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Autism spectrum disorder (ASD) is a developmental disorder with a prevalence of around 1% children worldwide and characterized by patient behaviour (communication, social interaction, and personal development). Data on the efficacy of diagnostic tests using copy number variations (CNVs) in candidate genes in ASD is currently around 10% but it is overrepresented by patients of Caucasian background. We report here that the diagnostic success of de novo candidate CNVs in Vietnamese ASD patients is around 6%. We recruited one hundred trios (both parents and a child) where the child was clinically diagnosed with ASD while the parents were not affected. We performed genetic screening to exclude RETT syndrome and Fragile X syndrome and performed genome-wide DNA microarray (aCGH) on all probands and their parents to analyse for de novo CNVs. We detected 1708 non-redundant CNVs in 100 patients and 118 (7%) of them were de novo. Using the filter for known CNVs from the Simons Foundation Autism Research Initiative (SFARI) database, we identified six CNVs (one gain and five loss CNVs) in six patients (3 males and 3 females). Notably, 3 of our patients had a deletion involving the SHANK3 gene-which is the highest compared to previous reports. This is the first report of candidate CNVs in ASD patients from Vietnam and provides the framework for building a CNV based test as the first tier screening for clinical management.

Nowadays, environmental issues and cleaner products are interest to many customers, considering whether to buy or continue using a product. This will affect the perception, attitude of upper managers in the process of strategic choices and operational management behavior. This study is based on the Upper Echelon Theory, research under the influence of customer pressure to attitude toward the environmental, decision choices for cleaner production strategies, implementation of environmental management accounting towards achieving green competitive advantage of Vietnamese manufacturing enterprises. In addition, the role of two moderator variables: (1) perceived benefit of cleaner production strategies and (2) perceived benefit of environmental management accounting in the research model is also considered. This study surveyed 234 CEOs of Vietnamese manufacturing enterprises. This study employed PLS-SEM, version 3 for data analysis. Results have shown that all relationships are statistically significant. Moderator variables have a statistically significant and positive impact in relationships in which they play a moderator role. This study helps CEOs realize the importance of producing products that are customer-oriented, environmentally friendly, and the implementation of environmental management accounting will have a strong impact on achieving a sustainable competitive advantage.

Organ-on-a-chip platforms are utilized in global bioanalytical and toxicological studies as a way to reduce materials and increase throughput as compared to in vivo based experiments. These platforms bridge the infrastructure and regulatory gaps between in vivo animal work and human systems, with models that exemplify active biological pathways. In conjunction with the advent of increased capabilities associated with next generation sequencing and mass spectrometry based ‘-omic’ technologies, organ-on-a-chip platforms provide an excellent opportunity to investigate the global changes at multiple biological levels, including the transcriptome, proteome and metabolome. When investigated concurrently, a complete profile of cellular and regulatory perturbations can be characterized following treatment with specific agonists. In this study, global effects were observed and analyzed following liver chip exposure to the chemical warfare agent, VX. Even though the primary mechanism of action of VX (i.e. acetylcholinesterase inhibition) is well characterized, recent in vivo studies suggest additional protein binding partners that are implicated in metabolism and cellular energetic pathways. In addition, secondary toxicity associated with peripheral organ systems, especially in human tissues, is not well defined. Our results demonstrate the potential of utilizing an organ-on-a-chip platform as a surrogate system to traditional in vivo studies. This is realized by specifically indicating significant dysregulation of several cellular processes in response to VX exposure including but not limited to amino acid synthesis, drug metabolism, and energetics pathways.

Background Short sleep duration (SSD) (Methods We used national Behavioral Risk Factor Surveillance System surveys to conduct a cross-sectional study examining SSD estimates for US women with prediabetes by rural/urban residence between 2016–2020. We applied logistic regression models to the BRFSS dataset to ascertain associations between rural/urban residence status and SSD prior to and following adjustment for sociodemographic factors (age, race, education, income, health care coverage, having a personal doctor). Results Our study included 20,997 women with prediabetes (33.7% rural). SSD prevalence was similar between rural (35.5%, 95% CI: 33.0%-38.0%) and urban women (35.4%, 95% CI: 33.7%-37.1). Rural residence was not associated with SSD among US women with prediabetes prior to adjustment (Odds Ratio: 1.00, 95% CI: 0.87–1.14) or following adjustment for sociodemographic factors (Adjusted Odds Ratio: 1.06, 95% CI: 0.92–1.22). Among women with prediabetes, irrespective of rural/urban residence status, being Black, aged Conclusions Despite the finding that SSD estimates among women with prediabetes did not vary by rural/urban residence status, 35% of rural women with prediabetes had SSD. Efforts to reduce diabetes burden in rural areas may benefit from incorporating strategies to improve sleep duration along with other known diabetes risk factors among rural women with prediabetes from certain sociodemographic backgrounds.

Hyperbilirubinemia is more frequently seen in low and middle-income countries like Indonesia. One of the contributing factors is a substandard dose of Phototherapy irradiance. This research aims to design a phototherapy intensity meter called PhotoInMeter using readily available low-cost components. PhotoInMeter is designed by using a microcontroller, light sensor, color sensor, and an ND (neutral-density) filter. We use machine learning to create a mathematical model that converts the emission from the color sensor and light sensor into light intensity measurements that are close to Ohmeda Biliblanket's measurements. Our prototype collects sensor reading data and pairs them with Ohmeda Biliblanket Light Meter to create a training set for our machine learning algorithm. We create a multivariate linear regression, random forest, and XGBoost model based on our training set to convert sensor readings to Ohmeda Biliblanket Light Meter measurement. We successfully devised a prototype that costs 20 times less to produce compared to our reference intensity meter while still having high accuracy. Compared to Ohmeda Biliblanket Light Meter, our PhotoInMeter has a Mean Absolute Error (MAE) of 0.83 and achieves more than a 0.99 correlation score in all six different devices for intensity in the range of 0-90 μW/cm2/nm. Our prototypes show consistent reading between PhotoInMeter devices, having an average difference of 0.435 among all six devices.

BackgroundThis study sought to examine the relationship between rural residence and physical activity levels among US myocardial infarction (MI) survivors.MethodsWe conducted a cross-sectional study using nationally representative Behavioral Risk Factor Surveillance System surveys from 2017 and 2019. We determined the survey-weighted percentage of rural and urban MI survivors meeting US physical activity guidelines. Logistic regression models were used to examine the relationship between rural/urban residence and meeting physical activity guidelines, accounting for sociodemographic factors.ResultsOur study included 22,732 MI survivors (37.3% rural residents). The percentage of rural MI survivors meeting physical activity guidelines (37.4%, 95% CI: 35.1%-39.7%) was significantly less than their urban counterparts (45.6%, 95% CI: 44.0%-47.2%). Rural residence was associated with a 28.8% (95% CI: 20.0%-36.7%) lower odds of meeting physical activity guidelines, with this changing to a 19.3% (95% CI: 9.3%-28.3%) lower odds after adjustment for sociodemographic factors.ConclusionsA significant rural/urban disparity in physical activity levels exists among US MI survivors. Our findings support the need for further efforts to improve physical activity levels among rural MI survivors as part of successful secondary prevention in US high-MI burden rural areas.

