Harnessing artificial intelligence-driven approach for enhanced indole-3-acetic acid from the newly isolated Streptomyces rutgersensis AW08.

Indole-3-acetic acid (IAA) derived from Actinobacteria fermentations on agro-wastes constitutes a safer and low-cost alternative to synthetic IAA. This study aims to select a high IAA-producing Streptomyces-like strain isolated from Lake Oubeira sediments (El Kala, Algeria) for further investigations (i.e., 16S rRNA gene barcoding and process optimization). Subsequently, artificial intelligence-based approaches were employed to maximize IAA bioproduction on spent coffee grounds as high-value-added feedstock. The specificity was the novel application of the Limited-Memory Broyden-Fletcher-Goldfarb-Shanno Box (L-BFGS-B) optimization algorithm. The new strain AW08 was a significant producer of IAA (26.116 ± 0.61 µg/mL) and was identified as Streptomyces rutgersensis by 16S rRNA gene barcoding and phylogenetic inquiry. The empirical data involved the inoculation of AW08 in various cultural conditions according to a four-factor Box Behnken Design matrix (BBD) of Response surface methodology (RSM). The input parameters and regression equation extracted from the RSM-BBD were the basis for implementing and training the L-BFGS-B algorithm. Upon training the model, the optimal conditions suggested by the BBD and L-BFGS-B algorithm were, respectively, L-Trp (X1) = 0.58 %; 0.57 %; T° (X2) = 26.37 °C; 28.19 °C; pH (X3) = 7.75; 8.59; and carbon source (X4) = 30 %; 33.29 %, with the predicted response IAA (Y) = 152.8; 169.18 µg/mL). Our findings emphasize the potential of the multifunctional S. rutgersensis AW08, isolated and reported for the first time in Algeria, as a robust producer of IAA. Validation investigations using the bioprocess parameters provided by the L-BFGS-B and the BBD-RSM models demonstrate the effectiveness of AI-driven optimization in maximizing IAA output by 5.43-fold and 4.2-fold, respectively. This study constitutes the first paper reporting a novel interdisciplinary approach and providing insights into biotechnological advancements. These results support for the first time a reasonable approach for valorizing spent coffee grounds as feedstock for sustainable and economic IAA production from S. rutgersensis AW08.

Blockage of KHSRP-NLRP3 by MCC950 Can Reverse the Effect of Manganese-Induced Neuroinflammation in N2a Cells and Rat Brain.

Manganese neurotoxicity has been reported to cause a neurodegenerative disease known as parkinsonism. Previous reports have shown that the expression of the KH-type splicing regulatory protein (KHSRP), a nucleic acid-binding protein, and NLRP3 is increased upon Mn exposure. However, the relation between these two during Mn toxicity has not been fully deduced. The mouse neuroblastoma (N2a) and SD rats are treated with LPS and MnCl2 to evaluate the expression of KHSRP and NLRP3. Further, the effect of the NLRP3 inhibitor MCC950 is checked on the expression of NLRP3, KHSRP and pro-inflammatory markers (TNFα, IL-18 and IL-1ß) as well as the caspase-1 enzyme. Our results demonstrated an increment in NLRP3 and KHSRP expression post-MnCl2 exposure in N2a cells and rat brain, while on the other hand with LPS exposure only NLRP3 expression levels were elevated and KHSRP was found to be unaffected. An increased expression of KHSRP, NLRP3, pro-inflammatory markers and the caspase-1 enzyme was observed to be inhibited with MCC950 treatment in MnCl2-exposed cells and rats. Manganese exposure induces NLRP3 and KHSRP expression to induce neuroinflammation, suggesting a correlation between both which functions in toxicity-related pathways. Furthermore, MCC950 treatment reversed the role of KHSRP from anti-inflammatory to pro-inflammatory.

Production and Purification of Pectinase from Bacillus subtilis 15A-B92 and Its Biotechnological Applications.

