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Background In the situation where COVID-19 pandemic has placed unprecedented demands and pressure on the health care system, we wanted to analyze how the medical microbiologists of our country were affected. Was it actually an opportunity to showcase the specialty or was it a doom? A debate was organized as a key session in the national e-conference of the Indian Association of Medical Microbiologists, held on 10 December 2020. Objectives The objective of the debate was to examine and analyze the various positive as well as negative impacts of COVID-19 on the discipline of the medical microbiology of our country. Content Before the debate a voting session was conducted to assess the opinion of the audience followed by a very interesting debate where both the speakers presented their view points. The points in favor of the discipline were, mainly up-gradation of the specialty of microbiology in terms of learning, skill development, infrastructure, networking & research opportunities related to COVID-19. While the main points against were, nerve wracking work load without much acknowledgement, performance pressure from hospital administration to maintain rapid turnaround time, and a forceful neglect of all other infectious diseases like tuberculosis and antimicrobial resistance which were the key battle fields of the medical microbiologists. Postgraduate & even undergraduate training programs got completely derailed to their disadvantage. By the end of the debate, it was concluded that COVID-19 was neither a boon nor a bane to the microbiologists. A balanced approach to the problem in hand is required without ignoring the pre-existing infectious diseases in our country. The post debate voting swayed the audience considerably for it to be a bane & the faculty debating for boon had a huge margin to begin with but finally won with a whisker indicating the intensity of the debate.

Background Immune system aberrations have been postulated to play a role in the pathophysiology of Obsessive-compulsive disorder (OCD). This study was aimed to examine the profile of immune cell subsets in peripheral blood of un-medicated OCD patients. Method Thirteen drug-naive/free OCD patients and twenty-six age & sex matched healthy controls were recruited. Immunophenotyping was carried out by staining the whole blood specimens with fluorescent monoclonal antibodies against the cell surface markers such as CD45, CD3, CD16, CD56, CD8, CD4, CD28, CD25 and CD127, followed by data acquisition on BD FACSVerse™ flow cytometer. The proportions of CD4 and CD8 T cells; T regulatory (Tregs), Natural Killer (NK) cells and NK-T cells were compared between patients with OCD and healthy control subjects. Results Significantly reduced percentage of T regulatory (Treg) cells was observed in individuals with OCD compared to healthy control subjects [1.0 ± 0.7 vs. 1.9 ± 1.4; p = 0.03, r = 0.33]. Conclusion Treg cells play a crucial role in regulating the immune response, especially by suppressing the functional activities of T cells. In this study, decreased population of Treg cells essentially indicates a dysregulated T cell and/or T cell mediated immune activation in drug-naive OCD patients. This preliminary observation might form the basis of further studies examining the immuno-inflammatory/autoimmune origin of OCD.

The immune hypothesis of schizophrenia has gained significant popularity in recent years in schizophrenia research. Evidence suggests that the peripheral immune system communicates with central nervous system and the effect propagates through microglial and lymphocyte crosstalk, especially during neuro-inflammation. Although, there is previous literature indicating changes in lymphocyte population in schizophrenia, detailed studies with respect to T and B cells are scarce. Mucosal associated invariant T (MAIT) cells are functionally associated with the gut microbiome. The gut microbiome has been implicated in the pathogenesis of schizophrenia. However, there is no information on the frequency of MAIT cells in schizophrenia. Hence, we investigated changes in proportions of T cells, B cells and MAIT cells in peripheral blood mononuclear cells derived from antipsychotic-free patients with schizophrenia in comparison to healthy controls. In line with earlier reports, we noted perturbations in Th17 cells. This study for the first time reports changes in frequencies of MAIT cells in a homogenous population of antipsychotic-free patients with schizophrenia. These changes, though not common across all patients nevertheless point to the fact that inflammation is prevalent in a significant subset of schizophrenia cases.

Guillain-Barre syndrome (GBS) is the commonest post-infectious inflammatory peripheral neuropathy with undiscerned aetiology. The commonly reported antecedent infections implicated in India include Campylobacter jejuni, chikungunya, dengue, and Japanese encephalitis (JE). In this study from south India, we investigated the role of these four agents in triggering GBS. This case-control study was performed on 150 treatment-naive patients with GBS and 150 age and sex-matched controls from the same community. IgM immunoreactivity for C. jejuni, chikungunya, and dengue was detected by enzyme-linked immunosorbent assay (ELISA) in serum of patients with GBS and control subjects. Immunoreactivity against JE was detected in serum as well as cerebrospinal fluid (CSF) from patients (n = 150) and orthopaedic control (n = 45) subjects. The immunoreactivity against infections was compared between demyelinating and axonal subtypes of GBS. Overall, 119/150 patients with GBS had serological evidence of antecedent infection. Amongst those with evidence of antecedent infection, 24 (16%), 8 (5%), and 9 (6%) patients were exclusively immunoreactive to chikungunya, JE, and C. jejuni, respectively. In the remaining patients (78/119), immunoreactivity to multiple pathogens was noted. Immunoreactivity to C. jejuni infection was found in 32% of GBS patients compared to 2.7% controls (P

Chikungunya virus (CHIKV) is primarily transmitted by Aedes spp. mosquitoes. The present study investigated vector competence for CHIKV in Aedes aegypti and Aedes albopictus mosquitoes found in Madurai, South India. The role of receptor proteins on midguts contributing to permissiveness of CHIKV to Aedes spp. mosquitoes was also undertaken. Mosquitoes were orally infected with CHIKV DRDE-06. Infection of midguts and dissemination to heads was confirmed by immunofluorescence assay at different time points. A plaque assay was performed from mosquito homogenates at different time points to study CHIKV replication. Presence of putative CHIKV receptor proteins on mosquito midgut epithelial cells was detected by virus overlay protein binding assay (VOPBA). The identity of these proteins was established using mass spectrometry. CHIKV infection of Ae. aegypti and Ae. albopictus midguts and dissemination to heads was observed to be similar. A plaque assay performed with infected mosquito homogenates revealed that CHIKV replication dynamics was similar in Aedes sp. mosquitoes until 28 days post infection. VOPBA performed with mosquito midgut membrane proteins revealed that prohibitin could serve as a putative CHIKV receptor on Aedes mosquito midguts, whereas an absence of CHIKV binding protein/s on Culex quinquefasciatus midguts can partially explain the non-permissiveness of these mosquitoes to infection.