The diagnoses of retroviruses are essential for controlling the rapid spread of pandemics. However, the real-time Reverse Transcriptase quantitative Polymerase Chain Reaction (RT-qPCR), which has been the gold standard for identifying viruses such as SARS-CoV-2 in the early stages of infection, is associated with high costs and logistical challenges. To innovate in viral RNA detection a novel molecular approach for detecting SARS-CoV-2 viral RNA, as a proof of concept, was developed. This method combines specific viral gene analysis, trans-acting ribozymes, and Fluorescence Resonance Energy Transfer (FRET)-based hybridization of fluorescent DNA hairpins. In this molecular mechanism, SARS-CoV-2 RNA is specifically recognized and cleaved by ribozymes, releasing an initiator fragment that triggers a hybridization chain reaction (HCR) with DNA hairpins containing fluorophores, leading to a FRET process. A consensus SARS-CoV-2 RNA target sequence was identified, and specific ribozymes were designed and transcribed in vitro to cleave the viral RNA into fragments. DNA hairpins labeled with Cy3/Cy5 fluorophores were then designed and synthesized for HCR-FRET assays targeting the RNA fragment sequences resulting from ribozyme cleavage. The results demonstrated that two of the three designed ribozymes effectively cleaved the target RNA within 10 minutes. Additionally, DNA hairpins labeled with Cy3/Cy5 pairs efficiently detected target RNA specifically and triggered detectable HCR-FRET reactions. This method is versatile and can be adapted for use with other viruses. Furthermore, the design and construction of a DIY photo-fluorometer prototype enabled us to explore the development of a simple and cost-effective point-of-care detection method based on digital image analysis. [ABSTRACT FROM AUTHOR]

The concerted action of DNA replication and cell division has been extensively investigated in eukaryotes. Well demarcated checkpoints have been identified in the cell cycle, which provides the correct DNA stoichiometry and appropriate growth in the progeny. In bacteria, which grow faster and less concerted than eukaryotes, the linkages between cell elongation and DNA synthesis are unclear. dTTP, one of the canonical nucleotide-building blocks of DNA, is also used for cell wall biosynthesis in mycobacteria. We hypothesize that the interconnection between DNA and cell wall biosynthesis through dTTP may require synchronization of these processes by regulating dTTP availability. We investigated growth, morphology, cellular dNTP pool, and possible signs of stress in Mycobacterium smegmatis upon perturbation of rhamnose biosynthesis by the overexpression of RmlA. RmlA is a cell wall synthetic enzyme that uses dTTP as the precursor for cross-linking the peptidoglycan with the arabinogalactan layers by a phosphodiester bond in the mycobacterial cell wall. We found that RmlA overexpression results in changes in cell morphology, causing cell elongation and disruption of the cylindrical cell shape. We also found that the cellular dTTP pool is reduced by half in RmlA overexpressing cells and that this reduced dTTP availability does not restrict cell growth. We observed 2-6-fold increases in the gene expression of replication and cell wall biosynthesis stress factors upon RmlA overexpression. Using super-resolution microscopy, we found that RmlA, acting to crosslink the nascent layers of the cell wall, localizes throughout the whole cell length in a helical pattern in addition to the cellular pole.

Nowadays, environmental issues and cleaner products are interest to many customers, considering whether to buy or continue using a product. This will affect the perception, attitude of upper managers in the process of strategic choices and operational management behavior. This study is based on the Upper Echelon Theory, research under the influence of customer pressure to attitude toward the environmental, decision choices for cleaner production strategies, implementation of environmental management accounting towards achieving green competitive advantage of Vietnamese manufacturing enterprises. In addition, the role of two moderator variables: (1) perceived benefit of cleaner production strategies and (2) perceived benefit of environmental management accounting in the research model is also considered. This study surveyed 234 CEOs of Vietnamese manufacturing enterprises. This study employed PLS-SEM, version 3 for data analysis. Results have shown that all relationships are statistically significant. Moderator variables have a statistically significant and positive impact in relationships in which they play a moderator role. This study helps CEOs realize the importance of producing products that are customer-oriented, environmentally friendly, and the implementation of environmental management accounting will have a strong impact on achieving a sustainable competitive advantage. [ABSTRACT FROM AUTHOR]

ObjectiveEffects of stroke (i.e., memory loss, paralysis) may make effective diabetes care difficult which can in turn contribute to additional diabetes related complications and hospitalization. However, little is known about US post-stroke diabetes care levels. This study sought to examine diabetes care levels among US adults with diabetes by stroke status.MethodsUsing 2015-2018 Behavioral Risk Factor Surveillance System surveys, the prevalence of nonadherence with the American Diabetes Association's diabetes care measures (ResultsOur study included 72,630 individuals, with 9.8% having had a stroke. Nonadherence levels varied for each diabetes care measure ranging from 20.4-42.2% for stroke survivors and 22.8-44.0% for those who had never had stroke. By stroke status, nonadherence with diabetes management measures was comparable except for stroke survivors having both a lower prevalence (30.2% versus 40.1%) and odds of nonadherence (OR: 0.73, 95% CI: 0.65, 0.82) with daily blood glucose check than those who had never had stroke.ConclusionWhile nonadherence with diabetes management does not vary by stroke status, considerable nonadherence still exists among stroke survivors with diabetes. Additional interventions to improve diabetes care may help to reduce risk of further diabetes complications in this population.

ObjectivesTo conduct a cross-sectional nationwide study examining how exclusion of nursing home COVID-19 cases influences the association between county level social distancing behavior and COVID-19 cases throughout the US during the early phase of the pandemic (February 2020-May 2020).MethodsUsing county-level COVID-19 data and social distancing metrics from tracked mobile devices, we investigated the impact social distancing had on a county's total COVID-19 cases (cases/100,000 people) between when the first COVID-19 case was confirmed in a county and May 31st, 2020 when most statewide social distancing measures were lifted, representing the pandemic's exponential growth phase. We created a mixed-effects negative binomial model to assess how implementation of social distancing measures when they were most stringent (March 2020-May 2020) influenced total COVID-19 cases while controlling for social distancing and COVID-19 related covariates in two scenarios: (1) when COVID-19 nursing home cases are not excluded from total COVID-19 cases and (2) when these cases are excluded. Model findings were compared to those from February 2020, a baseline when social distancing measures were not in place. Marginal effects at the means were generated to further isolate the influence of social distancing on COVID-19 from other factors and determine total COVID-19 cases during March 2020-May 2020 for the two scenarios.ResultsRegardless of whether nursing home COVID-19 cases were excluded from total COVID-19 cases, a 1% increase in average % of mobile devices leaving home was significantly associated with a 5% increase in a county's total COVID-19 cases between March 2020-May 2020 and about a 2.5% decrease in February 2020. When the influence of social distancing was separated from other factors, the estimated total COVID-19 cases/100,000 people was comparable throughout the range of social distancing values (25%-45% of mobile phone devices leaving home between March 2020-May 2020) when nursing home COVID-19 cases were not excluded (25% of mobile phones leaving home: 163.84 cases/100,000 people (95% CI: 121.81, 205.86), 45% of mobile phones leaving home: 432.79 cases/100,000 people (95% CI: 256.91, 608.66)) and when they were excluded (25% of mobile phones leaving home: 149.58 cases/100,000 people (95% CI: 111.90, 187.26), 45% of mobile phones leaving home: 405.38 cases/100,000 people (95% CI: 243.14, 567.62)).ConclusionsExclusion of nursing home COVID-19 cases from total COVID-19 case counts has little impact when estimating the relationship between county-level social distancing and preventing COVID-19 cases with additional research needed to see whether this finding is also observed for COVID-19 growth rates and mortality.

