The chloroplast genome of Farsetia hamiltonii Royle, phylogenetic analysis, and comparative study with other members of Clade C of Brassicaceae.

BACKGROUND: Farsetia hamiltonii Royle is a medicinally important annual plant from the Cholistan desert that belongs to the tribe Anastaticeae and clade C of the Brassicaceae family. We provide the entire chloroplast sequence of F.hamiltonii, obtained using the Illumina HiSeq2500 and paired-end sequencing. We compared F. hamiltonii to nine other clade C species, including Farsetia occidentalis, Lobularia libyca, Notoceras bicorne, Parolinia ornata, Morettia canescens, Cochlearia borzaeana, Megacarpaea polyandra, Biscutella laevigata, and Iberis amara. We conducted phylogenetic research on the 22 Brassicaceae species, which included members from 17 tribes and six clades. RESULTS: The chloroplast genome sequence of F.hamiltonii of 154,802 bp sizes with 36.30% GC content and have a typical structure comprised of a Large Single Copy (LSC) of 83,906 bp, a Small Single Copy (SSC) of 17,988 bp, and two copies of Inverted Repeats (IRs) of 26,454 bp. The genomes of F. hamiltonii and F. occidentalis show shared amino acid frequencies and codon use, RNA editing sites, simple sequence repeats, and oligonucleotide repeats. The maximum likelihood tree revealed Farsetia as a monophyletic genus, closely linked to Morettia, with a bootstrap score of 100. The rate of transversion substitutions (Tv) was higher than the rate of transition substitutions (Ts), resulting in Ts/Tv less than one in all comparisons with F. hamiltonii, indicating that the species are closely related. The rate of synonymous substitutions (Ks) was greater than non-synonymous substitutions (Ka) in all comparisons with F. hamiltonii, with a Ka/Ks ratio smaller than one, indicating that genes underwent purifying selection. Low nucleotide diversity values range from 0.00085 to 0.08516, and IR regions comprise comparable genes on junctions with minimal change, supporting the conserved status of the selected chloroplast genomes of the clade C of the Brassicaceae family. We identified ten polymorphic regions, including rps8-rpl14, rps15-ycf1, ndhG-ndhI, psbK-psbI, ccsA-ndhD, rpl36-rps8, petA-psbJ, ndhF-rpl32, psaJ-rpl3, and ycf1 that might be exploited to construct genuine and inexpensive to solve taxonomic discrepancy and understand phylogenetic relationship amongst Brassicaceae species. CONCLUSION: The entire chloroplast sequencing of F. hamiltonii sheds light on the divergence of genic chloroplast sequences among members of the clade C. When other Farsetia species are sequenced in the future, the full F. hamiltonii chloroplast will be used as a source for comprehensive taxonomical investigations of the genus. The comparison of F. hamiltonii and other clade C species adds new information to the phylogenetic data and evolutionary processes of the clade. The results of this study will also provide further molecular uses of clade C chloroplasts for possible plant genetic modifications and will help recognise more Brassicaceae family species.

Identification of putative genetic variants in major depressive disorder patients in Pakistan.

BACKGROUND: Major depressive disorder (MDD) is a polygenic, and highly prevalent disorder affecting 322 million people globally. It results in several psychological changes which adversely affect different dimensions of life and may lead to suicide. METHODS: Whole exome sequencing of 15 MDD patients, enrolled at the Dr. A. Q. Khan Institute of Behavioral Sciences, Karachi, was performed using NextSeq500. Different bioinformatics tools and databases like ANNOVAR, ALoFT, and GWAS were used to identify both common and rare variants associated with the pathogenesis of MDD. RESULTS: A total of 1985 variations were identified in 479 MDD-related genes. Several SNPs including rs1079610, rs11750538, rs1799913, rs1801131, rs2230267, rs2231187, rs3819976, rs4314963, rs56265970, rs587780434, rs6330, rs75111588, rs7596487, and rs9624909 were prioritized due to their deleteriousness and frequency difference between the patients and the South Asian population. A non-synonymous variation rs56265970 (BCR) had 26% frequency in patients and was not found in the South Asian population; a multiallelic UTR-5' insertion rs587780434 (RELN) was present with an allelic frequency of 70% in patients whereas 22% in the SAS population. Genetic alterations in PABPC1 genes, a stress-associated gene also had higher allele frequency in the cases than in the normal population. CONCLUSION: This present study identifies both common and rare variants in the genes associated with the pathogenesis of MDD in Pakistani patients. Genetic variations in BCR, RELN, and stress-associated PABPC1 suggest potential roles in the pathogenesis of MDD.

