to monitor temporal changes in AAPs in the alpine lake Gossenköllesee, located in the Tyrolean Alps, Austria. AAP abundance was low (103cells ml−1) until mid-July and reached a maximum of ∼1.3 × 105cells ml−1(29% of all prokaryotes) in mid-September. We compared the studied lake with other mountain lakes located across an altitudinal gradient (913 to 2,799 m above sea level). The concentration of dissolved organic carbon and water transparency seem to be the main factors influencing AAP abundance during the seasonal cycle as well as across the altitudinal gradient. While the AAP populations inhabiting the alpine lakes were composed of intensely pigmented large rods (5 to 12 μm), the lakes below the tree line were inhabited by a variety of smaller morphotypes. Analysis ofpufMdiversity revealed that AAPs in Gossenköllesee were almost exclusivelySphingomonadalesspecies, which indicates that AAP communities inhabiting alpine lakes are relatively homogeneous compared to those in low-altitude lakes.
A Fungal Insecticide Engineered for Fast
Per Os
Killing of Caterpillars Has High Field Efficacy and Safety in Full-Season Control of Cabbage Insect Pests
Authors: Liu Yong-Jie , Liu Jing , Ying Sheng-Hua , Liu Shu-Sheng , Feng Ming-Guang
Published: August 2013

Fungal insecticides developed from filamentous pathogens of insects are notorious for their slow killing action through cuticle penetration, depressing commercial interest and practical application. Genetic engineering may accelerate their killing action but cause ecological risk. Here we show that a
Beauveria bassiana
formulation, HV8 (BbHV8), engineered for fast
per os
killing of caterpillars by an insect midgut-acting toxin (Vip3Aa1) overexpressed in conidia has both high field efficacy and safety in full-season protection of cabbage from the damage of an insect pest complex dominated by
Pieris rapae
larvae, followed by
Plutella xylostella
larvae and aphids. In two fields repeatedly sprayed during summer, BbHV8 resulted in overall mean efficacies of killing of 71% and 75%, which were similar or close to the 70% and 83% efficacies achieved by commercially recommended emamectin benzoate but much higher than the 31% and 48% efficacies achieved by the same formulation of the parental wild-type strain (WT). Both BbHV8 and WT sprays exerted no adverse effect on a nontarget spider community during the trials, and the sprays did not influence saprophytic fungi in soil samples taken from the field plots during 4 months after the last spray. Strikingly, BbHV8 and the WT showed low fitness when they were released into the environment because both were decreasingly recovered from the field lacking native
B. bassiana
strains (undetectable 5 months after the spray), and the recovered isolates became much less tolerant to high temperature and UV-B irradiation. Our results highlight for the first time that a rationally engineered fungal insecticide can compete with a chemical counterpart to combat insect pests at an affordable cost and with low ecological risk.
The MAP Kinase Slt2 Is Involved in Vacuolar Function and Actin Remodeling in Saccharomyces cerevisiae Mutants Affected by Endogenous Oxidative Stress
Authors: Pujol-Carrion Nuria , Petkova Mima I. , Serrano Luis , de la Torre-Ruiz Maria Angeles
Published: August 2013

Oxidative stress causes transient actin cytoskeleton depolarization and also provokes vacuole fragmentation in wild-type cells. Under conditions of oxidative stress induced by hydrogen peroxide, the Slt2 protein is required to repolarize the actin cytoskeleton and to promote vacuole fusion. In this study, we show that
grx3 grx4
and
grx5
mutants are cellular models of endogenous oxidative stress. This stress is the result of alterations in iron homeostasis that lead to impairment of vacuolar function and also to disorganization of the actin cytoskeleton. Slt2 overexpression suppresses defects in vacuolar function and actin cytoskeleton organization in the
grx3 grx4
mutant. Slt2 exerts this effect independently of the intracellular levels of reactive oxygen species (ROS) and of iron homeostasis. The deletion of
SLT2
in the
grx3 grx4
mutant results in synthetic lethality related to vacuolar function with substantial vacuole fragmentation. The observation that both Vps4 and Vps73 (two proteins related to vacuole sorting) suppress vacuole fragmentation and actin depolarization in the
grx3 grx4 slt2
triple mutant strengthens the hypothesis that Slt2 plays a role in vacuole homeostasis related to actin dynamics. Here we show that in
sod1
,
grx5
, and
grx3 grx4 slt2
mutants, all of which are affected by chronic oxidative stress, the overexpression of Slt2 favors vacuole fusion through a mechanism dependent on an active actin cytoskeleton.
Methanosarcinaceae and Acetate-Oxidizing Pathways Dominate in High-Rate Thermophilic Anaerobic Digestion of Waste-Activated Sludge
Authors: Ho Dang P. , Jensen Paul D. , Batstone Damien J.
Published: August 2013

