A Missense Pathogenic Variant in a Conserved Region of CNTNAP2 Is Associated with Obesity, Seizures, and Language Impairment in a Pakistani Family.

Introduction: In a consanguineous family, seven siblings born in three sibships showed a syndromic disorder characterized by obesity, seizures, and language impairment phenotypes, which appeared at early age or developed during early childhood. Methods: By whole-exome sequencing and subsequent Sanger sequencing, a novel homozygous missense variant (c.3371 T>A [p.Ile1124Asn]) in exon 20 of the CNTNAP2 gene was identified. Results: The pathogenic variant in this family is located within one of the laminin G-like 4 domains of CASPR2 and may cause loss of hydrophobic interactions of CASPR2 with its partner proteins. Single nucleotide and copy number variants in this gene have previously been related to Gilles de la Tourette syndrome, cortical dysplasia-focal epilepsy syndrome, schizophrenia, Pitt-Hopkins syndrome, and autism spectrum, attention deficit hyperactivity, and obsessive compulsive disorders. Yet, few studies described patients with CNTNAP2 variants showing diet-induced obesity. Conclusion: This report expands the phenotypic spectrum of this rare syndrome and provides deeper insights by documenting the clinical features and genetic findings of the patients.

Production of commercially important enzymes from Bacillus licheniformis KIBGE-IB3 using date fruit wastes as substrate.

BACKGROUND: Pakistan is one of the top five date fruit-producing countries and produced more than 30% wastes in picking, packing, storage, and commercialization stages. The date fruit wastes are usually considered inedible for humans and only used for livestock feed. In current research, Bacillus licheniformis KIBGE-IB3 was screened for pectinase, xylanase, cellulase, and amylase production using date fruit wastes as substrate through solid state fermentation. RESULTS: The B. licheniformis KIBGE-IB3 produced higher concentration of pectinase using date fruit wastes as substrate as compared to amylase, cellulase, and xylanase. B. licheniformis KIBGE-IB3 produced maximum pectinase using 5.0 g/dl date fruit wastes and 0.5 g/dl yeast extract. B. licheniformis KIBGE-IB3 required pH 7.0, 37 °C incubation temperature, and 72 h incubation period for maximum production of pectinase. CONCLUSION: It has been concluded that date fruit waste is a good source of biomass and can be utilized for the commercial production of pectinase.

Effect of Aloe vera gel coating enriched with Fagonia indica plant extract on physicochemical and antioxidant activity of sapodilla fruit during postharvest storage.

The effect of Aloe vera (AV) gel (at 50% or 100%) alone or enriched with Fagonia indica (FI) plant extract at 1% on physiological and biochemical responses of sapodilla fruit were studied during storage at 20 °C for 12 days. Sapodilla fruit treated with AV 100% and FI 1% significantly reduced weight loss, decay incidence, soluble solids concentration, and kept a high level of firmness and titratable acidity compared to the untreated fruit. FI 1% added to AV 50% or AV 100% efficiently maintained higher ascorbic acid, total flavonoids, total phenolics and radical scavenging activity of sapodilla fruit. The panelists did not detect any negative effect of AV gel and FI plant extract on the sensory attributes of sapodilla fruit. Therefore, the addition of FI plant extract to AV gel coating could be a promising approach to prolong the shelf life and preserve the quality of sapodilla fruit during storage.

Past, present and future of hepatitis E virus infection: Zoonotic perspectives.

The origin of hepatitis E virus (HEV) is not fully understood, but it is considered an emerging zoonotic pathogen. To date, HEV has been isolated from many animal species. The family Hepeviridae consists of two genera. The genus Orthohepevirus includes four distinct species (A, B, C, and D), each with distinct genotypes. Within the Orthohepevirus A species, HEV-1 and HEV-2 host ranges are restricted to humans, whereas genotypes 3 and 4 primarily infect a wide range of diverse animal species, in addition to being zoonotic to humans. Swine and wild boar species were previously thought to be the primary natural HEV reservoir, but recently rabbits have also been identified as major carriers. Moreover, increasing the number of HEV infections within the food supply chain underscore the important role of farming and food processing practices in limiting virus transmission. Notably, a Chinese commercial vaccine has the potential to protect humans and possibly animal reservoirs from HEV infection. This review summarizes the status of HEV infection worldwide in different animal species and outlines various modes of zoonotic transmission, with reference to cross-species transmission and recent vaccine developments.