Pan-genomics, drug candidate mining and ADMET profiling of natural product inhibitors screened against Yersinia pseudotuberculosis.

Yersinia pseudotuberculosis belongs to the family Enterobacteriaceae and is responsible for scarlatinoid fever, food poisoning, post-infectious complications like erythema nodosum/reactive arthritis as well as pseudoappendicitis in children. Genome sequences of the 23 whole genomes from NCBI were utilized for conducting the pan-genomic analysis. Essential proteins from the core region were obtained and drug targets were identified using a hierarchal in silico approach. Among these, multidrug resistance protein sub-unit mdtC was chosen for further analysis. This protein unit confers resistance to antibiotics upon forming a tripartite complex with units A and B in Escherichia coli. Details of the function have not yet been elucidated experimentally in Yersinia spp. Computational structure modeling and validation were followed by screening against phytochemical libraries of traditional Indian (Ayurveda), North African, and traditional Chinese flora using Molecular Operating Environment software version 2019.0102. ADMET profiling and descriptor study of best docked compounds was studied. Since phytotherapy is the best resort to antibiotic resistance so these compounds should be tested experimentally to further validate the results. The obtained information could aid wet-lab scientists to work on the scaffold of screened drug-like compounds from natural resources. This could be useful in our quest for antibiotic-resistant therapy against Y. pseudotuberculosis.

Comprehensive comparative genomic and microsatellite analysis of SARS, MERS, BAT-SARS, and COVID-19 coronaviruses.

The coronavirus disease 2019 (COVID-19) pandemic has spread around the globe very rapidly. Previously, the evolution pattern and similarity among the COVID-19 causative organism severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and causative organisms of other similar infections have been determined using a single type of genetic marker in different studies. Herein, the SARS-CoV-2 and related ß coronaviruses Middle East respiratory syndrome coronavirus (MERS-CoV), SARS-CoV,  bat coronavirus (BAT-CoV) were comprehensively analyzed using a custom-built pipeline that employed phylogenetic approaches based on multiple types of genetic markers including the whole genome sequences, mutations in nucleotide sequences, mutations in protein sequences, and microsatellites. The whole-genome sequence-based phylogeny revealed that the strains of SARS-CoV-2 are more similar to the BAT-CoV strains. The mutational analysis showed that on average MERS-CoV and BAT-CoV genomes differed at 134.21 and 136.72 sites, respectively, whereas the SARS-CoV genome differed at 26.64 sites from the reference genome of SARS-CoV-2. Furthermore, the microsatellite analysis highlighted a relatively higher number of average microsatellites for MERS-CoV and SARS-CoV-2 (106.8 and 107, respectively), and a lower number for SARS-CoV and BAT-CoV (95.8 and 98.5, respectively). Collectively, the analysis of multiple genetic markers of selected ß viral genomes revealed that the newly born SARS-COV-2 is closely related to BAT-CoV, whereas, MERS-CoV is more distinct from the SARS-CoV-2 than BAT-CoV and SARS-CoV.

Distinct genetic landscape and a low response to doxorubicin in a luminal-A breast cancer cell line of Pakistani origin.