This study investigated the process of high-rate, high-temperature methanogenesis to enable very-high-volume loading during anaerobic digestion of waste-activated sludge. Reducing the hydraulic retention time (HRT) from 15 to 20 days in mesophilic digestion down to 3 days was achievable at a thermophilic temperature (55°C) with stable digester performance and methanogenic activity. A volatile solids (VS) destruction efficiency of 33 to 35% was achieved on waste-activated sludge, comparable to that obtained via mesophilic processes with low organic acid levels (<200 mg/liter chemical oxygen demand [COD]). Methane yield (VS basis) was 150 to 180 liters of CH
4
/kg of VS
added
. According to 16S rRNA pyrotag sequencing and fluorescence in situ hybridization (FISH), the methanogenic community was dominated by members of the
Methanosarcinaceae
, which have a high level of metabolic capability, including acetoclastic and hydrogenotrophic methanogenesis. Loss of function at an HRT of 2 days was accompanied by a loss of the methanogens, according to pyrotag sequencing. The two acetate conversion pathways, namely, acetoclastic methanogenesis and syntrophic acetate oxidation, were quantified by stable carbon isotope ratio mass spectrometry. The results showed that the majority of methane was generated by nonacetoclastic pathways, both in the reactors and in off-line batch tests, confirming that syntrophic acetate oxidation is a key pathway at elevated temperatures. The proportion of methane due to acetate cleavage increased later in the batch, and it is likely that stable oxidation in the continuous reactor was maintained by application of the consistently low retention time.
Fluorescence-Based Bacterial Bioreporter for Specific Detection of Methyl Halide Emissions in the Environment
Authors: Farhan Ul Haque Muhammad , Nadalig Thierry , Bringel Françoise , Schaller Hubert , Vuilleumier Stéphane
Published: August 2013

Methyl halides are volatile one-carbon compounds responsible for substantial depletion of stratospheric ozone. Among them, chloromethane (CH
3
Cl) is the most abundant halogenated hydrocarbon in the atmosphere. Global budgets of methyl halides in the environment are still poorly understood due to uncertainties in their natural sources, mainly from vegetation, and their sinks, which include chloromethane-degrading bacteria. A bacterial bioreporter for the detection of methyl halides was developed on the basis of detailed knowledge of the physiology and genetics of
Methylobacterium extorquens
CM4, an aerobic alphaproteobacterium which utilizes chloromethane as the sole source of carbon and energy. A plasmid construct with the promoter region of the chloromethane dehalogenase gene
cmuA
fused to a promotorless yellow fluorescent protein gene cassette resulted in specific methyl halide-dependent fluorescence when introduced into
M. extorquens
CM4. The bacterial whole-cell bioreporter allowed detection of methyl halides at femtomolar levels and quantification at concentrations above 10 pM (approximately 240 ppt). As shown for the model chloromethane-producing plant
Arabidopsis thaliana
in particular, the bioreporter may provide an attractive alternative to analytical chemical methods to screen for natural sources of methyl halide emissions.
Use of Aliphatic
n
-Alkynes To Discriminate Soil Nitrification Activities of Ammonia-Oxidizing Thaumarchaea and Bacteria
Authors: Taylor Anne E. , Vajrala Neeraja , Giguere Andrew T. , Gitelman Alix I. , Arp Daniel J. , Myrold David D. , Sayavedra-Soto Luis , Bottomley Peter J.
Published: August 2013

