bluuebunny/crossref_metadata_2025 huggingface dataset:
The effect of implant placement in patients with either Kennedy class II and III on oral health‐related quality of life: a prospective clinical trial
Authors: van Eekeren P. J. A. , Aartman I. H. A. , Tahmaseb A. , Wismeijer D.
Published: November 2015

There is little evidence of the effect of implants restored with fixed partial dentures on oral health‐related quality of life (OHRQoL) in partially edentulous Kennedy class II and III patients. The aim of this study was to determine the change in OHRQoL in Kennedy classification II and III patients treated with a two‐implant‐supported fixed dental prosthesis (FDP). Kennedy class II and III patients received dental implants and an FDP. Oral health‐related quality of life was measured by administration of the Oral Health Impact Profile‐14 (OHIP‐14NL) questionnaire at intake (T1), 2 weeks after surgery (T2) and after 1 year of loading (T3). The mean OHIP score at T1 was 6·5 ± 1·2, 2·4 ± 1·0 at T2, and 0·9 ± 0·3 at T3. There was a statistically significant difference between T1 and T2 (P = 0·002) and T1 and T3 (P < 0·001) but not between T2 and T3 (P = 0·407). The OHIP score in Kennedy II patients decreased from 4·8 ± 3·2 at T1 to 1·5 ± 2·0 at T2 and 1·1 ± 1·8 at T3, and that in Kennedy III patients decreased from 8·9 ± 9·6 at T1 to 3·6 ± 8·9 at T2 and 0·8 ± 2·2 at T3. There were no statistically significant differences in the reductions in Kennedy II and III patients. Oral health‐related quality of life changed positively in patients treated with implants and an FDP in both groups. There was no change in OHRQoL between the times of implant placement and FDP placement.
Parents’ and caregivers’ perspectives on the Manitoba Dental Association's Free First Visit program
Authors: Schroth Robert J. , Ndayisenga Stephanie , Guenther Katelyn , Marchessault Gail , Prowse Sarah , Hai‐Santiago Khalida , Edwards Jeanette M. , Moffatt Michael E.K. ,   the Manitoba Dental Association
Published: November 2015

Objectives

The Free First Visit (FFV) program was implemented in 2010 to promote early preventive dental visits for children <36 months of age in Manitoba, Canada. The purpose was to understand parents’ and caregivers’ perspectives on the program.

Methods

Three focus groups with 21 participants were conducted in Winnipeg, Canada using an interview guide in this qualitative study.

Results

Most participants were aware of the FFV program and the appropriate age for a child's first visit. Almost all agreed with the recommendation to see a dentist by one year of age. Some reported that general dentists advised them to bring their child after three years of age. Participants appreciated that the program was free, and some noted that the program made them aware of the appropriate age for a first visit. About half of the participants had taken their child for a FFV. Reasons for not taking their child for a FFV included: nothing wrong with their child's teeth, they believed there was still time as their child was not three years old, they had government insurance, child's temperament, and feelings of apathy. There were mixed opinions regarding whether the program was helping those who needed it the most.

Conclusions

The majority of participants liked the FFV program and believed that it should continue. Parents would benefit from further education and encouragement to seek oral care for their child by age one. Some general dentists may need further training and skills to meet the recommendations for first dental visits.
Effect of Intravaginal Prasterone on Sexual Dysfunction in Postmenopausal Women with Vulvovaginal Atrophy
Authors: Labrie Fernand , Derogatis Leonard , Archer David F. , Koltun William , Vachon Andrée , Young Douglas , Frenette Louise , Portman David , Montesino Marlene , Côté Isabelle , Parent Julie , Lavoie Lyne , Beauregard Adam , Martel Céline , Vaillancourt Mario , Balser John , Moyneur Érick ,   Members of the VVA Prasterone Research Group
Published: November 2015

Introduction

Previous data have shown that intravaginal dehydroepiandrosterone (DHEA, prasterone) improved all the domains of sexual function, an effect most likely related to the local formation of androgens from DHEA.

Aims

To confirm in a placebo-controlled, prospective, double-blind and randomized study the benefits of daily intravaginal DHEA for 12 weeks on sexual function using the Female Sexual Function Index (FSFI) questionnaire.

Methods

Placebo was administered daily to 157 women while 325 women received 0.50% (6.5 mg) DHEA daily for 12 weeks. All women were postmenopausal meeting the criteria of vulvovaginal atrophy (VVA), namely moderate to severe dyspareunia as their most bothersome symptom of VVA in addition to having ≤5% of vaginal superficial cells and vaginal pH > 5.0. The FSFI questionnaire was filled at baseline (screening and day 1), 6 weeks and 12 weeks. Comparison between DHEA and placebo of the changes from baseline to 12 weeks was made using the analysis of covariance test, with treatment group as the main factor and baseline value as the covariate.

Main Outcome Measures

The six domains and total score of the FSFI questionnaire were evaluated.

Results

The FSFI domain desire increased over placebo by 0.24 unit (+49.0%, P  = 0.0105), arousal by 0.42 unit (+56.8%, P  = 0.0022), lubrication by 0.57 unit (+36.1%, P  = 0.0005), orgasm by 0.32 unit (+33.0%, P  = 0.047), satisfaction by 0.44 unit (+48.3%, P  = 0.0012), and pain at sexual activity by 0.62 unit (+39.2%, P  = 0.001). The total FSFI score, on the other hand, has shown a superiority of 2.59 units in the DHEA group over placebo or a 41.3% greater change than placebo (P  = 0.0006 over placebo).

