Mutations in phosphodiesterase 6 identified in familial cases of retinitis pigmentosa.

To delineate the genetic determinants associated with retinitis pigmentosa (RP), a hereditary retinal disorder, we recruited four large families manifesting cardinal symptoms of RP. We localized these families to regions on the human genome harboring the α and β subunits of phosphodiesterase 6 and identified mutations that were absent in control chromosomes. Our data suggest that mutations in PDE6A and PDE6B are responsible for the retinal phenotype in these families

Pathogenic mutations in TULP1 responsible for retinitis pigmentosa identified in consanguineous familial cases.

PurposeTo identify pathogenic mutations responsible for autosomal recessive retinitis pigmentosa (arRP) in consanguineous familial cases.MethodsSeven large familial cases with multiple individuals diagnosed with retinitis pigmentosa were included in the study. Affected individuals in these families underwent ophthalmic examinations to document the symptoms and confirm the initial diagnosis. Blood samples were collected from all participating members, and genomic DNA was extracted. An exclusion analysis with microsatellite markers spanning the TULP1 locus on chromosome 6p was performed, and two-point logarithm of odds (LOD) scores were calculated. All coding exons along with the exon-intron boundaries of TULP1 were sequenced bidirectionally. We constructed a single nucleotide polymorphism (SNP) haplotype for the four familial cases harboring the K489R allele and estimated the likelihood of a founder effect.ResultsThe ophthalmic examinations of the affected individuals in these familial cases were suggestive of RP. Exclusion analyses confirmed linkage to chromosome 6p harboring TULP1 with positive two-point LOD scores. Subsequent Sanger sequencing identified the single base pair substitution in exon14, c.1466A>G (p.K489R), in four families. Additionally, we identified a two-base deletion in exon 4, c.286_287delGA (p.E96Gfs77*); a homozygous splice site variant in intron 14, c.1495+4A>C; and a novel missense variation in exon 15, c.1561C>T (p.P521S). All mutations segregated with the disease phenotype in the respective families and were absent in ethnically matched control chromosomes. Haplotype analysis suggested (p<10(-6)) that affected individuals inherited the causal mutation from a common ancestor.ConclusionsPathogenic mutations in TULP1 are responsible for the RP phenotype in seven familial cases with a common ancestral mutation responsible for the disease phenotype in four of the seven families

Loss of function mutations in RP1 are responsible for retinitis pigmentosa in consanguineous familial cases.

PurposeThis study was undertaken to identify causal mutations responsible for autosomal recessive retinitis pigmentosa (arRP) in consanguineous families.MethodsLarge consanguineous families were ascertained from the Punjab province of Pakistan. An ophthalmic examination consisting of a fundus evaluation and electroretinography (ERG) was completed, and small aliquots of blood were collected from all participating individuals. Genomic DNA was extracted from white blood cells, and a genome-wide linkage or a locus-specific exclusion analysis was completed with polymorphic short tandem repeats (STRs). Two-point logarithm of odds (LOD) scores were calculated, and all coding exons and exon-intron boundaries of RP1 were sequenced to identify the causal mutation.ResultsThe ophthalmic examination showed that affected individuals in all families manifest cardinal symptoms of RP. Genome-wide scans localized the disease phenotype to chromosome 8q, a region harboring RP1, a gene previously implicated in the pathogenesis of RP. Sanger sequencing identified a homozygous single base deletion in exon 4: c.3697delT (p.S1233Pfs22*), a single base substitution in intron 3: c.787+1G>A (p.I263Nfs8*), a 2 bp duplication in exon 2: c.551_552dupTA (p.Q185Yfs4*) and an 11,117 bp deletion that removes all three coding exons of RP1. These variations segregated with the disease phenotype within the respective families and were not present in ethnically matched control samples.ConclusionsThese results strongly suggest that these mutations in RP1 are responsible for the retinal phenotype in affected individuals of all four consanguineous families

Identification of novel transcripts and peptides in developing murine lens.

