Background and aims Familial hypercholesterolemia (FH) is a genetic disorder characterized by elevated low-density lipoprotein cholesterol (LDL-C) levels from birth, significantly increasing the risk of premature cardiac events and mortality. In Pakistan, despite the potential burden of FH, comprehensive studies evaluating its genetic characteristics, cascade screening significance, and lipoprotein (a) [Lp(a)] levels remain scarce. Understanding these factors is crucial for effective diagnosis, risk assessment, and management of FH in the Pakistani population. Methods After the identification of index case with clinical homozygous FH, characterized by high LDL-C and high Lp(a) levels together with a positive personal and family history of cardiovascular disease, a cascade screening of 66 relatives from a consanguineous family was performed. Blood samples were obtained from all subjects for biochemical and genetic analysis. Simon Broome criteria was applied on children for clinical FH diagnosis. Dutch Lipid Clinic Network scores were calculated for individuals aged ≥16years. Genetic screening was performed using next-generation sequencing to analyse all coding regions and exon-intron borders of the following genes: ALMS1, APOA1, APOB, APOA5, APOC2, APOC3, APOE, ABCA1, ABCG5, ABCG8, CREB3L3, GPIHBP1, LDLR, LDLRAP1, LIPA, LMF1, LPL, and PCSK9. The identified variants were confirmed using Sanger sequencing. Results Cascade screening identified seven homozygous and 25 heterozygous FH patients with pathogenic variant in the LDLR gene (NM_000527.5: c.2416dupG: p. Val806GlyfsTer11). Additionally, heterozygous variants of uncertain significance were identified in 4 other subjects. Conclusion This study underscores the high effectiveness of cascade screening in consanguineous families and societies that could lead to early detection and prevention.

Dyslipidaemia is a significant risk factor for cardiovascular diseases, which can manifest early in life. Despite its importance, the prevalence of dyslipidaemia in the paediatric population of Pakistan remains poorly understood. This study uses laboratory data to determine the prevalence of dyslipidaemia and lipid testing practices among Pakistani children and adolescents. This retrospective cohort study analysed the laboratory data from children and adolescents, aged up to 19 years, who underwent lipid testing. The data was obtained from two centres with collection points all over Pakistan for five years (March 2019–March 2024). Logistic regression models were used to assess relationships between demographic factors (age, sex and regions/provinces) and lipid profile parameters. Over five years, 9,787 children and adolescents with a mean age of 13.8 ± 5.1 years underwent lipid testing. Boys accounted for 59.7

The most prevalent type of acquired depigmentation disorder is vitiligo, which is characterized by the development of white spots on the skin, hair, and mucous membranes due to melanocyte loss. It is a polygenic and multifactorial disease with complicated genetic and environmental pathogenesis. The onset and progression of vitiligo are strongly influenced by both genetic and environmental variables, including neurological effects, oxidative-antioxidative stress, and immune dysfunction. The polymorphism (rs2670660) in the NLRP1 gene was analyzed in 110 patients with vitiligo and 110 healthy controls matched for age and sex. Tetra-ARMS PCR was used to determine the genotypes, and Sanger sequencing was employed to validate the three genotypes. Initial genotype analysis revealed significant differences in genotype distribution between cases and control (p = 0.035). Further analysis of the genotype data showed that the frequency of the GG genotypes was significantly higher in vitiligo cases than in controls under the recessive model (odds ratio [OR] 2.647; 95