Conclusion

The present data show that all the six domains of the FSFI are improved over placebo (from P  = 0.047 to 0.0005), thus confirming the previously observed benefits of intravaginal DHEA on female sexual dysfunction by an action exerted exclusively at the level of the vagina, in the absence of biologically significant changes of serum steroids levels.
Quantifying the reefscape transformation of a coastal Caribbean coral reef during a phase shift and the associated coastal landscape change
Authors: Rendis Abigail Martínez‐ , Acosta González Gilberto , Hernández‐Stefanoni José Luis , Arias González Jesús Ernesto
Published: November 2015

Caribbean coral reefs are under strong natural and human pressures and many have undergone phase shifts as a result of local and global change. However few studies have quantified the extent of these phase shifts and the potential impacts that they have on coral reef communities. A temporal comparative analysis of several coral reefscape and landscape metrics was conducted for data from Mahahual, Quintana Roo, Mexico, between the years 2000 and 2006, when a phase shift occurred. Landscape metrics were calculated from coral reef and coastal cover maps obtained by multispectral satellite image classification using IKONOS satellite imagery. These metrics showed that the coastal landscape and reefscape of Mahahual lost 85 ha of vegetation cover and 43 ha of coral cover over 6 years, respectively. Coastal landscape transformation was induced by the construction of a cruise ship pier and multiple associated tourist developments along the coast, such as hotels and restaurants, meanwhile alteration of the reefscape may be associated with potential stressors such as coastal development, bleaching events and hurricanes.
Population genomic footprints of fine‐scale differentiation between habitats in Mediterranean blue tits
Authors: Szulkin M. , Gagnaire P.‐A. , Bierne N. , Charmantier A.
Published: November 2015

Linking population genetic variation to the spatial heterogeneity of the environment is of fundamental interest to evolutionary biology and ecology, in particular when phenotypic differences between populations are observed at biologically small spatial scales. Here, we applied restriction‐site associated DNA sequencing (RAD‐Seq) to test whether phenotypically differentiated populations of wild blue tits (Cyanistes caeruleus) breeding in a highly heterogeneous environment exhibit genetic structure related to habitat type. Using 12 106 SNPs in 197 individuals from deciduous and evergreen oak woodlands, we applied complementary population genomic analyses, which revealed that genetic variation is influenced by both geographical distance and habitat type. A fine‐scale genetic differentiation supported by genome‐ and transcriptome‐wide analyses was found within Corsica, between two adjacent habitats where blue tits exhibit marked differences in breeding time while nesting < 6 km apart. Using redundancy analysis (RDA), we show that genomic variation remains associated with habitat type when controlling for spatial and temporal effects. Finally, our results suggest that the observed patterns of genomic differentiation were not driven by a small proportion of highly differentiated loci, but rather emerged through a process such as habitat choice, which reduces gene flow between habitats across the entire genome. The pattern of genomic isolation‐by‐environment closely matches differentiation observed at the phenotypic level, thereby offering significant potential for future inference of phenotype‐genotype associations in a heterogeneous environment.
Red fluorescence in coral larvae is associated with a diapause‐like state
Authors: Strader Marie E. , Aglyamova Galina V. , Matz Mikhail V.
Published: November 2015

Effective dispersal across environmental gradients is the key to species resilience to environmental perturbation, including climate change. Coral reefs are among the most sensitive ecosystems to global warming, but factors predicting coral dispersal potential remain unknown. In a reef‐building coral Acropora millepora, larval fluorescence emerged as a possible indicator of dispersal potential since it correlates with responsiveness to a settlement cue. Here, we show that gene expression in red fluorescent larvae of A. millepora is correlated with diapause‐like characteristics highly likely to be associated with extended dispersal. We compared gene expression among three larval fluorescent morphs under three coloured light treatments. While colour morphs did not differ in their gene expression responses to light colour, red larvae demonstrated gene expression signatures of cell cycle arrest and decreased transcription accompanied by elevated ribosome production and heightened defenses against oxidative stress. A meta‐analysis revealed that this profile was highly similar to the signatures of elevated thermal tolerance in the same coral species, and moreover, functionally resembled diapause states in an insect and a nematode. Our results support a connection between red fluorescence and long‐range dispersal, which offers a new perspective on the molecular underpinnings of coral larval dispersal and the biological function of GFP‐like fluorescent proteins.
A systematic review to determine the most effective interventions to increase water intake
Authors: Chua Teresa X.W. , Prasad Neha S. , Rangan Gopala K. , Allman‐Farinelli Margaret , Rangan Anna M.
Published: November 2015

Background

Maintaining adequate fluid intake has been hypothesized to be beneficial for the progression of chronic kidney disease (CKD). The aim of this study was to undertake a systematic review to determine the most effective interventions to increase water intake.

Methods

Six electronic databases were searched from 1910 until March 2015 in the English language. Additional sources through hand‐searches, expert recommendations and reviews were checked. Intervention studies increasing water intake in adults through non‐pharmacological methods were eligible for inclusion. The quality of included studies was assessed.

