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This research investigated the interpersonal impact of self-reported mental health diagnoses and/or perceptions on undergraduate students’ current or most recent romantic relationship. Analysis of data from a 43-item online questionnaire completed by 267 undergraduates revealed that 68.3% of women and 52.5% of men reported having either been professionally diagnosed with a mental illness or perceive themselves to be mentally ill based on DSM criteria, with women and white students reporting significantly higher levels. Sociologically speaking, mental illness was found to influence relationship initiation, maintenance, and dissolution in this study. The mental health of the respondents’ potential partners was an important consideration in deciding to form a relationship, particularly for male, white, heterosexuals. When the respondents reported relationship problems, men were more likely to blame such problems on mental health issues than women. Finally, though more respondents reported having broken up with a romantic partner who had mental health issues than had romantic partners break up with them, there were no significant gender, race, or sexual orientation differences in the termination of these romantic relationships. Study findings emphasize the importance of acknowledging and providing mental health resources—particularly interpersonal options—for emerging adults in the college setting.

In January 2021, a new academic unit (School of Engineering Design and Teaching Innovation) was created in the Faculty of Engineering for the first time in over 20 years. After identifying needs in the job market for programs with a focus on multidisciplinary skills, the academic unit’s first task was to develop a three-year nonaccredited program, titled “Bachelor’s of Multidisciplinary Design (Internship)”. Fundamentally, the program will include core technical expertise expected from an engineering faculty but will also offer flexibility for students to explore other interests. This type of flexibility and openness may seem daunting for first year students. Therefore, example learning paths were created based on current job market trends, with more learning paths in development. This paper focuses not only on the development of such a unique program in Canada, considering insight provided through stakeholder and focus group meetings, but also the challenges associated with submitting this program proposal through all levels of university bureaucracy, from the faculty committees through to provincial review. While there have been questions, and sometimes doubts, that such a program could be fully developed, the program is on track to pass all levels of approval and welcome its first cohort of students for the Fall 2023 term.

Creutzfeldt-Jakob disease (CJD) is a rapidly progressive, fatal degenerative encephalopathy caused by a pathologically altered form of the prion protein (PrP). CJD is rare, with 1 and 2 cases per million per year reported in the general population, mostly in individuals over 50 years of age. It is almost unknown in the pediatric population. Sporadic CJD with unusually long survival (sCJD-LS), an unusual clinicopathological variant of CJD, has been described mostly in Japanese patients. We present here the first case report of pediatric CJD-LS occurring sporadically in a teenage girl of European descent, with initially rapid neurocognitive decline followed by a prolonged (∼10 years) clinical course. Neuropathological findings at autopsy included generalized cerebral and cerebellar atrophy with relative sparing of the hippocampi, cerebral and cerebellar white and gray matter involvement, minimal spongiform change, PrP deposits in the neocortex, striatum and cerebellum by immunohistochemistry, and protease-resistant PrP by Western immunoblot. With its longer disease duration and atypical manifestations of white matter loss, CJD-LS can be clinically mistaken for other neurodegenerative diseases, or in the pediatric setting for metabolic/genetic conditions. This case clearly demonstrates that with rapid-onset encephalopathy, prion disease should be carefully considered, even in younger patients with slower disease progression.

This research investigated the relationship consequences of disclosing sexual secrets to a romantic partner. Analyses of data from a 39-item Internet questionnaire completed by 195 undergraduate students showed that revealing sex secrets to a romantic partner was associated with either neutral or positive relationship outcomes. Disclosure of sexual secrets almost never (

Background Although HIV pre-exposure prophylaxis (PrEP) substantially diminishes the likelihood of HIV acquisition, poor adherence can decrease the HIV-protective benefits of PrEP. The present investigation sought to identify the extent to which alcohol consumption, substance use, and depression were linked to PrEP nonadherence among gay, bisexual, and other men-who-have-sex-with-men (gbMSM). Methods gbMSM (age ≥ 18, prescribed PrEP for ≥3 months) were recruited from two clinics in Toronto, Canada for an e-survey assessing demographics; PrEP nonadherence (4-day PrEP-focused ACTG assessment); hazardous and harmful alcohol use (AUDIT scores of 8–15 and 16+, respectively); moderate/high risk substance use (NIDA M-ASSIST scores > 4); depression (CESD-10 scores ≥10); and other PrEP-relevant factors. The primary outcome, PrEP nonadherence, entailed missing one or more PrEP doses over the past 4 days. A linear-by-linear test of association assessed whether increasing severity of alcohol use (i.e., based on AUDIT categories) was linked to a greater occurrence of PrEP nonadherence. Univariate logistic regression was employed to determine factors associated with PrEP nonadherence, and factors demonstrating univariate associations at the p Results A total of 141 gbMSM (Mean age = 37.9, white = 63.1%) completed the e-survey. Hazardous/harmful drinking (31.9%), moderate/high risk substance use (43.3%), and depression (23.7%) were common; and one in five participants (19.9%) reported PrEP nonadherence. Increasing alcohol use level was significantly associated with a greater likelihood of nonadherence (i.e., 15.6, 25.0, and 44.4% of low-risk, hazardous, and harmful drinkers reported nonadherence, respectively (χ2(1) = 4.79, p = .029)). Multivariate logistic regression demonstrated that harmful alcohol use (AOR = 6.72, 95%CI = 1.49–30.33, p = .013) and moderate/high risk cocaine use (AOR = 3.11, 95%CI = 1.01–9.59, p = .049) independently predicted nonadherence. Furthermore, an additive association emerged, wherein the likelihood of PrEP nonadherence was highest among those who were hazardous/harmful drinkers and moderate/high risk cocaine users (OR = 2.25, 95%CI = 1.19–4.25, p = .013). Depression was not associated with nonadherence. Conclusions Findings highlight the need to integrate alcohol- and substance-focused initiatives into PrEP care for gbMSM. Such initiatives, in turn, may help improve PrEP adherence and reduce the potential for HIV acquisition among this group.