BackgroundHigher availability of firearms has been connected to higher rates of interpersonal violence in previous studies. Yet, those studies have focused mainly on the United States, or used aggregated international data to study firearm violence. Whether those aggregated findings are applicable to understanding the phenomenon in continental Europe specifically remains unclear. The aim of this systematic review is to bring together all studies that exclusively use European data.MethodsNine databases were searched, resulting in more than 1900 individual studies. These studies were assessed on relevance and eligibility for this study, based on their title, abstract and full text. Information on study characteristics, operationalizations of main concepts and study results were extracted from the six eligible studies.ResultsFour studies assessed the impact of firearm restrictive regulations on the rate of firearm homicides. Two other studies correlated rates of firearm availability and -violence. Results vary: some studies show a clear decline once availability of firearms is restricted, while others indicate a limited effect on only a very specific subgroup, such as female victims, or national guards with weapons at home. Moreover, studies used various operationalizations for firearm availability, thereby decreasing the comparability of findings.ConclusionEmpirical research exclusively using European data is still lacking. To increase comparability of future studies, methodological inconsistencies and regional gaps need to be overcome. Assessing how firearm availability can be measured with reliable and valid proxies across countries will be a crucial first step to improve future research on the link between firearms and firearm violence.

We investigate a no-boarding policy in a system of N buses serving M bus stops in a loop, which is an entrainment mechanism to keep buses synchronised in a reasonably staggered configuration. Buses always allow alighting, but would disallow boarding if certain criteria are met. For an analytically tractable theory, buses move with the same natural speed (applicable to programmable self-driving buses), where the average waiting time experienced by passengers waiting at the bus stop for a bus to arrive can be calculated. The analytical results show that a no-boarding policy can dramatically reduce the average waiting time, as compared to the usual situation without the no-boarding policy. Subsequently, we carry out simulations to verify these theoretical analyses, also extending the simulations to typical human-driven buses with different natural speeds based on real data. Finally, a simple general adaptive algorithm is implemented to dynamically determine when to implement no-boarding in a simulation for a real university shuttle bus service.

ObjectivesTo evaluate the clinical and radiographic outcome of low-demand patients with massive rotator cuff tears undergoing arthroscopic debridement in mid- and long-term follow-up, as well as the rate of conversion to reverse shoulder arthroplasty.MethodsWe performed a retrospective analysis of 19 patients with a mean age at surgery of 68 years (range, 55-80 years) from a previously described consecutive cohort and after a mean follow up of 47 month (FU1) and 145 month (FU2). The functional outcome was evaluated with the VAS score, the American Shoulder and Elbow Surgeons (ASES) score, and the age- and gender-adjusted Constant (aCS) score. The radiographic outcome was classified according to the Hamada classification. Non-parametric analyses were carried out with the Mann-Whitney U for independent samples and the Wilcoxon signed-rank test for related samples.ResultsFive patients (26%) developed symptomatic cuff tear arthropathy and underwent reverse shoulder arthroplasty after a mean time of 63 months (range, 45-97 months). These patients were excluded from further analyses. The mean VAS score of the remaining 14 patients at FU1 was significantly lower compared to preoperatively (P = .041), while there were no significant differences between the VAS score at FU1 and FU2 (P = 1.0). The ASES score of the affected shoulder at FU1 was significantly higher compared to prior to surgery (P = .028), while there were no significant differences between the scores of the affected shoulder between FU1 and FU2 (P = .878). While the ASES score of the contralateral shoulder at FU1 was significantly higher than the score of the affected shoulder (P = .038), there were no significant differences in the ASES scores of the affected and the healthy shoulder at FU2 (P = .575). The evaluation of the aCS produced similar results. A progression of the Hamada grade was documented in 6 patients.ConclusionsArthroscopic debridement is a safe and valid option for low-demand middle-age or elderly patients with symptomatic massive rotator cuff tears, leading to a significant pain relief and significantly improved functional outcome at mid- and long-term follow up. However, about a quarter of the patients in our cohort had to undergo reverse shoulder arthroplasty due to symptomatic cuff tear arthropathy. Furthermore, some of the remaining patients continued to undergo radiographic progression. This might be due to the natural history of their disease and/or the surgical procedure, and the clinical relevance of this finding should be evaluated in further studies.

The brown planthopper (BPH), Nilaparvata lugens, is a serious threat to rice production in Vietnam and insecticides are widely used for its control. Migration of the BPH have one of its roots in tropical Vietnam in the Mekong River Delta and the insecticide resistance status of BPH populations from Vietnam is thus important for East Asia. In the present investigation, we evaluate the susceptibility of BPH populations from nine provinces from the Red River Delta, the Central Coastal region and the Mekong River Delta of eight insecticides during 2015-17. BPH field populations of Vietnam have developed a low to moderate level of resistance to the neonicotinoids dinotefuran, nitenpyram and imidacloprid, the pyrethroid etofenprox, the anticholinesterase fenobucarb, as well as fipronil and pymetrozine, and the growth regulator buprofezin. There was a correlation of in toxicology of fipronil, dinotefuran, etofenprox, buprofezin, which represents four different modes of action. The neonicotinoid nitenpyram, pymetrozine and fenobucarb did not show correlation in toxicology to any of the investigated insecticides. For most insecticides, a gradient of susceptibility was established from the Red River Delta in the north, through the Central Coastal region and to the Mekong River Delta in the south of Vietnam. The most susceptible populations were from the north. Insecticide resistance of the BPH populations in Vietnam is not at an alarming level and they are not the direct origin of high insecticide resistance found in East Asia. The cross-resistance pattern of BPH populations in Vietnam, where insecticides with different modes of action correlated, indicate that insecticides should be used with caution. There could be a buildup of a general metabolic resistance, which alone or in combination with the emergence of target-site resistance mutations will cause control problems. The results will be beneficial for development of resistance management strategies to prevent and delay development of insecticide resistance in BPH not only for Vietnam, but also for more northern Asian regions due the migration of BPH from tropical Vietnam.

