Expression of cytokine-mRNA in peripheral blood mononuclear cell of human immunodeficiency virus-1 subtype C infected individuals with opportunistic viral infections from India (South).

Human immunodeficiency virus (HIV) disease progression is associated with a marked change in the level of plasma cytokines. The study reported here investigated the level of mRNA expression of different cytokines: Tumour necrosis factor-alpha (TNF-α), interferon (INF)-gamma, interleukin-10 (IL-10) and IL-21 in the peripheral blood mononuclear cell among the antiretroviral therapy naive subtype C HIV-1 infected individuals and normal healthy controls by real time polymerase chain reaction. The mRNA expressions of all the 4 cytokines in HIV-1 infected individuals were significantly higher compared to healthy controls (P value range 0.0004-0.01). The mean level of IL-10, INF-gamma and TNF-α were higher in HIV infected individuals with low CD4 counts (<300 cells/µl). The IL-10 expression showed a significant negative correlation with CD4 counts (r=-0.25, P=0.04) while IL-21 showed a positive correlation with CD4 counts (r=0.26, P=0.03). There was a significant negative correlation between the cytomegalovirus (CMV) viral load and IL-21 expression. Cytokine levels by mRNA detection avoids the inherent problem of measuring plasma level and this study also provide information on the cytokine levels and CD4+ T cell level among HIV-1 subtype C infected individuals with opportunistic viral infections like CMV.

Frequency of Epstein--Barr virus infection as detected by messenger RNA for EBNA 1 in histologically proven gastric adenocarcinoma in patients presenting to a tertiary care center in South India.

BACKGROUND: Epstein-Barr virus (EBV)-associated gastric carcinoma is a relatively uncommon entity detected in approximately 10% of gastric adenocarcinoma. OBJECTIVE: The purpose of this study is to estimate the frequency of EBV-associated gastric carcinoma and also to assess the nature of presentation, any significant difference between this subgroup and EBV-negative gastric adenocarcinomas with respect to age and sex predilection, lymph nodal status, site of presentation. MATERIALS AND METHODS: We prospectively analyzed 100 cases of gastric adenocarcinoma who underwent either a partial or total gastrectomy during the period from March 2010 to August 2011. The tumour and the corresponding normal gastric tissue from the same patient were analyzed for the presence of Epstein-Barr nuclear antigen 1 (EBNA1) messenger ribonucleic acid (mRNA) by real-time polymerase chain reaction (PCR). RESULT: EBV was detected in 6% cases of gastric adenocarcinoma. All the positive patients were males. The majority of cases involved the proximal stomach and there was variable lymph nodal involvement. CONCLUSION: Our study endorses that there is an association between EBV infection and gastric adenocarcinoma in the Indian population. There was no significant difference between this subgroup and EBV-negative gastric adenocarcinomas with respect to age and sex predilection, lymph nodal status and site of presentation. Short-term follow-up of this subgroup of patients seems to indicate a good overall prognosis after appropriate treatment. However, a larger study with long-term follow-up is needed to further establish the role of EBV in gastric adenocarcinoma in this study population.

Emergence of HIV-1 drug-resistant variants in women following antiretroviral prophylaxis for the prevention of mother to child transmission.

PURPOSE: Emergence of drug resistance following HIV prophylaxis has an important impact on ART program. OBJECTIVE: To investigate the emergence of drug resistance in HIV-1 infected pregnant women. MATERIALS AND METHODS: Fifty-three HIV-1 infected pregnant women who had received 4-12 weeks of antenatal AZT followed by Nevirapine during delivery and Combivir [AZT + 3TC] for 1 week postpartum (group-1, n = 48) or who come at the time of delivery and received Nevirapine followed by Combivir for 1 week (group-2, n = 5) were recruited. Samples were collected prior to the start of the prophylaxis and 5-8 weeks postpartum. In addition, a third sample was collected between 26-65 weeks postpartum from 7 women. Amplification of HIV-1 pol gene and drug resistance analysis was done. RESULT: Two (3.8%) women in group-1 showed transmitted drug resistance and they continued to show this even at 6 weeks postpartum. One (2%) woman from group-1 showed a mutation after 6-8 weeks of prophylaxis. Among the samples collected between 26-65 weeks postpartum, 3/7 (43%) showed mutations and all these women belong to group-1. Women belonging to group-2 didn't show mutation prior to or following prophylaxis. CONCLUSION: In contrast to the available data among pregnant women with ART prophylaxis, our data showed reduced frequency of mutations following 5-8 weeks of postpartum but an emergence of mutation later (26-65 weeks). The addition of Combivir with the single dose Nevirapine during delivery and the early stage of the disease with higher CD4 counts could be the reasons for this.

