Genome-wide association study of soybean (Glycine max [L.] Merr.) germplasm for dissecting the quantitative trait nucleotides and candidate genes underlying yield-related traits.

Soybean (Glycine max [L.] Merr.) is one of the most significant crops in the world in terms of oil and protein. Owing to the rising demand for soybean products, there is an increasing need for improved varieties for more productive farming. However, complex correlation patterns among quantitative traits along with genetic interactions pose a challenge for soybean breeding. Association studies play an important role in the identification of accession with useful alleles by locating genomic sites associated with the phenotype in germplasm collections. In the present study, a genome-wide association study was carried out for seven agronomic and yield-related traits. A field experiment was conducted in 2015/2016 at two locations that include 155 diverse soybean germplasm. These germplasms were genotyped using SoySNP50K Illumina Infinium Bead-Chip. A total of 51 markers were identified for node number, plant height, pods per plant, seeds per plant, seed weight per plant, hundred-grain weight, and total yield using a multi-locus linear mixed model (MLMM) in FarmCPU. Among these significant SNPs, 18 were putative novel QTNs, while 33 co-localized with previously reported QTLs. A total of 2,356 genes were found in 250 kb upstream and downstream of significant SNPs, of which 17 genes were functional and the rest were hypothetical proteins. These 17 candidate genes were located in the region of 14 QTNs, of which ss715580365, ss715608427, ss715632502, and ss715620131 are novel QTNs for PH, PPP, SDPP, and TY respectively. Four candidate genes, Glyma.01g199200, Glyma.10g065700, Glyma.18g297900, and Glyma.14g009900, were identified in the vicinity of these novel QTNs, which encode lsd one like 1, Ergosterol biosynthesis ERG4/ERG24 family, HEAT repeat-containing protein, and RbcX2, respectively. Although further experimental validation of these candidate genes is required, several appear to be involved in growth and developmental processes related to the respective agronomic traits when compared with their homologs in Arabidopsis thaliana. This study supports the usefulness of association studies and provides valuable data for functional markers and investigating candidate genes within a diverse germplasm collection in future breeding programs.

Genetic diversity and population structure analysis in cultivated soybean (Glycine max [L.] Merr.) using SSR and EST-SSR markers.

Soybean (Glycine max) is an important legume that is used to fulfill the need of protein and oil of large number of population across the world. There are large numbers of soybean germplasm present in the USDA germplasm resources. Finding and understanding genetically diverse germplasm is a top priority for crop improvement programs. The current study used 20 functional EST-SSR and 80 SSR markers to characterize 96 soybean accessions from diverse geographic backgrounds. Ninety-six of the 100 markers were polymorphic, with 262 alleles (average 2.79 per locus). The molecular markers had an average polymorphic information content (PIC) value of 0.44, with 28 markers ≥ 0.50. The average major allele frequency was 0.57. The observed heterozygosity of the population ranged from 0-0.184 (average 0.02), while the expected heterozygosity ranged from 0.20-0.73 (average 0.51). The lower value for observed heterozygosity than expected heterozygosity suggests the likelihood of a population structure among the germplasm. The phylogenetic analysis and principal coordinate analysis (PCoA) divided the total population into two major groups (G1 and G2), with G1 comprising most of the USA lines and the Australian and Brazilian lines. Furthermore, the phylogenetic analysis and PCoA divided the USA lines into three major clusters without any specific differentiation, supported by the model-based STRUCTURE analysis. Analysis of molecular variance (AMOVA) showed 94% variation among individuals in the total population, with 2% among the populations. For the USA lines, 93% of the variation occurred among individuals, with only 2% among lines from different US states. Pairwise population distance indicated more similarity between the lines from continental America and Australia (189.371) than Asia (199.518). Overall, the 96 soybean lines had a high degree of genetic diversity.

Analysis of genotype × environment interactions for agronomic traits of soybean (Glycine max [L.] Merr.) using association mapping.

