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In the present work, degradation of reactive orange 4 dye (RO4) has been investigated using hydrodynamic cavitation (HC) and in combination with other AOP's. In the hybrid techniques, combination of hydrodynamic cavitation and other oxidizing agents such as H2O2 and ozone have been used to get the enhanced degradation efficiency through HC device. The hydrodynamic cavitation was first optimized in terms of different operating parameters such as operating inlet pressure, cavitation number and pH of the operating medium to get the maximum degradation of RO4. Following the optimization of HC parameters, the degradation of RO4 was carried out using the combination of HC with H2O2 and ozone. It has been found that the efficiency of the HC can be improved significantly by combining it with H2O2 and ozone. The mineralization rate of RO4 increases considerably with 14.67% mineralization taking place using HC alone increases to 31.90% by combining it with H2O2 and further increases to 76.25% through the combination of HC and ozone. The synergetic coefficient of greater than one for the hybrid processes of HC+H2O2 and HC+Ozone has suggested that the combination of HC with other oxidizing agents is better than the individual processes for the degradation of dye effluent containing RO4. The combination of HC with ozone proves to be the most energy efficient method for the degradation of RO4 as compared to HC alone and the hybrid process of HC and H2O2., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Japanese encephalitis (JE) remains a major public health threat with vaccination as the only measure for its prevention. Epitope-based vaccination is a promising approach for achieving protective immunity and avoid immunopathology in Japanese encephalitis virus (JEV) infection due to flavivirus cross-reactivity. We have mapped B-cell epitopes from JEV envelope protein, responsible for elicitation of neutralizing antibodies. Incorporation of T helper (T(H)) epitopes, along with these, imparted protective immunity to the host. In the present study, based on in silico epitope selection we optimized and proposed a polytope DNA construct (P-JEV) consisting B-cell and T(H) epitopes from the JEV envelope (E) protein as well as non-structural protein-1 (NS1). The immunogenicity and protective efficacy of P-JEV was assessed by in vitro and in vivo experiments. The expressed P-JEV showed reactivity in in vitro assays with JEV monoclonal antibodies. Protective efficacy of P-JEV was assessed in BALB/c mice. Our findings indicate that P-JEV may be a candidate vaccine for the prevention of JEV infection., (Copyright © 2012 Elsevier B.V. All rights reserved.)

West Nile virus (WNV) and Japanese encephalitis virus (JEV), the members of JEV serocomplex group are pathogens of global health concern. The co-circulation of these viruses poses challenges in effective diagnostics due to antigenic similarity between the E-protein of these viruses. The present study aimed to design chimeric peptides and study the immune response against the same. B-cell epitopes were predicted on structural proteins of WNV and JEV based on bioinformatics tools. The peptides representing to these B-cell epitopes were synthesized and subjected to ELISA. Two peptides, one each from WNV (named WE147) and JEV (named JE40) E-protein, showed virus-specific and strong reactivity to the immune mice sera and human clinical samples. The chimeric peptides for WNV and JEV were constructed by synthesizing the B-cell epitope of WNV (WE147) or JEV (JE40) with T-helper epitope (JM17) separated by diglycine spacer in between. The immune response generated against these chimeric peptides was found to be specific to the respective B-cell epitopes. The anti-peptide sera showed virus-specific reactivity in ELISA and in immunofluorescence assay with no cross-reactivity. Also, the anti-peptide sera could neutralize JE and WN viruses in an in vitro virus neutralization assay. The B-cell epitopes identified in the present study may be used as diagnostic markers for differentiating between WN and JE virus infections. The present study can form a basis for future design of vaccines., (Copyright © 2011 Elsevier B.V. All rights reserved.)

In order to understand the factors influencing pathogenicity of a virus, two neutralization escape (NE) variants were selected from wild type lineage 1 West Nile virus (WNV) 68856 strain pathogenic by intra-peritoneal (i.p.) route using monoclonal antibodies (MAbs) against envelope (E) protein. Both NE IF1A7 1.1 and NE IVC3F10 1.2 were resistant to neutralization and were neurovirulent by intra-cranial (i.c.) inoculation. Growth kinetics in porcine stable (PS) kidney and baby hamster kidney (BHK) cells was unchanged. In contrast to parent WNV only NE IF1A7 1.1 failed to cause lethal encephalitis on i.p. inoculation and was non pathogenic. NE IF1A7 1.1 variant showed delayed replication kinetics in murine peritoneal exudate cells (PEC) and Neuro 346 cells in vitro. In comparison with parent WNV and NE IVC3F10 1.2 variant, non pathogenic variant exhibited significantly reduced tumour necrosis factor α (TNF-α) induction in infected animals and PEC. Other cytokines like Interleukin (IL)-10, IL-6 and Interferon (IFN)-β remained unchanged. However, IL-1β did not follow the pattern and was higher only in parent WNV-infected PEC. The E gene sequences of these NE variants showed three common amino acid substitutions at residues E50, E89 and E242. A unique E156 (ser→pro) substitution in NE IF1A7 1.1, was absent in NE IVC3F10 1.2 variant suggested probable virulence marker. Our data indicates possible role of WNV E protein in induction of TNF-α and IL-1β and its association with WNV pathogenesis., (Copyright © 2011 Elsevier B.V. All rights reserved.)

The Envelope glycoprotein (E-protein) of Japanese encephalitis virus (JEV) is the major structural component on the virion surface and is a primary target for the host immune system. Two monoclonal antibodies (MAbs) NHA-I (IgG2b) and NHA-II (IgM) against JEV (Indian strain 733913) were earlier developed in the authors' laboratory and found to be cross-reactive to nuclear histones. However, the epitope specificity of these MAbs has remained unknown. The present study was carried out to delineate the epitopes recognised by these MAbs on the E-protein of JEV strain 733913. The variable regions of the NHA-I and NHA-II were sequenced and the tertiary structures predicted. Molecular docking of the MAbs with the structural model of the JEV E-protein demonstrated that NHA-I binds to a predicted antigenic determinant (residue position 18-33) in domain-I. To understand the epitope specificity and check for possible cross-reactivity of these MAbs, comparative analysis of interactions with the known crystallographic structure of the West Nile virus (WNV) E-protein was also carried out. The studies predicted a differential binding of NHA-I but not of NHA-II between JEV and WNV. Mutagenesis studies could help analyse the specificity of NHA-I. The NHA-II appears to be cross-reactive as it docked in the groove region between domains I and III of both the JEV and WNV E-proteins. In laboratory assays, namely, ELISA and immunofluorescence assay both the MAbs reacted equally with JEV while the NHA-I did not show any reactivity with WNV. In silico results were thus validated by laboratory experiments. The present study would help in better understanding of virus-host interactions at the molecular level, and also be useful for the future design of vaccines as well as peptide based diagnostics., (Copyright © 2011 Elsevier B.V. All rights reserved.)