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1.  16S rRNA gene-based identification of bacteria in postoperative endophthalmitis by PCR-Denaturing Gradient Gel Electrophoresis (PCR-DGGE) fingerprinting 
Brazilian Journal of Microbiology  2012;43(1):283-287.
Conventional microbiological culture techniques are frequently insufficient to confirm endophthalmitis clinical cases which could require urgent medical attention because it could lead to permanent vision loss. We are proposing PCR-DGGE and 16S rRNA gene libraries as an alternative to improve the detection and identification rate of bacterial species from endophthalmitis cases.
PMCID: PMC3768976  PMID: 24031830
endophthalmitis; Staphylococcus epidermidis; PCR; DGGE
2.  Molecular and clinical analysis in a series of patients with Pyknodysostosis reveals some uncommon phenotypic findings 
Pyknodysostosis is a rare autosomal recessive skeletal dysplasia characterized by short stature, deformity of the skull, osteosclerosis, hypoplasia of the clavicle, and bone fragility. Radiographs show increased bone density, osteosclerosis, and acroosteolysis of the terminal phalanges. The pycnodysostosis gene is located on chromosome 1q21 and encodes an enzyme called Cathepsin K. Cathepsin K is a cysteine protease lysosomal protein associated with the degradation of bone and cartilage. In the current study, the authors described the clinical, radiological and molecular features of a group of six Mexican patients, including two familial and two sporadic cases, with Pyknodysostosis. One of the patients presented hypoacusia, an unusual finding in this disease.
PMCID: PMC4276157  PMID: 25550899
Pyknodysostosis; sclerosing; bone; dysplasia; Cathepsin K
3.  An epidemiological investigation of a Forkhead box protein E3 founder mutation underlying the high frequency of sclerocornea, aphakia, and microphthalmia in a Mexican village 
Molecular Vision  2013;19:1866-1870.
To investigate the molecular epidemiological basis for the unusually high incidence of sclerocornea, aphakia, and microphthalmia in a village in the Tlaxcala province of central Mexico.
A population census was performed in a village to identify all sclerocornea, aphakia, and microphthalmia cases. Molecular analysis of the previously identified Forkhead box protein E3 (FOXE3) mutation, c.292T>C (p.Y98H), was performed with PCR amplification and direct DNA sequencing. In addition, DNA from 405 randomly selected unaffected villagers was analyzed to establish the carrier frequency of the causal mutation. To identify the number of generations since the mutation arose in the village, 17 polymorphic markers distributed in a region of 6 Mb around the mutated locus were genotyped in the affected individuals, followed by DMLE software analysis to calculate mutation age.
A total of 22 patients with sclerocornea, aphakia, and microphthalmia were identified in the village, rendering a disease prevalence of 2.52 cases per 1,000 habitants (1 in 397). The FOXE3 homozygous mutation was identified in all 17 affected subjects who consented to molecular analysis. Haplotype analysis indicated that the mutation arose 5.0–6.5 generations ago (approximately 106–138 years). Among the 405 unaffected villagers who were genotyped, ten heterozygote carriers were identified, yielding a population carrier frequency of approximately 1 in 40 and a predicted incidence of affected of 1 in 6,400 based on random marriages between two carriers in the village.
This study demonstrates that a cluster of patients with sclerocornea, aphakia, and microphthalmia in a small Mexican village is due to a FOXE3 p.Y98H founder mutation that arose in the village just over a century ago at a time when a population migrated from a nearby village because of land disputes. The actual disease incidence is higher than the calculated predicted value and suggests non-random marriages (i.e., consanguinity) within the population. We can now offer the community more informed genetic counseling based on an accurate genetic test, thus increasing the likelihood of a better outcome for the families.
PMCID: PMC3762562  PMID: 24019743
4.  Polymorphism analysis and new JAG1 gene mutations of Alagille syndrome in Mexican population☆ 
Meta Gene  2013;2:32-40.
Alagille syndrome is a multisystem disorder with an autosomic dominant pattern of inheritance that affects the liver, heart, eyes, kidneys, skeletal system and presents characteristic facial features. Mutations of the JAG1 gene have been identified in 20–89% of the patients with Alagille syndrome, this gene encodes for a ligand that activates the Notch signaling pathway. In the present study we analyzed 9 Mexican patients with Alagille syndrome who presented the clinical criteria for the classical presentation of the disease. By using the denaturing high performance liquid chromatography mutation analysis we were able to identify different mutations in 7 of the patients (77.77%), importantly, we found 5 novel mutations in JAG1 gene. The allelic frequency distribution of 13 polymorphisms in Mexican population is also reported. The overall results demonstrated an expanding mutational spectrum of JAG1 gene in the Mexican population.