Ammonia (NH
3
)-oxidizing bacteria (AOB) and thaumarchaea (AOA) co-occupy most soils, yet no short-term growth-independent method exists to determine their relative contributions to nitrification
in situ
. Microbial monooxygenases differ in their vulnerability to inactivation by aliphatic
n
-alkynes, and we found that NH
3
oxidation by the marine thaumarchaeon
Nitrosopumilus maritimus
was unaffected during a 24-h exposure to ≤20 μM concentrations of 1-alkynes C
8
and C
9
. In contrast, NH
3
oxidation by two AOB (
Nitrosomonas europaea
and
Nitrosospira multiformis
) was quickly and irreversibly inactivated by 1 μM C
8
(octyne). Evidence that nitrification carried out by soilborne AOA was also insensitive to octyne was obtained. In incubations (21 or 28 days) of two different whole soils, both acetylene and octyne effectively prevented NH
4
+
-stimulated increases in AOB population densities, but octyne did not prevent increases in AOA population densities that were prevented by acetylene. Furthermore, octyne-resistant, NH
4
+
-stimulated net nitrification rates of 2 and 7 μg N/g soil/day persisted throughout the incubation of the two soils. Other evidence that octyne-resistant nitrification was due to AOA included (i) a positive correlation of octyne-resistant nitrification in soil slurries of cropped and noncropped soils with allylthiourea-resistant activity (100 μM) and (ii) the finding that the fraction of octyne-resistant nitrification in soil slurries correlated with the fraction of nitrification that recovered from irreversible acetylene inactivation in the presence of bacterial protein synthesis inhibitors and with the octyne-resistant fraction of NH
4
+
-saturated net nitrification measured in whole soils. Octyne can be useful in short-term assays to discriminate AOA and AOB contributions to soil nitrification.
Comparison of PCR versus Culture for Detection of Mycobacterium bovis after Experimental Inoculation of Various Matrices Held under Environmental Conditions for Extended Periods
Authors: Adams Angela P. , Bolin Steven R. , Fine Amanda E. , Bolin Carole A. , Kaneene John B.
Published: August 2013

The purpose of this study was to compare the performance of a molecular detection technique (nested PCR) with that of mycobacterial culture in the detection of
Mycobacterium bovis
DNA in a set of 687 samples of experimentally inoculated environmental substrates (hay, soil, corn, water) exposed to natural weather conditions in Michigan. Four replicates of each substrate were used; half were autoclaved for sterilization, all were inoculated with 50,000 CFU of
M. bovis
isolated from Michigan livestock, and all were placed in outdoor enclosures, with half under shade and the other half exposed to direct sunlight. Samples were tested for the presence of
M. bovis
during one 12-month period, with monthly sample testing and during three 12-week periods (winter, spring, summer) with weekly sample testing. Samples were subjected to mycobacterial culture for isolation of
M. bovis
and a nested PCR with two primer sets targeting IS
6110
to detect
M. bovis
DNA. In 128 samples tested during the 12-month period,
M. bovis
was not detectable by culture after 2 months but
M. bovis
DNA was detectable by PCR for at least 7 months. Of the 559 samples tested during the 12-week periods, PCR detected
M. bovis
DNA for up to 88 days in all of the sample types. There were no significant differences in the detection of
M. bovis
between shade and sun samples or between sterile and unsterilized samples, regardless of the detection method (PCR or culture). For use in epidemiologic investigations, the PCR assay was more rapid than mycobacterial culture, was not hindered by contaminating organisms, and detected
M. bovis
DNA in environment samples much longer after initial contamination than mycobacterial culture did.
Benchmarking Various Green Fluorescent Protein Variants in Bacillus subtilis, Streptococcus pneumoniae, and Lactococcus lactis for Live Cell Imaging
Authors: Overkamp Wout , Beilharz Katrin , Detert Oude Weme Ruud , Solopova Ana , Karsens Harma , Kovács Ákos T. , Kok Jan , Kuipers Oscar P. , Veening Jan-Willem
Published: August 2013