Malnutrition and poor oral health status are major risks among primary school children at Lasbela, Balochistan, Pakistan.

BACKGROUND: This survey was focusing on health and oral hygiene status of primary school children at Lasbela district considering the comparatively less developed and socio demographically deprived part of the Country. METHODS: A cross sectional survey was conducted to determine the health status of primary school children in seven tehseels of district Lasbela, Balochistan after applying proportionate sampling technique from March 2015 to July 2015. Field teams visited assigned schools to screen children and collect health related data on predesigned and pre coded proforma. RESULTS: Out of 200 schools, 196 schools found opened, while 2% of schools (04) remained closed. A total of 6363 students were clinically screened. About 45% of the school children had normal body mass index (BMI) and rest were falling in different categories of malnutrition. More than 19% had ear, nose and throat (ENT) problems and around 19% presented with clinical anemia. Less than 50% of children had scar of BCG vaccination and 4% informed about use of gutka/supari chewing (smokeless tobacco use). CONCLUSION: In conclusion, we estimated high prevalence of malnutrition, poor oral health including smokeless tobacco use, and low BCG coverage among primary school children at Lasbela. Current scenario suggests immediate and contextually focused interventions to confine existing public health risks and avoid future burden of disease.

Novel Nonsense Variants c.58C>T (p.Q20X) and c.256G>T (p.E85X) in the CHEK2 Gene Identified in Breast Cancer Patients from Balochistan.

Breast cancer is very common and the leading cause of cancer deaths among women globally. Hereditary cases account for 510% of the total burden and CHEK2, which plays crucial role in response to DNA damage to promote cell cycle arrest and repair or induce apoptosis, is considered as a moderate penetrance breast cancer risk gene. Our objective in the current study was to analyze mutations in related to breast cancer. A total of 271 individuals including breast cancer patients and normal subjects were enrolled and all 14 exons of CHEK2 were amplified and sequenced. The majority of the patients (>95%) were affected with invasive ductal carcinoma (IDC), 52.1% were diagnosed with grade III tumors and 56.2% and 27.5% with advanced stages III and IV. Two novel nonsense variants i.e. c.58C>T (P.Q20X) and c.256G>T (p.E85X) at exon 1 and 2 in two breast cancer patients were identified, both novel and not reported elsewhere.

Novel Nonsense Variants c.58C>T (p.Q20X) and c.256G>T (p.E85X) in the CHEK2 Gene Identified dentified in Breast Cancer Patients from Balochistan.

Breast cancer is the most commonly occurring and leading cause of cancer deaths among women globally. Hereditary cases account 5-10% of all the cases and CHEK2 is considered as a moderate penetrance breast cancer risk gene. CHEK2 plays a crucial role in response to DNA damage to promote cell cycle arrest and repair DNA damage or induce apoptosis. Our objective in the current study was to analyze mutations in the CHEK2 gene related to breast cancer in Balochistan. A total of 271 individuals including breast cancer patients and normal subjects were enrolled. All 14 exons of CHEK2 were amplified and sequenced. The majority of the patients (>95%) had invasive ductal carcinomas (IDCs), 52.1% were diagnosed with tumor grade III and 56.1% and 27.5% were diagnosed with advance stages III and IV. Two novel nonsense variants i.e. c.58C>T (P.Q20X) and c.256G>T (p.E85X) at exon 1 and 2 in two breast cancer patients were identified in the current study. Both the variants identified were novel and have not been reported elsewhere.

Immobilization of pectin depolymerising polygalacturonase using different polymers.