Enzymes that degrade pectin are called pectinases. Pectinases of microbial origin are used in juice clarification as the process is cost-effective. This study screened a pectinase-producing bacterium isolated from soil and identified as Bacillus subtilis 15A B-92 based on the 16S rRNA molecular technique. The purified pectinase from the isolate showed 99.6 U/mg specific activity and 11.6-fold purity. The molecular weight of the purified bacterial pectinase was 14.41 ± 1 kD. Optimum pectinase activity was found at pH 4.5 and 50 °C, and the enzyme was 100% stable for 3.5 h in these conditions. No enzymatic inhibition or activation effect was seen with Fe2+, Ca2+, or Mg2+. However, a slight inhibition was seen with Cu2+, Mn2+, and Zn2+. Tween 20 and 80 slightly inhibited the pectinase, whereas iodoacetic acid (IAA), ethylenediaminetetraacetate (EDTA), urea, and sodium dodecyl sulfate (SDS) showed potent inhibition. The bacterial pectinase degraded citrus pectin (100%); however, it was inactive in the presence of galactose. With citrus pectin as the substrate, the Km and Vmax were calculated as 1.72 mg/mL and 1609 U/g, respectively. The high affinity of pectinase for its substrate makes the process cost-effective when utilized in food industries. The obtained pectinase was able to clarify orange and apple juices, justifying its application in the food industry.

In Silico Molecular Docking and Simulation Studies of Protein HBx Involved in the Pathogenesis of Hepatitis B Virus-HBV.

Current drug discovery involves finding leading drug candidates for further development. New scientific approaches include molecular docking, ADMET studies, and molecular dynamic simulation to determine targets and lead compounds. Hepatitis B is a disease of concern that is a life-threatening liver infection. The protein considered for the study was HBx. The hepatitis B X-interacting protein crystal structure was obtained from the PDB database (PDB ID-3MSH). Twenty ligands were chosen from the PubChem database for further in silico studies. The present study focused on in silico molecular docking studies using iGEMDOCK. The triethylene glycol monoethyl ether derivative showed an optimum binding affinity with the molecular target HBx, with a high negative affinity binding energy of -59.02 kcal/mol. Lipinski's rule of five, Veber, and Ghose were followed in subsequent ADMET studies. Molecular dynamic simulation was performed to confirm the docking studies and to analyze the stability of the structure. In these respects, the triethylene glycol monoethyl ether derivative may be a promising molecule to prepare future hepatitis B drug candidates. Substantial research effort to find a promising drug for hepatitis B is warranted in the future.

In Vitro Antibacterial Activity of Green Synthesized Silver Nanoparticles Using Azadirachta indica Aqueous Leaf Extract against MDR Pathogens.

Rice is the most important staple food crop feeding more than 50% of the world's population. Rice blast is the most devastating fungal disease, caused by Magnaporthe oryzae (M. oryzae) which is widespread in rice growing fields causing a significant reduction in the yield. The present study was initiated to evaluate the effect of green synthesized silver nanoparticles (AgNPs) on the biochemical constituents of rice plants infected with blast. AgNPs were synthesized by using Azadirachta indica leaf extract and their characterization was performed using UV-visible spectroscopy, particle size analyser (PSA), scanning electron microscope (SEM), and X-ray diffraction (XRD) which confirmed the presence of crystalline, spherical shaped silver nanoparticles with an average size of 58.9 nm. After 45 days of sowing, artificial inoculation of rice blast disease was performed. After the onset of disease symptoms, the plants were treated with AgNPs with different concentrations. Application of nanoparticles elevated the activity of antioxidative enzymes such as superoxide dismutase, catalase, peroxidase, glutathione reductase, and phenylalanine ammonia-lyase compared to control plants, and total phenol and reducing sugars were also elevated. The outcome of this study showed that an increase in all biochemical constituents was recorded for A. indica silver nanoparticles-treated plants. The highest values were recorded in 30 ppm and 50 ppm AgNPs-treated plants, which showed the highest resistance towards the pathogen. Green synthesized AgNPs can be used in future for disease control in susceptible varieties of rice. The synthesized AgNPs using A. indica leaf extract have shown promising antibacterial activity when tested against 14 multidrug-resistant (MDR) bacteria comprising Gram-negative bacteria Escherichia coli (n = 6) and Klebsiella pneumoniae (n = 7) with a good zone of inhibition diameter, tested with the disc diffusion method. Based on these findings, it appears that A. indica AgNPs have promise as an antibacterial agent effective against MDR pathogens.

Characterization of Bioactive Compounds from Acacia concinna and Citrus limon, Silver Nanoparticles' Production by A. concinna Extract, and Their Biological Properties.