BACKGROUND: Breast cancers exhibit genetic heterogeneity which causes differential responses to various chemotherapy agents. Given the unique demographic and genomic background in South Asia, genetic architecture in breast cancers is not fully explored. METHODS AND RESULTS: In this study, we determined the genetic landscape of our previously established luminal-A subtype breast cancer cell line (BC-PAK1), and compared it with a Caucasian origin MCF7 breast cancer cell line of the same molecular subtype. Deep whole-exome sequencing (100X) was performed from early passages of the primary cancer cells using the Illumina NextSeq500. Data analysis with in silico tools showed novel non-silent somatic mutations previously not described in breast cancers, including a frameshift insertion (p.Ala1591AlafsTer28) in CIC, and a frameshift deletion (p.Lys333LysfsTer21) in PABPC1. Five genes CDC27, PIK3CG, ARAP3, RAPGEF1, and EFNA3, related with cell cycle pathway (hsa04110), ErbB signaling pathway (hsa04012), Ras signaling pathway (hsa04014), and Rap1 signaling pathway (hsa04015) were found to have recurrent non-silent somatic mutations. Further, the major contribution of COSMIC signatures 3 (failure of DNA double-strand break repair by homologous recombination), and 12 (transcriptional strand-bias for T>C substitutions) was observed. Also, the somatic mutations landscape in BC-PAK1 was found to be different as compared to the MCF7 cell line. The unique genetic landscape of BC-PAK1 might be responsible for significantly reduced response to doxorubicin than the MCF7 cell line. CONCLUSION: This study presents a distinct genetic architecture in luminal-A breast cancer potentially responsible for differential response to chemotherapy. Further studies on large cohorts of breast cancer patients are suggested for implementation in personalized medicine.

An Overview of Algorithms and Associated Applications for Single Cell RNA-Seq Data Imputation.

Single cell RNA-Seq technology enables the assessment of RNA expression in individual cells. This makes it popular in experimental biology for gleaning specifications of novel cell types as well as inferring heterogeneity. Experimental data conventionally contains zero counts or dropout events for many single cell transcripts. Such missing data hampers the accurate analysis using standard workflows, designed for massive RNA-Seq datasets. Imputation for single cell datasets is done to infer the missing values. This was traditionally done with ad-hoc code but later customized pipelines, workflows and specialized software appeared for this purpose. This made it easy to benchmark and cluster things in an organized manner. In this review, we have assembled a catalog of available RNA-Seq single cell imputation algorithms/workflows and associated softwares for the scientific community performing single-cell RNA-Seq data analysis. Continued development of imputation methods, especially using deep learning approaches, would be necessary for eradicating associated pitfalls and addressing challenges associated with future large scale and heterogeneous datasets.

Whole-genome sequencing of extensively drug-resistant Salmonella enterica serovar Typhi clinical isolates from the Peshawar region of Pakistan.

BACKGROUND: Typhoid fever, caused by Salmonella enterica serovar Typhi, is a significant public health concern due to the escalating of antimicrobial resistance (AMR), with limited treatment options for extensively drug-resistant (XDR) S. Typhi strains pose a serious threat to disease management and control. This study aimed to investigate the genomic characteristics, epidemiology and AMR genes of XDR S. Typhi strains from typhoid fever patients in Pakistan. METHODOLOGY: We assessed 200 patients with enteric fever symptoms, confirming 65 S. Typhi cases through culturing and biochemical tests. Subsequent antimicrobial susceptibility testing revealed 40 cases of extensively drug-resistant (XDR) and 25 cases of multi-drug resistance (MDR). Thirteen XDR strains were selected for whole-genome sequencing, to analyze their sequence type, phylogenetics, resistance genes, pathogenicity islands, and plasmid sequences using variety of data analysis resources. Pangenome analysis was conducted for 140 XDR strains, including thirteen in-house and 127 strains reported from other regions of Pakistan, to assess their genetic diversity and functional annotation. RESULTS: MLST analysis classified all isolates as sequence type 1 (ST-1) with 4.3.1.1. P1 genotype characterization. Prophage and Salmonella Pathogenicity Island (SPI) analysis identified intact prophages and eight SPIs involved in Salmonella's invasion and replication within host cells. Genome data analysis revealed numerous AMR genes including dfrA7, sul1, qnrS1, TEM-1, Cat1, and CTX-M-15, and SNPs associated with antibiotics resistance. IncY, IncQ1, pMAC, and pAbTS2 plasmids, conferring antimicrobial resistance, were detected in a few XDR S. Typhi strains. Phylogenetic analysis inferred a close epidemiological linkage among XDR strains from different regions of Pakistan. Pangenome was noted closed among these strains and functional annotation highlighted genes related to metabolism and pathogenesis. CONCLUSION: This study revealed a uniform genotypic background among XDR S. Typhi strains in Pakistan, signifying a persistence transmission of a single, highly antibiotic-resistant clone. The closed pan-genome observed underscores limited genetic diversity and highlights the importance of genomic surveillance for combating drug-resistant typhoid infections.

