The South Asian region has the second highest risk of maternal death in the world. To prevent maternal deaths due to sepsis and to decrease the maternal mortality ratio as per the World Health Organization Millenium Development Goals, a better understanding of the etiology of endometritis and related sepsis is required. We describe microbiological laboratory methods used in the maternal Postpartum Sepsis Study, which was conducted in Bangladesh and Pakistan, two populous countries in South Asia.Postpartum maternal fever in the community was evaluated by a physician and blood and urine were collected for routine analysis and culture. If endometritis was suspected, an endometrial brush sample was collected in the hospital for aerobic and anaerobic culture and molecular detection of bacterial etiologic agents (previously identified and/or plausible).The results emanating from this study will provide microbiologic evidence of the etiology and susceptibility pattern of agents recovered from patients with postpartum fever in South Asia, data critical for the development of evidence-based algorithms for management of postpartum fever in the region. Puerperal pyrexia and post-partum puerperal (PP) sepsis, an infection acquired during the first 42 days postpartum, remain leading causes of maternal morbidity and mortality in South Asia . It is eThis paper describes the samples collected and diagnostic tests used to determine and characterize the bacterial etiologies of severe postpartum infections in study centers in Pakistan (Matiari and Karachi) and Bangladesh (Sylhet). Quality assurance procedures at each site are also described.The PP sepsis study is a prospective cohort study that expands on the Aetiology of Neonatal Infection in South Asia (ANISA) study in three sites of Sylhet in Bangladesh and Karachi and Matiari in Pakistan. Methods for field evaluation are described by Bartlett et al. . BrieflyThe study protocol was approved after full ethical review by the Johns Hopkins University, Baltimore, USA; International Centre for Diarrheal Disease Research, Bangladesh; the Bangladesh Institute for Child Health for the Child Health Research Foundation in Bangladesh, and the Aga Khan University, Karachi, Pakistan.Microbiological and other laboratory tests were carried out at research laboratories in Karachi, Sylhet, and Dhaka. The Infectious Disease Research Laboratory (IDRL) at the Aga Khan University received and processed specimens from the Karachi and Matiari field sites, whereas, the Child Health Research Foundation (CHRF) Microbiology Laboratory at the Dhaka Shishu Hospital received specimens from the Sylhet site.The WHO advises culture of midstream urine specimens and blood for evaluation of the cause of post-partum maternal fever . TransvaIn keeping with these recommendations, we collected the following specimens to evaluate the cause of postpartum fever, following informed consent.Blood was collected for complete blood count, culture, and to determine the presence or absence of malarial parasites. The purpose of each of these tests was unique and has been described in Table To evaluate for leukocytosis and leukopenia as possible markers of sepsis , blood cThin and thick blood smears were made and examined at central laboratories to identify malarial parasites and to quantify and speciate Plasmodia, if present.Aerobic and anaerobic blood cultures were performed in the BACTEC 9240® instrument with BACTEC Aerobic Plus® and Anaerobic F Lytic bottles. Five to ten ml of blood was collected in each bottle and transported to respective central laboratories for incubation and processing. Those flagged positive by the instrument were removed and gram stain and agar subculture was performed.Endometrial tissue was collected using Tao sampling brushes by Cook Medical . SterileS.aureus and bacitracin disk and Lancefield grouping for S.pyogenes and S.agalactiae. Commercial systems such as API Strep and API-NH (BioMerieux) were used for fastidious organisms, such as alpha haemolytic, nutritionally variant Streptococci, and Haemophilus spp.One half of the brush was aseptically cut and placed into a thioglycolate broth medium with cooked meat to facilitate recovery of strict anaerobes. This tube was transported to the central laboratory at 25 °C within 6 h of collection. Upon receipt in the laboratory, the broth was vortexed at high speed to dislodge tissue from the brush and used to inoculate chocolate, MacConkey, and 5 % sheep blood agar (SBA) media. To recover strict anaerobes, pre-reduced Brucella agar with kanamycin and vancomycin supplements was inoculated with the thioglycolate broth . Agar inThe other half of the endometrial brush was introduced into a Universal Transport Medium (UTM) tube (Copan) and transported to the laboratory at 2–8 °C.Chlamydia trachomatis, Mycoplasma spp, and Ureaplasma spp was detected by molecular methods. For specimens collected in Pakistan, the Sacace Biotechnologies  kit [C.trachomatis, M.genitalium, M.hominis, and U.urealyticum at Aga Khan University’s Infectious Disease Research Laboratory. Endometrial specimens obtained in Bangladesh were shipped to University of Alabama (UAB) for culture of mycoplasma and ureaplasma and PCR detection for Mycoplasma and Ureaplasma using previously published methods [The presence of aly) kit  was used methods .In Pakistan, high vaginal swabs were collected after endometrial brush specimens in an attempt to exclude vaginal contaminating flora and to evaluate for presence of vaginitis on the basis of presence or absence of clue cells. Samples were collected from the high vaginal wall in Ames transport medium  and sent to the laboratory. Samples were plated on chocolate agar, Gardnerella agar (with 5 % human blood), Maconkey agar, and Sabouraud’s agar. Results were recorded semi-quantitatively as light, medium, or heavy growth.Clean catch urine was collected to determine the etiology of urinary tract infection (UTI) in patients with likely UTI (significant growth of uropathogen with positive nitrite and leukocyte esterase on dipstick test) .3 colony-forming unit (CFU) of bacteria per mL of urine were considered significant. Identification of uropathogens was carried out using either the minimal sulfide-indole-motility-citrate-urease-triple sugar iron (SIMCUT) method or commercial API (BioMerieux) strips.Dipstick tests, indicating the presence or absence of both leukocyte esterase and nitrite  were perS.aureus, vancomycin MICs were performed by the broth dilution method) [Susceptibility testing by the disk diffusion method was carried out only for aerobic or facultatively anaerobic pathogens in accordance with CLSI guidelines. Where disk diffusion was not the preferred method, broth dilution minimum inhibitory concentrations (MICs) were carried out to confirm susceptibility (eg for methicillin resistant  method) .At the Karachi site, dengue IgM and Rapid ICT malaria (PanBio) tests were additionally performed to detect dengue fever, and to improve detection of submicroscopic malaria infections. The city suffered major dengue outbreaks since 1994 , and conThe Pakistan site research laboratory regularly undergoes biannual proficiency testing (PT) for bacteriologic culture and identification and molecular detection of respiratory pathogens by the College of American Pathologists (CAP). The laboratory achieved 98 % satisfactory results for culture and identification and 95 % satisfactory scores for molecular detection and identification of pathogens in 2013.At the beginning of the study, the lead technologist at the Bangladesh site visited Johns Hopkins Medical Institutions clinical microbiology laboratory to review procedures and for additional specialized training. Research laboratories in Sylhet and Dhaka carried out isolation and identification and sent 10 % of bacterial pathogens isolated from blood to Infectious Disease Research Laboratory in Karachi to confirm identification.Procedures for and results of laboratory tests have been reviewed by experienced and certified clinical microbiologists at both sites (SS and MER). Results were also reviewed mid-study in November 2013 and at end-of-study in November 2014 at the PP sepsis Technical Advisory Group Meeting.Although there were minor variations in test menus (ancillary tests carried out for Karachi sites) and differences in quality assurance procedures , tests at either site laboratory were carried out using the same Standard Operation Procedures (SOPs) developed by SS and MER and reviewed by other members of the team.In implementing protocols and carrying out tests that had to be reported to physicians in time, the role of the laboratory in reporting timely and reliable results cannot be overemphasized. Laboratory managers played key roles in contacting study physicians and principal investigators with timely results. Among the challenges faced were difficult maintenance of transport temperatures of samples , transport of laboratory samples over long distances, and timely transport. Upon interim review of laboratory data, it was noted that earlier processing of cultures resulted in fewer contaminants and desirable colony counts. Thus, management of employee shifts to adjust culture processing times from 4 to 6 h lags to immediate processing was carried out.In conclusion, laboratory protocols for the etiology of postpartum fever study, as described, performed smoothly and efficiently despite challenges of implementation. Routine collection of endometrial samples in the community must be complemented by efficient laboratory services for optimal recovery of pathogens.