The applications of bioactive compounds from medicinal plants as therapeutic drugs are largely increasing. The present study selected the bioactive compounds from Acacia concinna (A. concinna) and Citrus limon (C. limon) to assess their phytochemicals, proteins, and biological activity. The plant material was collected, and extraction performed as per the standard procedure. Qualitative analysis was undertaken, and identification of functional organic groups was performed by FTIR and HPLC. Antibacterial, anticancer, antioxidant, antihyperglycemic, antihyperlipidemic, and inhibition kinetics studies for enzymes were performed to assess the different biological activities. Flavonoids and phenols were present in a significant amount in both the selected plants. A. concinna showed significant antimicrobial activity against Z. mobilis, E. coli, and S. aureus, with minimum inhibition zones (MIZ) of 24, 22, and 20 mm, respectively. C. limon strongly inhibited all the tested pathogenic bacteria with maximum and minimum MIZ of 32 and 17 mm. A. concinna silver nanoparticles also exhibited potent antimicrobial activity. Both extracts showed substantial antioxidant, antihyperlipidemic, antidiabetic, anticancer (MCF-7), and anti-urease (antiulcer) properties. To conclude, these plants can be used to treat hyperlipidemia, diabetes, cancer, and gastrointestinal ulcers. They can also serve as antimicrobial and antioxidant agents. Thus, the studied plants must be exploited cost-effectively to generate therapeutic drugs for various diseases.

Plant-Based Synthesis of Gold Nanoparticles and Theranostic Applications: A Review.

Bionanotechnology is a branch of science that has revolutionized modern science and technology. Nanomaterials, especially noble metals, have attracted researchers due to their size and application in different branches of sciences that benefit humanity. Metal nanoparticles can be synthesized using green methods, which are good for the environment, economically viable, and facilitate synthesis. Due to their size and form, gold nanoparticles have become significant. Plant materials are of particular interest in the synthesis and manufacture of theranostic gold nanoparticles (NPs), which have been generated using various materials. On the other hand, chemically produced nanoparticles have several drawbacks in terms of cost, toxicity, and effectiveness. A plant-mediated integration of metallic nanoparticles has been developed in the field of nanotechnology to overcome the drawbacks of traditional synthesis, such as physical and synthetic strategies. Nanomaterials' tunable features make them sophisticated tools in the biomedical platform, especially for developing new diagnostics and therapeutics for malignancy, neurodegenerative, and other chronic disorders. Therefore, this review outlines the theranostic approach, the different plant materials utilized in theranostic applications, and future directions based on current breakthroughs in these fields.

Effect of curcumin on growth, biofilm formation and virulence factor gene expression of Porphyromonas gingivalis.

Porphyromonas gingivalis is a keystone pathogen and major colonizer in host tissue which plays a pivotal role in periodontitis among the other polymicrobial infections. Increasing facts demonstrate that curcumin has antibacterial activity and anti-biofilm effect against the periodontopathogens through diverse mechanisms that have a positive impact on periodontal health. The present study was aimed to elucidate the effect of curcumin on biofilm formation and virulence factor gene expression of P. gingivalis. By using gene expression studies, we exploited the mechanism of anti-biofilm effects of curcumin on P. gingivalis. The minimum inhibitory concentration and minimum bactericidal concentration of curcumin for both ATCC and clinical strains of P. gingivalis were found to be 62.5 and 125 µg ml-1 respectively. Curcumin prevented bacterial adhesion and biofilm formation in a dose-dependent manner. Further, curcumin attenuated the virulence of P. gingivalis by reducing the expression of genes coding for major virulence factors, including adhesions (fimA, hagA, and hagB) and proteinases (rgpA, rgpB, and kgp). The results indicated that curcumin has shown anti-biofilm as well as antibacterial activity against P. gingivalis. Further, curcumin because of its pleiotropic actions could be a simple and inexpensive therapeutic strategy in the treatment of periodontal disease.

In vitro and in vivo inhibitory effects of Tabernaemontana alternifolia against Naja naja venom.

