bluuebunny/crossref_metadata_2025 huggingface dataset:
Cathepsin K modulates invasion, migration and adhesion of oral squamous cell carcinomas in vitro
Authors: Yamashita K , Iwatake M , Okamoto K , Yamada S‐i , Umeda M , Tsukuba T
Published: January 2017

Objective

Cathepsin K was initially discovered as an osteoclast‐specific cysteine proteinase, but the enzyme is also expressed in various cancers including oral squamous cell carcinomas. This study aimed to clarify the function of cathepsin K in oral squamous cell carcinomas.

Materials and Methods

Expression levels of cathepsin K were examined in six types of cell carcinomas. Carcinomas overexpressing cathepsin K were constructed. Effects of cathepsin K overexpression and treatment with odanacatib, a specific cathepsin K inhibitor, on cell invasion, migration and adhesion were analysed.

Results

Different levels of cathepsin K were expressed in carcinomas. Cathepsin K was predominantly localised in lysosomes. Cathepsin K overexpression impaired the proliferation of carcinomas. Invasion analysis showed that cathepsin K overexpression enhanced invasion and migration of carcinomas, whereas inhibition of cathepsin K by odanacatib caused the opposite effects in carcinomas. Cathepsin K overexpression also increased cell adhesion and slightly increased surface expression of the adhesion receptor CD29/integrin β1.

Conclusions

The enhanced invasion of carcinomas resulting from cathepsin K overexpression is probably due to the increased cell migration and adhesion. Thus, cathepsin K is implicated not only in protein degradation but also in invasion, migration and adhesion of oral squamous cell carcinomas.
Institutional Constraints, Rule‐Following, and Circumvention: Tax and Expenditure Limits and the Choice of Fiscal Tools During a Budget Crisis
Authors: Jimenez Benedict S.
Published: January 2017

One of the critical questions in local public budgeting and finance is how the confluence of two major developments in the local public sector—the proliferation of tax and expenditure limits (TELs) and the widespread budget crisis precipitated by the Great Recession—has affected local governments across the country. This research examines how TELs shape the fiscal behavior of municipal governments, specifically the pattern of use of different fiscal tools to respond to a major budget crisis. The fiscal tools available to local governments to address fiscal shocks include personnel‐related budget cuts, service‐related cuts, revenue‐raising responses, efficiency reforms, fund balance and inter‐fund transfers, external borrowing, and inter‐governmental approaches. To what extent did cities rely on specific tools and how did TELs influence cities’ fiscal choices? Using the results of a 2015 national survey of municipal governments in the United States with a population of 50,000 or more, this study shows that TELs have shaped the fiscal choices of cities in ways that can potentially redefine the relationships among city governments, their workforce, and citizens.
Improvement in phosphate acquisition and utilization by a secretory purple acid phosphatase (OsPAP21b) in rice
Authors: Mehra Poonam , Pandey Bipin Kumar , Giri Jitender
Published: January 2017

Phosphate (Pi) deficiency in soil system is a limiting factor for rice growth and yield. Majority of the soil phosphorus (P) is organic in nature, not readily available for root uptake. Low Pi‐inducible purple acid phosphatases (PAPs) are hypothesized to enhance the availability of Pi in soil and cellular system. However, information on molecular and physiological roles of rice PAPs is very limited. Here, we demonstrate the role of a novel rice PAP, OsPAP21b in improving plant utilization of organic‐P. OsPAP21b was found to be under the transcriptional control of OsPHR2 and strictly regulated by plant Pi status at both transcript and protein levels. Biochemically, OsPAP21b showed hydrolysis of several organophosphates at acidic pH and possessed sufficient thermostability befitting for high‐temperature rice ecosystems with acidic soils. Interestingly, OsPAP21b was revealed to be a secretory PAP and encodes a distinguishable major APase (acid phosphatase) isoform under low Pi in roots. Further, OsPAP21b‐overexpressing transgenics showed increased biomass, APase activity and P content in both hydroponics supplemented with organic‐P sources and soil containing organic manure as sole P source. Additionally, overexpression lines depicted increased root length, biomass and lateral roots under low Pi while RNAi lines showed reduced root length and biomass as compared to WT. In the light of these evidences, present study strongly proposes OsPAP21b as a useful candidate for improving Pi acquisition and utilization in rice.
Progression of mean pulmonary arterial pressure in idiopathic pulmonary fibrosis with mild to moderate restriction
Authors: Teramachi Ryo , Taniguchi Hiroyuki , Kondoh Yasuhiro , Ando Masahiko , Kimura Tomoki , Kataoka Kensuke , Suzuki Atsushi , Furukawa Taiki , Sakamoto Koji , Hasegawa Yoshinori
Published: January 2017

ABSTRACT

Background and objective

Elevation of mean pulmonary arterial pressure (MPAP) is associated with poor prognosis in patients with idiopathic pulmonary fibrosis (IPF), yet the progression of MPAP in patients with IPF has not been sufficiently elucidated. We evaluated serial changes in MPAP and its determinants in patients with IPF with mild to moderate restriction.

Methods

We retrospectively reviewed patients with IPF who underwent initial evaluations including right heart catheterization (RHC) in our institute from May 2007 to December 2013 with follow‐up RHC at least 1 year later. Patients with forced vital capacity (FVC) < 50% predicted or those with pulmonary artery wedge pressure >15 mm Hg were excluded.

