Enter and Discuss Orders and Prescriptions (EPA 4): A Curriculum for Fourth-Year Medical Students.

Introduction: Order entry, entrustable professional activity (EPA) 4, is one of several EPAs that residency program directors identify as a weakness for PGY 1 residents. A multispecialty survey of program directors indicated that only 69% of interns could be trusted to enter and discuss orders and prescriptions without supervision. To address this gap, we developed a formative workshop for fourth-year medical students. Methods: Prior to the start of their subinternships, 366 fourth-year medical students engaged in an order entry workshop. Students performed chart reviews on electronic standardized patients within an educational electronic health record (EHR), placed admission orders, customized order sets, responded to safety alerts, utilized decision support tools, and incorporated high-value care considerations. Students used expert-validated rubrics to assess the quality of their admission orders and participated in a facilitated group discussion on key learning points. Finally, students participated in order entry, with all orders requiring cosignature by a supervising physician, during their clinical rotations. Students reported their confidence with order entry before and after the workshop and after the clinical rotation. Results: One hundred seventeen students completed the pre- and postworkshop surveys, and 99 went on to complete the postcourse evaluation. Students showed a statistically significant increase in their confidence level following the workshop. Discussion: Order entry is a critical, complex skill that requires deliberate instruction. This curriculum, which leverages the features of an educational EHR, can facilitate instruction, practice, and confidence gains regarding order entry prior to further application of these skills in the clinical environment.

An Initiative to Increase Residency Program Diversity.

A physician workforce that reflects the patient population is associated with improved patient outcomes and promotes health equity. Notwithstanding, racial and ethnic disparities persist within US medical schools, making some individuals underrepresented in medicine (URM). We sought to increase the percentage of URM residents who matched into our pediatric residency programs from a baseline of 5% to 35% to achieve demographic parity with our patients. We developed a multifaceted approach using multiple iterative tests of change, with the primary strategy being increased visibility of URM trainees and faculty to residency applicants. Strategies included applicant interviews with URM faculty, interview dinners with URM residents, visibility at academic conferences for URM trainees, development of targeted marketing materials, and a visiting student program supported by networking with URM residents. The primary outcome measure was the percentage of matched residents in the categorical pediatrics, child neurology, and medical genetics training programs who identified as URM. The percentage of URM residents increased to 16% (6 of 37) in 2018, 26% (11 of 43) in 2019, 19% (8 of 43) in 2020, and 21% (9 of 43) in 2021 (a four-year average of 22% URM residents; P = .0002). This progress toward a more representative residency program was met by challenges, such as pipeline concerns, the minority tax, and recruitment during a pandemic. We were able to implement small, low-resource strategies that had a large cumulative impact and could be implemented in other residency programs. Specific tactics and challenges encountered are discussed in this special article.

Effective methods of clinical education.

OBJECTIVE: To provide an overview of key points and helpful tips related to engaging learners in different medical education settings, including the virtual education environment. DATA SOURCES: Searches were conducted in PubMed and Google Scholar for articles and review papers on the topic of medical education. STUDY SELECTIONS: There was a focus on articles that reviewed best practices on a variety of topics related to medical education. RESULTS: Medical education is a critical part of any students' or trainees' educational curriculum or training program. There are different settings where medical education is provided, and both general and type-specific best practices can be considered. Those who spend time in medical education are continuously working on innovative ideas to enhance and improve how educators teach in both large and small group settings. More recently, the presenter must also consider whether the event will occur in a live or virtual setting as e-learning has become more popular and at times required, which has both benefits and challenges in medical education. CONCLUSION: It is the responsibility of all those who work in medical education to provide interactive and engaging education to learners at all levels of training.

Advanced Multimodal Communication Curriculum for Pediatric Residents.

