bluuebunny/crossref_metadata_2025 huggingface dataset:
Evaluation of Daphnid Grazing on Microscopic Zoosporic Fungi by Using Comparative Threshold Cycle Quantitative PCR
Authors: Maier Michelle A. , Uchii Kimiko , Peterson Tawnya D. , Kagami Maiko
Published: April 2016

Lethal parasitism of large phytoplankton by chytrids (microscopic zoosporic fungi) may play an important role in organic matter and nutrient cycling in aquatic environments by shunting carbon away from hosts and into much smaller zoospores, which are more readily consumed by zooplankton. This pathway provides a mechanism to more efficiently retain carbon within food webs and reduce export losses. However, challenges in accurate identification and quantification of chytrids have prevented a robust assessment of the relative importance of parasitism for carbon and energy flows within aquatic systems. The use of molecular techniques has greatly advanced our ability to detect small, nondescript microorganisms in aquatic environments in recent years, including chytrids. We used quantitative PCR (qPCR) to quantify the consumption of zoospores by
Daphnia
in laboratory experiments using a culture-based comparative threshold cycle (

C
T

) method. We successfully quantified the reduction of zoospores in water samples during
Daphnia
grazing and confirmed the presence of chytrid DNA inside the daphnid gut. We demonstrate that comparative

C
T

qPCR is a robust and effective method to quantify zoospores and evaluate zoospore grazing by zooplankton and will aid in better understanding how chytrids contribute to organic matter cycling and trophic energy transfer within food webs.

IMPORTANCE
The study of aquatic fungi is often complicated by the fact that they possess complex life cycles that include a variety of morphological forms. Studies that rely on morphological characteristics to quantify the abundances of all stages of the fungal life cycle face the challenge of correctly identifying and enumerating the nondescript zoospores. These zoospores, however, provide an important trophic link between large colonial phytoplankton and zooplankton: that is, once the carbon is liberated from phytoplankton into the parasitic zoospores, the latter are consumed by zooplankton and carbon is retained in the aquatic food web rather than exported from the system. This study provides a tool to quantify zoospores and evaluate the consumption of zoospores by zooplankton in order to further our understanding of their role in food web dynamics.
Molecular Profiling of Shiga Toxin-Producing Escherichia coli and Enteropathogenic E. coli Strains Isolated from French Coastal Environments
Authors: Balière C. , Rincé A. , Delannoy S. , Fach P. , Gourmelon M.
Published: April 2016

Shiga toxin-producing
Escherichia coli
(STEC) and enteropathogenic
E. coli
(EPEC) strains may be responsible for food-borne infections in humans. Twenty-eight STEC and 75 EPEC strains previously isolated from French shellfish-harvesting areas and their watersheds and belonging to 68 distinguishable serotypes were characterized in this study. High-throughput real-time PCR was used to search for the presence of 75
E. coli
virulence-associated gene targets, and genes encoding Shiga toxin (
stx
) and intimin (
eae
) were subtyped using PCR tests and DNA sequencing, respectively. The results showed a high level of diversity between strains, with 17 unique virulence gene profiles for STEC and 56 for EPEC. Seven STEC and 15 EPEC strains were found to display a large number or a particular combination of genetic markers of virulence and the presence of
stx
and/or
eae
variants, suggesting their potential pathogenicity for humans. Among these, an O26:H11
stx
1a
eae
-β1 strain was associated with a large number of virulence-associated genes (
n
= 47), including genes carried on the locus of enterocyte effacement (LEE) or other pathogenicity islands, such as OI-122, OI-71, OI-43/48, OI-50, OI-57, and the high-pathogenicity island (HPI). One O91:H21 STEC strain containing 4
stx
variants (
stx
1a
,
stx
2a
,
stx
2c
, and
stx
2d
) was found to possess genes associated with pathogenicity islands OI-122, OI-43/48, and OI-15. Among EPEC strains harboring a large number of virulence genes (
n
, 34 to 50), eight belonged to serotype O26:H11, O103:H2, O103:H25, O145:H28, O157:H7, or O153:H2.

IMPORTANCE
The species
E. coli
includes a wide variety of strains, some of which may be responsible for severe infections. This study, a molecular risk assessment study of
E. coli
strains isolated from the coastal environment, was conducted to evaluate the potential risk for shellfish consumers. This report describes the characterization of virulence gene profiles and
stx
/
eae
polymorphisms of
E. coli
isolates and clearly highlights the finding that the majority of strains isolated from coastal environment are potentially weakly pathogenic, while some are likely to be more pathogenic.
Strain-Specific Features of Extracellular Polysaccharides and Their Impact on Lactobacillus plantarum-Host Interactions
Authors: Lee I-Chiao , Caggianiello Graziano , van Swam Iris I. , Taverne Nico , Meijerink Marjolein , Bron Peter A. , Spano Giuseppe , Kleerebezem Michiel
Published: April 2016