Results

A total of 95 patients were included. Median follow‐up time of second RHC was 1.8 years. MPAP increased significantly at follow‐up (from 16.8 to 20.2 mm Hg; P < 0.001), and annual change in MPAP (ΔMPAP) was 1.8 mm Hg/year. In multiple regression analysis, the lowest oxygen saturation (SpO2
) at 6‐min walk test (6MWT) was an independent predictor of ΔMPAP. When adjusted for age, sex, baseline MPAP and FVC % predicted, ΔMPAP was a significant predictor of mortality (hazard ratio: 1.21; P = 0.001).

Conclusion

ΔMPAP was significantly associated with desaturation in the 6MWT, and with increased mortality in patients with IPF with mild to moderate restriction.
Addressing tuberculosis patients’ medical and socio‐economic needs: a comprehensive programmatic approach
Authors: Contreras Carmen C. , Millones Ana K. , Santa Cruz Janeth , Aguilar Margot , Clendenes Martin , Toranzo Miguel , Llaro Karim , Lecca Leonid , Becerra Mercedes C. , Yuen Courtney M.
Published: January 2017

Objective

For a cohort of patients with tuberculosis in Carabayllo, Peru, we describe the prevalence of medical comorbidities and socio‐economic needs, the efforts required by a comprehensive support programme (‘TB Cero’) to address them and the success of this programme in linking patients to care.

Methods

Patients diagnosed with tuberculosis in Carabayllo underwent evaluations for HIV, diabetes, mental health and unmet basic needs. For patients initiating treatment during 14 September, 2015–15 May, 2016, we abstracted data from evaluation forms and a support request system. We calculated the prevalence of medical comorbidities and the need for socio‐economic support at the time of tuberculosis diagnosis, as well as the proportion of patients successfully linked to care or support.

Results

Of 192 patients, 83 (43%) had at least one medical comorbidity other than tuberculosis. These included eight (4%) patients with HIV, 12 (6%) with diabetes and 62 (32%) deemed at risk for a mental health condition. Of patients who required follow‐up for a comorbidity, 100% initiated antiretroviral therapy, 71% attended endocrinology consultations and 66% attended psychology consultations. Of 126 (65%) patients who completed the socio‐economic evaluation, 58 (46%) reported already receiving food baskets from the municipality, and 79 (63%) were given additional support, most commonly food vouchers and assistance in accessing health care.

Conclusion

Carabayllo tuberculosis patients face many challenges in addition to tuberculosis. A collaborative, comprehensive treatment support programme can achieve high rates of linkage to care for these needs.
Teachers’ Opinions Regarding the Usage of Action Research in Professional Development
Authors: Yigit Celal , Bagceci Birsen
Published: January 2017

The aim of the study is to investigate the contribution of action research to teachers’ professional development. In line with this goal, a group of teachers were asked their opinions. The working group of the study is comprised of six teachers working at a state primary and middle school. The study is an example of “Case Study”, one of qualitative research methods. The content analysis technique was utilized in the analysis of the data. To sum up, the teachers expressed that action research had a positive impact on their professional development in various aspects.
Microtubule mechanics in the working myocyte
Authors: Robison Patrick , Prosser Benjamin L.
Published: January 2017

The mechanical role of cardiac microtubules (MTs) has been a topic of some controversy. Early studies, which relied largely on pharmacological interventions that altered the MT cytoskeleton as a whole, presented no consistent role. Recent advances in the ability to observe and manipulate specific properties of the cytoskeleton have strengthened our understanding. Direct observation of MTs in working myocytes suggests a spring‐like function, one that is surprisingly tunable by post‐translational modification (PTM). Specifically, detyrosination of MTs facilitates an interaction with intermediate filaments that complex with the sarcomere, altering myocyte stiffness, contractility, and mechanosignalling. Such results support a paradigm of cytoskeletal regulation based on not only polymerization, but also associations with binding partners and PTMs that divide the MT cytoskeleton into functionally distinct subsets. The evolutionary costs and benefits of tuning cytoskeletal mechanics remain an open question, one that we discuss herein. Nevertheless, mechanically distinct MT subsets provide a rich new source of therapeutic targets for a variety of phenomena in the heart.

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In vitro models of the cardiac microenvironment to study myocyte and non‐myocyte crosstalk: bioinspired approaches beyond the polystyrene dish
Authors: Kofron Celinda M. , Mende Ulrike
Published: January 2017

The heart is a complex pluricellular organ composed of cardiomyocytes and non‐myocytes including fibroblasts, endothelial cells and immune cells. Myocytes are responsible for electrical conduction and contractile force generation, while the other cell types are responsible for matrix deposition, vascularization, and injury response. Myocytes and non‐myocytes are known to communicate and exert mutual regulatory effects. In concert, they determine the structural, electrical and mechanical characteristics in the healthy and remodelled myocardium. Dynamic crosstalk between myocytes and non‐myocytes plays a crucial role in stress/injury‐induced hypertrophy and fibrosis development that can ultimately lead to heart failure and arrhythmias. Investigations of heterocellular communication in the myocardium are hampered by the intricate interspersion of the different cell types and the complexity of the tissue architecture. In vitro models have facilitated investigations of cardiac cells in a direct and controllable manner and have provided important functional and mechanistic insights. However, these cultures often lack regulatory input from the other cell types as well as additional topographical, electrical, mechanical and biochemical cues from the cardiac microenvironment that all contribute to modulating cell differentiation, maturation, alignment, function and survival. Advancements in the development of more complex pluricellular physiological platforms that incorporate diverse cues from the myocardial microenvironment are expected to lead to more physiologically relevant cardiac tissue‐like in vitro models for mechanistic biological research, disease modelling, therapeutic target identification, drug testing and regeneration.

