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Purpose: Current sequencing strategies can genetically solve 55-60% of inherited retinal degeneration (IRD) cases, despite recent progress in sequencing. This can partially be attributed to elusive pathogenic variants (PVs) in known IRD genes, including copy-number variations (CNVs), which have been shown as major contributors to unsolved IRD cases.Methods: Five hundred IRD patients were analyzed with targeted next-generation sequencing (NGS). The NGS data were used to detect CNVs with ExomeDepth and gCNV and the results were compared with CNV detection with a single-nucleotide polymorphism (SNP) array. Likely causal CNV predictions were validated by quantitative polymerase chain reaction (qPCR).Results: Likely disease-causing single-nucleotide variants (SNVs) and small indels were found in 55.6% of subjects. PVs in USH2A (11.6%), RPGR (4%), and EYS (4%) were the most common. Likely causal CNVs were found in an additional 8.8% of patients. Of the three CNV detection methods, gCNV showed the highest accuracy. Approximately 30% of unsolved subjects had a single likely PV in a recessive IRD gene.Conclusion: CNV detection using NGS-based algorithms is a reliable method that greatly increases the genetic diagnostic rate of IRDs. Experimentally validating CNVs helps estimate the rate at which IRDs might be solved by a CNV plus a more elusive variant. (2020) 22: Genetics in Medicine

Pericentral retinitis pigmentosa (RP) is an atypical form of RP that affects the near-peripheral retina first and tends to spare the far periphery. This study was performed to further define the genetic basis of this phenotype. We identified a cohort of 43 probands with pericentral RP based on a comprehensive analysis of their retinal phenotype. Genetic analyses of DNA samples from these patients were performed using panel-based next-generation sequencing, copy number variations, and whole exome sequencing (WES). Mutations provisionally responsible for disease were found in 19 of the 43 families (44%) analyzed. These include mutations in RHO (five patients), USH2A (four patients), and PDE6B (two patients). Of 28 putatively pathogenic alleles, 15 (54%) have been previously identified in patients with more common forms of typical RP, while the remaining 13 mutations (46%) were novel. Burden testing of WES data successfully identified HGSNAT as a cause of pericentral RP in at least two patients, suggesting it is also a relatively common cause of pericentral RP. While additional sequencing might uncover new genes specifically associated with pericentral RP, the current results suggest that genetically pericentral RP is not a separate clinical entity, but rather is part of the spectrum of mild RP phenotypes.

In Mendelian disease diagnosis, variant analysis is a repetitive, error-prone, and time consuming process. To address this, we have developed the Mendelian Analysis Toolkit (MATK), a configurable, automated variant ranking program. Methods: MATK aggregates variant information from multiple annotation sources and uses expert-designed rules with parameterized weights to produce a ranked list of potentially causal solutions. MATK performance was measured by a comparison between MATK-aided and human-domain expert analyses of 1060 families with inherited retinal degeneration (IRD), analyzed using an IRD-specific gene panel (589 individuals) and exome sequencing (471 families). Results: When comparing MATK-assisted analysis with expert curation in both the IRDspecific gene panel and exome sequencing (1060 subjects), 97.3% of potential solutions found by experts were also identified by the MATK-assisted analysis (541 solutions identified with MATK of 556 solutions found by conventional analysis). Furthermore, MATK-assisted analysis identified 114 additional potential solutions from the 504 cases unsolved by conventional analysis. Conclusion: MATK expedites the process of identification of likely solving variants in Mendelian traits, and reduces variability stemming from human error and researcher bias. MATK facilitates data reanalysis to keep up with the constantly improving annotation sources and nextgeneration sequencing processing pipelines. The software is open source and available at https:// gitlab.com/matthew_maher/mendelanalysis.

Unstable blood pressure after spinal cord injury (SCI) is not routinely examined but rather predicted by level and completeness of injury (i.e., American Spinal Injury Association Impairment Scale AIS classification). Our aim was to investigate hemodynamic response to a sit‐up test in a large cohort of individuals with chronic SCI to better understand cardiovascular function in this population. Continuous blood pressure and ECG were recorded from individuals with SCI (n = 159) and non‐injured individuals (n = 48). We found orthostatic hypotension occurred within each level and AIS classification (n = 36). Moreover, 45 individuals with chronic SCI experienced a drop in blood pressure that did not meet the criteria for orthostatic hypotension, but was accompanied by dramatic increases in heart rate, reflecting orthostatic intolerance. A cluster analysis of hemodynamic response to a seated position identified eight distinct patterns of interaction between blood pressure and heart rate during orthostatic stress indicating varied autonomic responses. Algorithmic cluster analysis of heart rate and blood pressure is more sensitive to diagnosing orthostatic cardiovascular dysregulation. This indicates blood pressure instability cannot be predicted by level and completeness of SCI, and the consensus statement definition of orthostatic hypotension is insufficient to characterize the variability of blood pressure and heart rate responses during orthostatic stress. Both blood pressure and heart rate responses are needed to characterize autonomic function after SCI.

Purpose: In Mendelian disease diagnosis, variant analysis is a repetitive, error-prone, and time-consuming process. To address this, we have developed the Mendelian Analysis Toolkit (MATK), a configurable automated variant ranking program. Methods: MATK aggregates variant information from multiple annotation sources and uses expert-designed rules with parameterized weights to produce a ranked list of potentially causal solutions. MATK performance was measured by a comparison of MATK-aided versus human domain-expert analyses of 1060 inherited retinal degeneration (IRD) families investigated with an IRD-specific gene panel (589 families) and exome sequencing (471 families). Results: When comparing MATK-assisted analysis to expert curation, we found that 97.3% (541/556) of potential solutions found by experts were also identified by the MATK-assisted analysis. Furthermore, MATK-assisted analysis identified 114 additional potential solutions. The software also showed utility in data reanalysis after remapping to the GRCh38 genome build. Conclusion: MATK expedites the process of identifying likely solving variants in Mendelian traits and helps to remove variability coming from human error and researcher bias. MATK facilitates data re-analysis to keep up with the constantly improving annotation sources and NGS processing pipelines. The software is open source and available at https://gitlab.partners.org/meei-ogi-bioinformatics/MendelAnalysis

Transcutaneous spinal cord stimulation (TSCS) has demonstrated potential to beneficially modulate spinal cord motor and autonomic circuitry. We are interested in pairing cervical TSCS with other forms of nervous system stimulation to enhance synaptic plasticity in circuits serving hand function. We use a novel configuration for cervical TSCS in which the anode is placed anteriorly over ~C4–C5 and the cathode posteriorly over ~T2–T4. We measured the effects of single pulses of TSCS paired with single pulses of motor cortex or median nerve stimulation timed to arrive at the cervical spinal cord at varying intervals. In 13 participants with and 15 participants without chronic cervical spinal cord injury, we observed that subthreshold TSCS facilitates hand muscle responses to motor cortex stimulation, with a tendency toward greater facilitation when TSCS is timed to arrive at cervical synapses simultaneously or up to 10 milliseconds after cortical stimulus arrival. Single pulses of subthreshold TSCS had no effect on the amplitudes of median H-reflex responses or F-wave responses. These findings support a model in which TSCS paired with appropriately timed cortical stimulation has the potential to facilitate convergent transmission between descending motor circuits, segmental afferents, and spinal motor neurons serving the hand. Studies with larger numbers of participants and repetitively paired cortical and spinal stimulation are needed.

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