Results

A total of 950 studies were found of which 16 met the inclusion criteria. Eight studies were randomized controlled trials, and seven studies spanned 6 months or longer. The study populations varied and included patients with recurrent nephrolithiasis (n = 6), autosomal dominant polycystic kidney disease (n = 3), CKD (n = 1), urinary tract infection (n = 1) and other miscellaneous conditions (n = 5). The quality of the studies was mostly neutral (63%) with no studies of high quality. Interventions ranged from instruction alone to self‐monitoring tools, providing water bottles and counselling and education. Most interventions successfully increased water intake with 13 studies reporting an increase of at least 500 mL. The most effective strategies were instruction and self‐monitoring using urine dipstick or 24 h urine volume.

Conclusion

All interventions carried out in the studies succeeded in increasing water intake, with none leading to decreases in intake, and these could be implemented in potential clinical trials in CKD. However, more high quality long‐term intervention studies are required to further validate findings.
Hypodontia, a prospective predictive marker for tumor?
Authors: Yin W , Bian Z
Published: November 2015

Tooth agenesis and tumor are two totally different diseases occurring at different ages. In the past 10 years, more and more evidences suggested there was a relationship between them. High prevalence of breast, colon, lung, and ovary tumor was observed in tooth agenesis patients. But it is still controversial. Therefore, to have a greater understanding of the possible association, a critical review on molecular association for genes involving tooth agenesis and tumorigenesis is necessary. In this current review, we summarized the reported cases of tooth agenesis with different kinds of tumors and the molecular relationship between these two diseases through causative genes. The results indicated tooth agenesis might be a prospective predictive marker for tumor. Through this review, we want to draw more attention on this topic and hope it will be an effective way to predict the risk of tumor.
Biogeographic patterns of nutrient resorption from Quercus variabilis Blume leaves across China
Authors: Sun X. , Kang H. , Chen H. Y. H. , Björn B. , Samuel B. F. , Liu C.
Published: November 2015

The variation in nutrient resorption has been studied at different taxonomic levels and geographic ranges. However, the variable traits of nutrient resorption at the individual species level across its distribution are poorly understood. We examined the variability and environmental controls of leaf nutrient resorption of Quercus variabilis, a widely distributed species of important ecological and economic value in China. The mean resorption efficiency was highest for phosphorus (P), followed by potassium (K), nitrogen (N), sulphur (S), magnesium (Mg) and carbon (C). Resorption efficiencies and proficiencies were strongly affected by climate and respective nutrients concentrations in soils and green leaves, but had little association with leaf mass per area. Climate factors, especially growing season length, were dominant drivers of nutrient resorption efficiencies, except for C, which was strongly related to green leaf C status. In contrast, green leaf nutritional status was the primary controlling factor of leaf nutrient proficiencies, except for C. Resorption efficiencies of N, P, K and S increased significantly with latitude, and were negatively related to growing season length and mean annual temperature. In turn, N, P, K and S in senesced leaves decreased with latitude, likely due to their efficient resorption response to variation in climate, but increased for Mg and did not change for C. Our results indicate that the nutrient resorption efficiency and proficiency of Q. variabilis differed strongly among nutrients, as well as growing environments. Our findings provide important insights into understanding the nutrient conservation strategy at the individual species level and its possible influence on nutrient cycling.
Genotypic characterisation of human papillomavirus infections among persons living with HIV infection; a case–control study in Kumasi, Ghana
Authors: Yar Denis Dekugmen , Salifu Samson Pandam , Darko Samuel Nkansah , Annan Augustina Angelina , Gyimah Akosua Adumea , Buabeng Kwame Ohene , Owusu‐Dabo Ellis
Published: November 2015

Objectives

The objective of this study is to describe the burden of human papillomavirus (HPV) infection among women living with HIV and non‐infected women in Ghana.

Methods

A case–control study was conducted involving 107 women living with HIV aged between 18 and 59 years (cases) and 100 non‐HIV‐infected apparently healthy women (controls) who were recruited from the Kumasi South Hospital, from July to December, 2014. Cervicovaginal swabs were taken from study participants to characterise 28 high‐ and low‐risk HPV genotypes using a multiplex real‐time PCR.

Results

The overall mean age for the participants was 40.10 ± 9.76 years. The prevalence of high‐risk (hr)‐HPV genotypes was significantly higher among the cases than the controls (77.4% vs. 41.6%, P < 0.0001). Overall, HPV 58 and 54 were the most predominant high‐risk (18.8%) and low‐risk (15.0%) genotypes detected. The two most common hr‐HPV genotype isolates were 58 (18.8%) and 35 (15.9%) with 58 being the most prevalent among age group 35–44 years compared with hr‐HPV 16, 18, 35 and 45, found predominantly among 18–34 age group.

Conclusions

Significant variations exist in HPV genotypes among HIV‐infected and uninfected women.
Cord blood banking and quality issues
Authors: Murphy Amanda , McKenna David , McCullough Jeffrey
Published: November 2015

BACKGROUND

Food and Drug Administration guidelines are designed to assure the quality and safety of the cord blood product used for transplantation. It is valuable to determine whether the actions called for in these guidelines are effective.

STUDY DESIGN AND METHODS

We applied our cell therapy quality system to all cord blood units shipped into our cellular therapy laboratory for transplant at the University of Minnesota between 2011 and 2013. The quality issues were categorized as likely, potentially, or unlikely to have a clinical impact.

RESULTS

A total of 249 units of umbilical cord blood (UCB) were received from 16 cord blood banks. A total of 159 units (64%) had a total of 245 issues. Of these, 117 (48%) pertained to medical history, 120 (49%) to quality control, and eight (3%) to labeling and documentation. Units with quality issues were no more likely to fail to engraft, and no specific kind of quality issue was associated with failure to engraft. Compared to a similar study 10 years ago, there was a decrease in the number of issues per unit.