BACKGROUND:Severe hypertriglyceridemia (HTG) is associated with increased risk of cardiovascular disease and acute pancreatitis. Previously, there were no studies or data available from Pakistan regarding this condition. This study aimed to analyze the trends and prevalence of severe HTG in Pakistan over a 5-year period from 2 healthcare centers. METHODS:A retrospective analysis of laboratory data from 2 major healthcare centers across Pakistan's 4 provinces was conducted. Demographic information and lipid profiles of patients were collected. HTG was categorized as HTG (≥150 mg/dL), mild HTG (150-199), moderate HTG (200-999), severe HTG (1000-1999 mg/dL), and very severe HTG (≥2000 mg/dL). RESULTS:A study of 552,719 individuals (mean age 45.9 ± 12.6 years, 58.1% males and 41.9% females) revealed that 53.7% (1:2) were hypertriglyceridemic. Severe HTG was observed in 0.3% (1:286) of the population, while very severe HTG was found in 0.1% (1:861). Both severe and very severe HTG were more prevalent in males, individuals aged under 18 years, and those aged 36 to 45 years. Very severe HTG was most prevalent in Khyber Pakhtunkhwa (0.2%). Patients with severe HTG presented with mean triglyceride levels of 1308.2 ± 254.7 mg/dL, while those with very severe HTG showed levels of 3293.0 ± 1889.9 mg/dL. CONCLUSION:This study revealed that 53.7% of the population had HTG, with severe and very severe HTG affecting 0.3% and 0.1%, respectively. Males, younger individuals, and residents of Khyber Pakhtunkhwa had higher prevalence. These findings emphasize the need for targeted interventions in high-risk groups.

IntroductionHypertriglyceridemia (HTG) is a complex disorder caused by genetic and environmental factors that frequently results from loss-of-function variants in the gene encoding lipoprotein lipase (LPL). Heterozygous patients have a range of symptoms, while homozygous LPL deficiency presents with severe symptoms including acute pancreatitis, xanthomas, and lipemia retinalis.MethodsWe described the clinical characteristics of three Slovenian patients (an 8-year-old female, an 18-year-old man, and a 57-year-old female) and one Pakistani patient (a 59-year-old male) with LPL deficiency. We performed next-generation sequencing (NGS) targeting all coding exons and intron-exon boundaries of the LPL gene, and Sanger sequencing for variant confirmation. In addition, we performed a systematic literature review of all cases with three identified variants and described their clinical characteristics.ResultsTwo Slovenian patients with a heterozygous pathogenic variant NM_000237.3:c.984G>T (p.Met328Ile) were diagnosed within the first three years of life and had triglyceride (TG) values of 16 and 20 mmol/L. An asymptomatic Pakistani patient with TG values of 36.8 mmol/L until the age of 44 years, was identified as heterozygous for a pathogenic variant NM_000237.3:c.724G>A (p.Asp242Asn). His TG levels dropped to 12.7 mmol/L on dietary modifications and by using fibrates. A Slovenian patient who first suffered from pancreatitis at the age of 18 years with a TG value of 34 mmol/L was found to be homozygous for NM_000237.3:c.337T>C (p.Trp113Arg).ConclusionsPatients with LPL deficiency had high TG levels at diagnosis. Homozygous patients had worse outcomes. Good diet and medication compliance can reduce severity.

Background: Hereditary sensory and autonomic neuropathies (HSANs) are rare heterogeneous group of neurological disorders caused by peripheral nerve deterioration. The HSANs sub-clinical classes have clinical and genetic overlap which often lead to misdiagnosis. In the present study a Pakistani family with five affected members suffering from severe neuropathy were genetically analyzed to identify the disease causative element in the family. Methods: Genome wide high-density single nucleotide polymorphism (SNP) microarray analysis was carried out followed by whole exome sequencing of the affected proband and another affected sibling. Shared homozygous regions in all severely affected members were identified through homozygosity mapping approach. Results: The largest homozygous region of 14.1 Mb shared by the five severely affected members of the family was identified on chromosome 2. Subsequent exome sequencing identified a novel single nucleotide deletion c.2658del; p.(Ser887Profs*64) in KIF1A. Segregation analysis revealed that this mutation was homozygous in all five affected individuals of the family with severe clinical manifestation, while members of the family that were heterozygous carriers shared abnormal skin features (scaly skin) only with the homozygous affected members. Conclusions: A novel frameshift mutation p.(Ser887Profs*64) in KIF1A is the potential cause of severe HSANIIC in a Pakistani family along with incomplete penetrance in mutation carriers. We demonstrate that using a combination of different techniques not only strengthens the gene finding approach but also helps in proper sub-clinical characterization along with identification of mutated alleles exhibiting incomplete penetrance leading to intrafamilial clinical variability in HSAN group of inherited diseases.