Identification of Sulfamoylbenzamide derivatives as selective Cathepsin D inhibitors.

Aspartic proteases play very important role in post translational processing of proteins and several of them are essential for organism's viability. Here we present the enzyme inhibition activities of different Sulfamoylbenzamide derivatives against two aspartic proteases cathepsin D and plasmepsin II. Cathepsin D is an aspartic protease that degrades proteins at acidic pH in the lysosomes, or extracellular matrix. It is overexpressed by epithelial breast cancer cells and hence hyper-secreted. On the other hand plasmepsin II is an essential enzyme of Plasmodium falciperum. Cathepsin D and Plasmepsin II are pivotal drug targets for treatment of breast cancer and malaria respectively. Virtual screening of Sulfamoylbenzamide compounds followed by enzyme inhibition assays revealed these compounds as selective Cathepsin D inhibitors while inactive against Plasmepsin-II. IC50 values of five Sulfamoylbenzamide compounds tested are in range of 1.25-2.0 µM. N-(3-chlorophenyl)-2-sulfamoylbenzamide is identified as the most potent of all tested Sulfamoylbenzamide compounds with IC50 1.25 µM. It was also noted that the docking score of theses compounds was better in case of Cathepsin D as compared to Plasmepsin-II. Docking score ranges from -29.9±1.16 to -35.1±0.13 in case of Cathepsin D, while from -24.0±0.10 to -29.5±0.10 in case of Plasmepsin-II.

Association of Mitochondrial HVS-I Region Variants with Type 2 Diabetes in Pakistani Diabetic Subjects.

OBJECTIVE: To analyse mitochondrial hypervariable segment I (HVS-I) variations in Pakistani type 2 diabetic subjects. STUDY DESIGN: Case-control study. Place and Duration of the Study: National Institute of Diabetes and Endocrinology, Dow University of Health Sciences, Karachi, Pakistan, between January 2019 to January 2021. METHODOLOGY: DNA from whole blood was isolated, and mitochondrial HVS-I region (16024-16370) of 92 individuals, including 47 controls and 45 diabetics, was amplified, sequenced, and analysed. RESULTS: Ninety-two variable sites in the sequenced region were identified and individuals were classified into 56 different haplotypes according to phylotree 17.0 classifications, where major haplotype M5 was nearly 2-fold higher in diabetes. Fischer's exact test revealed variant 16189T>C significantly associated with diabetes (Odds ratio = 12.9, 95% CI = 0.6917 - 2400248) as compared to controls. The authors further analysed 1000 Genomes Project data of Pakistani Control subjects (i.e. PJL, n=96) and found that besides 16189T>C (Odds ratio = 5.875, 95% CI = 1.093 - 31.57, p <0.0339), 16264C>T (Odds ratio = 16, 95% CI = 0.8026 - 314.7, p <0.0310) also showed significant association with diabetic subjects. Comparing diabetic subject data with global control population data of the 1000 Genomes Project, significant associations of eight variants in the studied region were found. CONCLUSION: Based on the results of this case-control study, it can be concluded that specific variations in the mitochondrial hypervariable segment I (HVS-I) region are significantly associated with type 2 diabetes in the Pakistani population. The major haplotype M5 was found to be higher in diabetic subjects and variants 16189T>C and 16264C>T were significantly associated with diabetes. These findings suggest that mitochondrial DNA variations may play a role in the development of type 2 diabetes in the Pakistani population. KEY WORDS: Diabetes Mellitus, HVS-1 region, Diabetic subjects, Mitochondrial genomics, Pakistani population.

Study of drug resistance-associated genetic mutations, and phylo-genetic analysis of HCV in the Province of Sindh, Pakistan.