We previously investigated the transcriptome and proteome profiles of the murine ocular lens at six developmental time points including two embryonic (E15 and E18) and four postnatal time points (P0, P3, P6, and P9). Here, we extend our analyses to identify novel transcripts and peptides in developing  mouse lens. We identified a total of 9,707 novel transcripts and 325 novel fusion genes in developing mouse lens. Additionally, we identified 13,281 novel alternative splicing (AS) events in mouse lens including 6,990 exon skipping (ES), 2,447 alternative 3' splice site (A3SS), 1,900 alternative 5' splice site (A5SS), 1,771 mutually exclusive exons (MXE), and 173 intron retention (IR). Finally, we integrated our OMIC (Transcriptome and Proteome) datasets identifying 20 novel peptides in mouse lens. All 20 peptides were validated through matching MS/MS spectra of synthetic peptides. To the best of our knowledge, this is the first report integrating OMIC datasets to identify novel peptides in developing murine lens

Frequency distribution of hepatitis C virus genotypes in different geographical regions of Pakistan and their possible routes of transmission

<p>Abstract</p> <p>Background</p> <p>Information regarding hepatitis C virus genotypes and subtypes circulating in Pakistan and various risk factors for their transmission are not known well. The specific objective of this study was to find out the frequency of various HCV genotypes present in well-characterized Pakistani HCV isolates and their possible routes of transmission.</p> <p>Methods</p> <p>A total of 3351 serum samples were tested by type-specific genotyping assay. Out of 3351 HCV RNA positive patients, 2039 were males and 1312 were females. As regard as genotyped samples, 2165 belonged to Punjab region, 823 belonged to N.W.F.P., 239 to Sindh and 124 patients were from Balochistan.</p> <p>Results</p> <p>Out of the total 3351 tested serum samples, type-specific PCR fragments were observed in 3150 (94.00%) serum samples. The distribution of genotypes of the typeable samples as determined by this assay, was as follows: 1664 (49.05%) genotype 3a; 592 (17.66%) genotype 3b; 280 (8.35%) genotype 1a; 252 (7.52%) genotype 2a; 101 (3.01%) genotype 1b; 50 (1.49%) with genotype 4; 25 (0.75%) with 3c; 27 (0.80%) genotype 2b; 6 (0.18%) with subtype 5a; 5 (0.15%) genotype 1c; 4 (0.12%) with subtype 6a; 3 (0.09%) genotype 2c; and 161 (4.80%) patients were infected with mixed infection. Two hundred and one (5.99%) serum samples were found untypeable by the present genotyping system. More than 86% and 72% patients with genotypes 3a and 3b respectively had received multiple injections in past. For genotypes 1a and 1b the route of transmission was major/minor surgery along with unknown reasons. Majority of the cases with type 2a, 2b and indeterminate genotypes were sporadic. Mixed infections were common in thalassaemic patients.</p> <p>Conclusion</p> <p>The most common HCV genotype in Pakistan is type 3a. Regional difference in genotypes was observed only in Balochistan province of Pakistan. More than 70% of the cases were acquired in hospitals through reuse of needles/syringes and major/minor surgery that is very common in this country.</p

A study of best positive predictors for sustained virologic response to interferon alpha plus ribavirin therapy in naive chronic hepatitis C patients

<p>Abstract</p> <p>Background</p> <p>The aim of this study was to determine the rate of sustained virological response (SVR) and various factors associated with response rates in chronic hepatitis C infected patients treated with interferon alpha and ribavirin combination therapy.</p> <p>Methods</p> <p>A retrospective review of patients data collected at this Centre from 2001 to 2007 was performed. Out of 731 consecutive patients 400 patients that fulfilled the study criteria were evaluated and subsequently treated with a combination of interferon alpha 2b (3 MU subcutaneously three injections weekly) and ribavirin (800–1200 mg orally daily). Treatment were administered for either 24 weeks or 48 weeks and patients were followed for an additional 6 months thereafter. End of the treatment response (ETR), SVR and side effects were recorded.</p> <p>Results</p> <p>Out of 400 patients, 394 completed the entire treatment course and six patients discontinued treatment at month 2. Over 67% responded at the end of treatment and 16% suffered relapse. Among all treated patients, 47.6% males and 56.7% females had sustained viral response with a total combined sustained viral response rate of 50.5%. Rapid response was seen in 46.5% patients. In a multivariate logistic regression analysis, slow virological responders (adjusted OR 2.6 [95% CI 1.9–3.7]), HCV genotype 1&4 (adjusted OR 2.4 [95% CI 1.7–3.5]), pre-treatment viral load > 0.2 MIU/mL (adjusted OR 2.2 [95% CI 1.8–4.2]), Panjabi ethnic group (adjusted OR 1.6 [95% CI 1.0–3.2]) and Age > 40 years (adjusted OR 1.5 [95% CI 0.9–2.4]) were independent risk factors for non response. Side effects were usual and tolerable and only 1.5% discontinued the treatment.</p> <p>Conclusion</p> <p>The best positive predictor for SVR in this country are: rapid virologic response, HCV genotype 2 & 3, age < 40 years, ethnic race Pashtoons and pre-treatment viral load < 0.2 million IU/mL.</p

Homozygosity Mapping and Genetic Analysis of Autosomal Recessive Retinal Dystrophies in 144 Consanguineous Pakistani Families.

