[Show abstract][Hide abstract] ABSTRACT: Intradermal (ID) vaccination with modern cell culture rabies vaccines is a means to significantly reduce the cost of post-exposure prophylaxis as compared to intramuscular vaccination. In this study we evaluated the efficacy, immunogenicity and tolerability of PCECV and PVRV administered ID in doses of 0.1 mL per site according to the 2-site Thai Red Cross (TRC) regimen. Patients with WHO category III exposure to suspect or laboratory proven rabid animals were administered either PCECV (n=58) or PVRV (n=52) ID at a dose of 0.1 mL per site on days 0, 3 and 7 and at one site on days 30 and 90. Serum samples were withdrawn on days 0, 14, 30, 90, and 180 and rabies virus neutralizing antibody (RVNA) titers were determined by rapid fluorescent focus inhibition test (RFFIT). Patients who were exposed to laboratory confirmed rabid animals were followed up for one year after exposure. All 110 patients developed RVNA titers above 0.5 IU/mL by day 14. Adequate titers >0.5 IU/mL were maintained up to day 180. Both vaccines induced equivalent RVNA titers at all time points and were well tolerated. Five subjects who were bitten by laboratory confirmed rabid dogs were alive and healthy one year after exposure. As demonstrated, PCECV and PVRV are both immunogenic, efficacious and well tolerated when administered in the TRC post-exposure prophylaxis regimen in ID doses of 0.1 mL as recommended by WHO guidelines. The use of PCECV in this regimen may prove more economical in developing countries like India.
Human vaccines 10/2014; 2(5):200-204. DOI:10.4161/hv.2.5.3197 · 3.64 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Purpose: Opportunistic viral infections are one of the major causes of morbidity and mortality in HIV infection and their molecular detection in the whole blood could be a useful diagnostic tool. Objective: The frequency of opportunistic DNA virus infections among HIV-1-infected individuals using multiplex real-time PCR assays was studied. Materials and Methods: The subjects were in two groups; group 1: Having CD4 counts <100 cells/µl (n = 118) and the group 2: counts >350 cells/µl (n = 173). Individuals were classified by WHO clinical staging system. Samples from 70 healthy individuals were tested as controls. In-house qualitative multiplex real-time PCR was standardised and whole blood samples from 291 were tested, followed by quantitative real-time PCR for positives. In a proportion of samples genotypes of Epstein-Barr virus (EBV) and CMV were determined. Results: The two major viral infections observed were EBV and CMV. The univariate analysis of CMV load showed significant association with cryptococcal meningitis, oral hairy leukoplakia (OHL), CMV retinitis, CD4 counts and WHO staging (P < 0.05) while the multivariate analysis showed an association with OHL (P = 0.02) and WHO staging (P = 0.05). Univariate analysis showed an association of EBV load with CD4 counts and WHO staging (P < 0.05) and multivariate analysis had association only with CD4 counts. The CMV load was significantly associated with elevated SGPT and SGOT level (P < 0.05) while the EBV had only with SGOT. Conclusion: This study showed an association of EBV and CMV load with CD4+ T cell counts, WHO staging and elevated liver enzymes. These viral infections can accelerate HIV disease and multiplex real-time PCR can be used for the early detection. Genotype 1 and 2 of EBV and genotype gB1 and gB2 of CMV were the prevalent in the HIV-1 subtype C-infected south Indians.