Pre-exposure prophylaxis (PrEP) is traditionally prescribed by HIV specialist physicians. Given finite specialist resources, there is a need to scale up PrEP delivery by decentralizing services via other healthcare professionals. We aimed to assess the feasibility of delivering PrEP to men who have sex with men (MSM) through primary care physicians and sexual health clinic nurses. We piloted a multi-component, implementation and dissemination research program to increase provision of PrEP through primary care physicians and sexual health clinic nurses in Toronto, Canada. Community-based organizations (CBOs) provided prospective participants with information cards that contained links to an online module on engaging providers in a conversation about PrEP. In our patient-initiated continuing medical education (PICME) strategy, participants saw their family doctors and gave them the card, which also contained a link to a Continuing Medical Education module. In the nurse-led strategy, participants visited one of two participating clinics to obtain PrEP. We administered an optional online questionnaire to patients and providers at baseline and six months. CBOs distributed 3043 cards. At least 339 men accessed the online module and 196 completed baseline questionnaires. Most (55%) intended to visit nurses while 21% intended to consult their physicians. Among 45 men completing follow-up questionnaires at 6 months, 31% reported bringing cards to their physicians and obtaining PrEP through them; sexual health clinics delivered PrEP to 244 patients. Participants who went through the PICME approach reported no changes in relationships with their providers. Nurses showed fidelity to PrEP prescribing guidelines. Nurse-led PrEP and patient-initiated continuing medical education (PICME) for primary care physicians are feasible strategies to increase PrEP uptake. Nurse-led PrEP delivery was preferred by most patients.

Prospectively acquired Canadian cerebrospinal fluid samples were used to assess the performance characteristics of three ante-mortem tests commonly used to support diagnoses of Creutzfeldt–Jakob disease. The utility of the end-point quaking-induced conversion assay as a test for Creutzfeldt–Jakob disease diagnoses was compared to that of immunoassays designed to detect increased amounts of the surrogate markers 14-3-3γ and hTau. The positive predictive values of the end-point quaking-induced conversion, 14-3-3γ, and hTau tests conducted at the Prion Diseases Section of the Public Health Agency of Canada were 96%, 68%, and 66%, respectively.

Background Gay, bisexual and other men who have sex with men (gbMSM) in Canada continue to experience high rates of incident HIV. Pre-exposure prophylaxis (PrEP, the regular use of anti-HIV medication) reduces HIV acquisition and could reduce incidence. However, there are too few physicians with expertise in HIV care to meet the projected demand for PrEP. To meet demand and achieve greater public health impact, PrEP delivery could be ‘decentralized’ by incorporating it into front-line prevention services provided by family physicians (FPs) and sexual health clinic nurses. Methods This PrEP decentralization project will use two strategies. The first is an innovative knowledge dissemination approach called ‘Patient-Initiated CME’ (PICME), which aims to empower individuals to connect their family doctors with online, evidence-based, continuing medical education (CME) on PrEP. After learning about the project through community agencies or social/sexual networking applications, gbMSM interested in PrEP will use a uniquely coded card to access an online information module that includes coaching on how to discuss their HIV risk with their FP. They can provide their physician a link to the accredited CME module using the same card. The second strategy involves a pilot implementation program, in which gbMSM who do not have a FP may bring the card to designated sexual health clinics where trained nurses can deliver PrEP under a medical directive. These approaches will be evaluated through quantitative and qualitative methods, including: questionnaires administered to patients and physicians at baseline and at six months; focus groups with patients, FPs, and sexual health clinic staff; and review of sexual health clinic charts. The primary objective is to quantify the uptake of PrEP achieved using each decentralization strategy. Secondary objectives include a) characterizing barriers and facilitators to PrEP uptake for each strategy, b) assessing fidelity to core components of PrEP delivery within each strategy, c) measuring patient-reported outcomes including satisfaction with clinician-patient relationships, and d) conducting a preliminary costing analysis. Discussion This study will assess the feasibility of a novel strategy for disseminating knowledge about evidence-based clinical interventions, and inform future strategies for scale-up of an underutilized HIV prevention tool.