Provision and scale-up of high quality, evidence-based services is essential for successful international HIV prevention interventions in order to generate and maintain intervention uptake, study integrity and participant trust, from both health service delivery and diplomatic perspectives.We developed quality assurance (QAC) procedures to evaluate staff fidelity to a cluster-randomized trial of the NIMH Project Accept (HPTN 043) assessing the effectiveness of a community-based voluntary counseling and testing strategy. The intervention was comprised of three components—Mobile Voluntary Counseling and Testing (MVCT), Community Mobilization (CM) and Post-Test Support Services (PTSS). QAC procedures were based on standardized criteria, and were designed to assess both provider skills and adherence to the intervention protocol. Supervisors observed a random sample of 5% to 10% of sessions each month and evaluated staff against multiple criteria on scales of 1–5. A score of 5 indicated 100% adherence, 4 indicated 95% adherence, and 3 indicated 90% adherence. Scores below 3 were considered unsatisfactory, and protocol deviations were discussed with the respective staff.During the first year of the intervention, the mean scores of MVCT and CM staff across the 5 study sites were 4 (95% adherence) or greater and continued to improve over time. Mean QAC scores for the PTSS component were lower and displayed greater fluctuations. Challenges to PTSS staff were identified as coping with the wide range of activities in the PTSS component and the novelty of the PTSS process. QAC fluctuations for PTSS were also associated with new staff hires or changes in staff responsibilities. Through constant staff monitoring and support, by Year 2, QAC scores for PTSS activities had reached those of MVCT and CM.The implementation of a large-sale, evidence based HIV intervention requires extensive QAC to ensure implementation effectiveness. Ongoing appraisal of study staff across sites ensures consistent and high quality delivery of all intervention components, in keeping with the goals of the study protocol, while also providing a forum for corrective feedback, additional supervision and retraining of staff. QAC ensures staff fidelity to study procedures and is critical to the successful delivery of multi-site HIV prevention interventions, as well as the delivery of services scaled up in programmatic situations. The heightened profile of global health efforts in the international relations and political spheres in turn requires enhanced efforts to ensure that programs are delivered in the most adaptable, appropriate and diplomatic manner manner. In this context, there remains no more severe global health crisis in the world today than the HIV epidemic in sub-Saharan Africa. There are approximately 25 million people living with HIV in sub-Saharan Africa, with 1.6 million new infections occurring in 2012 alone... Efforts QAC roles and responsibilities at the site level are presented in For project-specific QAC measures to be effectively interpreted and generalized, it is essential that reliability and validity of related scales be considered. In the case of Project Accept, measurement of key constructs (e.g. PTSS protocol adherence) was validated during QAC tool development, based on a collaborative process with both service delivery staff and the input and approval of clients . For exaThroughout the intervention, a random sample of VCT, CM and PTSS sessions were selected for QAC evaluation by team leaders. For VCT, during the first 6 months of the intervention 10% of counseling sessions were evaluated, with a minimum of 2 evaluations per counselor per month. Thereafter, 5% of sessions delivered per month were evaluated with a minimum of 1 evaluation per counselor per month. For CM, during the first six months 15% of outreach sessions were evaluated, with a minimum of 2 evaluations per CM staff member per month. Thereafter, 5% of outreach sessions were evaluated, with a minimum of 1 evaluation per CM staff member per month. For PTSS, during the first 6 months of the intervention all CET and stigma reduction sessions, 2 information sharing sessions, 20% of support sessions and 15% of crisis counseling sessions were evaluated. Thereafter, 1 CET session, 1 stigma reduction session, 4 information sharing sessions, 5% of support sessions and 5% of crisis counseling sessions were evaluated.For the VCT component, VCT team leaders assessed counselors’ adherence and skill levels in 10 essential component areas, including (1) general counselor skills in keeping with the client-centered, personalized risk reduction model recommended by the U.S. Centers for Disease Control and Prevention (CDC) and World Health Organization (WHO) and (2) For the CM component, CM team leaders assessed outreach workers’ adherence and skill levels for 10 essential areas of community interaction and 6 areas of community referral and follow-up. These included (1) basic interaction skills, (2) provision of accurate information, and (3) referrals to further care. Interaction skills included empathy, being non-judgmental, maintaining appropriate boundaries, and maintaining session cohesion. In addition to adherence to manualized guidelines, CM activities were also rated, using the same scales, on the skill with which the staff member delivered the session. In particular, community mobilization staff were assessed on their success in adapting related activities to observe local social, cultural and religious norms, thereby For PTSS, team leaders assessed facilitators’ adherence and skill levels in individual crisis counseling, group information sessions, coping effectiveness training, and stigma reduction training (each with approximately 12 activity or skills criteria). Staff were evaluated on (1) basic support service delivery skills, (2) skills in guiding goal-setting, and (3) adherence to the PTSS curriculum. PTSS activities were evaluated and scored separately. As with CM and VCT assessment procedures, PTSS staff and service delivery was assessed with reference to acceptable standards of service delivery from the diplomatic perspective, including the provision of long-term, sustainable and effective coping strategies for patients diagnosed with HIV.For each session, project staff were informed in advance of QAC assessment in advance. While this generated a risk that QAC scores would be higher when staff were aware that the specific sessions were being assessed, given staffing and spatial constraints, alternative blinded or covert assessment systems were considered unfeasible by the Intervention Core. Team leaders then recorded the relevant QAC score for each evaluation criterion on the appropriate QAC form. Evaluation criteria were thematically grouped by essential component area, and the average score for each group was recorded at the end of each section. Individual item scores within an area were averaged to create a summary score and those summary scores were again averaged for an overall QAC score. At the end of the form, a single overall score for the entire session was calculated based on an average of these scores. For each intervention component, these scores were then aggregated and averaged to generate a single QAC score for VCT, CM, and PTSS each month. At the end of each month, all QAC records for each intervention component across sites were transmitted to the intervention coordinating center via scanned, security-protected e-mail documents. These reports were accompanied by a narrative report from the site project director describing QAC activity over the reporting period. Receipt of QAC data was acknowledged, entered into a password-protected Microsoft Access database, and cross-checked through Microsoft Excel.QAC performance charts across intervention components were produced for each study site by the intervention coordinating center and returned to site project directors with commentary and questions on performance on a monthly basis. Feedback reports also described emerging QAC issues across study sites in order to maintain communication and interactive learning between sites. Study sites were encouraged to use these reports as a training tool to focus attention on specific QAC issues, provide early feedback, and foster preventive measures. QAC data was also presented and examined on a monthly basis with US and site-based principal investigators via conference calls with the study steering committee.Also at the site level, feedback mechanisms included (1) weekly field staff meetings and (2) periodical written feedback to field staff by team leaders, component coordinators and project directors. QAC issues were also discussed at component-specific staff meetings, study-wide staff meetings, and in one-on-one meetings with project staff. The frequency and structure of these meetings was left to the discretion of the site team leaders and coordinators, who were judged to have the best assessment of staff needs. Team leaders and component coordinators used the QAC scores to (1) identify strengths and weaknesses of individual staff and overall areas and components of the intervention and (2) create action plans for improvement, retraining, or more frequent supervision and support. Staff failing to meet minimum QAC requirements were retrained and only allowed to resume their role after conducting 2 observed and evaluated sessions rated at 90% adherence or higher.In addition, all study sites received support and monitoring visits from the intervention coordinating center at UCSF and designated National Institutes of Mental Health (NIMH) staff. In this context, monitors accessed and inspected study facilities and documentation, as well as observing the performance of study procedures. In addition, the intervention coordination director visited each study site bi-annually to evaluate the quality and consistency of the implementation of the intervention. These visits allowed firsthand observation of the intervention components and maintained open relations with field staff. Study site visits generally took one week to conduct and included: (1) Observing sessions and rating performance using the QC evaluation form; (2) reviewing files and forms for completeness and accuracy; (3) observing daily operations; (4) ensuring that the intervention was being implemented as prescribed in the Protocol and SOP guidelines; and (5) individual and group meetings with the site teams  to provide direct supervision, feedback and answer questions. Study site visits were supported by monthly conference calls with each site.QAC scores across all components are presented in CM QAC scores are presented in PTSS QAC scores are presented in Project Directors identified challenges experienced by study staff as (1) coping with the wide range of field activities and (2) the novelty of the CBVCT intervention. QAC score fluctuations were also associated with new staff hires or changes in staff responsibilities. The major effort put into developing and implementing the in-depth quality-assurance methodology has resulted in essential information on the successes and challenges of implementing this complex intervention. The overall 95% adherence to essential intervention delivery components indicates that the wide range of activities contained within the intervention can be successfully and faithfully implemented in resource limited settings. The QAC process has also been able to highlight challenges to implementation, especially around PTSS activities, and provided solutions to those challenges, primarily in the early identification of enhanced training needs. QAC data also serves an important purpose by monitoring consistency of component implementation. Of note, the application of feedback systems  were considered to be highly effective in remedying QAVC issues, though these could not be scientifically linked to changes in QAC results at the site level.While the goal of QAC is to identify and correct non-adherence to the study protocol, it is also important to acknowledge elements that were performed particularly well and include these in the summaries as a way of motivating study staff. It is important that staff be given positive reinforcement in the areas where they are doing well and to be acknowledged for their valuable contribution to the project and to the team. These QAC procedures allow for the identification of skills training needs for VCT, PTSS, and CM staff, and relevant in-service skills trainings are discussed with the individual staff. In particular, VCT has been scaled up through-out African countries although very few report any attempts at QAC. Our experience in this large prevention trial is that such QAC is both (1) feasible and acceptable to both staff and participants and (2) associated with higher quality programmatic implementation.The “diplomatic” QAC model presented under the Project Accept aegis is not without limitations. Given different social contexts, it may entirely appropriate to have greater flexibility across sites, as the present study did with permitting site team leaders and coordinators to set the frequency and structure of meetings to address QAC issues. However, this also introduces variability in how QAC issues are addressed, which could have some bearing on actual QAC across sites . Addressing some of these complexities and the need for additional research on how to assure QAC before implementing multi-site, multi-level interventions and assuring QAC once in the field remain key tasks for future related interventions. A further limitation to these results relates to the presentation of mean fidelity scores across the sites without associated data on competencies and specific adherence challenges or that seemed to explain fluctuations over time and differences across contexts. Such information, given the range of components and geographical areas covered by the current manuscript, was considered to be beyond the scope of this paper. Finally, there are numerous factors in addition to QAC that influence fidelity and the ultimate success of interventions and implementation strategies: although QAC is likely a useful tool for supporting and promoting high fidelity when HIV prevention interventions are implemented, one cannot make definitive causal statements in this regard.The implementation of a large-sale, evidence based HIV intervention requires extensive QAC to ensure implementation effectiveness, building on a ‘feedback loop’ whereby improvements in service delivery become both an iterative and an intuitive process. During Project Accept, ongoing appraisal of study staff by intervention component across sites helped to ensure consistent and high quality delivery of all intervention components, in keeping with the goals of the study protocol. In addition, for the first time in any kind of global health intervention, attempts were made to assess standards of service delivery from a diplomatic perspective. The development of the field of global health diplomacy has helped to drive recognition of the importance of culturally appropriate and culturally sensitive global health interventions, which both international and domestic Project Accept QAC staff monitored on a routine basis throughout the intervention. The primary findings of Project Accept also suggest that a community-wide multicomponent intervention of mobilization, HIV testing and support services can be both safe and feasible and can S1 Fig(XLS)Click here for additional data file.S2 Fig(XLS)Click here for additional data file.S3 Fig(XLS)Click here for additional data file.S4 Fig(XLS)Click here for additional data file.S5 Fig(XLS)Click here for additional data file.