PMCID: PMC4287796  PMID: 25606387
AA, African American; ALGS, Alagille syndrome; CEU, Utah Residents with Northern and Western European Ancestry; CHB, Han Chinese in Beijing, China; CI, confidence interval; dbSNP, The Single Nucleotide Polymorphism Database; DHPLC, Denaturing high performance liquid chromatography; DSL, Delta-Serrate-Lag2 domain; EA, European American; ESP, Exome Sequencing Project; HGMD, The Human Gene Mutation Database; HWE, Hardy–Weinberg Equilibrium; JAG1, Gene coding for JAGGED1 protein; JPT, Japanese in Tokyo, Japan; kb, kilobase(s) or 1000 bp; LOVD, Leiden Open Variation Database; MAF, minor allele frequency; MEX, Mexican population; MIM, Mendelian Inheritance in Man; mutDB, mutDB Polymorphism Database; NA, not applicable; ND, not determined; NMD, Nonsense Mediated mRNA Decay; NOTCH2, gene coding for NOTCH2 protein; OR, odds ratio; PCR, polymerase chain reaction; YRI, Yoruba in Ibadan, Nigeria; Alagille syndrome; Mexican patients; JAG1 mutations
5.  Identification and expression analysis of a novel intragenic EFNB1 mutation causing craniofrontonasal syndrome☆ 
Meta Gene  2013;2:25-31.
Craniofrontonasal syndrome (CFNS) is an X-linked disorder caused by mutations in the EFNB1 gene and characterized by distinctive craniofacial and digital malformations. In contrast with most X-linked traits, female patients with CFNS display a more severe phenotype than males. In this report, the clinical, molecular and RNA expression analyses of a female subject with CFNS are described. A novel c.445_449delGAGGG deletion in exon 3 of EFNB1 was demonstrated in this patient. To assess the effect of this novel mutation at the transcript level, the expression of EFNB1 mRNA was studied by quantitative RT-PCR. To our knowledge, this is the first time that an EFNB1 transcript carrying a truncating mutation in exon 3 is demonstrated to undergo degradation by nonsense-mediated mRNA decay. Our results expand the mutational spectrum of CFNS and add to the functional consequences of truncating EFNB1 mutations.
•A novel EFNB1 truncating mutation in craniofrontonasal syndrome (CFNS) is reported.•Expression analyses suggest that nonsense-mediated RNA decay acts on this mutation.•Our report expands the EFNB1 mutational and functional spectrum in CFNS.
PMCID: PMC4287793  PMID: 25606386
Craniofrontonasal syndrome; CFNS; EFNB1; Nonsense-mediated decay
6.  Gene Therapy for Retinitis Pigmentosa Caused by MFRP Mutations: Human Phenotype and Preliminary Proof of Concept 
Human Gene Therapy  2011;23(4):367-376.
Autosomal recessive retinitis pigmentosa (RP), a heterogeneous group of degenerations of the retina, can be due to mutations in the MFRP (membrane-type frizzled-related protein) gene. A patient with RP with MFRP mutations, one of which is novel and the first splice site mutation reported, was characterized by noninvasive retinal and visual studies. The phenotype, albeit complex, suggested that this retinal degeneration may be a candidate for gene-based therapy. Proof-of-concept studies were performed in the rd6 Mfrp mutant mouse model. The fast-acting tyrosine-capsid mutant AAV8 (Y733F) vector containing the small chicken β-actin promoter driving the wild-type mouse Mfrp gene was used. Subretinal vector delivery on postnatal day 14 prevented retinal degeneration. Treatment rescued rod and cone photoreceptors, as assessed by electroretinography and retinal histology at 2 months of age. This AAV-mediated gene delivery also resulted in robust MFRP expression predominantly in its normal location within the retinal pigment epithelium apical membrane and its microvilli. The clinical features of MFRP-RP and our preliminary data indicating a response to gene therapy in the rd6 mouse suggest that this form of RP is a potential target for gene-based therapy.
In this proof-of-concept study, Dinculescu and colleagues demonstrate that subretinal delivery of a self-complementary tyrosine-capsid mutant AAV serotype 8 (AAV8) (Y733F) vector carrying the mouse Mfrp gene prevents retinal degeneration and rescues rod and cone photoreceptors in a mouse model of autosomal recessive retinitis pigmentosa.