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Relationship between cortical state and spiking activity in the lateral geniculate nucleus of marmosets
Authors: Pietersen Alexander N.J. , Cheong Soon Keen , Munn Brandon , Gong Pulin , Martin Paul R. , Solomon Samuel G.
Published: January 2017

Key points

How parallel are the primate visual pathways? In the present study, we demonstrate that parallel visual pathways in the dorsal lateral geniculate nucleus (LGN) show distinct patterns of interaction with rhythmic activity in the primary visual cortex (V1).

In the V1 of anaesthetized marmosets, the EEG frequency spectrum undergoes transient changes that are characterized by fluctuations in delta‐band EEG power.

We show that, on multisecond timescales, spiking activity in an evolutionary primitive (koniocellular) LGN pathway is specifically linked to these slow EEG spectrum changes. By contrast, on subsecond (delta frequency) timescales, cortical oscillations can entrain spiking activity throughout the entire LGN.

Our results are consistent with the hypothesis that, in waking animals, the koniocellular pathway selectively participates in brain circuits controlling vigilance and attention.

The major afferent cortical pathway in the visual system passes through the dorsal lateral geniculate nucleus (LGN), where nerve signals originating in the eye can first interact with brain circuits regulating visual processing, vigilance and attention. In the present study, we investigated how ongoing and visually driven activity in magnocellular (M), parvocellular (P) and koniocellular (K) layers of the LGN are related to cortical state. We recorded extracellular spiking activity in the LGN simultaneously with local field potentials (LFP) in primary visual cortex, in sufentanil‐anaesthetized marmoset monkeys. We found that asynchronous cortical states (marked by low power in delta‐band LFPs) are linked to high spike rates in K cells (but not P cells or M cells), on multisecond timescales. Cortical asynchrony precedes the increases in K cell spike rates by 1–3 s, implying causality. At subsecond timescales, the spiking activity in many cells of all (M, P and K) classes is phase‐locked to delta waves in the cortical LFP, and more cells are phase‐locked during synchronous cortical states than during asynchronous cortical states. The switch from low‐to‐high spike rates in K cells does not degrade their visual signalling capacity. By contrast, during asynchronous cortical states, the fidelity of visual signals transmitted by K cells is improved, probably because K cell responses become less rectified. Overall, the data show that slow fluctuations in cortical state are selectively linked to K pathway spiking activity, whereas delta‐frequency cortical oscillations entrain spiking activity throughout the entire LGN, in anaesthetized marmosets.
A Three-Dimensional Comprehensive Numerical Investigation of Different Operating Parameters on the Performance of a Photovoltaic Thermal System With Pancake Collector
Authors: Nahar Afroza , Hasanuzzaman M. , Rahim N. A.
Published: January 2017

Performance of photovoltaic (PV) module decreases significantly with increasing cell temperature due to its overheating. Photovoltaic thermal (PVT) is an optimized technology that facilitates effective removal and utilization of this excess heat leading to enhanced electrical performance. In this article, a 3D numerical model has been developed and analyzed to investigate the PVT performance with a new pancake-shaped flow channel design. This flow channel is attached directly to the backside of PV module by using thermal paste. The governing equations are solved numerically by using Galerkin's weighted residual finite-element method (FEM), which has been developed using COMSOL Multiphysics® software. The numerical results show that the cell temperature reduces on an average 42 °C, and the electrical efficiency and output power increase by 2% and 20 W, respectively, for both aluminum and copper channels with an increase in inlet velocity from 0.0009 to 0.05 m/s. On the other hand, overall efficiency of the PVT system drops about 13% in both cases as the inlet temperature increases from 20 °C to 40 °C. Cell temperature is found to increase approximately by 5.4 °C and 9.2 °C for every 100 W/m2 increase in irradiation level of the PV module with and without cooling system, respectively. Regarding flow channel material, it has been observed that use of either copper or aluminum produces almost similar performance of the PVT module.
Maximizing stoichiometry control in reactive sputter deposition of TiO2
Authors: Hoskins Brian D. , Strukov Dmitri B.
Published: January 2017

Thin films of amorphous TiO2 are grown by direct current (DC) reactive magnetron sputtering. Using modern models of DC reactive sputtering, conditions were established to maximize control of the O:Ti ratio by indirectly monitoring the change in ion-induced secondary electron emission of the Ti target. The Ti resistivity was continuously varied through over 12 orders of magnitude, and changes in stoichiometry were observed by Rutherford backscattering. Combining observed changes in stoichiometry with a predictive model of the composition, a percolative transition could be observed exhibiting universal and nonuniversal scaling parameters.
Rifampin Enhances the Activity of Amphotericin B against Fusarium solani Species Complex and Aspergillus flavus Species Complex Isolates from Keratitis Patients
Authors: He Yi , Zhou Lutan , Gao Chuanwen , Han Lei , Xu Yan
Published: January 2017