Polygalacturonase catalyses the hydrolysis of pectin substances and widely has been used in food and textile industries. In current study, different polymers such as calcium alginate beads, polyacrylamide gel and agar-agar matrix were screened for the immobilization of polygalacturonase through entrapment technique. Polyacrylamide gel was found to be most promising one and gave maximum (89%) immobilization yield as compared to agar-agar (80%) and calcium alginate beads (46%). The polymers increased the reaction time of polygalacturonase and polymers entrapped polygalacturonases showed maximum pectinolytic activity after 10 min of reaction as compared to free polygalacturonase which performed maximum activity after 5.0 min of reaction time. The temperature of polygalacturonase for maximum enzymatic activity was increased from 45°C to 50°C and 55°C when it was immobilized within agar-agar and calcium alginate beads, respectively. The optimum pH (pH 10) of polygalacturonase was remained same when it was immobilized within polyacrylamide gel and calcium alginate beads, but changed from pH 10 to pH 9.0 after entrapment within agar-agar. Thermal stability of polygalacturonase was improved after immobilization and immobilized polygalacturonases showed higher tolerance against different temperatures as compared to free enzyme. Polymers entrapped polygalacturonases showed good reusability and retained more than 80% of their initial activity during 2nd cycles.

Purification and characterization of thiol dependent, oxidation-stable serine alkaline protease from thermophilic Bacillus sp.

Alkaline serine protease was purified to homogeneity from culture supernatant of a thermophilic, alkaliphilic Bacillus sp. by 80% ammonium sulphate precipitation followed by CM-cellulose and DEAE-cellulose ion exchange column chromatography. The enzyme was purified up to 16.5-fold with 6900 U/mg activity. The protease exhibited maximum activity towards casein at pH 8.0 and at 80 °C. The enzyme was stable at pH 8.0 and 80 °C temperature up to 2 h. The Ca2+ and Mn2+ enhanced the proteolytic activity up to 44% and 36% as compared to control, respectively. However, Zn2+, K+, Ba2 +, Co2 +, Hg2+ and Cu2+ significantly reduced the enzyme activity. PMSF (phenyl methyl sulphonyl fluoride) completely inhibited the protease activity, whereas the activity of protease was stimulated up to two folds in the presence of 5 mM 2-mercaptoethanol. The enzyme was also stable in surfactant (Tween-80) and other commercial detergents (SDS, Triton X-100).

Morphological and molecular based identification of pectinase producing Bacillus licheniformis from rotten vegetable.

Pectinase catalyzed the degradation of pectin substances and has been used in various biotechnological industries. In the current study, 23 bacterial strains were isolated from rotten vegetables, soil and air. The isolated bacterial strains were qualitatively screened for pectinase production on pectin agar medium and only three strains HR 4, HR 21 and HR 23 were observed to produce extracellular pectinase. These strains were further screened quantitatively for pectinase production through submerged fermentation technology in pectin containing fermentation medium. Strain HR 4 from rotten brinjal (Solanum melongena) was found to produce higher pectinase as compared to others. The maximum pectinase producing bacterial strain was identified as Bacillus licheniformis on the basis of morphological, physiological and biochemical characteristics. For further confirmation of identification, 16S rDNA sequence analysis was performed. The 16S rDNA sequences were aligned and the phylogenetic tree was constructed. The phylogenetic tree confirmed that the strain was belonging to B. licheniformis. The 16S rDNA sequences of this new strain were submitted to GenBank and designated as B. licheniformis KIBGE-IB21 with the GenBank accession number JQ 411812. The newly isolated pectinase producing B. licheniformis used apple pectin as carbon and yeast extract as nitrogen source for maximum pectinase production.

Missense mutations (p.H371Y, p.D438Y) in gene CHEK2 are associated with breast cancer risk in women of Balochistan origin.