Lactobacilli are found in diverse environments and are widely applied as probiotic, health-promoting food supplements. Polysaccharides are ubiquitously present on the cell surface of lactobacilli and are considered to contribute to the species- and strain-specific probiotic effects that are typically observed. Two
Lactobacillus plantarum
strains, SF2A35B and Lp90, have an obvious ropy phenotype, implying high extracellular polysaccharide (EPS) production levels. In this work, we set out to identify the genes involved in EPS production in these
L. plantarum
strains and to demonstrate their role in EPS production by gene deletion analysis. A model
L. plantarum
strain, WCFS1, and its previously constructed derivative that produced reduced levels of EPS were included as reference strains. The constructed EPS-reduced derivatives were analyzed for the abundance and sugar compositions of their EPS, revealing
cps2
-like gene clusters in SF2A35B and Lp90 responsible for major EPS production. Moreover, these mutant strains were tested for phenotypic characteristics that are of relevance for their capacity to interact with the host epithelium in the intestinal tract, including bacterial surface properties as well as survival under the stress conditions encountered in the gastrointestinal tract (acid and bile stress). In addition, the Toll-like receptor 2 (TLR2) signaling and immunomodulatory capacities of the EPS-negative derivatives and their respective wild-type strains were compared, revealing strain-specific impacts of EPS on the immunomodulatory properties. Taken together, these experiments illustrate the importance of EPS in
L. plantarum
strains as a strain-specific determinant in host interaction.

IMPORTANCE
This study evaluates the role of extracellular polysaccharides that are produced by different strains of
Lactobacillus plantarum
in the determination of the cell surface properties of these bacteria and their capacity to interact with their environment, including their signaling to human host cells. The results clearly show that the consequences of removal of these polysaccharides are very strain specific, illustrating the diverse and unpredictable roles of these polysaccharides in the environmental interactions of these bacterial strains. In the context of the use of lactobacilli as health-promoting probiotic organisms, this study exemplifies the importance of strain specificity.
Reducing Salt in Raw Pork Sausages Increases Spoilage and Correlates with Reduced Bacterial Diversity
Authors: Fougy Lysiane , Desmonts Marie-Hélène , Coeuret Gwendoline , Fassel Christine , Hamon Erwann , Hézard Bernard , Champomier-Vergès Marie-Christine , Chaillou Stéphane
Published: April 2016

Raw sausages are perishable foodstuffs; reducing their salt content raises questions about a possible increased spoilage of these products. In this study, we evaluated the influence of salt reduction (from 2.0% to 1.5% [wt/wt]), in combination with two types of packaging (modified atmosphere [50% mix of CO
2
-N
2
] and vacuum packaging), on the onset of spoilage and on the diversity of spoilage-associated bacteria. After 21 days of storage at 8°C, spoilage was easily observed, characterized by noticeable graying of the products and the production of gas and off-odors defined as rancid, sulfurous, or sour. At least one of these types of spoilage occurred in each sample, and the global spoilage intensity was more pronounced in samples stored under modified atmosphere than under vacuum packaging and in samples with the lower salt content. Metagenetic 16S rRNA pyrosequencing revealed that vacuum-packaged samples contained a higher total bacterial richness (
n
= 69 operational taxonomic units [OTUs]) than samples under the other packaging condition (
n
= 46 OTUs). The core community was composed of 6 OTUs (
Lactobacillus sakei
,
Lactococcus piscium
,
Carnobacterium divergens
,
Carnobacterium maltaromaticum
,
Serratia proteamaculans
, and
Brochothrix thermosphacta
), whereas 13 OTUs taxonomically assigned to the
Enterobacteriaceae
,
Enterococcaceae
, and
Leuconostocaceae
families comprised a less-abundant subpopulation. This subdominant community was significantly more abundant when 2.0% salt and vacuum packaging were used, and this correlated with a lower degree of spoilage. Our results demonstrate that salt reduction, particularly when it is combined with CO
2
-enriched packaging, promotes faster spoilage of raw sausages by lowering the overall bacterial diversity (both richness and evenness).

IMPORTANCE
Our study takes place in the context of raw meat product manufacturing and is linked to a requirement for salt reduction. Health guidelines are calling for a reduction in dietary salt intake. However, salt has been used for a very long time as a hurdle technology, and salt reduction in meat products raises the question of spoilage and waste of food. The study was conceived to assess the role of sodium chloride reduction in meat products, both at the level of spoilage development and at the level of bacterial diversity, using 16S rRNA amplicon sequencing and raw pork sausage as a meat model.
Detection and Molecular Characterization of Hepatitis A Virus from Tunisian Wastewater Treatment Plants with Different Secondary Treatments
Authors: Ouardani Imen , Turki Syrine , Aouni Mahjoub , Romalde Jesús L.
Published: April 2016

Hepatitis A virus (HAV) is the main causative agent of hepatitis infection associated with waterborne outbreaks worldwide. In Tunisia, there is no specific surveillance system for HAV and current secondary wastewater treatment processes are unable to remove viral particles, which present a potential public health problem. Qualitative and quantitative analysis of HAV in 271 raw and treated wastewater samples from five sewage treatment plants (STPs) during 13 months was performed. Moreover, the efficiency of three secondary wastewater treatment processes (conventional activated sludge, extended aeration, and oxidation ditch activated sludge) was evaluated. Data obtained demonstrated that HAV is endemic in Tunisia and circulates with high prevalence in both raw (66.9%) and treated (40.7%) wastewater. HAV circulates throughout the year in the coastal areas, with the highest rates found during summer and autumn, whereas in central Tunisia, high levels were shown in autumn and winter. Total virus removal was not achieved, since no difference in mean HAV loads was observed in effluents (6.0 × 10
3
genome copies [GC]/ml) and influents (2.7 × 10
3
GC/ml). The comparison of the HAV removal values of the three different wastewater treatment methods indicates that extended aeration and oxidation ditch activated sludge had better efficiency in removing viruses than conventional activated sludge did. Molecular characterization revealed that the vast majority of HAV strains belonged to subgenotype IA, with the cocirculation of subgenotype IB in wastewater treatment plants that collect tourism wastewater.