Current management of HCV infection is based on Direct-Acting Antiviral Drugs (DAAs). However, resistance-associated mutations, especially in the NS3 and NS5B regions are gradually decreasing the efficacy of DAAs. The aim of the current study was to identify such mutations in the NS3, and NS5B genes in DAAs treatment-naïve Pakistani chronic HCV 3a patients. Peripheral blood samples were collected from 233 chronic HCV 3a patients at different tertiary care hospitals in Karachi, Pakistan, between August 2020 to September 2021. PCR-amplified target regions of the NS3/NS5B gene were subjected to Sanger sequencing to identify resistance-associated mutations. Phylogenetic analysis of the identified amino acid sequences was performed using HCV3a sequences of the global population in the virus pathogen resource (VIPR) database. Sequence analysis identified five amino acid mutations, Leu36Pro, Gln41His, Gln80Lys/Arg, Ala156Tyr, and Gln168Arg in the NS3 region, and two mutations Leu159Phe and Cys316Arg in the NS5B region. Phylogenetic analysis revealed a high genetic diversity in the studied isolates. Overall, the prevalence of resistance-associated substitutions was almost similar to other geographic regions worldwide. This data could be helpful in selecting the most effective treatment regimen for HCV chronically infected people in Pakistan.

Genomic analysis of Chryseobacterium indologenes and conformational dynamics of the selected DD-peptidase.

Chrysobacterium indologenes is an emerging MDR pathogen that belongs to the family Flavobacteriaceae. The genome of the C. indologenes, isolated from the nephrotic patient, was sequenced through Illumina MiSeq. The pangenomics of available 56 C. indologenes strains using BPGA revealed an open pangenome (n=5553 CDS), core genome (2141), and accessory genome (2013). The CEG/DEG database identified 662 essential genes that drastically reduced to 68 genes after non-homology analyses towards human and gut microbiome. Further filtering the data for other drug target prioritizing parameters resulted in 32 putative targets. Keeping in view the crucial role played in cell wall biosynthesis, dacB was selected as the final target that encodes D-alanyl-d-alanine carboxypeptidase/endopeptidase (DD-peptidase). The 3D structure of dacB was modelled and rendered to docking analyses against two compound libraries of African plants (n=6842) and Tibetan medicines (n=52). The ADMET profiling exhibited the physicochemical properties of final compounds. The MD simulations showed the stability of inhibitor-DD-peptidase complex and interactions in terms of RMSD, RMSF, binding free energy calculation and H-bonding. We propose that the novel compounds Leptopene and ZINC95486338 from our findings might be potent DD-peptidase inhibitors that could aid in the development of new antibiotic-resistant therapy for the emerging MDR C. indologenes.

Genome sequencing and analysis of genomic diversity in the locally transmitted SARS-CoV-2 in Pakistan.

Surveillance of genetic diversity of the SARS-CoV-2 is extremely important to detect the emergence of more infectious and deadly strains of the virus. In this study, we evaluated mutational events in the SARS-CoV-2 genomes through whole genome sequencing. The samples were collected from COVID-19 patients in different major cities of Pakistan during the four waves of the pandemic (May 2020 to July 2021) and subjected to whole genome sequencing. Using in silico and machine learning tools, the viral mutational events were analyzed, and variants of concern and of interest were identified during each of the four waves. The overall mutation frequency (mutations per genome) increased during the course of the pandemic from 12.19 to 23.63, 31.03, and 41.22 in the first, second, third, and fourth waves, respectively. We determined that the viral strains rose to higher frequencies in local transmission. The first wave had three most common strains B.1.36, B.1.160, and B.1.255, the second wave comprised B.1.36 and B.1.247 strains, the third wave had B.1.1.7 (Alpha variant) and B.1.36 strains, and the fourth waves comprised B.1.617.2 (Delta). Intriguingly, the B.1.36 variants were found in all the waves of the infection indicating their survival fitness. Through phylogenetic analysis, the probable routes of transmission of various strains in the country were determined. Collectively, our study provided an insight into the evolution of SARS-CoV-2 lineages in the spatiotemporal local transmission during different waves of the pandemic, which aided the state institutions in implementing adequate preventive measures.

Peelu (Salvadora oleoides Decne.): An Unexplored Medicinal Fruit with Minerals, Antioxidants, and Phytochemicals.