Changing paradigm of cryptococcal meningitis: an eight-year experience from a tertiary hospital in South India.

BACKGROUND: Cryptococcal meningitis (CM) is a common opportunistic fungal infection causing sub-acute meningitis with the potential for complications and significant mortality. We conducted this study to describe the difference in presentation and outcome between HIV-infected and HIV-uninfected patients. MATERIALS AND METHODS: Patients admitted to a tertiary care centre between 2005 and 2013 with confirmed CM were included in the analysis. Details of the clinical presentation, laboratory findings, treatment details, risk factors for infection and outcome were documented and analysed. RESULTS: During the study period, 102 (87.2%) cases of CM occurred among HIV infected individuals, whereas 15 (12.8%) occurred among HIV-uninfected patients. HIV-infected patients with CM were younger compared with HIV-uninfected patients (38.2 ± 8.5 years vs. 45 ± 11.5 years; P = 0.07). The median duration of symptoms prior to presentation was shorter in the HIV-infected group (20 ± 32 vs. 30 ± 42; P = 0.03). There was no difference between the cerebrospinal fluid (CSF) lymphocyte counts, CSF protein counts, and CSF sugar levels in both the groups. The diagnostic yield of Cryptococcus was similar with CSF India ink smear (89% vs. 87%), CSF fungal culture (95% vs. 87%), and blood culture (100% vs. 75%) in both the groups. Case fatality rate in the HIV-infected group was 30.6%, whereas there were no deaths in the HIV-uninfected group. CONCLUSION: HIV-infected patients with CM have a worse outcome compared to HIV-uninfected patients. The overall trend over 3 decades shows increasingly successful rates of treatment and hence early diagnosis and treatment are of paramount importance.

Significance of Epstein-Barr virus (HHV-4) and CMV (HHV-5) infection among subtype-C human immunodeficiency virus-infected individuals.

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.

Antiviral efficacy of adefovir dipivoxil in the treatment of chronic hepatitis B subjects from Indian subcontinent.

Adefovir is one of the therapeutic options for the treatment of chronic hepatitis B. A total of 30 adefovir-experienced subjects with the median treatment duration of 12 (interquartile range (IQR) 6-18) months were studied. Virological response was measured by hepatitis B virus deoxyribonucleic acid (HBV DNA) levels. HBV reverse transcriptase (rt) domains were sequenced for the identification of resistance mutations. Among the 30 subjects, two (7%) showed virological response and 19 (63%) were non-responders. The virological response for the remaining nine (30%) subjects was not determined. On sequence analysis, two subjects were identified with rtI169L and rtA181V mutation after 9 months and 18 months of adefovir treatment, respectively. Though the frequencies of adefovir resistance mutations are low, a large majority of subjects showed non-response. Therefore, adefovir in the management of HBV should be used judiciously.

Human herpes virus-8 infections among subjects with human immunodeficiency virus infection and normal healthy individuals in India.

Human herpes virus-8 (HHV-8) is etiologically associated with Kaposi's sarcoma. There is insufficient information on the epidemiology of HHV-8 infection from India. Blood samples from 87 human immunodeficiency virus (HIV)-infected individuals and 84 normal healthy blood donors were tested for the HHV-8 IgG antibodies. Further, a total of 309 whole blood samples from treatment-naïve HIV-1-infected individuals and from 70 normal healthy individuals were also collected and tested for HHV-8 DNA. The seroprevalence of HHV-8 was 4.7% in the South Indian population. There was no significant difference in the seroprevalence of HHV-8 in the HIV-infected and uninfected patients. None of the 379 samples tested were positive for HHV-8 DNA. Our study revealed a very low exposure of the South Indian patient population to HHV-8 and multicentric epidemiological studies are needed to understand the prevalence of HHV-8 in different regions of India and to confirm any gender-specific differences in seroprevalence.

