Case Report: Precision genetic diagnosis in a case of Dyggve-Melchior-Clausen syndrome reveals paternal isodisomy and heterodisomy of chromosome 18 with imprinting clinical implications.

A twelve-year-old patient with a previous clinical diagnosis of spondylocostal skeletal dysplasia and moderate intellectual disability was genetically analyzed through next generation sequencing of a targeted gene panel of 179 genes associated to skeletal dysplasia and mucopolysaccharidosis in order to stablish a precision diagnosis. A homozygous nonsense [c.62C>G; p.(Ser21Ter)] mutation in DYM gene was identified in the patient. Null mutations in DYM have been associated to Dyggve-Melchior-Clausen syndrome, which is a rare autosomal-recessive disorder characterized by skeletal dysplasia and mental retardation, compatible with the patient´s phenotype. To confirm the pathogenicity of this mutation, a segregation analysis was carried out, revealing that the mutation p(Ser21Ter) was solely inherited from the father, who is a carrier of the mutation, while the mother does not carry the mutation. With the suspicion that a paternal disomy could be causing the disease, a series of microsatellite markers in chromosome 18, where the DYM gene is harbored, was analyzed in all the members of the family. Haplotype analysis provided strong evidence of paternal isodisomy and heterodisomy in that chromosome, confirming the pathological effect of this mutation. Furthermore, the patient may have a compromised expression of the ELOA3 gene due to modifications in the genomic imprinting that may potentially increase the risk of digestive cancer. All these results highlight the importance of obtaining a precision diagnosis in rare diseases.

Brief Report: Evidence of Autism Spectrum Disorder Caused by a Mutation in ATRX Gene: A Case Report.

ATRX mutations are commonly associated with alpha-thalassaemia mental retardation syndrome (ATR-X syndrome) with a notable variable expressivity. This X-linked disorder is characterized by intellectual disability (ID) in a higher or lesser degree, in which the alpha-thalassaemia feature is not always present. Other phenotypic manifestations like facial dimorphism, hypotonia, microcephaly, skeletal abnormalities or urogenital malformations have been frequently observed in ATR-X syndrome. Herein, we report a missense ATRX mutation (Thr1621Met) in a patient with an autism spectrum disorder (ASD) diagnosis. Except for ID, no typical signs of ATR-X syndrome were found in the patient. These results confirm the extensive phenotypic variability associated to ATRX mutations and show the involvement of this gene in the ASD.

Acute Anisakiasis: Pharmacological Evaluation of Various Drugs in an Animal Model.

BACKGROUND: The accidental ingestion of the third larval stage of Anisakis can cause acute clinical symptoms, which are relieved via extraction of the larvae. Although this is a highly effective technique, it can only be practiced when the larvae are found in accessible areas of the gastrointestinal tract, and therefore instead the condition has often been treated using various different drugs. AIMS: This study evaluates the effectiveness of gastric acid secretion inhibitors (omeprazole and ranitidine), gastric mucosal protectants (sucralfate) and anthelmintics (mebendazole and flubendazole) in treating anisakiasis in Wistar rats. METHODS: Rats were infected with Anisakis-type I larvae and administered the drugs via a gastric probe. Data were recorded regarding the number of live and dead larvae, their location both within the animal and in its feces, and the presence of gastrointestinal lesions. Additionally, gastric pH was measured and histology performed. RESULTS: While rats in all experimental groups exhibited lesions; those treated with ranitidine and mebendazole showed significantly fewer lesions (50% and 35% of rats exhibited lesions, respectively). Histological examination of the gastric lesions revealed infection-induced changes, but no significant differences were observed between the treated and untreated rats. CONCLUSIONS: Mebendazole was found to be most efficacious in preventing gastrointestinal lesions, followed by ranitidine, which was the most effective antacid of those studied. Both these drugs could thus be considered as part of the conservative management of anisakiasis.