The Peelu (Salvadora oleoides Decne.) fruit is well known for its nutritional and medicinal values. The current study analyzed the chemical composition of Salvadora oleoides fruit. Fresh Peelu fruits were harvested, and physicochemical properties, proximate composition, macro- and micronutrients, and phytochemical properties were determined. Moreover, ethanol and methanol fruit extract was analyzed for physicochemical properties. The Peelu fruit seemed to be a potential source of essential macro- ((nitrogen (N), phosphorus (P), potassium (K), calcium (Ca), and magnesium (Mg)) and micronutrients (zinc (Zn), manganese (Mn), iron (Fe), and copper (Cu)). The fruit had significant biochemical properties (total soluble solids (TSS), total acidity (TA), and TSS : TA ratio) with appreciable moisture, crude fiber, and ash contents. The fruit extracts demonstrated significantly higher antioxidants and phenolics, ascorbic acid contents, and carotenoids. Phytochemical screening of fruit revealed the presence of coumarins, flavonoids, phlobatannins, tannins, and terpenoids. Physicochemical and sensory evaluation of extracts indicated its potential for further in vivo study trials. The Peelu fruit was found to be a good source of mineral nutrients, proximate contents, vitamins (ascorbic acid and carotenoid), phytochemicals (total phenolic sand antioxidant contents), and pharmaceutically important metabolites that can be used as functional drink.

Association between Dietary Habits and Type 2 Diabetes Mellitus in Yangon, Myanmar: A Case-Control Study.

In Myanmar, the escalating prevalence of type 2 diabetes mellitus (T2DM) and impaired glucose tolerance among adults was recently reported, with the highest prevalence in the Yangon Region. The aim of the present study was to identify the risk factors in dietary habits and their relationship with T2DM in urban Myanmar residents. We conducted a case-control study recruiting 300 individuals aged 25-74 years living in the Yangon Region, consisting of 150 newly diagnosed cases attending a diabetes clinic, and 150 controls, who were community residents and free of diabetes. The case group had a significantly higher consumption of noodles, fish, beans, fermented food and pickles, dried food, topping seasonings, and non-dairy milk products than the control group, whereas they had a lower vegetable intake (more than three servings/day) and fruit intake (more than three servings/day) than the control group. Furthermore, the case group exhibited a higher frequency of some dietary behaviors than the control group, such as (1) having meals with family, (2) skipping breakfast, and (3) eating out. The final model showed that topping seasonings (adjusted odds ratio (aOR) 11.23, 95% confidence interval (CI) 3.08-40.90), more than three servings/day of vegetable intake (aOR 0.18, 95% CI 0.05-0.67), and having meals with family (aOR 2.23, 95% CI 1.05-4.71) were associated with diabetes. The study suggests that Myanmar's characteristic dietary culture of topping their meals with salty seasonings and sauces and eating multiple dishes together as a family are risk factors associated with T2DM. Our findings may contribute recommendations and opportunities for the primary prevention of T2DM in urban Myanmar.

Metaproteomics reveals the structural and functional diversity of Dermatocarpon miniatum (L.) W. Mann. Microbiota.

Metaproteomics is a strategy to understand the taxonomy, functionality and metabolic pathways of the microbial communities. The relationship among the symbiotic microbiota in the entire lichen thallus, Dermatocarpon miniatum, was evaluated using the metaproteomic approach. Proteomic profiling using one-dimensional SDS-PAGE followed by LC-MS/MS analysis resulted in a total of 138 identified proteins via Mascot search against UniRef100 and Swiss-Prot databases. In addition to the fungal and algal partners, D. miniatum proteome encompasses proteins from prokaryotes, which is a multifarious community mainly dominated by cyanobacteria and proteobacteria. While proteins assigned to fungus were the most abundant (55 %), followed by protists (16 %), bacterial (13 %), plant (11 %), and viral (1 %) origin, whereas 4 % remained undefined. Various proteins were assigned to the different lichen symbionts by using Gene Ontology (GO) terms, e.g. fungal proteins involved in the oxidation-reduction process, protein folding and glycolytic process, while protists and bacterial proteins were involved in photosynthetic electron transport in photosystem II (PS II), ATP synthesis coupled proton transport, and carbon fixation. The presence of bacterial communities extended the traditional concept of fungal-algal lichen symbiotic interaction.

Novel Genetic Variations in Acute Myeloid Leukemia in Pakistani Population.

