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Biomedical subjects

G Sridharan

Publications and source records attributed to G Sridharan.

At least 19 recordsLinked to original sources
PubMed2006

Development and evaluation of a rapid IgM capture ELISA (JEV-Chex) for the diagnosis of Japanese encephalitis.

BACKGROUND: The IgM capture ELISA has been the most widely used diagnostic method for Japanese encephalitis. However, the lack of availability of validated commercial kits as well as the short shelf life of the kit reagents has limited the use of this technique to very few centres in Asia. OBJECTIVES: Development and evaluation of a rapid IgM capture ELISA (JEV Chex) in comparison to the conventional IgM capture ELISA. Produce key reagents such as cell culture derived JEV antigen and biotinylated monoclonal antibody which are stable at room temperature. STUDY DESIGN: The conventional IgM capture ELISA was modified to reduce the total assay time and two key reagents used in the assay JEV antigen and biotinylated anti-JEV monoclonal antibodies were rendered stable at room temperature using a special procedure. A multi-centric evaluation of this rapid ELISA was carried out using well characterized stored CSF and serum samples. Long term stability of the key reagents was also assessed over a period of 6 months. RESULTS: The rapid IgM capture ELISA developed by us showed complete concordance with the results obtained using the conventional ELISA at all the three centres where it was evaluated. In addition, the stability studies carried out with the inactivated cell culture antigen and the biotinylated monoclonal antibodies stored at room temperature for a period of 180 days revealed that both these reagents yielded consistent optical density values in the ELISA. CONCLUSIONS: The rapid ELISA format of the IgM capture ELISA (JEV-Chex) developed by us as well as the stability of reagents achieved by us in this study is what renders this rapid IgM capture ELISA very robust and user friendly since reagents can be stored at 4 degrees C by peripheral labs.

Aedes↗
PubMed2006

Total quality management in clinical virology laboratories.

The diagnostic laboratories in India are progressively promoting higher standards and are moving towards accreditation and international acceptance. Hence, the concept of "Quality" will need to be understood and implemented. Total quality management (TQM) in a laboratory is an integrated program involving all laboratory staff and management. TQM is a framework to operate and it is aiming for integration, consistency, increase in efficiency and a continuous drive for improvement. A well structured clinical virology service will include serology setup, cell culture facility and capacity for molecular diagnosis. The quality of results from the laboratory is significantly influenced by many pre-analytical and post-analytical factors which needed attention. The end goal of the TQM should be to provide the best care possible for the patient.

Accreditation↗
PubMed2005

Rapid identification of Burkholderia pseudomallei in blood culture supernatants by a coagglutination assay.

In total, 309 blood culture supernatants were tested for the presence of Burkholderia pseudomallei antigen using an in-house coagglutination test prepared by sensitising Cowan I staphylococcal cells with B. pseudomallei polyclonal antiserum. The coagglutination test gave a sensitivity, specificity, positive predictive value and negative predictive value of 100% in comparison with blood culture. A subset of 102 supernatants was also tested for B. pseudomallei antigen using a monoclonal antibody-based latex agglutination test. The sensitivity, specificity, positive predictive value and negative predictive values of this test were 100%, 90%, 75% and 100%, respectively.

Agglutination Tests↗
PubMed2005

Comparison of Microcapillary Cytometry Technology and Flow Cytometry for CD4+ and CD8+ T-Cell Estimation.

An alternative technology for the estimation of T cells based on a microcapillary technique (Guava Technologies, Hayward, CA) was compared to FACSCount (Becton Dickinson, San Jose, CA). Samples from 51 human immunodeficiency virus-infected and 21 healthy individuals were tested. The correlation (r) of the two systems for CD4(+) T cells was 0.994, and the coefficient of variation was 6.5%, establishing equable performance between the two technologies.

CD4-CD8 Ratio↗
PubMed2005

Human immunodeficiency virus infection and levels of Dehydroepiandrosterone sulfate in plasma among Indians.

The shift in cytokine profile during human immunodeficiency virus (HIV) disease progression is influenced by dehydroepiandrosterone sulfate (DHEAS) level. Radioimmunoassay was used to measure plasma DHEAS for 30 treatment-naïve HIV-infected and 30 uninfected individuals. There was a significant negative correlation of viral load with DHEAS level (P<0.05). Further studies of the use of DHEAS levels for monitoring HIV patients economically are warranted.

Adult↗
PubMed2005

Subtype & cytokine profiles of HIV infected individuals from south India.

