L'hémisphérotomie péri-insulaire: technique chirurgicale, monitoring EEG intraopératoire et résultats sur le contrôle de l'épilepsie [Periinsular hemispherotomy: surgical technique, intraoperative EEG monitoring and results on seizure outcome]

Peri-insular hemispherotomy is a surgical technique used in the treatment of drug-resistant epilepsy of hemispheric origin. It is based on the exposure of insula and semi-circular sulci, providing access to the lateral ventricle through a supra- and infra-insular window. From inside the ventricle, a parasagittal callosotomy is performed. The basal and medial portion of the frontal lobe is isolated. Projections to the anterior commissure are interrupted at the time of amygdala resection. The hippocampal tail and fimbria-fornix are disrupted posteriorly. We report our experience of 18 cases treated with this approach. More than half of them presented with congenital epilepsy. Neuronavigation was useful in precisely determining the center and extent of the craniotomy, as well as the direction of tractotomies and callosotomy, allowing minimal exposure and blood loss. Intra-operative monitoring by scalp EEG on the contralateral hemisphere was used to follow the progression of the number of interictal spikes during the disconnection procedure. Approximately 90% of patients were in Engel's Class I. We observed one case who presented with transient postoperative neurological deterioration probably due to CSF overdrainage and documented one case of incomplete disconnection in a patient presenting with hemimegalencephaly who needed a second operation. We observed a good correlation between a significant decrease in the number of spikes at the end of the procedure and seizure outcome. Peri-insular hemispherotomy provides a functional disconnection of the hemisphere with minimal resection of cerebral tissue. It is an efficient technique with a low complication rate. Intra-operative EEG monitoring might be used as a predictive factor of completeness of the disconnection and consequently, seizure outcome.

Host genetic and epigenetic factors in toxoplasmosis

Analysing human genetic variation provides a powerful tool in understanding risk factors for disease. Toxoplasma gondii acquired by the mother can be transmitted to the fetus. Infants with the most severe clinical signs in brain and eye are those infected early in pregnancy when fetal immunity is least well developed. Genetic analysis could provide unique insight into events in utero that are otherwise difficult to determine. We tested the hypothesis that propensity for T. gondii to cause eye disease is associated with genes previously implicated in congenital or juvenile onset ocular disease. Using mother-child pairs from Europe (EMSCOT) and child/parent trios from North America (NCCCTS), we demonstrated that ocular and brain disease in congenital toxoplasmosis associate with polymorphisms in ABCA4 encoding ATP-binding cassette transporter, subfamily A, member 4 previously associated with juvenile onset retinal dystrophies including Stargardt's disease. Polymorphisms at COL2A1 encoding type II collagen, previously associated with Stickler syndrome, associated only with ocular disease in congenital toxoplasmosis. Experimental studies showed that both ABCA4 and COL2A1 show isoform-specific epigenetic modifications consistent with imprinting, which provided an explanation for the patterns of inheritance observed. These genetic and epigenetic risk factors provide unique insight into molecular pathways in the pathogenesis of disease.

Parental occupational exposure to organic solvents and anencephaly in Mexico

Objective: To assess the relationship between parental occupational exposure to organic solvents, and the risk of anencephaly in Mexico. Methods: A case-control study was conducted based on the registers of the Epidemiological Surveillance System for Neural Tube Defects in Mexico; 151 cases of anencephaly of ≥20 weeks’ gestation were included. A control, born alive and without any apparent congenital malformations at birth, was selected for each case in the same maternity service in which the case was born. Information on occupational exposures, lifestyle habits, reproductive history, use of medicines, supplementation with multivitamins and folic acid, was obtained by a general questionnaire; a food frequency questionnaire was also applied to obtain information of daily intake of folate and other B vitamins. Occupational exposure to organic solvents was based on job title as a proxy for exposure and analysed considering two critical periods around conception. Results: In logistic regression analysis, the odds of having a child with anencephaly was higher if the mother or the father was occupationally exposed to organic solvents during the periconceptional period, or when both parents or at least one of them were occupationally exposed during this period with an adjusted odds ratio of 2.97 (95% CI 1.36 to 6.52). Conclusions: The results support the hypothesis that both maternal and paternal occupational exposure to organic solvents can increase the probability of having a child with anencephaly.