BACKGROUND: Tabernaemontana alternifolia root is traditionally used and practiced among few Indian tribes as an antidote for snakebites. OBJECTIVE: To combat and neutralize Naja naja venom using methanolic root extract of Tabernaemontana alternifolia and to explore its efficacy on venom biomarkers in search of newer herbal antidote or first-aid-point of care for therapeutics.Materialization.Pharmacological activities such as fibrinogenolytic, direct and indirect hemolytic activities for the neutralization of the venom were evaluated. Lethal toxicity annulation studies were performed using the murine model by pre-incubation and post-treatment protocols. Further, the neutralization of edema and myotoxicity were also evaluated. RESULTS: Electrophoretic analysis revealed that the complete neutralization of fibrinogen degradation was observed at 1:10 (w/w) (venom to extract). T. alternifolia exhibited an effective dose (ED50) value of 87.20 µg/mL for venom-induced hemolysis. Venom at 2 µg concentration produced 11 mm of hemolytic radiance and was neutralized at 1:20 (w/w) venom to extract concentration. The survival time and the neurotoxic symptoms in mice were concluded to be delayed by both the methods of lethal toxicity inhibition using methanol extract. The edema ratio reduced the venom to extract ratio of 1:20 (w/w) from 173 ± 45% to 133.61% when subjected to 5 µg of venom concentration. The plant extract significantly neutralized the myotoxic activity. CONCLUSION: T. alternifolia methanolic root extract could be a potent contributor in the effective treatment of N. naja venom-induced toxicity.

Identification of Deleterious SNPs and Their Effects on Structural Level in CHRNA3 Gene.

The aim of our study is to identify probable deleterious genetic variations that can alter the expression and the function of the CHRNA3 gene using in silico methods. Of the 2305 SNPs identified in the CHRNA3 gene, 115 were found to be non-synonymous and 12 and 15 nsSNPs were found to be in the 5' and 3' UTRs, respectively. Further, out of the 115 nsSNPs investigated, eight were predicted to be deleterious by both SIFT and PredictSNP servers. The major mutations predicted to affect the structure of the protein are phenylalanine to valine (Y43V) and lysine to asparagine (K216N) as shown by the trajectory run in molecular dynamics studies. The random transition of the protein structures over the simulation period caused by these mutations hints at how the native state is distorted which could lead to the loss of structural stability and functionality of the nicotinic acetylcholine receptors subunit α-3 protein. Based on this work, we propose that the nsSNP with SNP id of rs75495285 and rs76821682 will have comparatively more deleterious effects than the other predicted mutations in destabilizing the protein structure.

Molecular modeling approach to identify inhibitors of Rv2004c (rough morphology and virulent strain gene), a DosR (dormancy survival regulator) regulon protein from Mycobacterium tuberculosis.

Being a part of dormancy survival regulator (DosR) regulon, Rv2004c (rough morphology and virulent strain gene) has been identified in earlier experimental studies as an indispensable protein required for the growth and survival of Mycobacterium tuberculosis. This protein was predicted to have a role in inhibition of phospholipase A2 activity related to immuno-defence and other membrane-related events. Thus, considering significance of Rv2004c protein, a structure-based drug designing strategy was followed to identify potential inhibitors to this novel target. Initially, to validate the target, absence of homologous proteins in the host was verified through sequence and structure similarity search against human proteome. Then, a potential ligand binding site on the target was identified and virtual screening against Zinc database molecules was carried out. The top scoring hits along with their analogs were taken for docking studies with Glide. The binding free energy of the docked complexes of the Glide hits were predicted by Prime program from Schrodinger and molecules ZINC57990006, ZINC33605742, ZINC71773467 and ZINC34198774 were recognized as potential hits against this target. Analyzing the predicted pharmacokinetic properties of the molecules from QikProp and admetSAR tool, ZINC34198774 was identified as a valid molecule. Molecular dynamics simulation studies ascertained that ZINC34198774 could be a potential inhibitor against Rv2004c. Thus, results acquired from this study could be of use to design new therapeutics against tuberculosis.Communicated by Ramaswamy H. Sarma.

In Vitro Antibacterial Activity of Green Synthesized Silver Nanoparticles Using Mangifera indica Aqueous Leaf Extract against Multidrug-Resistant Pathogens.