Dopamine β-hydroxylase (DBH) is a copper-containing enzyme that has an important role in maintaining the cellular homeostasis between the two neurotransmitters, dopamine (DA) and nor-adrenaline (NA). DBH functional polymorphisms are associated with multiple neuro-psychiatric conditions and are found to alter the DBH protein levels in serum, affecting DBH enzymatic activity. The current study was conducted to determine the genetic association of DBH functional polymorphism rs1611115 and its effect on DBH levels in serum in major depressive disorder (MDD), bipolar disorder (BPD) and schizophrenia (SHZ) in the Pakistani population. In total n = 1097 subjects including MDD (n = 427), BPD (n = 204), SHZ (n = 134) and healthy controls (n = 332), were screened for rs1611115 by polymerase chain reaction-restriction fragment length polymorphism. Univariate logistic regression was applied and results were adjusted for age and sex (multivariant analysis). The DBH levels in serum were determined through enzyme-linked immunosorbent assay (ELISA) and the Mann Whitney U test was applied. The results showed a significant association of minor allele (-1021C > T) with a higher risk of developing BPD and SHZ in both univariable and multivariable analyses. Moreover, the overall total serum concentration of DBH was comparatively raised in MDD, however, in cross-comparison DBH serum levels were found markedly higher in CC homozygotes compared to TT homozygotes within the BPD group. Thus the present study suggested a significant association of DBH rs1611115 with BPD and SHZ and also the effect of rs1611115 on DBH serum levels in MDD and BPD, for the first time in the Pakistani population.

The retrospective study was conducted at the Faisalabad Institute of Cardiology from January 2022 to January 2023 to evaluate the association between apolipoprotein A5 single nucleotide polymorphisms (APOA5 SNPS) and hypertriglyceridemia in the Pakistani population and assess the efficacy of fibrate therapy in hypertriglyceridemic patients. Blood samples of patients were collected for DNA extraction by the standard inorganic method. Extracted DNA was analyzed biochemically and genetically following the selection and genotyping of the APOA5 gene to know SNP variants with risk alleles. We analyzed lipid profiles, including TG (triglyceride), HDL (high-density lipoprotein), and LDL (low-density lipoprotein) levels of hypertriglyceridemic patients, then compared them to normal and fibrate-treated individuals. We compared APOA5 (rs662799) SNP (single nucleotide polymorphism) among 50 hypertriglyceridemic, 50 healthy controls, and among the same 50 hypertriglyceridemic patients who were given fibrate. Results showed that fibrate decreased TG level and LDL by about 30-45% and 20-25%, respectively; increased high-density lipoprotein by 10-15% and total cholesterol decreased by 6-8% in Pakistani. Hypertriglyceridemia risk was significantly increased by a minor allele of APOA5 rs662799 polymorphism. Minor allele carriers of rs662799 had an odd ratio of (95% CI) 1.5 (1.03-2.18) (P = 0.032). Risk allele frequency differed among hypertriglyceridemic patients, healthy controls, and fibrate-treated hypertriglyceridemic individuals. We analyzed that 87.5% of healthy controls exhibited no risk allele, while 78% did not show this risk allele among hypertriglyceridemic patients. It was concluded that APOA5 rs662799 polymorphism is a genetic determinant of hypertriglyceridemia. Fibrate caused a reduction in TG and LDL and caused an elevation of HDL-C among hypertriglyceridemic patients.