BACKGROUND: Medical educators juggle competing demands as they seek to integrate medical advancements and new technology with the call for earlier introduction to clinical experiences. Newer models of medical education place even greater emphasis on the importance of deliberate training of providers who can deliver compassionate patient-centered care. The need for adaptable, effective communication skills training has never been more relevant than now, in our high-tech and ever-evolving learning climate. METHODS: At a large pediatric academic center, we used Kern's six-step approach to complete a needs assessment, identify goals and learning preferences of trainees, and ultimately develop and evaluate a multimodal communication curriculum. This curriculum was rooted in the Accreditation Council for Graduate Medical Education's Pediatric Milestones, with the goal of enhancing knowledge, skills, and competency. Pediatric interns were randomized to either the new curriculum (n = 19) or the existing didactic-based communication training (n = 17) from 2019 to 2020. We evaluated the impact of the new and traditional curriculum through evaluations by expert facilitators, learner surveys, and faculty-assigned resident milestones. RESULTS: Many trainees self-identified performance gaps in communication skills at baseline. Eighteen residents attended 1 to 6 in-person deliberate practice workshops. Workshop participation by the residents varied over time due to a variety of factors. All residents, regardless of curricular exposure, showed statistically significant improvement in communication milestones from first to second year and those enrolled in the deliberate practice curriculum highly valued the workshops and coaching. DISCUSSION: Our curriculum demonstrates the value of deliberate practice opportunities for residents to gain skills in advanced communication. Residents appreciated this type of training and the time devoted to expand these skills. Lessons learned and barriers to implementation from this curriculum can be helpful for educators interested in integrating active, deliberate practice, simulation-based communication training into their current educational model.

Validation of the Second Victim Experience and Support Tool-Revised in the Neonatal Intensive Care Unit.

OBJECTIVE: The aim of the study was to validate a revised version of the Second Victim Experience and Support Tool (SVEST-R). The SVEST survey instrument was developed to measure the emotional and professional impact of medical errors and adverse patient events on healthcare providers and can help healthcare organizations evaluate the effectiveness of support resources. METHODS: An SVEST-R was completed by 316 healthcare providers from seven neonatal intensive care units affiliated with a large, pediatric hospital. The original 29-item measure was expanded to 43 items to assess eight psychosocial domains (psychological distress, physical distress, colleague support, supervisor support, institutional support, nonwork-related support, professional self-efficacy, resilience) and two employment-related domains (turnover intentions, absenteeism) associated with the second victim experience. Seven additional items assessed desired forms of support (e.g., time away from the unit). A confirmatory factor analysis evaluated the factor structure of the modified measure. RESULTS: The initial confirmatory factor analysis did not reveal an acceptable factor structure; thus, eight items were removed because of inadequate factor loadings or for conceptual reasons. This resulted in an acceptable model for the final 35-item measure. The final version included nine factors (i.e., psychological distress, physical distress, colleague support, supervisor support, institutional support, professional self-efficacy, resilience, turnover intentions, and absenteeism), with Cronbach α ranging from 0.66 to 0.86. CONCLUSIONS: The SVEST-R is a valid measure for assessing the impact of errors or adverse events on healthcare providers. Importantly, the SVEST-R now includes positive outcomes (i.e., resilience) that may result from the second victim experience.

Pediatric Inpatient Problem List Review and Accuracy Improvement.

