Whole-exome sequencing of T- B+ severe combined immunodeficiency in Egyptian infants, JAK3 predominance and novel variants.

Severe combined immunodeficiency (SCID) is fatal if not treated with immune reconstitution. In Egypt, T- B+ SCID accounts for 38·5% of SCID diagnoses. An accurate genetic diagnosis is essential for choosing appropriate treatment modalities and for offering genetic counseling to the patient's family. The objectives of this study were to describe the clinical, immunological and molecular characteristics of a cohort of twenty Egyptian patients with T- B+ SCID. The initial diagnosis (based on clinical features and flow cytometry) was followed by molecular investigation (whole-exome sequencing). All patients had the classic clinical picture for SCID, including failure to thrive (n = 20), oral candidiasis (n = 17), persistent diarrhea (n = 14), pneumonia (n = 13), napkin dermatitis (n = 10), skin rash (n = 7), otitis media (n = 3) and meningitis (n = 2). The onset of manifestations was at the age of 2·4 ± 1·6 months and diagnosis at the age of 6·7 ± ·5 months, giving a diagnostic delay of 4·3 months. JAK3 gene variants were most frequent (n = 12) with three novel variants identified, followed by IL2Rγ variants (n = 6) with two novel variants. IL7Rα and CD3ε variants were found once, with a novel variant each. T- B+ NK- SCID accounted for approximately 90% of the Egyptian patients with T- B+ SCID. Of these T- B+ NK- SCID cases, 60% were autosomal recessive syndromes caused by JAK3 mutations and 30% were X-linked syndromes. It might be useful to sequence the JAK3 gene (i.e. targeted Sanger sequencing) in all T- B+ SCID patients, especially after X-linked SCID has been ruled out. Hence, no more than 10% of T- B+ SCID patients might require next-generation for a molecular diagnosis.

Evaluations of N-(Isobutoxymethyl) acrylamide gel dosimeter by NMR technique for radiotherapy and uncertainty in dose measurements.

N-(Isobutoxymethyl) acrylamide (NIBMA) monomer in gelatin, named NIBMAGAT gel dosimeter, was prepared and investigated by nuclear magnetic imaging (NMR) for radiotherapy in the dose range of 0-30 Gy. NIBMA monomer polymerizes upon irradiation, increasing spin-spin relaxation rate R2. The addition of glycerol as a co-solvent in the gel matrix improved its radiation sensitivity better than the co-solvents of acetone and methanol. The increase of glycerol content by 1% wt/wt enhanced the sensitivity by ˜ 3.1%. This gel has better radiation sensitivity as compared to the polyacrylamide gel (PAG) dosimeter; the sensitivities of NIBMAGAT gel and normoxic polyacrylamide gel (nPAG) are ≈0.13 and ≈0.1 s-1.Gy-1, respectively. By comparing NIBMAGAT gel dosimeter with PAG, nMAG and nPAG gel dosimeters, NIBMAGAT gel dosimeter is less influenced by scanning temperature than the last three dosimeters. The gel is water equivalent and has an energy-independent response from 80 keV to 20 MeV. The overall uncertainty of dose measurement using NIBMAGAT gel is 5.46% at 2σ. Our findings suggest the applicability of using NIBMAGAT gel dosimeter by NMR technique for dose verification/planning in the practice of clinical radiotherapy.

Phenotypical heterogeneity in RAG-deficient patients from a highly consanguineous population.

Mutations affecting recombination activation genes RAG1 and RAG2 are associated with variable phenotypes, depending on the residual recombinase activity. The aim of this study is to describe a variety of clinical phenotypes in RAG-deficient patients from the highly consanguineous Egyptian population. Thirty-one patients with RAG mutations (from 28 families) were included from 2013 to 2017. On the basis of clinical, immunological and genetic data, patients were subdivided into three groups; classical T- B- severe combined immunodeficiency (SCID), Omenn syndrome (OS) and atypical SCID. Nineteen patients presented with typical T- B- SCID; among these, five patients carried a homozygous RAG2 mutation G35V and five others carried two homozygous RAG2 mutations (T215I and R229Q) that were detected together. Four novel mutations were reported in the T- B- SCID group; three in RAG1 (A565P, N591Pfs*14 and K621E) and one in RAG2 (F29S). Seven patients presented with OS and a novel RAG2 mutation (C419W) was documented in one patient. The atypical SCID group comprised five patients. Two had normal B cell counts; one had a previously undescribed RAG2 mutation (V327D). The other three patients presented with autoimmune cytopaenias and features of combined immunodeficiency and were diagnosed at a relatively late age and with a substantial diagnostic delay; one patient had a novel RAG1 mutation (C335R). PID disorders are frequent among Egyptian children because of the high consanguinity. RAG mutations stand behind several variable phenotypes, including classical SCID, OS, atypical SCID with autoimmunity and T- B+ CID.

