Old tools, new applications: Use of environmental bacteriophages for typhoid surveillance and evaluating vaccine impact.

Typhoid-conjugate vaccines (TCVs) provide an opportunity to reduce the burden of typhoid fever, caused by Salmonella Typhi, in endemic areas. As policymakers design vaccination strategies, accurate and high-resolution data on disease burden is crucial. However, traditional blood culture-based surveillance is resource-extensive, prohibiting its large-scale and sustainable implementation. Salmonella Typhi is a water-borne pathogen, and here, we tested the potential of Typhi-specific bacteriophage surveillance in surface water bodies as a low-cost tool to identify where Salmonella Typhi circulates in the environment. In 2021, water samples were collected and tested for the presence of Salmonella Typhi bacteriophages at two sites in Bangladesh: urban capital city, Dhaka, and a rural district, Mirzapur. Salmonella Typhi-specific bacteriophages were detected in 66 of 211 (31%) environmental samples in Dhaka, in comparison to 3 of 92 (3%) environmental samples from Mirzapur. In the same year, 4,620 blood cultures at the two largest pediatric hospitals of Dhaka yielded 215 (5%) culture-confirmed typhoid cases, and 3,788 blood cultures in the largest hospital of Mirzapur yielded 2 (0.05%) cases. 75% (52/69) of positive phage samples were collected from sewage. All isolated phages were tested against a panel of isolates from different Salmonella Typhi genotypes circulating in Bangladesh and were found to exhibit a diverse killing spectrum, indicating that diverse bacteriophages were isolated. These results suggest an association between the presence of Typhi-specific phages in the environment and the burden of typhoid fever, and the potential of utilizing environmental phage surveillance as a low-cost tool to assist policy decisions on typhoid control.

Effectiveness of ten-valent pneumococcal conjugate vaccine on invasive pneumococcal disease among children <2 years old: A prospective population-based study in rural Bangladesh.

The 10-valent pneumococcal conjugate vaccine (PCV10) was introduced in March 2015 in Bangladesh. In this study, we aimed to estimate the impact of PCV10 on invasive pneumococcal disease (IPD) identified by blood cultures and severe pneumonia identified clinically and its effectiveness on invasive disease caused by vaccine serotypes. We conducted population-based surveillance among children aged 2- <24 months between April 2012 through March 2019 in Mirzapur, a rural sub-district of Bangladesh. We compared incidence of IPD and severe pneumonia before (April 2012 to March 2015) and after (April 2015 to March 2019) the introduction of PCV10. Vaccine effectiveness was measured using an indirect cohort analysis of data from four sentinel sites in which PCV10 vaccination status was compared between children with IPD caused by vaccine serotype vs. non-vaccine serotypes. We identified 24 IPD cases by blood culture and 1,704 severe pneumonia hospitalizations during the surveillance period. IPD incidence in under-2-year-old children fell 25 % (95 % CI: -1.2 % to 76 %; p-value = 0.59) from 106 cases per 100,000 child-years at baseline to 79.3 in April 2018- March 2019. Vaccine serotype-IPD incidence was lower (77 % reduction, 95 % CI: -0.45 % to 96 %; p-value = 0.068) in April 2018 - March 2019 than in the pre-vaccine period (85.7 cases to 19.8/100,000 child-years). A significant decline of 54.0 % (95 % CI: 47.0 % to 59.0 %; p-value < 0.001) was observed in hospitalizations due to severe pneumonia. From indirect cohort analysis, the effectiveness of PCV10 against vaccine serotype IPD was 37 % (95 % CI: -141.0 % to 83.5 %; p = 0.5) after the 1st dose and 63.1 % (95 % CI: -3.3 % to 85.9 %, p = 0.0411) after the 2nd or the 3rd dose. This study demonstrates that PCV10 introduction prevented hospitalizations with severe pneumonia and provided individual protection against vaccine serotypes.

Peripheral artery disease causes consistent gait irregularities regardless of the location of leg claudication pain.