Background Bacterially-produced recombinant prion protein (rPrP) has traditionally been used for in vitro fibrillation assays and reagent development for prion disease research. In recent years, it has also been used as a substrate for real-time quaking-induced conversion (RT-QuIC), a very sensitive method of detecting the presence of the misfolded, disease-associated isoform of the prion protein (PrP d ). Multi-centre trials have demonstrated that RT-QuIC is a suitably reliable and robust technique for clinical practice; however, in the absence of a commercial supplier of rPrP as a substrate for RT-QuIC, laboratories have been required to independently generate this key component of the assay. No harmonized method for producing the protein has been agreed upon, in part due to the variety of substrates that have been applied in RT-QuIC. Methods This study examines the effects of two different rPrP refolding protocols on the production, QuIC performance, and structure characteristics of two constructs of rPrP commonly used in QuIC: full length hamster and a sheep-hamster chimeric rPrP. Results Under the described conditions, the best performing substrate was the chimeric sheep-hamster rPrP produced by shorter guanidine-HCl exposure and faster gradient elution. Conclusions The observation that different rPrP production protocols influence QuIC performance indicates that caution should be exercised when comparing inter-laboratory QuIC results.

Atlantic salmon (Salmo salar L.) populations from different river origins mix in the North Atlantic during the marine life stage. To facilitate marine stock identification, we developed a genetic baseline covering the European component of the species’ range excluding the Baltic Sea, from the Russian River Megra in the north-east, the Icelandic Ellidaar in the west, and the Spanish Ulla in the south, spanning 3737 km North to South and 2717 km East to West. The baseline encompasses data for 14 microsatellites for 26 822 individual fish from 13 countries, 282 rivers, and 467 sampling sites. A hierarchy of regional genetic assignment units was defined using a combination of distance-based and Bayesian clustering. At the top level, three assignment units were identified comprising northern, southern, and Icelandic regions. A second assignment level was also defined, comprising eighteen and twenty-nine regional units for accurate individual assignment and mixed stock estimates respectively. The baseline provides the most comprehensive geographical coverage for an Atlantic salmon genetic data-set, and a unique resource for the conservation and management of the species in Europe. It is freely available to researchers to facilitate identification of the natal origin of European salmon.

Creutzfeldt-Jakob disease (CJD) is a fatal neurological illness for which accurate diagnosis is paramount. Real-time quaking-induced conversion (RT-QuIC) is a prion-specific assay with high sensitivity and specificity for CJD. The Canadian endpoint quaking-induced conversion (EP-QuIC) test is similar, but unlike RT-QuIC there is little data regarding its diagnostic utility in clinical practice. In this exploratory predictive value analysis of EP-QuIC in CJD, the negative predictive value (NPV) and positive predictive value (PPV) was 100% and 83%, respectively, with one false-positive result identified. Re-testing this sample with an optimized EP-QuIC protocol eliminated this false-positive result, leading to a PPV of 100%.La valeur prédictive de la méthode diagnostique dite de conversion provoquée par tremblement au point final dans le cas de la maladie de Creutzfeldt-Jakob. La maladie de Creutzfeldt-Jakob (MCJ) est une maladie neurologique qui entraîne à terme un décès et pour laquelle l’établissement d’un diagnostic précis est primordial. La conversion provoquée par tremblement en temps réel (RT-QuIC en anglais) est une méthode diagnostique qui repose sur la détection de la protéine prion. Dans le cas de la MCJ, cette méthode est réputée posséder une sensibilité et une spécificité élevées. La méthode canadienne dite de conversion provoquée par tremblement au point final (EP-QuIC en anglais) se veut similaire mais, à la différence de la RT-QuIC, il existe peu de données en ce qui concerne son utilité diagnostique dans le cadre d’une pratique clinique. Dans cette analyse prédictive exploratoire de la EP-QuIC en lien avec la MCJ, la valeur prédictive négative (VPN) et la valeur prédictive positive (VPP) ont été respectivement de 100 % et de 83 %, un seul résultat faux-positif ayant été identifié. Le fait de soumettre notre échantillon à un nouveau test effectué à l’aide d’un protocole d’EP-QuIC optimisé a permis d’éliminer ce résultat faux-positif, ce qui a débouché sur une VPP de 100 %.

Background More providers are needed to deliver PrEP at scale. We examined intentions to seek PrEP from family physicians (FPs) and sexual health clinic nurses (RNs) within an implementation science study on decentralizing PrEP delivery to gay, bisexual and other men who have sex with men (gbMSM). Methods Strategy A was a knowledge dissemination intervention in which community organizations distributed info-cards to gbMSM considering PrEP. Men used the cards to view an online module and meet with their FPs, who could use the card to complete an accredited e-module about PrEP. Strategy B was an implementation intervention in which gbMSM could instead access PrEP from sexual health RNs. Participants completed an optional survey at baseline and 6 months. We used descriptive statistics to characterize the sample and logistic regression to identify characteristics associated with intentions to seek PrEP from FPs vs RNs. Results From 3013 cards distributed, 339 men accessed the module; 179 completed the baseline survey and are included in this analysis. Median (IQR) age was 31 (26,40) years, 97.7% were cisgender males and 46.4% had a prior bacterial STI. Most (n=119, 66.5%) had a FP, of which only 59.5% were ‘out’ to them. Of 97 respondents with a FP and wanting to start PrEP, 35.1% vs 65.0% intended to use Strategy A vs B respectively (p=0.003). In univariable analyses, characteristics associated with intent to approach FPs included being ‘out’ to that doctor (OR=10.67, 95%CI=3.35, 33.96), very good/excellent physician skills in general communication (OR=3.42, 95%CI=1.38, 8.48) and participatory decision-making (OR=3.33, 95%CI=1.14, 9.79). In multivariable analysis, being ‘out’ was the only significant predictor (aOR=14.35, 95%CI=1.59, 129.83). Conclusion Among gbMSM with a FP, sexual health clinic RNs were preferred over FPs for PrEP by most. Multiple strategies are needed to increase numbers of PrEP providers, including interventions to help gbMSM feel comfortable disclosing sexual orientation. Disclosure No significant relationships.