In light of the unprecedented momentum to control or eliminate ten neglected tropical diseases (NTDs) by 2020, the NTD community is at a crossroads of opportunity. Efforts to eliminate lymphatic filariasis (LF) using mass drug administration (MDA) platforms have dramatically and simultaneously increased the number of individuals treated for soil-transmitted helminths (STHs) in many areas, owing to overlaps in endemicity and drug efficacy. A concomitant reduction in STH infection raises the possibility of moving beyond current World Health Organization (WHO) morbidity control guidelines to actual disease elimination for STHs . We idenAscaris lumbricoides), whipworm (Trichuris trichiura), and hookworm species . STHs are transmitted when infected persons shed eggs in feces that then contaminate soil or water and are ingested by or penetrate the skin of others. An individual’s worm burden, or the intensity of their infection, is directly related to their risk of STH-associated morbidity. Moderate to heavy STH infections are associated with diarrhea, malnutrition, chronic malaise, impaired cognitive development, disrupted linear growth, chronic inflammation, and anemia [Over 1,500,000,000 people globally are infected with STHs, including roundworm  or mebendazole, often donated to endemic countries by the pharmaceutical industry. Target populations include pre-school age children (PSAC), school age children (SAC), and women of childbearing age, including pregnant women in their second and third trimesters. The WHO strategy for SAC recommends that MDA programs target SAC annually where prevalence is 20%–50% and biannually where prevalence is over 50%. These programs are primarily delivered using school-based platforms with teachers serving as drug distributors. In alignment with the WHO NTD Roadmap, this sustained control strategy aims to decrease worm burden and reduce STH-associated morbidity by ensuring that 100% of endemic countries are reaching PSAC and SAC with at least 75% coverage by 2020 .Ascaris and Trichuris worms are harbored in adult populations [However, in 2013, only 39% of SAC and 49% of PSAC in need of treatment were reached with MDA, and only 27% of endemic countries currently attain London Declaration coverage targets [ulations –7. Recenulations .Wuchereria bancrofti, Brugia malayi, and Brugia timori species) is a disease caused by mosquito-transmitted parasitic worms that damage the human lymph system, resulting, most recognizably, in lymphedema, elephantiasis, and hydrocele. The WHO strategy for LF recommends annual MDA of ALB and either ivermectin or diethylcarbamazine (DEC) to all individuals living in endemic areas beginning at two years of age (for the ALB and DEC regimen) or five years of age (for ALB and ivermectin). Following four to six annual rounds of MDA, the density of circulating microfilaria in infected individuals (and thus the community-level force of infection) may be reduced and LF transmission disrupted.A potential strategy for broadening the reach of STH programs is to leverage the existing community-based infrastructure of LF elimination programs. LF  and success of LF programs may be a unique opportunity to facilitate a smooth transition to a community-based STH platform. At the same time, LF programs that have achieved post-MDA surveillance status will continue to benefit from an ongoing infrastructure to facilitate the surveillance activities necessary for monitoring potential recrudescence or collecting data for WHO elimination dossiers. There are a number of potential challenges to integrating LF and STH platforms. Most notably, utilizing community-based LF platforms to serve as the primary method to deliver STH services will require political as well as logistical changes. Where STH programs are primarily administered by ministries of education, there will need to be close coordination with ministries of health. Methodological innovations will also need to accompany the proposed programmatic shift to ensure that coordinated sampling methodologies are validated and field deployable diagnostics are available for relevant coordinated surveys. This presents a major challenge that will require technologies and processes to enable the simultaneous sampling, storage, and testing of fecal and serum specimens.The design of relevant field studies will need to account for complex issues such as randomization of communities on various LF elimination trajectories and with varying access to co-interventions such as water and sanitation activities. Hybrid trials that blend efficacy and effectiveness  and implementation research studies will be necessary for implementing simultaneous process evaluations that provide context for clinical findings and inform future guidelines. Perhaps the largest undertaking on the research agenda will be defining, testing, and validating STH transmission breakpoints. Meanwhile, ministries of health will be challenged by partnering with researchers to facilitate the intensified trials in select areas while continuing STH control policies broadly.In light of these research needs, the Bill & Melinda Gates Foundation recently launched the DeWorm3 Project in partnership with the Natural History Museum (NHM) in London. The purpose of DeWorm3 is to test the feasibility of interrupting transmission of STHs by leveraging existing LF elimination efforts. The project will require cluster randomized trials of a number of intensified STH interventions, including community-wide MDA, cost-effectiveness analyses, qualitative studies of compliance and stakeholder satisfaction, mathematical modeling exercises, drug resistance monitoring, and a number of other research initiatives that explore the unique opportunity to innovatively transition from STH control to elimination strategies.The prospect of leveraging LF platforms to benefit STH programs in co-endemic areas has been promoted since the initial establishment of the Global Alliance to Eliminate Lymphatic Filariasis (GAELF) in 2000 . As an i


RGD maintains and updates datasets for genomic elements such as genes, transcripts and increasingly in recent years, sequence variations, as well as map positions for multiple assemblies and sequence information. Functional annotations for genomic elements are curated from published literature, submitted by researchers and integrated from other public resources. Complementing the genomic data catalogs are those associated with phenotypes and disease, including strains, QTL and experimental phenotype measurements across hundreds of strains. Data are submitted by researchers, acquired through bulk data pipelines or curated from published literature. Innovative software tools provide users with an integrated platform to query, mine, display and analyze valuable genomic and phenomic datasets for discovery and enhancement of their own research. This update highlights recent developments that reflect an increasing focus on: (i) genomic variation, (ii) phenotypes and diseases, (iii) data related to the environment and experimental conditions and (iv) datasets and software tools that allow the user to explore and analyze the interactions among these and their impact on disease.The Rat Genome Database (RGD,  The Rat Genome Database (RGD) has been the premier resource for rat genetic, genomic and phenotypic data since its creation in 1999. The rat is an important animal model for pharmacology, toxicology, physiology and pathology  and Ensembl (http://www.ensembl.org/index.html). These include tracks from similar substrains, allowing users to compare results from different platforms and analysis systems. Variant density plots are available for larger regions as well as individual variation tracks with popups outlining details for a variant such as depth, conservation, location, zygosity and prediction of effect  and automated pipelines incorporate variants identified as clinically important from ClinVar (http://www.ncbi.nlm.nih.gov/clinvar/). Rat gene report pages provide links to lists of variants reported in various strains as well as one-click access to those data in the Variant Visualizer (described below), while human gene reports detail clinically significant variants with links to individual variant reports . Adjustments to QTL and strain reports and the creation of SNP report pages accommodate the increasing use of SNPs as position markers for identification of QTL and the generation of congenic strains.RGD has expanded its capacity to accommodate the ever-increasing amount of data generated by researchers using next-generation sequencing technologies and has incorporated copy number variations, single nucleotide polymorphisms (SNPs), indels and mutations into its reports and tools. To provide timely access to strain-specific variants generated from whole genome sequencing (WGS) projects, RGD developed a pipeline for data submitted by researchers for display on its Genome Browser and to date this has resulted in 40 single nucleotide variation (SNV) tracks, nine indel tracks, a copy number variation track and tracks for variation data from dbSNP   provides a genome-wide view of mapped genomic elements such as genes, QTL and congenic strains related to simple and complex ontology searches. Users can also layer on additional data returned from their initial searches using data types or ontology terms. With the re-annotation of QTL using ontologies for strain, trait and experimental data and the inclusion of gene–drug/chemical interactions in RGD, users can now create Boolean searches with terms from the following: Gene Ontology , MEDIC dThe Gene Annotator or GA Tool is a one-stop functional analysis tool for rat, human and mouse genes. For any of the three organisms, users can upload a list of common identifiers from RGD, EntrezGene, GenBank, Ensembl and Affymetrix or search by a chromosomal region or functional ontology identifiers to retrieve comprehensive reports for each gene. The output  and driven off the RGD Oracle database. The tools can be run on any Java container that implements the Java Servlet and JSP (JavaServer Pages) specification. The popular Spring (All tools mentioned are built in-house by RGD developers except GBrowse . RGD toor Spring  frameworftp://rgd.mcw.edu/pub/). All of the data found on the RGD website can be downloaded in bulk from the FTP site. Files available for download include descriptive information for genes, QTL and strains, functional gene annotations, SSLPs, SNPs, ontology term files and more. Bulk downloads of data allow researchers to perform analyses with their own software tools or other tools that are not available at RGD.An important avenue for RGD data access is the FTP site (RGD continues to acquire, curate and integrate data of critical importance to disease research communities. These include datasets associated with the interacting factors in disease processes—genomic variation, environment, molecular pathways and the resulting phenotypes that define disease. Increasing emphasis on the development of tools that integrate, analyze and visualize multiple types of data from rat, human and mouse studies has made RGD a comprehensive research platform for disease and cross-species investigations.Supplementary Data are available at NAR Online.