CHEK2 encodes a serine/threonine-protein kinase which plays a critical role in DNA damage signaling pathways. CHEK2 directly phosphorylates and regulates the functions of p53 and BRCA1. Most women with breast and/or ovarian cancer are not carriers of mutant BRCA1 or BRCA2. Multiple studies have shown that a CHEK2*1100delC confers about a two-fold increased risk of breast cancer in unselected females and a tenfold increase in males. Moreover, studies have shown that first-degree relatives of bilateral breast cancer cases who carried the CHEK2*1100delC allele had an eight-fold increased risk of breast cancer. It has been suggested that CHEK2 functions as a low-penetrance susceptibility gene for cancers and multiplies the risks associated with other gene(s) to increase cancer risk. The main goal of this study was to evaluate and to compare the role of truncating mutations, splice junction mutations and rare missense substitutions in breast cancer susceptibility gene CHEK2. Present study was performed on 140 individuals including 70 breast cancer patients both with and without family history and 70 normal individuals. Written consent was obtained and 3 ml intravenous blood was drawn from all the subjects. DNA was extracted from all the samples through inorganic method published already. Primers were synthesized for all the 14 exons of CHEK2 gene. Coding and adjacent intronic sequences of CHEK2 gene were amplified and sequenced. Two genetic variants (p.H371Y, p.D438Y) were found in exon 10 and exon 11 of gene CHEK2 which were not found in any of the 70 control individuals from same geographical area and ethnic group. The genetic variant c.1312G>T (p.D438Y) identified in a patient with a family history of breast cancer. To our knowledge, this is first mutation scanning study of gene CHEK2 from Balochistan population.

Various aspects, patterns and risk factors in breast cancer patients of Balochistan.

PURPOSE: Breast cancer is the commonest malignancy of females throughout the world with one million new cases each year. In Pakistan, the burden of breast cancer disease is high with late stage presentation being a common feature, more than half being stage III or stage IV. The objective of this study was to study various aspects, patterns and risk factors in breast cancer patients of Balochistan. METHOD: Present study was performed on 134 patients of breast cancer who were registered in CENAR. The patients were interviewed by providing a questionnaire. Informed consent was taken from all the patients who took part in this study after explanation of the study aims. Body mass index (BMI) was calculated andbiopsy reports were obtained from patients files. All the cases were classified with respect to age, gender, ethnic group (Baloch, Pashtoon, Punjabi, Afghani, Hazara) BMI, cancer type, cancer grade, hormonal status, side of the cancer, fertility and marital status. RESULTS: Out of 134 patients, the most common ethnic group was Pashtoon with a total of 42 and the common age group was 41-50 years with a total of 51. Invasive ductal carcinoma (IDC) was the most common type, accounting for in 128 patients (95.5%) followed by invasive lobular carcinoma (ILC). CONCLUSION: Pashtoon was the most common ethnic group, IDC was common type and most of the patients had an ER/PR positive hormonal status.

A missense mutation (p.G274R) in gene ASPA causes Canavan disease in a Pakistani family.

Canavan disease (OMIM 271900) is an autosomal recessive lethal neurodegenerative disorder characterized by spongy degeneration of the brain. A highly consanguineous Pakistani family with Canavan disease was enrolled on the basis of diagnosis. All the affected individuals have mental retardation, megalocephaly and degradation of motor skills, poor head control, partial vision loss, weakness of the muscles and raised urinary concentration of N-acetyl aspartic acid in the urine. Blood samples were collected from affected as well as normal siblings and processed for DNA purification. Linkage analysis was performed by typing three short tandem repeat markers D17S1583 (7.19 cM), D17S1828 (10.02 cM) and D17S919 (14.69 cM) for an already-reported gene/locus ASPA at chromosome 17p13.2 causing Canavan disease. During linkage analysis, all the affected individuals were homozygous for short tandem repeat markers while the normal siblings were heterozygous showing co-segregation of the disease. Gene ASPA (NM_000049) was undertaken to sequence for mutation analysis. As a result of sequence analysis, we found missense substitution 740A→G (p.G274R) in exon 6 of gene ASPA. To our knowledge, this is the first report about Canavan disease on a Pakistani family.