The soybean yield is a complex quantitative trait that is significantly influenced by environmental factors. G × E interaction (GEI), which derives the performance of soybean genotypes differentially in various environmental conditions, is one of the main obstacles to increasing the net production. The primary goal of this study is to identify the outperforming genotypes in different latitudes, which can then be used in future breeding programs. A total of 96 soybean genotypes were examined in two different ecological regions: Faisalabad and Tando Jam in Pakistan. The evaluation of genotypes in different environmental conditions showed a substantial amount of genetic diversity for grain yield. We identified 13 environment-specific genotypes showing their maximum grain yield in each environment. Genotype G69 was found to be an ideal genotype with higher grain yield than other genotypes tested in this study and is broadly adapted for environments E1 and E2 and also included in top-yielding genotypes in E3, E4, and E5. G92 is another genotype that is broadly adapted in E1, E3, and E4. In the case of environments, E3 is suggested to be a more ideal environment as it is plotted near the concentric circle and is very informative for the selection of genotypes with high yield. Despite the presence of GEI, advances in DNA technology provided very useful tools to investigate the insight of advanced genotypes. Association mapping is a useful method for swiftly and efficiently investigating the genetic basis of significant plant traits. A total of 26 marker-trait associations were found for six agronomic traits in five environments, with the highest significance (p-value = 2.48 × 10-08) for plant height and the lowest significance (1.03 × 10-03) for hundred-grain weight. Soybean genotypes identified in the present study could be a valuable source for future breeding programs as they are adaptable to a wide range of environments. Genetic selection of genotypes with the best yields can be used for gross grain production in a wide range of climatic conditions, and it would give an essential reference in terms of soybean variety selection.

Procedures, Visits, and Procedure Costs in the Management of Microbial Keratitis.

PURPOSE: To investigate the burden of procedures, visits, and procedure costs in the management of microbial keratitis (MK). METHODS: Medical records of patients from an academic hospital outpatient facility between December 2013 and May 2018 were retrospectively reviewed. Patients were included if they were older than the age of 18 years, recruited for study of likely MK, and not concurrently undergoing treatment for other acute eye conditions. For procedural costs, Medicare data for billing were obtained using the Center for Medicare and Medicaid Services Physician Fee Lookup tool. RESULTS: A total of 68 patients were included for analysis. Patients were on average 51.3 years (SD = 19.5), 55.9% women (n = 38), and 89.7% White (n = 61). Per person, the average number of procedures was 2.9 (SD = 4.2). The average number of visits was 13.9 (SD = 9.2) over an average of 26.9 weeks (SD = 24.3). Age (P < 0.0001), positive Gram stain (P = 0.03), and mixed Gram stain (P = 0.002) were positively associated with the number of procedures. Age (P = 0.0003), fungal keratitis (P = 0.02), and mixed Gram stain (P = 0.01) were positively associated with the number of visits. Race was inversely associated with the number of procedures (P = 0.045) and visits (0.03). Patients with bacterial keratitis were more likely to have amniotic membrane grafts (P = 0.01) and tarsorrhaphies (P = 0.03) than fungal patients. Across all procedures performed for the management of MK, the mean cost per patient was $1788.7 (SD = $3324.62). CONCLUSIONS: Patients incur many procedural costs and attend many visits during the management of MK. These findings emphasize the importance of patient-provider communication for frequent follow-up care and the potential need to perform procedures for disease management.

Measurement Reliability for Keratitis Morphology.

PURPOSE: To evaluate the reliability of manual annotation when quantifying cornea anatomical and microbial keratitis (MK) morphological features on slit-lamp photography (SLP) images. METHODS: Prospectively enrolled patients with MK underwent SLP at initial encounter at 2 academic eye hospitals. Patients who presented with an epithelial defect (ED) were eligible for analysis. Features, which included ED, corneal limbus (L), pupil (P), stromal infiltrate (SI), white blood cell (WBC) infiltration at the SI edge, and hypopyon (H), were annotated independently by 2 physicians on SLP images. Intraclass correlation coefficients (ICCs) were applied for reliability assessment; dice similarity coefficients (DSCs) were used to investigate the area overlap between readers. RESULTS: Seventy-five MK patients with an ED received SLP. DSCs indicate good to fair annotation overlap between graders (L = 0.97, P = 0.80, ED = 0.94, SI = 0.82, H = 0.82, WBC = 0.83) and between repeat annotations by the same grader (L = 0.97, P = 0.81, ED = 0.94, SI = 0.85, H = 0.84, WBC = 0.82). ICC scores showed good intergrader (L = 0.98, P = 0.78, ED = 1.00, SI = 0.67, H = 0.97, WBC = 0.86) and intragrader (L = 0.99, P = 0.92, ED = 0.99, SI = 0.94, H = 0.99, WBC = 0.92) reliabilities. When reliability statistics were recalculated for annotated SI area in the subset of cases where both graders agreed WBC infiltration was present/absent, intergrader ICC improved to 0.91 and DSC improved to 0.86 and intragrader ICC remained the same, whereas DSC improved to 0.87. CONCLUSIONS: Manual annotation indicates usefulness of area quantification in the evaluation of MK. However, variability is intrinsic to the task. Thus, there is a need for optimization of annotation protocols. Future directions may include using multiple annotators per image or automated annotation software.