The peptidylarginine-deiminase 4 (PADI4) is involved in the post-translational catalytic conversion of arginine into citrulline. The autoantibodies including anti-citrullinated protein antibodies (ACPAs) produced in response to hypercitrullinated proteins are a hallmark of rheumatoid arthritis (RA) autoimmunity. Therefore, the role of a missense variant rs874881 (Gly112Ala) of PADI4 in RA susceptibility was analyzed, along with in-silico analysis of structural and functional impacts of this substitution.We did a case-control association study and in-silico analysis. For the case-control study, confirmed RA cases and healthy controls were recruited. Genotyping for rs874881 (n = 750) was performed through polymerase chain reaction-restriction fragment length polymorphism. Multivariate logistic regression analysis was employed to determine association. The in-silico analysis was carried out through HOPE, VarMap, MutationAssessor, MutPred2, SIFT, PolyPhen, CADD, REVEL and MetaLR.In the case-control study, the rs874881 exhibited a strong association with increased RA susceptibility (G vs C odds ratio = 3.85, 95 % confidence interval = 2.81-5.27). Interaction analysis revealed significant interaction of genotype with smoking and gender (p < 0.05). Significant results (p < 0.05) were also obtained in stratified analysis by presence/absence of comorbidities and radiographic damage. According to in-silico pathogenicity prediction analysis, this Gly112Ala substitution does not exert a major effect on protein structure and function including its enzymatic activity.We report a significant association of PADI4 rs874881 with overall RA susceptibility. To our knowledge, this is the first study to do the interaction and stratified analyses on the PADI4 rs874881 in RA. Similar detailed studies should also be performed in other populations.

Background and Objectives: Hereditary multiple exostoses (HME) is a disease characterized by cartilage-capped bony protuberances at the site of growth plates of long bones. Functional mutations in the exostosin genes (EXT1 and EXT2) are reported to affect the hedgehog signalling pathways leading to multiple enchondromatosis. However, the exact role of each EXT protein in the regulation of heparan sulphate (HS) chain elongation is still an enigma. In this study, a Pakistani family with HME is investigated to find out the genetic basis of the disease. Materials and Methods: Genotyping of eight members of the family by amplifying microsatellite markers, tightly linked to the EXT1 and EXT2 genes. Results: The study revealed linkage of the HME family to the EXT1 locus 8q24.1. Sanger sequencing identified a heterozygous deletion (c.247Cdel) in exon 1 of EXT1, segregating with the disease phenotype in the family. In silico analysis predicted a shift in the frame causing an early stop codon (p.R83GfsX52). The predicted dwarf protein constituting 134 amino acids was functionally aberrant with a complete loss of the catalytic domain at the C-terminus. Interestingly, an alternative open reading frame 3 (ORF3) caused by the frame shift is predicted to encode a protein sequence, identical to the wild type and containing the catalytic domain, but lacking the first 100 amino acids of the wild-type EXT1 protein. Conclusion: Consequently, haploinsufficiency could be the cause of HME in the investigated family as the mutated copy of EXT1 is ineffective for EXT-1/2 complex formation. The predicted ORF3 protein could be of great significance in understanding several aspects of HME pathogenesis.

Spreading of multidrug-resistant bacteria has become a growing and significant threat to environmental and public health. Our understanding of environmental variables such as nutrients which contribute to the dissemination of antibiotic resistance and pathogenesis is still limited. In this study, we operated outdoor mesocosm experiments to evaluate how nitrate and phosphate ions with different concentrations influence on an adaptation of Enterococcus faecalis to aquatic environments. E. faecalis were isolated from the mesocosms at 8 sampling events for 96 days to evaluate biofilm production, quorum-sensing-related genes expression, and sensitivity to oxytetracycline, erythromycin, ciprofloxacin, ampicillin, vancomycin, and chloramphenicol. Quantitative real-time PCR was used to compare mRNA levels of E. faecalis quorum-sensing-related genes. E. faecalis isolates exhibited resistance to oxytetracycline, ampicillin, and ciprofloxacin, respectively. We observed that the biofilm production of E. faecalis isolates on day 60 and 96 was significantly increased (p < 0.01). The expressions of quorum-sensing-related genes were significantly up-regulated (∼tenfold) at the transcriptional level in nutrient-enriched treatments. Our findings indicate that nitrate and phosphate ions facilitate resistance to commonly used antibiotics, increase biofilm production and intra-species communication, which could be a major reviving strategy of E. faecalis in the aquatic environments.