Acute myeloid leukemia (AML) is a hematological malignancy characterized by clonal expansion of blast cells that exhibit great genetic heterogeneity. In this study, we describe the mutational landscape and its clinico-pathological significance in 26 myeloid neoplasm patients from a South Asian population (Pakistan) by using ultra-deep targeted next-generation DNA sequencing of 54 genes (∼5000×) and its subsequent bioinformatics analysis. The data analysis indicated novel non-silent somatic mutational events previously not reported in AML, including nine non-synonymous and one stop-gain mutations. Notably, two recurrent somatic non-synonymous mutations, i.e., STAG2 (causing p.L526F) and BCORL1 (p.A400V), were observed in three unrelated cases each. The BCOR was found to have three independent non-synonymous somatic mutations in three cases. Further, the SRSF2 with a protein truncating somatic mutation (p.Q88X) was observed for the first time in AML in this study. The prioritization of germline mutations with ClinVar, SIFT, Polyphen2, and Combined Annotation Dependent Depletion (CADD) highlighted 18 predicted deleterious/pathogenic mutations, including two recurrent deleterious mutations, i.e., a novel heterozygous non-synonymous SNV in GATA2 (p.T358P) and a frameshift insertion in NPM1 (p.L258fs), found in two unrelated cases each. The WT1 was observed with three independent potential detrimental germline mutations in three different cases. Collectively, non-silent somatic and/or germline mutations were observed in 23 (88.46%) of the cases (0.92 mutation per case). Furthermore, the pharmGKB database exploration showed a missense SNV rs1042522 in TP53, exhibiting decreased response to anti-cancer drugs, in 19 (73%) of the cases. This genomic profiling of AML provides deep insight into the disease pathophysiology. Identification of pharmacogenomics markers will help to adopt personalized approach for the management of AML patients in Pakistan.

Adaptation mechanism of mango fruit (Mangifera indica L. cv. Chaunsa White) to heat suggest modulation in several metabolic pathways.

Climate change is becoming a global problem because of its harmful effects on crop productivity. In this regard, it is crucial to carry out studies to determine crops' response to heatwave stress. Response molecular mechanisms during the development and ripening of mango fruit (Mangifera indica L. cv. Chaunsa White) under extreme heatwaves were studied. Mango flowers were tagged and fruits 18, 34, 62, 79, 92 days after flowering (DAF) as well as fruits on 10 and 15 days of postharvest shelf life were studied through RNA-Seq and metabolome of the fruit mesocarp. The environmental temperature was recorded during the experiment. Roughly, 2 000 000 clean reads were generated and assembled into 12 876 redundant transcripts and 2674 non-redundant transcripts. The expression of genes playing a role in oxidative stress, circadian rhythm, senescence, glycolysis, secondary metabolite biosynthesis, flavonoid biosynthesis and monoterpenoid biosynthesis was quantified as well as reactive oxygen species. Higher expressions of six abiotic stress genes and a senescent associated gene was found at 79 DAF (recorded temperature 44 °C). Higher expressions of nucleoredoxin and glutathione S-transferase 1 family protein were also recorded. Activation of the GABA-shunt pathway was detected by the glutamate decarboxylase transcript expression at 79 DAF. Larger energy demands at the beginning of fruit ripening were indicated by an increase in fructose-bisphosphate aldolase gene expression. Finally, the radical-scavenging effect of mango fruit inflorescence and fruit pulp extracts showed decline upon heatwave exposure. We recorded a broad genetic response of mango fruit suggesting the activation of several metabolic pathways which indicated the occurrence of genetic and metabolic crosstalks in response to intense heatwaves. Collectively, this study presents experimental evidence to help in the elucidation of the molecular mechanism of crops response to heat stress which in turn will help in the designing of protocols to increase crop productivity in the face of climate change.

Metagenomic analysis of drinking water samples collected from treatment plants of Hyderabad City and Mehran University Employees Cooperative Housing Society.