Brief AbstractWe analysed SARS-CoV-2 surveillance and contact tracing data from Karnataka, India up to 21 July 2020. We estimated metrics of infectiousness and the tendency for superspreading (overdispersion), and evaluated potential determinants of infectiousness and symptomaticity in COVID-19 cases. Among 956 cases confirmed to be forward-traced, 8.7% of index cases had 14.4% of contacts but caused 80% of all secondary cases, suggesting significant heterogeneity in individual-level transmissibility of SARS-CoV-2 which could not be explained by the degree of heterogeneity in underlying number of contacts. Secondary attack rate was 3.6% among 16715 close contacts. Transmission was higher when index case was aged >18 years, or was symptomatic (adjusted risk ratio, aRR 3.63), or was lab-confirmed ≥4 days after symptom onset (aRR 3.01). Probability of symptomatic infection increased with age, and symptomatic infectors were 8.16 times more likely to generate symptomatic secondaries. This could potentially cause a snowballing effect on infectiousness and clinical severity across transmission generations; further studies are suggested to confirm this. Mean serial interval was 5.4 days. Adding backward contact tracing and targeting control measures to curb super-spreading may be prudent. Due to low symptomaticity and infectivity, interventions aimed at children might have a relatively small impact on reducing transmission.Structured AbstractBackgroundIndia has experienced the second largest outbreak of COVID-19 globally, yet there is a paucity of studies analysing contact tracing data in the region. Such studies can elucidate essential transmission metrics which can help optimize disease control policies.MethodsWe analysed contact tracing data collected under the Integrated Disease Surveillance Programme from Karnataka, India between 9 March and 21 July 2020. We estimated metrics of disease transmission including the reproduction number (R), overdispersion (k), secondary attack rate (SAR), and serial interval. R and k were jointly estimated using a Bayesian Markov Chain Monte Carlo approach. We evaluated the effect of age and other factors on the risk of transmitting the infection, probability of asymptomatic infection, and mortality due to COVID-19.FindingsUp to 21 July, we found 111 index cases that crossed the super-spreading threshold of ≥8 secondary cases. R and k were most reliably estimated at R 0.75 (95% CI, 0.62-0.91) and k 0.12 (0.11-0.15) for confirmed traced cases (n=956); and R 0.91 (0.72-1.15) and k 0.22 (0.17-0.27) from the three largest clusters (n=394). Among 956 confirmed traced cases, 8.7% of index cases had 14.4% of contacts but caused 80% of all secondary cases. Among 16715 contacts, overall SAR was 3.6% (3.4-3.9) and symptomatic cases were more infectious than asymptomatic cases (SAR 7.7% vs 2.0%; aRR 3.63 [3.04-4.34]). As compared to infectors aged 19-44 years, children were less infectious (aRR 0.21 [0.07-0.66] for 0-5 years and 0.47 [0.32-0.68] for 6-18 years). Infectors who were confirmed ≥4 days after symptom onset were associated with higher infectiousness (aRR 3.01 [2.11-4.31]). Probability of symptomatic infection increased with age, and symptomatic infectors were 8.16 (3.29-20.24) times more likely to generate symptomatic secondaries. Serial interval had a mean of 5.4 (4.4-6.4) days with a Weibull distribution. Overall case fatality rate was 2.5% (2.4-2.7) which increased with age.ConclusionWe found significant heterogeneity in the individual-level transmissibility of SARS-CoV-2 which could not be explained by the degree of heterogeneity in the underlying number of contacts. To strengthen contact tracing in over-dispersed outbreaks, testing and tracing delays should be minimised, retrospective contact tracing should be considered, and contact tracing performance metrics should be utilised. Targeted measures to reduce potential superspreading events should be implemented. Interventions aimed at children might have a relatively small impact on reducing SARS-CoV-2 transmission owing to their low symptomaticity and infectivity. There is some evidence that symptomatic cases produce secondary cases that are more likely to be symptomatic themselves which may potentially cause a snowballing effect on infectiousness and clinical severity across transmission generations; further studies are needed to confirm this finding.FundingGiridhara R Babu is funded by an Intermediate Fellowship by the Wellcome Trust DBT India Alliance (Clinical and Public Health Research Fellowship); grant number: IA/CPHI/14/1/501499.

Growing evidence in the last decade suggest significant role of immune alterations in the pathogenesis of obsessive–compulsive disorder (OCD). Cytokines, mediators of inflammation, alter the neurotransmitter concentration and result in a hyposerotonergic and hyperglutamatergic state implicated in pathogenesis of OCD. However, only few studies have examined cytokine abnormalities in OCD with inconsistent results possibly due to confounding effects of medications and comorbid anxiety–depression. We examined 20 comorbidity free, drug free OCD patients and 20 age and sex matched healthy controls. Clinical severity was assessed using Yale Brown Obsessive Compulsive Scale, Hamilton anxiety rating scale, Hamilton depression rating scale and Clinical Global Impression. Levels of different cytokines, Interleukin (IL)-2, IL-4, IL-6, IL-10, Tumor necrosis factor (TNF)-α and Interferon (IFN)-γ were assessed using Cytometric Bead Array. OCD patients had significantly greater plasma levels of IL-2, IL-4, IL-6, IL-10 and TNF-α levels than controls but not IFN-γ. Reanalysis of data with only drug naive patients (excluding 4 drug free patients) did not alter the results. Presence of these abnormalities in drug-naive patients suggests the possible role of cytokines in the pathogenesis of OCD. Study findings have potential clinical utility in development of novel therapeutic options targeting cytokine aberrations in OCD.