Multiple regression adjusted by the covariates participants’ age and education level showed a positive association between odour identification and the cognitive inhibition response . The odour discrimination capacity was not a predictor of the cognitive executive performance. Our results suggest that both hot and cold executive functions seem to be associated with higher-order olfactory functioning in humans. These results robustly support the hypothesis that olfaction and executive measures have a common neural substrate in PFC and OFC, and suggest that olfaction might be a reliable cognitive marker in psychiatric and neurologic disorders.The prefrontal (PFC) and orbitofrontal cortex (OFC) appear to be associated with both executive functions and olfaction. However, there is little data relating olfactory processing and executive functions in humans. The present study aimed at exploring the role of olfaction on executive functioning, making a distinction between primary and more cognitive aspects of olfaction. Three executive tasks of similar difficulty were used. One was used to assess hot executive functions (Iowa Gambling Task-IGT), and two as a measure of cold executive functioning (Stroop Colour and Word Test-SCWT and Wisconsin Card Sorting Test-WCST). Sixty two healthy participants were included: 31 with normosmia and 31 with hyposmia. Olfactory abilities were assessed using the ‘‘Sniffin’ Sticks’’ test and the olfactory threshold, odour discrimination and odour identification measures were obtained. All participants were female, aged between 18 and 60. Results showed that participants with hyposmia displayed worse performance in decision making (IGT; Cohen’s- The olfactory system is directly associated with brain areas involved in cognitive and emotional processes and the evidence of an association between olfactory and some cognitive functions is strong –4. PreviThe main components of olfaction are odour threshold (OT), odour identification (OI) and odour discrimination (OD) . OT is mHowever, the relation between olfaction with more complex cognitive functions such as executive functions has not been properly addressed, despite the overlap between their brain circuits. Executive functions (EF) are higher order cognitive capacities that allow people to orient towards the future, display self-control and effectively have goal oriented behaviour . EF can Nevertheless, and in spite of this interesting hypothesis, there are no studies linking the different components of olfaction and EF in healthy young population, demonstrating the physiological role of olfaction on higher cognitive functions in humans. This study aimed to assess the association between olfactory functioning and executive functions while evaluating hot and cold executive functions in healthy subjects with hyposmia and normosmia. We made a distinction between primary and more cognitive aspects of olfaction and expected an association between executive processing and the higher-order components of olfaction, mainly the identification capacity.All participants were informed about the research procedures and gave informed consent in writing. Procedures were approved by the Ethical Committee of each of the following institutions: University Hospital of Bellvitge-IDIBELL; University Hospital of Santiago (Santiago de Compostela); Clinic University Hospital Virgen de Victoria, Málaga; University of Navarra, Pamplona; Biomedical Research Institute of Girona ; IMIM , the Department of Endocrinology at the University Hospital of Santiago (Santiago de Compostela); the Department of Diabetes, Endocrinology and Nutrition ; the Department of Endocrinology and Nutrition ; the Diabetes, Endocrinology and Nutrition Department, Biomedical Research Institute of Girona ; the Human Pharmacology and Clinical Neurosciences Research Group at IMIM  and the Department of Basic Psychology, Clinic and Psychobiology .Sixty two consecutive healthy subjects (n = 62) were included. 31 had normosmia and 31 had hyposmia that was detected in the TDI-score of the Sniffin’ Sticks . All parThe exclusion criteria were: (1) History of chronic medical illness or neurological condition that might affect cognitive function and/or olfactory functioning; (2) Head trauma with loss of consciousness for more than 2 min, intellectual disability; (3) Use of psychoactive medication or drugs (4) Being male; (5) Age under 18 or over 60 ; (6) Having diabetes type I or II. The lifetime history of health or mental illnesses profile was based on the general health questionnaire GHQ-28.As described in previous studies ,23, all Wisconsin Card Sorting Test : This isStroop Color and Word Test : This paIowa Gambling Task : This coOT, OD and OI were investigated using the Sniffin’ Sticks . A trainOlfactory threshold test: Three pens were presented in a randomized order, two contained odourless samples and the third contained an odorant sample. The task of the subject was to indicate the pen with the odorant. Concentration was augmented if the subject chose an odourless pen and reduced if the correct pen was recognized twice. A total of 16 odour concentrations were tested. The mean of the last four of seven trials was used, ranging from 1 to 16. The higher the score the higher the olfactory capacity.Olfactory discrimination test: The subjects were asked to discriminate between 16 triplets of odours. In each group 2 odours were identical and 1 odour was different and the task was to recognize the odour that was different. The total score was the sum of correct responses ranging from 0 to 16. Higher scores were indicative of a better discrimination capacity.Olfactory identification test: A pen with an odour was presented to the subject. The pens contain common fragrances, such as peppermint, orange, leather, cinnamon, banana, garlic, lemon, rose, coffee, apple, clove, pineapple, aniseed and fish. Subjects were asked to identify the odour, by choosing it in a card listing four terms, only one of which correctly identified the odour. For this purpose the eye mask was removed (only for reading the card). The total score ranged from 0 to 16, with higher scores indicating a better identification capacity.TDI-score: The sum of the scores from the three subtests resulted in the TDI-score  with a maximum of 48 points. As defined in , a scoreAnalyses were carried out with SPSS20 for Windows. First, T-TEST proceduresd compared the mean scores of the cognitive measures between participants into the normosmia and hyposmia groups. To control the potential inflation of Type-I error due to multiple statistical comparisons (one for each cognition measure) and since Bonferroni's correction method has largely been criticized for being too conservative, an alternative procedure was considered: Simes’ procedure , an imprNext, multiple regression analyses valued the contribution of olfactory scores on the cognitive measures. These regressions were modelled in two steps: first step entered and fixed the covariates participants’ age and years of education, and second step added the independent variables odour discrimination and odour identification.This study set out to examine the association between different components of olfaction and executive functions in humans. A significant association was observed between higher-order component of olfaction (identification) and cognitive flexibility, inhibition response and decision making. Our results suggested that subjects with normosmia showed a better capacity of set shifting and less sensibility to immediate reward (IGT) compared with subjects with hyposmia. Although olfactory modulation was previously related to cognitive performance in humans –4, this The main result of our study is the differentiation between the different components of olfaction on their effects on the executive functioning. According to our results only the identification component is linked to inhibition response cognitive flexibility and decision making. These results are in line with those showing that the ability to identify odours is linked to prefrontal, particularly OFC functioning , brain rSpecifically, the association between the odour identification capacity and the IGT performance, found by means of the regression analysis, is in line with the hypothesis of the OFC implication in both processes. The IGT is a classic neuropsychological task for measuring the capacity of making decision and the sensibility to immediate reward. Different profiles in IGT performance were found between groups: Subjects with normosmia performed better and learned to keep away from not advantageous decks, while the performance of hyposmia participants did not improve along the task. This group went for choices that result in elevated immediate gains despite important future losses. Thus, subjects with lower olfactory capacity have an incapacity to successfully regulate reward and punishment, which might be translated into deficit in planning capacity. The cognitive mechanism underlying the decision making performance in these subjects might be associated with an elevated level of impulsivity. It has been established that impulsive subjects have marked limitations for learning suitable associations between reward and punishment ,33. As aOur results are also in line with neuroimaging studies supporting the hypothesis of the PFC and OFC as common neural substrates of olfaction and executive functioning, although a direct association cannot be addressed considering that we did not use neuroimaging techniques ,35. TakeOur study has several important strengths including the relatively large sample size of healthy young subjects with hyposmia. Additionally, we have used a standardized method for testing the olfactory capacity differentiating between three components of olfaction. Most of the previous studies on the topic had been conducted in psychiatric or neurologic populations using cognitive tasks that measure memory or attention. Conversely, our study was specifically designed to test the executive functioning, by using three well validated executive tests.However, the results of this study should be interpreted within the context of some limitations. First, only females were included in the study, thus the results are not applicable to males. Further studies including both males and females should be desirable. Second, measures of intelligence quotient, which is an important variable to take into account in cognitive studies, were not included. Nevertheless, as a cognitive level measure, years of education, has been considered, and no significant differences were observed between experimental groups. Third, neuroimaging data was not collected, thus future studies employing these techniques are desirable in order to confirm a common neural substrate for olfactory and executive functioning. Finally, future studies should also consider including additional executive tasks  in order to shed more light on their interactions and possible brain mechanisms.To summarize, this study demonstrates that in humans, the olfactory capacity plays an important role on prefrontal-dependent cognitive functions, almost certainly by common cerebral circuits. This study might have implications in the research of the executive profile of psychiatric and neurologic disorders, including abnormal eating behaviours  and schi