Abstract The dopaminergic pathways control neural signals that modulate mood and behaviour along and have a vital role in the aetiology of major depression (MDD), schizophrenia (SHZ) and bipolar disorder (BD). Genome-wide association studies (GWAS) have reported several dopaminergic pathway’s and other genetic loci’s association with these disorders, therefore, the present study was conducted to analyse the GWAS and candidate gene loci of the dopaminergic and cognitive system genes in MDD, SHZ, and BD, in the Pakistani population. A total of 1237 subjects [MDD n = 479; BD n = 222; SHZ n = 146; and controls n = 390], were screened for eleven genetic variants through polymerase chain reaction (PCR) techniques. Univariant followed by multivariant logistic regression analysis were applied to determine the genetic association. Significant risk associations were observed for rs4532 and rs1799732 with MDD; and rs1006737 and rs2238056 with BD. However, after applying multiple test corrections rs4532 and rs1799732 association did not remain significant for MDD. Moreover, a protective association was found for three variants DRD4-120bp, rs10033951 and rs2388334 in the current cohort. Thus, in conclusion, the current study revealed the risk association of rs1006737 and rs2238056 with BD and the protective effect of DRD4-120bp in MDD and BD, of rs2388334 in BD and of rs10033951 in MDD, BD, and SHZ in the current Pakistani cohort.

The convolutional neural network showed considerable success in medical imaging with explainable AI for cancer detection and recognition. However, the irrelevant and large number of features increases the computational time and decreases the accuracy. This work proposes a deep learning and fuzzy entropy slime mould algorithm-based architecture for multiclass skin lesion classification. In the first step, we employed the data augmentation technique to increase the training data and further utilized it for training two fine-tuned deep learning models such as Inception-ResNetV2 and NasNet Mobile. Then, we used transfer learning on augmented datasets to train both models and obtained two feature vectors from newly fine-tuned models. Later, we applied a fuzzy entropy slime mould algorithm on both vectors to get optimal features that are finally fused using the Serial-Threshold fusion technique and classified using several machine learning classifiers. Eventually, the explainable AI technique named Gradcam opted for the visualization of the lesion region. The experimental process was conducted on two datasets, such as HAM10000 and ISIC 2018, and achieved 97.1 and 90.2% accuracy, better than the other techniques.

Osteopetrosis is a genetically heterogenous, fatal bone disorder characterized by increased bone density. Globally, various genetic causes are reported for osteopetrosis with all forms of inheritance patterns. A precise molecular diagnosis is necessary for prognosis and for prescribing treatment paradigms in osteopetrosis. Here we report on thirteen individuals diagnosed with infantile malignant osteopetrosis coming from ten unrelated Pakistani families; nine of whom are consanguineous. We performed whole exome sequencing and Sanger sequencing in all families and identified homozygous variants in genes previously reported for autosomal recessive inheritance of osteopetrosis. All the identified variants are expected to affect the stability or length of gene products except one nonsynonymous missense variant. TCIRG1 was found as a candidate causal gene in majority of the families. We report six novel variants; four in TCIRG1 and one each in CLCN7 and OSTM1. Our combined findings will be helpful in molecular diagnosis and genetic counselling of patients with osteopetrosis particularly in populations with high consanguinity.