BACKGROUND & OBJECTIVE: The global surveillance of human immunodeficiency virus (HIV) subtypes (clades) helps understand the global distribution and incidence of different HIV subtypes. As knowledge about subtypes circulating in an area is needed for developing a candidate vaccine, prevalence of the subtypes HIV-1 and HIV-2 were studied in south India. The profile of cytokines interleukin 10 (IL10) and interferon gamma (IFNgamma) in both types of infection were also analysed as these are considered indicators of disease progression. METHODS: Patients who belonged to the 4 south Indian States i.e. Tamil Nadu, Kerala, Karnataka and Andhra Pradesh were included. HIV-1 subtyping was carried out by the heteroduplex mobility analysis (HMA) while that of HIV-2 was done by direct sequencing. The quantitation of IFNgamma and IL-10 was carried out using commercial ELISA kits. RESULTS: Among the 82 HIV-1 infected individuals subtyped, 78 (95.1%) were subtype C while all 12 HIV-2 strains were subtype A. IL-10 concentration was significantly higher among HIV infected individuals compared to normal healthy controls. IFNgamma was significantly higher among symptomatic and AIDS groups compared to asymptomatic HIV-1 infected individuals. INTERPRETATION & CONCLUSION: HIV-1 subtype C and the HIV-2 subtype A are the major subtypes circulating in south India. The study showed a trend towards a shifting of the cytokine profile from Th1 to Th2/Th0 in HIV-1, HIV-2 infections, and HIV-1 and HIV-2 dual infected individuals as the disease progresses. This trend observed is not unlike that reported from the West, despite the difference in subtype profile.

Base Sequence↗
PubMed2005

Molecular epidemiology of HIV.

The phylogenic analysis of the nucleotide sequences of the env gene has enabled classification of HIV-1 into three groups. The group M of HIV-1 infection has been classified into 9 different genetic subtypes A-K, with E and I being classified as circulating recombinants forms (CRFs). The groups O and N are less frequently encountered in human infections. Presently group M of HIV-1 globally causes 99.6 per cent of all human infections. The epidemiological trends suggest that subtype C strains would dominate the HIV pandemic in the coming years. The geographic spread of subtype C strains is also very diverse with prevalence in Africa, Latin America and Asia. Data from India show a high prevalence of subtype C. In north and western India, 78.4 and 96 per cent of HIV-1 strains respectively were shown to be subtype C. Among female sex workers in Kolkata 95 per cent of the HIV-1 strains were subtype C. The south Indian subtype data are very similar to the data from the rest of India. The HIV-2 groups (subtypes) recognized are A-H. Unlike HIV-1, HIV-2 strains are predominantly found in Africa. The Indian HIV-2 strains identified till date are subtype A. This is also the predominant strain circulating in western African countries. This group (subtype) is estimated to cause 0.11 per cent of all HIV infections in humans.

Genotype↗
PubMed2005

A pilot study for serological evidence of hantavirus infection in human population in south India.

BACKGROUND AND OBJECTIVES: Hantaviruses are rodent-borne viruses of the family Bunyaviridae that have been identified as aetiological agents of two human diseases, haemorrhagic fever with renal syndrome (HFRS) and hantavirus pulmonary syndrome (HPS). There are no reports of hantavirus infections in humans from India, hence this pilot study was undertaken to provide the serological evidence of hantavirus infections in humans in south India. METHODS: Serum samples were obtained from individuals with acute febrile illness and from voluntary blood donors, majority of whom were from south India. Serum samples were tested for anti-hantavirus IgM using a commercial enzyme immunoassay (EIA). Samples found positive by the EIA were tested by an indirect immunofluorescence assay (IFA) using slides coated with Seoul virus (SEOV) infected cells as substrate. RESULTS: Of the 152 serum samples from individuals with pyrexic illness, 23 (14.7%) were positive for anti-hantavirus IgM by EIA. In contrast, only 5.7 per cent of healthy blood donors were positive by this assay. Eighteen of the 22 (82%) EIA-positive samples from patients were positive by the IFA assay. In contrast, only 2 of the 5 (40%) blood donor EIA positive samples were positive in the IFA assay. INTERPRETATION AND CONCLUSION: The finding of this study indicated the possible presence of hantavirus infections in the human population of India presenting both as asymptomatic and symptomatic infections. Further studies need to be done to confirm the findings on a larger sample using molecular techniques.

Fluorescent Antibody Technique, Indirect↗
PubMed2004

Detection of virulence attributes of Burkholderia pseudomallei.