Zea mays) has been well-characterized, but the impact on root systems and the rhizosphere remain unclear. The transition from wild ecosystems to modern agriculture has focused on selecting traits that yielded the largest aboveground production with increasing levels of crop management and nutrient inputs. Root morphology, anatomy, and ecophysiological processes may have been affected by the substantial environmental and genetic shifts associated with this transition. As a result, root and rhizosphere traits that allow more efficient foraging and uptake in lower synthetic input environments might have been lost. The development of modern maize has led to a shift in microbiome community composition, but questions remain as to the dynamics and drivers of this change during maize evolution and its implications for resource acquisition and agroecosystem functioning under different management practices. Better understanding of how domestication and breeding affected root and rhizosphere microbial traits could inform breeding strategies, facilitate the sourcing of favorable alleles, and open new frontiers to improve resource use efficiency through greater integration of root development and ecophysiology with agroecosystem functioning.The effect of domestication and modern breeding on aboveground traits in maize ( Zea mays ssp. parviglumis), the ancestor of modern maize, originates from a mountainous environment with seasonal nutrient fluxes and high interspecific competition with diverse deciduous trees, grasses, and annual dicots  and phosphorus (P), contribute to eutrophication, climate forcing, and loss of biodiversity, with serious impacts on human and ecosystem health , environment-driven (E), and genotype-by-environment (G × E) mechanisms and highlight research gaps to be addressed in the future.The suite of traits selected during the domestication of crop wild relatives to increase yield or facilitate agronomic cultivation are collectively described as “domestication syndrome” Hammer, . Domestiteosinte branched1 (tb1) and barren stalk1 (ba1) , accounting for the naked kernels , controlling kernel row regularity  has fewer seminal roots, possibly related to smaller seed size, but a greater number of narrower, shorter, more branched nodal roots, which may be beneficial for early P acquisition  Lynch,  must be Rhizosphere functioning is shaped by the combined influence of host genotype (G), soil environment as affected by agroecosystem management and inherent soil characteristics (E), and their interaction (G × E). Shifts in these determinants during the course of maize evolution resulted in profoundly different selection environments that may have altered ecological functions of the rhizosphere, with consequences for foraging and acquisition of soil resources Figure .Maize genotypes differ in recruitment ability, resource provision, and responsiveness to beneficial rhizosphere microorganisms  differs from that of its wild ancestors in function as well as diversity, with genes affecting host-microbe interactions showing evidence of positive selection  have higher rhizobacterial diversity than their wild ancestor L. serriola, but diversity indices do not differ significantly between L. sativa cultivars  tends to be higher in landraces than either wild ancestors or modern cultivars  and tolerance of high planting density. Pinpointing the loss of favorable traits on an evolutionary timeline can help identify germplasm for use in creating new, resource-efficient varieties suited to low-input systems. For instance, QTL mapping of teosinte × maize crosses has been used to increase aerenchyma formation  decreased as a result of compositional changes, or has it been maintained by functional redundancy in the microbiome? An extracellular enzyme involved in N cycling differed between maize phylogenetic groups in one study (Szoboszlay et al., All authors listed have made substantial, direct and intellectual contribution to the work, and approved it for publication.The authors would like to acknowledge University of California and the College of Agricultural and Environmental Sciences for a Graduate Scholars Fellowship to JS and seed research funds from CAES Programmatic Initiative to AG.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The reviewer LC and handling Editor declared their shared affiliation, and the handling Editor states that the process nevertheless met the standards of a fair and objective review.


Practice-based research networks (PBRNs) are organizations that involve practicing clinicians in asking and answering clinically relevant research questions. This review explores the origins, characteristics, funding, and lessons learned through practice-based research in the United States. Primary care PBRNs emerged in the USA in the 1970s. Early studies explored the etiology of common problems encountered in primary care practices , demonstrating the gap between research conducted in controlled specialty settings and real-world practices. Over time, national initiatives and an evolving funding climate have shaped PBRN development, contributing to larger networks, a push for shared electronic health records, and the use of a broad range of research methodologies . Today, there are over 160 active networks registered with the Agency for Healthcare Research and Quality’s PBRN Resource Center that engage primary care clinicians, pharmacists, dentists, and other health care professionals in research and quality-improvement initiatives. PBRNs provide an important laboratory for encouraging collaborative research partnerships between academicians and practices or communities to improve population health, conduct comparative effectiveness and patient-centered outcomes research, and study health policy reform. PBRNs continue to face critical challenges that include: (1) adapting to a changing landscape; (2) recruiting and retaining membership; (3) securing infrastructure support; (4) straddling two worlds (academia and community) and managing expectations; and (5) preparing for workforce transitions. The Agency for Healthcare Research and Quality (AHRQ), one government agency that supports PBRN research in the United States, defines a primary care PBRN as “a group of ambulatory practices devoted principally to the primary care of patients, and affiliated in their mission to investigate questions related to community-based practice and to improve the quality of primary care.”2 By linking questions from practicing clinicians with rigorous research methods, PBRNs can produce research findings that are relevant to clinicians and, in theory, are more easily assimilated into everyday practice. Clinicians are motivated to participate in PBRNs for many reasons: to contribute new knowledge, to reduce feelings of isolation, and to improve the quality of care provided to patients.6 Network members and PBRN structure are meant to foster a sense of commitment that transcends individual research projects.1Networks are usually formal collaborations between community-based practices and academic institutions.In this paper, we review the origins and development, characteristics and functions, funding, and lessons learned through research conducted in US PBRNs. We highlight how PBRNs emerged in response to the needs of practicing primary care clinicians and have adapted in parallel with changes in the health care and funding landscape. We end by exploring the role PBRNs play in building practice-based evidence, supporting research translation, and providing important infrastructure to enable change and improvement in health care delivery.8 These US networks involved partnerships with community clinics, and they emerged at approximately the same time family medicine identified itself as a medical specialty.9 The Ambulatory Sentinel Practice Network (ASPN) was established in 1981; it included both US and Canadian practices. Frequently considered the seminal US PBRN, the ASPN’s research and leadership played a critical role in early PBRN development.10 A second national PBRN, Pediatric Research in Office Settings, was established in 1986 by the American Academy of Pediatrics, demonstrating the merit of these networks.10The first regional PBRNs in the USA, the Dartmouth Primary Care Cooperative Information Project in New Hampshire and the Family Medicine Information System in Colorado, were created in 1978, slightly later than those emerging in the Netherlands, Canada, and Great Britain in the late 1960s and early 1970s.11 the number of networks in the USA has grown substantially in the past few decades. According to a survey published in the Journal of Family Practice, there were 28 active PBRNs in North America by 1994.12 Early networks tended to be regional in scope and to focus on the epidemiology, natural history, and diagnosis of common problems encountered in ambulatory care.9 In 2002, the AHRQ supported the establishment of a PBRN Resource Center, which had over 100 primary care networks registered by 2004.11 Currently, the AHRQ PBRN Resource Center lists 162 registered PBRNs.13 The emergence of new networks has occurred in parallel with the dissolution of existing networks .Although the emergence of PBRNs in the USA was slower than in European countries,14 Despite the focus on primary care, networks of dentists, pharmacists, naturopaths, and palliative medicine clinicians have also emerged in recent years.2 PBRNs continue to evolve in response to the needs of practicing clinicians, policy changes, and current funding opportunities.11 Today’s PBRNs are regional (30%), state-based (28%), local (23%), and national (20%) in membership and scope.14 Additionally, a PBRN may have a specific mission or focus that shapes its membership requirements, such as practicing in a rural setting or in a Federally Qualified Health Center or using a specific electronic health record (EHR). Of the PBRNs registered with the AHRQ resource center, 66% use EHRs and 71% have or plan to collaborate with another PBRN.1416PBRNs vary in a number of ways, including member composition , affiliation , and size. A recent survey of PBRNs found that 76% were affiliated with a university; most others were affiliated with a nonprofit or professional organization.18 focused on developing the capacity to extract or modify data from EHRs19 or developed expertise in conducting comparative-effectiveness research.20The diversity in PBRN mission, size, and geographic area presented in 21 To develop and sustain member relations, PBRNs may actively maintain a membership roster, support multiple methods of communication with key stakeholders, and host regular meetings .22 PBRNs must also keep abreast of member needs, match academic investigators with constituents who have similar topical interests, and respond to funding announcements in a timely fashion.Certain functions appear essential across PBRNs. These include supporting project development, building sustainable relationships with principal investigators and funders, recruiting and retaining voluntary clinicians and practices, managing staff and governance groups, and performing research activities .22 Core infrastructure frequently includes a network director (often an MD or PhD) and a coordinator who are operationally responsible for the PBRN and support the day-to-day operations of the network and research initiatives.22 Network leadership may also sustain an advisory board composed of representative members of the PBRN to guide network activities and inform research. To accomplish network goals and support research studies, PBRNs may hire project managers, research assistants, and statistical experts.22 Networks may also employ practice facilitators to assist primary care teams with quality-improvement studies, community outreach, or other shared goals.24 In some PBRNs, facilitators are regionally based, and they develop long-term relationships with member clinicians and staff that go beyond specific studies.25 Due to the affiliation of many PBRNs with academic institutions, some networks collaborate with university departments to hire core and study-specific staff for portions of their time. This can be economically beneficial for both developed and developing networks.PBRNs create organizational structures to accomplish these functions.22 Early US PBRNs, as well as those today, are constantly balancing and rebalancing the infrastructure needed to maintain basic network functions and achieve their missions. The closure of the ASPN in 1999 due to inadequate infrastructure support underscores the equilibrium PBRNs must maintain.26Variation in the structure and function of PBRNs in the USA exists because there is a dynamic interaction between these elements and the network’s mission and available resources. Green et al argued that PBRNs should establish their mission and purpose first and then design the infrastructure to support it.22 Today, many US PBRNs receive funding from diverse sources, including Federal Agencies such as the AHRQ and the National Institutes of Health ([NIH] 84%), home institutions (74%), foundations (56%), professional organizations (24%), and other sources.27Practicing primary care clinicians and academic faculty have contributed much in-kind effort to the development of PBRNs. However, networks also receive financial support from a number of sources, including state and federal research grants, network membership fees, and institutional and organizational affiliations. To develop as a PBRN, many networks have secured funding earmarked specifically for infrastructure development through grants or awards provided at the local or national level. Although exact figures vary with each PBRN’s research mission, estimates for annual infrastructure costs range from $69,700 for a basic network to $287,600 for a moderately complex network.1 Foundations  have also played a critical role with their support of PBRNs.30 For example, awards from the WK Kellogg Foundation allowed the ASPN to hire its first full-time staff member in 1984 and to appoint its first full-time medical director in 1985.10 Infrastructure and development grants, coupled with support for specific research projects, facilitated the establishment of many PBRNs and helped to sustain core operations.31 The first roadblock was taking new knowledge about disease mechanisms identified through basic research and applying it to the diagnosis, treatment, and prevention of these diseases in people . The NIH called this “T1 research.” The second roadblock identified was translating the results of clinical studies into clinical decision making and treatment in everyday practice . The NIH called this “T2 research.” Many PBRNs have reframed their mission to provide community-based laboratories for T2 research. The NIH now includes dissemination and implementation research in its portfolio, which encourage applications from research and practice networks, demonstrating how PBRNs have shaped the funding landscape.33The funding landscape has shaped how PBRNs frame the work they