Traditional healers working with primary care and mental health for early intervention in psychosis in young persons: protocol for the feasibility cluster randomised controlled trial.

OBJECTIVES: In low/middle-income countries (LMICs), more than half of patients with first-episode psychosis initially seek treatment from traditional and religious healers as their first care. This contributes to an excessively long duration of untreated psychosis (DUP). There is a need for culturally appropriate interventions to involve traditional and spiritual healers to work collaboratively with primary care practitioners and psychiatrists through task-shifting for early detection, referral and treatment of first episode of psychosis. METHODS: To prevent the consequences of long DUP in adolescents in LMICs, we aim to develop and pilot test a culturally appropriate and context-bespoke intervention. Traditional HEalers working with primary care and mental Health for early interventiOn in Psychosis in young pErsons (THE HOPE) will be developed using ethnographic and qualitative methods with traditional healers and caregivers. We will conduct a randomised controlled cluster feasibility trial with a nested qualitative study to assess study recruitment and acceptability of the intervention. Ninety-three union councils in district Peshawar, Pakistan will be randomised and allocated using a 1:1 ratio to either intervention arm (THE HOPE) or enhanced treatment as usual and stratified by urban/rural setting. Data on feasibility outcomes will be collected at baseline and follow-up. Patients, carers, clinicians and policymakers will be interviewed to ascertain their views about the intervention. The decision to proceed to the phase III trial will be based on prespecified stop-go criteria. ETHICS AND DISSEMINATION: Ethical approval has been obtained from Keele University Ethical Review Panel (ref: MH210177), Khyber Medical University Ethical Review Board (ref: DIR/KMU-EB/IG/001005) and National Bioethics Committee Pakistan (ref no. 4-87/NBC-840/22/621). The results of THE HOPE feasibility trial will be reported in peer-reviewed journals and academic conferences and disseminated to local stakeholders and policymakers. TRIAL REGISTRATION NUMBER: ISRCTN75347421.

Molecular characterization of Mycobacterium tuberculosis through MPT64 gene polymorphism using next-generation sequencing technology.

Aim: To characterize and analyze polymorphism of the MPT64 gene and evaluate AgMPT64-based immunochromatographic assay (ICA) specificity associated with polymorphism. Materials & methods: A total of 1449 suspected samples were tested for tuberculosis (TB), and the MPT64/rv1980c gene was sequenced using next-generation sequencing for polymorphism analysis. Results: Of the TB-positive individuals, 200 (13.80%), 186 (12.84%) and 129 (8.90%) were positive using the liquid culture, GeneXpert and fluorescence microscopy assays, respectively. Liquid culture medium-based samples were confirmed using ICA, in which 193 (96.5%) were positive while seven (3.5%) were negative. Out of 14 sequenced samples, seven were positive and seven negative; 13 were identical to the reference and just one (ICA positive) showed a C477A point mutation (F159L). Conclusion: The results indicate that AgMPT64 can be considered as a potent vaccine candidate.

Genomic Characterization of SARS-CoV2 from Peshawar Pakistan Using Next-Generation Sequencing.

This study aimed to characterize the whole genome of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV2) isolated from an oropharyngeal swab specimen of a Pashtun Pakistani patient using next-generation sequencing. Upon comparing the SARS-CoV2 genome to the reference genome, a total of 10 genetic variants were identified. Among the 10 genetic variants, 1 missense mutation (c.1139A > G, p.Lys292Glu) in the Open Reading Frame 1ab (ORF1ab) positioned at 112 in the non-structural protein 2 (NSP2) was found to be unique. Phylogenetic analysis (n = 84) revealed that the current SARS-CoV2 genome was closely clustered with 8 Pakistani strains belonging to Punjab, Federal Capital, Azad Jammu and Kashmir (AJK), and Khyber Pakhtunkhwa (KP). In addition, the current SARS-CoV2 genome was very similar to the genome of SARS-CoV2 reported from Guam, Taiwan, India, the USA, and France. Overall, this study reports a slight mismatch in the SARS-CoV2 genome, indicating the presence of a single unique missense mutation. However, phylogenetic analysis revealed that the current SARS-CoV2 genome was closely clustered with 8 other Pakistani strains.

Drug repositioning against COVID-19: a first line treatment.

