Predisposing deleterious variants in the cancer-associated human kinases in the global populations.

Human kinases play essential and diverse roles in the cellular activities of maintaining homeostasis and growth. Genetic mutations in the genes encoding the kinases (or phosphotransferases) have been linked with various types of cancers. In this study, we cataloged mutations in 500 kinases genes in >65,000 individuals of global populations from the Human Genetic Diversity Project (HGDP) and ExAC databases, and assessed their potentially deleterious impact by using the in silico tools SIFT, Polyphen2, and CADD. The analysis highlighted 35 deleterious non-synonymous SNVs in the ExAC and 5 SNVs in the HGDP project. Notably, a higher number of deleterious mutations was observed in the Non-Finnish Europeans (26 SNVs), followed by the Africans (14 SNVs), East Asians (13 SNVs), and South Asians (12 SNVs). The gene set enrichment analysis highlighted NTRK1 and FGFR3 being most significantly enriched among the kinases. The gene expression analysis revealed over-expression of NTRK1 in liver cancer, whereas, FGFR3 was found over-expressed in lung, breast, and liver cancers compared to their expression in the respective normal tissues. Also, 13 potential drugs were identified that target the NTRK1 protein, whereas 6 potential drugs for the FGFR3 target were identified. Taken together, the study provides a framework for exploring the predisposing germline mutations in kinases to suggest the underlying pathogenic mechanisms in cancers. The potential drugs are also suggested for personalized cancer management.

Increased antimicrobial resistance against azithromycin during COVID: role of irrational utilization.

Objectives: To assess the use of azithromycin during coronavirus disease-2019 pandemic and its impact on antimicrobial resistance in an urban setting. METHODS: The retrospective, cross-sectional study was conducted on two different data sets. The first data set was of inpatients (N = 300) during the first wave of COVID 19 i.e. January to December, 2020. Data was collected from tertiary care hospitals in Karachi between October 2021 and November 2022 after approval from the ethics review committee of Ziauddin University, Karachi. Drug utilisation evaluation was done using a structured and validated tool. The treatment protocols were evaluated by comparing against the coronavirus disease-2019 treatment protocol 2020 and the guidelines issued by the Medical Microbiology and Infectious Diseases Society of Pakistan. Second data set comprised of the consumption data (obtained from pharmacies for both inpatients and outpatients) as well as the antimicrobial resistance, (obtained from antibiogram collected from the microbiology departments of the participating hospitals). This data was taken for the period of three years i.e. 2019 (Pre-COVID) to 2021 (Post-first wave of COVID) to establish trends of both consumption and antibiotic resistance. Data was analysed using SPSS 20. RESULTS: Of the 300 patients, 207(69%) were males and 93(31%) were females. There were 162(54%) adults with mean age 40.06±10.48 years, followed by 120(40%) geriatrics with mean age 70.37±6.94 years, 18(6%) paediatrics with mean age 13.5±3.60 years. All patients were given Azithromycin empirically followed by culture sensitivity test in 21% cases only. Comparison with COVID treatment protocols revealed the non-compliance of just 3%. However, in case of Community Acquired Pneumonia (CAP), sinusitis, typhoid and urethritis, comparison with MMIDSP guidelines revealed non-compliance of 95%, 22%, 75% and 100% respectively. Moreover, in 11% of patients, it was administered for conditions not recommended by guidelines. Furthermore, the Antibiogram exhibited percent increase in resistance against azithromycin. CONCLUSIONS: Enhanced consumption and irrational use of azithromycin during the coronavirus disease-2019 pandemic most likely contributed to increase in antimicrobial resistance.

Role of Alanine Transaminase and Transient Elastography in Categorising Nonalcoholic Fatty Liver Disease Subgroups.