Comparison of HIV-1 RNA level estimated with plasma and DBS samples: a pilot study from India (South).

PURPOSE: The use of dried blood spots (DBS) for HIV-1 viral load determination could greatly enhance the management of HIV infected individuals in resource-limited countries. OBJECTIVE: To compare the HIV-1 viral load values obtained between parallel collected plasma and DBS. MATERIALS AND METHODS: DBS and plasma samples were collected from 62 HIV-1 infected individuals and were used for determination of HIV-1 RNA concentrations using the Abbot real-time HIV-1 PCR. RESULT: Mean of the log difference of viral load values between plasma and DBS was -0.41 log. DBS viral load values significantly correlated with plasma viral load (r = 0.9818, P < 0.0001). CONCLUSION: These results suggest that DBS samples can be used as an alternative to plasma for the estimation of HIV-1 viral load if samples are appropriately stored.

Human immunodeficiency virus type-2-A milder, kinder virus: an update.

Human immunodeficiency virus type-2 (HIV-2) belongs to the family retroviridae which is phylogenetically clusters with SIV SM from sooty mangabeys. This virus is morphologically similar to human immunodeficiency virus type-1 (HIV-1) but has got only a 40% homology at the nucleotide level. There is a distinct geographical distribution of HIV-2 unlike HIV-1. There are currently eight subtypes/groups identified with subtype/group A responsible for the majority of infections. HIV-2 shows a considerable difference in the course of the disease. Clinical, haematological and immunological evaluation of individuals infected with HIV-2 has shown the virus to be less pathogenic than HIV-1 although the exact mechanism underlying this difference is not well defined. Similar to HIV-1, the HIV-2 isolates also showed distinct replicative and cytopathic characteristics. The transmission rate for HIV-2 compared to HIV-1 is very low both by heterosexual route and mother to child transmission. The clinical signs and symptoms of immunodeficiency associated with HIV-2 are similar to the ones seen among the HIV-1-infected individuals and they can also progress to AIDS. It is naturally resistant to NNRTI and hence the diagnosis become important as it affects the treatment strategy. Similar to HIV-1, HIV-2 strains of infected individuals also show mutations that can cause drug resistance. The current evidence suggests that there is no protective effective for HIV-2 against HIV-1.

Antiviral resistance mutations and genotype-associated amino acid substitutions in treatment-naïve hepatitis B virus-infected individuals from the Indian subcontinent.

BACKGROUND/AIMS: Antiviral resistance is a major challenge to the treatment currently available for hepatitis B virus (HBV). In this study, mutations that may affect the antiviral efficacy in treatment-naïve HBV-infected individuals were analyzed. METHODS: Ninety-seven treatment-naïve HBV-infected individuals were included in this study. HBV reverse transcriptase (rt) domains were sequenced and nucleotide differences were compared to GenBank wild-type sequences. Furthermore, HBV genotypes, subgenotypes and subtypes were determined by analyzing surface gene sequences. RESULTS: An adefovir-related rtI233V mutation was identified in 4 subjects. The rtS213T lamivudine and entecavir refractory mutant was presented in 3 individuals. Altogether, drug-related, atypical and novel HBVrt amino acid substitutions were seen in 73 positions. The HBV genotypes A, C, D and G were depicted in 15, 21, 60 and 1 individuals, respectively. There were 17 HBVrt amino acid substitutions that are associated with certain genotypes of HBV. Mutations in HBVrt corresponded to established surface gene mutations in 9 patients. CONCLUSION: This data shows that antiviral-resistant HBV strains do exist in treatment-naïve individuals in this region. Further studies are essential to characterize the role of HBVrt amino acid substitutions in response to anti-HBV therapy.