The genus Flavivirus contains many mosquito-borne human pathogens of global epidemiological importance such as dengue virus, West Nile virus, and Zika virus, which has recently emerged at epidemic levels. Infections with these viruses result in divergent clinical outcomes ranging from asymptomatic to fatal. Myriad factors influence infection severity including exposure, immune status and pathogen/host genetics. Furthermore, pre-existing infection may skew immune pathways or divert immune resources. We profiled immune cells from dengue virus-infected individuals by multiparameter mass cytometry (CyTOF) to define functional status. Elevations in IFNβ were noted in acute patients across the majority of cell types and were statistically elevated in 31 of 36 cell subsets. We quantified response to in vitro (re)infection with dengue or Zika viruses and detected a striking pattern of upregulation of responses to Zika infection by innate cell types which was not noted in response to dengue virus. Significance was discovered by statistical analysis as well as a neural network-based clustering approach which identified unusual cell subsets overlooked by conventional manual gating. Of public health importance, patient cells showed significant enrichment of innate cell responses to Zika virus indicating an intact and robust anti-Zika response despite the concurrent dengue infection., Author summary Mosquitoes carry many globally important human pathogens including a family of related viruses: dengue virus, West Nile virus, Yellow Fever virus, and recently of critical significance, Zika virus. The Zika virus epidemic emerged very rapidly in the susceptible South American population and in many cases immune responses were unable to control the infection. Immune history is a key element of susceptibility or resistance to severe disease. We examined whether pre-existing infection would skew or divert immune resources and might play a role in the severity of Zika infection in the Americas. Using samples from dengue patients and healthy controls from India, we tested functional responses to Zika virus in the context of pre-existing dengue infection. We quantified frequency and functional status of 36 individual cell subsets in depth using advanced profiling techniques and a novel deep learning algorithm. We showed an intact response to new infection with Zika virus which was enriched for early innate immune pathways and robust even during existing dengue infection. Thus, our study suggests that concurrent dengue infection would not be expected to impair immune responses to new infection with Zika virus.

Inflammation is considered to be relevant in pathophysiology of schizophrenia. Existing literature indicates that controlling inflammation may be helpful in patient management. Procalcitonin (PCT) is an established marker of inflammation which has not been well studied in context with schizophrenia. The study recruited 34 schizophrenia patients free of antipsychotic treatment and 24 healthy controls without any signs of inflammation. Plasma C reactive protein was quantified using a high sensitivity turbidimetric assay. Plasma PCT levels was estimated by sandwich ELISA. The study ruled out autoimmune antibodies by ANA and RF tests which exclude confounding factors contributing to inflammation. The data shows a subgroup of patients 17/34 (50%) have either elevated PCT or CRP levels. This study is the first to report PCT values in antipsychotic drug-free patients with schizophrenia.

Multiple studies have identified the presence of peripheral immune aberrations in subjects with Autism Spectrum Disorder (ASD). However, comprehensive assessment of these peripheral immune aberrations, in the cellular and systemic compartments, in a single group of subjects with ASD is lacking. We assessed proportions of various subsets of immune cells in peripheral blood (T helper cells, T regulatory cells, B cells, monocytes, Natural Killer cells, dendritic cells) by multi-parametric flow cytometry in 50 children with ASD and compared it with thirty healthy controls matched for age, gender, socio-economic status and body mass index. There were no significant differences noted in the proportion of T regulatory cells, B cells, monocytes and Natural Killer cells, between ASD subjects and controls. On the contrary, the proportion of activated Th17 and myeloid dendritic cells were significantly higher in children with ASD. Based on these findings, group comparison of serum levels of Th17 cytokines (interleukin-6, interleukin-17A) was performed. Elevated serum levels of interleukin-6 and interleukin-17A in children with ASD corroborated our immunophenotyping findings. We did not find any significant differences among the pro-inflammatory (interleukin-1 beta), Th1 (interferon-gamma) and Th2 (interleukin-4) cytokines. This is the first evidence with concurrent findings from immunophenotyping and cytokine data demonstrating activation of the Th17 pathway in subjects with ASD. This finding assumes significance in the light of recent maternal immune activation mouse model study that has highlighted the role of Th17 pathway in the pathophysiology of ASD. Future longitudinal studies are needed to clarify the role of this dysregulated immune pathway in the development of ASD.

Accelerated ageing processes are postulated to underlie schizophrenia pathogenesis. This postulate is supported by observations of reduced telomere length in schizophrenia patients. Hippocampus, one of the most important brain regions implicated in schizophrenia, is shown to atrophy at a faster rate in aging. In this study, telomere length (TL) was measured in 30 antipsychotic-naive/free schizophrenia patients and 60 healthy controls using quantitative PCR assay. Hippocampus volume was measured using voxel-based morphometry. Schizophrenia was associated with differential TL between sexes [Status × Sex; F(1,85) = 5.9, p = 0.017, η2 = 0.065]. Male schizophrenia patients had significantly lower relative TL than female patients [F(1,85) = 7.38, p = 0.008], while such sex difference was not observed in healthy controls [F(1,85) = 0.16, p = 0.69]. Schizophrenia patients showed a significant sex-by-telomere interaction with both right & left hippocampus, with male patients showing positive association of telomere length with volume, while female patients showed negative association. Telomere shortening and the positive association of telomere length with hippocampus volume was observed only in male patients with schizophrenia. Since correlational observations in this cross-sectional study does not necessarily support definitive causal relationship, further longitudinal studies examining hippocampus volume and telomere in schizophrenia patients are needed.

In schizophrenia, plasma cytokines abnormalities offer vital support for immunopathogenetic basis. However, most of the previous studies on plasma cytokines are confounded by examination of antipsychotic-treated schizophrenia patients. In this study, we examined a large sample of antipsychotic-naive/free schizophrenia patients (N = 75) in comparison with healthy controls (N = 102). Plasma cytokines (Interleukins ([IL] 2, 4, 6, 10, 17), Tumor necrosis factor [TNF] and Interferon gamma [IFN-g]) were assessed using cytometric bead array assay. Schizophrenia patients showed significantly greater levels of IL-6 and lower levels of IL-17 as well as IFN-g in comparison to healthy controls. However, after taking censoring into account and adjusting for potential confounders (sex, age, BMI and smoking), only IL-6 was found to be elevated in patients. Cytokine profile showed differential and pathogenetically relevant correlation with clinical symptoms. Together, these observations offer further support to immunological component in schizophrenia pathogenesis.