OBJECTIVE: To investigate the association of alanine transaminase (ALT) with transient elastography grades to define various nonalcoholic fatty liver disease (NAFLD) groups for disease status. STUDY DESIGN: Cross-sectional, descriptive study. Place and Duration of the Study: Gastroenterology Outpatient Clinic, Ziauddin Hospital, from January to December 2022. METHODOLOGY: This study included 194 NAFLD patients. Demographic data, body mass index, enzymes, and transient elastography (TE) findings were recorded. NAFLD patients were categorised as nonalcoholic fatty liver (NAFL), nonalcoholic steatohepatitis (NASH), steatofibrosis (significant fibrosis F2-F3 with normal ALT), and cirrhosis using TE grades. RESULTS: The median age of the patients was 44 [IQR 18.25] years; 146 (75.3%) were males. Out of 194 NAFLD patients, 21 (10.8%) were NAFL, 116 (59.8%) were NASH, 14 (7.2%) showed steatofibrosis, and 43 (22.2%) were cirrhotic. On transient elastography, the majority were with S3 steatosis (n=107, 55.2%) and 59 (30.2%) had F0-F1 fibrosis. There was a statistically significant difference in the mean rank of age, ALT, AST, and GGT levels within 4 groups of NAFLD (p <0.001). Most of the patients with all the stages of fibrosis had increased ALT levels (p=0.034). CONCLUSION: This study concluded that a combination of ALT levels and transient elastography findings could be considered for differentiating uncomplicated steatosis from NASH, steatofibrosis, and cirrhosis, hence limiting the use of liver biopsy. This may prove a reliable way to measure the severity of the disease. KEY WORDS: Nonalcoholic fatty liver disease, Cirrhosis, Transient elastography.

Emerging novel sequence types of Staphylococcus aureus in Pakistan.

BACKGROUND: Despite an increasing incidence of Staphylococcus aureus infection and dissemination in Pakistan, the epidemiology of different Staphylococcus aureus research clones has been the subject of only a small number of investigations. By analyzing the collected data sequence, this study was designed to study the epidemiology of Staphylococcus aureus in the area using multilocus sequence typing (MLST). METHODS: A total of 1015 staphylococcus strains collected from the city's tertiary care facilities were biochemically screened, followed by antimicrobial susceptibility testing against a panel of 13 antibiotics. Analyzed methicillin-resistant Staphylococcus aureus (MRSA) was subjected to molecular characterization using multilocus sequence typing (MLST), clonal complex analysis, recombination testing, and phylogenetic analysis. RESULTS: Approximately 421 bacteria were verified as Staphylococcus aureus by biochemical analysis. 57% of the isolates exhibited multidrug resistance, of which 89% were found to be methicillin-resistant Staphylococcus aureus (MRSA). MLST results in a total of 39 sequence types (ST) and 5 clonal complexes (CC), out of which twenty-two STs were newly documented worldwide. The most common CC identified was CC8. The direct sequencing data also revealed significant shifts at MLST loci, with point mutations resulting in the aroE-343 and tpi-278 alleles. CONCLUSIONS: This study concludes that there is high diversity in the locally circulating clones of Staphylococcus aureus present in nature and that they are defined by their geographic epidemiology. These findings have practical implications for public health, including the need for tailored infection control strategies, antibiotic stewardship, global surveillance, and a deeper understanding of bacterial evolution.

Inflammatory markers and COVID-19 disease progression.

BACKGROUND: The COVID-19 pandemic has resulted in a global humanitarian crisis. Despite ongoing research, transmission risks and many disease characteristics remained unclear. Most patients have displayed elevated levels of certain inflammatory markers, which we sought to investigate further in relation to disease severity. The aim of this study was to examine the correlation between inflammatory markers and the severity of COVID-19 among patients. METHODS: We conducted a cross-sectional study from April to September 2020, involving 143 COVID-19 PCR-positive patients from Ziauddin Hospital. Electronic patient records provided data on demographics, clinical status, and laboratory results. RESULTS: The majority of PCR-positive patients were elderly males with comorbidities such as diabetes and hypertension. Almost all patients exhibited increased levels of various inflammatory markers, with procalcitonin (97.2%) being the most common. Statistically significant differences were observed in the levels of TLC (p = 0.005), CRP (p = 0.001), LDH (p = 0.001), Ferritin (p = 0.001), D-dimer (p = 0.001), and procalcitonin (p = 0.028), in relation to COVID-19 severity. CONCLUSIONS: The data suggest a significant association between levels of inflammatory markers and COVID-19 severity. All markers, except procalcitonin, demonstrated a significant correlation with disease severity. These results could enhance our understanding of COVID-19 pathogenesis and help predict and manage severe cases.