COVID-19 disease caused by the SARS-CoV-2 virus has shaken our health and wealth foundations. Although COVID-19 vaccines will become available allowing for attenuation of disease progression rates, distribution of vaccines can create other challenges and delays. Hence repurposed drugs against SARS-CoV-2 can be an attractive parallel strategy that can be integrated into routine clinical practice even in poorly-resourced countries. The present study was designed using knowledge of viral pathogenesis and pharmacodynamics of broad-spectrum antiviral agents (BSAAs). We carried out the virtual screening of BSAAs against the SARS-CoV-2 spike glycoprotein, RNA dependent RNA polymerase (RdRp), the main protease (Mpro) and the helicase enzyme of SARS-CoV-2. Imatinib (a tyrosine kinase inhibitor), Suramin (an anti-parasitic), Glycyrrhizin (an anti-inflammatory) and Bromocriptine (a dopamine agonist) showed higher binding affinity to multiple targets. Further through molecular dynamics simulation, critical conformational changes in the target protein molecules were revealed upon drug binding which illustrates the favorable binding conformations of antiviral drugs against SARS-CoV-2 target proteins. The resulting drugs from the present study in combination and in cocktails from the arsenal of existing drugs could reduce the translational distance and could offer substantial clinical benefit to decrease the burden of COVID-19 illness. This also creates a roadmap for subsequent viral diseases that emerge.Communicated by Ramaswamy H. Sarma.

Association of ATM, CDH1 and TP53 genes polymorphisms with familial breast cancer in patients of Khyber Pakhtunkhwa, Pakistan.

Background: Genetic studies play a significant role in understanding the underlying risk factors of breast cancer. Polymorphism in the tumor suppressor gene TP 53, CDH1 and ATM genes are found to increase susceptibility for breast cancer globally. Objective: This study aimed to identify/analyze the contribution of genetic polymorphisms in the breast cancer candidate genes ATM, TP53 and CDH1 that may be associated with familial breast cancer risk in the Khyber Pakhtunkhwa population. Subjects and Methods: In the present case-control study, Whole Exome Sequencing (WES) of the 100 breast cancer patients and 100 ethnic controls were performed for the selected genes in the target population. Results: Of the studied variants rs3743674 of the CDH1 gene (crude P=0.014 and adjusted p=0.000) evident significant association with breast cancer in Pakistani Pashtun population. Whereas TP53rs1042522 (crude P=0.251 and adjusted P=0.851) and ATM rs659243 (crude p=0.256 and adjusted p=0.975) showed no or negative association with breast cancer in study population. Conclusion: The present study demonstrates that CDH1rs3743674 polymorphism is associated with elevated breast cancer risk in the Pashtun ethic population of Khyber Pakhtunkhwa.

Detection of SARs-CoV-2 in wastewater using the existing environmental surveillance network: A potential supplementary system for monitoring COVID-19 transmission.

The ongoing COVID-19 pandemic is caused by SARs-CoV-2. The virus is transmitted from person to person through droplet infections i.e. when infected person is in close contact with another person. In January 2020, first report of detection of SARS-CoV-2 in faeces, has made it clear that human wastewater might contain this virus. This may illustrate the probability of environmentally facilitated transmission, mainly the sewage, however, environmental conditions that could facilitate faecal oral transmission is not yet clear. We used existing Pakistan polio environment surveillance network to investigate presence of SARs-CoV-2 using three commercially available kits and E-Gene detection published assay for surety and confirmatory of positivity. A Two-phase separation method is used for sample clarification and concentration. An additional high-speed centrifugation (14000Xg for 30 min) step was introduced, prior RNA extraction, to increase viral RNA yield resulting a decrease in Cq value. A total of 78 wastewater samples collected from 38 districts across Pakistan, 74 wastewater samples from existing polio environment surveillance sites, 3 from drains of COVID-19 infected areas and 1 from COVID 19 quarantine center drainage, were tested for presence of SARs-CoV-2. 21 wastewater samples (27%) from 13 districts turned to be positive on RT-qPCR. SARs-COV-2 RNA positive samples from areas with COVID 19 patients and quarantine center strengthen the findings and use of wastewater surveillance in future. Furthermore, sequence data of partial ORF 1a generated from COVID 19 patient quarantine center drainage sample also reinforce our findings that SARs-CoV-2 can be detected in wastewater. This study finding indicates that SARs-CoV-2 detection through wastewater surveillance has an epidemiologic potential that can be used as supplementary system to monitor viral tracking and circulation in cities with lower COVID-19 testing capacity or heavily populated areas where door-to-door tracing may not be possible. However, attention is needed on virus concentration and detection assay to increase the sensitivity. Development of highly sensitive assay will be an indicator for virus monitoring and to provide early warning signs.