Algorithm Variability in Quantification of Epithelial Defect Size in Microbial Keratitis Images.

PURPOSE: To investigate the sources of measurement variability when quantifying the morphology of microbial keratitis (MK) from slit-lamp photography (SLP) images using a semiautomated, image-analysis algorithm. METHODS: Prospectively enrolled patients with MK underwent SLP to obtain images of their epithelial defects (ED). Eyes were stained with fluorescein and imaged multiple times under blue light, at low and high magnifications. A masked research assistant chose the 3 best images and annotated each 3 times to provide seed regions corresponding to ED and healthy cornea. The algorithm returned the ED area for each seeded image. Eyes without EDs and algorithm failures were excluded. Variance components were estimated with a random effects model and intraclass correlation coefficients estimated with intragrader reliability. RESULTS: A total of 42 eyes from 42 MK participants were photographed. After excluding poor quality images, eyes with no EDs, and algorithm failures, 34 patients with 92 images and 274 seeds were analyzed. No significant differences in the average ED area were found between seedings or high- versus low-SLP magnifications (all P > 0.5, paired t tests). Minimal measurement variability was because of image (0.9%), magnification (0.2%), or seed (0.1%). Most variability was attributable to differences in ED sizes between patients (85.2%). 13.7% of variability was unexplained. Multiple iterations of the algorithm on the same image showed good consistency (intraclass correlation coefficient = 0.98, 95% confidence interval, 0.97-0.99). CONCLUSIONS: Image-analysis algorithms showed good reliability for measuring the ED area from SLP images. Most measurement variability was because of between-patient differences, not imaging settings or application of the algorithm by the user.

Ultrasonic debridement with stem cell therapy of suspensory branch desmitis in an equine patient.

Ultrasonic debridement as a treatment for tendinopathy and desmitis is a relatively new approach in orthopedic surgery. Previously only used in limited cases, this procedure shows promise for treating ligament-bone and tendon-bone interface injuries. We present a case study of a 2-yr-old thoroughbred male horse, unable to train due to recalcitrant symptoms after extensive conservative management of suspensory branch desmitis. It was then treated with ultrasonic debridement and concurrent manubrial stem cell autograft injection, to treat the ultrasound visualized lesion. Post-surgically, the patient recovered quickly, began training within 16 wk, and went onto win several races. Repeat ultrasound imaging reveals a complete restoration of the internal fiber architecture of the ligament. With a 3-yr follow-up, there has been consistent training and race performance with no re-injury. This study is the first to document the successful outcome of ultrasonic debridement with concurrent stem cell injection in the treatment of equine desmitis.

Medication Accuracy in Electronic Health Records for Microbial Keratitis.

IMPORTANCE: Electronic health records (EHRs) contain an abundance of health information. However, researchers need to understand data accuracy to ask appropriate research questions. OBJECTIVE: To investigate the concordance of the names of medications for microbial keratitis in the structured, formal EHR medication list and the text of clinicians' progress notes. DESIGN, SETTING, AND PARTICIPANTS: This cross-sectional study, conducted in the cornea section of an ophthalmology department in a tertiary care, referral academic medical center, examined the medications of 53 patients with microbial keratitis treated until disease resolution from July 1, 2015, to August 1, 2018. Documentation of medications was compared between the structured medication list extracted from the EHR server and medications written into the clinical progress note and transcribed by the study team. EXPOSURE: Medication treatment for microbial keratitis. MAIN OUTCOMES AND MEASURES: Medication mismatch frequency. RESULTS: The study sample included 24 men and 29 women, with a mean (SD) age of 51.8 (19.6) years. Of the 247 medications identified, 57 (23.1%) of prescribed medications differed between the progress notes and the formal EHR-based medication list. Reasons included medications not prescribed via the EHR ordering system (25 [43.9%]), outside medications not reconciled in the internal EHR medication list (23 [40.4%]), and medications prescribed via the EHR ordering system and in the formal list, but not described in the clinical note (9 [15.8%]). Fortified antimicrobials represented the largest category for medication mismatch between modalities (17 of 70 [24.3%]). Nearly one-third of patients (17 [32.1%]) had at least 1 medication mismatch in their record. CONCLUSIONS AND RELEVANCE: Almost 1 in 4 medications were mismatched between the progress note and formal medication list in the EHR. These findings suggest that EHR data should be checked for internal consistency before use in research.