Angiotensin-converting enzyme 2 (ACE2) polymorphisms and susceptibility of severe SARS-CoV-2 in a subset of Pakistani population.

Science is digging for the varied presentation of COVID-19 patients exposed to the same risk factors, and medical conditions may be influenced by the presence of polymorphic genetic variants. This study investigated the link between ACE2 gene polymorphisms and the severity of SARS-CoV-2. This cross-sectional study recruited COVID-19 PCR-positive patients by consecutive sampling from Ziauddin Hospital from April to September 2020. DNA was extracted from whole blood, followed by gene amplification and Sanger's sequencing. Most of the patients, 77: 53.8%, were serious. Males were higher (80; 55.9%) with age more than 50 years (106: 74.1%). We found 22 ACE2 SNPs. rs2285666 SNP was most prevalent with 49.2% CC, 45.2% TT, 4.8% CT heterozygosity, and 0.8% AA genotypes. Variants with multiple genotypes were also insignificantly associated with the severity of COVID-19 in the analysis of the dominant model. Only rs2285666 had a significant statistical link with gender (p-value 0.034, OR; 1.438, CI; 1.028-2.011) while rs768883316 with age groups (p-value 0.026, OR; 1.953, CI; 1.085-3.514). Haplotypes ATC of three polymorphisms (rs560997634, rs201159862, and rs751170930) commonly found in 120 (69.77%) and TTTGTAGTTAGTA haplotype consisting of 13 polymorphisms (rs756737634, rs146991645, rs1601703288, rs1927830489, rs1927831624, rs764947941, rs752242172, rs73195521, rs781378335, rs756597390, rs780478736, rs148006212, rs768583671) in 112 (90.32%) had statistically significant association with the severity having p = value 0.029 and 0.001 respectively. Males of old age and diabetics are found to have more severe COVID-19 infection in the current study. We also found that common ACE2 polymorphism rs2285666 influences the susceptibility of acquiring the severe SARS-CoV-2 infection.

Analysis Of Common Somatic Mutations In Colorectal Carcinoma And Associated Dysregulated Pathwaysarts.

BACKGROUND: Identification of gene targets and biological pathways involved in colorectal carcinoma (CRC) is essential for better management of patients. Our study aims to highlight common somatic mutations in colorectal carcinoma and to identify dysregulated pathways and gene enrichment based on KRAS and BRAF interaction network analysis. METHODS: By using cancer browser tool in COSMIC database, mutation frequencies of the top 20 mutated genes listed for colorectal adenocarcinoma were identified. The most frequent variants of selected genes were explored with ClinVar database which led to identification of protein change along with its cytogenic location, variant type, variant length and the associated single nucleotide polymorphism (SNP). These identified SNPs were searched in Pakistani database using 1000genome in an attempt to identify common polymorphisms. Using the database ClinicalTrial.gov the number of clinical trials based upon these selected mutations was explored. Enrichment and protein interaction (PI) analysis of KRAS and BRAF was carried out to reveal significant biological pathways associated with these genes. RESULTS: In cumulative data, among all variants about 57% of substitution mutations are observed to be G>A including mutations in KRAS, Tp53, SMAD4, PI3K and NRAS. The mutations of KRAS (c.35G>A), TP53 (c.524G>A) and APC (c.4348C>T) were found to be pathogenic with single nucleotide variation and variant length of 1bp. Searching 1000genome database revealed that 100 % of alleles found in East Asian population studied are 'C'(frequency=1). Significant biological pathways (<0.05) identified by our search include Trk receptor signalling mediated by the MAPK pathway, signalling to p38 via RIT and RIN, signalling to ERKs, Frs2-mediated activation, ARMS-mediated activation and prolonged ERK activation events. CONCLUSIONS: Our study highlights the role of genetic profiling in CRC, with emphasis on mutations which may define treatment outcome. Targeting several collateral pathways simultaneously may be further explored to improve colorectal cancer therapeutics.