DISCUSSION

The cost of collecting, testing, processing, and storing UCB is very high. However, there may be activities that do not contribute to the quality or safety of the cord blood. The guidelines could be reviewed to determine their value based on years of experience.
Sympathetic‐mediated activation versus suppression of the immune system: consequences for hypertension
Authors: Case Adam J. , Zimmerman Matthew C.
Published: November 2015

It is generally well‐accepted that the immune system is a significant contributor in the pathogenesis of hypertension. Specifically, activated and pro‐inflammatory T‐lymphocytes located primarily in the vasculature and kidneys appear to have a causal role in exacerbating elevated blood pressure. It has been proposed that increased sympathetic nerve activity and noradrenaline outflow associated with hypertension may be primary contributors to the initial activation of the immune system early in the disease progression. However, it has been repeatedly demonstrated in many different human and experimental diseases that sympathoexcitation is immunosuppressive in nature. Moreover, human hypertensive patients have demonstrated increased susceptibility to secondary immune insults like infections. Thus, it is plausible, and perhaps even likely, that in diseases like hypertension, specific immune cells are activated by increased noradrenaline, while others are in fact suppressed. We propose a model in which this differential regulation is based upon activation status of the immune cell as well as the resident organ. With this, the concept of global immunosuppression is obfuscated as a viable target for hypertension treatment, and we put forth the concept of focused organ‐specific immunotherapy as an alternative option.

image
Weakly nonlinear acoustic wave propagation in a nonlinear orthotropic circular cylindrical waveguide
Authors: Prakash S Vijay , Sonti Venkata R.
Published: November 2015

Nonlinear acoustic wave propagation is considered in an infinite orthotropic thin circular cylindrical waveguide. The modes are non-planar having small but finite amplitude. The fluid is assumed to be ideal and inviscid with no mean flow. The cylindrical waveguide is modeled using the Donnell's nonlinear theory for thin cylindrical shells. The approximate solutions for the acoustic velocity potential are found using the method of multiple scales (MMS) in space and time. The calculations are presented up to the third order of the small parameter. It is found that at some frequencies the amplitude modulation is governed by the Nonlinear Schrödinger Equation (NLSE). The first objective is to study the nonlinear term in the NLSE, as the sign of the nonlinear term determines the stability of the amplitude modulation. On the other hand, at other specific frequencies, interactions occur between the primary wave and its higher harmonics. Here, the objective is to identify the frequencies of the higher harmonic interactions. Lastly, the linear terms in the NLSE obtained using the MMS calculations are validated. All three objectives are met using an asymptotic analysis of the dispersion equation.
The effects of ipsilateral, contralateral, and bilateral broadband noise on the mid-level hump in intensity discrimination
Authors: Roverud Elin , Strickland Elizabeth A.
Published: November 2015

Previous psychoacoustical and physiological studies indicate that the medial olivocochlear reflex (MOCR), a bilateral, sound-evoked reflex, may lead to improved sound intensity discrimination in background noise. The MOCR can decrease the range of basilar-membrane compression and can counteract effects of neural adaptation from background noise. However, the contribution of these processes to intensity discrimination is not well understood. This study examined the effect of ipsilateral, contralateral, and bilateral noise on the “mid-level hump.” The mid-level hump refers to intensity discrimination Weber fractions (WFs) measured for short-duration, high-frequency tones which are poorer at mid levels than at lower or higher levels. The mid-level hump WFs may reflect a limitation due to basilar-membrane compression, and thus may be decreased by the MOCR. The noise was either short (50 ms) or long (150 ms), with the long noise intended to elicit the sluggish MOCR. For a tone in quiet, mid-level hump WFs improved with ipsilateral noise for most listeners, but not with contralateral noise. For a tone in ipsilateral noise, WFs improved with contralateral noise for most listeners, but only when both noises were long. These results are consistent with MOCR-induced WF improvements, possibly via decreases in effects of compression and neural adaptation.
Tone-burst auditory brainstem response wave V latencies in normal-hearing and hearing-impaired ears
Authors: Lewis James D. , Kopun Judy , Neely Stephen T. , Schmid Kendra K. , Gorga Michael P.
Published: November 2015

The metric used to equate stimulus level [sound pressure level (SPL) or sensation level (SL)] between ears with normal hearing (NH) and ears with hearing loss (HL) in comparisons of auditory function can influence interpretation of results. When stimulus level is equated in dB SL, higher SPLs are presented to ears with HL due to their reduced sensitivity. As a result, it may be difficult to determine if differences between ears with NH and ears with HL are due to cochlear pathology or level-dependent changes in cochlear mechanics. To the extent that level-dependent changes in cochlear mechanics contribute to auditory brainstem response latencies, comparisons between normal and pathologic ears may depend on the stimulus levels at which comparisons are made. To test this hypothesis, wave V latencies were measured in 16 NH ears and 15 ears with mild-to-moderate HL. When stimulus levels were equated in SL, latencies were shorter in HL ears. However, latencies were similar for NH and HL ears when stimulus levels were equated in SPL. These observations demonstrate that the effect of stimulus level on wave V latency is large relative to the effect of HL, at least in cases of mild-to-moderate HL.
Mutant Alleles of
lptD
Increase the Permeability of Pseudomonas aeruginosa and Define Determinants of Intrinsic Resistance to Antibiotics
Authors: Balibar Carl J. , Grabowicz Marcin
Published: November 2015