IMPORTANCE
This report provides important data on the incidence, behavior, seasonality, and genotype distribution of HAV in the environment in Tunisia, as well as the risk of infection derived from its occurrence in effluents due to inadequate wastewater treatment. In addition, these findings seem to confirm that the prevalence of HAV depends on socioeconomic level, sanitary conditions in the communities, sewage facilities, the locality, and the climate. The wide dispersion of HAV in effluents proves the inefficacity of the current wastewater treatment processes used in Tunisia to remove virus; therefore, establishment of tertiary treatment processes or replacement of the medium-charge activated sludge (conventional activated sludge) by the low-charge version (oxidation ditch activated sludge) is absolutely needed. Rapid detection of the HAV genome in wastewater may provide a timely warning sign to health authorities to implement population protection measures.
Transcriptome and Secretome Analyses of the Wood Decay Fungus Wolfiporia cocos Support Alternative Mechanisms of Lignocellulose Conversion
Authors: Gaskell Jill , Blanchette Robert A. , Stewart Philip E. , BonDurant Sandra Splinter , Adams Marie , Sabat Grzegorz , Kersten Phil , Cullen Dan
Published: April 2016

Certain wood decay basidiomycetes, collectively referred to as brown rot fungi, rapidly depolymerize cellulose while leaving behind the bulk of cell wall lignin as a modified residue. The mechanism(s) employed is unclear, but considerable evidence implicates the involvement of diffusible oxidants generated via Fenton-like chemistry. Toward a better understanding of this process, we have examined the transcriptome and secretome of
Wolfiporia cocos
when cultivated on media containing glucose, purified crystalline cellulose, aspen (
Populus grandidentata
), or lodgepole pine (
Pinus contorta
) as the sole carbon source. Compared to the results obtained with glucose, 30, 183, and 207 genes exhibited 4-fold increases in transcript levels in cellulose, aspen, and lodgepole pine, respectively. Mass spectrometry identified peptides corresponding to 64 glycoside hydrolase (GH) proteins, and of these, 17 corresponded to transcripts upregulated on one or both woody substrates. Most of these genes were broadly categorized as hemicellulases or chitinases. Consistent with an important role for hydroxyl radical in cellulose depolymerization, high transcript levels and upregulation were observed for genes involved in iron homeostasis, iron reduction, and extracellular peroxide generation. These patterns of regulation differ markedly from those of the closely related brown rot fungus
Postia placenta
and expand the number of enzymes potentially involved in the oxidative depolymerization of cellulose.

IMPORTANCE
The decomposition of wood is an essential component of nutrient cycling in forest ecosystems. Few microbes have the capacity to efficiently degrade woody substrates, and the mechanism(s) is poorly understood. Toward a better understanding of these processes, we show that when grown on wood as a sole carbon source the brown rot fungus
W. cocos
expresses a unique repertoire of genes involved in oxidative and hydrolytic conversions of cell walls.
A β-Alanine Catabolism Pathway Containing a Highly Promiscuous ω-Transaminase in the 12-Aminododecanate-Degrading Pseudomonas sp. Strain AAC
Authors: Wilding Matthew , Peat Thomas S. , Newman Janet , Scott Colin
Published: April 2016

We previously isolated the transaminase KES23458 from
Pseudomonas
sp. strain AAC as a promising biocatalyst for the production of 12-aminododecanoic acid, a constituent building block of nylon-12. Here, we report the subsequent characterization of this transaminase. It exhibits activity with a broad substrate range which includes α-, β-, and ω-amino acids, as well as α,ω-diamines and a number of other industrially relevant compounds. It is therefore a prospective candidate for the biosynthesis of a range of polyamide monomers. The crystal structure of KES23458 revealed that the protein forms a dimer containing a large active site pocket and unusual phosphorylated histidine residues. To infer the physiological role of the transaminase, we expressed, purified, and characterized a dehydrogenase from the same operon, KES23460. Unlike the transaminase, the dehydrogenase was shown to be quite selective, catalyzing the oxidation of malonic acid semialdehyde, formed from β-alanine transamination via KES23458. In keeping with previous reports, the dehydrogenase was shown to catalyze both a coenzyme A (CoA)-dependent reaction to form acetyl-CoA and a significantly slower CoA-independent reaction to form acetate. These findings support the original functional assignment of KES23458 as a β-alanine transaminase. However, a seemingly well-adapted active site and promiscuity toward unnatural compounds, such as 12-aminododecanoic acid, suggest that this enzyme could perform multiple functions for
Pseudomonas
sp. strain AAC.