Comprehensive analysis of RET common and rare variant patientsin a series of Spanish Hirschsprung patients confirms a synergistic effect of both kinds of events

Background. RET is the major gene associated to Hirschsprung disease (HSCR) with differential contributions of its rare and common, coding and noncoding mutations to the multifactorial nature of this pathology. In the present study, we have performed a comprehensive study of our HSCR series evaluating the involvement of both RET rare variants (RVs) and common variants (CVs) in the context of the disease. Methods. RET mutational screening was performed by dHPLC and direct sequencing for the identification of RVs. In addition Taqman technology was applied for the genotyping of 3 RET CVs previously associated to HSCR, including a variant lying in an enhancer domain within RET intron 1 (rs2435357). Statistical analyses were performed using the SPSS v.17.0 to analyze the distribution of the variants. Results. Our results confirm the strongest association to HSCR for the "enhancer" variant, and demonstrate a significantly higher impact of it in male versus female patients. Integration of the RET RVs and CVs analysis showed that in 91.66% of cases with both kinds of mutational events, the enhancer allele is in trans with the allele bearing the RET RV. Conclusions. A gender effect exists on both the transmission and distribution of rare coding and common HSCR causing mutations. In addition, these RET CVs and RVs seem to act in a synergistic way leading to HSCR phenotype.

Immunopathology in ocular toxoplasmosis: facts and clues

Although parasite-mediated host cell lysis is deemed to be an important cause of tissue destruction in ocular toxoplasmosis (OT), the severity of the disease is probably correlated with hypersensitivity and inflammation. Notwithstanding, the mechanisms that regulate the inflammatory process in recurrent OT are poorly understood. Recent evidence has identified interleukin (IL) 17 as a marker for disease severity. The ocular and cerebral presence of this cytokine is generally associated with the induction of autoimmune responses in the brain and the eye. Indeed, there are indications that autoimmunity may contribute to clinical variability in the activity of OT. IL-23, which induces the proliferation of IL-17-producing cells and IL-27, which is a counterplayer to IL-17, may regulate T(H)-1-cell-mediated responses in OT. The importance of these cytokines in experimental models of uveitis and encephalitis has been recently reported. CD25(+) regulatory T-cells may control the local inflammatory response and protect the host against collateral inflammatory tissue damage. The responses of these cells to OT may be suitably tailored to cope with either an acquired or a congenital aetiology. Knowledge relating to immunoreactivity in OT has grown impressively during the past few years. Its characteristic and variable features have been identified and the potential relevance of autoimmunity has been assessed. In light of this knowledge, potential future treatment options have been considered.

Veterinary vaccines against Toxoplasma gondii

Toxoplasma gondii has a very wide intermediate host range and is thought to be able to infect all warm blooded animals. The parasite causes a spectrum of different diseases and clinical symptoms within the intermediate hosts and following infection most animals develop adaptive humoral and cell-mediated immune responses. The development of protective immunity to T. gondii following natural infection in many host species has led researchers to look at vaccination as a strategy to control disease, parasite multiplication and establishment in animal hosts. A range of different veterinary vaccines are required to help control T. gondii infection which include vaccines to prevent congenital toxoplasmosis, reduce or eliminate tissue cysts in meat producing animals and to prevent oocyst shedding in cats. In this paper we will discuss some of the history, challenges and progress in the development of veterinary vaccines against T. gondii.

Ocular toxoplasmosis: the influence of patient age

The influence of patient age on various features of ocular toxoplasmosis has been a subject of study for many years. The age at which Toxoplasma gondii infection occurs in different populations is related to socioeconomic factors and studies suggest that ocular toxoplasmosis is a more severe disease at the extremes of age. The prevalence of ocular involvement is markedly different between individuals with congenital and those with post-natally acquired infections. Even among those with post-natally acquired infections, age influences the risk and timing of ocular involvement. The severity of toxoplasmic retinochoroiditis (in terms of lesion size, location and associated inflammation) is also affected by patient age at the time of initial infection or recurrence. The risk of recurrent toxoplasmic retinochoroiditis is influenced by age at the time of initial infection and age at most recent episode of active disease. Understanding of relationships between ocular toxoplasmosis and patient age is incomplete; evidence has often been indirect and in some cases conflicting. The influence of patient age on ocular toxoplasmosis should be studied in a systematic manner to provide a better understanding of disease mechanisms and to provide clinical information that can used to establish better strategies for disease treatment and prevention.

Contributions to the history of ocular toxoplasmosis in Southern Brazil

The evolution of knowledge regarding ocular toxoplasmosis over the last 30 years is described based on studies and observations performed in Southern Brazil. The isolation ofToxoplasma gondii established the definitive diagnosis of the disease. It was proven that in most cases, the disease was acquired after birth, a concept supported by the description of numerous familial cases and observation of the disease many years after primary infection. Epidemiological studies showed important regional variations in the prevalence of the disease due to different factors, including the types of strains involved, of which type I predominates. The large number of patients also enabled detailed study of the different forms of clinical presentation of the disease and its complications. New parameters have been established for the use of steroids and the management of pregnant women with active lesions. Studies on the epidemiology of toxoplasmic infection in pregnant women and newborns showed a high prevalence of infection. The different factors of exposure to infection have also been studied. Gradually, preventive actions have been developed in the sphere of public health, although they have not been sufficiently effective. Trends for future research over the next few years are also outlined.