Indian Journal of Medical Microbiology 07/2014; 32(3):261-9. DOI:10.4103/0255-0857.136558 · 1.04 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Abstract Very little information is available on the role of innate immune mechanisms in Japanese encephalitis virus (JEV) infection. This study was designed to investigate the role of all Pattern Recognition Receptors (PRRs) in JEV infection in a mouse neuronal cell line in comparison to events that occur in vivo, using JEV infected suckling and adult mice. Analysis of mRNA expression was carried out using RT-PCR for detection of PRR genes and their downstream pathway genes, while a PCR array technique was used to examine the complete transcription analysis. Amongst the various innate immune receptors, TLR3 gene exhibited differential expression in JEV-infected Neuro2a, in suckling mice and adult mouse brain cells but not in uninfected control cells. The downstream events of TLR3 were confirmed by increased mRNA expression of IRF3 and interferon-β in JEV-infected Neuro2a cells and suckling mice brain tissue. To confirm the functional significance of this observation, TLR3 gene silencing experiments were carried using specific siRNA in Neuro2a cells. The results revealed a significant enhancement of JEV replication in TLR3 gene silenced JEV-infected Neuro2a cells, thereby suggesting that TLR3 serves a protective role against JEV. The expression levels of other PRRs varied. JEV-infected adult mice showed significant upregulation of TLR2 and MDA5 as compared to JEV-infected suckling mice and Neuro2a cells. In addition, upregulation of Myd88 and IRF7 was also noted in adult mice. These observations, coupled with the fact that adult mice infected with JEV exhibited longer survival rates, suggests that the host antiviral TLR2 response seen in adult mice was eventually countered by the virus by using MDA5 receptor. Our findings suggest that different PRRs appear to be involved in JEV infection in Neuro2a cells and brains of suckling and adult mice.
[Show abstract][Hide abstract] ABSTRACT: Progressive multifocal leukoencephalopathy (PML) is a rare, subacute, demyelinating disease of the central nervous system caused by JC virus. Studies of PML from HIV Clade C prevalent countries are scarce. We sought to study the clinical, neuroimaging, and pathological features of PML in HIV Clade C patients from India. This is a prospective cum retrospective study, conducted in a tertiary care Neurological referral center in India from Jan 2001 to May 2012. Diagnosis was considered "definite" (confirmed by histopathology or JCV PCR in CSF) or "probable" (confirmed by MRI brain). Fifty-five patients of PML were diagnosed between January 2001 and May 2012. Complete data was available in 38 patients [mean age 39 ± 8.9 years; duration of illness-82.1 ± 74.7 days). PML was prevalent in 2.8 % of the HIV cohort seen in our Institute. Hemiparesis was the commonest symptom (44.7 %), followed by ataxia (36.8 %). Definitive diagnosis was possible in 20 cases. Eighteen remained "probable" wherein MRI revealed multifocal, symmetric lesions, hypointense on T1, and hyperintense on T2/FLAIR. Stereotactic biopsy (n = 11) revealed demyelination, enlarged oligodendrocytes with intranuclear inclusions and astrocytosis. Immunohistochemistry revelaed the presence of JC viral antigen within oligodendroglial nuclei and astrocytic cytoplasm. No differences in clinical, radiological, or pathological features were evident from PML associated with HIV Clade B. Clinical suspicion of PML was entertained in only half of the patients. Hence, a high index of suspicion is essential for diagnosis. There are no significant differences between clinical, radiological, and pathological picture of PML between Indian and Western countries.
Journal of NeuroVirology 05/2013; DOI:10.1007/s13365-013-0168-8 · 3.32 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The frequencies of 10 opportunistic DNA viruses were determined by multiplex real-time PCR in paired cerebrospinal fluid (CSF) and brain tissue of HIV-infected individuals. In the CSF, viruses were detectable in 45/55 cases: JC virus (JCV) in 62%, Epstein-Barr virus (EBV) in 44%, cytomegalovirus (CMV) in 25%, varicella-zoster virus (VZV) in 3.6%, herpes simplex virus 1 (HSV-1) in 1.8%, and human herpesvirus 6 (HHV-6) in 1.8% of cases. A single virus was detectable in 20 cases, 19 cases had coinfection with two viruses, and 6 cases were positive for three viruses. JCV was detectable in the CSF of 62% of cases and in 42% of brain tissues, with higher loads in progressive multifocal leukoencephalopathy (PML) (P < 0.05).
