Najeeb Ullah

Helicoverpa armigera (cotton bollworm) is a globally distributed lepidopteran pest that causes estimated annual agricultural losses exceeding 5 billion USD. While chemical pesticides remain the primary control strategy, their prolonged use has led to significant environmental contamination, development of widespread insecticide resistance, and non-target organism toxicity. These limitations underscore the critical need for plant-derived biopesticides that offer target specificity, environmental biodegradability, and sustainable pest management solutions without promoting resistance development. Here, we elucidate the insecticidal potential of Nelumbo nucifera Chitinase (NnChi) against the insect H. armigera through structural informatics and in-vivo insecticidal bioassays. SDS-PAGE showed a single band of ~32 kDa, and LC-MS/MS analysis depicted a fragment of 10 amino acids with 100% identity with Family 19 Class I Chitinase of Mangifera indica. NnChi-predicted structure revealed its two domains (ChB D, Cat D) connected through linker region and docking analysis of both these domains with (GlcNAc)₄ showing binding affinities of -5.6 kcal/mol and -7.0 kcal/mol, respectively. MD simulation (100 ns) showed that 4 residues (RQCR) of ChB D and 4 residues (NRIP) of Cat D contributed to binding with (GlcNAc)₄. To the best of our knowledge, we are reporting the molecular interactions of both domains (ChB D and Cat D) with (GlcNAc)₄ via simulation studies for the first time. These computational findings were further verified through insecticidal assay. Significant larval mortality of H. armigera was observed from 3^rd-6^th instar against 15 µg/g NnChi treatment. Among life cycle parameters, larval and pupal duration, adult eclosion, larval and pupal weight are significantly decreased in a concentration-dependent manner as compared to control. Our integrated structural-functional characterization demonstrates that NnChi exhibits significant insecticidal activity against Helicoverpa armigera. These findings establish NnChi as a promising biopesticide candidate worthy of further investigation.

HIV (human immunodeficiency virus) presents a global health crisis, causing significant AIDS-related deaths and over one million new infections annually. The curbing of HIV is an intricate and continuously evolving domain, marked by numerous challenges, including drug resistance and the absence of a significant cure or vaccine because of its mutating ability and diverse antigens in its envelope, prompting research for functional cures and long-term remission strategies. The endeavor to devise an HIV vaccine capable of eliciting robust and broadly cross-reactive humoral and cellular immune responses is a formidable undertaking, primarily due to the pronounced genetic heterogeneity of HIV-1, the variances observed in virus subtypes (clades) across distinct geographic regions, and the polymorphic nature of human leukocyte antigens (HLA). The viral envelope protein (gp120) selectively interacts with CD4 and chemokine receptors on the surface of target cells. It serves as the key initiator in the intricate viral entry into host cells, rendering it a compelling candidate for vaccine development. This study used bioinformatic tools to design a safe, hypoallergenic, and non-toxic mRNA HIV-1 vaccine by assembling immunogenic B- and T-cell epitopes from the gp120 protein. We identified antigenic, non-toxic, and non-allergic B-cell epitopes (IEPLGIAPTRAKRRVVER) and T-cell epitopes (QQKVHALFY, ITIGPGQVF, WQGVGQAMY, APTRAKRRV, KQQKVHALFYRLDIV, QQKVHALFYRLDIVQ, QKVHALFYRLDIVQI, SLAEEEIIIRSENLT, and IRSENLTNNVKTIIV). For designing the mRNA vaccine against HIV-1 gp120, we assembled the epitopes with 5′ m7G cap, 5′ UTR (untranslated region), Kozak sequence, signal peptide (tPA), RpfE (resuscitation-promoting factor E) adjuvant at N-terminal and MITD (MHC class I trafficking domain) adjuvant, stop codon, 3′ UTR, and 120-nucleotide long poly(A) tail at the C-terminal with immunogenic robustness linkers. The mRNA vaccine is translated into a protein-based vaccine by the host body’s ribosomes. Their comprehensive computational findings, including physicochemical, structural, and 3D refinement analyses, substantiated the stability and quality of the translated vaccine. Molecular docking and simulation revealed a strong and stable binding affinity of vaccine immunization with immune cells’ pattern recognition receptors (TLR4). Immune simulations demonstrated a potent primary immune response characterized by a gradual increase in immunoglobulins and a corresponding decline in antigen concentration. This bioinformatics-driven study presents a promising HIV-1 mRNA vaccine candidate, underscoring the need for further experimental validation through preclinical and clinical trials. At the same time, its methodologies hold the potential for addressing other challenging infectious diseases, thereby impacting vaccinology broadly.