EDA, EDAR, EDARADD and WNT10A allelic variants in patients with ectodermal derivative impairment in the Spanish population.

BACKGROUND: Ectodermal dysplasias (ED) are a group of genetic conditions affecting the development and/or homeostasis of two or more ectodermal derivatives. An attenuated phenotype is considered a non-syndromic trait when the patient is affected by only one impaired ectodermal structure, such as in non-syndromic tooth agenesis (NSTA) disorder. Hypohidrotic ectodermal dysplasia (HED) is the most highly represented ED. X-linked hypohidrotic ectodermal dysplasia (XLHED) is the most common subtype, with an incidence of 1/50,000-100,000 males, and is associated with the EDA gene (Xq12-q13.1); the dominant and recessive subtypes involve the EDAR (2q13) and EDARADD (1q42.3) genes, respectively. The WNT10A gene (2q35) is associated more frequently with NSTA. Our goal was to determine the mutational spectrum in a cohort of 72 Spanish patients affected by one or more ectodermal derivative impairments referred to as HED (63/72) or NSTA (9 /72) to establish the prevalence of the allelic variants of the four most frequently associated genes. Sanger sequencing of the EDA, EDAR, EDARADD and WNT10A genes and multiplex ligation-dependent probe amplification (MLPA) were performed. RESULTS: A total of 61 children and 11 adults, comprising 50 males and 22 females, were included. The average ages were 5.4 and 40.2 years for children and adults, respectively. A molecular basis was identified in 51/72 patients, including 47/63 HED patients, for whom EDA was the most frequently involved gene, and 4/9 NSTA patients, most of whom had variants of WNT10A. Among all the patients, 37/51 had variants of EDA, 8/51 had variants of the WNT10A gene, 4/51 had variants of EDAR and 5/51 had variants of EDARADD. In 42/51 of cases, the variants were inherited according to an X-linked pattern (27/42), with the remaining showing an autosomal dominant (10/42) or autosomal recessive (5/42) pattern. Among the NSTA patients, 3/9 carried pathogenic variants of WNT10A and 1/9 carried EDA variants. A total of 60 variants were detected in 51 patients, 46 of which were different, and out of these 46 variants, 12 were novel. CONCLUSIONS: This is the only molecular study conducted to date in the Spanish population affected by ED. The EDA, EDAR, EDARADD and WNT10A genes constitute the molecular basis in 70.8% of patients with a 74.6% yield in HED and 44.4% in NSTA. Twelve novel variants were identified. The WNT10A gene has been confirmed as the second molecular candidate that has been identified and accounts for one-half of non-EDA patients and one-third of NSTA patients. Further studies using next generation sequencing (NGS) will help to identify other contributory genes in the remaining uncharacterized Spanish patients.

Tonic Seizure Status Epilepticus Triggered by Valproate in a Child with Doose Syndrome.

Antiepileptic drugs may occasionally increase seizure frequency or eliciting de novo seizure occurrence; the underlying mechanism of these effects is not known. The potential adverse effects of valproic acid in myoclonic astatic epilepsy have been noted by experienced clinicians in various different regions of the world, but this important observation has not been sufficiently reported. We present the case of tonic status epilepticus in an 8-year-old boy with Doose syndrome related to valproic acid. Valproic acid, such as others antiepileptic drugs, is liable to produce paradoxical effects such as the atypical seizures we report. We emphasize the importance for the management of acute seizures in an intensive care unit setting and increase awareness of the acute toxic effects of antiepileptic drugs.

Experimental demonstration of pathogenic potential of Anisakis physeteris and Anisakis paggiae in Wistar rats.