[Show abstract][Hide abstract] ABSTRACT: BACKGROUND: Focal brain lesions (FBL) in HIV/AIDS frequently pose a diagnostic dilemma as the etiology varies from infective (tuberculoma, toxoplasmosis and tuberculous abscesses) to neoplastic lesions like lymphoma. For determining etiology, advanced neuroimaging techniques, serological and molecular biological tests have been evolved with varying sensitivities/specificities. Stereotactic biopsy (STB) of the lesions is reserved for lesions unresponsive to appropriate therapy. OBJECTIVE AND METHODS: In this study, the diagnostic yield of neuroimaging [cranial CT (n=25), MRI (n=24), and Th201/99Tc SPECT scan (n=18)] is compared with histopathological diagnosis obtained by STB (n=21) or autopsy (n=4) in 25 HIV-1 subtype C seropositive individuals with FBL identified by neuroimaging with special reference to cerebral toxoplasmosis in an eighteen month study period (2006-2007). RESULTS AND CONCLUSION: Cerebral toxoplasmosis was the most frequent cause of FBL (21/25, 84%), followed by one case each of tuberculoma, progressive multifocal leukoencephalopathy (PML), primary central nervous system lymphoma (PCNSL) and measles inclusion body encephalitis (MIBE), the last two diagnosed at autopsy. Of the 21 cases of cerebral toxoplasmosis, definitive diagnosis with histopathological confirmation was available in 14/21 (66.6%), with indirect evidence suggesting probable toxoplasmosis in seven, all of whom responded to antitoxoplasma therapy. CT and MRI had comparable specificities (75%), while MRI had marginally higher sensitivity (85% versus 80.9%) in detecting multiple lesions. The positive predictive value of both CT and MRI was identical (94.4%), suggesting that CT maybe a cost effective screening tool in resource restricted settings, for evaluating FBL. Sensitivity of 99Tc SPECT scan for diagnosing inflammatory lesions was 75% but failed to differentiate PCNSL from toxoplasmosis. This study is the first of its kind from India analyzing FBL with specific focus on cerebral toxoplasmosis in the setting of HIV-1 subtype C.
[Show abstract][Hide abstract] ABSTRACT: Nervous system lesions are characterized by the loss of neuronal numbers and types. The neurotrophic factor levels in an injured tissue reflect their potential for regeneration. This hypothesis was investigated in olfactory bulb (OB), where olfactory tract was surgically transected disrupting neuronal migration and turnover. The effects were followed with quantification of mitral cells and three neurotrophic factors mRNA levels for 6 weeks. The neuronal numbers decreased by 3rd- and 4th-week in transected OBs followed by their restoration, comparable with that of controls at 5th- and 6th-week. The endogenous levels of three neurotrophic factors - (brain derived neurotrophic factor, insulin growth factor-1 and fibroblast growth factor-2) using qPCR showed increase at 2nd-week by 136-, 8- and 2-fold respectively. Also, there was a significant increase in specific neurotrophic factors at 5th-week and 6th-weeks. The results propose a temporal link between deployment of neurotrophic factors and the plausible restorative events for mitral cell numbers in OB.
Indian journal of experimental biology 11/2012; 50(11):765-70. · 0.75 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: To investigate the drug to drug interaction of N-methylisatin-β-thiosemicarbazone (MIBT) derivative (SCH16) with ribavirin, mycophenolic acid and pentoxifylline against Japanese encephalitis virus in vitro. Our earlier studies have reported significant antiviral activity of these compounds against Japanese encephalitis virus in vitro and in vivo.
An in vitro drug to drug combination analysis was carried out to investigate whether or not the direct antiviral effect shown by the individual MIBT derivative could be effectively increased when lower concentrations of two compounds in combination were used. The results of this study showed that the combination of MIBT derivative (SCH16) with ribavirin or mycophenolic acid significantly enhanced the antiviral activity of SCH16 against JEV in vitro. In contrast, the combination of SCH16 and pentoxifylline resulted in antagonism.
The antiviral activity showed by SCH16 was enhanced in the presence of ribavirin and mycophenolic acid.
Studying the synergistic/additive interaction of the compounds in combination would help in lowering the effective concentration so as to overcome the concern of toxicity.