Disparate lines of evidence including epidemiological and case-control studies have increasingly implicated vitamin D in the pathogenesis of schizophrenia. Vitamin D deficiency can lead to dysfunction of the hippocampus--a brain region hypothesized to be critically involved in schizophrenia. In this study, we examined for potential association between serum vitamin D level and hippocampal gray matter volume in antipsychotic-naïve or antipsychotic-free schizophrenia patients (n = 35). Serum vitamin D level was estimated using 25-OH vitamin D immunoassay. Optimized voxel-based morphometry was used to analyze 3-Tesla magnetic resonance imaging (MRI) (1-mm slice thickness). Ninety-seven percent of the schizophrenia patients (n = 34) had sub-optimal levels of serum vitamin D (83%, deficiency; 14%, insufficiency). A significant positive correlation was seen between vitamin D and regional gray matter volume in the right hippocampus after controlling for age, years of education and total intracranial volume (Montreal Neurological Institute (MNI) coordinates: x = 35, y = -18, z = -8; t = 4.34 pFWE(Corrected) = 0.018). These observations support a potential role of vitamin D deficiency in mediating hippocampal volume deficits, possibly through neurotrophic, neuroimmunomodulatory and glutamatergic effects.

Reports of magnetic resonance imaging (MRI) findings in subacute sclerosing panencephalitis (SSPE) with clinical and electroencephalography (EEG) correlation are few. To describe the MRI features of SSPE and correlate with its clinical features and EEG. Clinical, MRI and EEG data of 33 patients (male/female 19/14; age: 11.4 ± 5.3 yr) were reviewed. All patients had raised IgG anti-measles antibody (≥ 1:625) in cerebrospinal fluid. Their clinical features were myoclonus (100%), seizures (42%), visual disturbances (18%), pyramidal signs (30%), extrapyramidal symptoms (27%), and cognitive impairment (79%) and behavioral changes (21%). EEGs were universally abnormal: diffuse slowing of background activity-all; periodic complexes-27 (symmetrical): High voltage delta waves (55%) and sharp & slow wave discharges (45%). MRI findings included – normal (9%), cerebral atrophy (78%), white matter (76%) and basal ganglia (6%) changes. Restricted diffusion in periventricular/subcortical region was noted in 4/23 patients (stage 2). Magnetic resonance spectroscopy showed decreased N-acetyl aspartate peaks and elevated choline peaks in 75% and lactate peak in 25% (stage 3) of patients. Correlation of background delta activity among patients with MRI signal abnormalities and those without any signal change was noted (P=0.02). There was significant correlation between visual deficits and bilateral parieto-occipital white matter signal changes (P=0.03). There was no other EEG-MRI or clinical-MRI correlation. MRI abnormalities in SSPE are diverse and may or may not reflect underlying pathology. Visual deficits were observed in patients with parieto-occipital white matter signal change. Restricted diffusion of signal abnormalities and lactate peak in magnetic resonance spectroscopy were uncommon and it may add to better understanding.

Chikungunya (CHIKV) is an emerging arboviral infection of public health concern in India contributing to widespread morbidity. The precise molecular events occurring early in the infection have not been well understood. Cytokines/chemokines are suspected to play a key role in its pathogenesis. Very few studies have correlated the plasma levels of cytokines/chemokines with diagnostic markers such as viral loads and presence of CHIKV IgM antibodies. Understanding these dynamics in the early phase of CHIKV infection is likely to provide an insight into the evolution of the immune response, identify biomarkers for assessing severity, and for development of newer therapeutic strategies. This study was therefore undertaken to estimate the levels of various cytokines/chemokines in plasma samples of patients infected with CHIKV and correlate to viral load and CHIKV IgM antibodies. Cytokine/chemokine levels and viral loads in plasma were measured using cytometric bead array and TaqMan real time PCR assay, respectively. The findings revealed that acute phase of CHIKV infection is characterized by predominant inflammatory responses mediated by IL-6, IL-8, IP-10, MCP-1, and MIG (P

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).

Healthy donors exposed to Japanese encephalitis (JE) virus show a CD8+ T cell response that cross reacts with other flaviviruses. Patients that recovered from JE show a CD4+ T cell response that targets structural proteins of JE virus., Japanese encephalitis (JE) virus (JEV) is an important cause of encephalitis in children of South and Southeast Asia. However, the majority of individuals exposed to JEV only develop mild symptoms associated with long-lasting adaptive immunity. The related flavivirus dengue virus (DENV) cocirculates in many JEV-endemic areas, and clinical data suggest cross-protection between DENV and JEV. To address the role of T cell responses in protection against JEV, we conducted the first full-breadth analysis of the human memory T cell response using a synthetic peptide library. Ex vivo interferon-γ (IFN-γ) responses to JEV in healthy JEV-exposed donors were mostly CD8+ and targeted nonstructural (NS) proteins, whereas IFN-γ responses in recovered JE patients were mostly CD4+ and targeted structural proteins and the secreted protein NS1. Among patients, a high quality, polyfunctional CD4+ T cell response was associated with complete recovery from JE. T cell responses from healthy donors showed a high degree of cross-reactivity to DENV that was less apparent in recovered JE patients despite equal exposure. These data reveal divergent functional CD4+ and CD8+ T cell responses linked to different clinical outcomes of JEV infection, associated with distinct targeting and broad flavivirus cross-reactivity including epitopes from DENV, West Nile, and Zika virus.

Chikungunya fever a re-emerging infection with expanding geographical boundaries, can mimic symptoms of other infections like dengue, malaria which makes the definitive diagnosis of the infection important. The present study compares the utility of four laboratory diagnostic methods viz. IgM capture ELISA, an in house reverse transcription PCR for the diagnosis of Chikungunya fever, TaqMan real-time PCR, and a one step reverse transcription-loop mediated isothermal amplification assay (RT-LAMP). Out of the 70 serum samples tested, 29 (41%) were positive for Chikungunya IgM antibody by ELISA and 50 (71%) samples were positive by one of the three molecular assays. CHIKV specific nucleic acid was detected in 33/70 (47%) by reverse transcription PCR, 46/70 (66%) by TaqMan real-time PCR, and 43/70 (62%) by RT-LAMP assay. A majority of the samples (62/70; 89%) were positive by at least one of the four assays used in the study. The molecular assays were more sensitive for diagnosis in the early stages of illness (2-5 days post onset) when antibodies were not detectable. In the later stages of illness, the IgM ELISA is a more sensitive diagnostic test. In conclusion we recommend that the IgM ELISA be used as an initial screening test followed one of the molecular assays in samples that are collected in the early phase of illness and negative for CHIKV IgM antibodies. Such as approach would enable rapid confirmation of the diagnosis and implementation of public health measures especially during outbreaks.