An estimated 35% of the world's population depends on wheat as their primary crop. One fifth of the world's wheat is utilized as animal feed, while more than two thirds are used for human consumption. Each year, 17-18% of the world's wheat is consumed by China and India. In wheat, spot blotch caused by Bipolaris sorokiniana is one of the major diseases which affects the wheat crop growth and yield in warmer and humid regions of the world. The present work was conducted to evaluate the effect of green synthesized silver nanoparticles on the biochemical constituents of wheat crops infected with spot blotch disease. Silver nanoparticles (AgNPs) were synthesized using Mangifera indica leaf extract and their characterization was performed using UV-visible spectroscopy, SEM, XRD, and PSA. Characterization techniques confirm the presence of crystalline, spherical silver nanoparticles with an average size of 52 nm. The effect of green synthesized nanoparticles on antioxidative enzymes, e.g., Superoxide dismutase (SOD), Catalase (CAT), Glutathione Reductase (GR), Peroxidase (POX), and phytochemical precursor enzyme Phenylalanine Ammonia-Lyase (PAL), and on primary and secondary metabolites, e.g., reducing sugar and total phenol, in Bipolaris sorokiniana infected wheat crop were studied. Inoculation of fungal spores was conducted after 40 days of sowing. Subsequently, diseased plants were treated with silver nanoparticles at different concentrations. Elevation in all biochemical constituents was recorded under silver nanoparticle application. The treatment with a concentration of nanoparticles at 50 pp min diseased plants showed the highest resistance towards the pathogen. The efficacy of the green synthesized AgNPs as antibacterial agents was evaluated against multi drug resistant (MDR) bacteria comprising Gram-negative bacteria Escherichia coli (n = 6) and Klebsiella pneumoniae (n = 7) and Gram-positive bacteria Methicillin resistant Staphylococcus aureus (n = 2). The results show promising antibacterial activity with significant inhibition zones observed with the disc diffusion method, thus indicating green synthesized M. indica AgNPs as an active antibacterial agent against MDR pathogens.

Curcumin-Encapsulated Nanomicelles Improve Cellular Uptake and Cytotoxicity in Cisplatin-Resistant Human Oral Cancer Cells.

Oral cancer has a high mortality rate, which is mostly determined by the stage of the disease at the time of admission. Around half of all patients with oral cancer report with advanced illness. Hitherto, chemotherapy is preferred to treat oral cancer, but the emergence of resistance to anti-cancer drugs is likely to occur after a sequence of treatments. Curcumin is renowned for its anticancer potential but its marred water solubility and poor bioavailability limit its use in treating multidrug-resistant cancers. As part of this investigation, we prepared and characterized Curcumin nanomicelles (CUR-NMs) using DSPE-PEG-2000 and evaluated the anticancer properties of cisplatin-resistant cancer cell lines. The prepared CUR-NMs were sphere-shaped and unilamellar in structure, with a size of 32.60 ± 4.2 nm. CUR-NMs exhibited high entrapment efficiency (82.2%), entrapment content (147.96 µg/mL), and a mean zeta potential of -17.5ζ which is considered moderately stable. The cellular uptake and cytotoxicity studies revealed that CUR-NMs had significantly higher cytotoxicity and cellular uptake in cisplatin drug-resistant oral cancer cell lines and parental oral cancer cells compared to plain curcumin (CUR). The DAPI and FACS analysis corroborated a high percentage of apoptotic cells with CUR-NMs (31.14%) compared to neat CUR (19.72%) treatment. Conclusively, CUR-NMs can potentially be used as an alternative carrier system to improve the therapeutic effects of curcumin in the treatment of cisplatin-resistant human oral cancer.

Molecular modelling and simulation techniques to investigate the effects of fungal metabolites on the SARS-CoV-2 RdRp protein inhibition.