PurposeThe Pakistan Punjab population has been a rich source for identifying genes causing or contributing to autosomal recessive retinal degenerations (arRD). This study was carried out to delineate the genetic architecture of arRD in the Pakistani population.MethodsThe genetic origin of arRD in a total of 144 families selected only for having consanguineous marriages and multiple members affected with arRD was examined. Of these, causative mutations had been identified in 62 families while only the locus had been identified for an additional 15. The remaining 67 families were subjected to homozygosity exclusion mapping by screening of closely flanking microsatellite markers at 180 known candidate genes/loci followed by sequencing of the candidate gene for pathogenic changes.ResultsOf these 67 families subjected to homozygosity mapping, 38 showed homozygosity for at least one of the 180 regions, and sequencing of the corresponding genes showed homozygous cosegregating mutations in 27 families. Overall, mutations were detected in approximately 61.8 % (89/144) of arRD families tested, with another 10.4% (15/144) being mapped to a locus but without a gene identified.ConclusionsThese results suggest the involvement of unmapped novel genes in the remaining 27.8% (40/144) of families. In addition, this study demonstrates that homozygosity mapping remains a powerful tool for identifying the genetic defect underlying genetically heterogeneous arRD disorders in consanguineous marriages for both research and clinical applications

Whole genome sequencing data of multiple individuals of Pakistani descent

Here we report whole genome sequencing of four individuals (H3, H4, H5, and H6) from a family of Pakistani descent. Whole genome sequencing yielded 1084.92, 894.73, 1068.62, and 1005.77 million mapped reads corresponding to 162.73, 134.21, 160.29, and 150.86 Gb sequence data and 52.49x, 43.29x, 51.70x, and 48.66x average coverage for H3, H4, H5, and H6, respectively. We identified 3,529,659, 3,478,495, 3,407,895, and 3,426,862 variants in the genomes of H3, H4, H5, and H6, respectively, including 1,668,024 variants common in the four genomes. Further, we identified 42,422, 39,824, 28,599, and 35,206 novel variants in the genomes of H3, H4, H5, and H6, respectively. A major fraction of the variants identified in the four genomes reside within the intergenic regions of the genome. Single nucleotide polymorphism (SNP) genotype based comparative analysis with ethnic populations of 1000 Genomes database linked the ancestry of all four genomes with the South Asian populations, which was further supported by mitochondria based haplogroup analysis. In conclusion, we report whole genome sequencing of four individuals of Pakistani descent

Monitoring of toxic metals (cadmium, lead, arsenic and mercury) in vegetables of Sindh, Pakistan

A monitoring study was carried out with the aim to assess the level of toxic metals i.e., lead (Pb), cadmium (Cd), arsenic (As) and mercury (Hg) in different vegetables grown in Sindh province of Pakistan during 2007-2008. Two hundred ten samples of twenty one vegetables were collected from farmers' field of Sindh and exporters at Karachi. These samples were grouped into four categories viz., leafy, root and tuberous, cucurbits and fruity. The samples in duplicate were digested with nitric and perchloric acid mixture with 3:1 ratio. Cadmium and Pb were analyzed with Graphite Furnace Atomic Absorption Spectrophotometer and As and Hg on Atomic Absorption using Vapor and Hydride Generation Assembly. Average concentration of Cd, Pb, As and Hg in leafy vegetables was found 0.083 μgg-1, 0.05 μgg-1, 0.042 μgg-1 and 0.008 μgg-1 respectively, in roots and tuberous vegetables was 0.057 μgg-1, 0.03 μgg-1, 0.045 μgg-1 &amp; 0.004 μgg-1 respectively, in cucurbit vegetables was 0.021 μgg-1, 0.051 μgg-1, 0.056 μgg-1 and 0.0089 μgg-1 respectively and in fruity vegetables was 0.035 μgg-1, 0.067 μgg-1, 0.054 μgg-1 and 0.007 μgg-1 respectively. In leafy vegetables, the concentration of cadmium, lead and mercury were found comparatively higher than other three groups of vegetables. However, concentration of heavy metals found in the samples of all four categories of vegetables, was within the permissible limits and safe to consume