Undergraduates (196) at a large southeastern university completed a 59 item Internet survey on threesome attitudes and behaviors. Fifteen percent of the sample reported having engaged in a threesome. Men evidenced more interest in having a threesome than women and persons most likely to report having had a threesome held hedonistic compared to relativistic or absolutist sexual values. The motivation for a threesome was primarily curiosity and the third person invited to join the couple in a threesome was most often a friend/acquaintance of the woman. The outcome of a threesome for the couple was primarily no effect with a about a fifth reporting negative outcomes and a similar percentage reporting positive outcomes. Finally, the event happened only once for 83% of the respondents who had a threesome. Symbolic interaction, operant learning, social learning, affective theory and feminist theory were used to interpret the data. Implications and limitations are identified.

Mass spectrometry-based phenotypic H-antigen typing (MS-H) combined with whole-genome-sequencing-based genetic identification of H antigens, O antigens, and toxins (WGS-HOT) was used to type 60 clinical Escherichia coli isolates, 43 of which were previously identified as nonmotile, H type undetermined, or O rough by serotyping or having shown discordant MS-H and serotyping results. Whole-genome sequencing confirmed that MS-H was able to provide more accurate data regarding H antigen expression than serotyping. Further, enhanced and more confident O antigen identification resulted from gene cluster based typing in combination with conventional typing based on the gene pair comprising wzx and wzy and that comprising wzm and wzt . The O antigen was identified in 94.6% of the isolates when the two genetic O typing approaches (gene pair and gene cluster) were used in conjunction, in comparison to 78.6% when the gene pair database was used alone. In addition, 98.2% of the isolates showed the existence of genes for various toxins and/or virulence factors, among which verotoxins (Shiga toxin 1 and/or Shiga toxin 2) were 100% concordant with conventional PCR based testing results. With more applications of mass spectrometry and whole-genome sequencing in clinical microbiology laboratories, this combined phenotypic and genetic typing platform (MS-H plus WGS-HOT) should be ideal for pathogenic E. coli typing.

BACKGROUND Escherichia coli H antigen typing with antisera, a useful method for flagella clinical identification and classification, is a time-consuming process because of the need to induce flagella growth and the occurrence of undetermined strains. We developed an alternative rapid and analytically sensitive mass spectrometry (MS) method, termed MS-based H antigen typing (MS-H), and applied it at the protein sequence level for H antigen typing. We also performed a comparison with traditional serotyping on reference strains and clinical isolates. METHODS On the basis of international guidelines, the analytical selectivity and sensitivity, imprecision, correlation, repeatability, and reproducibility of the MS-H platform was evaluated using reference strains. Comparison of MS-H typing and serotyping was performed using 302 clinical isolates from 5 Canadian provinces, and discrepant results between the 2 platforms were resolved through whole genome sequencing. RESULTS Repeated tests on reference strain EDL933 demonstrated a lower limit of the measuring interval at the subsingle colony (16.97 μg or 1.465 × 107 cells) level and close correlation (r2 > 0.99) between cell culture biomass and sequence coverage. The CV was CONCLUSIONS MS-H typing of E. coli is useful for fast and accurate flagella typing and could be very useful during E. coli outbreaks.

The Prion Laboratory Section of the Public Health Agency of Canada supports heath care professionals dealing with patients suspected to have Creutzfeldt-Jakob disease (CJD) by testing cerebrospinal fluid (CSF) for protein markers of CJD. To better serve Canadian diagnostic requirements, a quaking-induced conversion (QuIC)-based assay has been added to the test panel. The QuIC tests exploit the ability of disease-associated prion protein, found in the CSF of a majority of CJD patients, to convert a recombinant prion protein (rPrP) into detectable amounts of a misfolded, aggregated form of rPrP. The rPrP aggregates interact with a specific dye, causing a measurable change in the dye's fluorescence emission spectrum. Optimal test and analysis parameters were empirically determined. Taking both practical and performance considerations into account, an endpoint QuIC (EP-QuIC) configuration was chosen. EP-QuIC uses a thermo-mixer to perform the shaking necessary to produce the quaking-induced conversions. Fluorescence readings are obtained from a microwell fluorescence reader only at the beginning and the end of EP-QuIC reactions. Samples for which the relative fluorescence unit ratio between the initial and final readings represent a ≥4 increase in signal intensity in at least two of the three replicates are classified as positive. A retrospective analysis of 91 CSF samples that included 45 confirmed cases of CJD and 46 non-CJD cases was used to estimate the performance characteristics of the EP-QuIC assay. The diagnostic sensitivity and specificity of the EP-QuIC test of this set of samples were 98 and 91%, respectively.