[Show abstract][Hide abstract] ABSTRACT: Despite an elaborate armamentarium to tackle microbes, emerging infectious diseases remain a crucial global challenge. Emerging infections can be defined as “infections that have newly appeared in a population or have existed previously but are rapidly increasing in incidence or geographic range.” Several factors like increase in international travel and trade, human encroachment on wild-life habitats, changes in agricultural practices and wild-life trade have contributed to the emergence of pathogens. Emergence/re-emergence of several viral infections has been reported from India in the past few decades; some of the important emerging viral infections are discussed in this review. They include infection due to Nipah, Hantaviruses, Chikungunya, Human Enterovirus-71, Influenza, Chandipura, Crimean Congo, SARS Coronavirus, Buffalopox, Dengue and Japanese Encephalitis viruses. Creating increased awareness and training of clinical microbiologists/virologists for identification of new/emerging pathogens, and prompt reporting and management of outbreaks is essential to tackle the threat posed by emerging/re-emerging infections.
Proceedings of the National Academy of Sciences, India - Section B: Biological Sciences 03/2012; 82(1). DOI:10.1007/s40011-011-0001-1 · 0.40 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: To investigate if two important epidemic viral encephalitis in children, Enterovirus 71 (EV71) encephalomyelitis and Japanese encephalitis (JE) whose clinical and pathological features may be nonspecific and overlapping, could be distinguished.
Tissue sections from the central nervous system of infected cases were examined by light microscopy, immunohistochemistry and in situ hybridization.
All 13 cases of EV71 encephalomyelitis collected from Asia and France invariably showed stereotyped distribution of inflammation in the spinal cord, brainstem, hypothalamus, cerebellar dentate nucleus and, to a lesser extent, cerebral cortex and meninges. Anterior pons, corpus striatum, thalamus, temporal lobe, hippocampus and cerebellar cortex were always uninflamed. In contrast, the eight JE cases studied showed inflammation involving most neuronal areas of the central nervous system, including the areas that were uninflamed in EV71 encephalomyelitis. Lesions in both infections were nonspecific, consisting of perivascular and parenchymal infiltration by inflammatory cells, oedematous/necrolytic areas, microglial nodules and neuronophagia. Viral inclusions were absent.
Immunohistochemistry and in situ hybridization assays were useful to identify the causative virus, localizing viral antigens and RNA, respectively, almost exclusively to neurones. The stereotyped distribution of inflammatory lesions in EV71 encephalomyelitis appears to be very useful to help distinguish it from JE.
[Show abstract][Hide abstract] ABSTRACT: Substantial evidence exists for HLA and other host genetic factors being determinants of susceptibility or resistance to infectious diseases. However, very little information is available on the role of host genetic factors in HIV-TB coinfection. Hence, a longitudinal study was undertaken to investigate HLA associations in a cohort of HIV seropositive individuals with and without TB in Bangalore, South India.
A cohort of 238 HIV seropositive subjects were typed for HLA-A, B, and DR by PCR-SSP and followed up for 5 years or till manifestation of Tuberculosis. HLA data of 682 HIV Negative healthy renal donors was used as control.
The ratio of males and females in HIV cohort was comparable (50.4% and 49.6%). But the incidence of TB was markedly lower in females (12.6%,) than males (25.6%). Further, HLA-B*57 frequency in HIV cohort was significantly higher among females without TB (21.6%, 19/88) than males (1.7%, 1/59); P = 0.0046; OR = 38. CD4 counts also were higher among females in this cohort.
This study suggests that HIV positive women with HLA-B*57 have less occurrence of TB as compared to males.
[Show abstract][Hide abstract] ABSTRACT: Japanese encephalitis virus (JEV) requires the presence of an inexplicable cellular receptor on the surface of the host cell for its entry into the cell. The JEV envelope (E) protein has been shown to play an important role in attachment to cells. By using a widely accepted technique, virus overlay protein binding assay (VOPBA), a protein molecule of approximately 60 kDa, identified as vimentin by matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI TOF), was recognized on porcine kidney (PS) cells as a possible receptor for JEV. Further, anti-vimentin monoclonal antibodies were able to block JEV entry into the PS cells. Additionally, co-immunoprecipitation assay confirmed that vimentin protein present on the PS cells interacts with the JEV-E protein. These observations indicate that vimentin serves as a putative receptor for JEV in porcine kidney cells.