Human placental PPAR-γ and SOX-2 expression in serologically proved toxoplasmosis.

To explore PPAR-γ and SOX-2 transcription factors expression in placenta according to maternal anti-Toxoplasma gondii serological profile during pregnancy and pregnancy outcome. The study included 240 placentas, grouped according to IgM and IgG serostatus and then subgrouped according to pregnancy outcome that varied between miscarriages, premature labour, stillbirth and giving birth to CNS anomaly or apparently healthy neonates. Samples were H&E stained and histopathologically scored blindly. PPAR-γ expression was measured by ELISA, while SOX-2-positive nuclei were stained immunohistochemically to be calculated by ImageJ. The mean pathological score was significantly higher in IgM+ve and IgG rising than IgG-ve and persistent low groups. Former groups showed significantly higher PPAR-γ (mean = 258.63, 227.11). However, PPAR-γ was higher in apparently healthy neonate subgroups. SOX-2 was significantly lower in IgM+ve and IgG rising groups (mean = 12.87, 43.13) and associated with obvious fibrosis. SOX-2 lowest count was in CNS anomaly subgroup. PPAR-γ and SOX-2 changes may give clues of how Toxoplasma induces pathogenesis during vertical transmission. Triggering PPAR-γ expression may be a tool to downregulate the inflammatory response and establish a metabolically permissive cellular environment for Toxoplasma persistence. Low SOX-2 is suspected to disturb placental mesenchymal stem cells pluripotency and neuroectoderm development.

Characterization of microsatellite markers in mandarin orange (Citrus reticulata Blanco).

A dinucleotide-enriched genomic library was obtained from mandarin orange (Citrus reticulata Blanco). A subset of 101 positive clones was sequenced and primers were designed. The loci were screened for levels of variation using 26-29 wild mandarin oranges collected in Vietnam. Forty-three loci were polymorphic with the number of alleles ranging from two to 18. The observed heterozygosity (H(O) ) and expected heterozygosity (H(E) ) were from 0.03 to 0.96 and from 0.03 to 0.92, respectively.

Quality and flavour stability of coffee substitute prepared by extrusion of wheat germ and chicory roots.

A mixture of roasted chicory roots and wheat germ (1:1 w/w) was subjected to extrusion processing for preparation of coffee substitute. Comparative studies concerning sensory characteristics and headspace volatiles were carried out between genuine coffee and a freshly prepared coffee substitute. The sensory evaluation revealed similarities between the two samples. The comparative odour profile analysis showed that the sweetish/caramel-like note scored higher in our coffee substitute sample than in real coffee, whereas the other odour quality attributes showed an opposite trend. The high quality of the fresh coffee substitute was correlated to the presence of volatiles that are responsible for the fresh coffee aroma, such as: 2-methylbutanal, 3-methylbutanal, 2-methylfuran and 2,3-butanedione in high concentration. Storage of coffee substitute samples revealed a noticeable decrease in concentration of the Strecker aldehydes and diketones and a remarkable increase in phenolic compounds, whereas pyrazine and furan derivatives showed no linear changes during storage. The ratio of 2,3-butanedione/2-methylfuran (B/M) was used as an indicator for aging of coffee substitute samples. The variation in this ratio (B/M) during storage for 6 months was consistent with that of the odour profile analysis.

Serum levels of bcl-2 and cellular oxidative stress in patients with viral hepatitis.

PURPOSE: This study was conducted to investigate the presence of bcl-2 protein in the serum of patients with viral hepatitis and to find out if there is any correlation between bcl-2 protein levels and cellular oxidative stress in the pathogenesis of viral hepatitis. METHODS: This study was carried out on 130 patients with viral hepatitis, 70 with chronic hepatitis, 30 with liver cirrhosis and 30 with hepatocellular carcinoma (HCC) in addition to 20 healthy persons as the control. Serum bcl-2 protein was estimated by enzyme-linked immunosorbent assay, serum malondialdehyde (MDA), nitric oxide (NO) and antioxidant enzymes (GSH, GSH-px, GR and SOD) were measured using spectrophotometric analysis. RESULTS: bcl-2 protein level was significantly elevated in the serum of HCC, cirrhosis and chronic hepatitis groups as compared to control group. There were significant positive correlations between higher bcl-2 protein level and viral hepatitis markers (HBsAg, anti-HCV antibodies) in HCC and cirrhotic patients as compared to chronic hepatitis group. An increase in oxidative stress markers (MDA, NO) and a decrease in antioxidant enzyme activities (SOD, GSH and GSH-px) were observed. However, there was a negative correlation between bcl-2 levels and GR in all studied patient groups. CONCLUSIONS: The release of oxidative free radicals, deficiency in antioxidant enzymes and the expression of bcl-2 protein might play a role in the pathogenesis of viral hepatitis. The ability to measure bcl-2 protein in the serum could be useful as a prognostic marker of cancer patients.