BACKGROUND & OBJECTIVES: Melioidosis caused by Burkholderia pseudomallei is an emerging disease in India. This study examined the toxin activity of bacteria-free culture filtrate in three different cell lines (cytotoxic assay) and its effect on Caenorhabditis elegans (nematode toxicity assay). Endotoxic activity of the viable bacteria was also studied in C. elegans (co-culture killing assay). METHODS: For toxin studies, serial doubling dilutions of unheated, heated crude and ultra filtrate of bacteria-free culture supernatants of B. pseudomallei were tested in 96-well microtitre plate containing confluent mono layers of McCoy, Hep-2 and HeLa cell lines. For the effects on C. elegans, the worms were exposed to heated and unheated bacteria-free culture supernatants in 24-well microtitre plate for 24h and then transferred to OP50 Escherichia coli lawn culture. The endotoxic activity of the live bacterium was studied by feeding the worms in the lawn culture of B. pseudomallei. RESULTS: All the clinical isolates (n=38) produced cytotoxic changes in all the cell lines. No difference was observed in the cytotoxicity of unheated, heated and ultra-filtered culture supernatant. The septicaemic isolates were observed to produce cytotoxic changes in high dilutions (1:160) of culture filtrate. None of the unheated and heated crude filtrate had deleterious effect on C. elegans, while all the live bacteria were found to be lethal to the nematode. INTERPRETATION & CONCLUSION: The culture supernatants, though produced cytopathic effect in various tissue cultures, failed to have any deleterious effect on the worms. However, live bacteria were lethal to the worms B. pseudomallei. Use of C. elegans model to detect virulence attributes of B. pseudomallei is recommended as an alternative to tissue culture methods as this can be carried out in laboratories where a tissue culture set up is not available.

Animals↗
PubMed2004

Cytotoxin testing of environmental Aeromonas spp. in Vero cell culture.

BACKGROUND & OBJECTIVES: Aeromonas spp. are known to cause a variety of infections in humans and this organism has been isolated from a variety of sources including environmental sources. The pathogenicity of the environmental isolates in and around Vellore has not been studied. This study was conducted to determine the cytotoxicity of the Aeromonas spp. isolated from water bodies, soil sediments, plankton and sewers in and around Vellore. METHODS: Aeromonas spp. isolated from environmental sources were identified by standard procedures. Representative isolates of Aeromonas spp. were tested for cell free cytotoxic factor in tissue culture system. Undiluted and diluted cell free filtrates of isolates and known toxigenic and non-toxigenic bacteria were added to Vero cell monolayer in microtitre plates. After appropriate incubation in 5 per cent CO2 atmosphere, the microtitre plate was examined for cytopathic effect. Cell detachment and shrinkage of Vero cells were recorded as toxic changes. RESULTS: All 36 environmental isolates demonstrated cytopathic effect of which 41.7, 50 and 8.3 per cent belonged to A. hydrophila, A. veronii biotype sobria and A. caviae respectively. INTERPRETATION & CONCLUSION: The results demonstrated the presence of potentially pathogenic environmental aeromonads in and around Vellore and they produced cytotoxin.

Aeromonas↗
PubMed2003

Invasive properties of south Indian strains of Streptococcus pyogenes in a HEp-2 cell model.

The objective of this study was to consider the invasive properties of Streptococcus pyogenes in human pharyngeal epithelial cells, and to correlate these with their clinical significance. Clinical isolates of S. pyogenes obtained from blood cultures over a period of 10 years, and throat and skin isolates from a community-based study, were used in this investigation. The S. pyogenes isolates were inoculated in HEp-2 cells and subsequently treated with antibiotics to kill the extracellular bacteria. The cells were then lyzed, and a colony count was carried out to check for invasion. The throat and skin isolates had 45.7%, 25.7% and 28.5% of low, intermediate and high invasion efficiencies, respectively, while 80%, 8.6% and 11.4% of the blood isolates had low, intermediate and high invasion efficiencies. We concluded that the throat and the skin isolates from superficial infections were more invasive than the blood isolates, which is an interesting and paradoxical feature.

Colony Count, Microbial↗
PubMed2003

Prevalence of HIV infection in psychiatric patients attending a general hospital in Tamil Nadu, south India.

