Effect of the Histone Methyltransferase Specific Probe BRD4770 on Metabolic Profiling of the Endophytic Fungus Diaporthe longicolla

The endophytic fungus Diaporthe longicolla was isolated from the stem of Saraca asoca (Roxb.) Willd., commonly known as Ashok plant in India and Sri Lanka. Since no reports are available regarding epigenetic modulations by BRD4770 in microbial entities, D. longicolla was treated with different concentrations of BRD4770 for this purpose and evaluated for its antioxidant and antibacterial potential against five human pathogenic bacteria, Staphylococcus aureus, methicillin-resistant Staphylococcus aureus (MRSA), Shigella boydii, Klebsiella pneumoniae, and Escherichia coli. The crude extract obtained from cultures treated with 100 nM concentration of BRD4770 showed increased antioxidant activity and inhibition zone against S. aureus and MRSA, compared to the non-treated control. The composition of the non-treated and treated crude extract was analyzed, and induced compounds were identified with the help of Gas chromatography–mass spectrometry (GC-MS) and LC-ESI-MS/MS. LC-ESI-MS/MS analysis showed that berberine (antibacterial)-, caffeine-, and theobromine (antioxidant)-like compounds were induced in the BRD4770-treated crude extract. The presence of particular absorbance at a wavelength of 346.5 nm for berberine, 259.4 nm for caffeine, and 278.4 nm for theobromine in the reverse-phase high-performance liquid chromatography (HPLC) analysis of both BRD4770-treated crude metabolites and standard solution of the above compounds strongly supported the increased antibacterial and antioxidant activities that may be due to inducing the alterations in bioactivities of the BRD4770-treated culture

Formulation and Evaluation of once daily sustained release matrix tablets of Aceclofenac using natural gums

In present study, an attempt has been made to evaluate the effect of natural gums on the release profile of drug from matrix system for once daily sustained release tablets formulations. Aceclofenac NSAIDs was used as a model drug to evaluate it release characteristics from different matrices. Matrix tablets of Aceclofenac were prepared by direct compression process using natural gums (xanthan gum and karaya gum) in different ratios drug: gum ratios of FX, FK and FXK (FX and FK in 1:1 ratios). The tablets were evaluated for physical characteristic like hardness, weight variation, friability, swelling index and drug content, in-vitro release of drug was performed in Phosphate buffer pH 7.4 for 24 hours. All the physical characteristic of fabricated tablet was within acceptable limits. The release of Aceclofenac from a gelatinous swollen mass, which controls the diffusion of drug molecules through the polymeric materials in to aqueous medium. The FXK matrices show  prices controlled release than FX and FK matrices because of burst effect and fast release in case of FX and FK matrices respectively and there was no chemical interaction between drug and polymers in FXK formulation as confirmed by FTIR studies. The release mechanism was explained with zero order, first order, higuchi and korsmeyer equations via swelling and non fickian diffusion mechanism. The FXK matrices leads to more  prices result than FX and FK alone by utilisation of synergistic interaction between two biopolymers and uniformity in the hydration layer in dissolution media

DENDRIMERS IN DRUG DELIVERY, DIAGNOSIS AND THERAPY: BASICS AND POTENTIAL APPLICATIONS

This review gives concise information about the dendrimers, properties, synthesis and application in drug delivery, diagnosis and therapy. Due to their unique architecture these have improved physical and chemical properties. They show high solubility, miscibility and reactivity due to their terminal groups. Dendrimers have well defined size, shape, molecular weight and monodispersity. These properties make the dendrimers a suitable carrier in drug delivery application. Dendrimers are unimolecular miceller in nature and due to this enhances the solubility of poorly soluble drugs. Their compatibility with DNA, heparin and polyanions make them more versatile. Dendrimers, also referred as modern day polymers, they offer much more good properties than the conventional polymers. Due to their multivalent and mono disperse character dendrimers have stimulated wide interest in the field of chemistry biology, especially in applications like drug delivery, gene therapy and chemotherapy. Self assembly produces a faster means of generating nanoscopic functional and structural systems. But their actual utility in drug delivery can be assessed only after deep understanding of factors affecting their properties and their behavior in vivo. Key words: Dendrimers, PAMAM, monodispersity, Divergent-Convergent synthesis, carrier for drug deliveryÂ