BackgroundWhile overall rates of meningococcal disease have been declining in the United States for the past several decades, New York City (NYC) has experienced two serogroup C meningococcal disease outbreaks in 2005-2006 and in 2010-2013. The outbreaks were centered within drug use and sexual networks, were difficult to control, and required vaccine campaigns.MethodsWhole Genome Sequencing (WGS) was used to analyze preserved meningococcal isolates collected before and during the two outbreaks. We integrated and analyzed epidemiologic, geographic, and genomic data to better understand transmission networks among patients. Betweenness centrality was used as a metric to understand the most important geographic nodes in the transmission networks. Comparative genomics was used to identify genes associated with the outbreaks.ResultsNeisseria meningitidis serogroup C (ST11/ET-37) was responsible for both outbreaks with each outbreak having distinct phylogenetic clusters. WGS did identify some misclassifications of isolates that were more distant from the outbreak strains, as well as those that should have been included based on high genomic similarity. Genomes for the second outbreak were more similar than the first and no polymorphism was found to either be unique or specific to either outbreak lineage. Betweenness centrality as applied to transmission networks based on phylogenetic analysis demonstrated that the outbreaks were transmitted within focal communities in NYC with few transmission events to other locations.ConclusionsNeisseria meningitidis is an ever changing pathogen and comparative genomic analyses can help elucidate how it spreads geographically to facilitate targeted interventions to interrupt transmission.

OBJECTIVES:To investigate the prevalence of shoulder-related acute and overuse injuries in triathletes and examine the role of possible risk factors, in order to identify potential preventive measures. METHODS:We performed a retrospective epidemiologic study of 193 amateur triathletes between June and August 2013 and evaluated their competition and training habits, as well as the presence of acute and overuse injuries of the shoulder sustained during the past 12 months. Contingency tables were analyzed using Pearson's chi-squared test. Normally distributed data were compared with the independent samples t-test, while non-parametric analyses were performed with the Mann-Whitney U test. Binary logistic regression was used to identify important predictors of injuries. RESULTS:12 participants (6%) sustained acute injuries and 36 athletes experienced an overuse injury. The acute injury rate amounted to 0.11 per 1000 hours of training and the overuse injury rate to 0.33 per 1000 hours of training. There was no association between athletes' age, height, weight, BMI, a history of shoulder complaints or triathlon experience in years and acute or overuse injuries. Male athletes had a trend for sustaining more acute injuries then female athletes (8% vs. 2%, p = 0.079). Athletes with acute injuries spent a significantly higher amount of time per week doing weight training (p = 0.007) and had a trend for a higher weekly duration of cycling training (p = 0.088). Athletes with overuse injuries participated in a significantly higher number of races compared to athletes without overuse injuries (p = 0.005). The regular use of paddles was associated with a significantly higher rate of overuse injuries (24% vs. 10%, p = 0.014). CONCLUSION:The regular use of paddles during swimming training appears to be a risk factor for the development of overuse injuries, while an increased duration of weight and cycling training seems to be associated with a higher rate of acute injuries.

Stunting remains a major public health concern worldwide. Although its global prevalence is slowly decreasing, the actual number of affected children is still rising in Sub-Saharan Africa. In the Central African Republic (CAR), about one third of all children below the age of five are stunted. Stunting is correlated with many long-term consequences, including poor cognitive development and a higher rate of morbidity and mortality, making stunting a major contributor to poverty. In CAR, little is known about the factors that contribute to stunting. This study aimed at analysing, in a cross-sectional study, the main factors associated with stunting in a group of 414 children recruited between December 2011 and November 2013, aged five years or less and living in Bangui. For all children, demographic, socio-economic and anthropometric data were recorded and asymptomatic enteropathogen carriage was assessed in stool samples using classical microbiological assays. The study group had a mean age of 14.2±10 months. Fifty-eight percent (292/414) were boys, and 36 percent (148/414) exhibited stunted growth. Of the stunted children, 51% (75/148) showed a moderate delay in linear growth for their age group [height-for-age z-score (HAZ) between -2 and -3 SD] while 49% (73/148) presented a severe delay (HAZ < -3). Factors significantly associated with stunting included gender (aOR: 1.67; 95% CI: 1.07; 2.62 for boys compared to girls) and age (aOR of 3.98 (95% CI: 2.45; 6.46) for toddlers and aOR 4.42 (95% CI: 2.36; 8.28) for children compared to infants). Most importantly, we identified being overweight [weight-for-height z-score (WHZ) > 2 SD; aOR: 3.21; 95% CI: 1.50; 6.90 of overweight compared to normal weight] as also being significantly associated with stunting. This is the first study showing that even in the poorest countries of the world there is an association of stunting with being overweight.

OBJECTIVE:To describe the cause-specific morbidity and mortality, and referral patterns of all neonates admitted to a tertiary referral hospital in the northern provinces of Vietnam. DESIGN:A prospective hospital based observational study. SETTING:The Neonatal Department, National Hospital of Pediatrics, Hanoi, Vietnam. PATIENTS:All admissions to the Neonatal Department over a 12 month period. MAIN OUTCOME MEASURES:Cause-specific morbidity and mortality; deaths. RESULTS:There were 5064 admissions with the commonest discharge diagnoses being infection (32%) and prematurity (29%). The case fatality ratio (CFR) was 13.9% (n = 703). Infection (38%), cardio/respiratory disorders (27%), congenital abnormalities (20%) and neurological conditions (10%) were the main causes of death. Of all the deaths, 38% had an admission weight ≥2500g. Higher CFR were associated with lower admission weights. Very few deaths (3%) occurred in the first 24 hours of life. Most referrals and deaths came from Hanoi and neighbouring provincial hospitals, with few from the most distant provinces. Two distant referral provinces had the highest CFR. CONCLUSIONS:The CFR was high and few deaths occurred in neonates

Charcot-Marie-Tooth (CMT) disease or hereditary motor and sensory neuropathy is the most prevalent inherited peripheral neuropathy and is associated with over 90 causative genes. Mutations in neurofilament light polypeptide gene, NEFL cause CMT2E, an axonal form of CMT that results in abnormal structures and/or functions of peripheral axons in spinal cord motor neurons and dorsal root ganglion neurons. We have previously generated and characterized a knock-in mouse model of CMT2E with the N98S mutation in Nefl that presented with multiple inclusions in spinal cord neurons. In this report, we conduct immunofluorescence studies of cultured dorsal root ganglia (DRG) from NeflN98S/+ mice, and show that inclusions found in DRG neurites can occur in embryonic stages. Ultrastructural analyses reveal that the inclusions are disordered neurofilaments packed in high density, segregated from other organelles. Immunochemical studies show decreased NFL protein levels in DRG, cerebellum and spinal cord in NeflN98S/+ mice, and total NFL protein pool is shifted toward the triton-insoluble fraction. Our findings reveal the nature of the inclusions in NeflN98S/+ mice, provide useful information to understand mechanisms of CMT2E disease, and identify DRG from NeflN98S/+ mice as a useful cell line model for therapeutic discoveries.