Opisthorchis viverrini is the etiological agent of the disease opisthorchiasis and related cholangiocarcinoma (CCA). It infects fish-eating mammals and more than 10 million people in Southeast Asia suffered from opisthorchiasis with a high fatality rate. The only effective drug against this parasite is Praziquantel, which has significant side effects. Due to the lack of appropriate treatment options and the high death rate, there is a dire need to develop novel therapies against this pathogen. In this study, we designed a multi-epitope chimeric vaccine design against O. viverrini by using immunoinformatics approaches. Non-allergenic and immunogenic MHC-1, MHC-2, and B cell epitopes of three candidate proteins thioredoxin peroxidase (Ov-TPx-1), cathepsin F1 (Ov-CF-1) and calreticulin (Ov-CALR) of O. viverrini, were predicted to construct a potent multiepitope vaccine. The coverage of the HLA-alleles of these selected epitopes was determined globally. Four vaccine constructs made by different adjuvants and linkers were evaluated in the context of their physicochemical properties, antigenicity, and allergenicity. Protein-protein docking and MD simulation found that vaccines 3 was more stable and had a higher binding affinity for TLR2 and TLR4 immune receptors. In-silico restriction cloning of vaccine model led to the formation of plasmid constructs for expression in a suitable host. Finally, the immune simulation showed strong immunological reactions to the engineered vaccine. These findings suggest that the final vaccine construct has the potential to be validated by in vivo and in vitro experiments to confirm its efficacy against the CCA causing O. viverrini.

SARS-CoV-2 papain-like protease (PLpro) covers multiple functions. Beside the cysteine-protease activity, facilitating cleavage of the viral polypeptide chain, PLpro has the additional and vital function of removing ubiquitin and ISG15 (Interferon-stimulated gene 15) from host-cell proteins to support coronaviruses in evading the host’s innate immune responses. We identified three phenolic compounds bound to PLpro, preventing essential molecular interactions to ISG15 by screening a natural compound library. The compounds identified by X-ray screening and complexed to PLpro demonstrate clear inhibition of PLpro in a deISGylation activity assay. Two compounds exhibit distinct antiviral activity in Vero cell line assays and one inhibited a cytopathic effect in non-cytotoxic concentration ranges. In the context of increasing PLpro mutations in the evolving new variants of SARS-CoV-2, the natural compounds we identified may also reinstate the antiviral immune response processes of the host that are down-regulated in COVID-19 infections.

Biofilm synthesizing multi-drug resistantStaphylococcus pseudintermediusbacteria has been recognized as the human infectious agent. It has been detected in the diseases of skin, ear, and postoperative infections. Its infections are becoming a major health problem due to its multi-drug resistance capabilities. However, no commercial vaccine for the treatment of its infections is currently available in the market. Here we employed the subtractive proteomics and reverse vaccinology approach to determine the potential novel drug and vaccine targets againstS. pseudintermediusinfections in humans. After screening the core-proteome of the 39 complete genomes ofS. pseudintermedius, 2 metabolic pathways dependent and 34 independent proteins were determined as novel potential drug targets. Two proteins were found and used as potential candidates for designing the chimeric vaccine constructs. Depending on the properties such as antigenicity, toxicity and solubility, multi-epitope based vaccines constructs were designed. For immunogenicity enhancement, different specific sequences like linkers, PADRE sequences and molecular adjuvants were added. Molecular docking and molecular dynamic simulation analyses were performed to evaluate the prioritized vaccine construct’s interactions with human immune cells HLA and TLR4. Finally, the cloning and expression ability of the vaccine construct was determined in the bacterial cloning system and human body immune response was predicted through immune simulation analysis. In conclusion, this study proposed the potential drug and vaccine targets and also designed a chimera vaccine to be tested and validated against infectiousS. pseudintermediusspecies.