Drug Susceptibility in Leishmania Isolates Following Miltefosine Treatment in Cases of Visceral Leishmaniasis and Post Kala-Azar Dermal Leishmaniasis

Resistance to antimonials has emerged as a major hurdle to the treatment and control of VL and led to the introduction of Miltefosine as first line treatment in the Indian subcontinent. MIL is an oral drug with a long half-life, and it is feared that resistance may emerge rapidly, threatening control efforts under the VL elimination program. There is an urgent need for monitoring treatment efficacy and emergence of drug resistance in the field. In a set of VL/PKDL cases recruited for MIL treatment, we observed comparable drug susceptibility in pre- and post-treatment isolates from cured VL patients while MIL susceptibility was significantly reduced in isolates from VL relapse and PKDL cases. The PKDL isolates showed higher tolerance to MIL as compared to VL isolates. Both VL and PKDL isolates were uniformly susceptible to PMM. MIL transporter genes LdMT/LdRos3 were previously reported as potential resistance markers in strains in which MIL resistance was experimentally induced. The point mutations and the down-regulated expression of these transporters observed in vitro could, however, not be verified in natural populations of parasites. LdMT/LdRos3 genes therefore, do not appear to be suitable markers so far for monitoring drug susceptibility in clinical leishmanial isolates

Circulating MicroRNAs: Potential and emerging biomarkers for diagnosis of human infectious diseases

MicroRNAs (miRNAs) are evolutionary conserved, small non-coding RNA with size ranging from 19-24 nucleotides. They endogenously regulate the gene expression at the post transcriptional level either through translation repression or mRNA degradation. MiRNAs have shown the potential to be used as a biomarker for the diagnosis, prognosis and therapy of infectious diseases. Many miRNAs have shown significantly altered expression during infection. The altered expression of miRNA level in an infected human can be identified by the use of advanced diagnostic tools. In this review, we have highlighted the use of miRNA as an emerging tool for the identification of the human infectious disease. Till date, several miRNAs have been reported as a molecular biomarker in infectious diseases, such as miRNA-150 and miRNA-146b-5p in human immunodeficiency virus (HIV); miRNA-122, miRNA-21 and miRNA-34a in hepatitis; miRNA-361-5p and miRNA-29c in tuberculosis; miRNA-16 and miRNA-451 in malaria and miRNA-181 in Helicobacter pylori infection. The diagnosis of infection with the help of a biomarker is a non-invasive tool that has shown to have a key role in early diagnosis of infection. The discovery of circulating miRNA in the blood of infected patients has the potential to become a powerful non-invasive biomarker in coming future

Novel Immunoinformatics Approaches to Design Multi-epitope Subunit Vaccine for Malaria by Investigating Anopheles Salivary Protein

Abstract Malaria fever has been pervasive for quite a while in tropical developing regions causing high morbidity and mortality. The causal organism is a protozoan parasite of genus Plasmodium which spreads to the human host by the bite of hitherto infected female Anopheles mosquito. In the course of biting, a salivary protein of Anopheles helps in blood feeding behavior and having the ability to elicit the host immune response. This study represents a series of immunoinformatics approaches to design multi-epitope subunit vaccine using Anopheles mosquito salivary proteins. Designed subunit vaccine was evaluated for its immunogenicity, allergenicity and physiochemical parameters. To enhance the stability of vaccine protein, disulfide engineering was performed in a region of high mobility. Codon adaptation and in silico cloning was also performed to ensure the higher expression of designed subunit vaccine in E. coli K12 expression system. Finally, molecular docking and simulation study was performed for the vaccine protein and TLR-4 receptor, to determine the binding free energy and complex stability. Moreover, the designed subunit vaccine was found to induce anti-salivary immunity which may have the ability to prevent the entry of Plasmodium sporozoites into the human host