Aim: 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. Methods and Results: 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. Conclusion: The antiviral activity showed by SCH16 was enhanced in the presence of ribavirin and mycophenolic Acid. Significance and Impact of the Study: 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.

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.

Japanese encephalitis virus (JEV) envelope (E) protein has been shown to play a critical role in attachment to cells. However, the receptor interacting with envelope protein has not been conclusively identified. Using mouse neuroblastoma (Neuro2a) cells and purified JEV-E protein in `Virus Overlay Protein Binding Assay' followed by MALDI-TOF analysis, we identified `heat shock protein 70' (Hsp70) as a possible receptor for JEV. Indirect immunofluorescence and flow-cytometry analysis demonstrated localization of Hsp70 on Neuro2a cell surface. Co-immunoprecipitation followed by Western blot analysis reconfirmed the interaction between Hsp70 and JEV-E protein. Further, anti-Hsp70 polyclonal-antibodies were able to block JEV entry into Neuro2a cells. Additionally, using the bioinformatic tool - FTDOCK, clocking between the proteins was performed. Amongst six interacting structural poses studied one pose involving RGD motif on JEV-E and leucine(539) on Hsp70 displayed stable interaction. These observations indicate that Hsp70 serves as putative receptor for JEV in Neuro2A cells.

Several investigations have shown that pentoxifylline possesses broad-spectrum antiviral activity against a range of RNA and DNA viruses. However, its ability to inhibit Japanese encephalitis virus (JEV) replication has not yet been studied. The present study was designed to investigate the antiviral activity of pentoxifylline against JEV in vitro and in vivo. The activity of pentoxifylline against JEV was evaluated in vitro using cytopathic effect inhibition and plaque reduction assays. Pentoxifylline was able to inhibit JEV replication in a dose-dependent manner at a 50% inhibitory concentration (IC 50 ) of 50.3 μg/mL (0.00018 μM) and a therapeutic index (TI) of 10. Experiments to study the mechanism of antiviral action of pentoxifylline using in vitro translation of viral mRNA suggested that the drug did not interfere either with early or late protein synthesis but most likely exerted its action on virus assembly and/or release. Furthermore, the in vivo study showed that pentoxifylline at a concentration of 100 mg/kg and 200 mg/kg body weight was able to protect completely mice challenged with 50 × 50% lethal dose (LD 50 ) of JEV.

Passive immunization is an important parameter of post exposure rabies prophylaxis. Two types of rabies immunoglobulin (RIGs) are currently available for Passive immunization against rabies i.e., human rabies immunoglobulin (HRIG) and equine rabies immunoglobulin (ERIG). The former is very expensive and not easily available and the latter causes side effects because of which its utility is limited. In the present study we have produced murine monoclonal antibodies (Mabs) to rabies glycoprotein (G) and studied their utility in passive immunization against rabies using animal models. Their efficacy was compared to commercially available ERIG both in terms of neutralizing titer and effective protein concentration. The neutralizing titers of these Mabs ranged from 1650-75,000 IU/mL by RFFIT. They belonged to the IgG 2 a subclass. The Mabs were able to protect 70-100% of mice and guinea pigs inoculated with rabies viruses, depending on the strain of the virus. These Mabs were found to be 2,000 times more potent than commercial ERIG in terms of effective protein concentration and neutralizing titer. Further studies are required to study their utility in humans exposed to rabies.

A PCR was standardized for amplifying three different mycobacterial--IS6110, MPB-64, 65 kDa DNA sequences. A comparative evaluation of the three PCR assays was carried out for the rapid diagnosis of tuberculous meningitis (TBM) using cerebrospinal fluid (CSF) specimens. While the IS6110 PCR was a single-step amplification reaction, the MPB-64 and 65 kDa antigen PCR assays were nested reactions. A total of 176 cerebrospinal fluid (CSF) samples from 176 patients were subjected to amplification of the three different mycobacterial sequences. Amongst them, 45 samples were obtained from confirmed cases of TBM (culture positive) and 56 samples were obtained from clinically suspected cases of TBM which were culture-negative. The remaining 75 CSF samples were categorized under the non-infectious and infectious illness of the central nervous system (CNS). Against a gold standard of culture, a sensitivity of 98% (NPV=99%) and a specificity of 100% (PPV=100%) was observed with the IS6110 PCR. Among the nested PCRs, a sensitivity of 91% (NPV=94%) and a specificity of 91% (PPV=85%) was observed with the MPB-64 assay, while the 65 kDa protocol had an associated sensitivity of 51% (NPV=76%) and a specificity of 92% (PPV=79%). These findings suggest that among the nested PCR assays, the MPB-64 PCR assay was associated with an enhanced degree of sensitivity and was comparable in terms of specificity. Our study also demonstrates that the IS6110 assay, while being a single-step PCR had the advantage of being a rapid test for the diagnosis of TBM, with increased sensitivity and enhanced specificity as compared to the nested PCR protocols.

Most studies of cognitive functioning in human immunodeficiency virus type 1 (HIV-1)-seropositive (HIV-1+) subjects have been done in the United States and Europe, where clade B infections predominate. However, in other parts of the world such as South India, where clade C HIV is most common, the prevalence of HIV-1 is increasing. Standardized neuropsychological tests were used to assess cognitive functioning in a sample of 119 adults infected with clade C HIV-1 who were not on antiretroviral medications. The subjects did not have neurological or psychiatric illness and were functioning adequately. Neuropsychological test performance was compared with gender-, age-, and education-matched normative data derived from a sample of 540 healthy volunteers and a matched cohort of 126 healthy, HIV-1-seronegative individuals. Among the seropositive subjects, 60.5% had mild to moderate cognitive deficits characterized by deficits in the domains of fluency, working memory, and learning and memory. None of the subjects had severe cognitive deficits. The HIV-1+ sample was classified into groups according to the level of immune suppression as defined by CD4 count (200, 201-499, and500 cells/mm3) and viral load (5000, 5001-30,000, 30,001-99,999, 100,000-1,000,000, and1,000,001 copies). Although the most immunosuppressed group (CD4 count200 cells/mm3 or viral load1,000,001 copies) was small, their rate of impairment in visual working memory was greater when compared to groups with better immune functioning. Mild to moderate cognitive deficits can be identified on standardized neuropsychological tests in clade C-infected HIV-1+ adults who do not have any clinically identifiable functional impairment. The prevalence of cognitive deficits is similar to that reported in antiretroviral treatment-naïve individuals infected with clade B virus in the western world.