do – and it has been shaped by this research. For example, in the early 2000s, the Institute of Medicine’s Clinical Research Roundtable identified two major roadblocks to moving research into practice.35 Additionally, recent funding calls from the Patient-Centered Outcomes Research Institute emphasized the importance of comparative clinical-effectiveness research to help patients and health care providers make more-informed decisions.37 PBRN infrastructure provides a critical framework for supporting research like this in real-world practice and community settings.There is also synergy between the NIH’s Clinical and Translational Science Award (CTSA) program and the newly formed Patient-Centered Outcomes Research Institute, which share many of the same priorities as PBRNs. The emergence of the CTSA program in 2006 elevated the importance of incorporating community-based research into academic health science institutions across the USA. Some PBRNs used this as an opportunity to emphasize the work they did supporting community-engaged research with both practicing clinicians and community partners. As such, CTSAs at some institutions have partnered with PBRNs to support these efforts.11 Some networks also use a collaborative approach by which academics and community partners  work together in a participatory fashion to codevelop the research agenda.11 We present a brief review of PBRN research over time to highlight critical contributions, describe expanding approaches and methodologies, and explore the opportunities ahead. We identified studies using a search for “practice-based research” and “practice-based research networks” in PubMed and selected a sample of articles from early, middle, and the current time periods to inform us about the lessons learned. This approach was not intended to be systematic or comprehensive, but rather to highlight patterns in the PBRN research landscape.PBRN research helped establish knowledge vital to the delivery of high-quality health care in ambulatory-care and community settings. PBRNs conduct research on topics that emerge from practicing clinicians (bottom-up research), and from individual investigators (top-down research).38 Results from these initial studies were generally presented at conferences and published in the Journal of the American Board of Family Medicine and Journal of Family Practice. This research demonstrated the misalignment between evidence-based, published guidelines and the manifestation of symptoms and disease in general practice.9 For example, an observational study of usual primary care indicated that 40% of spontaneous abortions were managed completely in the office or at home. This finding raised questions about text recommendations for dilation and curettage.39 This and other studies highlight how the context in which you study a question  shapes the answers you discover and can have a major impact on how care is rendered.40Early PBRN studies explored everyday clinical problems , and many of them engaged physicians directly in data collection using the card study methodology.These early findings made an important contribution to the evidence base in health care, and leaders in primary care used the results of these early studies to advocate for practice-based research. They did so by pointing out the limitations of randomized controlled trials that narrowly defined the study sample were conducted in controlled environments and were frequently led by researchers and specialists unfamiliar with general practice. While studies with these attributes may carry weight in the field and inform guideline-setting organizations and standards of care, they may not accurately portray the effectiveness of new treatments in the general population.9 Moreover, the impact of these applied studies on clinicians and patients had the potential to be immediate and far-reaching. Green et al wrote, “The new knowledge that comes from practice-based research will not find application to only a few with fully developed or perhaps unusual disease. It will benefit virtually everyone.”41 Bringing practices together into a network (1) created the infrastructure to quickly get enough power or practices and people to study a problem, (2) enhanced the generalizability of the studies, and (3) increased research productivity. A single PBRN could provide the practices needed to study a range of phenomena simultaneously. Moreover, PBRNs created the infrastructure to generate practice-based evidence – evidence that is relevant to clinicians and the settings in which they practice.27In addition to demonstrating a vital knowledge gap, early practice-based research established the feasibility of conducting research in networks of community practices.42 geriatric,43 pediatric,44 and family medicine settings.46 In addition to expanding research topics, PBRNs were just beginning to diversify the research methods used in studies including cross-sectional survey research,45 cohort studies,44 observational studies and interviews,43 and mixed-methods research.46 Much of the research coming from PBRNs at this time was published in the Journal of Family Practice, but findings were also beginning to gain traction in a wider range of journals, with manuscripts appearing in the Archives of Family Medicine, the Western Journal of Medicine, and the Journal of the American Geriatrics Society.By the late 1990s, a variety of research was being done in PBRNs to look at clinical issues in obstetric,47 Studies involving multiple PBRNs,48 or data mined from EHRs to provide a generous sample size,49 are not uncommon. PBRNs can also support system-level interventions, and randomization can be done at the practice or patient level. A 2007 mixed-method study of primary care PBRN directors and administrative officers found that common research foci included prevention, diabetes, cardiovascular risk factors, and mental health.11 Additionally, some PBRNs play an active role in supporting health-reform initiatives and quality-improvement projects, such as assisting practices as they pursue patient-centered medical home status.51 Networks have also begun to address community-level health needs by using participatory research methods52 and conducting studies that link clinics with community-based resources to foster health behavior change.55Just as early PBRN research studied everyday clinical problems, today’s networks explore a diverse range of phenomenon experienced in daily practice, including service delivery and health care redesign. This trend toward the expansion of PBRN research methods and research impact continues. Studies have become more complex, they involve an increasing number of participating sites, and they occur in a broader range of PBRNs, including dentistry.56 This change highlights the increasing role of team-based care in health care settings. Additionally, articles from PBRNs now regularly appear in the Journal of the American Board of Family Medicine, the Annals of Family Medicine, and non-primary care journals, including those with a focus on medical informatics,48 pharmacology,49 health disparities,57 health care management,58 and dentistry.47 Top-tier journals such as the American Journal of Preventive Medicine have dedicated entire issues exclusively to PBRN research.29As network foci expanded, study participants and publication targets diversified. PBRN studies now engage various frontline health care providers, including nonphysician clinicians (nurse practitioners and physician assistants), nurses, social workers, and behavioral health specialists.51 expanding partnerships between primary care and public health organizations,60 and participating in the emergence of accountable care organizations.61 Networks also have the opportunity to engage in comparative-effectiveness research.20 Networks continue to play an important role in both the study of care delivery and the application of these approaches to daily practice.62 We highlight five critical challenges that may inform future PBRN work.PBRNs are poised to continue to play a critical role in health reform initiatives such as supporting clinic redesign,PBRNs have responded to the changing health care landscape by widening their membership  and embracing diverse research methodologies . This expansion provides greater opportunities to partner with academic researchers, meet the needs of practicing members, and stay flexible in light of funding opportunities. Moreover, it provides the breadth and infrastructure to address critically relevant questions for practitioners, academicians, policymakers, and other stakeholders. However, network leadership may be challenged to identify and sustain a shared vision that can motivate the participation and secure the infrastructure capacity needed to respond to more diverse stakeholders.PBRN leadership has played a critical role in developing network membership, locating funding opportunities, and implementing research studies. Clinicians were initially drawn to PBRNs for the camaraderie and opportunity to improve care for their patients. However, changing practice structures  and growing competing demands may make it more difficult to recruit and sustain practice membership. PBRNs must learn how to negotiate their roles within larger health care systems, reframe and renew the benefits PBRNs offer individual clinicians , and engage nonclinician practice and systems’ administrative and executive staffs.Securing and sustaining funding to support network infrastructure will continue to be a challenge for PBRNs. Although the emergence of CTSAs provided some networks with resources to build critical, foundational relationships for research, many PBRNs still struggle to finance core infrastructure. Building a robust research capacity is difficult when networks are dependent on soft money (grant funds) for core staff support. In an increasingly competitive grant environment, networks may need to pursue nontraditional sources of funding by building partnerships with state governments, insurance companies, and health care systems. The tension between working with new partners to secure financing will need to be carefully balanced with the mission and credibility of the PBRN.PBRNs play an important role in spanning the boundaries between clinical and community practice and the academic establishment. The time demands, focal concerns, and indicators of quality and achievement are different in clinical and academic environments. Practices and communities operate at a fast pace and are often focused on providing services at the individual level. Academics have accommodated a schedule of delayed gratification, where the traditional research timeline from idea to funding to project completion can last for years. Networks may struggle to balance a clinic’s and community’s needs for action with a researcher’s timeline. Developing methods to straddle these worlds for their mutual benefit may require compromise and trade-offs. For example, academics may need protected time to collaborate with clinic and community partners, an aspect of community-engaged research that is not often accounted for in traditional academic promotion calculations; practices and communities may also benefit from taking time to reflect upon research results and questions, although this is something they may have neither the patience nor the luxury to embrace.As with the general US population, PBRN leadership is aging. Developing transition plans so that networks can be sustained as current network directors retire will be crucial. Because many networks have a lean infrastructure, there are often no people in the ranks  who have been mentored over time to take on leadership roles. Such transitions provide opportunities for innovation and challenge network stability.PBRNs have been identified as research laboratories essential to advancing the science of medical care. They are a venue both for describing clinical problems encountered in everyday practice and for speeding the translation of research into routine care. PBRNs have expanded from regional affiliations to national and international organizations that use multiple methods to address the needs of practicing clinicians and communities. Because PBRNs have demonstrated their effectiveness as laboratories for clinical research and knowledge translation, they have become central players in health services research. PBRNs are well poised to play important roles in implementing and exploring areas critical to health care reform, such as facilitating better integration between primary care and public health services or assisting with the development of accountable care organizations.The sustainability and effectiveness of PBRNs have been, and may continue to be, predicated on PBRNs’ ability to negotiate their mission in light of current funding priorities and a dynamic health care environment. PBRNs must attend to five critical challenges as they move forward: (1) adapting to a changing landscape; (2) recruiting and retaining membership; (3) securing infrastructure support; (4) straddling two worlds (academia and community) and managing expectations; and (5) preparing for workforce transitions.