Differential Expression of miRNA Regulates T Cell Differentiation and Plasticity during Visceral Leishmaniasis Infection

Visceral leishmaniasis (VL) is a tropical neglected disease caused by Leishmania donovani, results in significant mortality in the Indian subcontinent. The plasticity of T cell proliferation and differentiation depends on microRNA mediated gene regulation which leads Th1/Th2 or Th17/Treg type of immune response during human VL. This study depicts the identification of target immune signaling molecule and transcription factors, which play a role in T-cell proliferation and differentiation followed by the identification of miRNA controlling their gene expression using three web servers’ viz., TargetScan, miRPath and miRDB. This study provides the bioinformatics evidences that seed region present in the miRNAs miR-29-b, miR-29a, have the putative binding site in the 3’-UTR region of TBX21 transcription factor of CD4+ T helper (Th1), which may suppress the Th1 specific protective immune response. Development of Th2 type specific immune response can be suppressed by binding of miR-135 and miR-126 miRNAs over the 3’-UTR region of GATA-3 transcription factor of Th2 specific CD4+ T helper cells. MiRNA identified against Th2/Treg immune cells are important and their over expression or administration can be used for developing the Th1/Th17 type of protective immune response during VL infection. This study indicates that miRNAs have the capacity to regulate immune signaling, cytokine production and immune cell migration to control the VL infection in human. This observation warrants further investigation for the development of miRNA based therapy controlling T cell differentiation in human VL

Prediction of an immunogenic peptide ensemble and multi-subunit vaccine for Visceral leishmaniasis using bioinformatics approaches

Visceral Leishmaniasis (VL) is a neglected tropical disease of public health importance in the Indian subcontinent. Despite consistent elimination initiatives, the disease has not yet been eliminated and there is an increased risk of resurgence from active VL reservoirs including asymptomatic, post kala azar dermatitis leishmaniasis (PKDL) and HIV-VL co-infected individuals. To achieve complete elimination and sustain it in the long term, a prophylactic vaccine, which can elicit long lasting immunity, is desirable. In this study, we employed immunoinformatic tools to design a multi-subunit epitope vaccine for the Indian population by targeting antigenic secretory proteins screened from the Leishmania donovani proteome. Out of 8014 proteins, 277 secretory proteins were screened for their cellular location and proteomic evidence. Through NCBI BlastP, unique fragments of the proteins were cropped, and their antigenicity was evaluated. B-cell, HTL and CTL epitopes as well as IFN-ɣ, IL-17, and IL-10 inducers were predicted, manually mapped to the fragments and common regions were tabulated forming a peptide ensemble. The ensemble was evaluated for Class I MHC immunogenicity and toxicity. Further, immunogenic peptides were randomly selected and used to design vaccine constructs. Eight vaccine constructs were generated by linking random peptides with GS linkers. Synthetic TLR-4 agonist, RS09 was used as an adjuvant and linked with the constructs using EAAK linkers. The predicted population coverage of the constructs was ∼99.8 % in the Indian as well as South Asian populations. The most antigenic, nontoxic, non-allergic construct was chosen for the prediction of secondary and tertiary structures. The 3D structures were refined and analyzed using Ramachandran plot and Z-scores. The construct was docked with TLR-4 receptor. Molecular dynamic simulation was performed to check for the stability of the docked complex. Comparative in silico immune simulation studies showed that the predicted construct elicited humoral and cell-mediated immunity in human host comparable to that elicited by Leish-F3, which is a promising vaccine candidate for human VL

Evaluation of Blood Agar Microtiter Plates for Culturing Leishmania Parasites To Titrate Parasite Burden in Spleen and Peripheral Blood of Patients with Visceral Leishmaniasis▿

Serial dilution of blood and spleen biopsy specimens, plated on Novy-MacNeal-Nicolle (NNN) blood agar using microtiter culture plates, is a sensitive and reproducible method for detection and growth of Leishmania parasites. Plates could be easily monitored, and growth could be rapidly detected. Moreover, parasite number may be estimated using this technique