It remains unknown whether the administration of a deep neuromuscular block (NMB) during bariatric surgery improves surgical conditions and patient outcome. The authors studied the effect of deep versus moderate NMB in laparoscopic bariatric surgery on surgical conditions and postoperative pain.One hundred patients scheduled to undergo elective bariatric surgery were randomized to a deep NMB (post-tetanic-count 2-3) or a moderate NMB (train-of-four 1-2). The quality of the surgical field was scored using the Leiden-Surgical Rating Scale (L-SRS), a 5-point scale ranging from 1 (extremely poor conditions) to 5 (optimal conditions). Three surgeons scored the L-SRS at 10-min intervals during surgery; postoperative pain scores were obtained in the postanesthesia-care-unit (PACU) and on the ward. Mean (95% confidence interval) L-SRS scores in moderate NMB 4.2 (4.0-4.4) versus 4.8 (4.7-4.9) in deep NMB (p < 0.001). Moderate NMB resulted in 17% of scores at L-SRS scores of 1-3, while deep NMB resulted in 100% scores at the high end of the L-SRS (4-5). Deep NMB led to improved pain scores in the PACU (4.6 (4.2-4.9) versus 3.9 (3.6-4.4), p = 0.03) and reduced shoulder pain on the ward (1.8 (1.5-2.1) versus 1.3 (1.1-1.5), p = 0.03). A composite score of pain and opioid use in the PACU favoured deep NMB (p = 0.001).In bariatric surgery, deep relaxation has advantages for surgeon and patient. Compared to moderate NMB, deep NMB produced stable and improved surgical conditions with less postoperative pain.

Mutational processes and signatures that drive early tumorigenesis are centrally important for early cancer prevention. Yet, to date, biomarkers and risk factors for polyps (adenomas) that inordinately and rapidly develop into colon cancer remain poorly defined. Here, we describe surprisingly high mutational profiles through whole-genome sequence (WGS) analysis in 2 of 4 pairs of benign colorectal adenoma tissue samples. Unsupervised hierarchical clustered transcriptomic analysis of a further 7 pairs of adenomas reveals distinct mutational signatures regardless of adenoma size. Transitional single nucleotide substitutions of C:G>T:A predominate in the adenoma mutational spectrum. Strikingly, we observe mutations in the TGF-β pathway and CEA-associated genes in 4 out of 11 adenomas, overlapping with the Wnt pathway. Immunohistochemical labeling reveals a nearly 5-fold increase in CEA levels in 23% of adenoma samples with a concomitant loss of TGF-β signaling. We also define a functional role by which the CEA B3 domain interacts with TGFBR1, potentially inactivating the tumor suppressor function of TGF-β signaling. Our study uncovers diverse mutational processes underlying the transition from early adenoma to cancer. This has broad implications for biomarker-driven targeting of CEA/TGF-β in high-risk adenomas and may lead to early detection of aggressive adenoma to CRC progression.

BACKGROUND:Inter-observer delineation variation has been detailed for many years in almost every tumor location. Inadequate delineation can impair the chance of cure and/or increase toxicity. The aim of our original work was to prospectively improve the homogeneity of delineation among all of the senior radiation oncologists in the Nord-Pas de Calais region, irrespective of the conditions of practice. METHODS:All 11 centers were involved. The first studied cancer was prostate cancer. Three clinical cases were studied: a low-risk prostate cancer case (case 1), a high-risk prostate cancer case (pelvic nodes, case 2) and a case of post-operative biochemical elevated PSA (case 3). All of the involved physicians delineated characteristically the clinical target volume (CTV) and organs at risk. The volumes were compared using validated indexes: the volume ratio (VR), common and additional volumes (CV and AV), volume overlap (VO) and Dice similarity coefficient (DSC). A second delineation of the same three cases was performed after discussion of the slice results and the choice of shared guidelines to evaluate homogenization. A comparative analysis of the indexes before and after discussion was conducted using the Wilcoxon test for paired samples. A p-value less than 0.05 was considered to indicate statistical significance. RESULTS:The indexes were not improved in case 1, for which the inter-observer agreement was considered good after the first comparison (DSC = 0.83 ± 0.06). In case 2, the second comparison showed homogenization of the CTV delineation with a significant improvement in CV (81.4 ± 11.7 vs. 88.6 ± 10.26, respectively, p = 0.048), VO (0.41 ± 0.09 vs. 0.47 ± 0.07, respectively; p = 0.009) and DSC (0.58 ± 0.09 vs. 0.63 ± 0.07, respectively; p = 0.0098). In case 3, VR and AV were significantly improved: VR: 1.71(± 0.6) vs. 1.34(± 0.46), respectively, p = 0.0034; AV: 46.58(± 14.50) vs. 38.08(± 15.10), respectively, p = 0.0024. DSC was not improved, but it was already superior to 0.6 in the first comparison. CONCLUSION:Our prospective work showed that a collaborative discussion about clinical cases and the choice of shared guidelines within an established framework improved the homogeneity of CTV delineation among the senior radiation oncologists in our region.

We have reported that high-hydrostatic-pressure (HHP) technology is safe and useful for producing various kinds of decellularized tissue. However, the preparation of decellularized or inactivated skin using HHP has not been reported. The objective of this study was thus to prepare inactivated skin from human skin using HHP, and to explore the appropriate conditions of pressurization to inactivate skin that can be used for skin reconstruction. Human skin samples of 8 mm in diameter were packed in bags filled with normal saline solution (NSS) or distilled water (DW), and then pressurized at 0, 100, 150, 200 and 1000 MPa for 10 minutes. The viability of skin after HHP was evaluated using WST-8 assay. Outgrowth cells from pressurized skin and the viability of pressurized skin after cultivation for 14 days were also evaluated. The pressurized skin was subjected to histological evaluation using hematoxylin and eosin staining, scanning electron microscopy (SEM), immunohistochemical staining of type IV collagen for the basement membrane of epidermis and capillaries, and immunohistochemical staining of von Willebrand factor (vWF) for capillaries. Then, human cultured epidermis (CE) was applied on the pressurized skin and implanted into the subcutis of nude mice; specimens were subsequently obtained 14 days after implantation. Skin samples pressurized at more than 200 MPa were inactivated in both NSS and DW. The basement membrane and capillaries remained intact in all groups according to histological and immunohistological evaluations, and collagen fibers showed no apparent damage by SEM. CE took on skin pressurized at 150 and 200 MPa after implantation, whereas it did not take on skin pressurized at 1000 MPa. These results indicate that human skin could be inactivated after pressurization at more than 200 MPa, but skin pressurized at 1000 MPa had some damage to the dermis that prevented the taking of CE. Therefore, pressurization at 200 MPa is optimal for preparing inactivated skin that can be used for skin reconstruction.