The quality assessment of water, supplied to the end user, is an essential part to assess the physical, chemical, and biological status of water, which impacts on human health. For the quality assessment of drinking water treatment plants and distribution systems of Hyderabad City and Mehran University of Engineering and Technology, Jamshoro, Pakistan, 13 surface drinking water samples were collected from three treatment plants, two of Hyderabad City, including WASA treatment plant and its distribution system (n = 5), Hala Nakka treatment plant and its distribution system (n = 6), and Mehran University Employees Cooperative Housing Society (MUECHS) treatment plant and its distribution system (n = 2). Physicochemical parameters of all drinking water samples were in the range compared to EPA and WHO guidelines, except in L-12 sample. Notably, no free-chlorine was detected in all samples. In metagenomics analysis, targeting V3-V4 hypervariable region of 16S rRNA gene, in QIIME2 environment, high bacterial prevalence was observed in all samples. On average, 348 OTUs were observed per sample. Among all samples, treated water sample from the Hala Nakka Treatment Plant (HNTR) was the most diverse sample in bacterial composition (Shannon 7.51 and Simpsons reciprocal indices 0.98). Overall, Proteobacteria, Bacteroidetes, Cyanobacteria, Verrucomicrobia, and Actinobacteria were the five most abundant phyla (relative abundances of 43.6, 37.9, 8.5, 2.5, and 2.4 percent, respectively). Notably, Cyanobacteria are well-known toxin producers which effect the human, and animal health. At genus level, Flavobacterium (4.86%) and Aquirestis (3.77%) were the most abundant genera. Functional predictions, based on 16S rRNA gene by PICRUSt, predicted 6909 KEGG orthologies, relating to 245 KEGG pathways. Among the predicted pathways of KEGG orthologies, pathways to human infections were also found. In conclusion, this study gave a deep insight into bacterial contamination in drinking water samples of Hyderabad City and MUECHS treatment plants and water quality status in Hyderabad and Mehran University of Engineering and Technology.

Thiourea derivatives induce fetal hemoglobin production in-vitro: A new class of potential therapeutic agents for ß-thalassemia.

Fetal hemoglobin (HbF) induction is a cost-effective therapeutic approach for the treatment of ß-hemoglobinopathies like ß-thalassemia and sickle cell anemia. The present study discusses the potential of thiourea derivatives as new class of compounds that induce the fetal hemoglobin production. HbF inducing effect of thiourea derivatives was studied using experimental cell system, the human erythroleukemic K562 cell line. Erythroid induction of K562 cells was studied by the benzidine/H2O2 reaction, total hemoglobin production was estimated by plasma hemoglobin assay kit, and γ-globin gene expression by RT-qPCR, whereas fetal hemoglobin production was estimated by flow cytometry and immunofluorescence microscopy. The results indicated that newly synthesized thiourea derivative are potent inducers of erythroid differentiation of K562 cells with an increased γ-globin gene expression and fetal hemoglobin production. Moreover, these compounds showed no cytotoxic effect and inhibition on K562 cells at HbF inducing concentrations. It is important to note that hydroxyurea is a cytotoxic chemotherapeutic agent and have deleterious side effects, reflecting the need to identify new safe and effective HbF induces. These results signify thiourea derivatives as promising HbF inducers, with the potential to be studied against hematological disorders, including ß-thalassemia and sickle cell anemia.

Estimating the mutational load for cardiovascular diseases in Pakistani population.

The deleterious genetic variants contributing to certain diseases may differ in terms of number and allele frequency from population to population depending on their evolutionary background. Here, we prioritize the deleterious variants from Pakistani population in manually curated gene list already reported to be associated with common, Mendelian, and congenital cardiovascular diseases (CVDs) using the genome/exome sequencing data of Pakistani individuals publically available in 1000 Genomes Project (PJL), and Exome Aggregation Consortium (ExAC) South Asia. By applying a set of tools such as Combined Annotation Dependent Depletion (CADD), ANNOVAR, and Variant Effect Predictor (VEP), we highlighted 561 potentially detrimental variants from PJL data, and 7374 variants from ExAC South Asian data. Likewise, filtration from ClinVar for CVDs revealed 03 pathogenic and 02 likely pathogenic variants from PJL and 112 pathogenic and 42 likely pathogenic variants from ExAC South Asians. The comparison of derived allele frequencies (DAF) revealed many of these prioritized variants having two fold and higher DAF in Pakistani individuals than in other populations. The highest number of deleterious variants contributing to common CVDs in descending order includes hypertension, atherosclerosis, heart failure, aneurysm, and coronary heart disease, and for Mendelian and congenital CVDs cardiomyopathies, cardiac arrhythmias, and atrioventricular septal defects.