BACKGROUND: The most common symptom of peripheral artery disease (PAD) is intermittent claudication that involves the calf, thigh, and/or buttock muscles. How the specific location of this leg pain is related to altered gait, however, is unknown. OBJECTIVES: We hypothesized that because the location of claudication symptoms uniquely affects different leg muscle groups in people with PAD, this would produce distinctive walking patterns. METHODS: A total of 105 participants with PAD and 35 age-matched older volunteers without PAD (CTRL) were recruited. Participants completed walking impairment questionnaires (WIQ), Gardner-Skinner progressive treadmill tests, the six-minute walk test, and we performed an advanced evaluation of the biomechanics of their overground walking. Participants with PAD were categorized into 4 groups according to their stated pain location(s): calf only (C, n = 43); thigh and calf (TC, n = 18); buttock and calf (BC, n = 15); or buttock, thigh, and calf (BTC, n = 29). Outcomes were compared between CTRL, C, TC, BC and BTC groups using a one-way ANOVA with post-hoc comparisons to identify and assess statistically significant differences. RESULTS: There were no significant differences between CTRL, C, TC, BC and BTC groups in distances walked or walking speed when either pain-free or experiencing claudication pain. Each participant with PAD had significantly dysfunctional biomechanical gait parameters, even when pain-free, when compared to CTRL (pain-free) walking data. During pain-free walking, out of the 18 gait parameters evaluated, we only identified significant differences in hip power generation during push-off (in C and TC groups) and in knee power absorption during weight acceptance (in TC and BC groups). There were no between-group differences in gait parameters while people with PAD were walking with claudication pain. CONCLUSIONS: Our data demonstrate that PAD affects the ischemic lower extremities in a diffuse manner irrespective of the location of claudication symptoms. DATABASE REGISTRATION: ClinicalTrials.gov NCT01970332.

Assessing implementation outcomes for launching balanced energy protein supplementation: A formative study in rural Bangladesh.

Balanced energy protein (BEP) supplementation is an efficacious intervention in pregnancy for improving birthweight and is recommended by World Health Organization (WHO) in countries with high maternal undernutrition. Few countries have implemented BEP programmes due in part to high cost, lack of data on acceptability and feasibility, and complexity of delivery. We sought to address implementation gaps in BEP interventions through a formative study designed to understand implementation outcomes. We conducted 52 in-depth interviews and 8 focus-group discussions with married women of reproductive age, family members, health care providers and pharmacists in three unions of the Gaibandha district in rural Bangladesh. Interviews were translated and transcribed in English and analysed using an analytic framework for implementation science in nutrition. BEP was viewed as an acceptable and appropriate intervention to combat undernutrition in this setting. There was a lack of clarity on who should or could be responsible for providing/distributing BEP in a way convenient to mothers. Many participants preferred door-to-door delivery and thought this approach could address social and gender inequities, but providers mentioned already being overworked and worried about adding new tasks. Participants were concerned about the affordability of BEP and opportunity costs associated with travel to proposed distribution sites such as ANC or pharmacies. Women in these communities do not always have the agency to travel without supervision or make purchasing decisions. BEP supplementation is a complex intervention; future trials seek to assess ways to overcome these implementation challenges and inform a long-term systems-owned BEP intervention.

Trends in antimicrobial resistance amongst Salmonella Paratyphi A isolates in Bangladesh: 1999-2021.

