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An epidemic of Chikungunya fever of unprecedented magnitude occurred in many parts of India in early 2006 after an interval of 33 years, and there has been a resurgence in some parts of South India since June 2007. The article highlights clinical manifestations of infection and various molecular tests that were used for diagnoses of Chikungunya virus infection. Of particular interest is the real-time loop-mediated isothermal amplification (RT LAMP) assay, which is rapid and cost-effective and can be adopted at ill-equipped laboratories. Clinical symptoms were characterized by a triad of fever, rash, and severe rheumatic manifestations. RT LAMP identified 20 additional Chikungunya virus—positive cases, compared with reverse-transcriptase polymerase chain reaction. Chikungunya virus was isolated from 20 randomly selected samples. Genotyping of the virus isolates revealed that the East Central South African genotype of Chikungunya virus was the etiologic agent of this epidemic. Molecular diagnosis is an important tool to identify such new vectorborne viral illnesses.

Chikungunya fever is an important arboviral infection prevalent throughout Africa and Southeast Asia. Recently, in 2006, it has reemerged in many parts of India, affecting more than a million persons. A detail serological, virological, and molecular investigation of this unprecedented outbreak was carried out by collecting and studying 540 samples from all the affected regions of India during this epidemic. An in-depth investigation revealed the presence of anti-Chikungunya antibodies in 68% of the samples and genomic RNA in 49% of them. In addition 32 Chikungunya viruses were isolated from 45 representative polymerase chain reaction-positive samples. The nucleotide sequences of partial E1 gene of 25 representative Chikungunya viruses were deciphered. The sequence analysis indicated that all the isolates of this epidemic belonged to the new Indian Ocean island clade of East Central South (ECS) African genotype. This study conclusively proved the genotype shift from Asian to ECS African as the major factor in the reemergence of Chikungunya in an unprecedented outbreak in India after a gap of 32 years.

Dengue, Japanese encephalitis, West Nile encephalitis, yellow fever are the common flaviviral diseases associated with high morbidity and mortality. The initial symptoms of most of the flaviviral infections are similar to each other as well as to some other viral diseases. Making clinical diagnosis, therefore, becomes a challenging task for the clinician. Several studies have been reported on using detection of serum antibodies against flavivirus for the diagnosis of specific flaviviral disease; no field-based pan-flavi virus detection system is available, which can be used in low-endemicity areas for differentiation of flaviviral disease from other viral diseases.To identify a conserved amino acid sequence among all flaviviruses and evaluate the antibody formed against the conserved peptide to develop pan-flavivirus detection system.In the present study we have compared amino acid sequences of several flaviviruses and identified a conserved amino acid sequence lying in domain II of envelope protein.A peptide having the conserved amino acid sequence was used to generate polyclonal antibodies and these antibodies were used to detect several flaviviruses. Anti-peptide polyclonal antibodies selectively recognized flaviviruses and did not detect non-flaviviruses. Anti-peptide antibodies detected presence of virus in serum spiked with pure virus preparations.The study offers a rationale for development of pan-flavivirus capture assay suitable for low endemic areas.

An outbreak of febrile illness occurred between September to November 2001 in Gwalior, Madhya Pradesh affecting individuals mostly in the age group30 yr. A total of 312 febrile indoor patients suspected to have dengue infection were investigated.The investigation included examination of blood samples from patients for dengue specific IgM and IgG antibodies, isolation of virus in suckling mouse pups and in C(6/36) cell line followed by confirmation and typing through reverse transcriptase-PCR and nested PCR.The serological analysis of the 312 samples indicated 65 per cent positivity of which 21 per cent are of recent infection as indicated by the presence of IgM antibody and 78 per cent are found to be secondary in nature by showing the presence of IgG and/or IgM antibodies. The RT-PCR analysis of patients' sera employing dengue virus group specific conserved amplimer confirmed the etiological agent as dengue complex by showing the characteristic 511 bp amplicons. None of the antibody positive samples were found to be positive by RT-PCR. A total of 13 (6%) samples positive by RT-PCR, were processed for virus isolation in mouse pups and in C(6/36) cells. Of these 9 samples (80%) were confirmed positive for virus isolation as identified by RT-PCR.The typing of isolates by nested PCR employing serotype specific amplimer revealed 119 bp amplicon characteristic of dengue virus type-2 and thus confirming the outbreak attributed to dengue virus type-2.

Here we report standardization of a dipstick enzyme-linked immunosorbent assay (ELISA, Dipstick ELISA) and its comparative evaluation with a commercial Rapid PanBio Immunochromatographic test (IC test) for detection of Dengue (DEN) virus-specific IgM and IgG antibodies in patient sera. Among crude and purified viral antigens prepared from mouse brains or cell cultures, a DEN virus type 2 antigen purified from cell cultures by sucrose density gradient centrifugation was found superior in terms of the signal/ noise (S/N) ratio in the assay system. The sensitivity of detection of the virus by specific IgM antibody was improved by removal of IgG from patient sera prior to testing. The evaluation of the Dipstick ELISA by use of 156 serum samples revealed an overall accordance of 96% and 93% with the IC test in detection of IgM antibodies to DEN viruses (IgM antibodies) and IgG antibodies to DEN viruses (IgG antibodies), respectively. The sensitivity of the Dipstick ELISA and the IC test with reference to the mu-capture ELISA was 83% and 87%, respectively, with a specificity of 98% in both cases. The sensitivity of the Dipstick ELISA with reference to the IC test in detecting IgM and IgG antibodies was 84% and 94%, respectively, and the specificity of the Dipstick ELISA was 98% and 92%, respectively.

A strain of Clostridium botulinum type E has been isolated from soil samples of Gwalior, India. The isolated strain shows curved vegetative cells with oval, bulging, and terminal spores. The production of toxin was detected by immunodiffusion test, symptomatic death of mice and mouse protection assay with trivalent antitoxin (A+B+E). The culture supernatant gave 10(3) MLD (minimum lethal dose) per ml without any protease treatment. Group specific and serotype specific primers amplified the DNA fragments of 260 bp and 445 bp, respectively, indicating Clostridium botulinum type 'E.'

Efficacy of two colorimetric assays, viz. MTT (3-4,5-dimethylthiazol-2-(yl-2,5-diphenyl tetrazolium bromide) and neutral red (NR) assays, performed by integrating them to micro culture virus titration (MCVT), was compared with the conventional MCVT method in terms of percentages of infectivity and 50% infectivity end points by employing Polio virus type-3 and Dengue virus type 4 as the candidate viruses. The results suggested that MTT assay has an edge over NR assay as well as conventional MCVT method. For the first time, NR assay has been successfully employed for the determination of virus infectivity titre.

Caprinised rinderpest virus GTV strain (GTV) was adapted to grow in Vero cells (vGTV) by polyethylene glycol-mediated fusion of infected goat spleenocytes with non-infected Vero cells. The usual methods of infection of cell culture, i.e. virus adsorption or co-cultivation, were not successful. vGTV-induced cytopathic changes in Vero cells were similar to those reported for rinderpest virus. Virus titers increased with the passage number but the virulence for goats decreased. Immunoblot analysis did not reveal any difference between vGTV and vRBOK, the tissue culture rinderprest virus RBOK strain adapted to Vero cells.