The research work presented in this study is mainly concerned with the bioactivity-directed phytochemical and biological evaluation of Persea duthiei. Persea duthiei is a typical medicinal plant used to treat a variety of ailments such as asthma, edema, and bronchitis. Ethyl acetate, n-hexane, n-butanol, and compounds that are soluble in water were used to examine the antibacterial as well as antifungal capacities of the plant. The antibacterial activity of the soluble parts of ethyl acetate and n-hexane against Escherichia coli, Staphylococcus aureus, Salmonella typhi, and Bacillus subtilis was high, even though there was no activity against Pseudomonas aeruginosa. Likewise, the n-hexane and ethyl acetate fractions were found to have substantial efficacy against several fungal strains such as Aspergillus flavus, Aspergillus fumigates, Fusarium solani, and Aspergillus niger, but not against Candida glabrata. Among the studied fractions, the ethyl acetate soluble fraction had potent antibacterial activity against all of the tested species. This fraction was submitted to phytochemical analysis utilizing various chromatographic methods for the extraction of various pure components. As a consequence, four compounds were isolated, and their structures were elucidated using various spectroscopic methods such as IR, EIMS, HR-EIMS, ¹H-NMR, ¹³C-NMR, NOESY, COSY, HMBC, and HMQC. Urs-12-en-3β-ol (α-amyrine) (1), Urs-12-ene-2α-3β-diol (chamaedrydiol) (2), 3β-hydroxyurs-12-en-28-aldehyde (ursolic aldehyde) (3), and 12-oleanex-3β-ol (β-amyrine) (4) were extracted. Compounds 1, 2, 3, and 4 were examined for antibacterial and antifungal activity and found to have zones of inhibition ranging from 0 to 11mm against tested bacteria strains and percent inhibition ranging from 0 to 25 percent against fungus strains. Compounds 1 and 4 showed strong efficacy against the investigated fungal species, with a 25 inhibition rate. In the case of antibacterial activity, compounds 4 and 1 showed potent activity with zones of inhibition of 11mm and 10mm, respectively. Compounds 2 and 3 were observed to have nonsignificant antimicrobial activity. However, docking studies reflected the complex formation of compound 1 with beta-hydroxyacyl-ACP dehydratase HadAB and S. aureus tyrosyl-tRNA synthetase and compound 2 with topoisomerase II DNA gyrase complex, and they were reported to have antibacterial properties. Similarly, compound 4 was discovered to be well compatible with the lanosterol 14-demethylase (fungal enzyme) and is thus regarded as having antifungal capabilities. Chimera software was used to identify the binding pockets of these complexes. These results indicated that Persea duthiei is a valuable source of medicinal compounds for medication development.

Deaths related to drug errors are common in Pakistan, but these are not accurately reported. Since medication management is the main responsibility of nurses, it is vital that they have a good understanding of high alert of medication error. Patient health care, particularly drug fortification, is the main exertion and the challenge for healthcare professionals around the world. The profession of a pharmacist is world-renowned for providing medical care to patients. Herein, we aim to assess the role of pharmacist according to medications error in Government sectors hospitals located in district Bannu Kp Pakistan. We collected a total number of 368 outdoor prescriptions from July 2011 to December 2011 from District Head Quarter Hospital, Bannu (DHQB). We found 71% of drug-drug interactions (DDIs), 32% of inapt dosage faults and 35% of management errors among them. Male patients with angina pectoris and myocardial infarction had a higher MEs ratio than female patients. According to the findings, clinical pharmacists in hospital wards must provide prompt counseling to primary care doctors during the prescription process, as well as management recommendations to nursing staff and other auxiliary medical employees.