In this prospective cross-sectional study on young premenopausal breast cancer patients, the objectives were to: determine the incidences of chemotherapy-related amenorrhea (CRA) and menopause (CRM); identify associated factors; and assess plasma levels of estradiol (E2) and follicular stimulating hormone (FSH) among patients who developed menopause.Eligibility criteria include Chinese stage I-III breast cancer patients, premenopausal, age ≤45 at breast cancer diagnosis, having received adjuvant chemotherapy, within 3-10 years after breast cancer diagnosis. Detailed menstrual history prior to and after adjuvant treatment was taken at study entry. Patients' background demographics, tumor characteristics and anti-cancer treatments were collected. The rates of CRA and CRM were determined. Analysis was conducted to identify factors associated with CRM. For postmenopausal patients, levels of E2 and FSH were analyzed.286 patients were recruited; the median time from breast cancer diagnosis to study entry was 5.0 years. 255 patients (91.1%) developed CRA. Of these, 66.7% regained menstruation. At the time of study entry, 137 (48.9%) had developed CRM, amongst whom 84 were age ≤45. On multivariate analysis, age was the only associated factor. Among patients with CRM, the median FSH was 41.0 IU/L; this was significantly lower in those who were taking tamoxifen compared to those who were not (20.1 vs. 59.7 IU/L, p

BPAG1a and BPAG1b (BPAG1a/b) constitute two major isoforms encoded by the dystonin (Dst) gene and show homology with MACF1a and MACF1b. These proteins are members of the plakin family, giant multi-modular proteins able to connect the intermediate filament, microtubule and microfilament cytoskeletal networks with each other and to distinct cell membrane sites. They also serve as scaffolds for signaling proteins that modulate cytoskeletal dynamics. To gain better insights into the functions of BPAG1a/b, we further characterized their C-terminal region important for their interaction with microtubules and assessed the role of these isoforms in the cytoskeletal organization of C2.7 myoblast cells. Our results show that alternative splicing does not only occur at the 5' end of Dst and Macf1 pre-mRNAs, as previously reported, but also at their 3' end, resulting in expression of additional four mRNA variants of BPAG1 and MACF1. These isoform-specific C-tails were able to bundle microtubules and bound to both EB1 and EB3, two microtubule plus end proteins. In the C2.7 cell line, knockdown of BPAG1a/b had no major effect on the organization of the microtubule and microfilament networks, but negatively affected endocytosis and maintenance of the Golgi apparatus structure, which became dispersed. Finally, knockdown of BPAG1a/b caused a specific decrease in the directness of cell migration, but did not impair initial cell adhesion. These data provide novel insights into the complexity of alternative splicing of Dst pre-mRNAs and into the role of BPAG1a/b in vesicular transport, Golgi apparatus structure as well as in migration in C2.7 myoblasts.

High bilirubin/albumin (B/A) ratios increase the risk of bilirubin neurotoxicity. The B/A ratio may be a valuable measure, in addition to the total serum bilirubin (TSB), in the management of hyperbilirubinemia. We aimed to assess whether the additional use of B/A ratios in the management of hyperbilirubinemia in preterm infants improved neurodevelopmental outcome.In a prospective, randomized controlled trial, 615 preterm infants of 32 weeks' gestation or less were randomly assigned to treatment based on either B/A ratio and TSB thresholds (consensus-based), whichever threshold was crossed first, or on the TSB thresholds only. The primary outcome was neurodevelopment at 18 to 24 months' corrected age as assessed with the Bayley Scales of Infant Development III by investigators unaware of treatment allocation. Secondary outcomes included complications of preterm birth and death.Composite motor (100 ± 13 vs. 101 ± 12) and cognitive (101 ± 12 vs. 101 ± 11) scores did not differ between the B/A ratio and TSB groups. Demographic characteristics, maximal TSB levels, B/A ratios, and other secondary outcomes were similar. The rates of death and/or severe neurodevelopmental impairment for the B/A ratio versus TSB groups were 15.4% versus 15.5% (P = 1.0) and 2.8% versus 1.4% (P = 0.62) for birth weights ≤ 1000 g and 1.8% versus 5.8% (P = 0.03) and 4.1% versus 2.0% (P = 0.26) for birth weights of >1000 g.The additional use of B/A ratio in the management of hyperbilirubinemia in preterm infants did not improve their neurodevelopmental outcome.Controlled-Trials.com ISRCTN74465643.

We aimed to identify a prostate cancer DNA hypermethylation microarray signature (denoted as PHYMA) that differentiates prostate cancer from benign prostate hyperplasia (BPH), high from low-grade and lethal from non-lethal cancers. This is a non-randomized retrospective study in 111 local Asian men (87 prostate cancers and 24 BPH) treated from 1995 to 2009 in our institution. Archival prostate epithelia were laser-capture microdissected and genomic DNA extracted and bisulfite-converted. Samples were profiled using Illumina GoldenGate Methylation microarray, with raw data processed by GenomeStudio. A classification model was generated using support vector machine, consisting of a 55-probe DNA methylation signature of 46 genes. The model was independently validated on an internal testing dataset which yielded cancer detection sensitivity and specificity of 95.3% and 100% respectively, with overall accuracy of 96.4%. Second validation on another independent western cohort yielded 89.8% sensitivity and 66.7% specificity, with overall accuracy of 88.7%. A PHYMA score was developed for each sample based on the state of methylation in the PHYMA signature. Increasing PHYMA score was significantly associated with higher Gleason score and Gleason primary grade. Men with higher PHYMA scores have poorer survival on univariate (p = 0.0038, HR = 3.89) and multivariate analyses when controlled for (i) clinical stage (p = 0.055, HR = 2.57), and (ii) clinical stage and Gleason score (p = 0.043, HR = 2.61). We further performed bisulfite genomic sequencing on 2 relatively unknown genes to demonstrate robustness of the assay results. PHYMA is thus a signature with high sensitivity and specificity for discriminating tumors from BPH, and has a potential role in early detection and in predicting survival.

Established cancers are frequently associated with a lymphocytic infiltrate that fails to clear the tumour mass. In contrast, the importance of recruited lymphocytes during premalignancy is less well understood. In a mouse model of premalignant skin epithelium, transgenic mice that express the human papillomavirus type 16 (HPV16) E7 oncoprotein under a keratin 14 promoter (K14E7 mice) display epidermal hyperplasia and have a predominant infiltrate of lymphocytes consisting of both CD4 and CD8 T cells. Activated, but not naïve T cells, were shown to preferentially traffic to hyperplastic skin with an increased frequency of proliferative CD8+ T cells and CD4+ T cells expressing CCR6 within the tissue. Disruption of the interaction between E7 protein and retinoblastoma tumour suppressor protein (pRb) led to reduced epithelial hyperplasia and T cell infiltrate. Finally, while K14E7 donor skin grafts are readily accepted onto syngeneic, non-transgenic recipients, these same skin grafts lacking skin-resident lymphocytes were rejected. Our data suggests that expression of a single oncoprotein in the epidermis is sufficient for lymphocyte trafficking (including immunosuppressive lymphocytes) to premalignant skin.