IMPORTANCE
We describe the characterization of an industrially relevant transaminase able to metabolize 12-aminododecanoic acid, a constituent building block of the widely used polymer nylon-12, and we report the biochemical and structural characterization of the transaminase protein. A physiological role for this highly promiscuous enzyme is proposed based on the characterization of a related gene from the host organism. Molecular dynamics simulations were carried out to compare the conformational changes in the transaminase protein to better understand the determinants of specificity in the protein. This study makes a substantial contribution that is of interest to the broad biotechnology and enzymology communities, providing insights into the catalytic activity of an industrially relevant biocatalyst as well as the biological function of this operon.
Alternative Biotransformation of Retinal to Retinoic Acid or Retinol by an Aldehyde Dehydrogenase from Bacillus cereus
Authors: Hong Seung-Hye , Ngo Ho-Phuong-Thuy , Nam Hyun-Koo , Kim Kyoung-Rok , Kang Lin-Woo , Oh Deok-Kun
Published: April 2016

A novel bacterial aldehyde dehydrogenase (ALDH) that converts retinal to retinoic acid was first identified in
Bacillus cereus
. The amino acid sequence of ALDH from
B. cereus
(
Bc
ALDH) was more closely related to mammalian ALDHs than to bacterial ALDHs. This enzyme converted not only small aldehydes to carboxylic acids but also the large aldehyde all-
trans
-retinal to all-
trans
-retinoic acid with NAD(P)
+
. We newly found that
Bc
ALDH and human ALDH (ALDH1A1) could reduce all-
trans
-retinal to all-
trans
-retinol with NADPH. The catalytic residues in
Bc
ALDH were Glu266 and Cys300, and the cofactor-binding residues were Glu194 and Glu457. The E266A and C300A variants showed no oxidation activity. The E194S and E457V variants showed 15- and 7.5-fold higher catalytic efficiency (
k
cat
/

K
m

) for the reduction of all-
trans
-retinal than the wild-type enzyme, respectively. The wild-type, E194S variant, and E457V variant enzymes with NAD
+
converted 400 μM all-
trans
-retinal to 210 μM all-
trans
-retinoic acid at the same amount for 240 min, while with NADPH, they converted 400 μM all-
trans
-retinal to 20, 90, and 40 μM all-
trans
-retinol, respectively. These results indicate that
Bc
ALDH and its variants are efficient biocatalysts not only in the conversion of retinal to retinoic acid but also in its conversion to retinol with a cofactor switch and that retinol production can be increased by the variant enzymes. Therefore,
Bc
ALDH is a novel bacterial enzyme for the alternative production of retinoic acid and retinol.

IMPORTANCE
Although mammalian ALDHs have catalyzed the conversion of retinal to retinoic acid with NAD(P)
+
as a cofactor, a bacterial ALDH involved in the conversion is first characterized. The biotransformation of all-
trans
-retinal to all-
trans
-retinoic acid by
Bc
ALDH and human ALDH was altered to the biotransformation to all-
trans
-retinol by a cofactor switch using NADPH. Moreover, the production of all-
trans
-retinal to all-
trans
-retinol was changed by mutations at positions 194 and 457 in
Bc
ALDH. The alternative biotransformation of retinoids was first performed in the present study. These results will contribute to the biotechnological production of retinoids, including retinoic acid and retinol.
Activation of an Otherwise Silent Xylose Metabolic Pathway in Shewanella oneidensis
Authors: Sekar Ramanan , Shin Hyun Dong , DiChristina Thomas J.
Published: April 2016

Shewanella oneidensis
is unable to metabolize the sugar xylose as a carbon and energy source. In the present study, an otherwise silent xylose catabolic pathway was activated in
S. oneidensis
by following an adaptive evolution strategy. Genome-wide scans indicated that the
S. oneidensis
genome encoded two proteins similar to the xylose oxido-reductase pathway enzymes xylose reductase (SO_0900) and xylulokinase (SO_4230), and purified SO_0900 and SO_4230 displayed xylose reductase and xylulokinase activities, respectively. The
S. oneidensis
genome was missing, however, an
Escherichia coli
XylE-like xylose transporter. After 12 monthly transfers in minimal growth medium containing successively higher xylose concentrations, an
S. oneidensis
mutant (termed strain XM1) was isolated for the acquired ability to grow aerobically on xylose as a carbon and energy source. Whole-genome sequencing indicated that strain XM1 contained a mutation in an unknown membrane protein (SO_1396) resulting in a glutamine-to-histidine conversion at amino acid position 207. Homology modeling demonstrated that the Q207H mutation in SO_1396 was located at the homologous xylose docking site in XylE. The expansion of the
S. oneidensis
metabolic repertoire to xylose expands the electron donors whose oxidation may be coupled to the myriad of terminal electron-accepting processes catalyzed by
S. oneidensis
. Since xylose is a lignocellulose degradation product, this study expands the potential substrates to include lignocellulosic biomass.

IMPORTANCE
The activation of an otherwise silent xylose metabolic system in
Shewanella oneidensis
is a powerful example of how accidental mutations allow microorganisms to adaptively evolve. The expansion of the
S. oneidensis
metabolic repertoire to xylose expands the electron donors whose oxidation may be coupled to the myriad of terminal electron-accepting processes catalyzed by
S. oneidensis
. Since xylose is a lignocellulose degradation product, this study expands the potential substrates to include lignocellulosic biomass.
Developing guidance for pregnancy testing of adolescents participating in research: ethical, legal and practical considerations
Authors: Larcher Vic , Brierley Joe
Published: April 2016