Various protein/receptor targets have been discovered through in-silico research. They are expanding rapidly due to their extensive advantage of delivering new drug candidates more quickly, efficiently, and at a lower cost. The automation of organic synthesis and biochemical screening will lead to a revolution in the entire research arena in drug discovery. In this research article, a few fungal metabolites were examined through an in-silico approach which involves major steps such as (a) Molecular Docking Analysis, (b) Drug likeness and ADMET studies, and (c) Molecular Dynamics Simulation. Fungal metabolites were taken from Antibiotic Database which showed antiviral effects on severe viral diseases such as HIV. Docking, Lipinski's, and ADMET analyses investigated the binding affinity and toxicity of five metabolites: Chromophilone I, iso; F13459; Stachyflin, acetyl; A-108836; Integracide A (A-108835). Chromophilone I, iso was subjected to additional analysis, including a 50 ns MD simulation of the protein to assess the occurring alterations. This molecule's docking data shows that it had the highest binding affinity. ADMET research revealed that the ligand might be employed as an oral medication. MD simulation revealed that the ligand-protein interaction was stable. Finally, this ligand can be exploited to develop SARS-CoV-2 therapeutic options. Fungal metabolites that have been studied could be a potential source for future lead candidates. Further study of these molecules may result in creating an antiviral drug to battle the SARS-CoV-2 virus.

Cancer Stem Cell Traits in Tumor Spheres Derived from Primary Laryngeal Carcinoma Cell Lines.

OBJECTIVE: Cancer stem cells (CSCs) belong to a subpopulation of undifferentiated cells present within tumors that have the potential to regenerate, differentiate, maintenance of pluripotency, drug resistance, and tumorigenicity when transplanted into an innate host. These can influence the growth and behavior of these tumors and are used to investigate the initiation, progression, and treatment strategies of laryngeal cancer. Research on CSC science and targeted therapies were hinge on their isolation and/or enrichment procedures. The object of the study is to isolate cancer stem cells from primary laryngeal carcinoma (CSCPLC) by tumor spheres enrichment. We checked the properties of self-renewal, stemness, clonogenicity, and chemotherapeutic resistance. MATERIALS AND METHODS: We performed tumor sphere formation assay (primary, secondary, and tertiary) chemotherapy resistance by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay were performed to evaluate the CSC cells. Immunofluorescence for stem cell markers (CD133+, CD44+) and gene expression of stem cell markers for CD133+, CD44+, OCT4, SOX2, and NANOG was done using the real-time polymerase chain reaction technique. RESULTS: We were able to isolated CSC subpopulations from PLC cell lines by the tumor sphere method. These cells exhibited good primary, secondary, and tertiary tumor sphere formation efficiency and also disclosed a resistant index of more than 2. Immunofluorescence for stem cell markers (CD133+ and CD44+) confirms the presence of CSC. There was significantly higher mRNA expression of stem cell markers in CSC enriched subpopulations compared to the parental cell lines. CONCLUSION: We conclude that tumor spheres enrichment is an efficient, economical, and reliable approach for the isolation and characterization of CSC from PLC cell lines. These cells demonstrated the properties of self-renewal, stemness, clonogenicity, and chemotherapeutic resistance.

Statistical optimization and characterization of bacterial cellulose produced by isolated thermophilic Bacillus licheniformis strain ZBT2.

Bacterial cellulose (BC) is an excellent natural biopolymer with wide range of applications. The present study reports a potential BC producing thermophile, identified as Bacillus licheniformis strain ZBT2. The thermophile produced pellicle form of BC (3.0 g/l) under static conditions. Statistical optimization of BC was carried out by Plackett-Burman and central composite design. Results suggest that BC yield (9.2 g/l) was enhanced with 6.6-fold after optimization. BC-gelatin hydrogels composites were developed to assess various properties. The water retention capability and moisture content properties of BC and composites were promising and also exhibited negligible protein adsorption. The composites also demonstrated to be consistent during controlled drug delivery profiling. Furthermore, the composites also demonstrated antibacterial efficiency against Escherichia coli and Micrococcus luteus. The structural, morphological and thermal properties were assessed by analytical techniques such as, fourier transform infrared spectroscopy, scanning electron microscopy with energy dispersive X-ray analysis, thermogravimetric analysis and differential scanning calorimetry analysis. The study reflects the exploitation of a thermophile for development of BC which can be a preferred choice as a scaffold for tissue engineering and drug-delivery systems.

Structural, molecular, functional and immunological characterization of Wuchereria bancrofti-galectin.