Microscopic [corrected] Observation Drug Susceptibility (MODS) has been shown to be an effective and rapid technique for early diagnosis of tuberculosis (TB). Thus far only a limited number of studies evaluating MODS have been performed in children and in extra-pulmonary tuberculosis. This study aims to assess relative accuracy and time to positive culture of MODS for TB diagnosis in children admitted to a general pediatric hospital in Vietnam.Specimens from children with suspected TB were tested by smear, MODS and Lowenstein-Jensen agar (LJ). 1129 samples from 705 children were analyzed, including sputum (n=59), gastric aspirate (n=775), CSF (n=148), pleural fluid (n=33), BAL (n=41), tracheal fluid (n=45), other (n=28). 113 TB cases were defined based on the "clinical diagnosis" (confirmed and probable groups) as the reference standard, in which 26% (n=30) were diagnosed as extra-pulmonary TB. Analysis by patient shows that the overall sensitivity and specificity of smear, LJ and MODS against "clinical diagnosis" was 8.8% and 100%, 38.9% and 100%, 46% and 99.5% respectively with MODS significantly more sensitive than LJ culture (P=0.02). When analyzed by sample type, the sensitivity of MODS was significantly higher than LJ for gastric aspirates (P=0.004). The time to detection was also significantly shorter for MODS than LJ (7 days versus 32 days, P

How pathogenic bacteria adapt and evolve in the complex and variable environment of the host remains a largely unresolved question. Here we have used whole genome sequencing of Salmonella enterica serovar Typhimurium LT2 populations serially passaged in mice to identify mutations that adapt bacteria to systemic growth in mice. We found unique pathoadaptive mutations in two global regulators, phoQ and stpA, which increase the competitive indexes of the bacteria 3- to 5-fold. Also, all mouse-adapted lineages had changed the orientation of the hin invertable element, resulting in production of a FliC type of flagellum. Competition experiments in mice with locked flagellum mutants showed that strains expressing the FliC type of flagellum had a 5-fold increase in competitive index as compared to those expressing FljB type flagellum. Combination of the flagellum cassette inversion with the stpA mutation increased competitive indexes up to 20-fold. These experiments show that Salmonella can rapidly adapt to a mouse environment by acquiring a few mutations of moderate individual effect that when combined confer substantial increases in growth.

BACKGROUND:Substituting sugar-free for sugar-sweetened beverages reduces weight gain. A possible explanation is that sugar-containing and sugar-free beverages cause the same degree of satiety. However, this has not been tested in long-term trials. METHODS:We randomized 203 children aged 7-11 years to receive 250 mL per day of an artificially sweetened sugar-free beverage or a similarly looking and tasting sugar-sweetened beverage. We measured satiety on a 5-point scale by questionnaire at 0, 6, 12 and 18 months. We calculated the change in satiety from before intake to 1 minute after intake and 15 minutes after intake. We then calculated the odds ratio that satiety increased by 1 point in the sugar-group versus the sugar-free group. We also investigated how much the children liked and wanted the beverages. RESULTS:146 children or 72% completed the study. We found no statistically significant difference in satiety between the sugar-free and sugar-sweetened group; the adjusted odds ratio for a 1 point increase in satiety in the sugar group versus the sugar-free group was 0.77 at 1 minute (95% confidence interval, 0.46 to 1.29), and 1.44 at 15 minutes after intake (95% CI, 0.86 to 2.40). The sugar-group liked and wanted their beverage slightly more than the sugar-free group, adjusted odds ratio 1.63 (95% CI 1.05 to 2.54) and 1.65 (95% CI 1.07 to 2.55), respectively. CONCLUSIONS:Sugar-sweetened and sugar-free beverages produced similar satiety. Therefore when children are given sugar-free instead of sugar-containing drinks they might not make up the missing calories from other sources. This may explain our previous observation that children in the sugar-free group accumulated less body fat than those in the sugar group. TRIAL REGISTRATION:ClinicalTrials.gov NCT00893529 http://clinicaltrials.gov/show/NCT00893529.

Current regimens for induction therapy of pediatric acute lymphoblastic leukemia (ALL), or for re-induction post relapse, use a combination of vincristine (VCR), a glucocorticoid, and L-asparaginase (ASP) with or without an anthracycline. With cure rates now approximately 80%, robust pre-clinical models are necessary to prioritize active new drugs for clinical trials in relapsed/refractory patients, and the ability of these models to predict synergy/antagonism with established therapy is an essential attribute. In this study, we report optimization of an induction-type regimen by combining VCR, dexamethasone (DEX) and ASP (VXL) against ALL xenograft models established from patient biopsies in immune-deficient mice. We demonstrate that the VXL combination was synergistic in vitro against leukemia cell lines as well as in vivo against ALL xenografts. In vivo, VXL treatment caused delays in progression of individual xenografts ranging from 22 to >146 days. The median progression delay of xenografts derived from long-term surviving patients was 2-fold greater than that of xenografts derived from patients who died of their disease. Pharmacokinetic analysis revealed that systemic DEX exposure in mice increased 2-fold when administered in combination with VCR and ASP, consistent with clinical findings, which may contribute to the observed synergy between the 3 drugs. Finally, as proof-of-principle we tested the in vivo efficacy of combining VXL with either the Bcl-2/Bcl-xL/Bcl-w inhibitor, ABT-737, or arsenic trioxide to provide evidence of a robust in vivo platform to prioritize new drugs for clinical trials in children with relapsed/refractory ALL.

Although gemcitabine is highly active in several cancer types, intrinsic and acquired drug resistance remains a major challenge. Overexpression of Bcl-2 has been associated with gemcitabine resistance. The aim of this study is to determine whether gossypol can overcome gemcitabine resistance in cell lines with high level of Bcl-2 expression in combination drug therapy. Our study demonstrated that in 10 cell lines derived from different cancers, high Bcl-2 baseline expression was observed in cell lines that were resistant to gemcitabine (GEM-R). Furthermore, synergistic effect of combination therapy was observed in gemcitabine-resistant (GEM-R) cell lines with high Bcl-2 expression, but not in a gemcitabine-sensitive (GEM-S) cell lines regardless of Bcl-2 expression. Gossypol treatment resulted in the decrease of anti-apoptotic genes such as Bcl-2 and Bcl-xl and an upregulation of the pro-apoptotic gene, Noxa. Furthermore, the addition of gossypol to gemcitabine resulted in lower expressions of anti-apoptotic genes compared to gemcitabine alone. Gene expression profiling in GEM-R and GEM-S cell lines suggest that anti-apoptotic genes such as pAkt and PI3KR2 may play important role in gemcitabine resistance, while pro-apoptotic Bcl-2 related genes (Bad, Caspase-6 and Calpain-1) may regulate synergistic interaction in combination therapy.