Why do mothers die in Dakahlia, Egypt?

AIM: To investigate the causes of maternal mortality in the Dakahlia Governorate in Egypt. METHODS: A confidential enquiry for each case of maternal mortality during the years 2004-2005 was carried out. RESULTS: One hundred and seventy-nine maternal deaths were reported, giving a maternal mortality rate (MMR) of 71.3/100,000 live births. 140 (78.2%) women died due to direct obstetric causes, 24 (13.4%) due to indirect causes and 15 (8.4%) due to accidental or unexplained causes. 44 (24.6%) women died during or following delivery by cesarean section and 91 (50.8%) during labor or within 24 h following delivery. Complications during cesarean delivery, postpartum hemorrhage and hypertensive disorders were the leading causes of maternal mortality. Death due to substandard care was encountered in 85% of cases. Obstetricians were responsible for 51% of causes of avoidable maternal death. CONCLUSION: Maternal mortality in Dakahlia, although declining, is still relatively high. To further reduce maternal mortality, deliveries should be conducted at well-equipped hospitals.

Insulin-like growth factor binding protein-3: a novel biomarker for the assessment of the synthetic capacity of hepatocytes in liver cirrhosis.

The liver is the major source of circulating insulin-like growth factor binding protein-3 (IGFBP-3). Because the hepatic tissue is deranged in cirrhotic patients, we measured serum IGFBP-3 concentrations by two-site immunoradiometric assay in sera from 37 cirrhotic patients with different stages of hepatic dysfunction. These were compared with IGFBP-3 levels from 11 healthy controls. Serum IGFBP-3 levels in patients with chronic liver disease were significantly lower than those of the control group (P < 0.0005). The mean percent decrease in cases of early liver cirrhosis, cirrhosis without, and cirrhosis with ascites were 44%, 59%, and 82% respectively, indicating that serum IGFBP-3 levels decrease as the severity of hepatic dysfunction increases. Moreover, the decrease was more pronounced in cases with hyperbilirubinemia, elevated serum transaminases, hypoalbuminemia, and prolonged prothrombin time. There was a significant positive correlation between serum IGFBP-3 and serum albumin, as well as a significant negative correlation between serum IGFBP-3 and prothrombin time. These results indicate the close correlation of IGFBP-3 levels to worsening of hepatic functions. The determination of serum IGFBP-3 level is a clinically useful marker for the assessment of the synthetic capacity of hepatocytes in cirrhotic patients and an early predictor of impending hepatic dysfunction as well.

Comparative results of the immunogenicity of Edmonston-Zagreb and Schwartz measles vaccines administered to 60 Egyptian infants.

The present study was conducted on sixty 9-11 month infants attending a primary care clinic in a rural Giza governorate area. Patients were divided into two groups: the first group comprised 42 infants who were vaccinated with the Edmonston Zagreb measles vaccine strain, whereas the second group comprised 18 infants who were vaccinated with the Schwartz measles vaccine strain. Estimation of measles antibody titer by neutralization testing was determined by the microtiter technique prior to and 4 weeks post vaccination. The overall serconversion rate was 85%. Three infants failed vaccination. The Edmonston-Zagreb strain was superior to the Schwartz strain in inducing immunity to non immune infants. The nutritional status of the study group was abnormal in almost 1/2 (29/60) infants and borderline in 1/3 (20/60).

Biosynthesis of phenolic compounds inVitis vinifera cell suspension cultures: Study on hydroxycinnamoyl CoA:ligase.

In cell suspensions cultures from grape berry pulp (Vitis vinifera cv. Gamay fréaux)hydroxycinnamoyl CoA ligase (CoAL) displayed maximum activity (100 %) forp-coumaric acid and then, in decreasing order, for ferulic acid (81.3 %) and caffeic acid (60.4 %). No activity was detected with sinapic and cinnamic acids. The changes in CoAL activity during the growth cycle of the culture displayed two peaks : the highest (6 h after subculturing) was linked with a strong increase in protein caused by dilution ; the second was weaker and occurred on the 7th day of culture.Grape cell suspension accumulated mainly peonidin (Pn) and cyanidin (Cy) glucosides (Pn 3-glucoside, Cy 3-glucoside, Pn 3-acetylglucoside, Pn 3-caffeylglucoside, Pn 3-p-coumarylglucoside, and Cy 3-p-coumarylglucoside). Maximum accumulation of anthocyanins was associated with the exponential growth phase of the culture and might be the result of the substantial increase in CoAL activity resulting from the effect of dilution. The second enzyme activity peak was probably oriented towards the acylation of anthocyanins since the percentage of acylated forms increased with time after subculturing.