Preterm birth complications are the leading cause of deaths for children under five years. Antenatal corticosteroids (ACS) are effective at reducing mortality and serious morbidity amongst infants born at <34 weeks gestation. WHO guidelines strongly recommend use of ACS for women at risk of imminent preterm birth where gestational age, imminent preterm birth, and risk of maternal infection can be assessed, and appropriate maternal/newborn care provided. However, coverage remains low in high-burden countries for reasons not previously systematically investigated.Every Newborn Action Plan process. Country workshops involved technical experts to complete the survey tool, which is designed to synthesise and grade health system "bottlenecks", factors that hinder the scale up, of maternal-newborn intervention packages. We used quantitative and qualitative methods to analyse the bottleneck data, combined with literature review, to present priority bottlenecks and actions relevant to different health system building blocks for ACS.The bottleneck analysis tool was applied in 12 countries in Africa and Asia as part of the Eleven out of twelve countries provided data in response to the ACS questionnaire. Health system building blocks most frequently reported as having significant or very major bottlenecks were health information systems (11 countries), essential medical products and technologies (9 out of 11 countries) and health service delivery (9 out of 11 countries). Bottlenecks included absence of coverage data, poor gestational age metrics, lack of national essential medicines listing, discrepancies between prescribing authority and provider cadres managing care, delays due to referral, and lack of supervision, mentoring and quality improvement systems.Analysis centred on health system building blocks in which 9 or more countries (>75%) reported very major or significant bottlenecks. Health information systems should include improved gestational age assessment and track ACS coverage, use and outcomes. Better health service delivery requires clarified policy assigning roles by level of care and cadre of provider, dependent on capability to assess gestational age and risk of preterm birth, and the implementation of guidelines with adequate supervision, mentoring and quality improvement systems, including audit and feedback. National essential medicines lists should include dexamethasone for antenatal use, and dexamethasone should be integrated into supply logistics. Each year, an estimated 15 million infants are born premature, representing over one in ten live births , with ovReducing the burden of preterm birth requires effective maternal care including comprehensive obstetric care , and spThere has been rapid adoption of ACS use in high-income countries (HICs) since the mid-1990s , with coACS is only one part of obstetric and preterm birth management  and shouLow coverage of ACS has been variously attributed to lack of guidelines, prescribing authority, provider awareness or skills, drug availability, and patient access to appropriate facilities . HoweverThis paper presents the results of a systematic multi-country analysis of barriers to uptake of ACS for preterm birth. In this analysis, we aim to identify priority health system building blocks facing common and critical bottlenecks to scaling up this life-saving intervention. We additionally discuss policy and programmatic implications and recommend priority actions drawn from the survey responses, existing evidence, and programme experience.Objectives of this paper are as follows:1. Use a 12-country analysis to explore health system bottlenecks affecting the scale-up of ACS.2. Present the solutions to overcome the most significant bottlenecks including insights from respondent countries, literature review and programme experience.3. Discuss policy and programmatic implications and propose priority actions.This study used quantitative and qualitative research methods to collect information, assess health system bottlenecks and identify solutions to scale-up of maternal and newborn care interventions in 12 countries: Afghanistan, Cameroon, Democratic Republic of Congo (DRC), Kenya, Malawi, Nigeria, Uganda, Bangladesh, India, Nepal, Pakistan and Vietnam. The methodology has been discussed in detail previously and in paper 1 of this supplement ,25.Every Newborn Action Plan (ENAP) process to assist countries in identifying context-specific bottlenecks to the scale-up and provision of maternal and newborn health interventions across seven health system building blocks [Every Newborn Steering Group between July 1st and December 31st 2013 . We first present the grading in heat maps according to the very major or significant health system bottlenecks as reported by all 12 countries, then by mortality contexts  <30 or NMR ≥30 deaths per 1000 live births) and by region (countries in Africa and countries in Asia). We developed a second heat map showing the specific grading of bottlenecks for each health system building block by individual country. Survey responses were analysed for common specific bottlenecks, defined to be those reported by at least 3 countries. Where the same specific bottleneck was reported under more than one building block, it was categorised in accordance with the relevant survey question, or under the building block where most countries reported the specific bottleneck.We graded bottlenecks for each health system building block using one of the following options: not a bottleneck (=1), minor bottleneck (=2), significant bottleneck (=3), or Finally, we categorised context-specific solutions for scaling up under the subcategories of corresponding bottlenecks within each health system building block.Eleven out of twelve country teams submitted their responses to the ACS questionnaire. Cameroon, Democratic Republic of the Congo (DRC), Kenya, Malawi, Nigeria, Uganda, Bangladesh, Nepal, and Vietnam returned national-level responses. Pakistan provided subnational data from all provinces, Gilgit-Baltistan, and Azad Jammu and Kashmir, excluding two tribal territories. Sindh province, Pakistan, did not provide a grade for community ownership and partnership. India returned data from three states: Andhra Pradesh, Odisha, and Rajasthan. Rajasthan state completed the questionnaire and listed specific bottlenecks, but did not provide ratings for any building block. India responded to the earlier version of the questionnaire with fewer questions. Afghanistan, the twelfth country, returned national-level survey data without responses to the ACS portion and therefore is not included in our analysis.An overview of grading for the health system building blocks, for all countries and by mortality setting and geography, is shown in Figure Building block grading for each respondent country is summarised in Figure Table Leadership and governance was graded as a very major or significant bottleneck by 7 of 11 country teams Figure , represeProposed solutions included development (or update) and dissemination of national guidelines and protocols on prevention and management of preterm labour, including ACS. Updated WHO guidelines will be useful to inform national guidelines.Health financing was graded as a very major or significant bottleneck by 8 of 11 country teams. The most commonly cited specific bottleneck was insufficient funding, reported by 9 country teams, with 8 indicating a lack of priority or policy as the underlying cause.Proposed solutions included increased advocacy and leadership, particularly with funding and budget allocation, for preterm birth prevention, management and care including ACS.Health workforce was graded as a very major or significant bottleneck by 7 of 11 country teams. All country teams questioned reported a shortage of health workers especially in higher cadres, inadequate training, and inadequate supervision or mentoring. India did not provide an answer to the question on health worker shortages.Proposed solutions included integration of ACS into pre-service and in-service training, development and dissemination of job aids, and consideration of expanded prescription authority for more cadres of health workers.Essential medical products and technologies was graded as a very major or significant bottleneck by 9 of 11 country teams. All 11 country teams cited as a specific bottleneck the lack of inclusion of ACS on the national essential medicines list (NEML) for fetal lung maturation, though all included dexamethasone for other indications. Of 8 country teams reporting inadequate procurement or distribution of ACS, 6 cited the lack of policy or NEML listing for fetal lung  as a direct cause of lack of integration into supply chain system. Additionally, no respondent country included ACS in quantification and forecasting, and none reported having an indicator for ACS drug availability.Proposed solutions included listing of ACS drugs on NEMLs for a fetal lung maturation indication, development and dissemination of guidelines, and software to support needs-based forecasting and procurement.Health service delivery was graded as a very major or significant bottleneck by 9 of 11 country teams. All country teams reported having inadequate or non-existent quality monitoring and improvement systems, and 9 country teams noted a lack of alignment between ACS prescribing authority and health worker cadres providing care for women at risk of preterm birth. Additionally, 7 countries cited delays due to referrals as a specific bottleneck. Table S3 and S4 of the Additional file Proposed solutions included development and dissemination of guidelines across facilities, creation of supervision and mentoring systems, integration into clinical audits and reviews, and strengthened referral systems.Health information system was identified as the most problematic building block, with all 11 country teams reporting very major or significant bottlenecks. Responses to health information systems questions revealed an almost complete absence of defined and standardised indicators for ACS use, with the exception of Nepal and one province in Pakistan, where country teams reported that ACS use was tracked on partographs. No countries reported having an indicator for ACS drug availability. Moreover, no countries included ACS use in clinical audits or perinatal reviews.Proposed solutions included creation of indicator(s) for ACS use and integration of ACS data into record-keeping systems and regular reviews.Community engagement was graded as a very major or significant bottleneck by 8 of 11 country teams. The most commonly cited specific bottlenecks were lack of awareness and initiatives targeted at raising awareness of preterm birth risks and management (9 