Creutzfeldt–Jakob disease (CJD) is a rapidly progressive neurodegenerative disease that can arise spontaneously, genetically, or be acquired through iatrogenic exposure. Most patients die within a year of symptom onset. It is rare, affecting 1–2 per million per year, and the majority of cases are sporadic. Primary angiitis of the central nervous system (PACNS) is also rare, affecting 2.4 per million per year. We present a case of an unusually long clinical course of CJD, almost five years, which began with symptoms of apraxia. The patient had biopsy-proven PACNS 16 years prior to clinical presentation, and the site of biopsy was the left parietal lobe. Autopsy revealed multicentric prion plaques in the cerebellum, in the setting of normal genetic testing. The presence of plaques in the cerebellum, and prior neurosurgery, raises the possibility of iatrogenic exposure. We present the details of this case, including pathology from the original biopsy and final autopsy, as well as a review of relevant cases in the literature.

Nontuberculous mycobacteria (NTM) are environmental bacteria that may cause chronic lung disease. Environmental factors that favor NTM growth likely increase the risk of NTM exposure within specific environments. We aimed to identify water-quality constituents (Al, As, Cd, Ca, Cu, Fe, Pb, Mg, Mn, Mo, Ni, K, Se, Na, Zn, and pH) associated with NTM disease across Colorado watersheds. We conducted a geospatial, ecological study, associating data from patients with NTM disease treated at National Jewish Health and water-quality data from the Water Quality Portal. Water-quality constituents associated with disease risk were identified using generalized linear models with Poisson-distributed discrete responses. We observed a highly robust association between molybdenum (Mo) in the source water and disease risk. For every 1- unit increase in the log concentration of molybdenum in the source water, disease risk increased by 17.0%. We also observed a statistically significant association between calcium (Ca) in the source water and disease risk. The risk of NTM varied by watershed and was associated with watershed-specific water-quality constituents. These findings may inform mitigation strategies to decrease the overall risk of exposure.

Introduction Evidence is lacking on the profile of gynaecological conditions affecting women in the Solomon Islands, including the availability and quality of surgical management. Methods Prospective analysis of hospital records was undertaken on all patients who underwent gynaecological surgery at Gizo Hospital, Western Province during a 6-day program led by volunteer Australian surgeons. Patient data on pre-operative history, investigation results, performed surgical procedures and postoperative recovery were collected. Results Of the 23 patients who presented with gynaecological problems requiring surgery, 20 underwent at least one surgical procedure during the study period. The most common presenting symptoms were pain and abnormal uterine bleeding. Median body mass index was 27, and 70% of patients were overweight or obese. Two surgeries were cancelled due to dengue fever. The surgeries performed were 12 vaginal operations, 8 laparoscopies and 9 laparotomies. Of surgical specimens collected, 61% were sent for histopathology testing. The median duration of postoperative hospital admission was 2 days (interquartile range, 1 day). Conclusion The Solomon Islands presents a unique profile of challenges to surgical practice, including the impact of dengue infection on fitness for surgery, a mobile patient population dispersed across the islands, difficult access to pathology services, and increased length of stay. Despite this, most patients had surgical outcomes equivalent to those in a developed setting.

Abstract –Collaborative Project Based Learning (CPBL) is known for enhancing deep learning, professional skills development, student engagement and motivation, cultivating interdependence in learning, thinking, problem solving, and creating interest and excitement in learning. This article describes the impact of an engineering design course on first-year engineering students. Student performance was evaluated before and after taking the course using a pre-and-post design skill assessment test, peer feedback evaluations, and thematic analysis of each student’s self-reflection of lessons learned. Initial analysis of the data indicates that a CPBL environment has a significantly positive impact on the development of engineering students’ ability to describe the engineering design process and relate it directly to real-world problems. Students also realized the importance of communication, team work, investigation and project management skills.

Nontuberculous mycobacteria (NTM) are ubiquitous environmental microorganisms. Infection is thought to result primarily from exposure to soil and/or water sources. NTM disease prevalence varies greatly by geographic region, but the geospatial factors influencing this variation remain unclear.To identify sociodemographic and environmental ecological risk factors associated with NTM infection and disease in Colorado.We conducted an ecological study, combining data from patients with a diagnosis of NTM disease from National Jewish Health's electronic medical record database and ZIP code-level sociodemographic and environmental exposure data obtained from the U.S. Geological Survey, the U.S. Department of Agriculture, and the U.S. Census Bureau. We used spatial scan methods to identify high-risk clusters of NTM disease in Colorado. Ecological risk factors for disease were assessed using Bayesian generalized linear models assuming Poisson-distributed discrete responses (case counts by ZIP code) with the log link function.We identified two statistically significant high-risk clusters of disease. The primary cluster included ZIP codes in urban regions of Denver and Aurora, as well as regions south of Denver, on the east side of the Continental Divide. The secondary cluster was located on the west side of the Continental Divide in rural and mountainous regions. After adjustment for sociodemographic, drive time, and soil variables, we identified three watershed areas with relative risks of 12.2, 4.6, and 4.2 for slowly growing NTM infections compared with the mean disease risk for all watersheds in Colorado. This study population carries with it inherent limitations that may introduce bias. The lack of complete capture of NTM cases in Colorado may be related to factors such as disease severity, education and income levels, and insurance status.Our findings provide evidence that water derived from particular watersheds may be an important source of NTM exposure in Colorado. The watershed with the greatest risk of NTM disease contains the Dillon Reservoir. This reservoir is also the main water supply for major cities located in the two watersheds with the second and third highest disease risk in the state, suggesting an important possible source of infection.