Jatropha curcas L. is a non-edible oilseed containing almost 40% of seed oil and is famous as the best source of raw material for biofuel production. J. curcas seeds contain three main tissues, such as inner integument, endosperm, and embryo. To best understand the physiological events related to specific tissues, it is important to perform the proteome analysis of these tissues. Previously we have explored the pattern of reserves deposition and tissue-specific biological pathways by analyzing the proteome of the inner integument and endosperm and organelles, such as plastids and gerontoplasts isolated from these tissues. The focus of the present study was to perform the proteomic analysis of embryo isolated from the mature seeds of J. curcas. This analysis resulted in the identification of 564 proteins of which 206 are not identified previously from any other tissue of this plant. The identified proteins were functionally classified using the MapMan classification system revealing various proteins involved in different functionalities. The proteins involved in transport functions and those with proteolytic activity were determined through the Transporter Classification Database (TCDB) and MEROPS database, respectively. In addition to identify a large number of proteins participating in various metabolic processes, we found several proteins involved in defense functions, such as the members of chaperones and the ubiquitin-proteasome system. Similarly, members of the legumin and vicilin family of seed storage proteins (SSPs) were identified which in addition to their storage function, are involved in defense. In addition, we have reported that proteases belonging to different mechanistic classes and are involved in diverse physiological functions. Last but not the least, several classes of transport-related proteins were identified that are discussed concerning their function in the transportation of different nutrients across the embryo. To the best of our knowledge, this study reported the highest number of proteins identified from the embryo of mature J. curcas seeds, most of which are essential for seed germination, reflecting the fact that many proteins required for germination are already present in the mature embryo.

Silymarin is proclaimed to be a blend of flavonolignans or phytochemicals. An era of new generation of direct‐acting antivirals (DAAs) has commenced to have facet effect in swaying of the hepatitis C virus (HCV). Nonetheless, this therapy has serious side effects that jeopardize its efficacy. This study is aimed at probing the effects of ribavirin (RBV) and sofosbuvir (SOF) along with silymarin as an adjunct therapy on hematological parameters and markers of obscured oxidative stress. The effect of DAAs along with silymarin was also examined on variable sex hormone level and liver function markers such as alanine aminotransferase (ALT), aspartate transaminase (AST), alkaline phosphatase (ALP), and bilirubin. The study was followed to determine viral load and viral genotypes. A total of 30 patients were randomly divided into two equal groups comprising the control group (n = 15) and treatment group (n = 15). The control group was solely administered with DAAs (SOF and RBV; 400 mg/800 mg each/day). Conversely, the treatment group was dispensed with DAAs, but with adjunct therapy of silymarin (400 mg/day) along with DAAs (400/800 mg/day) over period of 8 weeks. Sampling of blood was performed at pre‐ and posttreatment levels for the evaluation of different propound parameters. Our data showed that silymarin adjunct therapy enhances the efficiency of DAAs. A decrease in menace level of liver markers such as ALT, ALP, AST, and bilirubin was observed (p > 0.05). The adjunct therapy concurrently also demonstrated an ameliorative effect on hematological indices and oxidative markers, for instance, SOD, TAS, GSH, GSSG, and MDA (p < 0.05), diminishing latent viral load. The silymarin administration was also found to revamp the fluster level of sex hormones. Our outcomes provide evidence that systematic administration of silymarin effectively remits deviant levels of hematological, serological, hormonal, and antioxidant markers. This demonstrates a possibly unique role of silymarin in mitigating hepatitis C.