BACKGROUND: Typhoid and paratyphoid remain common bloodstream infections in areas with suboptimal water and sanitation infrastructure. Paratyphoid, caused by Salmonella Paratyphi A, is less prevalent than typhoid and its antimicrobial resistance (AMR) trends are less documented. Empirical treatment for paratyphoid is commonly based on the knowledge of susceptibility of Salmonella Typhi, which causes typhoid. Hence, with rising drug resistance in Salmonella Typhi, last-line antibiotics like ceftriaxone and azithromycin are prescribed for both typhoid and paratyphoid. However, unlike for typhoid, there is no vaccine to prevent paratyphoid. Here, we report 23-year AMR trends of Salmonella Paratyphi A in Bangladesh. METHODS: From 1999 to 2021, we conducted enteric fever surveillance in two major pediatric hospitals and three clinics in Dhaka, Bangladesh. Blood cultures were performed at the discretion of the treating physicians; cases were confirmed by culture, serological and biochemical tests. Antimicrobial susceptibility was determined following CLSI guidelines. RESULTS: Over 23 years, we identified 2,725 blood culture-confirmed paratyphoid cases. Over 97% of the isolates were susceptible to ampicillin, chloramphenicol, and cotrimoxazole, and no isolate was resistant to all three. No resistance to ceftriaxone was recorded, and >99% of the isolates were sensitive to azithromycin. A slight increase in minimum inhibitory concentration (MIC) is noticed for ceftriaxone but the current average MIC is 32-fold lower than the resistance cut-off. Over 99% of the isolates exhibited decreased susceptibility to ciprofloxacin. CONCLUSIONS: Salmonella Paratyphi A has remained susceptible to most antibiotics, unlike Salmonella Typhi, despite widespread usage of many antibiotics in Bangladesh. The data can guide evidence-based policy decisions for empirical treatment of paratyphoid fever, especially in the post typhoid vaccine era, and with the availability of new paratyphoid diagnostics.

Acceptability of a balanced energy protein (BEP) supplement for pregnant women in Bangladesh.

Balanced energy protein (BEP) supplementation in pregnancy is recommended in the context of undernutrition for the reduction of small-for-gestational age neonates and stillbirths. To inform an effectiveness trial, we evaluated the acceptability of a packaged, ready-to-eat fortified BEP product among women of reproductive age and their health care providers (HCPs) in rural Bangladesh and explored the feasibility of adhering to daily supplementation. We implemented a formative study using focus groups discussions with women (n = 29) and HCPs (n = 17) to introduce the product and investigate components of acceptability. A "trials of improved practice" activity was conducted in subset of women (n = 16) to evaluate adherence to BEP over a 2-week period, followed by focus group discussions to identify challenges with adherence and strategies employed. Contributors to BEP acceptability included the product's sensory attributes, such as taste, smell and texture; the attractive packaging and informative labelling; and the perceived benefits of use. Participants also identified household and community level factors influencing the adoption of BEP, such as trust in the provider, cultural beliefs on supplement use in pregnancy, and family member tasting and approval. Over the 2-week period, women consumed over 80% of the supplements provided to them and identified strategies for adherence, including visual aids and reminders from family members or providers. HCPs recommended targeted communication messages for mothers-in-law to foster a supportive home environment. Findings informed changes to the BEP product to improve acceptability and shaped the content of communication messages to optimise adherence in a forthcoming effectiveness trial.

COVID-19 vaccine eligibility of pregnant and lactating women in Bangladesh: Gap between policy and policy interpretation among policymakers and healthcare workers.

SARS-CoV-2 infection in pregnancy is associated with a greater risk of maternal and newborn morbidity and maternal death. Bangladesh confirmed its first COVID-19 case in March of 2020, and vaccination rollout started in January of 2021. In Bangladesh, pregnant women are allowed to receive COVID-19 vaccines during pregnancy with qualifications while lactating women are permitted to receive COVID-19 vaccines with no qualifications as of October 2021. There is limited evidence on how vaccine policies are disseminated, interpreted, and implemented from the national level to the community level in Bangladesh. We conducted in-depth interviews from April-August 2022 with policymakers and healthcare workers in Bangladesh to understand how different stakeholders understood and implemented COVID-19 vaccination policies related to pregnant and lactating women. We interviewed policymakers at three levels: national, divisional, and district, and interviewed healthcare workers from one one urban and three rural communities within one division. We found a gap between policies related to COVID-19 vaccination for pregnant and lactating women and policy interpretation among policymakers and healthcare workers. Policymakers and healthcare workers' perceptions differed related to policy dissemination, attitudes toward policies related to pregnant and lactating women, and eligibility of pregnant and lactating women. Our findings indicate the need for effective dissemination of and understanding of policies. Within the context of vaccine uptake and vaccine acceptance, policymakers play a critical role as they are charged with developing and disseminating policy related to vaccine eligibility. Healthcare workers rely on timely and accurate communication related to vaccine eligibility, including populations, timing, and locations. Efforts are needed to narrow the policy and policy implementation gap as doing so is crucial to controlling vaccine preventable disease.