Purpose The need for rapid and accurate H typing is evident during Escherichia coli outbreak situations. This study explores the transition of MS-H, a method originally developed for rapid H antigen typing of E. coli using LC-MS/MS of flagella digest of reference strains and some clinical strains, to E. coli isolates in clinical scenario through quantitative analysis and method validation. Experimental design Motile and nonmotile strains were examined in batches to simulate clinical sample scenario. Various LC-MS/MS batch run procedures and MS-H typing rules were compared and summarized through quantitative analysis of MS-H data output for a standard method development. Results Label-free quantitative data analysis of MS-H typing was proven very useful for examining the quality of MS-H result and the effects of some sample carryovers from motile E. coli isolates. Based on this, a refined procedure and protein identification rule specific for clinical MS-H typing was established and validated. Conclusions and clinical relevance With LC-MS/MS batch run procedure and database search parameter unique for E. coli MS-H typing, the standard procedure maintained high accuracy and specificity in clinical situations, and its potential to be used in a clinical setting was clearly established.

The expression of two or more discrete phenotypes amongst individuals within a species (morphs) provides multiple modes upon which selection can act semi-independently, and thus may be an important stage in speciation. In the present study, we compared two sympatric morph systems aiming to address hypotheses related to their evolutionary origin. Arctic charr in sympatry in Loch Tay, Scotland, exhibit one of two discrete, alternative body size phenotypes at maturity (large or small body size). Arctic charr in Loch Awe segregate into two temporally segregated spawning groups (breeding in either spring or autumn). Mitochondrial DNA restriction fragment length polymorphism analysis showed that the morph pairs in both lakes comprise separate gene pools, although segregation of the Loch Awe morphs is more subtle than that of Loch Tay. We conclude that the Loch Awe morphs diverged in situ (within the lake), whereas Loch Tay morphs most likely arose through multiple invasions by different ancestral groups that segregated before post-glacial invasion (i.e. in allopatry). Both morph pairs showed clear trophic segregation between planktonic and benthic resources (measured by stable isotope analysis) but this was significantly less distinct in Loch Tay than in Loch Awe. By contrast, both inter-morph morphological and life-history differences were more subtle in Loch Awe than in Loch Tay. The strong ecological but relatively weak morphological and life-history divergence of the in situ derived morphs compared to morphs with allopatric origins indicates a strong link between early ecological and subsequent genetic divergence of sympatric origin emerging species pairs. The emergence of parallel specialisms despite distinct genetic origins of these morph pairs suggests that the effect of available foraging opportunities may be at least as important as genetic origin in structuring sympatric divergence in post-glacial fishes with high levels of phenotypic plasticity. © 2012 The Linnean Society of London, Biological Journal of the Linnean Society, 2012, ••, ••–••.

N-methyl-d-aspartate (NMDA) receptors are glutamate-gated cation channels that mediate excitatory neurotransmission in the central nervous system. In addition to glutamate, NMDA receptors are also activated by coagonist binding of the gliotransmitter, d-serine. Neuronal NMDA receptors mediate activity-dependent blood flow regulation in the brain. Our objective was to determine whether NMDA receptors expressed by brain endothelial cells can induce vasodilation of isolated brain arteries. Adult mouse middle cerebral arteries (MCAs) were isolated, pressurized, and preconstricted with norepinephrine. N-methyl-d-aspartate receptor agonists, glutamate and NMDA, significantly dilated MCAs in a concentration-dependent manner in the presence of d-serine but not alone. Dilation was significantly inhibited by NMDA receptor antagonists, d-2-amino-5-phosphonopentanoate and 5,7-dichlorokynurenic acid, indicating a response dependent on NMDA receptor glutamate and d-serine binding sites, respectively. Vasodilation was inhibited by denuding the endothelium and by selective inhibition or genetic knockout of endothelial nitric oxide synthase (eNOS). We also found evidence for expression of the pan-NMDA receptor subunit, NR1, in mouse primary brain endothelial cells, and for the NMDA receptor subunit NR2C in cortical arteries in situ. Overall, we conclude that NMDA receptor coactivation by glutamate and d-serine increases lumen diameter in pressurized MCA in an endothelial and eNOS-dependent mechanism.

Three ecologically and morphologically distinct forms of Arctic charr (Salvelinus alpinus L.) have been identified in Loch Rannoch, Scotland, whose evolutionary status and origins are incompletely understood. A study was made of restriction fragment length polymorphism (RFLPs) detected variation in the D-loop, ND1 and cytochrome b regions of the mitochondrial genome, encompassing >3500 bp. Eight RFLP haplotypes were identified that clustered into three distinct clans based on restriction differences and into four clans based on sequence differences. Significant differences in RFLP frequencies were found among all morph groups. The pelagic morph was highly divergent from the two benthic forms, with the benthic forms having variants from only one genetic clan while the pelagic was dominated by a single variant from another clan. The relative divergence observed among benthic and pelagic forms is ~10 fold greater when nucleotide divergence among the haplotypes, as well as haplotype frequency differences, is taken into account. Sequence divergence between haplotypes in the two main clans is of a similar order to that between haplotypes in these clans and a charr from North America. In contrast, divergence among the two benthic morphs relates entirely to differences in haplotype frequencies. The study confirms the genetic distinctiveness of the pelagic and benthic forms as well as of the two benthic forms. It strongly supports previous evidence that the genetic divergence between the pelagic and benthic populations is allopatric in origin. Additionally, the results strongly suggest that the two benthic populations have undergone peripatric divergence through the sequential colonisation of the two basins by one lineage, followed by their spatial separation and reproductive isolation.