Gram-negative bacteria provide a particular challenge to antibacterial drug discovery due to their cell envelope structure. Compound entry is impeded by the lipopolysaccharide (LPS) of the outer membrane (OM), and those molecules that overcome this barrier are often expelled by multidrug efflux pumps. Understanding how efflux and permeability affect the ability of a compound to reach its target is paramount to translating
in vitro
biochemical potency to cellular bioactivity. Herein, a suite of
Pseudomonas aeruginosa
strains were constructed in either a wild-type or efflux-null background in which mutations were engineered in LptD, the final protein involved in LPS transport to the OM. These mutants were demonstrated to be defective in LPS transport, resulting in compromised barrier function. Using isogenic strain sets harboring these newly created alleles, we were able to define the contributions of permeability and efflux to the intrinsic resistance of
P. aeruginosa
to a variety of antibiotics. These strains will be useful in the design and optimization of future antibiotics against Gram-negative pathogens.
Mechanism of Action of ME1111, a Novel Antifungal Agent for Topical Treatment of Onychomycosis
Authors: Takahata Sho , Kubota Natsuki , Takei-Masuda Naomi , Yamada Tsuyoshi , Maeda Mari , Alshahni Mohamed Mahdi , Abe Shigeru , Tabata Yuji , Maebashi Kazunori
Published: November 2015

Despite the existing treatment options for onychomycosis, there remains a strong demand for potent topical medications. ME1111 is a novel antifungal agent that is active against dermatophytes, has an excellent ability to penetrate human nails, and is being developed as a topical agent for onychomycosis. In the present study, we investigated its mechanism of action.
Trichophyton mentagrophytes
mutants with reduced susceptibility to ME1111 were selected in our laboratory, and genome sequences were determined for 3 resistant mutants. The inhibitory effect on a candidate target was evaluated by a spectrophotometric enzyme assay using mitochondrial fractions. Point mutations were introduced into candidate genes by a reverse genetics approach. Whole-genome analysis of the 3 selected mutants revealed point mutations in the structural regions of genes encoding subunits of succinate dehydrogenase (complex II). All of the laboratory-generated resistant mutants tested harbored a mutation in one of the subunits of succinate dehydrogenase (SdhB, SdhC, or SdhD). Most of the mutants showed cross-resistance to carboxin and boscalid, which are succinate dehydrogenase inhibitors. ME1111 strongly inhibited the succinate-2,6-dichlorophenolindophenol reductase reaction in
Trichophyton rubrum
and
T. mentagrophytes
(50% inhibitory concentrations [IC
50
s] of 0.029 and 0.025 μg/ml, respectively) but demonstrated only moderate inhibition of the same reaction in human cell lines. Furthermore, the target protein of ME1111 was confirmed by the introduction of point mutations causing the amino acid substitutions in SdhB, SdhC, and SdhD found in the laboratory-generated resistant mutants, which resulted in reduced susceptibility to ME1111. Thus, ME1111 is a novel inhibitor of the succinate dehydrogenase of
Trichophyton
species, and its mechanism of action indicates its selective profile.
Evaluating 5-Nitrothiazoles as Trypanocidal Agents
Authors: O'Shea Ivan P. , Shahed Mohammed , Aguilera-Venegas Benjamín , Wilkinson Shane R.
Published: November 2015

The growth-inhibitory properties of a 5-nitrothiazole series were evaluated against
Trypanosoma brucei
. A subset of related compounds displayed the greatest potency toward the parasite while exhibiting little cytotoxic effect on mammalian cells, with this antiparasitic activity dependent on expression of a type I nitroreductase by the trypanosome. We conclude that the 5-nitrothiazole class of nitroheterocyclic drugs may represent a new lead in the treatment of human African trypanosomiasis.
Antagonism of Fluconazole and a Proton Pump Inhibitor against Candida albicans
Authors: Liu Ning-Ning , Köhler Julia R.
Published: November 2015

Hospitalized ill patients, at risk for invasive candidiasis, often receive multiple medications, including proton pump inhibitors (PPIs). The antifungal fluconazole perturbs the vacuolar proton ATPase. The PPI omeprazole antagonized
Candida albicans
growth inhibition by fluconazole. A
C. albicans
codon-adapted pHluorin, Ca.pHluorin, was generated to measure cytosolic pH. The fungal cytosol was acidified by omeprazole and realkalinized by coexposure to fluconazole. Vacuolar pH was alkalinized by fluconazole. Off-target effects of any medication on fungal pathogens may occur.
Exposing a β-Lactamase “Twist”: the Mechanistic Basis for the High Level of Ceftazidime Resistance in the C69F Variant of the Burkholderia pseudomallei PenI β-Lactamase
Authors: Papp-Wallace Krisztina M. , Becka Scott A. , Taracila Magdalena A. , Winkler Marisa L. , Gatta Julian A. , Rholl Drew A. , Schweizer Herbert P. , Bonomo Robert A.
Published: November 2015