The
in vitro
activities of amphotericin B in combination with rifampin were assessed against 95 ocular fungal isolates. The interactions between amphotericin B and rifampin at 4, 8, 16, and 32 μg/ml were synergistic for 11.8%, 51.0%, 90.2%, and 94.1%, respectively, of
Fusarium solani
species complex isolates and for 13.6%, 45.5%, 93.2%, and 95.5%, respectively, of
Aspergillus flavus
species complex isolates. Antagonism was never observed for the amphotericin B-rifampin combinations.
Perturbations of Phosphatidate Cytidylyltransferase (CdsA) Mediate Daptomycin Resistance in Streptococcus mitis/oralis by a Novel Mechanism
Authors: Mishra Nagendra N. , Tran Truc T. , Seepersaud Ravin , Garcia-de-la-Maria Cristina , Faull Kym , Yoon Alex , Proctor Richard , Miro Jose M. , Rybak Michael J. , Bayer Arnold S. , Arias Cesar A. , Sullam Paul M.
Published: January 2017

Streptococcus mitis/oralis
is an important pathogen, causing life-threatening infections such as endocarditis and severe sepsis in immunocompromised patients. The β-lactam antibiotics are the usual therapy of choice for this organism, but their effectiveness is threatened by the frequent emergence of resistance. The lipopeptide daptomycin (DAP) has been suggested for therapy against such resistant
S. mitis/oralis
strains due to its
in vitro
bactericidal activity and demonstrated efficacy against other Gram-positive pathogens. Unlike other bacteria, however,
S. mitis/oralis
has the unique ability to rapidly develop stable, high-level resistance to DAP upon exposure to the drug both
in vivo
and
in vitro
. Using isogenic DAP-susceptible and DAP-resistant
S. mitis/oralis
strain pairs, we describe a mechanism of resistance to both DAP and cationic antimicrobial peptides that involves loss-of-function mutations in
cdsA
(encoding a phosphatidate cytidylyltransferase). CdsA catalyzes the synthesis of cytidine diphosphate-diacylglycerol, an essential phospholipid intermediate for the production of membrane phosphatidylglycerol and cardiolipin. DAP-resistant
S. mitis/oralis
strains demonstrated a total disappearance of phosphatidylglycerol, cardiolipin, and anionic phospholipid microdomains from membranes. In addition, these strains exhibited cross-resistance to cationic antimicrobial peptides from human neutrophils (i.e., hNP-1). Interestingly, CdsA-mediated changes in phospholipid metabolism were associated with DAP hyperaccumulation in a small subset of the bacterial population, without any binding by the remaining larger population. Our results indicate that CdsA is the major mediator of high-level DAP resistance in
S. mitis/oralis
and suggest a novel mechanism of bacterial survival against attack by antimicrobial peptides of both innate and exogenous origins.
Interactions of Freshwater Cyanobacteria with Bacterial Antagonists
Authors: Osman Omneya Ahmed , Beier Sara , Grabherr Manfred , Bertilsson Stefan
Published: January 2017

Cyanobacterial and algal mass development, or blooms, have severe effects on freshwater and marine systems around the world. Many of these phototrophs produce a variety of potent toxins, contribute to oxygen depletion, and affect water quality in several ways. Coexisting antagonists, such as cyanolytic bacteria, hold the potential to suppress, or even terminate, such blooms, yet the nature of this interaction is not well studied. We isolated 31 cyanolytic bacteria affiliated with the generaPseudomonas,Stenotrophomonas,Acinetobacter, andDelftiafrom three eutrophic freshwater lakes in Sweden and selected four phylogenetically diverse bacterial strains with strong-to-moderate lytic activity. To characterize their functional responses to the presence of cyanobacteria, we performed RNA sequencing (RNA-Seq) experiments on coculture incubations, with an initial predator-prey ratio of 1:1. Genes involved in central cellular pathways, stress-related heat or cold shock proteins, and antitoxin genes were highly expressed in both heterotrophs and cyanobacteria. Heterotrophs in coculture expressed genes involved in cell motility, signal transduction, and putative lytic activity.l,d-Transpeptidase was the only significantly upregulated lytic gene inStenotrophomonas rhizophilaEK20. Heterotrophs also shifted their central metabolism from the tricarboxylic acid cycle to the glyoxylate shunt. Concurrently, cyanobacteria clearly show contrasting antagonistic interactions with the four tested heterotrophic strains, which is also reflected in the physical attachment to their cells. In conclusion, antagonistic interactions with cyanobacteria were initiated within 24 h, and expression profiles suggest varied responses for the different cyanobacteria and studied cyanolytes.

IMPORTANCEHere, we present how gene expression profiles can be used to reveal interactions between bloom-forming freshwater cyanobacteria and antagonistic heterotrophic bacteria. Species-specific responses in both heterotrophs and cyanobacteria were identified. The study contributes to a better understanding of the interspecies cellular interactions underpinning the persistence and collapse of cyanobacterial blooms.
Ursodeoxycholic Acid and Its Taurine- or Glycine-Conjugated Species Reduce Colitogenic Dysbiosis and Equally Suppress Experimental Colitis in Mice
Authors: Van den Bossche Lien , Hindryckx Pieter , Devisscher Lindsey , Devriese Sarah , Van Welden Sophie , Holvoet Tom , Vilchez-Vargas Ramiro , Vital Marius , Pieper Dietmar H. , Vanden Bussche Julie , Vanhaecke Lynn , Van de Wiele Tom , De Vos Martine , Laukens Debby
Published: January 2017