Sources of COVID-19 Vaccine Promotion for Pregnant and Lactating Women in Bangladesh.

COVID-19 vaccines are an effective public health intervention to reduce COVID-19-related morbidity and mortality. Given that pregnant and lactating women have a higher risk of severe COVID-19 complications, it is paramount to understand the factors that inform vaccine decision-making among this population. In this study, we sought to identify facilitators and barriers to COVID-19 vaccine acceptance and vaccine promotion in pregnant and lactating women in Bangladesh. We conducted 40 in-depth interviews with 12 pregnant women, 12 lactating women, and 16 health workers from one urban and four rural communities in Bangladesh. We used a grounded theory approach to identify emerging themes. Our results suggest that health workers and religious leaders played key roles in promoting COVID-19 vaccines in this population. Further, we found that the culture of trust in public health authorities and the existing vaccine infrastructure facilitated vaccine promotion. However, changes in vaccine eligibility and myths and rumors acted as both facilitators and barriers to vaccine promotion within our study. It is crucial that maternal immunization vaccine promotion efforts push pregnant and lactating women toward vaccine acceptance to protect the health of mothers and their babies. Additionally, as new maternal vaccines are developed and licensed, understanding how to best promote vaccines within this group is paramount.

Water-Tight Arthrotomy Joint Closure of Modified Intervastus Approach in Total Knee Arthroplasty.

BACKGROUND: The joint closure technique used for total knee arthroplasty cases can have an impact on outcomes, especially when considering accelerated rehabilitation programs that follow surgery. In this study, we describe the details of the technical steps involved in performing the water-tight arthrotomy joint closure technique that we developed and use. METHODS: A total of 536 patients (average age: 62 years, average body mass index: 34 kg/m2) with primary osteoarthritis of the knee underwent total knee arthroplasty using the modified intervastus approach between 2019 and 2021. We used the water-tight arthrotomy joint closure technique to close the knee arthrotomy incision. Any infections and complications, as well as the duration of surgery and cost related to this wound closure technique, are also reported. RESULTS: Few complications were noted with this closure technique. When we first started using it, there was one case of drainage through the proximal capsular repair which required a return to the operating room 5 days postoperatively for an irrigation and debridement. We also had two cases of superficial skin necrosis along a small part of the incision line which were observed on a weekly basis and which healed uneventfully with application of betadine once daily on the necrotic area. The average time for performing wound closure after total knee arthroplasty was 45 min. CONCLUSION: We conclude that the water-tight closure approach can achieve very durable, water-tight capsule repairs and results in a decrease in postoperative wound drainage.

Densely Convolutional Spatial Attention Network for nuclei segmentation of histological images for computational pathology.

Introduction: Automatic nuclear segmentation in digital microscopic tissue images can aid pathologists to extract high-quality features for nuclear morphometrics and other analyses. However, image segmentation is a challenging task in medical image processing and analysis. This study aimed to develop a deep learning-based method for nuclei segmentation of histological images for computational pathology. Methods: The original U-Net model sometime has a caveat in exploring significant features. Herein, we present the Densely Convolutional Spatial Attention Network (DCSA-Net) model based on U-Net to perform the segmentation task. Furthermore, the developed model was tested on external multi-tissue dataset - MoNuSeg. To develop deep learning algorithms for well-segmenting nuclei, a large quantity of data are mandatory, which is expensive and less feasible. We collected hematoxylin and eosin-stained image data sets from two hospitals to train the model with a variety of nuclear appearances. Because of the limited number of annotated pathology images, we introduced a small publicly accessible data set of prostate cancer (PCa) with more than 16,000 labeled nuclei. Nevertheless, to construct our proposed model, we developed the DCSA module, an attention mechanism for capturing useful information from raw images. We also used several other artificial intelligence-based segmentation methods and tools to compare their results to our proposed technique. Results: To prioritize the performance of nuclei segmentation, we evaluated the model's outputs based on the Accuracy, Dice coefficient (DC), and Jaccard coefficient (JC) scores. The proposed technique outperformed the other methods and achieved superior nuclei segmentation with accuracy, DC, and JC of 96.4% (95% confidence interval [CI]: 96.2 - 96.6), 81.8 (95% CI: 80.8 - 83.0), and 69.3 (95% CI: 68.2 - 70.0), respectively, on the internal test data set. Conclusion: Our proposed method demonstrates superior performance in segmenting cell nuclei of histological images from internal and external datasets, and outperforms many standard segmentation algorithms used for comparative analysis.