Evaluation of a dry format reagent for CD4+ and CD8+ T-cell enumeration with FACSCount and Guava PCA [corrected].

PURPOSE: In all CD4+/CD8+ T-cell estimation systems, the reagents used are liquid in nature and have to be transported and stored at 2°-8°C. This causes problems in countries where the ambient temperature is high for most parts of the year or where the laboratories are at remote places. MATERIALS AND METHODS: We evaluated a dry format of CD4/CD8 reagents from ReaMetrix (Bangalore, India) against the existing liquid reagents from Becton Dickinson (San Jose, CA, USA) and Guava PCA system (Guava Technologies, Hayward, CA, USA). Blood samples collected during March 2009 through May 2009 from 102 HIV-infected individuals and 31 normal healthy individuals in a tertiary care centre in India (south) were tested by Guava EasyCD4 System (PCA) and FACSCount using the respective reagents and the corresponding ReaMetrix reagents. RESULTS: Overall, the correlation (r) of the new Rea T Count and FACSCount reagents for the CD4+ T-cell estimation was 0.98, while with ReaPan 3 4 G reagent in the Guava PCA system with the Guava reagent was 0.97. The mean bias for CD4+ T-cell measurements between Rea T count and BD reagent was -6 cells/ml, while the same with ReaPan 3 4 G reagent in the Guava PCA system was 78 cells/ml. The mean bias for the Rea T count and the ReaPan 3 4 G reagent tested in the FACSCount and Guava PCA system was 17 cells. CONCLUSIONS: The dry reagents were found to be reliable and cheaper compared to the existing liquid reagents. This allows the transportation of reagents in the absence of cold chain and will facilitate a more user-friendly CD4+ T-cell testing system.

Comparison of a conventional polymerase chain reaction with real-time polymerase chain reaction for the detection of neurotropic viruses in cerebrospinal fluid samples.

PURPOSE: To compare a conventional polymerase chain reaction (PCR) and real-time PCR for the detection of neurotropic DNA viruses. MATERIALS AND METHODS: 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. RESULTS: Overall, in the entire set of samples, the real-time PCR yielded 88 (59.9%) positives and conventional PCR had six (4.1%) positives. CONCLUSION: 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%).

Association of HLA and TNF polymorphisms with the outcome of HBV infection in the South Indian population.

The role of host genetic factors in the pathogenesis and outcome of hepatitis B virus (HBV) infection is not well known. We assessed the association of HLA and TNF (rs361525, rs1800629, rs1799724, rs1800630 and rs1799964) polymorphisms with HBV outcome in the South Indian population. Association of HLA polymorphism was analyzed in 90 individuals from each group, that is, spontaneous recovery (SR) and chronic-HBV (C-HBV) infection. The role of TNF polymorphisms was evaluated in 150 subjects with SR and 137 patients with C-HBV infection. After adjusting for age and sex, HLA-DRB1*07:01 was strongly associated with chronicity (corrected P-value (pc) <0.005, odds ratio (OR) 3.76, 95% confidence interval (CI) 1.84-7.68). The rs1800630 genotype was associated with HBV outcome in codominant (pc<0.01, OR=1.99, 95% CI 1.30-3.05) and dominant (pc<0.01, OR=2.28, 95% CI 1.35-3.84) analyzing models after adjusting for age and sex. Similarly, the rs1799964 genotype was associated with HBV outcome in codominant (pc=0.01, OR=1.57, 95% CI 1.09-2.27) and dominant (pc<0.01, OR=2.21, 95% CI 1.27-3.83) analyzing models. Haplotype analysis (rs1799964/rs1800630/rs1799724/rs1800629/rs361525) revealed that the CACGG haplotype was strongly associated with C-HBV infection (P=0.0004). Our study suggests that inheritance of HLA and TNF polymorphisms might explain the outcome of HBV infection in the South Indian population.

Frequency of HIV type 2 infections among blood donor population from India: a 10-year experience.