Around the world,
Burkholderia
spp. are emerging as pathogens highly resistant to β-lactam antibiotics, especially ceftazidime. Clinical variants of
Burkholderia pseudomallei
possessing the class A β-lactamase PenI with substitutions at positions C69 and P167 are known to demonstrate ceftazidime resistance. However, the biochemical basis for ceftazidime resistance in class A β-lactamases in
B. pseudomallei
is largely undefined. Here, we performed site saturation mutagenesis of the C69 position and investigated the kinetic properties of the C69F variant of PenI from
B. pseudomallei
that results in a high level of ceftazidime resistance (2 to 64 mg/liter) when expressed in
Escherichia coli
. Surprisingly, quantitative immunoblotting showed that the steady-state protein levels of the C69F variant β-lactamase were ∼4-fold lower than those of wild-type PenI (0.76 fg of protein/cell versus 4.1 fg of protein/cell, respectively). However, growth in the presence of ceftazidime increases the relative amount of the C69F variant to greater than wild-type PenI levels. The C69F variant exhibits a branched kinetic mechanism for ceftazidime hydrolysis, suggesting there are two different conformations of the enzyme. When incubated with an anti-PenI antibody, one conformation of the C69F variant rapidly hydrolyzes ceftazidime and most likely contributes to the higher levels of ceftazidime resistance observed in cell-based assays. Molecular dynamics simulations suggest that the electrostatic characteristics of the oxyanion hole are altered in the C69F variant. When ceftazidime was positioned in the active site, the C69F variant is predicted to form a greater number of hydrogen-bonding interactions than PenI with ceftazidime. In conclusion, we propose “a new twist” for enhanced ceftazidime resistance mediated by the C69F variant of the PenI β-lactamase based on conformational changes in the C69F variant. Our findings explain the biochemical basis of ceftazidime resistance in
B. pseudomallei
, a pathogen of considerable importance, and suggest that the full repertoire of conformational states of a β-lactamase profoundly affects β-lactam resistance.
Potential of Tetracycline Resistance Proteins To Evolve Tigecycline Resistance
Authors: Linkevicius Marius , Sandegren Linus , Andersson Dan I.
Published: November 2015

Tigecycline is a glycylcycline antibiotic active against multidrug-resistant bacterial pathogens. The objectives of our study were to examine the potential of the Tet(A), Tet(K), Tet(M), and Tet(X) tetracycline resistance proteins to acquire mutations causing tigecycline resistance and to determine how this affects resistance to earlier classes of tetracyclines. Mutations in all four
tet
genes caused a significant increase in the tigecycline MIC in
Escherichia coli
, and strains expressing mutant Tet(A) and Tet(X) variants reached clinically relevant MICs (2 mg/liter and 3 mg/liter, respectively). Mutations predominantly accumulated in transmembrane domains of the efflux pumps, most likely increasing the accommodation of tigecycline as a substrate. All selected Tet(M) mutants contained at least one mutation in the functionally most important loop III of domain IV. Deletion of leucine 505 of this loop led to the highest increase of the tigecycline MIC (0.5 mg/liter) among Tet(M) mutants. It also caused collateral sensitivity to earlier classes of tetracyclines. A majority of the Tet(X) mutants showed increased activity against all three classes of tetracylines. All tested Tet proteins have the potential to acquire mutations leading to increased MICs of tigecycline. As
tet
genes are widely found in pathogenic bacteria and spread easily by horizontal gene transfer, resistance development by alteration of existing Tet proteins might compromise the future medical use of tigecycline. We predict that Tet(X) might become the most problematic future Tet determinant, since its weak intrinsic tigecycline activity can be mutationally improved to reach clinically relevant levels without collateral loss in activity to other tetracyclines.
Development and
In Vivo
Evaluation of a Novel Histatin-5 Bioadhesive Hydrogel Formulation against Oral Candidiasis
Authors: Kong Eric F. , Tsui Christina , Boyce Heather , Ibrahim Ahmed , Hoag Stephen W. , Karlsson Amy J. , Meiller Timothy F. , Jabra-Rizk Mary Ann
Published: November 2015

Oral candidiasis (OC), caused by the fungal pathogen
Candida albicans
, is the most common opportunistic infection in HIV
+
individuals and other immunocompromised populations. The dramatic increase in resistance to common antifungals has emphasized the importance of identifying unconventional therapeutic options. Antimicrobial peptides have emerged as promising candidates for therapeutic intervention due to their broad antimicrobial properties and lack of toxicity. Histatin-5 (Hst-5) specifically has exhibited potent anticandidal activity indicating its potential as an antifungal agent. To that end, the goal of this study was to design a biocompatible hydrogel delivery system for Hst-5 application. The bioadhesive hydroxypropyl methylcellulose (HPMC) hydrogel formulation was developed for topical oral application against OC. The new formulation was evaluated
in vitro
for gel viscosity, Hst-5 release rate from the gel, and killing potency and, more importantly, was tested
in vivo
in our mouse model of OC. The findings demonstrated a controlled sustained release of Hst-5 from the polymer and rapid killing ability. Based on viable
C. albicans
counts recovered from tongues of treated and untreated mice, three daily applications of the formulation beginning 1 day postinfection with
C. albicans
were effective in protection against development of OC. Interestingly, in some cases, Hst-5 was able to clear existing lesions as well as associated tissue inflammation. These findings were confirmed by histopathology analysis of tongue tissue. Coupled with the lack of toxicity as well as anti-inflammatory and wound-healing properties of Hst-5, the findings from this study support the progression and commercial feasibility of using this compound as a novel therapeutic agent.
Susceptibility of Meningococcal Strains Responsible for Two Serogroup B Outbreaks on U.S. University Campuses to Serum Bactericidal Activity Elicited by the MenB-4C Vaccine
Authors: Rossi Raffaella , Beernink Peter T. , Giuntini Serena , Granoff Dan M.
Published: November 2015