Adolescents need safe effective drugs that have undergone ethically approved testing in clinical trials; such studies often require pregnancy testing in ‘women of childbearing age’ which includes children/adolescents. There is a lack of consistent standard operating procedures for pregnancy testing in these individuals, in either research or clinical (ie, both preprocedure and clinical emergency) settings. Some harmonisation between a selective or universal testing approach based on a risk analysis of the trial drug or procedure would seem sensible. The need for pregnancy testing and the reasons for the method chosen (universal or selective) should be clearly defined in the research protocol. Research ethics committees (RECs) need to satisfy themselves that the selection of subjects to be tested, the procedures for obtaining consent and the respecting of the young person's confidentiality are appropriate and that management of any positive tests are in accord with local safeguarding policies and procedures. Researchers should have core competencies necessary to manage sensitive questioning and child safeguarding training. Clinical trials of medicinal product (CTIMP) pregnancy testing in females 13–15 years of age requires parental consent and the child's active involvement in the decision-making process (‘assent’) the implications of a positive test should be discussed in advance. Children under 13 years should not normally be subject to pregnancy testing in CTIMPs, unless there are exceptional circumstances, for example, a trial of contraceptive agents of a high teratogenicity risk, as reviewed by a specialist paediatric REC. We analyse the ethical, legal and practical aspects of this issues and supply guidance to support those involved.
Recreational drug use in the Oslo nightlife setting: study protocol for a cross-sectional time series using biological markers, self-reported and qualitative data
Authors: Nordfjærn Trond , Edland-Gryt Marit , Bretteville-Jensen Anne Line , Buvik Kristin , Gripenberg Johanna
Published: April 2016

Introduction

Recreational drug use in the nightlife setting carries the risk of many negative consequences, such as violence, injuries, aberrant driving and sexual risk-taking. The aim of this study is to investigate recreational drug use and user characteristics among people visiting licensed premises, for example, nightclubs and bars, by using self-reports and biological markers. Staff of licensed premises will be asked to report drug use observations. Further, by using qualitative data, we will examine the motives, consequences and culture associated with recreational drug use. An additional aim is to compare self-reported drug use with oral fluid test (OFT) results in order to validate the different measurement methods in this context.

Methods and analyses

Data collection will be conducted among patrons (n=1000) outside licensed premises. On consent, patrons will be asked to anonymously complete a questionnaire, a breath alcohol concentration test and an OFT. Patrons who report use of recreational drugs in the previous 12 months will be asked to leave their contact information for a subsequent qualitative in-depth interview (n=30–40). Staff from licensed premises (n=500) will be invited during Responsible Beverage Service Training to participate in an anonymous survey. Survey data will be analysed by univariate and multivariate statistical methods and the oral fluids will be analysed for a large number of drugs using biochemical methods. Cohen's κ will be used as a measure of agreement between self-reported drug use and OFT. In-depth interviews will be coded in HyperRESEARCH and analysed using an inductive approach. Data collection will be repeated on a biannual basis until at least 2020, allowing for examination of trends in recreational drug use.

Ethics and dissemination

This study has been approved by the Regional Committee for Medical and Health Research Ethics. Results will be disseminated in research journals, conferences and the media.
Assisted reproductive technology and risk of asthma and allergy in the offspring: protocol for a systematic review and meta-analysis
Authors: Nwaru Bright I , McCleary Nicola , Erkkola Maijaliisa , Kaila Minna , Virtanen Suvi M , Sheikh Aziz
Published: April 2016

Introduction

The use of assisted reproductive technology (ART) procedures has increased globally over the last three decades. Recent observational studies suggest that children conceived through ART may be at increased risk of asthma and atopic disease compared with children conceived naturally, but findings are mixed. We aim to synthesise the evidence on the impact of ART on the risk of asthma and atopic disease in the offspring.

Methods and analysis

We will identify relevant studies by searching MEDLINE, EMBASE, Cochrane Library, ISI Web of Science, CINAHL, Scopus, Google Scholar, AMED, Global Health, PsychINFO, CAB International and the WHO Global Health Library from 1978 to 2016. We will locate additional studies through searching databases of the proceedings of international conferences, contacting international experts in the field, and searching the references cited in identified studies. We will include analytic observational studies (cohort studies, case–control studies and cross-sectional studies) that have investigated the impact of any type of ART on offspring's asthma and atopic disease. Screening of identified records, data extraction from eligible studies and risk of bias assessment of eligible studies will be independently undertaken by two reviewers, with arbitration by a third reviewer. The Effective Public Health Practice Project will be employed for risk of bias assessment. Estimates from studies judged to be clinically, methodologically and statistically homogeneous will be synthesised using random-effects meta-analysis.

Ethics and dissemination

As this study is based solely on the published literature, no ethics approval is required. We will publish our findings in a peer-reviewed scientific journal and present the results at national and international scientific conferences.

Protocol registration

We will register a detailed protocol for the review with the International Prospective Register of Systematic Reviews (PROSPERO) prior to starting the review.
Protocol for systematic review and meta-analysis: hop (Humulus lupulusL.) for menopausal vasomotor symptoms
Authors: Abdi Fatemeh , Kazemi Farideh , Ramezani Tehrani Fahimeh , Roozbeh Nasibeh
Published: April 2016

Introduction

Menopause is a critical stage in every woman's life. It can cause a distressing time for women by creating various vasomotor symptoms (VMS). Phytoestrogens can potentially exert various favourable effects and alleviate VMS in postmenopausal women. The hop (Humulus lupulusL.) contains 8-prenylnaringenin (8-PN), the most potent phytoestrogen known to date. The hop is eight times stronger than any other herbal oestrogens. This study aims to conduct a comprehensive systematic review and a meta-analysis survey of the effects of hop in the management of VMS in postmenopausal women.