PURPOSE: In India, HIV-2 epidemic is alongside with HIV-1. Blood banks are introducing nucleic acid testing (NAT) for screening. The limitation of NAT systems is the inability to detect HIV-2. MATERIALS AND METHOD: An analysis of HIV screening of a blood bank at a tertiary care center from 1998 to 2007 was carried out. RESULTS: A total of 175026 donors were screened by serological assays and 789 were reactive for HIV antibody. Only 478 (61%) were confirmed positive by Western blot/immunoblot. There were 465 (97.2%) donations positive for HIV-1, 6 (1.3%) for HIV-2 (monotypic infection) and 7 (1.5%) for HIV-1 and HIV-2 (dual infection). CONCLUSION: We show the presence of HIV-2 infection among the blood donors and the need for incorporating HIV-2 detection also in the NAT systems.

Immune responses to Epstein-Barr virus in individuals with systemic and organ specific autoimmune disorders.

PURPOSE: Autoimmune diseases usually manifest in genetically predisposed individuals following an environmental trigger. There are several viral infections including Epstein-Barr virus (EBV) implicated in the pathogenesis of autoimmune disorders. The aim of this study was to look at the antibody pattern to EBV proteins in the plasma of both systemic and organ specific autoimmune disorders, estimate pro-inflammatory plasma cytokines (IL-8 and TNF-alpha) among these autoimmune patients and compare the observations with those in normal healthy controls. MATERIALS AND METHODS: Samples from 44 rheumatoid arthritis patients, 25 Hashimoto's thyroiditis patients, appropriately age and sex matched healthy controls were tested for EBV IgM antibodies by an immunoblot assay and two cytokines (IL-8 and TNF-alpha) by commercial assays. RESULTS: Among the rheumatoid arthritis patients, 23 (52%) were positive for EBNA1 antibody, while 13 (52%) of the Hashimoto's thyroiditis patients and 12 (30%) of the healthy controls showed similar bands. The intensity of the bands was high in the autoimmune patients when compared to the bands seen in control samples. The difference in the EBNA1 reactivity between rheumatoid arthritis patients and controls were significant (P = 0.038). There was a significant difference in the IgM reactivity to VCAp19 protein between patients and controls (P = 0.011). CONCLUSION: Our study showed an increased EBV activation among the autoimmune patient groups compared to the normal healthy controls. Further studies are required to delineate the association between the aetiology of autoimmune disorders and EBV.

Nanotechnology tools for single-virus particle detection.

The development and potential application of nanotechnology tools for single-virus particle detection by emergent nanotechnology is likely to revolutionize diagnosis and determining treatment endpoints for life threatening virus infections. Direct detection of biological macromolecules using semiconducting nanowires or carbon nanotubes for electrical field change measurements is a milestone application in this field. The promise of selective detection at a single particle level (stochastic sensing) with nanowire or nanotube field-effect transistor-based devices is a major breakthrough for outbreak situations, where a rapid and specific detection of the viral agent allows intervention at public health level. The same technology would be eminently suitable for bedside diagnosis and therapeutic intervention.

Association of mannose-binding lectin polymorphisms and HBV outcome in a South Indian population.

Mannose binding lectin (MBL) is an important innate immune system pattern recognition molecule. The MBL gene polymorphisms are reported to play a crucial role in outcome of hepatitis B virus (HBV) infection. In this study, we ascertained the association of MBL genotypes with HBV outcome in a South Indian population. The MBL gene polymorphisms at codons 52, 54 and 57 of exon I, and promoter polymorphisms at -221 were typed by polymerase chain reaction-sequence specific primer in spontaneously recovered and in chronic HBV group. The allele frequency of codon 52 'C' was significantly higher in chronic HBV group than in the recovered group (98.5% vs. 93.6%; P = 0.003) and codon 52 'T' was significantly higher in recovered group than in the chronic group (6.4% vs. 1.5%; P = 0.003). In multivariate analysis, after adjusting for age, sex and state of origin, codon 52 'CC' and 'CT' genotypes were significantly associated with chronicity and recovery respectively [odds ratio (OR), 0.25; 95% confidence interval (CI), 0.08-0.80, P = 0.02] in co-dominant analyzing models. This was re-affirmed in analysis performed exclusively on Tamil Nadu subjects (OR, 0.23; 95% CI, 0.06-0.93, P = 0.039). The frequency of low/none haplotype (XY/O) was significantly higher in recovered group than in chronic group (15.6% vs 7.5%) and associated with spontaneous recovery (OR, 2.28; 95% CI, 1.04-4.99, P = 0.035). Our results provide preliminary evidence that inheritance of codon 52 genotypes and XY/O haplotype associated with low MBL level substantially determine the outcome of HBV infection in a sympatrically isolated South Indian population.