COVID-19 vaccine decision-making among pregnant and lactating women in Bangladesh.

Pregnant and lactating women's vaccine decision-making process is influenced by many factors. Pregnant women were at increased risk for severe disease and poor health outcomes from COVID-19 at various time points during the pandemic. COVID-19 vaccines have been found to be safe and protective during pregnancy and while breastfeeding. In this study, we sought to examine key factors that informed the decision-making process among pregnant and lactating women in Bangladesh. We conducted 24 in-depth interviews, with 12 pregnant and 12 lactating women. These women were from three communities in Bangladesh: one urban community, and two rural communities. We used a grounded theory approach to identify emerging themes and organized emerging themes using a socio-ecological model. The socio-ecological model suggests that individuals are influenced by many levels, including individual-level influences, interpersonal-level influences, health care system-level influences, and policy-level influences. We found key factors at each socio-ecological level that influenced the decision-making process of pregnant and lactating women, including perceived benefits of vaccines and vaccine safety (individual-level), the influence of husbands and peers (interpersonal-level), health care provider recommendations and vaccine eligibility (health care system-level), and vaccine mandates (policy-level). As vaccination can reduce the effect of COVID-19 disease in mothers, infants, and unborn children, targeting critical factors that inform the decision-making process is paramount for improving vaccine acceptance. We hope the results of this study will inform vaccine acceptance efforts to ensure that pregnant and lactating women take advantage of this life-saving intervention.

Neonatal invasive candidiasis in low- and middle-income countries: Data from the NeoOBS study.

Neonatal invasive candidiasis (NIC) has significant morbidity and mortality. Reports have shown a different profile of those neonates affected with NIC and of fluconazole-resistant Candida spp. isolates in low- and middle-income countries (LMICs) compared to high-income countries (HICs). We describe the epidemiology, Candida spp. distribution, treatment, and outcomes of neonates with NIC from LMICs enrolled in a global, prospective, longitudinal, observational cohort study (NeoOBS) of hospitalized infants <60 days postnatal age with sepsis (August 2018-February 2021). A total of 127 neonates from 14 hospitals in 8 countries with Candida spp. isolated from blood culture were included. Median gestational age of affected neonates was 30 weeks (IQR: 28-34), and median birth weight was 1270 gr (interquartile range [IQR]: 990-1692). Only a minority had high-risk criteria, such as being born <28 weeks, 19% (24/127), or birth weight <1000 gr, 27% (34/127). The most common Candida species were C. albicans (n = 45, 35%), C. parapsilosis (n = 38, 30%), and Candida auris (n = 18, 14%). The majority of C. albicans isolates were fluconazole susceptible, whereas 59% of C. parapsilosis isolates were fluconazole-resistant. Amphotericin B was the most common antifungal used [74% (78/105)], followed by fluconazole [22% (23/105)]. Death by day 28 post-enrollment was 22% (28/127). To our knowledge, this is the largest multi-country cohort of NIC in LMICs. Most of the neonates would not have been considered at high risk for NIC in HICs. A substantial proportion of isolates was resistant to first choice fluconazole. Understanding the burden of NIC in LMIC is essential to guide future research and treatment guidelines.

Our study describes neonates from low- and middle-income countries with neonatal invasive candidiasis (NIC). Most of them were outside the groups considered at high risk for NIC described in high-income countries. Candida spp. epidemiology was also different. The mortality was high (22%). Further research in these settings is required.

Region Segmentation of Whole-Slide Images for Analyzing Histological Differentiation of Prostate Adenocarcinoma Using Ensemble EfficientNetB2 U-Net with Transfer Learning Mechanism.