PMCID: PMC3327606  PMID: 22142163
7.  Mutations in a novel serine protease PRSS56 in families with nanophthalmos 
Molecular Vision  2011;17:1850-1861.
Nanophthalmos is a rare genetic ocular disorder in which the eyes of affected individuals are abnormally small. Patients suffer from severe hyperopia as a result of their markedly reduced axial lengths, but otherwise are capable of seeing well unlike other more general forms of microphthalmia. To date one gene for nanophthalmos has been identified, encoding the membrane-type frizzled related protein MFRP. Identification of additional genes for nanophthalmos will improve our understanding of normal developmental regulation of eye growth.
We ascertained a cohort of families from eastern Canada and Mexico with familial nanophthalmos. We performed high density microsatellite and high density single nucleotide polymorphism (SNP) genotyping to identify potential chromosomal regions of linkage. We sequenced coding regions of genes in the linked interval by traditional PCR-based Sanger capillary electrophoresis methods. We cloned and sequenced a novel cDNA from a putative causal gene to verify gene structure.
We identified a linked locus on chromosome 2q37 with a peak logarithm (base 10) of odds (LOD) score of 4.7. Sequencing of coding exons of all genes in the region identified multiple segregating variants in one gene, recently annotated as serine protease gene (PRSS56), coding for a predicted trypsin serine protease-like protein. One of our families was homozygous for a predicted pathogenic missense mutation, one family was compound heterozygous for two predicted pathogenic missense mutations, and one family was compound heterozygous for a predicted pathogenic missense mutation plus a frameshift leading to obligatory truncation of the predicted protein. The PRSS56 gene structure in public databases is based on a virtual transcript assembled from overlapping incomplete cDNA clones; we have now validated the structure of a full-length transcript from embryonic mouse brain RNA.
PRSS56 is a good candidate for the causal gene for nanophthalmos in our families.
PMCID: PMC3137557  PMID: 21850159
8.  Clinical features associated with an I126M α2-chimaerin mutation in a family with autosomal dominant Duane retraction syndrome 
We describe the clinical phenotype of a Mexican family segregating Duane syndrome as an autosomal dominant trait linked to chromosome 2q31 (DURS2) and previously reported to harbor a heterozygous α2-chimaerinmissense mutation.
A five-generation Mexican family was analyzed. Ten affected subjects were available for clinical examination. Participating subjects were tested for visual acuity, ocular alignment by prism cover testing, ocular ductions and versions, and globe retraction. In children, alignment was measured with the Krimsky test in cardinal positions of gaze.
Ten cases were included, 6 females and 4 males. Five cases presented with bilateral and 5 with unilateral Duane syndrome. Right side was the most commonly affected side on unilateral cases. Five cases exhibited exotropia, 4 esotropia, and 1 hypotropia. Seven patients had important limitation of abduction; two, moderate limitation. Four patients had mild adduction limitation and 4 had moderate limitation. No additional anomalies such as fourth (trochlear) nerve palsy, blepharoptosis, or dense amblyopia, reported in previous Duane syndrome families, were observed. All 3 cases that exhibited vertical dysfunction had upgaze limitation. One instance of nonpenetrance was recorded.
Considerable intrafamilial clinical variability was observed in this Duane syndrome pedigree carrying a α2-chimaerin mutation. The presence of bilateral involvement and associated vertical movements, commonly observed in this and others DURS2 families, could suggest the occurrence of CHN1 mutations as the source of the disease in isolated or familial DURS cases.
PMCID: PMC2702258  PMID: 19541263
9.  Human CHN1 mutations hyperactivate α2-chimaerin and cause Duane’s retraction syndrome 
Science (New York, N.Y.)  2008;321(5890):839-843.
The RacGAP molecule α2-chimaerin is implicated in neuronal signaling pathways required for precise guidance of developing corticospinal axons. We now demonstrate that a variant of Duane’s retraction syndrome, a congenital eye movement disorder in which affected individuals show aberrant development of axon projections to the extraocular muscles, can result from gain-of-function heterozygous missense mutations in CHN1 that increase α2-chimaerin RacGAP activity in vitro. A subset of mutations enhances α2-chimaerin membrane translocation and/or α2-chimaerin’s previously unrecognized ability to form a complex with itself. In ovo expression of mutant CHN1 alters the development of ocular motor axons. These data demonstrate that human CHN1 mutations can hyperactivate α2-chimaerin and result in aberrant cranial motor neuron development.
PMCID: PMC2593867  PMID: 18653847

Results 1-9 (9)