An anonymous, unlinked study was conducted to detect antibodies to HIV-1 or HIV-2 infections in 1,160 consecutive, newly registered, adult psychiatric outpatients at a general hospital in South India to determine whether psychiatric patients presenting to general hospitals are a population at high risk for HIV infection and should be routinely screened. The seroprevalence of HIV infection (12/1160; 1.03%; 95% CI = 0.4-1.6%) did not approximate rates expected of a high-risk group compared to the national (0.7%) or regional community (1.8%) prevalence. It did not differ significantly from HIV seroprevalence in non-psychiatric patients (233/35450; 0.7%; 95% CI = 0.57-0.74%) who were systematically screened (relative risk = 1.57; 95% CI = 0.88-2.80) during the same period, but was greater than the seroprevalence in healthy blood donors (0.5%; p = 0.02; relative risk = 2.15 95% CI = 1.17-3.95). Non-psychiatric patients were also more likely to be HIV infected than blood donors (p = 0.02; relative risk = 1.37; 95% CI = 1.05-1.78). These findings have implications for HIV testing policies among psychiatric and non-psychiatric patients presenting to general hospitals in India.

Adolescent↗
PubMed2003

Seroepidemiological study of herpes simplex virus types 1 and 2 in Brazil, Estonia, India, Morocco, and Sri Lanka.

BACKGROUND: The association between herpes simplex virus type 2 (HSV-2) and human immunodeficiency virus (HIV) and the development of HSV vaccines have increased interest in the study of HSV epidemiology. OBJECTIVES: To estimate the age and sex specific seroprevalence of HSV-1 and HSV-2 infections in selected populations in Brazil, Estonia, India, Morocco, and Sri Lanka. METHODS: Serum samples were collected from various populations including children, antenatal clinic attenders, blood donors, hospital inpatients, and HIV sentinel surveillance groups. STD clinic attenders were enrolled in Sri Lanka, male military personnel in Morocco. Sera were tested using a common algorithm by type specific HSV-1 and HSV-2 antibody assay. RESULTS: 13,986 samples were tested, 45.0% from adult females, 32.7% from adult males, and 22.3% from children. The prevalence of HSV-1 varied by site ranging from 78.5%-93.6% in adult males and from 75.5%-97.8% in adult females. In all countries HSV-1 seroprevalence increased significantly with age (p<0.001) in both men and women. The prevalence of HSV-2 infection varied between sites. Brazil had the highest age specific rates of infection for both men and women, followed by Sri Lanka for men and Estonia for women, the lowest rates being found in Estonia for men and India for women. In all countries, HSV-2 seroprevalence increased significantly with age (p<0.01) and adult females had higher rates of infection than adult males by age of infection. CONCLUSIONS: HSV-1 and HSV-2 seroprevalence was consistently higher in women than men, particularly for HSV-2. Population based data on HSV-1 and HSV-2 will be useful for designing potential HSV-2 vaccination strategies and for focusing prevention efforts for HSV-1 and HSV-2 infection.

Adolescent↗
PubMed2002

Comparison of IgM capture ELISA with a commercial rapid immunochromatographic card test & IgM microwell ELISA for the detection of antibodies to dengue viruses.

BACKGROUND & OBJECTIVES: There is a need for a reliable test for the early diagnosis of dengue fever (DF), which is now active in many parts of India especially in the post monsoon months. This study evaluated two commercial tests with an assay available from a national laboratory in India to obtain information and to make a comparison among the three tests as to which will be the most suited for the detection of IgM antibodies to dengue virus. METHODS: An IgM capture ELISA (National Institute of Virology, Pune, India) was compared with two commercial tests, the PanBio Rapid Immunochromatographic Card Test (Brisbane, Australia) and the PanBio Microwell IgM ELISA for the detection of IgM antibodies to dengue virus. We tested 154 samples from individuals with febrile illnesses having DF--like symptoms. RESULTS: The NIV IgM capture ELISA (MAC-ELISA) showed a high positivity rate (38.9%) as compared to the PanBio Rapid (22.7%) and the PanBio IgM ELISA (20.7%). The true prevalence of disease, sensitivity and specificity of the three tests were estimated using 2LC latent class models using expectation-maximization (EM) algorithm. The NIV MAC-ELISA showed a high sensitivity (96%) as compared to PanBio Rapid (73%) and PanBio IgM ELISA (72%). A subset of 68 samples (of the 154 tested) were analyzed by the NIV MAC-ELISA for IgM antibodies additionally to Japanese encephalitis (JE) and West Nile (WN) of which 31 samples showed positivity to either one, two or all three flaviviruses. Out of the 8 samples which were positive for dengue IgM alone by the NIV MAC-ELISA, only 2 (25%) each were picked up by the other 2 tests. While out of 7 samples positive for IgM to all three flaviviruses IgM by the NIV MAC-ELISA, 5 (71%) were picked up by the other 2 tests. Of the 5 that were picked up by the PanBio tests, 3 had the highest absorbance values to WN by the NIV MAC-ELISA, indicating cross reactivity by PanBio tests. INTERPRETATION & CONCLUSION: The MAC-ELISA though a 3 day procedure, would be a valuable screening test for the detection of IgM to dengue in routine diagnostic laboratories because of its high sensitivity and specificity rates. The test uses specific viral antigens to detect IgM antibodies not only to dengue but also to JE and West Nile as a result of which IgM antibodies to all the 3 commonly encountered flaviviruses can be detected in a single run. It also has the advantage in that depending on the strength of the antibody units obtained to a specific flaviviral antigen, presumptive diagnosis as to which particular arboviral infection has occurred can be made in conjunction with clinical presentation.