Knowledge of mammalian diversity is still surprisingly disparate, both regionally and taxonomically. Here, we present a comprehensive assessment of the conservation status and distribution of the world's mammals. Data, compiled by 1700+ experts, cover all 5487 species, including marine mammals. Global macroecological patterns are very different for land and marine species but suggest common mechanisms driving diversity and endemism across systems. Compared with land species, threat levels are higher among marine mammals, driven by different processes (accidental mortality and pollution, rather than habitat loss), and are spatially distinct (peaking in northern oceans, rather than in Southeast Asia). Marine mammals are also disproportionately poorly known. These data are made freely available to support further scientific developments and conservation action.

The management of irreparable rotator cuff tears associated with osteoarthritis of the glenohumeral joint has long been challenging. Reverse total shoulder arthroplasty (RSA) was designed to provide pain relief and improve shoulder function in patients with severe rotator cuff tear arthropathy. While this procedure has been known to reduce pain, improve strength and increase range of motion in shoulder elevation, scapular notching, rotation deficiency, early implant loosening and dislocation have attributed to complication rates as high as 62 %. Patient selection, surgical approach and post-operative management are factors vital to successful outcome of RSA, with implant design and component positioning having a significant influence on the ability of the shoulder muscles to elevate, axially rotate and stabilise the humerus. Clinical and biomechanical studies have revealed that component design and placement affects the location of the joint centre of rotation and therefore the force-generating capacity of the muscles and overall joint mobility and stability. Furthermore, surgical technique has also been shown to have an important influence on clinical outcome of RSA, as it can affect intra-operative joint exposure as well as post-operative muscle function. This review discusses the behaviour of the shoulder after RSA and the influence of implant design, component positioning and surgical technique on post-operative joint function and clinical outcome.

Matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS) has gained popularity in recent years for rapid bacterial identification, mostly at the genus or species level. In this study, a rapid method to identify the Escherichia coli flagellar antigen (H antigen) at the subspecies level was developed using a MALDI-TOF MS platform with high specificity and sensitivity. Flagella were trapped on a filter membrane, and on-filter trypsin digestion was performed. The tryptic digests of each flagellin then were collected and analyzed by MALDI-TOF MS through peptide mass fingerprinting. Sixty-one reference strains containing all 53 H types and 85 clinical strains were tested and compared to serotyping designations. Whole-genome sequencing was used to resolve conflicting results between the two methods. It was found that DHB (2,5-dihydroxybenzoic acid) worked better than CHCA (α-cyano-4-hydroxycinnamic acid) as the matrix for MALDI-TOF MS, with higher confidence during protein identification. After method optimization, reference strains representing all 53 E. coli H types were identified correctly by MALDI-TOF MS. A custom E. coli flagellar/H antigen database was crucial for clearly identifying the E. coli H antigens. Of 85 clinical isolates tested by MALDI-TOF MS-H, 75 identified MS-H types (88.2%) matched results obtained from traditional serotyping. Among 10 isolates where the results of MALDI-TOF MS-H and serotyping did not agree, 60% of H types characterized by whole-genome sequencing agreed with those identified by MALDI-TOF MS-H, compared to only 20% by serotyping. This MALDI-TOF MS-H platform can be used for rapid and cost-effective E. coli H antigen identification, especially during E. coli outbreaks.

Small populations may be expected to harbour less genetic variation than large populations, but the relation between census size (N), effective population size (N e), and genetic diversity is not well understood. We compared microsatellite variation in four small peripheral Atlantic salmon populations from the Iberian peninsula and three larger populations from Scotland to test whether genetic diversity was related to population size. We also examined the historical decline of one Iberian population over a 50-year period using archival scales in order to test whether a marked reduction in abundance was accompanied by a decrease in genetic diversity. Estimates of effective population size (N e) calculated by three temporal methods were consistently low in Iberian populations, ranging from 12 to 31 individuals per generation considering migration, and from 38 to 175 individuals per generation if they were regarded as closed populations. Corresponding N e/N ratios varied from 0.02 to 0.04 assuming migration (mean=0.03) and from 0.04 to 0.18 (mean=0.10) assuming closed populations. Population bottlenecks, inferred from the excess of heterozygosity in relation to allelic diversity, were detected in all four Iberian populations, particularly in those year classes derived from a smaller number of returning adults. However, despite their small size and declining status, Iberian populations continue to display relatively high levels of heterozygosity and allelic richness, similar to those found in larger Scottish populations. Furthermore, in the R. Ason no evidence was found for a historical loss of genetic diversity despite a marked decline in abundance during the last five decades. Thus, our results point to two familiar paradigms in salmonid conservation: (1)␣endangered populations can maintain relatively high levels of genetic variation despite their small size, and (2) marked population declines may not necessarily result in a significant loss of genetic diversity. Although there are several explanations for such results, microsatellite data and physical tagging suggest that high levels of dispersal and asymmetric gene flow have probably helped to maintain genetic diversity in these peripheral populations, and thus to avoid the negative consequences of inbreeding.