In silico analysis of non-synonymous missense SNPs (nsSNPs) in CPE, GNAS genes and experimental validation in type II diabetes mellitus through Next Generation Sequencing.

Non-synonymous missense SNPs (nsSNPs) in CPE and GNAS genes were investigated computationally. In silico identified nsSNPs were experimentally validated in type II diabetes mellitus (T2DM) in Pakistani Pathan population using next generation sequencing (NGS). Sixty two high-risk nsSNPs in CPE and 44 in GNAS were identified. Only 12 in GNAS were clinically significant. Thirty six high-risk nsSNPs in CPE and 08 clinically significant nsSNPs in GNAS lies in the most conserved regions. I-mutant predicted that nsSNPs decrease the proteins stability and ModPred predicted 20 and 12 post-translational modification sites in CPE and GNAS proteins respectively. Ramachandran plot showed 88.7% residues are in the most favored region of protein models. By experimentation, none of the nsSNPs were found to be associated with T2DM. In conclusion, this study differentiates the deleterious nsSNPs from the neutral ones. Although nsSNPs are not associated with T2DM, they can be targeted in other CPE and GNAS genes related disorders.

Fecal Microbiome and Resistome Profiling of Healthy and Diseased Pakistani Individuals Using Next-Generation Sequencing.

In this paper, we aimed to characterize the fecal microbiome and its resistomes of healthy and diseased subjects infected with multidrug-resistant Escherichia coli using next-generation sequencing (NGS). After initial screening, 26 stools samples belonging to healthy (n = 13) and diseased subjects (n = 13) were selected and subjected to NGS. A total of 23 and 42 antibiotic-resistant genes (ARGs) conferring resistance to 6 and 9 classes of antibiotics were identified in the resistomes of healthy and diseased subjects, respectively. Bacteroidetes were found to be the major phylum in both healthy and diseased subjects; however, Proteobacteria was predominantly present in the diseased subjects only. Microbial dysbiosis and predominance of various ARGs in the resistome of diseased subjects reflect the excessive usage of antibiotics in Pakistan and warrants immediate attention to regulate the use of various antimicrobials.

Resistome and microbial profiling of pediatric patient's gut infected with multidrug-resistant diarrhoeagenic Enterobacteriaceae using next-generation sequencing; the first study from Pakistan.

A high prevalence of multidrug-resistant (MDR) pathogens has been reported in adult and pediatric populations of Pakistan. However, data describing the effect of MDR microbes on the gut microbiota is scarce. We designed a cross-sectional pediatric study to investigate the effect of MDR microbes' infection on the gut microbiome and its resistome of children using high-throughput next-generation sequencing (NGS). A cross-sectional study was conducted at a tertiary health care hospital in Peshawar Pakistan, between 5 September 2019 to 15 February 2020. Pediatric patients with acute gastroenteritis (n = 200) were enrolled. All the enrolled pediatric patients underwent initial antimicrobial resistance (AMR) screening using the disk diffusion method. Children with MDR infections were identified and selected for gut microbiome and its resistome profiling using NGS. Out of 200 enrolled pediatric patients, 80 (40%) were found infected with MDR diarrheagenic Enterobacteriaceae consisting of 50 (62.5%) infections caused by extended-spectrum beta-lactamase (ESBL) producing E. coli while 30 (37.5%) by MDR Enterobacter specie. A total of 63 and 17 antibiotic-resistant genes (ARGs) conferring resistance to 7 and 5 classes of antibiotics were identified in the resistomes of MDR diarrheagenic Enterobacteriaceae infected and healthy children, respectively. NGS-based gut microbial profiling of MDR Enterobacter spp., ESBL producing E. coli infected pediatric patients and healthy controls revealed the predominance of Proteobacteria and Actinobacteria, respectively. An increased abundance of several pathogenic gram-negative bacteria namely E. coli, Enterobacter cloacae, and Salmonella enterica was observed in the gut microbiota of children infected with MDR bacterial infections than that of the healthy controls. This work indicates that children with MDR infections have reduced microbial diversity and enriched ARGs than healthy controls. The emergence of MDR bacterial strains and their association with gut dysbiosis needs immediate attention to regulate antibiotics usage in Pakistani children.