countries), and the absence of an accessible facility or transportation to reach a facility where ACS could be administered (9 countries).Proposed solutions included leveraging existing community outreach infrastructure, such as community groups, opinion leaders, and media.This systematic analysis of ACS use in 11 high-burden countries, which together account for over half of maternal and newborn deaths, has identified commonly experienced health system bottlenecks to scaling up ACS for management of preterm birth. A comprehensive approach to preterm birth should include strategies for prevention as well as management; however, the menu for high-impact, evidence-based preterm birth prevention is currently limited . In highThe important principle of "do no harm" and potential risks of ACS have been highlighted recently through the ACT trial. The benefits of ACS are dependent on high coverage in preterm babies <34 weeks; benefit is not expected or is marginal after 34 weeks, and risks increase close to term. Targeting based on gestational age and more accurate diagnosis of imminent preterm birth are therefore key aspects of effective, safe use as underscored in WHO guidelines. In addition, ACS alone cannot be a magic bullet; the preterm infant still requires a minimum of supportive care including warmth and feeding support, and if <32 weeks gestation is more likely to require oxygen therapy and respiratory support. Approximately half of the births in the ACT trial occurred at home or low-level primary care facilities .Despite variation across the 11 countries that responded to the survey, the greatest barriers were consistent, highlighting three priority health system building blocks--health information systems, health service delivery and essential medical products and technologies--with the most critical common bottlenecks. Drawing from country programme experience, and evidence from literature Table , this paData on ACS coverage, use, and outcomes including safety were absent in nearly all countries. To enable continuous and quantitative assessment of ACS programmes, appropriate indicators need to be defined, and these data integrated into existing health information systems in order to allow tracking and comparison at facility, district, and national levels. ACS coverage data are one of the priority indicators identified in the Every Newborn Action Plan . CoveragUse of data for quality improvement would be aided if ACS were systematically included in safe childbirth checklists, partographs, clinical audits, and perinatal death reviews . Data arEstimates suggest ACS coverage in LMICs is generally low even in tertiary facilities Table , consistAvailable evidence supports focusing scale-up efforts in facilities with the capacity for gestational age assessment and diagnosis of women at risk of imminent preterm birth, as well as ongoing support for preterm infants and well-functioning maternity wards . The recWith clear evidence for use in hospitals in HICs with adequate neonatal support and reason for caution at lower-level facilities in LIC settings, programmatic scale-up is best focused on higher-level and high-volume facilities, where resources can also be used to produce significant impact cost-effectively and also improve targeting and safety tracking. Increased population coverage will then depend on increased and timely identification and referral of at-risk women, both in lower-level facilities and in homes. Systemic improvements are needed to encourage institutional deliveries with skilled birth attendants and to strengthen referral systems, while guidelines and training at lower levels of care will be critical to building capacity for timely identification. Expanded care such as a pre-referral dose may also improve care by allowing a longer time for ACS to take effect. However, such steps must be considered cautiously by each country based on the capacity of lower-level facilities to provide adequate care to ensure safety.Within hospitals, coverage is often limited by inadequate numbers of physicians or other providers present and also adequately trained to assess gestational age, diagnose high risk of preterm birth, and authorised to prescribe and administer ACS. The majority of countries reported both a shortage of health workers at higher cadres and a mismatch between provider cadres allowed to prescribe ACS and those cadres likely to be caring for women at risk of preterm birth  and are often assumed to be widely available. Yet most respondents indicated shortages at the country level, primarily attributed to a lack of supporting policy or effective logistics systems. NEML listing for the fetal lung maturation indication is essential to prioritisation for procurement as well as integration into supply chain, from forecasting to distribution.Adequate procurement further depends on development and dissemination of guidelines on ACS use, including clarity on the choice of corticosteroid and appropriate regimen. Two corticosteroids, dexamethasone and betamethasone, have been shown to be safe and effective to manage preterm birth. Betamethasone is sometimes preferred in HICs due to limited evidence suggesting better outcomes  and potentially lower risk of maternal infection [However, evidence on dexamethasone still conclusively supports its overall safe and effective use for management of preterm birth. A meta-analysis of 9 studies directly comparing dexamethasone with betamethasone further found no statistically significant differences apart from a greater reduction in intraventricular haemorrhage using dexamethasone . A large RCT directly comparing dexamethasone to betamethasone is currently underway .Critically, only dexamethasone is a feasible choice for scale-up in most LMICs. While dexamethasone is widely available from international suppliers for a variety of indications, the formulation of betamethasone supported by most evidence  has been subject to global shortages  and costDexamethasone also faces fewer policy hurdles to increased use for management of preterm birth. With a variety of uses, dexamethasone in the recommended formulation is already registered, listed on the NEML, and included in procurement and supply chain in all 11 countries, for other indications. Dexamethasone is the only ACS listed on the WHO EML with a fetal lung maturation indication .National policies, in line with the current WHO guideline, should therefore focus on promoting appropriate use of dexamethasone through procurement policy, clear guidelines, and integration into forecasting, procurement, and supply chain.Soliciting responses from a wide range of in-country partners and practitioners in maternal and newborn health captured context-specific challenges and generated collaborative solution ideas. The grading process also created consensus around priority bottlenecks and health system building blocks to be addressed. However, these consensus views are subjective. The quality and amount of information also varied depending on the level of knowledge of participants on health system issues and on workshop facilitation. In addition, bottlenecks were reported as perceived bottlenecks relative to the other health system building blocks. National-level assessment may mask regional disparities, particularly between urban and rural areas. This comprehensive questionnaire may have led to respondent fatigue, but this effect is least significant for ACS as the first intervention in the bottleneck analysis tool. For India and Pakistan, subnational responses may not mirror all national-level challenges. However, Pakistan submitted responses from all regions except two tribal territories, and although India returned data from only three of its 28 states, these areas are amongst the poorest states and include populations similar to those of Vietnam, Kenya, and DRC .Further research is needed to establish the effectiveness of the solutions described here, especially in the context of ACS use in LMIC hospital settings where most of the world's births now occur. To date the evidence on ACS is primarily from HIC settings with neonatal intensive care  or the AAnother important track of research is to improve the accuracy and also feasibility of gestational age assessment, including testing tools to improve accuracy of last menstrual period assessment, use of ultrasound dating in later pregnancy and ways to promote better recording in notes and use of the data by clinicians. Key messages and actions are summarised in Figure ACS for management of preterm birth has been a standard of care in wealthy countries for over 20 years, yet is greatly underused in low-resource countries, where complications of preterm birth have recently become the leading cause of death in children under 5. Preterm infants are over 12 times more likely to die in the poorest compared to the richest countries, and infants in the poorest countries have a survival rate at 32 weeks similar to that of infants born in the richest countries at 25 weeks. This systematic bottleneck analysis using data from 11 countries identifies critical areas of focus and suggests a set of actionable solutions for extending this inexpensive, high-impact intervention, whilst also promoting the tracking for safety, potentially saving hundreds of thousands of lives each year.ACS: Antenatal corticosteroids; ANM: Auxiliary nurse-midwife; DRC: Democratic Republic of Congo; EML: Essential medicines list; FLM: Fetal lung maturation; HICs: High income countries; LIC: Low income country; LMIC: Low- or middle-income country; LMIS: Logistics management and information system; MIC: Middle-income country; MOH: Ministry of Health; NEML: National essential medicines list; NMR: Neonatal mortality rate; RCT: Randomised controlled trial; RDS: Respiratory distress syndrome; WHO: World Health Organization.The authors have not declared any competing interests. The assessment of bottlenecks expressed during consultations reflects the perception of the technical experts and may not be national policy. The authors alone are responsible for the views expressed in this article and they do not necessarily represent the decisions, policy or views of the organisations listed, including WHO.GL analysed data and drafted the manuscript in collaboration with JS. KED, AS-K JEL and SGM contributed to conceptualisation of the paper, data analysis and interpretation, and review of drafts. FA and MG and MM contributed to data interpretation and discussion and reviewed drafts. JMS, JH, PB, MJ, and EM contributed to country case studies. All authors reviewed drafts and approved the final manuscript.Bottleneck tool questionnaire.Click here for fileSupplementary tables, figures and literature search strategy.Click here for file