Virus Research 06/2011; 160(1-2):404-8. DOI:10.1016/j.virusres.2011.06.001 · 2.83 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: To compare a conventional polymerase chain reaction (PCR) and real-time PCR for the detection of neurotropic DNA viruses.
A total of 147 cerebrospinal fluid (CSF) samples was collected from patients attending a tertiary care hospital in South India for a period from 2005 to 2008. All these samples were tested using a conventional multiplex/uniplex PCR and a real-time multiplex/uniplex PCR. This technique was used to detect a large number of herpes viruses responsible for central nervous system infections, including HSV-1, HSV-2, VZV, CMV and EBV and the polyoma virus JCV.
Overall, in the entire set of samples, the real-time PCR yielded 88 (59.9%) positives and conventional PCR had six (4.1%) positives.
Our results suggest that the real-time PCR assay was more sensitive compared with the conventional PCR. The advantage of real-time PCR is that it can be performed much faster than conventional PCR. Real-time PCR is less time-consuming, less labour-intensive and also reduces the chance of contamination as there is no post-amplification procedure. In the entire study population, the major viruses detected using real-time PCR were EBV (34%), HSV-2 (10.8%) and VZV (6.8%).
Indian journal of medical microbiology 04/2011; 29(2):102-9. DOI:10.4103/0255-0857.81777 · 1.04 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Japanese encephalitis is a major public health problem in several parts of Asia, particularly India, Nepal, Sri Lanka and Myanmar (Burma). Despite its public health implications, there are no effective antiviral drugs available.
The present study evaluated the effect of mycophenolic acid on Japanese encephalitis virus (JEV) using an in vitro cytopathic effect inhibition assay, plaque reduction assay and virus yield reduction assay, and its therapeutic potential was also assessed in vivo in a mouse model.
Analysis of the results obtained in the in vitro and in vivo experiments suggests that mycophenolic acid has significant antiviral activity against JEV, with an IC(50) of 3.1 μg/ml, a therapeutic index of 16 and a 75% protection against lethal challenge of JEV.
The study concludes that this compound significantly inhibited the replication of JEV in vitro and protected mice in vivo.
[Show abstract][Hide abstract] ABSTRACT: The recent emergence of the swine-origin influenza A H1N1 virus (S-OIV) poses a serious global health threat. Rapid detection and differentiation of S-OIV from seasonal influenza is crucial for patient management and control of the epidemics. A one-step, single-tube accelerated and quantitative S-OIV-specific H1 reverse transcription loop-mediated isothermal amplification (RTLAMP) assay for clinical diagnosis of S-OIV by targeting the H1 gene is reported in this article. A comparative evaluation of the H1-specific RTLAMP assay vis-à-vis the World Health Organization-approved real-time polymerase chain reaction (RTPCR), involving 239 acute-phase throat swab samples, demonstrated exceptionally higher sensitivity by picking up all of the 116 H1N1-positive cases and 36 additional positive cases among the negatives that were sequence-confirmed as S-OIV H1N1. None of the real-time RTPCR-positive samples were missed by the RTLAMP system. The comparative analysis revealed that S-OIV RTLAMP was up to tenfold more sensitive than the World Health Organization real-time RTPCR; it had a detection limit of 0.1 tissue culture infectious dosage of (50)/ml. One of the most attractive features of this isothermal gene amplification assay is that it seems to have an advantage in monitoring gene amplification by means of SYBR Green I dye-mediated naked-eye visualization within 30 minutes compared to 2 to 3 hours for a real-time reverse transcription polymerase chain reaction. This suggests that the RTLAMP assay is a valuable tool for rapid, real-time detection and quantification of S-OIV in acute-phase throat swab samples without requiring sophisticated equipment.
The Journal of molecular diagnostics: JMD 01/2011; 13(1):100-7. DOI:10.1016/j.jmoldx.2010.11.003 · 3.96 Impact Factor