Mutations in known disease genes account for the majority of autosomal recessive retinal dystrophies.

Retinal dystrophies (RDs) are hereditary blinding eye conditions that are highly variable in their clinical presentation. The remarkable genetic heterogeneity that characterizes RD was a major challenge in establishing the molecular diagnosis in these patients until the recent advent of next-generation sequencing. It remains unclear, however, what percentage of autosomal recessive RD remain undiagnosed when all established RD genes are sequenced. We enrolled 75 families in which RD segregates in an apparently autosomal recessive manner. We show that the yield of a multigene panel that contains known RD genes is 67.5%. The higher yield (82.3%) when whole exome sequencing was implemented instead was often due to hits in genes that were not included in the original design of the panel. We also show the value of homozygosity mapping even during the era of exome sequencing in uncovering cryptic mutations. In total, we describe 45 unique likely deleterious variants (of which 18 are novel including one deep intronic and one genomic deletion mutation). Our study suggests that the genetic heterogeneity of autosomal recessive RD is approaching saturation and that any new RD genes will probably account for only a minor role in the mutation burden.

Genetic investigation of 93 families with microphthalmia or posterior microphthalmos.

Microphthalmia is a developmental eye defect that is highly variable in severity and in its potential for systemic association. Despite the discovery of many disease genes in microphthalmia, at least 50% of patients remain undiagnosed genetically. Here, we describe a cohort of 147 patients (93 families) from our highly consanguineous population with various forms of microphthalmia (including the distinct entity of posterior microphthalmos) that were investigated using a next-generation sequencing multi-gene panel (i-panel) as well as whole exome sequencing and molecular karyotyping. A potentially causal mutation was identified in the majority of the cohort with microphthalmia (61%) and posterior microphthalmos (82%). The identified mutations (55 point mutations, 15 of which are novel) spanned 24 known disease genes, some of which have not or only very rarely been linked to microphthalmia (PAX6, SLC18A2, DSC3 and CNKSR1). Our study has also identified interesting candidate variants in 2 genes that have not been linked to human diseases (MYO10 and ZNF219), which we present here as novel candidates for microphthalmia. In addition to revealing novel phenotypic aspects of microphthalmia, this study expands its allelic and locus heterogeneity and highlights the need for expanded testing of patients with this condition.

Genome-wide linkage and sequence analysis challenge CCDC66 as a human retinal dystrophy candidate gene and support a distinct NMNAT1-related fundus phenotype.

To uncover the genotype underlying early-onset cone-rod dystrophy and central nummular macular atrophic lesion in 2 siblings from an endogamous Arab family, we performed targeted next-generation sequencing (NGS) of 44 retinal dystrophy genes, whole-exome sequencing (WES) and genome-wide linkage analysis. Targeted NGS and WES in the index patient highlighted 2 homozygous variants, a CCDC66 frameshift deletion and a novel missense NMNAT1 variant, c.500G>A (p.Asn167Ser). Linkage and segregation analysis excluded the CCDC66 variant and confirmed the NMNAT1 mutation. Biallelic NMNAT1 mutations cause Leber congenital amaurosis with a central nummular macular atrophic lesion (LCA9). The NMNAT1 mutation reported here underlied cone-rod dystrophy rather than LCA but the fundus lesion was compatible with that of LCA9 patients, highlighting that such a fundus appearance should raise suspicion for biallelic mutations in NMNAT1 when in the context of any retinal dystrophy. Although Ccdc66 mutations have been proposed to cause retinal disease in dogs, our results and public databases challenge CCDC66 as a candidate gene for human retinal dystrophy.

Identification of three novel ECEL1 mutations in three families with distal arthrogryposis type 5D.

Arthrogryposis refers to congenital contracture in at least two different body parts. When distal joints are primarily involved, the term distal arthrogryposis (DA) is used. The recognition of clinically distinct subtypes of DA has proven very useful in mapping the disease genes for this genetically heterogeneous condition. DA5D is characterized by ocular involvement usually in the form of ptosis and incomitant strabismus, but extraocular manifestations have also been reported. In a multiplex consanguineous family with DA5D, we combined autozygosity mapping and exome sequencing to identify a novel mutation in ECEL1. This was followed by targeted sequencing of this gene in another two extended consanguineous family with the same phenotype, which revealed two additional novel homozygous mutations. Our results support the recent identification of mutations in ECEL1 as a disease gene in DA5D and expand the clinical and allelic spectrum of this condition.