Assessment of genetic risk of type 2 diabetes among Pakistanis based on GWAS-implicated loci.

Genome-wide association studies (GWAS) have identified multiple type 2 diabetes (T2D) loci, mostly among populations of European descent. There is a high prevalence of T2D among Pakistanis. Both genetic and environmental factors may be responsible for this high prevalence. In order to understand the shared genetic basis of T2D among Pakistanis and Europeans, we examined 77 genome-wide significant variants previously implicated among European populations. We genotyped 77 single-nucleotide polymorphisms (SNPs) by iPLEX® Gold or TaqMan® assays in a case-control sample of 1,683 individuals. Association analysis was performed using logistic regression. A total of 16 SNPs (TCF7L2/rs7903146, GLIS3/rs7041847, CHCHD9/rs13292136, PLEKHA1/rs2292626, FTO/rs9936385, CDKAL1/rs7756992, KCNJ11/rs5215, LOC105372155/rs12970134, KCNQ1/rs163182, CTRB1/rs7202877, ST6GAL1/rs16861329, ADAMTS9-AS2/rs6795735, LOC105370275/rs1359790, C5orf67/rs459193, ZBED3-AS1/rs6878122 and UBE2E2/rs7612463) showed statistically significant associations after controlling for the false discovery rate. While KCNQ1/rs163182 and ZBED3-AS1/rs6878122 showed opposite allelic effects, the remaining significant SNPs had the same allelic effects as reported previously. Our data indicate that a selected number of T2D loci previously identified among populations of European descent also affect the risk of T2D in the Pakistani population.

Assessment of General Populations Knowledge, Attitude, and Perceptions Toward the Coronavirus Disease (COVID-19): A Cross-Sectional Study From Pakistan.

Background: Coronavirus disease 2019 (COVID-19) is a global health threat and caused a universal psychosocial impact on the general population. Therefore, the knowledge, attitude, and perceptions (KAPs) of the general population are critical for the development and effective implementation of standard operating procedures (SOP) to contain the contagion and minimize the losses. Therefore, the current study was conducted to understand and evaluate the KAPs of Pakistani populations toward the COVID-19. Methods: An online cross-sectional study was carried out among participants from 1 May to 30 July 2020 in different areas of Pakistan. The respondents of the study were the general population with age ≥ 18 years. The poll URL was posted on several channels after a call for participation. Other social media platforms such as WeChat, WhatsApp, Facebook, Twitter, Instagram, Messenger, and LinkedIn were engaged to maximize general population engagement. The questionnaire included details about sociodemographic, knowledge about COVID-19, perceptions toward universal safety precautions of COVID-19, and beliefs attitude toward the COVID-19. The obtained data were exported into a Microsoft Excel spreadsheet and SPSS software version 21 for windows. The descriptive statistics values were presented in frequencies and percentages. Binary logistic regression, Chi-square test, and one-way ANOVA were applied to analyze the participants' socio-demographic characteristics and variables related to KAPs. P-value < 0.05 was recorded as significant. Results: A total of 1,000 participants were invited of which 734 participated in this study. The response rate was 73.4% (734/1,000). The gender, marital status, education, and residence showed a significant association with the knowledge score. The majority of the study participants were thinking that COVID-19 may be more dangerous in elderly individuals 94.5% (n = 700), and individuals with chronic diseases or severe complications 96.7% (n = 710) (p = 0.00). More than half of the participants 52.5% (n = 385) showed their concern that either they or their family members might get the infection. More than 98% (n = 703), (P-value = 0.00) of the participants held that COVID-19 would be successfully controlled in Pakistan by following the standard SOPs and government guidelines. Conclusion: This study showed that the general population of Pakistan has good awareness and reasonable attitudes and perceptions toward the full features of the COVID-19. The current study suggests that mass-level effective health education programs are necessary for developing countries to improve and limit the gap between KAP toward COVID-19.

