Next-generation sequencing to genetically diagnose a diverse range of inherited eye disorders in 15 consanguineous families from Pakistan.

Inherited retinal dystrophies (IRDs) are characterized by photoreceptor dysfunction or degeneration. Clinical and phenotypic overlap between IRDs makes the genetic diagnosis very challenging and comprehensive genomic approaches for accurate diagnosis are frequently required. While there are previous studies on IRDs in Pakistan, causative genes and variants are still unknown for a significant portion of patients. Therefore, there is a need to expand the knowledge of the genetic spectrum of IRDs in Pakistan. Here, we recruited 52 affected and 53 normal individuals from 15 consanguineous Pakistani families presenting non-syndromic and syndromic forms of IRDs. We employed single molecule Molecular Inversion Probes (smMIPs) based panel sequencing and whole genome sequencing to identify the probable disease-causing variants in these families. Using this approach, we obtained a 93% genetic solve rate and identified 16 (likely) causative variants in 14 families, of which seven novel variants were identified in ATOH7, COL18A1, MERTK, NDP, PROM1, PRPF8 and USH2A while nine recurrent variants were identified in CNGA3, CNGB1, HGSNAT, NMNAT1, SIX6 and TULP1. The novel MERTK variant and one recurrent TULP1 variant explained the intra-familial locus heterogeneity in one of the screened families while two recurrent CNGA3 variants explained compound heterozygosity in another family. The identification of variants in known disease-associated genes emphasizes the utilization of time and cost-effective screening approaches for rapid diagnosis. The timely genetic diagnosis will not only identify any associated systemic issues in case of syndromic IRDs, but will also aid in the acceleration of personalized medicine for patients affected with IRDs.

Combined Single Gene Testing and Genome Sequencing as an Effective Diagnostic Approach for Anophthalmia and Microphthalmia Patients.

Anophthalmia and microphthalmia (A/M) are among the most severe congenital developmental eye disorders. Despite the advancements in genome screening technologies, more than half of A/M patients do not receive a molecular diagnosis. We included seven consanguineous families affected with A/M from Pakistani cohort and an unknown molecular basis. Single gene testing of FOXE3 was performed, followed by genome sequencing for unsolved probands in order to establish a genetic diagnosis for these families. All seven families were provided with a genetic diagnosis. The identified variants were all homozygous, classified as (likely) pathogenic and present in an A/M-associated gene. Targeted FOXE3 sequencing revealed two previously reported pathogenic FOXE3 variants in four families. In the remaining families, genome sequencing revealed a known pathogenic PXDN variant, a novel 13bp deletion in VSX2, and one novel deep intronic splice variant in PXDN. An in vitro splice assay was performed for the PXDN splice variant which revealed a severe splicing defect. Our study confirmed the utility of genome sequencing as a diagnostic tool for A/M-affected individuals. Furthermore, the identification of a novel deep intronic pathogenic variant in PXDN highlights the role of non-coding variants in A/M-disorders and the value of genome sequencing for the identification of this type of variants.

Comparison of fine needle aspiration cytology and thyroid scan in solitary thyroid nodule.

Objective. This was a comparative study between FNAC and thyroid scan used to diagnose the solitary thyroid nodule and histopathology was used as gold standard to compare the results of both modalities. We hypothesized that Fine needle aspiration cytology and thyroid scan diagnose solitary thyroid nodule (STN) as accurately as histopathology. Materials and Methods. This study comprised of 50 patients with solitary thyroid nodules (STN) presented to OPD. After clinical examination these patients were referred to Centre for Nuclear Medicine, Mayo Hospital Lahore for thyroid function tests and thyroid scan (TS). These patients underwent FNAC in the department of Pathology and surgery in Mayo Hospital. The cases were operated and evaluated for histopathological changes. Results. On thyroid scan, 40 patients (80%) having cold nodule were labeled as suspicious 10 patients (20%) had hot nodule. On FNAC 23 patients (46%) had benign lesion, 22 patients (44%) had indeterminate lesion and 5 patients (10%) had malignant lesions. On histopathology, 45 patients (90%) were confirmed to have benign lesions and 5 patients (10%), malignant lesions. After comparison of results of thyroid scan and FNAC with histopathology, the sensitivity, specificity, positive predictive value, negative predictive value and diagnostic accuracy of thyroid scan were 80%, 20%, 10%, 90% and 26%, respectively whereas those of FNAC were 80%, 97.7%, 80%, 97.7% and 96%, respectively. Conclusion. Fine needle aspiration was a significantly better predictor of malignancy than thyroid scan and resulted in a smaller proportion of excisions for benign nodules.