Mutations in ALDH1A3 cause microphthalmia.

Microphthalmia is an important inborn error of eye development that can be associated with multisystem involvement. Anophthalmia is more severe and rarer. Single mutations in an expanding list of genes are known to cause this spectrum of anomaly. In one branch of a multiplex family with microphthalmia and anophthalmia, autozygome analysis excluded all known microphthalmia genes at the time of doing this study. Exome sequencing and autozygome filtration identified a novel homozygous variant in ALDH1A3. Subsequently, we identified another homozygous variant in 2 of the 10 probands with microphthalmia we specifically screened for mutations in ALDH1A3. Interestingly, the other branch of the original family was found to segregate anophthalmia/syndactyly with a novel homozygous SMOC1 variant. Our data support the very recent and independent identification of ALDH1A3 as a disease gene in microphthalmia. Locus heterogeneity should be considered in consanguineous families even for extremely rare phenotypes.

Identification of a novel CRYAB mutation associated with autosomal recessive juvenile cataract in a Saudi family.

PURPOSE: To describe the first cataract-causing recessive mutation in the crystalline, alpha-b gene CRYAB. METHODS: Homozygosity mapping complemented by linkage analysis was performed in a family with autosomal recessive juvenile cataract. RESULTS: A homozygous missense mutation in CRYAB was identified. The mutation replaces a highly conserved amino acid residue in a dual function domain of the protein. None of the patients has clinically significant myopathy, but the oldest patient (the mother) has retinal pathology. CONCLUSIONS: This is the first report of a recessive mutation in CRYAB causing cataract. Based on recent knowledge of the structure and function of this small heat shock protein, we speculate on the potential mutational mechanism.

Comparison of polypropylene and silicone Ahmed valve survival 2 years following implantation in the first 2 years of life.

AIM: To compare the 2-year survival rates of polypropylene and silicone Ahmed glaucoma valves (AGVs) implanted during the first 2 years of life. METHODS: Retrospective analysis of polypropylene and silicone AGV implantation during the first 2 years of life in children with 2 years' postoperative follow-up. Patients with prior aqueous drainage device implantation were excluded. RESULTS: Forty-two eyes of 36 children were reviewed. Thirty-one eyes received a polypropylene AGV (six S1, 25 S2), and 11 eyes received a silicone AGV (11 FP7). The average survival time (maintenance of intraocular pressure < or =22 mm Hg with (or without) medications and without significant complications) was significantly longer (p = 0.001 by the logrank test) for the silicone group (23.36 months, standard error (SE) 1.64, 95% confidence interval (CI) 20.16 to 24.00 months) than for the polypropylene group (19.10 months, SE 1.53, 95% CI 16.1 to 22.12 months). Cumulative probabilities of survival at 2 years by Kaplan to Meier analysis were 90.9% (SE 8.7, 95% CI 70 to 100%) and 54.8% (SE 8.9, 95% CI 23 to 87%), respectively (p = 0.001 by the logrank test). All eyes implanted with silicone AGVs had the diagnosis of congenital glaucoma, which was independently associated with 2-year survival. CONCLUSIONS: Two years after surgery, silicone AGVs survived longer than polypropylene AGVs. However, all eyes that received silicone AGVs in our cohort had congenital glaucoma, which had a better survival than other paediatric glaucoma diagnoses. A study of silicone AGV implantation in other paediatric glaucoma diagnoses is needed to determine whether or not silicone AGVs independently have a better survival after implantation in the first 2 years of life.

Complications and 2-year valve survival following Ahmed valve implantation during the first 2 years of life.