Recent advances in computer-aided detection via deep learning (DL) now allow for prostate cancer to be detected automatically and recognized with extremely high accuracy, much like other medical diagnoses and prognoses. However, researchers are still limited by the Gleason scoring system. The histopathological analysis involved in assigning the appropriate score is a rigorous, time-consuming manual process that is constrained by the quality of the material and the pathologist's level of expertise. In this research, we implemented a DL model using transfer learning on a set of histopathological images to segment cancerous and noncancerous areas in whole-slide images (WSIs). In this approach, the proposed Ensemble U-net model was applied for the segmentation of stroma, cancerous, and benign areas. The WSI dataset of prostate cancer was collected from the Kaggle repository, which is publicly available online. A total of 1000 WSIs were used for region segmentation. From this, 8100 patch images were used for training, and 900 for testing. The proposed model demonstrated an average dice coefficient (DC), intersection over union (IoU), and Hausdorff distance of 0.891, 0.811, and 15.9, respectively, on the test set, with corresponding masks of patch images. The manipulation of the proposed segmentation model improves the ability of the pathologist to predict disease outcomes, thus enhancing treatment efficacy by isolating the cancerous regions in WSIs.

Machine Learning-Based Peripheral Artery Disease Identification Using Laboratory-Based Gait Data.

Peripheral artery disease (PAD) manifests from atherosclerosis, which limits blood flow to the legs and causes changes in muscle structure and function, and in gait performance. PAD is underdiagnosed, which delays treatment and worsens clinical outcomes. To overcome this challenge, the purpose of this study is to develop machine learning (ML) models that distinguish individuals with and without PAD. This is the first step to using ML to identify those with PAD risk early. We built ML models based on previously acquired overground walking biomechanics data from patients with PAD and healthy controls. Gait signatures were characterized using ankle, knee, and hip joint angles, torques, and powers, as well as ground reaction forces (GRF). ML was able to classify those with and without PAD using Neural Networks or Random Forest algorithms with 89% accuracy (0.64 Matthew's Correlation Coefficient) using all laboratory-based gait variables. Moreover, models using only GRF variables provided up to 87% accuracy (0.64 Matthew's Correlation Coefficient). These results indicate that ML models can classify those with and without PAD using gait signatures with acceptable performance. Results also show that an ML gait signature model that uses GRF features delivers the most informative data for PAD classification.

Management of mangled hand injuries with JESS fixator in a tertiary care hospital in North East India: A prospective study.

Introduction: Mangled hand injuries have proved to be a tough challenge for both the patients and surgeons alike. Severe bony and soft tissue injuries necessitate the requirement of a multidisciplinary approach. Materials: In our tertiary care facility, a prospective study was done on the use of the Joshi's external stabilization system (JESS) fixator in the treatment of 31 instances with mangled hand injuries from November 2019 to November 2020. The cases were evaluated for functional outcome at the end of 12 months of follow up. Results: 31 cases of mangled hand injuries with 43 fractures were treated with the said intervention. The mean age of the patients was 33.7 ± 4.2 years and almost 90% patients were males. Machinery injuries were the most frequent type of injury (55%). Cases were intervened at an emergency basis within 24 h of the injury. Functional evaluation was done by Modified Mayo Wrist Score with an average score of 84 ± 10. Good results were seen in about 60% of cases and there were no Poor results. Conclusion: The application of JESS fixator for the prompt treatment of mangling hand injuries yielded satisfactory results in the working population with an early return to function. The strict adherence to the principles of ligamentotaxis, appropriate wound care and post-operative physiotherapy are other variables in the final outcome of such injuries. Level of evidence: II.

Temporal and Spatial Differences between Symptomatic and Asymptomatic Malaria Infections in the Chittagong Hill Districts, Bangladesh.