BACKGROUND AND OBJECTIVES: The problem list (PL) is a meaningful use-incentivized criterion for electronic health record documentation. Inconsistent use or inaccuracy of the PL can create communication gaps among providers, potentially leading to diagnostic delays and serious safety events. The objective of the study was to increase the rate of PL review by attending physicians for inpatients discharged from hospital pediatrics and infectious disease services from a baseline of 70% to 80% by June 2018 and to sustain the rate for 6 months. The secondary aim was to improve PL accuracy by decreasing the rate of duplicate codes and red code diagnoses that should resolve before discharge from a baseline of 12% and 11%, respectively, to 5% and sustaining the rate for 6 months. METHODS: A quality improvement team used the Institute for Healthcare Improvement Model for Improvement. We tracked duplicate codes and red codes as surrogate markers of PL quality. Rates of PL review and PL quality were analyzed monthly via statistical process control charts (p-charts) with 3-σ control limits to identify special cause variation. RESULTS: PL review improved from a baseline of 70% to 90%, and the change was sustained for 1 year. PL quality improved as duplicate codes at the time of discharge decreased from 12% to 6% and as red codes decreased from a baseline of 11% to 6%. CONCLUSIONS: The PL is an important communication tool that is underused. By engaging and educating stakeholders, incentivizing compliance, standardizing PL management, leveraging electronic health record enhancements, and providing physician feedback, we improved PL meaningful use and quality.

The Pediatric Match Frenzy: An Overview and an Approach for Mentoring Medical Students.

Applying for a pediatric residency position has become an increasingly stressful event and recently medical educators have described it as a "Match Frenzy." Match statistics demonstrate increased competition for pediatric residency positions and a record number of applications. Faculty who mentor medical students are now challenged to counsel them through the intensified process and pediatric residency programs are now forced to navigate the rising number of applications. We define the Match Frenzy and its implications, review historical and current match statistics, and describe a data-driven approach to the problem. Through mitigating the frenzy surrounding this process, we can help students and residency programs better allocate their precious resources.

The emotional impact of errors or adverse events on healthcare providers in the NICU: The protective role of coworker support.

AIMS: To examine the impact of errors or adverse events on emotional distress and professional quality of life in healthcare providers in the neonatal intensive care unit, and the moderating role of coworker support. BACKGROUND: Errors or adverse events can result in negative outcomes for healthcare providers. However, the role of coworker support in improving emotional and professional outcomes has not been examined. DESIGN: A cross-sectional online survey from a quality improvement initiative to train peer supporters in a neonatal intensive care unit. METHODS: During 2015, 463 healthcare providers in a neonatal intensive care unit completed a survey assessing their experiences with an error or adverse event, anxiety, depression, professional quality of life and coworker support. RESULTS: Compared with those who did not experience an error or adverse event (58%), healthcare providers who observed (23%) or were involved (19%) in an incident reported higher levels of anxiety and secondary traumatic stress. Those who were involved in an event reported higher levels of depression and burnout. Differences between the three groups (no event, observation and involvement) for compassion satisfaction were non-significant. Perceived coworker support moderated the association between experiencing an event and both anxiety and depression. Specifically, experiencing an event was associated with higher levels of anxiety and depression when coworkers were perceived as low in supportiveness, but not when they were viewed as highly supportive. CONCLUSION: Findings suggest that errors or adverse events can have a harmful impact on healthcare providers and that coworker support may reduce emotional distress.

Deployment of a Second Victim Peer Support Program: A Replication Study.

BACKGROUND: The second victim phenomenon occurs when health-care providers experience significant professional distress (compassion dissatisfaction, burnout, secondary traumatic stress) and psychological distress (shame, anxiety, and depression) as a result of medical errors or adverse patient outcomes. Few hospitals have institution-wide systems in place to assist employees through the recovery process. METHODS: At Nationwide Children's Hospital (NCH), a peer-based support program called "YOU Matter" was executed and spread hospital-wide. The program emulated the framework and execution strategy designed by University of Missouri Health Care's (MUHC) "forYOU" Team. Strategic elements of the program's structure were reviewed and adapted for NCH with system-wide deployment and enhancement to include electronic peer support reporting. This article summarizes program implementation, management, and sustainment over the past 2 years. RESULTS: By following University of Missouri Health Care's model, we successfully deployed an institution-wide second victim program. Since the November 2013 initiation, we have documented 232 peer and 21 group encounters. High-risk clinical areas for second victimization at NCH included the emergency department (ED), pediatric intensive care unit (PICU), cardiothoracic intensive care unit (CTICU), and pharmacy department. Registered nurses (RNs) and licensed practical nurses (LPNs) have had the highest number of encounters necessitating second victim support (32%). Supported staff reported improved emotional state and improved return-to-work metrics. CONCLUSIONS: An organization's culture of patient safety can be enhanced by ensuring staff psychological safety. Programs like "YOU Matter" and the "forYOU" Team are essential building blocks to improve the overall safety culture and quality of care. Implementation of "YOU Matter" at NCH validates the MUHC program and demonstrates its generalizability to other health-care institutions.