The incidence of microvascular complications, including diabetic retinopathy (DR), increases with duration of type 2 diabetes (T2D). Meta-GWAS have reported numerous single-nucleotide polymorphisms (SNPs) associated with T2D; however, no loci, achieving genome-wide significance has been reported for DR. Vascular endothelial growth factor A (VEGFA) and insulin-like growth factor 1 (IGF1) are considered as potential genetic candidates involved in T2D and DR progression. Moreover, the association of serum levels of these proteins with diabetes-related traits is controversial. Therefore, the current study was designed to evaluate the possible genetic predisposition and role of these circulating growth factors in serum in the pathophysiology of T2D and DR. A cohort of 1126 individuals with T2D was collected including those without retinopathy (DNR = 573), non-progressive diabetic retinopathy (NPDR = 301) and progressive diabetic retinopathy (PDR = 252), and 348 healthy controls. Genomic DNA was isolated, and six SNPs: rs833061, rs13207351, rs1570360, rs2010963, rs5742632 and rs6214, were genotyped and results statistically analyzed. ELISA was performed on a subset of the samples to measure serum levels of IGF1 and VEGFA. The minor allele of rs6214 was associated with T2D [OR = 1.67 (95% CI  1.39–2.01, p = 4.9E−8)], rs13207351 was associated with NPDR [OR = 1.97 (95% CI  1.28–3.03, p = 9.0E−3)]when compared with DNR, and rs5742632 showed positive association with PDR [OR = 1.66 (95% CI  1.33–2.05, p = 1.0E−4)] compared to DNR. Lowered IGF1 serum levels were found to be associated with T2D, NPDR and PDR. IGF1 was found to increase the T2DM susceptibility as well as advanced DR, i.e., PDR, while VEGFA was found to be associated with early DR stage, i.e., NPDR.

Myocardial infarction (MI) is a major cause of morbidity and mortality in patients with coronary artery disease. Hypertension is independent risk factors for MI. Genetic studies have shown association of single nucleotide polymorphism (SNP) rs2272996 in Vanin1 ( VNN1 ) gene with hypertension and

Human activity recognition from the video is an important problem due to its potential applications in remote surveillance, content-based video retrieval, and in humanoid robots. Most of the visual activity recognition research in the last decade focus on recognizing basic human actions from well-constrained laboratory videos and depend upon fully annotated video dataset for model training. Whereas activity recognition from unconstrained videos is still very challenging due to large variations in object appearance and pose, occlusion, and inter and intraclass variations. It is an extremely laborious task to prepare large scale realistic video activity dataset with detailed annotations of the human, object, and their mutual interactions in each frame. Although it is intuitive to model contextual relationships form fully annotated dataset, however, it is unknown as to how reliably multilevel contextual features can be extracted in the absence of annotated dataset. To mitigate these challenges, we propose a weakly supervised approach for complex human activity recognition from realistic videos. The proposed approach requires only activity labels for each video to train the model. A novel multilevel contextual features and context estimation procedure from the un-annotated dataset is also introduced. Restricted Boltzman machine is used to systematically integrate multilevel contextual features. We evaluate the proposed approach on benchmark realistic surveillance video datasets for human-human and human-object interaction activity recognition. The experimental results show improved accuracies on benchmark datasets without using fully annotated datasets.

Objectives: This study was aimed at determining human T-lymphotropic virus 1/2 prevalence among apparently healthy, immunocompromised and haematologic malignant individuals in Nigeria’s capital, as well as meta-analysis of all Nigerian studies until date. Methods: A total of 200 participants were recruited into a cross-sectional study. In total, 1 mL each of sera and plasma were obtained from 5 mL blood of each participant and analysed for antibodies to human T-lymphotropic virus 1/2 using enzyme-linked immunosorbent assay; positive samples confirmed with qualitative real-time polymerase chain reaction, followed by statistical and meta-analysis. Sociodemographic characteristics and possible risk factors were assessed via questionnaires. Results: Enzyme-linked immunosorbent assay yielded 1% prevalence which was confirmed to be zero via polymerase chain reaction. A total of 119 (59.5%) of the participants were male, while the mean age was 35.28 ± 13.61 years. Apart from sex and blood reception/donation, there was generally a low rate of exposure to human T-lymphotropic virus–associated risk factors. Meta-analysis revealed pooled prevalence of human T-lymphotropic virus 1 and 2 to be 3% and 0%, respectively, from Nigerian studies. Conclusion: This study discovered zero prevalence of human T-lymphotropic virus 1/2 from five major hospitals in Nigeria’s capital, exposing the importance of confirmatory assays after positive antibody detection assay results. Meta-analysis highlighted the existence of very few reliable Nigerian studies compared to the demography of the nation. Large-scale epidemiological studies and routine screening of risk populations are therefore needed since Nigeria lies in the region of endemicity.