Galectins regulate growth and differentiation of immune cells and inflammation through their carbohydrate-binding function in humans, while also play a role in parasite survival. This study focused on the galectin of lymphatic filarial parasite Wuchereria bancrofti (Wb-Gal). The multiple sequence alignment with other galectins showed that the Wb-Gal belonged to galactoside binding lectin family, particularly tandem repeat type galectin-9. A homology model of Wb-Gal was developed in the I-TASser server using high similarity 3D structures with a quality score of 89.5. Molecular docking and dynamics studies revealed that the CCRD and NCRD of Wb-Gal bind with galactose and lactose. Further, Wb-Gal was cloned into the pET28 vector, expressed in E. coli Rosetta strain and purified by affinity chromatography. In the hemagglutination assays, the rWb-Gal bound to lactose, galactose, and glucose. Indirect Enzyme-Linked Immunosorbent Assay (ELISA) using different clinical filarial sera showed that the IgG and IgM response was against Wb-Gal x very high in all filarial clinical groups, whereas the IgA and IgG2 response was minimum to negligible. There was an enhanced response of IgG1 and IgG4 antibodies in Microfilaremics (MF) cases compared to Chronic Pathology (CP) and Endemic Normal (EN) individuals. Interestingly, the IgE response was comparatively higher in EN than MF and CP. These studies show that Wb-Gal is a member of the lectin family of proteins binding to different carbohydrates and may have an important role in the pathophysiology of filarial infection which needs to be investigated in greater detail.

Design, expression, and evaluation of novel multiepitope chimeric antigen of Wuchereria bancrofti for the diagnosis of lymphatic filariasis - A structure-based strategy.

The Global Program for Elimination Lymphatic Filariasis (GPELF) is in an advanced stage and requires tools for diagnosing infection, assessing transmission and certification. This study was aimed at developing an antibody-based assay using a chiemric antigen containing multi-B-cell epitopes from antigens highly expressed in different stages of Wuchereria bancrofti to detect LF infection and its transmission. The antigen was express cloned and two indirect ELISA based (IgG1 & IgG4 based) antibody assays were developed using the recombinant antigen. The chimeric antigen displayed 1 and 3-fold reactivity with IgG1 and IgG4 antibodies, respectively in microfilaraial (mf) positive sera when compared to that in sera samples of Non-endemic normal sera (NEN) (O.D, 0.13 ± 0.20 and 0.18 ± 0.07), thus differentiating infected from uninfected individuals. In IgG1 and IgG4 antibody assays, the multiepitope antigen also showed reactivity (O.D, 0.27 ± 0.18 and 0.16 ± 0.03) in a small proportion (18 and 30, respectively out of 156) endemic normal individuals and in IgG1 antibody in a few (4) chronic patients (CP). The antigen did not react with IgG1 or IgG4 antibodies in the sera samples of malaria, scrub typhus, dengue, hookworm, and roundworm helminth cases (0.139 ± 0.018, 0.144 ± 0.007 0.17804 ± 0.007 and 0.162 ± 0.006), thus showing its high specificity. The sensitivity (%) and specificity (%) of the multi-epitope antigen-based IgG1 and IgG4 antibody assays are 100, 98.1 and 100, 99.52, respectively. Thus, the recombinant multiepitope antigen appears to have good potential in detecting active LF infection and in assessing its transmission in endemic communities.

Variation in the Occurrence of fimA Genotypes of Porphyromonas gingivalis in Periodontal Health and Disease.

Porphyromonas gingivalis is regarded as a "keystone pathogen" in periodontitis. The fimbria assists in the initial attachment, biofilm organization, and bacterial adhesion leading to the invasion and colonization of host epithelial cells. The present study aimed to investigate the occurrence of fimA genotypes in patients with chronic periodontitis and healthy individuals in the Indian population, and to study their association with the number of P. gingivalis cells obtained in subgingival plaque samples of these subjects. The study comprised 95 samples from the chronic periodontitis (CP) group and 35 samples from the healthy (H) group, which were detected positive for P. gingivalis in our previous study. Fimbrial genotyping was done by PCR and PCR-restriction fragment length polymorphism (RFLP). The fimA type II was more prevalent in the CP group (55.89%), followed by type IV (30.52%), whereas in the H group, type I was the most prevalent fimbria (51.42%). The quantity of P. gingivalis cells increased with the presence of fimA types II and III. Our results suggest a strong relationship between fimA types II and IV and periodontitis, and between type I and the healthy condition. The colonization of organisms was increased with the occurrence of type II in deep periodontal sites, which could play an important role in the progression of the disease.