Mapping asymptomatic malaria infections, which contribute to the transmission reservoir, is important for elimination programs. This analysis compared the spatiotemporal patterns of symptomatic and asymptomatic Plasmodium falciparum malaria infections in a cohort study of ∼25,000 people living in a rural hypoendemic area of about 179 km2 in a small area of the Chittagong Hill Districts of Bangladesh. Asymptomatic infections were identified by active surveillance; symptomatic clinical cases presented for care. Infections were identified by a positive rapid diagnostic test and/or microscopy. Fifty-three subjects with asymptomatic P. falciparum infection were compared with 572 subjects with symptomatic P. falciparum between mid-October 2009 and mid-October 2012 with regard to seasonality, household location, and extent of spatial clustering. We found increased spatial clustering of symptomatic compared with asymptomatic infections, and the areas of high intensity were only sometimes overlapping. Symptomatic cases had a distinct seasonality, unlike asymptomatic infections, which were detected year-round. In a comparison of 42 symptomatic Plasmodium vivax and 777 symptomatic P. falciparum cases from mid-October 2009 through mid-March 2015, we found substantial spatial overlap in areas with high infection rates, but the areas with the greatest concentration of infection differed. Detection of both symptomatic P. falciparum and symptomatic P. vivax infections was greater during the May-to-October high season, although a greater proportion of P. falciparum cases occurred during the high season compared with P. vivax. These findings reinforce that passive malaria surveillance and treatment of symptomatic cases will not eliminate the asymptomatic reservoirs that occur distinctly in time and space.

Comprehensive In Silico Analysis of RNA Silencing-Related Genes and Their Regulatory Elements in Wheat (Triticum aestivum L.).

Dicer-like (DCL), Argonaute (AGO), and RNA-dependent RNA polymerase (RDR) are known as the three major gene families that act as the critical components of RNA interference or silencing mechanisms through the noncoding small RNA molecules (miRNA and siRNA) to regulate the expressions of protein-coding genes in eukaryotic organisms. However, most of their characteristics including structures, chromosomal location, subcellular locations, regulatory elements, and gene networking were not rigorously studied. Our analysis identified 7 TaDCL, 39 TaAGO, and 16 TaRDR genes as RNA interference (RNAi) genes from the wheat genome. Phylogenetic analysis of predicted RNAi proteins with the RNAi proteins of Arabidopsis and rice showed that the predicted proteins of TaDCL, TaAGO, and TaRDR groups are clustered into four, eight, and four subgroups, respectively. Domain, 3D protein structure, motif, and exon-intron structure analyses showed that these proteins conserve identical characteristics within groups and maintain differences between groups. The nonsynonymous/synonymous mutation ratio (Ka/Ks) < 1 suggested that these protein sequences conserve some purifying functions. RNAi genes networking with TFs revealed that ERF, MIKC-MADS, C2H2, BBR-BPC, MYB, and Dof are the key transcriptional regulators of the predicted RNAi-related genes. The cis-regulatory element (CREs) analysis detected some important CREs of RNAi genes that are significantly associated with light, stress, and hormone responses. Expression analysis based on an online database exhibited that almost all of the predicted RNAi genes are expressed in different tissues and organs. A case-control study from the gene expression level showed that some RNAi genes significantly responded to the drought and heat stresses. Overall results would therefore provide an excellent basis for in-depth molecular investigation of these genes and their regulatory elements for wheat crop improvement against different stressors.

Peripheral artery disease affects the function of the legs of claudicating patients in a diffuse manner irrespective of the segment of the arterial tree primarily involved.

Different levels of arterial occlusive disease (aortoiliac, femoropopliteal, multi-level disease) can produce claudication symptoms in different leg muscle groups (buttocks, thighs, calves) in patients with peripheral artery disease (PAD). We tested the hypothesis that different locations of occlusive disease uniquely affect the muscles of PAD legs and produce distinctive patterns in the way claudicating patients walk. Ninety-seven PAD patients and 35 healthy controls were recruited. PAD patients were categorized to aortoiliac, femoropopliteal and multi-level disease groups using computerized tomographic angiography. Subjects performed walking trials both pain-free and during claudication pain and joint kinematics, kinetics, and spatiotemporal parameters were calculated to evaluate the net contribution of the calf, thigh and buttock muscles. PAD patients with occlusive disease affecting different segments of the arterial tree (aortoiliac, femoropopliteal, multi-level disease) presented with symptoms affecting different muscle groups of the lower extremity (calves, thighs and buttocks alone or in combination). However, no significant biomechanical differences were found between PAD groups during the pain-free conditions with minimal differences between PAD groups in the claudicating state. All statistical differences in the pain-free condition occurred between healthy controls and one or more PAD groups. A discriminant analysis function was able to adequately predict if a subject was a control with over 70% accuracy, but the function was unable to differentiate between PAD groups. In-depth gait analyses of claudicating PAD patients indicate that different locations of arterial disease produce claudication symptoms that affect different muscle groups across the lower extremity but impact the function of the leg muscles in a diffuse manner generating similar walking impairments.