Naegleria fowleri from Pakistan Has Type-2 Genotype.

Background: Primary amoebic meningoencephalitis (PAM) is an acute and fulminant CNS infection caused by Naegleria fowleri. Recreational activities and ritual ablution with contaminated warm fresh water are the main reason of PAM. Pakistan ranked the second most affected country, where most of the PAM incidences were reported from Karachi, Pakistan. Methods: In May, 2019, a 28-yr-old suspected PAM patient came to the Imam Zain-Ul-Abdin Hospital, Karachi. Biochemical and cytological investigations of patient's CSF were carried out at Karachi Diagnostic Center and Molecular Biology Lab. Sequencing of Naegleria sp. specific (ITS) primer-based amplicons was performed from both patient's CSF and water samples followed by multiple sequence alignment and phylogenetic studies. Results: Biochemical and cytological investigations of patient's CSF showed 5 mg/dl glucose, 240 mg/dl total protein and 2260/mm3 TLC suggesting acute meningoencephalitis. PCR-based analyses of patient's CSF and his residential tap water samples using Naegleria sp. specific (ITS) and N. fowleri specific primers revealed the presence of N. fowleri DNA. Nucleotide sequences of ITS primer-based amplicons from both patient's CSF and water samples were submitted in GenBank under the accession numbers MT726981.1 and MT726226.1, respectively. According to phylogenetic analysis, N. fowleri isolate from Pakistan has shown the least node age of seven. Conclusion: Here, for the very first time in Pakistan, N. fowleri genotype has been identified as type-2. Phylogenetic analysis showed that N. fowleri isolate from Pakistan is among the latest descendants, i.e., evolved later in life.

Intensification in Genetic Information and Acquisition of Resistant Genes in Genome of Acinetobacter baumannii: A Pan-Genomic Analysis.

Acinetobacter baumannii (A. baumannii) attributes 26% of the mortality rate in hospitalized patients, and the percentage can rise to 46 in patients admitted to ICU as it is a major cause of ventilator-associated pneumonia. It has been nominated as the critical priority organism by WHO for which new therapeutic drugs are urgently required. To understand the genomic identification of different strains, antimicrobial resistance patterns, and epidemiological typing of organisms, whole-genome sequencing (WGS) analysis provides insight to explore new epitopes to develop new drugs against the organism. Therefore, the study is aimed at investigating the whole genome sequence of A. baumannii strains to report the new intensifications in its genomic profile. The genome sequences were retrieved from the NCBI database system. Pan-genome BPGA (Bacterial Pan-genome Analysis Tool) was used to analyze the core, pan, and species-specific genome analysis. The pan and core genome curves were extrapolated using the empirical power law equation f(x) = a.xb and the exponential equation f1(x) = c.e (d.x). To identify the resistant genes with resistant mutations against antibiotics, ResFinder and Galaxy Community hub bioinformatics tools were used. According to pan-genome analysis, there were 2227 core genes present in each species of the A. baumannii genome. Furthermore, the number of accessory genes ranged from 1182 to 1460, and the unique genes in the genome were 931. There were 325 exclusively absent genes in the genome of Acinetobacter baumannii. The pan-genome analysis showed that there is a 5-fold increase in the genome of A. baumannii in 5 years, and the genome is still open. There is the addition of multiple unique genes; among them, genes participating in the function of information and processing are increased.