Methods

Only randomised controlled clinical trials, with cluster randomisation and crossover, blinded and non-blinded designs, conducted between 2000 and 2015, will be included in this review. Quasi-experimental and observational studies as well as case reports will be excluded. The studies will be selected if their participants were aged 40–60 years, had elevated follicle-stimulating hormone (FSH) levels and/or menstrual irregularities, and experienced discomforting VMS (at least hot flashes or night sweats). The primary outcome will be the rate of response to treatment, such as changes in frequency and intensity of symptoms in the intervention and placebo groups. ‘Hop’, ‘Humulus’, ‘menopause’, ‘vasomotor’, ‘hot flashes’, ‘phytoestrogen’ and ‘night sweats’ will be used as search key words. Prior to their inclusion in the review, the selected papers will be assessed by two independent reviewers for methodological validity. Any disagreements will be resolved through a third reviewer. The risk of bias will be independently determined using the Cochrane Risk of Bias Tool. The quality of the papers will be assessed based on the CONSORT checklist.

Ethics and dissemination

Results will be disseminated through traditional academic literature. Dissemination of results will occur by peer-reviewed publications. The results of our project can help reproductive health researchers when evaluating the discomforts of research procedures described in study protocols or when designing a study. Information on experiences of menopausal women involved in previous studies may also help in future research. The expected dissemination actions are effective treatment in designing strategies that aim to develop women's health and healthcare providers when offering treatment for women with vasomotor symptoms.
Point-of-care lung ultrasound in young children with respiratory tract infections and wheeze
Authors: Varshney Terry , Mok Elise , Shapiro Adam J , Li Patricia , Dubrovsky Alexander S
Published: April 2016

Objective

Characterise lung ultrasound (LUS) findings, diagnostic accuracy and agreement between novice and expert interpretations in young children with respiratory tract infections and wheeze.

Methods

Prospective cross-sectional study in a paediatric ED. Patients ≤2 years with a respiratory tract infection and wheeze at triage were recruited unless in severe respiratory distress. Prior to clinical management, a novice sonologist performed the LUS using a six-zone scanning protocol. The treating physician remained blinded to ultrasound findings; final diagnoses were extracted from the medical record. An expert sonologist, blinded to all clinical information, assessed the ultrasound video clips at study completion. Positive LUS was defined as the presence of ≥1 of the following findings: ≥3 B-lines per intercostal space, consolidation and/or pleural abnormalities.

Results

Ninety-four patients were enrolled (median age 11.1 months). LUS was positive in 42% (39/94) of patients (multiple B-lines in 80%, consolidation in 64%, pleural abnormalities in 23%). The proportion of positive LUS, along with their diagnostic accuracy (sensitivity (95% CI), specificity (95% CI)), were as follows for children with bronchiolitis, asthma, pneumonia and asthma/pneumonia: 46% (45.8% (34.0% to 58.0%), 72.7% (49.8% to 89.3%)), 0% (0% (0.0% to 23.3%), 51.3% (39.8% to 62.6%)), 100% (100% (39.8% to 100.0%), 61.1% (50.3% to 71.2%)), 50% (50% (6.8% to 93.2%), 58.9% (48.0% to 69.2%)), respectively. There was good agreement between the novice and expert sonographers for a positive LUS (kappa 0.68 (95% CI 0.54 to 0.82)).

Conclusions

Among children with respiratory tract infections and wheeze, a positive LUS seems to distinguish between clinical syndromes by ruling in pneumonia and ruling out asthma. If confirmed in future studies, LUS may emerge as a point-of-care tool to guide diagnosis and disposition in young children with wheeze.
ASPECTS decay during inter-facility transfer in patients with large vessel occlusion strokes
Authors: Mokin Maxim , Gupta Rishi , Guerrero Waldo R , Rose David Z , Burgin William S , Sivakanthan Sananthan
Published: April 2016

Background

Favorable imaging profile according to the Alberta Stroke Program Early CT Score (ASPECTS) on non-contrast head CT is a key criterion for the selection of patients with ischemic stroke from large vessel occlusion (LVO) for IA revascularization therapies.

Objective

To analyze factors associated with changes in ASPECTS during inter-hospital transfer and to determine how deterioration of ASPECTS affects eligibility for endovascular procedures.

Methods

We analyzed factors associated with changes in ASPECTS during inter-hospital transfer and their potential impact on eligibility for IA stroke therapies in patients with anterior circulation ischemic strokes. Clinical and demographic characteristics between patients with favorable (ASPECTS ≥6) and unfavorable (ASPECTS <6) imaging on repeat CT were compared.

Results

Stroke evolution towards unfavorable ASPECTS occurred in 13/42 (31%) patients who initially had a favorable imaging profile at outside hospitals. A higher National Institutes of Health Stroke Scale (NIHSS) score was the only significant predictor of ASPECTS decay, whereas other clinical characteristics, such as the use of IV thrombolysis and site of LVO, were similar between the two groups.