The promising results seen in studies of secondary bile acids in experimental colitis suggest that they may represent an attractive and safe class of drugs for the treatment of inflammatory bowel diseases (IBD). However, the exact mechanism by which bile acid therapy confers protection from colitogenesis is currently unknown. Since the gut microbiota plays a crucial role in the pathogenesis of IBD, and exogenous bile acid administration may affect the community structure of the microbiota, we examined the impact of the secondary bile acid ursodeoxycholic acid (UDCA) and its taurine or glycine conjugates on the fecal microbial community structure during experimental colitis. Daily oral administration of UDCA, tauroursodeoxycholic acid (TUDCA), or glycoursodeoxycholic acid (GUDCA) equally lowered the severity of dextran sodium sulfate-induced colitis in mice, as evidenced by reduced body weight loss, colonic shortening, and expression of inflammatory cytokines. Illumina sequencing demonstrated that bile acid therapy during colitis did not restore fecal bacterial richness and diversity. However, bile acid therapy normalized the colitis-associated increased ratio of
Firmicutes
to
Bacteroidetes
. Interestingly, administration of bile acids prevented the loss of
Clostridium
cluster XIVa and increased the abundance of
Akkermansia muciniphila
, bacterial species known to be particularly decreased in IBD patients. We conclude that UDCA, which is an FDA-approved drug for cholestatic liver disorders, could be an attractive treatment option to reduce dysbiosis and ameliorate inflammation in human IBD.

IMPORTANCE
Secondary bile acids are emerging as attractive candidates for the treatment of inflammatory bowel disease. Although bile acids may affect the intestinal microbial community structure, which significantly contributes to the course of these inflammatory disorders, the impact of bile acid therapy on the fecal microbiota during colitis has not yet been considered. Here, we studied the alterations in the fecal microbial abundance in colitic mice following the administration of secondary bile acids. Our results show that secondary bile acids reduce the severity of colitis and ameliorate colitis-associated fecal dysbiosis at the phylum level. This study indicates that secondary bile acids might act as a safe and effective drug for inflammatory bowel disease.
Osmotolerance in Escherichia coli Is Improved by Activation of Copper Efflux Genes or Supplementation with Sulfur-Containing Amino Acids
Authors: Xiao Mengyong , Zhu Xinna , Fan Feiyu , Xu Hongtao , Tang Jinlei , Qin Ying , Ma Yanhe , Zhang Xueli
Published: January 2017

Improvement in the osmotolerance of
Escherichia coli
is essential for the production of high titers of various bioproducts. In this work, a
cusS
mutation that was identified in the previously constructed high-succinate-producing
E. coli
strain HX024 was investigated for its effect on osmotolerance. CusS is part of the two-component system CusSR that protects cells from Ag(I) and Cu(I) toxicity. Changing
cusS
from strain HX024 back to its original sequence led to a 24% decrease in cell mass and succinate titer under osmotic stress (12% glucose). When cultivated with a high initial glucose concentration (12%), introduction of the
cusS
mutation into parental strain Suc-T110 led to a 21% increase in cell mass and a 40% increase in succinate titer. When the medium was supplemented with 30 g/liter disodium succinate, the
cusS
mutation led to a 120% increase in cell mass and a 492% increase in succinate titer. Introducing the
cusS
mutation into the wild-type strain ATCC 8739 led to increases in cell mass of 87% with 20% glucose and 36% using 30 g/liter disodium succinate. The
cusS
mutation increased the expression of
cusCFBA
, and gene expression levels were found to be positively related to osmotolerance abilities. Because high osmotic stress has been associated with deleterious accumulation of Cu(I) in the periplasm, activation of CusCFBA may alleviate this effect by transporting Cu(I) out of the cells. This hypothesis was confirmed by supplementing sulfur-containing amino acids that can chelate Cu(I). Adding methionine or cysteine to the medium increased the osmotolerance of
E. coli
under anaerobic conditions.

IMPORTANCE
In this work, an activating Cus copper efflux system was found to increase the osmotolerance of
E. coli
. In addition, new osmoprotectants were identified. Supplementation with methionine or cysteine led to an increase in osmotolerance of
E. coli
under anaerobic conditions. These new strategies for improving osmotolerance will be useful for improving the production of chemicals in industrial bioprocesses.
Genome-Wide Identification of Host-Segregating Epidemiological Markers for Source Attribution in Campylobacter jejuni
Authors: Thépault Amandine , Méric Guillaume , Rivoal Katell , Pascoe Ben , Mageiros Leonardos , Touzain Fabrice , Rose Valérie , Béven Véronique , Chemaly Marianne , Sheppard Samuel K.
Published: January 2017

Campylobacter
is among the most common worldwide causes of bacterial gastroenteritis. This organism is part of the commensal microbiota of numerous host species, including livestock, and these animals constitute potential sources of human infection. Molecular typing approaches, especially multilocus sequence typing (MLST), have been used to attribute the source of human campylobacteriosis by quantifying the relative abundance of alleles at seven MLST loci among isolates from animal reservoirs and human infection, implicating chicken as a major infection source. The increasing availability of bacterial genomes provides data on allelic variation at loci across the genome, providing the potential to improve the discriminatory power of data for source attribution. Here we present a source attribution approach based on the identification of novel epidemiological markers among a reference pan-genome list of 1,810 genes identified by gene-by-gene comparison of 884 genomes of
Campylobacter jejuni
isolates from animal reservoirs, the environment, and clinical cases. Fifteen loci involved in metabolic activities, protein modification, signal transduction, and stress response or coding for hypothetical proteins were selected as host-segregating markers and used to attribute the source of 42 French and 281 United Kingdom clinical
C. jejuni
isolates. Consistent with previous studies of British campylobacteriosis, analyses performed using STRUCTURE software attributed 56.8% of British clinical cases to chicken, emphasizing the importance of this host reservoir as an infection source in the United Kingdom. However, among French clinical isolates, approximately equal proportions of isolates were attributed to chicken and ruminant reservoirs, suggesting possible differences in the relative importance of animal host reservoirs and indicating a benefit for further national-scale attribution modeling to account for differences in production, behavior, and food consumption.