This letter indicates that infection with HIV type 1 can occur despite preexposure prophylaxis with combination antiretroviral therapy.

S-Adenosylmethionine (AdoMet) is the methyl donor of numerous methylation reactions. The current model is that an increased concentration of AdoMet stimulates DNA methyltransferase reactions, triggering hypermethylation and protecting the genome against global hypomethylation, a hallmark of cancer. Using an assay of active demethylation in HEK 293 cells, we show that AdoMet inhibits active demethylation and expression of an ectopically methylated CMV-GFP (green fluorescent protein) plasmid in a dose-dependent manner. The inhibition of GFP expression is specific to methylated GFP; AdoMet does not inhibit an identical but unmethylated CMV-GFP plasmid. S-Adenosylhomocysteine (AdoHcy), the product of methyltransferase reactions utilizing AdoMet does not inhibit demethylation or expression of CMV-GFP. In vitro, AdoMet but not AdoHcy inhibits methylated DNA-binding protein 2/DNA demethylase as well as endogenous demethylase activity extracted from HEK 293, suggesting that AdoMet directly inhibits demethylase activity, and that the methyl residue on AdoMet is required for its interaction with demethylase. Taken together, our data support an alternative mechanism of action for AdoMet as an inhibitor of intracellular demethylase activity, which results in hypermethylation of DNA.

Current understanding of the postglacial colonization of Nearctic and Palearctic species relies heavily on inferences drawn from the phylogeographic analysis of contemporary generic variants. Modern postglacial populations are supposed to be representative of their Pleistocene ancestors, and their current distribution is assumed to reflect the different colonization success and dispersal patterns of refugial lineages. Yet, testing of phylogeographic models against ancestral genomes from glacial refugia has rarely been possible. Here we compare ND1 mitochondrial DNA variation in late Pleistocene (16,000-40,000 years before present), historical and contemporary Atlantic salmon (Salmo salar) populations from northern Spain and other regions of western Europe. Our study demonstrates the presence of Atlantic salmon in the Iberian glacial refugium during the last 40,000 years and points to the Iberian Peninsula as the likely source of the most common haplotype within the Atlantic lineage in Europe. However, our findings also suggest that there may have been significant changes in the genetic structure of the Iberian refugial stock since the last ice age, and question whether modern populations in refugial areas are representative of ice age populations. A common haplotype that persisted in the Iberian Peninsula during the Pleistocene last glacial maximum is now extremely rare or absent from European rivers, highlighting the need for caution when making phylogeographic inferences about the origin and distribution of modern genetic types.

Forty-three reference strains involving the 24 most common serovars of Salmonella enterica were examined by using a mass spectrometry-based H antigen typing platform (MS-H). The results indicate that MS-H can be used as a sensitive, rapid, and straightforward approach for the typing of Salmonella flagella at the molecular level without antiserum and phase inversion.

Previous lines of evidence have shown that inhibition of DNA methyltransferase (MeTase) can arrest tumor cell growth; however, the mechanisms involved were not clear. In this manuscript we show that out of 16 known tumor suppressors and cell cycle regulators, the cyclin-dependent kinase inhibitor p21 is the only tumor suppressor induced in the human lung cancer cell line, A549, following inhibition of DNA MeTase by a novel DNA MeTase antagonist or antisense oligonucleotides. The rapid induction of p21 expression points to a mechanism that does not involve demethylation of p21 promoter. Consistent with this hypothesis, we show that part of the CpG island upstream of the endogenous p21 gene is unmethylated and that the expression of unmethylated p21 promoter luciferase reporter constructs is induced following inhibition of DNA MeTase. These results are consistent with the hypothesis that the level of DNA MeTase in a cell can control the expression of a nodal tumor suppressor by a mechanism that does not involve DNA methylation.

Vertebrates have developed multiple mechanisms to coordinate the replication of epigenetic and genetic information. Dnmt1 encodes the maintenance enzyme DNA-methyltransferase, which is responsible for propagating the DNA methylation pattern and the epigenetic information that it encodes during replication. Direct sequence analysis and bisulfite mapping of the 5' region of DNA-methyltransferase 1 (dnmt1) have indicated the presence of many sequence elements associated with previously characterized origins of DNA replication. This study tests the hypothesis that the dnmt1 region containing these elements is an origin of replication in human cells. First, we demonstrate that a vector containing this dnmt1 sequence is able to support autonomous replication when transfected into HeLa cells. Second, using a gel retardation assay, we show that it contains a site for binding of origin-rich sequences binding activity, a recently purified replication protein. Finally, using competitive polymerase chain reaction, we show that replication initiates in this region in vivo. Based on these lines of evidence, we propose that initiation sites for DNA replication are located between the first intron and exon 7 of the human dnmt1 locus.