Green fluorescent protein (GFP) offers efficient ways of visualizing promoter activity and protein localization
in vivo
, and many different variants are currently available to study bacterial cell biology. Which of these variants is best suited for a certain bacterial strain, goal, or experimental condition is not clear. Here, we have designed and constructed two “superfolder” GFPs with codon adaptation specifically for
Bacillus subtilis
and
Streptococcus pneumoniae
and have benchmarked them against five other previously available variants of GFP in
B. subtilis
,
S. pneumoniae
, and
Lactococcus lactis
, using promoter-
gfp
fusions. Surprisingly, the best-performing GFP under our experimental conditions in
B. subtilis
was the one codon optimized for
S. pneumoniae
and
vice versa
. The data and tools described in this study will be useful for cell biology studies in low-GC-rich Gram-positive bacteria.
Integrating Terminal Truncation and Oligopeptide Fusion for a Novel Protein Engineering Strategy To Improve Specific Activity and Catalytic Efficiency: Alkaline α-Amylase as a Case Study
Authors: Yang Haiquan , Liu Long , Shin Hyun-dong , Chen Rachel R. , Li Jianghua , Du Guocheng , Chen Jian
Published: August 2013

In this work, we integrated terminal truncation and N-terminal oligopeptide fusion as a novel protein engineering strategy to improve specific activity and catalytic efficiency of alkaline α-amylase (AmyK) from
Alkalimonas amylolytica
. First, the C terminus or N terminus of AmyK was partially truncated, yielding 12 truncated mutants, and then an oligopeptide (AEAEAKAKAEAEAKAK) was fused at the N terminus of the truncated AmyK, yielding another 12 truncation-fusion mutants. The specific activities of the truncation-fusion mutants AmyKΔC500-587::OP and AmyKΔC492-587::OP were 25.5- and 18.5-fold that of AmyK, respectively. The
k
cat
/

K
m

was increased from 1.0 × 10
5
liters · mol
−1
· s
−1
for AmyK to 30.6 × and 23.2 × 10
5
liters · mol
−1
· s
−1
for AmyKΔC500-587::OP and AmyKΔC492-587::OP, respectively. Comparative analysis of structure models indicated that the higher flexibility around the active site may be the main reason for the improved catalytic efficiency. The proposed terminal truncation and oligopeptide fusion strategy may be effective to engineer other enzymes to improve specific activity and catalytic efficiency.
Environmental Detection of Genogroup I, II, and IV Noroviruses by Using a Generic Real-Time Reverse Transcription-PCR Assay
Authors: Miura Takayuki , Parnaudeau Sylvain , Grodzki Marco , Okabe Satoshi , Atmar Robert L. , Le Guyader Françoise S.
Published: August 2013

Norovirus is the most common agent implicated in food-borne outbreaks and is frequently detected in environmental samples. These viruses are highly diverse, and three genogroups (genogroup I [GI], GII, and GIV) infect humans. Being noncultivable viruses, real-time reverse transcription-PCR (RT-PCR) is the only sensitive method available for their detection in food or environmental samples. Selection of consensus sequences for the design of sensitive assays has been challenging due to sequence diversity and has led to the development of specific real-time RT-PCR assays for each genogroup. Thus, sample screening can require several replicates for amplification of each genogroup (without considering positive and negative controls or standard curves). This study reports the development of a generic assay that detects all three human norovirus genogroups on a qualitative basis using a one-step real-time RT-PCR assay. The generic assay achieved good specificity and sensitivity for all three genogroups, detected separately or in combination. At variance with multiplex assays, the choice of the same fluorescent dye for all three probes specific to each genogroup allows the levels of fluorescence to be added and may increase assay sensitivity when multiple strains from different genogroups are present. When it was applied to sewage sample extracts, this generic assay successfully detected norovirus in all samples found to be positive by the genogroup-specific RT-PCRs. The generic assay also identified all norovirus-positive samples among 157 archived nucleic acid shellfish extracts, including samples contaminated by all three genogroups.
Exoproteome Profiles of Clostridium cellulovorans Grown on Various Carbon Sources
Authors: Matsui Kazuma , Bae Jungu , Esaka Kohei , Morisaka Hironobu , Kuroda Kouichi , Ueda Mitsuyoshi
Published: August 2013