To study the association between quality of life (QOL) domains and biological markers of disease progression of HIV infection, i.e. viral load (VL) and CD4 counts among asymptomatic subjects with HIV subtype C infection in South India. Quality of life was measured using the locally validated version of the WHOQOL HIV-BREF. The subjects were neurologically asymptomatic, non psychiatrically ill HIV infected men and women participating in a cohort study. The results indicated mixed findings, with some QOL dimensions being associated with high VLs and low CD4 counts while several others did not show any associations. Significant associations were seen between low CD4 counts and the psychological and social relationships domain, with lower mean scores in these domains being reported by subjects having CD4 counts

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 at two sites 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.

Clinical and laboratory characteristics of 39 patients with adult onset subacute sclerosing panencephalitis (SSPE) are described and compared to those of juvenile onset patients regarding preceding measles, age at onset, gender, interval between onset and diagnosis, clinical profile, and course during follow up. Diagnosis was based on clinical and electroencephalographic findings and raised anti-measles antibody titres in cerebrospinal fluid. Mean age at SSPE symptom onset was 20.9+/-4.9 years and mean interval from onset to diagnosis was 6.3+/-9.6 months. Referral diagnosis was accurate in only 12 patients. Presenting symptoms included myoclonus, behavioural changes, seizures, and cognitive, visual, and extrapyramidal disturbance. All patients received symptomatic therapy; 19 also received disease modifying agents. Five of seven pregnant women had successful deliveries. The follow-up period varied widely (maximum 60 months, median 9 months). The profile of adult onset SSPE did not differ from the rest of the cohort, except for a longer interval between measles infection and symptom onset (p

Antibodies against Mycobacterium tuberculosis antigens were detected by enzyme-linked immunosorbent assay in cerebrospinal fluid (CSF) samples obtained from 442 patients with tuberculous meningitis (TBM) and 102 control patients. Antibodies were found in the CSF of 87% of patients with clinical (culture-negative) TBM, 72% of patients with culture-positive TBM, and 65% of patients with autopsy-proven TBM. That anti-M. tuberculosis antibodies were detected in the CSF of patients with clinically diagnosed cases more frequently than in patients with culture-positive cases suggests that the detection of antibodies in CSF tends to decrease as bacillary load increases. Of the patients with clinical TBM who were coinfected with human immunodeficiency virus (HIV), 70% exhibited anti-M. tuberculosis antibody in CSF, which suggests that antibody responses in this group were substantially weaker than those in HIV-negative patients with clinical TBM. Some groups showed a stronger response to certain antigens, which suggests that antigen recognition patterns may be specific for the stage of disease.

The etiopathogenesis of schizophrenia is poorly understood. Within the proposed “neurodegeneration paradigm”, observations have been put forth for “accelerated aging” in this disorder. This proposition is largely based on the neuroscience research that demonstrates progressive changes in brain as well as other systemic abnormalities supportive of faster aging process in patients with this disorder. In this review, we have summarized the literature related to the concept of early aging in schizophrenia. These studies include P300 abnormalities & visual motion discrimination, neuroimaging findings, telomere dynamics as well as neuropathology of related brain regions. We also propose a role of vitamin D, neuroimmunological changes and elevated oxidative stress as well as mitochondrial dysfunction in addition to the above factors with ‘vitamin-D deficiency’ as the central paradox. Put together, the evidence supporting early aging in schizophrenia is compelling and this requires further systematic studies.

The aim of the study was to study factors related to anxiety, depression, and suicidal ideation among HIV-seropositive heterosexuals soon after being tested for their HIV status for the first time. Anxiety, depression, and suicidal ideation were assessed among 51 HIV-seropositive heterosexual men and women with various stages of HIV infection. All assessments were done between 4 and 6 weeks after revelation of positive serostatus. Psychosocial variables such as quality of family relationships and substance use and sociodemographic details such as gender, income, education, and residence were studied for their association with psychiatric morbidity. Illness details studied for their association with psychiatric morbidity included stage of HIV infection, spouse's HIV status, presence of physical illness, and pain. Depression was present in 40% and anxiety in 36% of the sample. Serious suicidal intent was seen in 14%. Multiple regression analysis indicated that presence of pain, concurrent alcohol abuse, poor family relations, and presence of AIDS in the spouse were significant factors associated with depression, anxiety, and suicidal ideation.

Neurodevelopmental aberrations influenced by neurotrophic factors are among the important paradigms to understand schizophrenia pathogenesis. Among various neurotrophic factors, Brain-Derived Neurotrophic Factor (BDNF) is strongly implicated by previous research studies. Evaluating co-morbidity free, antipsychotic-naïve schizophrenia patients for BDNF levels and examining the correlates of this factor with symptoms might facilitate elucidation of its pathogenetic role without confounds of potential influencing factors. In this study, 59 co-morbidity free, antipsychotic-naïve schizophrenia patients were compared with 60 healthy controls for serum BDNF levels. In addition, the relationship between Schneiderian First Rank Symptoms (FRS) and BDNF level in patients was examined. As a group, schizophrenia patients (28.8 ± 11.7 ng/mL) had significantly lower serum BDNF than healthy controls (34.9 ± 8.2 ng/mL) after controlling for the potential confounding effects of age and sex (F = 7.8; p = 0.006). Further analyses revealed FRS status to have significant effect on plasma BDNF after controlling for the potential confounding effects of age and sex (F = 4.5; p = 0.01). Follow-up post hoc analyses revealed FRS(+) patients to have significant deficit in plasma BDNF level in comparison with healthy controls (p = 0.002); however, FRS(-) patients did not differ from healthy controls (p = 0.38). Our study observations add further support to the role for BDNF in schizophrenia pathogenesis and suggest a potential novel link between deficient BDNF and FRS.