Food allergies, and peanut allergy in particular, are leading causes of anaphylactic fatalities worldwide. The immune mechanisms that underlie food allergy remain ill-defined and controversial, in part because studies in humans typically focus on analysis of a limited number of prototypical Th1/Th2 cytokines. Here we determine the kinetics and prevalence of a broad panel of peanut-driven cytokine and chemokine responses in humans with current peanut allergy vs those with stable, naturally occurring clinical tolerance to peanut. Our primary focus is identification of novel indicators of immune dysregulation. Antigen-specific cytokine mRNA and protein responses were elicited in primary culture via peanut or irrelevant antigen (Leishmania extract, milk antigens) mediated stimulation of fresh peripheral blood cells from 40 individuals. Peanut extract exposure in vitro induced a broad panel of responses associated with Th2/Th9-like, Th1-like and Th17-like immunity. Peanut-dependent Type 2 cytokine responses were frequently found in both peanut allergic individuals and those who exhibit clinical tolerance to peanut ingestion. Among Th2/Th9-associated cytokines, IL-9 responses discriminated between allergic and clinically tolerant populations better than did commonly used IL-4, IL-5 or IL-13 responses. Comparison with responses evoked by unrelated control antigen-mediated stimulation showed that these differences are antigen-dependent and allergen-specific. Conversely, the intensity of IL-12, IL-17, IL-23 and IFN-γ production was indistinguishable in peanut allergic and peanut tolerant populations. In summary, the ability to generate and maintain cytokine responses to peanut is not inherently distinct between allergic and peanut tolerant humans. Quantitative differences in the intensity of cytokine production better reflects clinical phenotype, with optimally useful indicators being IL-9, IL-5, IL-13 and IL-4. Equivalent, and minimal, Ag-dependent pro-inflammatory cytokine levels in both healthy and peanut allergic volunteers argues against a key role for such cytokines in maintenance of clinical tolerance to food antigens in humans.

Plague disease caused by the gram-negative bacterium Yersinia pestis routinely affects animals and occasionally humans, in the western United States. The strains native to the North American continent are thought to be derived from a single introduction in the late 19(th) century. The degree to which these isolates have diverged genetically since their introduction is not clear, and new genomic markers to assay the diversity of North American plague are highly desired. To assay genetic diversity of plague isolates within confined geographic areas, draft genome sequences were generated by 454 pyrosequencing from nine environmental and clinical plague isolates. In silico assemblies of Variable Number Tandem Repeat (VNTR) loci were compared to laboratory-generated profiles for seven markers. High-confidence SNPs and small Insertion/Deletions (Indels) were compared to previously sequenced Y. pestis isolates. The resulting panel of mutations allowed clustering of the strains and tracing of the most likely evolutionary trajectory of the plague strains. The sequences also allowed the identification of new putative SNPs that differentiate the 2009 isolates from previously sequenced plague strains and from each other. In addition, new insertion points for the abundant insertion sequences (IS) of Y. pestis are present that allow additional discrimination of strains; several of these new insertions potentially inactivate genes implicated in virulence. These sequences enable whole-genome phylogenetic analysis and allow the unbiased comparison of closely related isolates of a genetically monomorphic pathogen.

BackgroundDespite the decades-long use of Bacillus atrophaeus var. globigii (BG) as a simulant for biological warfare (BW) agents, knowledge of its genome composition is limited. Furthermore, the ability to differentiate signatures of deliberate adaptation and selection from natural variation is lacking for most bacterial agents. We characterized a lineage of BGwith a long history of use as a simulant for BW operations, focusing on classical bacteriological markers, metabolic profiling and whole-genome shotgun sequencing (WGS).ResultsArchival strains and two "present day" type strains were compared to simulant strains on different laboratory media. Several of the samples produced multiple colony morphotypes that differed from that of an archival isolate. To trace the microevolutionary history of these isolates, we obtained WGS data for several archival and present-day strains and morphotypes. Bacillus-wide phylogenetic analysis identified B. subtilis as the nearest neighbor to B. atrophaeus. The genome of B. atrophaeus is, on average, 86% identical to B. subtilis on the nucleotide level. WGS of variants revealed that several strains were mixed but highly related populations and uncovered a progressive accumulation of mutations among the "military" isolates. Metabolic profiling and microscopic examination of bacterial cultures revealed enhanced growth of "military" isolates on lactate-containing media, and showed that the "military" strains exhibited a hypersporulating phenotype.ConclusionsOur analysis revealed the genomic and phenotypic signatures of strain adaptation and deliberate selection for traits that were desirable in a simulant organism. Together, these results demonstrate the power of whole-genome and modern systems-level approaches to characterize microbial lineages to develop and validate forensic markers for strain discrimination and reveal signatures of deliberate adaptation.

Rare (RVs) and common variants of the RET gene contribute to Hirschsprung disease (HSCR; congenital aganglionosis). While RET common variants are strongly associated with the commonest manifestation of the disease (males; short-segment aganglionosis; sporadic), rare coding sequence (CDS) variants are more frequently found in the lesser common and more severe forms of the disease (females; long/total colonic aganglionosis; familial).Here we present the screening for RVs in the RET CDS and intron/exon boundaries of 601 Chinese HSCR patients, the largest number of patients ever reported. We identified 61 different heterozygous RVs (50 novel) distributed among 100 patients (16.64%). Those include 14 silent, 29 missense, 5 nonsense, 4 frame-shifts, and one in-frame amino-acid deletion in the CDS, two splice-site deletions, 4 nucleotide substitutions and a 22-bp deletion in the intron/exon boundaries and 1 single-nucleotide substitution in the 5' untranslated region. Exonic variants were mainly clustered in RET the extracellular domain. RET RVs were more frequent among patients with the most severe phenotype (24% vs. 15% in short-HSCR). Phasing RVs with the RET HSCR-associated haplotype suggests that RVs do not underlie the undisputable association of RET common variants with HSCR. None of the variants were found in 250 Chinese controls.

BACKGROUND: Rhegmatogenous retinal detachment (RRD) is a major cause of visual loss in developed countries. Proliferative vitreoretinopathy (PVR), an eye-sight threatening complication of RRD surgery, resembles a wound-healing process with inflammation, scar tissue formation, and membrane contraction. This study was performed to determine the possible involvement of a wide range of cytokines in the future development of PVR, and to identify predictors of PVR and visual outcome. METHODOLOGY: A multiplex immunoassay was used for the simultaneous detection of 29 different cytokines in subretinal fluid samples from patients with primary RRD. Of 306 samples that were collected and stored in our BioBank between 2001 and 2008, 21 samples from patients who developed postoperative PVR were compared with 54 age-, sex-, and storage-time-matched RRD control patients who had an uncomplicated postoperative course during the overall follow-up period. FINDINGS: Levels of IL-1α, IL-2, IL-3, IL-6, VEGF, and ICAM-1 were significantly higher (P0.05). Multivariate logistic regression analysis revealed that IL-3 (P = 0.001), IL-6 (P = 0.047), ICAM-1 (P = 0.010), and preoperative visual acuity (P = 0.026) were independent predictors of postoperative PVR. Linear regression analysis showed that ICAM-1 (P = 0.005) and preoperative logMAR visual acuity (P = 0.001) were predictive of final visual outcome after primary RRD repair. CONCLUSIONS/SIGNIFICANCE: Our findings indicate that after RRD onset an exaggerated response of certain cytokines may predispose to PVR. Sampling at a time close to the onset of primary RRD may thus provide clues as to which biological events may initiate the development of PVR and, most importantly, may provide a means for therapeutic control.