In 2013 and 2014, two U.S. universities had meningococcal serogroup B outbreaks (a total of 14 cases) caused by strains from two different clonal complexes. To control the outbreaks, students were immunized with a serogroup B meningococcal vaccine (Novartis) that was not yet licensed in the United States. The vaccine (referred to as MenB-4C) contains four components capable of eliciting bactericidal activity. Both outbreak strains had high expression levels of two of the vaccine antigens (subfamily B factor H binding protein [FHbp] and neisserial heparin binding antigen [NHba]); the university B outbreak strain also had moderate expression of a third antigen, NadA. We investigated the bactericidal activity of sera from mice immunized with FHbp, NHba, or NadA and sera from MenB-4C-immunized infant macaques and an adult human. The postimmunization bactericidal activity of the macaque or human serum against isolates from university B with FHbp identification (ID) 1 that exactly matched the vaccine FHbp sequence variant was 8- to 21-fold higher than that against isolates from university A with FHbp ID 276 (96% identity to the vaccine antigen). Based on the bactericidal activity of mouse antisera to FHbp, NadA, or NHba and macaque or human postimmunization serum that had been depleted of anti-FHbp antibody, the bactericidal activity against both outbreak strains largely or entirely resulted from antibodies to FHbp. Thus, despite the high level of strain expression of FHbp from a subfamily that matched the vaccine antigen, there can be large differences in anti-FHbp bactericidal activity induced by MenB-4C vaccination. Further, strains with moderate to high NadA and/or NHba expression can be resistant to anti-NadA or anti-NHba bactericidal activity elicited by MenB-4C vaccination.
The Vaccine Candidate Substrate Binding Protein SBP2 Plays a Key Role in Arginine Uptake, Which Is Required for Growth of Moraxella catarrhalis
Authors: Otsuka Taketo , Kirkham Charmaine , Brauer Aimee , Koszelak-Rosenblum Mary , Malkowski Michael G. , Murphy Timothy F.
Published: November 2015

Moraxella catarrhalis
is an exclusively human pathogen that is an important cause of otitis media in children and lower respiratory tract infections in adults with chronic obstructive pulmonary disease. A vaccine to prevent
M. catarrhalis
infections would have an enormous global impact in reducing morbidity resulting from these infections. Substrate binding protein 2 (SBP2) of an ABC transporter system has recently been identified as a promising vaccine candidate antigen on the bacterial surface of
M. catarrhalis
. In this study, we showed that SBP1, -2, and -3 individually bind different basic amino acids with exquisite specificity. We engineered mutants that each expressed a single SBP from this gene cluster and showed in growth experiments that SBP1, -2, and -3 serve a nutritional function through acquisition of amino acids for the bacterium. SBP2 mediates uptake of arginine, a strict growth requirement of
M. catarrhalis
. Adherence and invasion assays demonstrated that SBP1 and SBP3 play a role in invasion of human respiratory epithelial cells, consistent with a nutritional role in intracellular survival in the human respiratory tract. This work demonstrates that the SBPs of an ABC transporter system function in the uptake of basic amino acids to support growth of
M. catarrhalis
. The critical role of SBP2 in arginine uptake may contribute to its potential as a vaccine antigen.
The Chromosome-Encoded Hypothetical Protein TC0668 Is an Upper Genital Tract Pathogenicity Factor of Chlamydia muridarum
Authors: Conrad Turner Allen , Gong Siqi , Yang Zhangsheng , Matulich Patrick , Keck Jonathon , Beltrami Noah , Chen Chaoqun , Zhou Zhou , Dai Jin , Zhong Guangming
Published: November 2015

We previously associated a missense mutation of the
tc0668
gene of serial
in vitro
-passaged
Chlamydia muridarum
, a murine model of human urogenital
C. trachomatis
, with severely attenuated disease development in the upper genital tract of female mice. Since these mutants also contained a TC0237 Q117E missense mutation that enhances their
in vitro
infectivity, an effort was made here to isolate and characterize a
tc0668
single mutant to determine its individual contribution to urogenital pathogenicity. Detailed genetic analysis of
C. muridarum
passages revealed a truncated variant with a G216* nonsense mutation of the 408-amino-acid TC0668 protein that does not produce a detectable product. Intracellular growth and infectivity of
C. muridarum in vitro
remain unaffected in the absence of TC0668. Intravaginal inoculation of the TC0668 null mutant into C3H/HeJ mice results in a typical course of lower genital tract infection but, unlike a pathogenic isogenic control, is unable to elicit significant chronic inflammation of the oviduct and fails to induce hydrosalpinx. Thus, TC0668 is demonstrated as an important chromosome-encoded urogenital pathogenicity factor of
C. muridarum
and the first with these characteristics to be discovered for a
Chlamydia
pathogen.
Enhanced Direct Major Histocompatibility Complex Class I Self-Antigen Presentation Induced by Chlamydia Infection
Authors: Cram Erik D. , Simmons Ryan S. , Palmer Amy L. , Hildebrand William H. , Rockey Daniel D. , Dolan Brian P.
Published: November 2015