The Kuwait Stitch: A novel surgical technique for surgical wound closures.

There are several stitch techniques used for surgical wound closing. Each stitch has its own benefits and drawbacks that a surgeon must balance before use. In this paper, we highlight some of the more common techniques utilized in operative wound closure and briefly discuss benefits and caveats one must be aware of before using. The rest of our paper will focus on a new technique, the Kuwait Stitch, with instructions on how to perform the technique followed by a discussion on the benefits and indications for use of this stitch.

Incidence of typhoid and paratyphoid fever in Bangladesh, Nepal, and Pakistan: results of the Surveillance for Enteric Fever in Asia Project.

BACKGROUND: Precise enteric fever disease burden data are needed to inform prevention and control measures, including the use of newly available typhoid vaccines. We established the Surveillance for Enteric Fever in Asia Project (SEAP) to inform these strategies. METHODS: From September, 2016, to September, 2019, we conducted prospective clinical surveillance for Salmonella enterica serotype Typhi (S Typhi) and Paratyphi (S Paratyphi) A, B, and C at health facilities in predetermined catchment areas in Dhaka, Bangladesh; Kathmandu and Kavrepalanchok, Nepal; and Karachi, Pakistan. Patients eligible for inclusion were outpatients with 3 or more consecutive days of fever in the last 7 days; inpatients with suspected or confirmed enteric fever; patients with blood culture-confirmed enteric fever from the hospital laboratories not captured by inpatient or outpatient enrolment and cases from the laboratory network; and patients with non-traumatic ileal perforation under surgical care. We used a hybrid surveillance model, pairing facility-based blood culture surveillance with community surveys of health-care use. Blood cultures were performed for enrolled patients. We calculated overall and age-specific typhoid and paratyphoid incidence estimates for each study site. Adjusted estimates accounted for the sensitivity of blood culture, the proportion of eligible individuals who consented and provided blood, the probability of care-seeking at a study facility, and the influence of wealth and education on care-seeking. We additionally calculated incidence of hospitalisation due to typhoid and paratyphoid. FINDINGS: A total of 34 747 patients were enrolled across 23 facilitates (six tertiary hospitals, surgical wards of two additional hospitals, and 15 laboratory network sites) during the study period. Of the 34 303 blood cultures performed on enrolled patients, 8705 (26%) were positive for typhoidal Salmonella. Adjusted incidence rates of enteric fever considered patients in the six tertiary hospitals. Adjusted incidence of S Typhi, expressed per 100 000 person-years, was 913 (95% CI 765-1095) in Dhaka. In Nepal, the adjusted typhoid incidence rates were 330 (230-480) in Kathmandu and 268 (202-362) in Kavrepalanchok. In Pakistan, the adjusted incidence rates per hospital site were 176 (144-216) and 103 (85-126). The adjusted incidence rates of paratyphoid (of which all included cases were due to S Paratyphi A) were 128 (107-154) in Bangladesh, 46 (34-62) and 81 (56-118) in the Nepal sites, and 23 (19-29) and 1 (1-1) in the Pakistan sites. Adjusted incidence of hospitalisation was high across sites, and overall, 2804 (32%) of 8705 patients with blood culture-confirmed enteric fever were hospitalised. INTERPRETATION: Across diverse communities in three south Asian countries, adjusted incidence exceeded the threshold for "high burden" of enteric fever (100 per 100 000 person-years). Incidence was highest among children, although age patterns differed across sites. The substantial disease burden identified highlights the need for control measures, including improvements to water and sanitation infrastructure and the implementation of typhoid vaccines. FUNDING: Bill & Melinda Gates Foundation.