Conclusions

In our cohort, one out of three patients became ineligible for IA thrombectomy because of unfavorable ASPECTS ‘decay’ following inter-hospital transfer. Except for NIHSS severity, baseline clinical factors could not identify which patients were at risk for ASPECTS deterioration.
Early detection of changes in lung mechanics with oscillometry following bariatric surgery in severe obesity
Authors: Peters Ubong , Hernandez Paul , Dechman Gail , Ellsmere James , Maksym Geoffrey
Published: April 2016

Obesity is associated with respiratory symptoms that are reported to improve with weight loss, but this is poorly reflected in spirometry, and few studies have measured respiratory mechanics with oscillometry. We investigated whether early changes in lung mechanics following weight loss are detectable with oscillometry. Furthermore, we investigated whether the changes in lung mechanics measured in the supine position following weight loss are associated with changes in sleep quality. Nineteen severely obese female subjects (mean body mass index, 47.2 ± 6.6 kg/m2) were evaluated using spirometry, oscillometry, plethysmography, and the Pittsburgh Sleep Quality Index before and 5 weeks after bariatric surgery. These tests were conducted in both the upright and the supine position, and pre- and postbronchodilation with 200 μg of salbutamol. Five weeks after surgery, weight loss of 11.5 ± 2.5 kg was not associated with changes in spirometry and plethysmography, with the exception of functional residual capacity. There were also no changes in upright respiratory system resistance (Rrs) or reactance following weight loss. Importantly, however, in the supine position, weight loss caused a substantial reduction in Rrs. In addition, sleep quality improved significantly and was highly correlated with the reduction in supine Rrs. Prior to weight loss, subjects did not respond to the bronchodilator when assessed in the upright position with either spirometry or oscillometry; however, with modest weight loss, bronchodilator responsiveness returned to the normal range. Improvements in lung mechanics occur very early after weight loss, mostly in the supine position, resulting in improved sleep quality. These improvements are detectable with oscillometry but not with spirometry.
Influence of habitat attributes on density of Virginia opossums (Didelphis virginiana) in agricultural ecosystems
Authors: Beatty William S. , Beasley James C. , Olson Zachary H. , Rhodes Olin E.
Published: April 2016

In agriculturally fragmented ecosystems, mesopredators play dominant roles in food webs through scavenging. We examined the influence of habitat attributes associated with carrion on local Virginia opossum (Didelphis virginiana Kerr, 1792) density in an agricultural landscape. We conducted opossum mark–recapture in 25 forest patches from 2005 to 2010, which represented the most extensive sampling of opossums to date. We analyzed mark–recapture data with a closed robust design and evaluated effects of landscape features linked to carrion on opossum density and female opossum density with generalized linear mixed-effects models. We included landscape-level (1481.6 m buffer) and patch-level covariates linked to carrion in addition to other covariates associated with high opossum densities. We developed a set of 19 candidate models and examined model fit with Akaike’s information criterion. The top model for opossum density included the density of adjoining roads, whereas the top model for female density included patch size, although the statistical null was a competing model in both cases. The long-distance dispersal capability and generalist diet of the opossum likely precluded us from detecting a definitive relationship between covariates and opossum density. The scale of effect for opossum density in agriculturally fragmented landscapes is likely larger than the spatial scales examined here.
A conceptual framework for homeostasis: development and validation
Authors: McFarland Jenny , Wenderoth Mary Pat , Michael Joel , Cliff William , Wright Ann , Modell Harold
Published: April 2016

We have developed and validated a conceptual framework for understanding and teaching organismal homeostasis at the undergraduate level. The resulting homeostasis conceptual framework details critical components and constituent ideas underlying the concept of homeostasis. It has been validated by a broad range of physiology faculty members from community colleges, primarily undergraduate institutions, research universities, and medical schools. In online surveys, faculty members confirmed the relevance of each item in the framework for undergraduate physiology and rated the importance and difficulty of each. The homeostasis conceptual framework was constructed as a guide for teaching and learning of this critical core concept in physiology, and it also paves the way for the development of a concept inventory for homeostasis.
ClueConnect: a word array game to promote student comprehension of key terminology in an introductory anatomy and physiology course
Authors: Burleson Kathryn M. , Olimpo Jeffrey T.
Published: April 2016

The sheer amount of terminology and conceptual knowledge required for anatomy and physiology can be overwhelming for students. Educational games are one approach to reinforce such knowledge. In this activity, students worked collaboratively to review anatomy and physiology concepts by creating arrays of descriptive tiles to define a term. Once guessed, students located the structure or process within diagrams of the body. The game challenged students to think about course vocabulary in novel ways and to use their collective knowledge to get their classmates to guess the terms. Comparison of pretest/posttest/delayed posttest data revealed that students achieved statistically significant learning gains for each unit after playing the game, and a survey of student perceptions demonstrated that the game was helpful for learning vocabulary as well as fun to play. The game is easily adaptable for a variety of lower- and upper-division courses.
Writing on the board as students' preferred teaching modality in a physiology course
Authors: Armour Chris , Schneid Stephen D. , Brandl Katharina
Published: April 2016