Transcriptome resources and functional characterization of monoterpene synthases for two host species of the mountain pine beetle, lodgepole pine (Pinus contorta) and jack pine (Pinus banksiana).

BACKGROUND: The mountain pine beetle (MPB, Dendroctonus ponderosae) epidemic has affected lodgepole pine (Pinus contorta) across an area of more than 18 million hectares of pine forests in western Canada, and is a threat to the boreal jack pine (Pinus banksiana) forest. Defence of pines against MPB and associated fungal pathogens, as well as other pests, involves oleoresin monoterpenes, which are biosynthesized by families of terpene synthases (TPSs). Volatile monoterpenes also serve as host recognition cues for MPB and as precursors for MPB pheromones. The genes responsible for terpene biosynthesis in jack pine and lodgepole pine were previously unknown. RESULTS: We report the generation and quality assessment of assembled transcriptome resources for lodgepole pine and jack pine using Sanger, Roche 454, and Illumina sequencing technologies. Assemblies revealed transcripts for approximately 20,000 - 30,000 genes from each species and assembly analyses led to the identification of candidate full-length prenyl transferase, TPS, and P450 genes of oleoresin biosynthesis. We cloned and functionally characterized, via expression of recombinant proteins in E. coli, nine different jack pine and eight different lodgepole pine mono-TPSs. The newly identified lodgepole pine and jack pine mono-TPSs include (+)-α-pinene synthases, (-)-α-pinene synthases, (-)-ß-pinene synthases, (+)-3-carene synthases, and (-)-ß-phellandrene synthases from each of the two species. CONCLUSION: In the absence of genome sequences, transcriptome assemblies are important for defence gene discovery in lodgepole pine and jack pine, as demonstrated here for the terpenoid pathway genes. The product profiles of the functionally annotated mono-TPSs described here can account for the major monoterpene metabolites identified in lodgepole pine and jack pine.

Draft genome of the mountain pine beetle, Dendroctonus ponderosae Hopkins, a major forest pest.

BACKGROUND: The mountain pine beetle, Dendroctonus ponderosae Hopkins, is the most serious insect pest of western North American pine forests. A recent outbreak destroyed more than 15 million hectares of pine forests, with major environmental effects on forest health, and economic effects on the forest industry. The outbreak has in part been driven by climate change, and will contribute to increased carbon emissions through decaying forests. RESULTS: We developed a genome sequence resource for the mountain pine beetle to better understand the unique aspects of this insect's biology. A draft de novo genome sequence was assembled from paired-end, short-read sequences from an individual field-collected male pupa, and scaffolded using mate-paired, short-read genomic sequences from pooled field-collected pupae, paired-end short-insert whole-transcriptome shotgun sequencing reads of mRNA from adult beetle tissues, and paired-end Sanger EST sequences from various life stages. We describe the cytochrome P450, glutathione S-transferase, and plant cell wall-degrading enzyme gene families important to the survival of the mountain pine beetle in its harsh and nutrient-poor host environment, and examine genome-wide single-nucleotide polymorphism variation. A horizontally transferred bacterial sucrose-6-phosphate hydrolase was evident in the genome, and its tissue-specific transcription suggests a functional role for this beetle. CONCLUSIONS: Despite Coleoptera being the largest insect order with over 400,000 described species, including many agricultural and forest pest species, this is only the second genome sequence reported in Coleoptera, and will provide an important resource for the Curculionoidea and other insects.