The frequency of HIV-I drug resistance mutations among treatment-naive individuals at a tertiary care centre in south India.

Antiretroviral treatment (ART) use in India requires information on baseline drug resistance mutations and polymorphisms in the protease (Pr) and reverse transcriptase (RT) genes of HIV-1 strains from treatment-naïve individuals. We report resistance predictor mutations and polymorphisms in the Pr and the RT sequence of non-clade B HIV-1 strains from ART naïve individuals. The genotypic resistance assay was done on 93 treatment-naïve individuals. The sequences were analysed by Stanford HIV drug resistance data for genotypic drug resistance analysis and REGA HIV-1 subtyping tool. Phylogenetic tree was generated with MEGA 4 for quality control. Ninety-two strains belonged to clade C and one to clade A (A1). Amino acid substitutions were seen at positions associated with drug resistance in Pr gene--10, 24, 74 (each 3%) and position 82 (11%). Substitutions were seen at positions 41 (1%), 100 (1%), 101 (6%), 103 (2%), 179 (6%) and 181 (1%) of the RT sequence known to confer drug resistance in clade B. Polymorphisms in HIV-1 pol gene among treatment-naïve individuals were similar when compared with previous data. One strain each had Y181C substitution, T74S and E35G substitutions in the Pr and one had A98G, K101R and L210FL substitutions in RT.

Detection of opportunistic DNA viral infections by multiplex PCR among HIV infected individuals receiving care at a tertiary care hospital in South India.

PURPOSE: Opportunistic viral infections cause increased morbidity and mortality among human immunodeficiency virus (HIV) infected individuals, especially those who are not on antiretroviral treatment. Early diagnosis of these opportunistic viruses will be able to reduce the risk of disease progression with appropriate intervention. MATERIALS AND METHODS: Multiplex PCR was attempted to detect the opportunistic herpes viruses (HSV-1, HSV-2, VZV, EBV, and CMV), adenovirus and polyoma viruses (JC and BK) in three cocktails of PCR reactions. Subsequently, all the viruses detected were quantitated by testing using monoplex real time PCR. Whole blood samples collected between 2006 and 2007 from 68 treatment naïve HIV-1 infected and 30 normal healthy individuals were tested for these eight viruses. Among the 68 HIV-1 infected individuals 35 had CD4+ T cell count less than or equal to 200 while the other 33 had greater than 200 CD4+ T cells. RESULTS: Among the 68 HIV-1 infected individuals, 49 (72%) were positive for EBV, 5 (7%) samples were positive for CMV. All the five CMV positive individuals had CD4+ T cell count of less than or equal to 200 cells/microL. The mean EBV load among the individuals with a CD4+ T cells of less than or equal to 200 cells/microL was 3.88 log(10) while among those with greater than 200 CD4+ T cells it was 3.75 log(10) . The mean CMV load was 6.98 log(10). Three samples were positive for both CMV & EBV. None of the samples was positive for HSV-1, HSV-2, VZV, Adenovirus, JC and BK viruses. CONCLUSIONS: In our study, multiplex PCR based detection system was found useful in detecting opportunistic viruses in HIV infected individuals. Though EBV is the most prevalent opportunistic viral infection among HIV infected individuals, there was no significant association between EBV load, CD4+ T cell counts and HIV-1 virus load. CMV was seen in HIV infected individuals with low CD4+ T cell counts (less than 200 cells/microL).