The cellulosome is a complex of cellulosomal proteins bound to scaffolding proteins. This complex is considered the most efficient system for cellulose degradation.
Clostridium cellulovorans
, which is known to produce cellulosomes, changes the composition of its cellulosomes depending on the growth substrates. However, studies have investigated only cellulosomal proteins; profile changes in noncellulosomal proteins have rarely been examined. In this study, we performed a quantitative proteome analysis of the whole exoproteome of
C. cellulovorans
, including cellulosomal and noncellulosomal proteins, to illustrate how various substrates are efficiently degraded.
C. cellulovorans
was cultured with cellobiose, xylan, pectin, or phosphoric acid-swollen cellulose (PASC) as the sole carbon source. PASC was used as a cellulose substrate for more accurate quantitative analysis. Using an isobaric tag method and a liquid chromatography mass spectrometer equipped with a long monolithic silica capillary column, 639 proteins were identified and quantified in all 4 samples. Among these, 79 proteins were involved in saccharification, including 35 cellulosomal and 44 noncellulosomal proteins. We compared protein abundance by spectral count and found that cellulosomal proteins were more abundant than noncellulosomal proteins. Next, we focused on the fold change of the proteins depending on the growth substrates. Drastic changes were observed mainly among the noncellulosomal proteins. These results indicate that cellulosomal proteins were primarily produced to efficiently degrade any substrate and that noncellulosomal proteins were specifically produced to optimize the degradation of a particular substrate. This study highlights the importance of noncellulosomal proteins as well as cellulosomes for the efficient degradation of various substrates.
Marked Synergistic Bactericidal Effects and Mode of Action of Medium-Chain Fatty Acids in Combination with Organic Acids against Escherichia coli O157:H7
Authors: Kim S. A. , Rhee M. S.
Published: August 2013

The aim of this study was to examine the synergistic bactericidal effects of medium-chain fatty acids (MCFAs; caprylic, capric, and lauric acid) and organic acids (OAs; acetic, lactic, malic, and citric acid) against
Escherichia coli
O157:H7 and to identify their underlying mechanism(s) of action.
E. coli
O157:H7 was treated with MCFAs, OAs, or different combinations of MCFAs and OAs. Membrane damage and cell morphology were examined by flow cytometry and transmission electron microscopy, respectively. Combined treatment resulted in an additional log-unit reduction compared with the sum of the reductions obtained after individual treatment. For example, caprylic acid (1.0 mM, or 0.016%) and citric acid (1.0 mM, or 0.012%) alone showed negligible bactericidal effects (0.30- and 0.06-log-unit reductions, respectively); however, a marked synergistic effect (>7.15-log-unit reduction) was observed when the two were combined. Although flow cytometry and microscopic analyses of bacteria treated with individual MCFAs and OAs showed evidence of membrane disruption, the bacteria were still able to form colonies; thus, the cell damage was recoverable. In contrast, cells exposed to combined treatments showed clear membrane disintegration and/or cell death (irreversible damage). The mechanism underlying the antimicrobial effects of combined treatment with MCFAs or OAs may involve disruption of the bacterial membrane, which then facilitates the entry of other antimicrobial compounds into the cytoplasm. The main advantage of combined treatment with very low concentrations of natural antimicrobial compounds is that it is very cost-effective. Thus, this approach may be an alternative to more conventional antimicrobial treatments, such as those currently used in public health, medical centers, and the food industry.
Simultaneous Near-Infrared Radiant Heating and UV Radiation for Inactivating Escherichia coli O157:H7 and Salmonella enterica Serovar Typhimurium in Powdered Red Pepper (Capsicum annuum L.)
Authors: Ha Jae-Won , Kang Dong-Hyun
Published: August 2013

This study was conducted to investigate the efficacy of the simultaneous application of near-infrared (NIR) heating and UV irradiation for reducing populations of food-borne pathogens, including
Salmonella enterica
serovar Typhimurium and
Escherichia coli
O157:H7 in red pepper powder and to clarify the mechanisms of the lethal effect of the NIR-UV combined treatment. Also, the effect of the combination treatment on quality was determined by measuring changes in color and pungency constituents. Simultaneous NIR-UV combined treatment for 5 min achieved 3.34- and 2.78-log CFU reductions in
S
. Typhimurium and
E. coli
O157:H7, respectively, which involved 1.86- and 1.31-log CFU reductions, respectively, which were attributed to the synergistic effect. Through qualitative and quantitative analyses, damage to the cell envelope was identified as the main factor contributing to the synergistic lethal effect of NIR-UV combined treatment. Color values and capsaicin and dihydrocapsaicin content of NIR-UV simultaneously treated red pepper powder were not significantly (
P
> 0.05) different from those of untreated samples. These results suggest that simultaneous application of NIR and UV treatment can be effectively used to control food-borne pathogens in powdered red pepper without affecting quality.