Anisakis morphotype I is the principal etiologic agent of human anisakiasis, with differences in pathogenicity found between the Anisakis simplex s.s. and A. pegreffii species; however, the role of morphotype II larvae in this illness is not well understood. The purpose of this study is to verify the ability of morphotype II larvae to invade tissues via the experimental infection of Wistar rats, an animal model which simulates infection in humans. In the in vivo assay, 7.1% (4/56 L3 morphotype II) showed pathogenic potential, defined as the capacity of the larvae to cause lesions, attach to the gastrointestinal wall or penetrate it. Two of these larvae, one of A. physeteris and one of A. paggiae, penetrated the stomach wall and were found within the abdominal cavity, with the first one producing a small lesion with blood vessel breakage. The majority of the L3 larvae of morphotype II were found in the intestine (51.8%; 29/56) with the caecum being the least frequent location (8.9%; 5/56). In contrast, 44.0% (11/25) of the morphotype I larvae demonstrated pathogenic potential. Isoenzyme electrophoresis, PCR-RFLP of ITS1-5.8 s-ITS2 and PCR-sequencing of the cox2 mitochondrial gene were used to identify these larvae as A. physeteris (42.9%), A. paggiae (30.3%) and A. brevispiculata (1.8%). Although the morphotype II larvae of A. physeteris and A. paggiae have lower pathogenic potential than morphotype I larvae of A. simplex s.s. (93 and 91% lower, respectively), they may still be implicated in human anisakiasis, as they are capable of attaching to and penetrating the gastrointestinal wall of animals, demonstrating a similar pathogenicity to that of A. pegreffii. The techniques used for the identification of species reveal a great genetic heterogeneity of A. paggiae and A. physeteris, suggesting the existence of sibling species.

A 2.84 Mb deletion at 21q22.11 in a patient clinically diagnosed with Marden-Walker syndrome.

We present a girl with the characteristic clinical picture associated with Marden-Walker syndrome (MWS; OMIM 248700), including mask-like face with blepharophimosis, joint contractures, intellectual disability, a multicystic dysplastic kidney and cerebral dysgenesis. The long-term follow-up allowed us to monitor the evolution of the phenotype in this patient, and among the main findings we highlight the following: demyelination of the pyramidal tract demonstrated by transcranial magnetic stimulation and the involvement of the levator muscles of angle of mouth in fixed facial expression with relative integrity of the rest of the facial expression muscles. A 244 k array comparative genomic hybridization (aCGH) was carried out and showed a de novo interstitial deletion of approximately 2.84 Mb affecting only the cytoband 21q22.11 (genome coordinates chr21:31,874,016-34,711,763). We selected 10 of the most recent published cases with either total or partial deletions of cytoband 21q22.11 that provided good characterization of the genomic size or the genes in the deleted regions. We observed that in nine of the 10 cases the deleted regions included the RUNX1 gene in 21q22.12, which is not affected in the current patient's deletion or in that of Patient 3 from Roberson et al. [2011]. After a comparison of shared deleted genes between cases, and correlation of their potential phenotypes, we concluded that the pattern of defects considered for a diagnosis of MWS may represent part of the phenotypic expression of a partial or total deletion of 21q22.11.

Periodic Lateralized Epileptiform Discharges (PLEDs) and pneumococcal meningoencephalitis.

Pneumococcal meningoencephalitis (PME) is a life-threatening condition of the central nervous system (CNS), and is often the result of a complicated upper airway infection. Periodic Lateralized Epileptiform Discharges (PLEDs) are a typical electroencephalographic (EEG) pattern found in some acutely acquired brain insults. Within the pediatric population they are frequently seen in association with herpetic encephalitis, a CNS infection with a high morbidity and mortality rate. We report the case of a 3-year-old girl with a bilateral ear infection who developed convulsions and coma. She had early PLEDs lateralized to the right on the EEG and microbiological criteria for Streptococcus pneumoniae infection. Concomitant herpetic encephalitis was ruled out. Intensive antibiotic and antiepileptic treatment resulted in a remarkable improvement, with the patient being able to resume her normal activities within months. To our knowledge, the association of PME and PLEDs has not been previously described in children. On the other hand, EEG has scarcely been used in the management of acute CNS infections. Hence, non-herpetic CNS encephalitis with potentially more favorable outcomes ought to be considered in the differential diagnosis of PLEDs. Continuous EEG monitoring should be considered in children with CNS infections presenting with altered sensorium, independent of the presence of seizures.