Antibodies, Viral↗
PubMed2002

A pilot study of seroprevalence of influenza virus type A in Vellore, south India.

BACKGROUND & OBJECTIVES: Influenza virus type A is active in many regions of the world. However, information from many parts of India is sparse. Hence we carried out a serological study on the prevalence of antibodies to influenza virus type A in Vellore, south India. METHODS: Antibodies to influenza virus type A (H1N1 and H3N2) were detected using a haemagglutination inhibition (HI) test. A total of 186 individuals comprising healthy blood donors and laboratory workers were tested. RESULTS: The titres ranged from < 20-2580, with a high geometric mean titre (GMT) of 200 for H3N2 and < 20-1280 with a GMT of 74 for H1N1 serotype of influenza virus type A. Among the 186 serum samples tested, 175 (94.04%) were positive for H1N1 and 185 (99.5%) for H3N2 with a titre > 20. INTERPRETATION & CONCLUSION: The prevalence of elevated level of antibodies in the individuals indicates a high exposure to the influenza A virus in our population with a seropositive status in 99.5 per cent of the individuals tested. Virus surveillance needs to be instituted in different parts of the country to monitor the activity of these viruses.

Antibodies, Viral↗
PubMed2001

A peptide enzyme linked immunosorbent assay (ELISA) for the detection of human immunodeficiency virus type-2 (HIV-2) antibodies: an evaluation on polymerase chain reaction (PCR) confirmed samples.

BACKGROUND: HIV-1 and HIV-2 infections differ in prognosis, and may also require different prevention and/or treatment approaches. Thus, estimating the true prevalence of HIV-1 and HIV-2 infections, as well as co-infections, is a critical step in controlling the disease. There are a few commercial ELISA and immunoblot kits, which can differentiate between HIV-1 and HIV-2 infections. However, some of these assays overestimate the prevalence of dual infection. Hence, it is necessary to develop assays capable of discriminating between the two infections. OBJECTIVES: To develop a synthetic HIV-2 env based peptide ELISA for the detection of HIV-2 specific antibodies and evaluate its performance on samples from HIV positive individuals previously tested by HIV-1 and HIV-2 PCR and HIV seronegative individuals. STUDY DESIGN: We studied 45 HIV seronegative and 63 HIV infected individuals, including 30 HIV-1 PCR and immunoblot positives, 19 HIV-2 PCR and immunoblot positives, five HIV-1 and two PCR and dual immunoblot positives, two PCR negative but positive for HIV-2 by immunoblot and seven dual immunoblot positives who were only positive for HIV-1 by PCR. RESULTS: All 24 HIV-2 PCR positive samples tested were positive by the peptide assay. Among 30 HIV-1 PCR and immunoblot positive samples, only one (3.3%) showed an absorbance value above the cut off level. The seven dual positive samples by immunoblot (only positive for HIV-1 by PCR) were negative by the HIV-2 peptide ELISA. There was a 100% concordance between HIV-2 PCR and peptide ELISA. The sensitivity, specificity, and the likelihood ratio for the peptide ELISA were 100,94.9, and 19.5, respectively when compared against the PCR findings. CONCLUSIONS: This ELISA, using a specific immunodominant epitope (11 amino acids) from the transmembrane (gp36) portion of the HIV-2 envelope glycoprotein showed a high concordance with PCR findings. This can be considered as a highly sensitive, specific and economically feasible assay for the discrimination of HIV-1 and HIV-2, and may serve as an alternative to HIV-2 PCR in epidemiological studies.

Enzyme-Linked Immunosorbent Assay↗