The direct major histocompatibility complex (MHC) class I antigen presentation pathway ensures intracellular peptides are displayed at the cellular surface for recognition of infected or transformed cells by CD8+cytotoxic T lymphocytes.Chlamydiaspp. are obligate intracellular bacteria and, as such, should be targeted by CD8+T cells. It is likely thatChlamydiaspp. have evolved mechanisms to avoid the CD8+killer T cell responses by interfering with MHC class I antigen presentation. Using a model system of self-peptide presentation which allows for posttranslational control of the model protein's stability, we tested the ability of variousChlamydiaspecies to alter direct MHC class I antigen presentation. Infection of the JY lymphoblastoid cell line limited the accumulation of a model host protein and increased presentation of the model-protein-derived peptides. Enhanced self-peptide presentation was detected only when presentation was restricted todefectiveribosomalproducts, or DRiPs, and total MHC class I levels remained unaltered. Skewed antigen presentation was dependent on a bacterial synthesized component, as evidenced by reversal of the observed phenotype upon preventing bacterial transcription, translation, and the inhibition of bacterial lipooligosaccharide synthesis. These data suggest thatChlamydiaspp. have evolved to alter the host antigen presentation machinery to favor presentation of defective and rapidly degraded forms of self-antigen, possibly as a mechanism to diminish the presentation of peptides derived from bacterial proteins.
Regulation of a Protein Acetyltransferase in Myxococcus xanthus by the Coenzyme NADP
+
Authors: Liu Xin-Xin , Liu Wei-bing , Ye Bang-Ce
Published: November 2015

NADP
+
is a vital cofactor involved in a wide variety of activities, such as redox potential and cell death. Here, we show that NADP
+
negatively regulates an acetyltransferase from
Myxococcus xanthus
, Mxan_3215 (
Mx
Kat), at physiologic concentrations.
Mx
Kat possesses an NAD(P)-binding domain fused to the Gcn5-type
N
-acetyltransferase (GNAT) domain. We used isothermal titration calorimetry (ITC) and a coupled enzyme assay to show that NADP
+
bound to
Mx
Kat and that the binding had strong effects on enzyme activity. The Gly11 residue of
Mx
Kat was confirmed to play an important role in NADP
+
binding using site-directed mutagenesis and circular dichroism spectrometry. In addition, using mass spectrometry, site-directed mutagenesis, and a coupling enzymatic assay, we demonstrated that
Mx
Kat acetylates acetyl coenzyme A (acetyl-CoA) synthetase (Mxan_2570) at Lys622 in response to changes in NADP
+
concentration. Collectively, our results uncovered a mechanism of protein acetyltransferase regulation by the coenzyme NADP
+
at physiological concentrations, suggesting a novel signaling pathway for the regulation of cellular protein acetylation.

IMPORTANCE
Microorganisms have developed various protein posttranslational modifications (PTMs), which enable cells to respond quickly to changes in the intracellular and extracellular milieus. This work provides the first biochemical characterization of a protein acetyltransferase (
Mx
Kat) that contains a fusion between a GNAT domain and NADP
+
-binding domain with Rossmann folds, and it demonstrates a novel signaling pathway for regulating cellular protein acetylation in
M. xanthus
. We found that NADP
+
specifically binds to the Rossmann fold of
Mx
Kat and negatively regulates its acetyltransferase activity. This finding provides novel insight for connecting cellular metabolic status (NADP
+
metabolism) with levels of protein acetylation, and it extends our understanding of the regulatory mechanisms underlying PTMs.
Biased Gene Conversion in Rhizobium etli Is Caused by Preferential Double-Strand Breaks on One of the Recombining Homologs
Authors: Yáñez-Cuna Fares Osam , Castellanos Mildred , Romero David
Published: November 2015

Gene conversion, the nonreciprocal transfer of information during homologous recombination, is the main process that maintains identity between members of multigene families. Gene conversion in the nitrogenase (
nifH
) multigene family of
Rhizobium etli
was analyzed by using a two-plasmid system, where each plasmid carried a copy of
nifH
. One of the
nifH
copies was modified, creating restriction fragment length polymorphisms (RFLPs) spaced along the gene. Once the modified plasmid was introduced into
R. etli
, selection was done for cointegration with a resident plasmid lacking the RFLPs. Most of the cointegrate molecules harbor gene conversion events, biased toward a gain of RFLPs. This bias may be explained under the double-strand break repair model by proposing that the
nifH
gene lacking the RFLPs suffers a DNA double-strand break, so the incoming plasmid functions as a template for repairing the homolog on the resident plasmid. To support this proposal, we cloned an SceI site into the
nifH
homolog that had the RFLPs used for scoring gene conversion.
In vivo
expression of the meganuclease I-SceI allowed the generation of a double-strand break on this homolog. Upon introduction of this modified plasmid into an
R. etli
strain lacking I-SceI, biased gene conversion still favored the retention of markers on the incoming plasmid. In contrast, when the recipient strain ectopically expressed I-SceI, a dramatic reversal in gene conversion bias was seen, favoring the preservation of resident sequences. These results show that biased gene conversion is caused by preferential double-strand breaks on one of the recombining homologs.

IMPORTANCE
In this work, we analyzed gene conversion by using a system that entails horizontal gene transfer followed by homologous recombination in the recipient cell. Most gene conversion events are biased toward the acquisition of the incoming sequences, ranging in size from 120 bp to 800 bp. This bias is due to preferential cutting of resident DNA and can be reversed upon introduction of a double-strand break on the incoming sequence. Since conditions used in this work are similar to those in horizontal gene transfer, it provides evidence that, upon transfer, the resident DNA preferentially acquires gene variants.