Prenatal BACs-on-Beads™: the prospective experience of five prenatal diagnosis laboratories.

OBJECTIVE: We previously reported on the validation of Prenatal BACs-on-Beads™ on retrospectively selected and prospective prenatal samples. This bead-based multiplex assay detects chromosome 13, 18, 21 and X/Y aneuploidies and the nine most frequent microdeletion syndromes. We demonstrated that Prenatal BACs-on-Beads(TM) is a new-generation, prenatal screening tool. Here, we describe the experience of five European prenatal diagnosis laboratories concerning the ongoing use of Prenatal BACs-on-Beads™ . METHODS: Some 1653 samples were analyzed. All results were confirmed by conventional karyotyping or another appropriate technique. All indications for invasive prenatal diagnosis were included. Amniotic fluid and chorionic villus samples were analyzed in equivalent proportions. RESULTS: The failure rate was 3.3% and the overall abnormality detection rate was ~1/10. Eighty-five percent of the detected abnormalities were common aneuploidies. Eleven microdeletions and duplications were identified, thus giving an overall yield for microdeletion and microduplication detection of 1/145. Compared with QF-PCR, Prenatal BACs-on-Beads™ provides an additional detection rate of ~1/250 for low-risk pregnancies. The false positive and negative rates were both <1%. CONCLUSION: When associated with conventional karyotyping, the Prenatal BACs-on-Beads™ assay combines a short turnaround time (typical of rapid aneuploidy detection tests) with valuable detection of the most frequent microdeletion syndromes that cannot be detected in cytogenetic analyses.

Atypical glycine encephalopathy in an extremely low birth weight infant: description of a new mutation and clinical and electroencephalographic analysis.

We present the clinical course and EEG evolution of an extreme low birth weight preterm neonate with an uncommon type of glycine encephalopathy. The patient presented with myoclonic jerks, apnea and encephalopathy three months after birth without satisfactory therapeutic response. During the first days of clinical symptoms the patient presented a paroxystic burst-attenuation EEG pattern which progressively evolved into an established typical burst-suppression pattern within a few days. West syndrome occurred four weeks later and the patient died at seven months of extra-uterine life due to a serious respiratory infection with cardio-respiratory arrest. Genetic analysis showed a non-previously described mutation affecting a consensus splice site (IVS2-1G > C 3) in the AMT gene encoding the T protein of the glycine cleavage system.

A soybean seed protein with carboxylate-binding activity.

The seed coat serves as a multifunctional organ with a role in protection and for the supply of nutrients to the embryo sac during development. The composition of the legume seed coat differs from other seed tissues in many ways including its protein composition. An abundant 24 kDa protein (SC24) has been purified and identified from soybean (Glycine max [L.] Merr) seed hulls. The corresponding cDNA and genomic DNA clones for SC24 were isolated and characterized, and expression patterns were determined. The deduced protein sequence of 219 amino acids included an N-terminal signal peptide. Transcripts encoding SC24 were present in the seed coat from 30 days after pollination (DAP) until maturity, but the protein was not detected until the final stages of seed maturation. In mature seeds, most of the SC24 protein was localized to the parenchyma and aleurone layers of the seed coat. The expression of SC24 was also induced in vegetative tissues by pathogen infection and by wounding. The SC24 protein bound to an affinity column containing an isophthalic acid ligand, and was eluted with 7 mM citrate. Polyclonal antibodies raised against recombinant SC24 cross-reacted with the seed coat peroxidase enzyme, suggesting that these two proteins may share an antigenic determinant. Overall, the results indicate that SC24 belongs to a novel class of plant defence proteins with carboxylate-binding activity.