Clinical and laboratory relevance of JAK2 V617F and BCR-ABL co-existence in Philadelphia positive CML patients.

Chronic Myeloid Leukaemia (CML) is characterized by BCR-ABL1 mutation. A number of research studies have published reports of concomitant JAK2-V617F mutation in BCR-ABL positive Chronic Myeloid Leukaemia. This study aims to investigate the frequency of JAK2-V617F mutation in BCR-ABL positive CML cases. After approval from ethical committee, participants were enrolled in the study. A total of 103 samples from CML patients were analysed for the presence of JAK2-V617F mutation using real-time polymerase chain reaction. Patients were monitored for treatment response using real-time quantitative PCR for BCR-ABL1 mutation. Out of 103 samples analysed, 2 patients tested positive for JAK2-V617F mutation. These two patients when treated with standard Tyrosine Kinase Inhibitors (TKI) therapy achieved molecular response and normalized the haemoglobin and white cell counts. However, one patient has sustained thrombocytosis. JAK2 remained positive throughout the treatment course. We could not follow the second patient till the end of the study. JAK2 mutation in BCR-ABL1 mutated CML appears to be rare. Treatment with TKI does not appear to reduce JAK2 mutation burden despite a decrease in BCR-ABL1 copy numbers.

Next-Generation Sequencing Based Gut Resistome Profiling of Broiler Chickens Infected with Multidrug-Resistant Escherichia coli.

The study was designed to investigate the fecal microbiome and resistome of broiler chickens infected with multidrug-resistant (MDR) Escherichia coli (E. coli). Fecal samples (n = 410) from broiler chickens were collected from thirteen randomly selected sites of Khyber Pakhtunkhwa and screened for the presence of MDR E. coli. Upon initial screening, thirteen (13) MDR E. coli isolates were then subjected to shotgun metagenome next-generation sequencing (NGS). NGS based resistome analysis identified the multidrug efflux pump system-related genes at the highest prevalence (36%) followed by aminoglycoside (26.1%), tetracycline (15.9%), macrolide-lincosamide-streptogramin (9.6%), beta-lactam (6.6%), rifampin (2%), sulphonamide (1.3%), phenicol (0.91%), vancomycin (0.62%), trimethoprim (0.34%), colistin (0.30%), and quinolone (0.33%). The most abundant virulence-associated genes (VAGs) identified were iroN, iutA, iss, and iucA. NGS based taxonomic profiling at the phylum level revealed the predominance of Proteobacteria (38.9%) followed by Firmicutes (36.4%), Bacteroidetes (15.8%), and Tenericutes (8.9%). Furthermore, pathobionts such as E. coli, Salmonella enterica, Klebsiella pneumoniae, and Shigella flexneri belonging to the family Enterobacteriaceae were predominantly found. This study revealed the widespread presence of MDR genes, diverse VAGs, and a dysbiotic gut in the broiler chickens infected with MDR E. coli of Khyber Pakhtunkhwa for the first time using NGS.

Viroinformatics approach to explore the inhibitory mechanism of existing drugs repurposed to fight against COVID-19.

The rapid breakout of the coronavirus disease of 2019 (COVID-19) has been declared pandemic with serious global concern due to high morbidity and mortality. As we enter the phase beyond limitations there is an urgent need for explicit treatment against COVID-19. To face this immediate global challenge, drug development from scratch is a lengthy process and unrealistic to conquer this battle. Drug repurposing is an emerging and practical approach where existing drugs, safe for humans, are redeployed to fight this harder to treat disease. A number of multi clinical studies have repurposed combined cocktail (remdesivir + chloroquine and favipiravir + chloroquine) to be effective against COVID-19. However, the exact mechanistic aspect has not yet been revealed. In the present study, we have tried to decipher the mechanistic aspects of existing medicines at the viral entry and replication stage via the structural viroinformatics approach. Here we implied the molecular docking and dynamic simulations with emphasis on the unique structural properties of host receptor angiotensin-converting enzyme 2 (ACE2), SARS-CoV2 spike protein and RNA dependent RNA polymerase enzyme (RdRp) of the SARS-CoV2. Deep structural analysis of target molecules exposed key binding residues and structural twists involved in binding with important pharmacophore features of existing drugs [(7-chloro-N-[5-(diethylamino)pentan-2-yl]quinolin-4-amine (chloroquine),N-[[4-(4-methylpiperazin-1-yl)phenyl]methyl]-1,2-oxazole-5-carboxamide N-[[4-(4-methylpiperazin-1-yl)phenyl]methyl]-1,2-oxazole-5-carboxamide) (SSAA09E2), 2-ethylbutyl (2S)-2-{[(S)-{[(2R,3S,4R,5R)-5-{4-aminopyrrolo[2,1-f][1,2,4]triazin-7-yl}-5-cyano-3 (remdesivir) and 6-Fluor-3-oxo-3,4-dihydro-2-pyrazincarboxamid (favipiravir)]. It is evident from this structural informatics study that combo of chloroquine + SSAA09E2 with remdesivir or favipiravir could significantly restrain the virus at the entry and replication stage. Thus, drug repurposition is an attractive approach with reduced time and cost to treat COVID-19, we don't have enough time as the whole world is lockdown and we are in urgent need of an obvious therapeutics' measures.