Factors associated with seropositivity for anti-Toxoplasma gondiiantibodies in pregnant women of Londrina, Paraná, Brazil

The aim of this study was to evaluate associations between seropositivity for IgG and IgM anti-Toxoplasma gondii antibodies and socio-economic and environmental variables in pregnant women of Londrina, state of Paraná, Brazil. We interviewed 492 pregnant women, each of whom answered an epidemiological questionnaire, and collected blood samples for measurement of IgG and IgM anti-T. gondii antibodies by chemiluminescence. A confirmatory diagnosis of acute infection was made by an IgG avidity test. Titres of specific IgG anti-T. gondii were obtained by IFAT. Seropositivity for IgG anti-T. gondii antibodies was observed in 242 women (49.2%) and, of these, six pregnant women (1.2%) showed seropositivity for IgM. Age group, level of education, per capita income, presence of a cat in the house and a habit of eating green vegetables were all factors associated with a greater chance of infection with T. gondii. This study showed that 250 (50.8%) pregnant women were susceptible to T. gondii and considered to be at high risk for toxoplasmosis during pregnancy. Based on the results obtained, is critical to establish a program of health surveillance for toxoplasmosis, in order to contribute to diagnosis and early treatment during the prenatal period. It is also necessary to introduce measures to prevent the Toxoplasma infection in seronegative pregnant women.

Prevalence of acute toxoplasmosis infection among 41,112 pregnant women and the mother-to-child transmission rate in a public hospital in South Brazil

Untreated acute toxoplasmosis among pregnant women can lead to serious sequelae among newborns, including neurological impairment and blindness. In Brazil, the risk of congenital toxoplasmosis (CTox) has not been fully evaluated. Our aim was to evaluate trends in acute toxoplasmosis prevalence from 1998-2005, the incidence of CTox and the rate of mother-to-child transmission (MTCT). A cross-sectional study was undertaken to dentify patients who fit the criteria for acute toxoplasmosis during pregnancy. Exposed newborns were included in a historical cohort, with a median follow-up time of 11 months, to establish definite diagnosis of CTox. Diagnoses for acute infection in pregnancy and CTox were based on European Research Network on Congenital Toxoplasmosis criteria. In 41,112 pregnant women, the prevalence of acute toxoplasmosis was 4.8/1,000 women. The birth prevalence of CTox was 0.6/1,000 newborns [95% confidence interval (CI): 0.4-0.9]. During the follow-up study, 12 additional cases were detected, increasing the CTox rate to 0.9/1,000 newborns (95% CI: 0.6-1.3). Among the 200 newborns exposed to Toxoplasma gondii,there were 37 babies presenting diagnostic criteria of CTox, leading to an MTCT rate of 18.5% (95% CI: 13.4-24.6%). The additional cases identified during follow-up reinforce the need for serological monitoring during the first year of life, even in the absence of evidence of congenital infection at birth.

Deficiencia nutricional de yodo en gestantes pertenecientes al distrito sanitario Sierra de Huelva-Andévalo, sur de España

Background. Iodine is an essential trace element implicated in synthesis of thyroid hormones. Iodine requirements vary throughout life. Yhis iodine requirement is increased during pregnancy and breastfeeding. In a previous study carried out by our group in 2008, we detected an iodine-deficient area in the province of Huelva, specially in dictrict Sierra de Huelva-Andévalo by means of neonatal TSH determinations. Objective. To reinforce the iodine supplementation campaign and its impact on their newborns in order to assess nutrition iodine status in 'pregnant women using questionnaire and ioduria determination. Material and methods. This study has been jointly carried out by Congenital Hypothiroidism Unit of the Clinical Biochemistry Department of the Virgen Macarena University Hosplital (Seville) and the Gynecology and Clinical Analysis Unit of the Río Tinto Hospital (Huelva) during two years. We studied 313 pregnant women. All of them filled out a personal questionnaire to know the iodine nutritional status in their area. Ioduria was determined by high-resolution liquid chromatography. Data from pregnant and results of the studied variables were analyzed with SPSS v.13.0. Conclusions. Pregnant women from the sanitary district Sierra de Huelva-Andévalo present a median for ioduria which corresponds to an insufficient iodine intake according to the WHO classification. The questionnaire suggest that this iodine deficiency is consequence of an insufficient iodine intake and a low adherence to the treatment.