IMPORTANCE
Accurately quantifying the relative contribution of different host reservoirs to human
Campylobacter
infection is an ongoing challenge. This study, based on the development of a novel source attribution approach, provides the first results of source attribution in
Campylobacter jejuni
in France. A systematic analysis using gene-by-gene comparison of 884 genomes of
C. jejuni
isolates, with a pan-genome list of genes, identified 15 novel epidemiological markers for source attribution. The different proportions of French and United Kingdom clinical isolates attributed to each host reservoir illustrate a potential role for local/national variations in
C. jejuni
transmission dynamics.
Development of
oriC
-Based Plasmids for Mesoplasma florum
Authors: Matteau Dominick , Pepin Marie-Eve , Baby Vincent , Gauthier Samuel , Arango Giraldo Mélissa , Knight Thomas F. , Rodrigue Sébastien
Published: January 2017

The near-minimal bacterium
Mesoplasma florum
constitutes an attractive model for systems biology and for the development of a simplified cell chassis in synthetic biology. However, the lack of genetic engineering tools for this microorganism has limited our capacity to understand its basic biology and modify its genome. To address this issue, we have evaluated the susceptibility of
M. florum
to common antibiotics and developed the first generation of artificial plasmids able to replicate in this bacterium. Selected regions of the predicted
M. florum
chromosomal origin of replication (
oriC
) were used to create different plasmid versions that were tested for their transformation frequency and stability. Using polyethylene glycol-mediated transformation, we observed that plasmids harboring both
rpmH-dnaA
and
dnaA-dnaN
intergenic regions, interspaced or not with a copy of the
dnaA
gene, resulted in a frequency of ∼4.1 × 10
−6
transformants per viable cell and were stably maintained throughout multiple generations. In contrast, plasmids containing only one
M. florum
oriC
intergenic region or the heterologous
oriC
region of
Mycoplasma capricolum
,
Mycoplasma mycoides
, or
Spiroplasma citri
failed to produce any detectable transformants. We also developed alternative transformation procedures based on electroporation and conjugation from
Escherichia coli
, reaching frequencies up to 7.87 × 10
−6
and 8.44 × 10
−7
transformants per viable cell, respectively. Finally, we demonstrated the functionality of antibiotic resistance genes active against tetracycline, puromycin, and spectinomycin/streptomycin in
M. florum
. Taken together, these valuable genetic tools will facilitate efforts toward building an
M. florum
-based near-minimal cellular chassis for synthetic biology.

IMPORTANCE
Mesoplasma florum
constitutes an attractive model for systems biology and for the development of a simplified cell chassis in synthetic biology.
M. florum
is closely related to the mycoides cluster of mycoplasmas, which has become a model for whole-genome cloning, genome transplantation, and genome minimization. However,
M. florum
shows higher growth rates than other
Mollicutes
, has no known pathogenic potential, and possesses a significantly smaller genome that positions this species among some of the simplest free-living organisms. So far, the lack of genetic engineering tools has limited our capacity to understand the basic biology of
M. florum
in order to modify its genome. To address this issue, we have evaluated the susceptibility of
M. florum
to common antibiotics and developed the first artificial plasmids and transformation methods for this bacterium. This represents a strong basis for ongoing genome engineering efforts using this near-minimal microorganism.
The Tat Substrate SufI Is Critical for the Ability of Yersinia pseudotuberculosis To Cause Systemic Infection
Authors: Avican Ummehan , Doruk Tugrul , Östberg Yngve , Fahlgren Anna , Forsberg Åke
Published: January 2017

The twin arginine translocation (Tat) system targets folded proteins across the inner membrane and is crucial for virulence in many important human-pathogenic bacteria. Tat has been shown to be required for the virulence of
Yersinia pseudotuberculosis
, and we recently showed that the system is critical for different virulence-related stress responses as well as for iron uptake. In this study, we wanted to address the role of the Tat substrates in
in vivo
virulence. Therefore, 22 genes encoding potential Tat substrates were mutated, and each mutant was evaluated in a competitive oral infection of mice. Interestingly, a Δ
sufI
mutant was essentially as attenuated for virulence as the Tat-deficient strain. We also verified that SufI was Tat dependent for membrane/periplasmic localization in
Y. pseudotuberculosis
.
In vivo
bioluminescent imaging of orally infected mice revealed that both the Δ
sufI
and Δ
tatC
mutants were able to colonize the cecum and Peyer's patches (PPs) and could spread to the mesenteric lymph nodes (MLNs). Importantly, at this point, neither the Δ
tatC
mutant nor the Δ
sufI
mutant was able to spread systemically, and they were gradually cleared. Immunostaining of MLNs revealed that both the Δ
tatC
and Δ
sufI
mutants were unable to spread from the initial infection foci and appeared to be contained by neutrophils, while wild-type bacteria readily spread to establish multiple foci from day 3 postinfection. Our results show that SufI alone is required for the establishment of systemic infection and is the major cause of the attenuation of the Δ
tatC
mutant.
Eyedrop Inoculation Causes Sublethal Leptospirosis in Mice
Authors: Sullivan Joseph Pierce , Nair Nisha , Potula Hari-Hara , Gomes-Solecki Maria
Published: January 2017