Development of the Next Generation Sequencing-Based Diagnostic Test for ß-Thalassemia and its Validation in a Pashtun Family.

ß-Thalassemia (ß-thal) is a common monogenic disease with ethnic-specific mutations on the HBB gene throughout the world. The reported mutations either reduce the expression or completely inactivate the HBB gene. In Pakistan, the prevalence of ß-thal is high due to consanguineous marriages. Accurate identification of mutations in carriers is imperative for prevention of ß-thal in subsequent generations. To overcome the limitations of traditional testing methods for ß-thal, a next-generation sequencing (NGS)-based diagnostic test was designed and validated by sequencing the entire HBB gene. The primer set covering the entire HBB gene was designed and validated in a Pashtun ß-thalassemic family. The polymerase chain reaction (PCR) product was sequenced using an Illumina MiSeq platform. A homozygous pathogenic insertion of A>AC/AC (rs35699606) was detected in an affected member of the family, while unaffected members were heterozygous for it. In addition, all family members were homozygous for the synonymous variant, A>G/G (rs713040), except the father who was heterozygous for it. We sequenced the entire HBB gene using the NGS-based test, which is highly sensitive, robust and specific for the diagnosis and screening of ß-thal in Pakistan, especially for families practicing consanguineous marriages.

Novel deleterious mutation in MYO7A, TH and EVC2 in two Pakistani brothers with familial deafness.

OBJECTIVE: In Pakistan, 74% of consanguineous marriages are among the first cousins. Continuity of consanguineous marriages over generations increases the risk of recessive diseases such as deafness. The objective of this study was to investigate genetic origin of Pakistani deaf brothers with parents of consanguineous marriage. METHODS: DNA was extracted from the blood through Qiagen kit. Paired-end sequencing library was prepared according to protocol of Illumina's TruSight Rapid Capture kit and TruSight Inherited Disease Panel. Library was normalized and used for Next Generation Sequencing through MiSeq. NGS data were analyzed using various bioinformatics tools. RESULTS: Both brothers were found to have novel deleterious mutation in MYO7A (c.2476G>A) while the younger brother had additional novel deleterious mutation in TH (c.43C>T) and EVC2 (c.2614C>T) genes. CONCLUSION: It is concluded that in addition to novel mutations in MYO7A, TH and EVC2, the CDH23 and GJB2 can also be responsible for deafness in the family with consanguineous marriages.

Synthesis of secondary metabolites by Cladosporium resinae (NRL-6437) under different growth media and chemical inducers and their pharmaceutical activity.

The role of different growth media and chemical enhancer on synthesis of secondary metabolites Cladosporium resinae (NRL-6437) was investigated for their in vitro biological activities. Cladosporium resinae (NRL-6437) were grown in various nutrient media (Czapeak-dox Broth (CB), Czapeak Yeast-extract Broth (CYB), Yeast Extract Sucrose (YES), Potato Dextrose Broth (PDB) and Czapeak-dox (supplemented with glucose and starch) Broth (CGSB) for the production of metabolites. Two chemical epigenetic modifiers (suberoyl-anilide hydroxamic acid (SAHA) and 5-azacytidine (5-AZA) were also used for the expression of silent genes for secondary metabolite production. Our results indicated that among different media, Czapeak yeast extract broth produced more secondary metabolites. Application of 15mM of both modifiers was effective for the expressions of silent genes resulting in an increased metabolites production. Secondary metabolites extracted in ethyl acetate and fractionized in n-Hexane were also tested for their biological activity. The secondary metabolites revealed varying degrees of growth inhibitions of the tested organisms. Similarly, these metabolites were also active against brine shrimps and Lemna.