The introduction of PowerPoint presentation software has generated a paradigm shift in the delivery of lectures. PowerPoint has now almost entirely replaced chalkboard or whiteboard teaching at the undergraduate and graduate levels. This study investigated whether undergraduate biology students preferred to have lectures delivered by PowerPoint or written on the board as well as the reasons behind their preference. Two upper-division physiology courses were surveyed over a period of 7 yr. A total of 1,905 students (86.7%) indicated they preferred lectures delivered by “writing on the board” compared to 291 students (13.3%) who preferred PowerPoint. Common themes drawn from explanations reported by students in favor of writing on the board included: 1) more appropriate pace, 2) facilitation of note taking, and 3) greater alertness and attention. Common themes in favor of PowerPoint included 1) increased convenience, 2) focus on listening, and 3) more accurate and readable notes. Based on the students' very strong preference for writing on the board and the themes supporting that preference, we recommend that instructors incorporate elements of the writing on the board delivery style into whatever teaching modality is used. If instructors plan to use PowerPoint, the presentation should be paced, constructed, and delivered to provide the benefits of lectures written on the board. The advantages of writing on the board can be also incorporated into instruction intended to occur outside the classroom, such as animated narrated videos as part of the flipped classroom approach.
Preclinical Evaluation of Fatty Acid Synthase and EGFR Inhibition in Triple-Negative Breast Cancer
Authors: Giró-Perafita Ariadna , Palomeras Sònia , Lum David H. , Blancafort Adriana , Viñas Gemma , Oliveras Glòria , Pérez-Bueno Ferran , Sarrats Ariadna , Welm Alana L. , Puig Teresa
Published: April 2016

Purpose: Triple-negative breast cancer (TNBC) lacks an approved targeted therapy. Despite initial good response to chemotherapy, 30% of the patients relapse within 5 years after treatment. EGFR overexpression is a common marker in TNBC, and its expression has been correlated with poor outcome. Inhibition of fatty acid synthase (FASN) activity leads to apoptosis of human carcinoma cells overexpressing FASN. We tested the hypothesis that blocking FASN in combination with anti-EGFR signaling agents would be an effective antitumor strategy in sensitive and chemoresistant TNBC.

Experimental Design: Several TNBC cell lines and 29 primary tumors were included to determine whether FASN is a potential target in TNBC. Doxorubicin-resistant TNBC cell lines (231DXR and HCCDXR) have been developed and characterized in our laboratory. Cellular and molecular interactions of anti-FASN compounds (EGCG and C75) with cetuximab were analyzed. In vivo tumor growth inhibition was evaluated after cetuximab, EGCG, or the combination in TNBC orthoxenograft models.

Results: TNBC cell lines showed overexpression of FASN enzyme and its inhibition correlated to FASN levels. FASN staining was observed in all of the 29 TNBC tumor samples. In vitro, EGCG and C75 plus cetuximab showed strong synergism in sensitive and chemoresistant cells. In vivo, the combination of EGCG with cetuximab displayed strong antitumor activity against the sensitive and chemoresistant TNBC orthoxenografts, without signs of toxicity.

Conclusions: Our results show that the simultaneous blockade of FASN and EGFR is effective in preclinical models of sensitive and chemoresistant TNBC. Clin Cancer Res; 22(18); 4687–97. ©2016 AACR.
MSK1-Mediated β-Catenin Phosphorylation Confers Resistance to PI3K/mTOR Inhibitors in Glioblastoma
Authors: Wu Shaofang , Wang Shuzhen , Zheng Siyuan , Verhaak Roel , Koul Dimpy , Yung W.K. Alfred
Published: April 2016

Glioblastoma (GBM) represents a compelling disease for kinase inhibitor therapy because most of these tumors harbor genetic alterations that result in aberrant activation of growth factor–signaling pathways. The PI3K/mammalian target of the rapamycin (mTOR) pathway is dysregulated in over 50% of human GBM but remains a challenging clinical target. Inhibitors against PI3K/mTOR mediators have limited clinical efficacy as single agents. We investigated potential bypass mechanisms to PI3K/mTOR inhibition using gene expression profiling before and after PI3K inhibitor treatment by Affymetrix microarrays. Mitogen- and stress-activated protein kinase 1 (MSK1) was markedly induced after PI3K/mTOR inhibitor treatment and disruption of MSK1 by specific shRNAs attenuated resistance to PI3K/mTOR inhibitors in glioma-initiating cells (GIC). Further investigation showed that MSK1 phosphorylates β-catenin and regulates its nuclear translocation and transcriptional activity. The depletion of β-catenin potentiated PI3K/mTOR inhibitor-induced cytotoxicity and the inhibition of MSK1 synergized with PI3K/mTOR inhibitors to extend survival in an intracranial animal model and decreased phosphorylation of β-catenin at Ser552. These observations suggest that MSK1/β-catenin signaling serves as an escape survival signal upon PI3K/mTOR inhibition and provides a strong rationale for the combined use of PI3K/mTOR and MSK1/β-catenin inhibition to induce lethal growth inhibition in human GBM. Mol Cancer Ther; 15(7); 1656–68. ©2016 AACR.
Personalized Oncology Meets Immunology: The Path toward Precision Immunotherapy
Authors: Mandal Rajarsi , Chan Timothy A.
Published: April 2016

Personalized oncology aims to tailor therapy by targeting the unique genetic characteristics of a patient's tumor, whereas cancer immunotherapy focuses on activating the patient's immune system to control the tumor. The fusion of these ostensibly separate strategies has created a new dimension for personalized cancer immunotherapy. This entails the development of next-generation cancer vaccines that target neoantigens as well as the use of mutational signatures as predictive biomarkers for clinical response. The optimal use of immunotherapeutic agents will hinge on a robust understanding of the mutational profile of a cancer's genome that significantly dictates antitumor immunity and immunotherapeutic response.

Significance: Cancer immunotherapy has provided substantial clinical benefit in a significant number of patients with advanced disease. However, the need for more precise immunotherapies and predictive biomarkers remains pressing. Recent progress in these areas has been promising and has created a framework for precision immune-oncology. Cancer Discov; 6(7); 703–13. ©2016 AACR.