Transcriptome and full-length cDNA resources for the mountain pine beetle, Dendroctonus ponderosae Hopkins, a major insect pest of pine forests.

Bark beetles (Coleoptera: Curculionidae: Scolytinae) are major insect pests of many woody plants around the world. The mountain pine beetle (MPB), Dendroctonus ponderosae Hopkins, is a significant historical pest of western North American pine forests. It is currently devastating pine forests in western North America--particularly in British Columbia, Canada--and is beginning to expand its host range eastward into the Canadian boreal forest, which extends to the Atlantic coast of North America. Limited genomic resources are available for this and other bark beetle pests, restricting the use of genomics-based information to help monitor, predict, and manage the spread of these insects. To overcome these limitations, we generated comprehensive transcriptome resources from fourteen full-length enriched cDNA libraries through paired-end Sanger sequencing of 100,000 cDNA clones, and single-end Roche 454 pyrosequencing of three of these cDNA libraries. Hybrid de novo assembly of the 3.4 million sequences resulted in 20,571 isotigs in 14,410 isogroups and 246,848 singletons. In addition, over 2300 non-redundant full-length cDNA clones putatively containing complete open reading frames, including 47 cytochrome P450s, were sequenced fully to high quality. This first large-scale genomics resource for bark beetles provides the relevant sequence information for gene discovery; functional and population genomics; comparative analyses; and for future efforts to annotate the MPB genome. These resources permit the study of this beetle at the molecular level and will inform research in other Dendroctonus spp. and more generally in the Curculionidae and other Coleoptera.

Genome sequence of Mycoplasma capricolum subsp. capripneumoniae strain M1601.

Mycoplasma capricolum subsp. capripneumoniae is the causative agent of contagious caprine pleuropneumonia, a devastating disease of goats listed by the World Organization for Animal Health. Here we report the first complete genome sequence of this organism (strain M1601, a clinically isolated strain from China).

Updated genome assembly and annotation of Paenibacillus larvae, the agent of American foulbrood disease of honey bees.

BACKGROUND: As scientists continue to pursue various 'omics-based research, there is a need for high quality data for the most fundamental 'omics of all: genomics. The bacterium Paenibacillus larvae is the causative agent of the honey bee disease American foulbrood. If untreated, it can lead to the demise of an entire hive; the highly social nature of bees also leads to easy disease spread, between both individuals and colonies. Biologists have studied this organism since the early 1900s, and a century later, the molecular mechanism of infection remains elusive. Transcriptomics and proteomics, because of their ability to analyze multiple genes and proteins in a high-throughput manner, may be very helpful to its study. However, the power of these methodologies is severely limited without a complete genome; we undertake to address that deficiency here. RESULTS: We used the Illumina GAIIx platform and conventional Sanger sequencing to generate a 182-fold sequence coverage of the P. larvae genome, and assembled the data using ABySS into a total of 388 contigs spanning 4.5 Mbp. Comparative genomics analysis against fully-sequenced soil bacteria P. JDR2 and P. vortex showed that regions of poor conservation may contain putative virulence factors. We used GLIMMER to predict 3568 gene models, and named them based on homology revealed by BLAST searches; proteases, hemolytic factors, toxins, and antibiotic resistance enzymes were identified in this way. Finally, mass spectrometry was used to provide experimental evidence that at least 35% of the genes are expressed at the protein level. CONCLUSIONS: This update on the genome of P. larvae and annotation represents an immense advancement from what we had previously known about this species. We provide here a reliable resource that can be used to elucidate the mechanism of infection, and by extension, more effective methods to control and cure this widespread honey bee disease.

Genome and transcriptome analyses of the mountain pine beetle-fungal symbiont Grosmannia clavigera, a lodgepole pine pathogen.