Absence of SNCA polymorphisms in Pakistani Parkinsons disease patients.

OBJECTIVE: To elucidate the genetic risk and role of alpha-synuclein gene in the pathogenesis of Parkinson's disease in Pakistani population. METHODS: This case-control study was conducted at Institute of Biomedical and Genetic Engineering (IBGE), Islamabad from May 2013 to May 2016, and comprised patients with Parkinson's disease and their ethnically-matched healthy controls. Allele-specific polymerase chain reaction was used for screening of three pathogenic single nucleotide polymorphisms in alpha-synuclein gene. Moreover, 20% samples were randomly selected for bidirectional Sanger sequencing to confirm the results. SPSS 13 was used for data analysis. RESULTS: Of the 374 participants, 174(46.5%) were patients and 200(53.5%) were controls. The mean age for the onset of the disease was 55±13 years. No polymorphism was observed for rs104893875(G>A), rs104893877(G>A) and rs104893878(C>G) in alpha-synuclein gene in samples of patients and controls. CONCLUSIONS: Alpha-synuclein gene mutations might not be relevant to all the populations in causing Parkinson's disease.

Genetic etiology of oral cancer.

Head and neck squamous cell carcinoma (HNSCC) is the sixth most common cancer worldwide. It accounts for 2.5% of all new cancer cases and 1.9% of all cancer deaths annually. More than 90% of oral cancers (occurring in the mouth, lip, and tongue) are oral squamous cell carcinoma. The incidence rate of oral cancer varies widely throughout the world, with an evident prevalence in South Asian countries. This high incidence occurs in correlation with oral cancer-associated behaviors such as alcohol, tobacco use. Researchers have reported that these behaviors lead to genetic variations in tumor suppressor genes (APC, p53), proto-oncogenes (Myc), oncogene (Ras) and genes controlling normal cellular processes (EIF3E, GSTM1). Processes such as segregation of chromosomes, genomic copy number, loss of heterozygosity, telomere stabilities, regulations of cell-cycle checkpoints, DNA damage repairs and defects in notch signaling pathways are involved in causing oral cancer. In order to develop preventive and therapeutic options, it is necessary to comprehend the basic molecular mechanisms forcing oral tumorigenesis. This review examines, in detail, the mechanisms of genetic alteration which are considered to be responsible for the initiation of oral cancer.

WaveCNV: allele-specific copy number alterations in primary tumors and xenograft models from next-generation sequencing.

MOTIVATION: Copy number variations (CNVs) are a major source of genomic variability and are especially significant in cancer. Until recently microarray technologies have been used to characterize CNVs in genomes. However, advances in next-generation sequencing technology offer significant opportunities to deduce copy number directly from genome sequencing data. Unfortunately cancer genomes differ from normal genomes in several aspects that make them far less amenable to copy number detection. For example, cancer genomes are often aneuploid and an admixture of diploid/non-tumor cell fractions. Also patient-derived xenograft models can be laden with mouse contamination that strongly affects accurate assignment of copy number. Hence, there is a need to develop analytical tools that can take into account cancer-specific parameters for detecting CNVs directly from genome sequencing data. RESULTS: We have developed WaveCNV, a software package to identify copy number alterations by detecting breakpoints of CNVs using translation-invariant discrete wavelet transforms and assign digitized copy numbers to each event using next-generation sequencing data. We also assign alleles specifying the chromosomal ratio following duplication/loss. We verified copy number calls using both microarray (correlation coefficient 0.97) and quantitative polymerase chain reaction (correlation coefficient 0.94) and found them to be highly concordant. We demonstrate its utility in pancreatic primary and xenograft sequencing data. AVAILABILITY AND IMPLEMENTATION: Source code and executables are available at https://github.com/WaveCNV. The segmentation algorithm is implemented in MATLAB, and copy number assignment is implemented Perl. CONTACT: lakshmi.muthuswamy@gmail.com SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.