Leptospirosis is potentially a fatal zoonosis acquired by contact of skin and mucosal surfaces with soil and water contaminated with infected urine. We analyzed the outcome of infection of C3H/HeJ mice withLeptospira interrogansserovar Copenhageni using an enzootic mode of transmission, the conjunctival route. Infection led to weight loss andL. interrogansdissemination from blood to urine, and spirochetes were detected in blood and urine simultaneously. The infectious dose that led to consistent dissemination to kidney after conjunctival infection was ∼108leptospires. Interestingly, a lower number of spirochetes appeared to colonize the kidney, given that we quantified ∼105and ∼10 leptospires per μl of urine and per μg of kidney, respectively. Leptospira-specific IgM and IgG were detected at 15 days postinfection, and isotyping of the Ig subclass showed that the total IgG response switched from an IgG1 response to an IgG3 response after infection withL. interrogans. Histological periodic acid-Schiff D staining of infected kidney showed interstitial nephritis, mononuclear cell infiltrates, and reduced size of glomeruli. Quantification of proinflammatory immunomediators in kidney showed that keratinocyte-derived chemokine, macrophage inflammatory protein 2, RANTES, tumor necrosis factor alpha, gamma interferon, and interleukin-10 were upregulated in infected mice. We show that the kinetics of disease progression after infection via the ocular conjunctiva is delayed compared with infection via the standard intraperitoneal route. Differences may be related to the number ofL. interrogansspirochetes that succeed in overcoming the natural defenses of the ocular conjunctiva and transit through tissue.
Genome-Wide Transcriptional Response to Varying RpoS Levels in Escherichia coli K-12
Authors: Wong Garrett T. , Bonocora Richard P. , Schep Alicia N. , Beeler Suzannah M. , Lee Fong Anna J. , Shull Lauren M. , Batachari Lakshmi E. , Dillon Moira , Evans Ciaran , Becker Carla J. , Bush Eliot C. , Hardin Johanna , Wade Joseph T. , Stoebel Daniel M.
Published: January 2017

The alternative sigma factor RpoS is a central regulator of many stress responses inEscherichia coli. The level of functional RpoS differs depending on the stress. The effect of these differing concentrations of RpoS on global transcriptional responses remains unclear. We investigated the effect of RpoS concentration on the transcriptome during stationary phase in rich media. We found that 23% of genes in theE. coligenome are regulated by RpoS, and we identified many RpoS-transcribed genes and promoters. We observed three distinct classes of response to RpoS by genes in the regulon: genes whose expression changes linearly with increasing RpoS level, genes whose expression changes dramatically with the production of only a little RpoS (“sensitive” genes), and genes whose expression changes very little with the production of a little RpoS (“insensitive”). We show that sequences outside the core promoter region determine whether an RpoS-regulated gene is sensitive or insensitive. Moreover, we show that sensitive and insensitive genes are enriched for specific functional classes and that the sensitivity of a gene to RpoS corresponds to the timing of induction as cells enter stationary phase. Thus, promoter sensitivity to RpoS is a mechanism to coordinate specific cellular processes with growth phase and may also contribute to the diversity of stress responses directed by RpoS.

IMPORTANCEThe sigma factor RpoS is a global regulator that controls the response to many stresses inEscherichia coli. Different stresses result in different levels of RpoS production, but the consequences of this variation are unknown. We describe how changing the level of RpoS does not influence all RpoS-regulated genes equally. The cause of this variation is likely the action of transcription factors that bind the promoters of the genes. We show that the sensitivity of a gene to RpoS levels explains the timing of expression as cells enter stationary phase and that genes with different RpoS sensitivities are enriched for specific functional groups. Thus, promoter sensitivity to RpoS is a mechanism that coordinates specific cellular processes in response to stresses.
ThiN as a Versatile Domain of Transcriptional Repressors and Catalytic Enzymes of Thiamine Biosynthesis
Authors: Hwang Sungmin , Cordova Bryan , Abdo Merna , Pfeiffer Friedhelm , Maupin-Furlow Julie A.
Published: January 2017

Thiamine biosynthesis is commonly regulated by a riboswitch mechanism; however, the enzymatic steps and regulation of this pathway in archaea are poorly understood.
Haloferax volcanii
, one of the representative archaea, uses a eukaryote-like Thi4 (thiamine thiazole synthase) for the production of the thiazole ring and condenses this ring with a pyrimidine moiety synthesized by an apparent bacterium-like ThiC (2-methyl-4-amino-5-hydroxymethylpyrimidine [HMP] phosphate synthase) branch. Here we found that archaeal Thi4 and ThiC were encoded by leaderless transcripts, ruling out a riboswitch mechanism. Instead, a novel ThiR transcription factor that harbored an N-terminal helix-turn-helix (HTH) DNA binding domain and C-terminal ThiN (TMP synthase) domain was identified. In the presence of thiamine, ThiR was found to repress the expression of
thi4
and
thiC
by a DNA operator sequence that was conserved across archaeal phyla. Despite having a ThiN domain, ThiR was found to be catalytically inactive in compensating for the loss of ThiE (TMP synthase) function. In contrast, bifunctional ThiDN, in which the ThiN domain is fused to an N-terminal ThiD (HMP/HMP phosphate [HMP-P] kinase) domain, was found to be interchangeable for ThiE function and, thus, active in thiamine biosynthesis. A conserved Met residue of an extended α-helix near the active-site His of the ThiN domain was found to be important for ThiDN catalytic activity, whereas the corresponding Met residue was absent and the α-helix was shorter in ThiR homologs. Thus, we provide new insight into residues that distinguish catalytic from noncatalytic ThiN domains and reveal that thiamine biosynthesis in archaea is regulated by a transcriptional repressor, ThiR, and not by a riboswitch.