Expression of PROKR1 and PROKR2 in Human Enteric Neural Precursor Cells and Identification of Sequence Variants Suggest a Role in HSCR

Background. The enteric nervous system (ENS) is entirely derived from neural crest and its normal development is regulated by specific molecular pathways. Failure in complete ENS formation results in aganglionic gut conditions such as Hirschsprung's disease (HSCR). Recently, PROKR1 expression has been demonstrated in mouse enteric neural crest derived cells and Prok-1 was shown to work coordinately with GDNF in the development of the ENS. Principal Findings. In the present report, ENS progenitors were isolated and characterized from the ganglionic gut from children diagnosed with and without HSCR, and the expression of prokineticin receptors was examined. Immunocytochemical analysis of neurosphere-forming cells demonstrated that both PROKR1 and PROKR2 were present in human enteric neural crest cells. In addition, we also performed a mutational analysis of PROKR1, PROKR2, PROK1 and PROK2 genes in a cohort of HSCR patients, evaluating them for the first time as susceptibility genes for the disease. Several missense variants were detected, most of them affecting highly conserved amino acid residues of the protein and located in functional domains of both receptors, which suggests a possible deleterious effect in their biological function. Conclusions. Our results suggest that not only PROKR1, but also PROKR2 might mediate a complementary signalling to the RET/GFRα1/GDNF pathway supporting proliferation/survival and differentiation of precursor cells during ENS development. These findings, together with the detection of sequence variants in PROKR1, PROK1 and PROKR2 genes associated to HSCR and, in some cases in combination with RET or GDNF mutations, provide the first evidence to consider them as susceptibility genes for HSCR.

Mutation screening of multiple genes in Spanish patients with autosomal recessive retinitis pigmentosa by targeted resequencing.

Retinitis Pigmentosa (RP) is a heterogeneous group of inherited retinal dystrophies characterised ultimately by the loss of photoreceptor cells. RP is the leading cause of visual loss in individuals younger than 60 years, with a prevalence of about 1 in 4000. The molecular genetic diagnosis of autosomal recessive RP (arRP) is challenging due to the large genetic and clinical heterogeneity. Traditional methods for sequencing arRP genes are often laborious and not easily available and a screening technique that enables the rapid detection of the genetic cause would be very helpful in the clinical practice. The goal of this study was to develop and apply microarray-based resequencing technology capable of detecting both known and novel mutations on a single high-throughput platform. Hence, the coding regions and exon/intron boundaries of 16 arRP genes were resequenced using microarrays in 102 Spanish patients with clinical diagnosis of arRP. All the detected variations were confirmed by direct sequencing and potential pathogenicity was assessed by functional predictions and frequency in controls. For validation purposes 4 positive controls for variants consisting of previously identified changes were hybridized on the array. As a result of the screening, we detected 44 variants, of which 15 are very likely pathogenic detected in 14 arRP families (14%). Finally, the design of this array can easily be transformed in an equivalent diagnostic system based on targeted enrichment followed by next generation sequencing.

Naevus Lentiginosus Linearis: A Distinct Skin Disorder

Congenital naevi of the melanocytic system include numerous types, which differ in their clinical appearance, pattern of distribution, and histopathological features (1). Examples are large congenital melanocytic naevus, macular naevus spilus, papular naevus spilus, café-au-lait macules of neurofibromatosis 1, café-au-lait macules arranged in broad bands as noted in McCune-Albright syndrome, partial unilateral lentiginosis, naevus achromicus (naevus depigmentosus), phylloid hypermelanosis, and phylloid hypomelanosis (1–3). We describe here two patients with a systematized pigmentary naevus that differed from all naevi reported so far.

Novel MLPA procedure using self-designed probes allows comprehensive analysis for CNVs of the genes involved in Hirschsprung disease.

Background: Hirschsprung disease is characterized by the absence of intramural ganglion cells in the enteric plexuses, due to a fail during enteric nervous system formation. Hirschsprung has a complex genetic aetiology and mutations in several genes have been related to the disease. There is a clear predominance of missense/nonsense mutations in these genes whereas copy number variations (CNVs) have been seldom described, probably due to the limitations of conventional techniques usually employed for mutational analysis. In this study, we have looked for CNVs in some of the genes related to Hirschsprung (EDNRB, GFRA1, NRTN and PHOX2B) using the Multiple Ligation-dependent Probe Amplification (MLPA) approach. Methods: CNVs screening was performed in 208 HSCR patients using a self-designed set of MLPA probes, covering the coding region of those genes. Results: A deletion comprising the first 4 exons in GFRA1 gene was detected in 2 sporadic HSCR patients and in silico approaches have shown that the critical translation initiation signal in the mutant gene was abolished. In this study, we have been able to validate the reliability of this technique for CNVs screening in HSCR. Conclusions: The implemented MLPA based technique presented here allows CNV analysis of genes involved in HSCR that have not been not previously evaluated. Our results indicate that CNVs could be implicated in the pathogenesis of HSCR, although they seem to be an uncommon molecular cause of HSCR.