AIM: To report complications and 2-year valve survival following Ahmed valve implantation during the first 2 years of life. METHODS: Retrospective institutional case series. RESULTS: Forty-two eyes of 36 patients with Ahmed valve implantation (without prior drainage device surgery) during the first 2 years of life and 2 years' postsurgical follow-up were identified. Most eyes had primary congenital glaucoma (28/42, 66.7%), aphakic glaucoma (5/42, 11.9%) or Peters anomaly (5/42, 11.9%). All but three eyes had prior ocular surgery. Surgery was at a mean age of 11.83 months (m) (SD 5.63). The most common significant postoperative complications were tube malpositioning requiring intervention (11/42, 26.2%), endophthalmitis (3/42, 7.1%; one with tube exposure) and retinal detachment (3/42, 7.1%). Thirty-six eyes (85.8%) required resumption of antiglaucoma medications to maintain intraocular pressure (IOP) < or =22 mm Hg a mean of 7.2 m (SD 6.8) postoperatively. Cumulative probabilities of valve survival (IOP< or =22 mm Hg with or without medication) by Kaplan-Meier analysis were 73.8% and 63.3% at 12 months and 24 months, respectively. CONCLUSIONS: Postoperative tube malpositioning that required surgical revision was common in this age group. Infectious endophthalmitis and retinal detachment are known potential complications following any incisional surgery for advanced buphthalmos; however, tube exposure is a unique potential problem following aqueous shunt implantation that can lead to intraocular infection. Cumulative valve survival 2 years following implantation was 63.3%.

Paediatric secondary intraocular lens estimation from the aphakic refraction alone: comparison with a standard biometric technique.

AIM: To compare the following two methods of paediatric secondary posterior chamber intraocular lens (PCIOL) determination with the Holladay formula: (1) estimation from the aphakic refraction alone (using assumed keratometry (K) of 44 diopters); and (2) calculation based on preoperative measured biometry. METHODS: (1) Retrospective medical record review in a referral eye hospital of children with aphakia aged < or =12 years who underwent secondary PCIOL implantation with an Alcon MA60BM lens; (2) PCIOL determination for a plano refraction by the above two methods (estimation and calculation); and (3) prediction of pseudophakic refraction for the PCIOL actually implanted by the above two methods compared with the actual pseudophakic refraction. RESULTS: 50 eyes of 30 children with aphakia were studied. The estimated (mean, 95% confidence interval (CI)) secondary PCIOL values (25.81, +/-1.65 D) and the calculated secondary PCIOL values (26.35, +/-1.50 D) were not significantly different (mean absolute value of the difference 1.86 D, 95% CI +/-0.41 D) by the two-tailed paired t test at alpha = 0.05 (p = 0.11). For each eye, the pseudophakic refractions predicted by the two methods for the PCIOL that was actually implanted differed, both from each other and from the actual pseudophakic refraction (repeated-measures analysis of variance, p<0.001; Tukey test, p<0.01). CONCLUSIONS: The method of PCIOL estimation from the aphakic refraction alone provides values similar to those obtained by a standard technique and can be useful if biometry is unavailable. Targeting a pseudophakic refraction in paediatric aphakia is prone to error.

Retinoscopic (refractive) estimation of axial length in paediatric aphakia: a comparison with ultrasonic measurement.

AIM: To compare paediatric axial length values estimated from the aphakic refraction alone with axial length values measured by ultrasound. METHODS: Retrospective institutional medical record review of paediatric aphakic patients 12 years of age and younger with documented ultrasonic axial length and objective refraction (retinoscopy) within 3 months of each other. An estimate of axial length was made from the aphakic refraction alone (with an assumed average keratometry value of 44 dioptres) for all patients. RESULTS: 149 eyes of 102 paediatric aphakic patients were identified. The ultrasonic axial length values (mean 22.47 mm, SD 1.69, 95% confidence interval (CI) 0.27) and estimated axial length values (mean 22.41 mm, SD 1.53, 95% CI 0.25) had an average difference of 0.05 mm (SD 1.04, 95% CI 0.17) and were not significantly different (p = 0.56) by the two tailed paired t test. A histogram of the differences that did exist between the two values resembled a normal distribution. The nine eyes with the largest differences between the two values had either low hyperopic aphakic refractions or abnormal average keratometry values. CONCLUSIONS: There was no significant difference between the two groups of axial length values, and the distribution of differences that did exist seemed random. The greatest differences between the two values occurred in longer (less hyperopic) eyes and in eyes with abnormally steep or flat keratometry. Estimation of axial length from the aphakic refraction alone seems to be a useful technique in the average paediatric eye, especially if biometry is unavailable.