In western North America, the current outbreak of the mountain pine beetle (MPB) and its microbial associates has destroyed wide areas of lodgepole pine forest, including more than 16 million hectares in British Columbia. Grosmannia clavigera (Gc), a critical component of the outbreak, is a symbiont of the MPB and a pathogen of pine trees. To better understand the interactions between Gc, MPB, and lodgepole pine hosts, we sequenced the ∼30-Mb Gc genome and assembled it into 18 supercontigs. We predict 8,314 protein-coding genes, and support the gene models with proteome, expressed sequence tag, and RNA-seq data. We establish that Gc is heterothallic, and report evidence for repeat-induced point mutation. We report insights, from genome and transcriptome analyses, into how Gc tolerates conifer-defense chemicals, including oleoresin terpenoids, as they colonize a host tree. RNA-seq data indicate that terpenoids induce a substantial antimicrobial stress in Gc, and suggest that the fungus may detoxify these chemicals by using them as a carbon source. Terpenoid treatment strongly activated a ∼100-kb region of the Gc genome that contains a set of genes that may be important for detoxification of these host-defense chemicals. This work is a major step toward understanding the biological interactions between the tripartite MPB/fungus/forest system.

Gene discovery for the bark beetle-vectored fungal tree pathogen Grosmannia clavigera.

BACKGROUND: Grosmannia clavigera is a bark beetle-vectored fungal pathogen of pines that causes wood discoloration and may kill trees by disrupting nutrient and water transport. Trees respond to attacks from beetles and associated fungi by releasing terpenoid and phenolic defense compounds. It is unclear which genes are important for G. clavigera's ability to overcome antifungal pine terpenoids and phenolics. RESULTS: We constructed seven cDNA libraries from eight G. clavigera isolates grown under various culture conditions, and Sanger sequenced the 5' and 3' ends of 25,000 cDNA clones, resulting in 44,288 high quality ESTs. The assembled dataset of unique transcripts (unigenes) consists of 6,265 contigs and 2,459 singletons that mapped to 6,467 locations on the G. clavigera reference genome, representing ~70% of the predicted G. clavigera genes. Although only 54% of the unigenes matched characterized proteins at the NCBI database, this dataset extensively covers major metabolic pathways, cellular processes, and genes necessary for response to environmental stimuli and genetic information processing. Furthermore, we identified genes expressed in spores prior to germination, and genes involved in response to treatment with lodgepole pine phloem extract (LPPE). CONCLUSIONS: We provide a comprehensively annotated EST dataset for G. clavigera that represents a rich resource for gene characterization in this and other ophiostomatoid fungi. Genes expressed in response to LPPE treatment are indicative of fungal oxidative stress response. We identified two clusters of potentially functionally related genes responsive to LPPE treatment. Furthermore, we report a simple method for identifying contig misassemblies in de novo assembled EST collections caused by gene overlap on the genome.

A regulatory toolbox of MiniPromoters to drive selective expression in the brain.

The Pleiades Promoter Project integrates genomewide bioinformatics with large-scale knockin mouse production and histological examination of expression patterns to develop MiniPromoters and related tools designed to study and treat the brain by directed gene expression. Genes with brain expression patterns of interest are subjected to bioinformatic analysis to delineate candidate regulatory regions, which are then incorporated into a panel of compact human MiniPromoters to drive expression to brain regions and cell types of interest. Using single-copy, homologous-recombination "knockins" in embryonic stem cells, each MiniPromoter reporter is integrated immediately 5' of the Hprt locus in the mouse genome. MiniPromoter expression profiles are characterized in differentiation assays of the transgenic cells or in mouse brains following transgenic mouse production. Histological examination of adult brains, eyes, and spinal cords for reporter gene activity is coupled to costaining with cell-type-specific markers to define expression. The publicly available Pleiades MiniPromoter Project is a key resource to facilitate research on brain development and therapies.