IMPORTANCE
Thiamine pyrophosphate (TPP) is a cofactor needed for the enzymatic activity of many cellular processes, including central metabolism. In archaea, thiamine biosynthesis is an apparent chimera of eukaryote- and bacterium-type pathways that is not well defined at the level of enzymatic steps or regulatory mechanisms. Here we find that ThiN is a versatile domain of transcriptional repressors and catalytic enzymes of thiamine biosynthesis in archaea. Our study provides new insight into residues that distinguish catalytic from noncatalytic ThiN domains and reveals that archaeal thiamine biosynthesis is regulated by a ThiN domain transcriptional repressor, ThiR, and not by a riboswitch.
CO2 diffusion into pore spaces limits weathering rate of an experimental basalt landscape
Authors: van Haren Joost , Dontsova Katerina , Barron-Gafford Greg A. , Troch Peter A. , Chorover Jon , Delong Stephen B. , Breshears David D. , Huxman Travis E. , Pelletier Jon D. , Saleska Scott R. , Zeng Xubin , Ruiz Joaquin
Published: January 2017

Basalt weathering is a key control over the global carbon cycle, though in situ measurements of carbon cycling are lacking. In an experimental, vegetation-free hillslope containing 330 m3 of ground basalt scoria, we measured real-time inorganic carbon dynamics within the porous media and seepage flow. The hillslope carbon flux (0.6–5.1 mg C m–2 h–1) matched weathering rates of natural basalt landscapes (0.4–8.8 mg C m–2 h–1) despite lacking the expected field-based impediments to weathering. After rainfall, a decrease in CO2 concentration ([CO2]) in pore spaces into solution suggested rapid carbon sequestration but slow reactant supply. Persistent low soil [CO2] implied that diffusion limited CO2 supply, while when sufficiently dry, reaction product concentrations limited further weathering. Strong influence of diffusion could cause spatial heterogeneity of weathering even in natural settings, implying that modeling studies need to include variable soil [CO2] to improve carbon cycling estimates associated with potential carbon sequestration methods.
The control of the tropical North Atlantic on Holocene millennial climate oscillations
Authors: Domínguez-Villar David , Wang Xianfeng , Krklec Kristina , Cheng Hai , Edwards R. Lawrence
Published: January 2017

Changes in ocean dynamics in the northern North Atlantic affect the thermohaline circulation that controls global climate. During glacial and deglaciation periods these dynamics are enhanced due to large variations in the surface ocean density caused by changes in glacier volumes. During full interglacial conditions, the dominant role of the northern North Atlantic on global climate is limited due to the reduced discharge of freshwater to the ocean, causing other regional dynamics to gain importance. Here we present a speleothem δ18O record from the Iberian Peninsula that supports that the northern North Atlantic and tropical North Atlantic were both source regions of millennial climate oscillations during the Holocene. The speleothem δ18O signal records millennial time-scale changes in the hydrological cycle as a result of persistent anomalies of the Gulf Stream–North Atlantic Current dynamics. In addition, the speleothem δ18O record shows synchronous variability with records from the eastern Pacific region though the entire Holocene, whereas records from western Pacific region have limited or no correlation beyond periods of major instability of the northern North Atlantic. The discontinuous climate connection among the studied records is the result of different mechanisms affecting the climate system that originated in distant regions. We suggest that two regions, the tropical North Atlantic and northern North Atlantic, alternate their dominance as source regions causing millennial climate anomalies in large planetary regions. The duration of these persistent climate changes and the extension of the regions affected depend on the region triggering the anomaly because different mechanisms affecting the climate system are involved.
Block-controlled hillslope form and persistence of topography in rocky landscapes
Authors: Glade Rachel C. , Anderson Robert S. , Tucker Gregory E.
Published: January 2017

Rocky hillslopes dotted with boulder-sized blocks and covered by a thin, nonuniform soil are common in both steep landscapes and arid environments on Earth, as well as on other planets. While the evolution of soil-mantled, convex-upward hillslopes in uniform lithology is reasonably well understood, the influence of heterogeneous lithology and geologic structure on hillslope form and evolution has yet to be properly addressed. Landscapes developed in layered sedimentary rocks feature sharp-edged landforms such as mesas and hogbacks that exhibit steep, linear to concave-upward ramps with scattered blocks calved from resistant rock layers overlying softer strata. Here we show that blocks can control the persistence of topography and the form and evolution of hillslopes in these landscapes. We present a numerical model demonstrating that incorporation of feedbacks between block release, interruption of soil creep by blocks, and sporadic downslope movement of blocks are necessary and sufficient to capture the morphology and evolution of these landscapes. Numerical results are reproduced by a simple analytical solution that predicts steady-state concave hillslope form and average slope angle from block size and spacing. Our results illuminate previously unrecognized hillslope feedbacks, advancing our understanding of the geomorphology of rocky hillslopes. On a landscape scale, our findings establish a quantitative method to address the migration of sharp edges and the persistence of topography in layered landscapes.