Background Various lines of evidence including epidemiological, genetic and foetal pathogenetic models suggest a compelling role for Interleukin-6 (IL-6) in the pathogenesis of schizophrenia. IL-6 mediated inflammatory response triggered by maternal infection or stress induces disruption of prenatal hippocampal development which might contribute towards psychopathology during adulthood. There is a substantial lack of knowledge on how genetic predisposition to elevated IL-6 expression effects hippocampal structure in schizophrenia patients. In this first-time study, we evaluated the relationship between functional polymorphism rs1800795 of IL-6 and hippocampal gray matter volume in antipsychotic-naïve schizophrenia patients in comparison with healthy controls. Methodology We examined antipsychotic-naïve schizophrenia patients [N = 28] in comparison with healthy controls [N = 37] group matched on age, sex and handedness. Using 3 Tesla – MRI, bilateral hippocampi were manually segmented by blinded raters with good inter-rater reliability using a valid method. Additionally, Voxel-based Morphometry (VBM) analysis was performed using hippocampal mask. The IL-6 level was measured in blood plasma using ELISA technique. SNP rs1800795 was genotyped using PCR and DNA sequencing. Psychotic symptoms were assessed using Scale for Assessment of Positive Symptoms and Scale for Assessment of Negative Symptoms. Results Schizophrenia patients had significantly deficient left and right hippocampal volumes after controlling for the potential confounding effects of age, sex and total brain volume. Plasma IL-6 levels were significantly higher in patients than controls. There was a significant diagnosis by rs1800795 genotype interaction involving both right and left hippocampal volumes. Interestingly, this effect was significant only in men but not in women. Conclusion Our first time observations suggest a significant relationship between IL-6 rs1800795 and reduced hippocampal volume in antipsychotic-naïve schizophrenia. Moreover, this relationship was antithetical in healthy controls and this effect was observed in men but not in women. Together, these observations support a “differential susceptibility” effect of rs1800795 in schizophrenia pathogenesis mediated through hippocampal volume deficit that is of possible neurodevelopmental origin.

The protective ability of cytotoxic T cells (CTL) raised in vitro against Japanese encephalitis virus (JEV) was examined by adoptive transfer experiments. Adoptive transfer of anti-JEV effecters by intracerebral (i.c.) but not by intraperitoneal (i.p.) or intravenous (i.v.) routes protected adult BALB/c mice against lethal i.c. JEV challenge. In contrast to adult mice, adoptive transfer of anti-JEV effecters into newborn (4-day-old) and suckling (8-14-day-old) mice did not confer protection. However, virus-induced death was delayed in suckling mice compared to newborn mice upon adoptive transfer. The specific reasons for lack of protection in newborn mice are not clear but virus load was found to be higher in newborn mice brains compared to those of adults and virus clearance was observed only in adult mice brains but not in newborn mice brains upon adoptive transfer. Specific depletion of Lyt 2.2(+), L3T4(+) or Thy-1(+) T cell populations before adoptive transfer abrogated the protective ability of transferred effecters. However, when Lyt 2.2(+) cell-depleted and L3T4(+) cell-depleted effecters were mixed and transferred into adult mice the protective activity was retained, demonstrating that both Lyt 2.2(+) and L3T4(+) T cells are necessary to confer protection. Although the presence of L3T4(+) T cells in adoptively transferred effector populations enhanced virus-specific serum neutralizing antibodies, the presence of neutralizing antibodies alone without Lyt 2.2(+) cells was not sufficient to confer protection.

We have previously reported that Lyt(2+) cytotoxic T lymphocytes (CTL) can be raised against Japanese encephalitis virus (JEV) in BALB/c mice. In order to confirm the presence of H-2K(d)-restricted CTL and to examine their cross-recognition of West Wile virus (WNV), we tested the capacity of anti-JEV CTL to lyse uninfected syngeneic target cells that were pulsed with synthetic peptides. The sequence of the synthetic peptides was predicted based upon the H-2K(d) binding consensus motif. We show here that preincubation of uninfected syngeneic targets (P388D1) with JEV NS1- and NS3-derived peptides [NS1 (891-899) and NS3 (1804-1812)], but not with JEV NS5-derived peptide [NS5 (3370-3378)], partially sensitized them for lysis by polyclonal anti-JEV CTL. These results indicate the CTL recognition of NS1- and NS3-derived peptides of JEV.

This study reports the pathological findings and the distribution of viral antigen in the brains of 13 confirmed and autopsied cases of Japanese encephalitis (JE) in correlation with other virus-specific immunological parameters measured in the cerebrospinal fluid (CSF) antemortem. Japanese encephalitis virus (JEV)-specific antibodies were detected in the CSF of 10 of 13 patients, JEV antigen was detected in the CSF of 7 of 13 and JEV-specific immune complexes were detected in the CSF of 3 of 11 patients. Viral antigen was localised immunocytochemically in the brain tissue of 11 of 13 cases, indicating, that viral antigen could not be cleared from the tissues by the antibody. The topographic distribution of the tissue-associated antigen in the thalamus, hippocampus, substantia nigra and medulla oblongata explain the evolution of post JE sequelae.

Immunological and virological evidence for persistence of Japanese encephalitis virus (JEV) in the human nervous system is described in 16/323 (5%) laboratory-confirmed cases of Japanese encephalitis. In 9/16 patients, JEV specific IgM antibodies were detected in the CSF even at 50-180 days after the onset of symptoms. Similarly, in 7/16 patients, apart from IgM antibodies, viral antigen was also present in the CSF beyond the third week of illness and in one patient it could be detected even at 117 days. Infectious virus could be isolated from the CSF beyond the third week of illness in 3/16 patients. In one patient, JEV was isolated from the CSF on three consecutive occasions at 90, 110, and 117 days after onset of clinical symptoms. These findings suggest that JEV persists in the nervous system of a small proportion of patients.