Salmo salar and Esox lucius full-length cDNA sequences reveal changes in evolutionary pressures on a post-tetraploidization genome.

BACKGROUND: Salmonids are one of the most intensely studied fish, in part due to their economic and environmental importance, and in part due to a recent whole genome duplication in the common ancestor of salmonids. This duplication greatly impacts species diversification, functional specialization, and adaptation. Extensive new genomic resources have recently become available for Atlantic salmon (Salmo salar), but documentation of allelic versus duplicate reference genes remains a major uncertainty in the complete characterization of its genome and its evolution. RESULTS: From existing expressed sequence tag (EST) resources and three new full-length cDNA libraries, 9,057 reference quality full-length gene insert clones were identified for Atlantic salmon. A further 1,365 reference full-length clones were annotated from 29,221 northern pike (Esox lucius) ESTs. Pairwise dN/dS comparisons within each of 408 sets of duplicated salmon genes using northern pike as a diploid out-group show asymmetric relaxation of selection on salmon duplicates. CONCLUSIONS: 9,057 full-length reference genes were characterized in S. salar and can be used to identify alleles and gene family members. Comparisons of duplicated genes show that while purifying selection is the predominant force acting on both duplicates, consistent with retention of functionality in both copies, some relaxation of pressure on gene duplicates can be identified. In addition, there is evidence that evolution has acted asymmetrically on paralogs, allowing one of the pair to diverge at a faster rate.

De novo genome sequence assembly of a filamentous fungus using Sanger, 454 and Illumina sequence data.

Sequencing-by-synthesis technologies can reduce the cost of generating de novo genome assemblies. We report a method for assembling draft genome sequences of eukaryotic organisms that integrates sequence information from different sources, and demonstrate its effectiveness by assembling an approximately 32.5 Mb draft genome sequence for the forest pathogen Grosmannia clavigera, an ascomycete fungus. We also developed a method for assessing draft assemblies using Illumina paired end read data and demonstrate how we are using it to guide future sequence finishing. Our results demonstrate that eukaryotic genome sequences can be accurately assembled by combining Illumina, 454 and Sanger sequence data.

Generation of ESTs in Vitis vinifera wine grape (Cabernet Sauvignon) and table grape (Muscat Hamburg) and discovery of new candidate genes with potential roles in berry development.

We report the generation and analysis of a total of 77,583 expressed sequence tags (ESTs) from two grapevine (Vitis vinifera L.) cultivars, Cabernet Sauvignon (wine grape) and Muscat Hamburg (table grape) with a focus on EST sequence quality and assembly optimization. The majority of the ESTs were derived from normalized cDNA libraries representing berry pericarp and seed developmental series, pooled non-berry tissues including root, flower, and leaf in Cabernet Sauvignon, and pooled tissues of berry, seed, and flower in Muscat Hamburg. EST and unigene sequence quality were determined by computational filtering coupled with small-scale contig reassembly, manual review, and BLAST analyses. EST assembly was optimized to better discriminate among closely related paralogs using two independent grape sequence sets, a previously published set of Vitis spp. gene families and our EST dataset derived from pooled leaf, flower, and root tissues of Cabernet Sauvignon. Sequence assembly within individual libraries indicated that those prepared from pooled tissues contributed the most to gene discovery. Annotations based upon searches against multiple databases including tomato and strawberry sequences helped to identify putative functions of ESTs and unigenes, particularly with respect to fleshy fruit development. Sequence comparison among the three wine grape libraries identified a number of genes preferentially expressed in the pericarp tissue, including transcription factors, receptor-like protein kinases, and hexose transporters. Gene ontology (GO) classification in the biological process aspect showed that GO categories corresponding to 'transport' and 'cell organization and biogenesis', which are associated with metabolite movement and cell wall structural changes during berry ripening, were higher in pericarp than in other tissues in the wine grape studied. The sequence data were used to characterize potential roles of new genes in berry development and composition.