Infection with Japanese encephalitis virus (JEV) is a major public health problem in Asia. Detection of JEV-specific IgM in serum and cerebrospinal fluid (CSF) by the IgM antibody capture enzyme-linked immunosorbent assay (MAC-ELISA) is currently the most widely used diagnostic method to detect JEV infection. Because of the possible presence of IgM cross-reactivity with other flaviviruses in serum and the high ratio of inapparent-to-apparent JEV infections, a positive result in serum only suggests a recent infection and not necessarily an encephalitic illness caused by JEV. Consequently, detection of JEV-specific IgM in CSF assumes great diagnostic relevance. We evaluated two commercial JEV MAC-ELISA kits using 60 CSF samples obtained from patients with acute encephalitis syndrome. The Panbio and XCyton kits had sensitivities of 65-80% and 95% and specificities of 90% and 97.5%, respectively. Performance information on these commercial JEV MAC-ELISA kits for CSF should assist in laboratory-based JE surveillance programs.

Background: 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. Methods: 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. Results: 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 IC50 of 3.1 µg/ml, a therapeutic index of 16 and a 75% protection against lethal challenge of JEV. Conclusion: The study concludes that this compound significantly inhibited the replication of JEV in vitro and protected mice in vivo.

Objective There are only a few studies correlating diverse radiological and EEG features of subacute sclerosing panencephalitis (SSPE). The objective of the study was to (a) describe EEG profile and (b) correlate it with the clinical and imaging data of patients with confirmed SSPE. Methods This study was conducted at a University teaching hospital in south India and involved 58 patients (M:F=37:21, age: 12.3, SD 4.8 years) of SSPE. Diagnosis of SSPE was based on the characteristic clinical manifestations, and raised IgG (⩾1:625) anti-measles antibody in cerebrospinal fluid (CSF) by ELISA in all the patients. Scalp EEGs were recorded on 16 channel machines using standard parameters and procedures. The EEG, clinical and imaging data were reviewed. Results EEGs were frequently abnormal: typical (37) and atypical (21). Diffuse slowing of background activity (BGA) was noted in 46 records being asymmetrical in six. Periodic complexes were periodic (32), quasi-periodic (21) or a-periodic (4). Periodic complexes (PC) (amplitude: 370.7, SD 171.2μV; duration – 1.7, SD 2.0s; inter-complex interval: 8.4, SD 9.2s) were symmetrical in 39 and asymmetrical in 19. CT (32) and MRI (23) scans were normal in 16 patients while others had white matter (15), cerebral edema (8), cerebral atrophy (8), basal ganglia (2), and thalamic (2) changes. There was an independent association of frontally dominant slowing of BGA ( p =0.04) and typical PCs ( p =0.03) with the diffuse cerebral edema on imaging. White matter changes correlated with slowing of BGA ( p =0.04), but not with typical PC ( p =0.16). Conclusions This study provides valuable insight into the structural and clinical correlates of EEG changes in SSPE. Significance Irrespective of the incidence of occurrence of SSPE in a community, a clinician should be aware of the wide spectra of EEG findings. This study also discusses the possible underlying structural and clinical correlates.

Electrophysiological studies in subjects with HIV/AIDS demonstrate subtle changes in the visual pathway even in the absence of visual symptoms. But the pathological correlate of the electrophysiological abnormalities is largely unknown. This study attempts to correlate pathological changes in the retina and intraorbital portion of optic nerve in four drug naive patients of AIDS caused by HIV-1 clade C, who had abnormalities in the visual evoked potentials recorded antemortem. Three had no visual complaints and one patient had sudden loss of vision in the right eye. In all four patients, the visual evoked potentials disclosed variable prolongation of P100 latencies. Histologically axonal cytoskeletal breakdown and depletion in the optic nerves was the cardinal finding with variable myelin loss, even in the absence of overt visual dysfunction, or infective retinitis. The axonal loss was maximal in the symptomatic case. Retinal ganglion cell depletion was seen in only two patients. Sectoral infiltration of the optic nerve by cryptococci and Cryptococcal choroiditis was the only opportunistic infection to involve the eye. Axonal pathology in the optic nerve appears to be related to the abnormalities recorded in visual evoked potentials even in the absence of overt clinical symptoms. Opportunistic infections could be contributing to the axonal pathology in the optic nerve in patients with AIDS.

Background: Rabies is an enzootic and fatal disease and is still a major problem in developing countries. Ante mortem diagnosis in human cases is necessary for medical management of the patient and to ensure appropriate post-exposure treatment of contacts. Both conventional RT-PCR and Real time PCR (TaqMan) have been described for the detection of rabies virus RNA from saliva and tissue respectively, however to date, there have been no studies comparing conventional and real time PCR assays for detection of rabies virus nucleic acid using saliva samples for ante mortem diagnosis. Objectives: In this study, we evaluated the utility of conventional RT-PCR and SYBR Green I Real time PCR in the ante mortem diagnosis of rabies using saliva samples. Study design: Saliva samples collected from twenty-four patients presenting with typical clinical manifestations of rabies were tested in the two assays. Results: Amongst the 24 samples tested, 21 samples (87.5%) were positive by either of the two molecular methods. Of these 21, rabies virus RNA was detected in 6/21 in the conventional RT-PCR assay while SYBR Green I Real time PCR could detect RNA in 18/21 samples. Conclusion: Real time PCR assay was more sensitive than conventional RT-PCR assay (sensitivity 75% versus 37%, p = 0.0189). This study highlights the utility of molecular diagnostic tests in establishing ante mortem diagnosis of rabies using saliva samples within a few hours.

Cell-mediated immune response to Japanese encephalitis virus (JEV) and its purified envelope (E) protein was measured in 45 laboratory confirmed JE patients using a proliferation assay of peripheral blood mononuclear cells (PBMC). In parallel, JEV-specific IgM antibodies were measured by ELISA. No correlation was observed between the antibody response and results of the lymphocyte proliferation assay. Only 11 of the 42 patients positive in the antibody test were positive in the proliferation assay, and PBMC from 14/45 (31%) patients did not respond to either phytohemagglutinin and to JEV and/or its purified E protein antigen. No correlation was observed between the cell-mediated immune response and the final clinical outcome (fatality vs. recovery).