Evaluation of three Polymerase chain reaction tests targeting morphological transforming region II, UL-83 gene and glycoprotein O gene for the detection of Human Cytomegalovirus genome in clinical specimens of immunocompromised patients in Chennai, India

BACKGROUND: Human Cytomegalovirus (HCMV) continues to be an important cause of morbidity and occasional mortality in immunocompromised patients. Polymerase chain reaction (PCR) is the most sensitive and commonly used method for the assessment of HCMV infection in the immunocompromised patients at risk from severe associated clinical manifestations. However, there is little consistency in the qualitative PCR used for different regions of HCMV genome. Therefore, the performance of three Qualitative PCR tests to detect HCMV genome in clinical specimens from immunocompromised patients was evaluated. With pp65 antigenemia assay as the "gold standard", nested PCR for morphological transforming region II (mtr II) and glycoprotein O (gO) gene and uniplex PCR for UL 83 gene were applied on 92 consecutive clinical specimens obtained from 74 immunocompromised patients with clinically suspected HCMV disease. Virus isolation was attempted on 12 clinical specimens from six pp65 antigenemia positive patients. Based on the pp 65 antigenemia results as "gold standard", the sensitivity, specificity, positive predictive value and negative predictive value for each PCR was calculated. RESULTS: The PCR targeting mtr II region showed a higher sensitivity (100%) and negative predictive value (100%) than the other two PCRs in detecting HCMV DNA from clinical specimens obtained from different immunocompromised patient population of Chennai region, India. CONCLUSION: The results suggests that the optimal method of detection of HCMV DNA could be achieved by PCR using primer sequences targeting mtr II region of genome of HCMV in Chennai region, India

DRAFT GENOME SEQUENCE OF HUMAN HERPES VIRUS-4 VRF_EBV_01, AN EPSTEIN BARR VIRUS OBTAINED FROM A PEDIATRIC POST TRANSPLANT LYMPHOPROLIFERATIVE DISORDER (PTLD) PATIENT

Objectives: To decode the sequence of Epstein-barr virus (EBV) genome isolated from a pediatric patient with post transplant lymphoproliferative disorder (PTLD).Methods: EBV culture harvested from the blood sample of a 4-year-old patient with post liver transplant lymphoproliferative disorder prior to treatment was subjected to whole genome sequencing using Illumina platform. Generated data were subjected to various quality analysis and the filtered sequences were submitted to NCBI and published under accession number KM269735-KM269744.Results: Annotation results of VRF_ EBV01 genome using Prokka tool and manual blast search infers 48 hypothetical proteins, each 3 genes coding for Epstein-Barr nuclear antigen 3A and 3B and each one gene coding for Epstein-Barr nuclear antigens 1, 4, 4B and DNA primase UL70 protein. Single copy of genes coding for each variant of Tegument protein BSRF1, BLRF2, BGLF2 mir-BART 1,2,3,5,7,12,15,17,20 and two copies of gene coding for primary envelopment factor BFRF1 gene was also found.Conclusion: To date, only eight EBV genome sequences have been reported worldwide and there is no genome sequence reported from India. This study is the first of its kind to report on EBV genome from a post–transplant lymphoproliferative disorder to the scientific community for the welfare of research against EBV diagnostic markers and drug discovery.Â

RESEARCH AND REVIEWS: JOURNAL OF MICROBIOLOGY AND BIOTECHNOLOGY Farewell, Chloramphenicol? Is this True?: A Review

ABSTRACT Chloramphenicol has remained as a potent broad spectrum antibiotic over decades and due to its side effects its usage has been limited. In an era of increasing resistance to many antibacterial agents, chloramphenicol might have a role in the treatment of intra abdominal infections and respiratory tract infections caused by multi drug-resistant pathogens. This review article focuses on the spectrum of activity of chloramphenicol, mechanism of action , side effects , its parentral and oral use, its resistance mechanisms, toxicity elucidated by enteric pathogens against it and also on the possibility of its therapeutic use in treatment of multi -drug resistant bacterial infections

INSIGHTS ON DRUG TARGETING OF TOXOPLASMA GONDII HOST INVASION PROTEINS: A REVIEW

Toxoplasma gondii is an obligate intracellular parasite that infects homoeothermic animals. It is also the major cause of retinochoroiditis in humans.Drugs targeting T. gondii proteins involved in the establishment of host-pathogen interactions is well documented to be an efficient way to combatthe infections. Basically, parasitic invasion of T. gondii occurs by the sequential secretion of apical membrane antigen 1 and rhoptry neck proteins onthe parasite and host cell surfaces, respectively. These proteins operate synergistically and form the moving junction (MJ) complex, thereby, enablingattachment and penetration of the parasite into the host cell. Better understanding of molecular interactions of these proteins is essential to develophighly efficient therapeutic modalities. Hence, by this review it is intended to update the current status of rhoptry and other MJ complex proteins asideal candidates for targeting T. gondii.Keywords: Toxoplasma gondii, Rhoptry proteins, Moving junction complex, Toxoplasmosis

Cost-Effectiveness of Interventions to Prevent Disability in Leprosy: A Systematic Review

Background: Prevention of disability (POD) is one of the key objectives of leprosy programmes. Recently, coverage and access have been identified as the priority issues in POD. Assessing the cost-effectiveness of POD interventions is highly relevant to understanding the barriers and opportunities to achieving universal coverage and access with limited resources. The purpose of this study was to systematically review the quality of existing cost-effectiveness evidence and discuss implications for future research and strategies to prevent disability in leprosy and other disabling conditions. Methodology/Principal Findings: We searched electronic databases (NHS EED, MEDLINE, EMBASE, and LILACS) and databases of ongoing trials (www.controlled-trials.com/mrct/, www.who.int/trialsearch). We checked reference lists and contacted experts for further relevant studies. We included studies that reported both cost and effectiveness outcomes of two or more alternative interventions to prevent disability in leprosy. We assessed the quality of the identified studies using a standard checklist for critical appraisal of economic evaluations of health care programmes. We found 66 citations to potentially relevant studies and three met our criteria. Two were randomised controlled trials (footwear, management of neuritis) and one was a generic model-based study (cost per DALY). Generally, the studies were small in size, reported inadequately all relevant costs, uncertainties in estimates, and issues of concern and were based on limited data sources. No cost-effectiveness data on self-care, which is a key strategy in POD, was found. Conclusion/Significance: Evidence for cost-effectiveness of POD interventions for leprosy is scarce. High quality research is needed to identify POD interventions that offer value for money where resources are very scarce, and to develop strategies aimed at available, affordable and sustainable quality POD services for leprosy. The findings are relevant for other chronically disabling conditions, such as lymphatic filariasis, Buruli ulcer and diabetes in developing countries

Brief Report - Diagnostic Value of Enzyme Linked Immuno-sorbent Assay for Cytomegalovirus Disease

Background: Since interpretation of results of enzyme linked immuno-sorbent assay (ELISA) for diagnosis of Cytomegalovirus (CMV) infection in India is difficult, its diagnostic value required evaluation. Aims: To evaluate the diagnostic value of ELISA against polymerase chain reaction (PCR) in CMV disease. Settings and Design: Results of ELISA test for CMV antibodies in CMV-DNA PCR positive and negative patients and normal healthy blood donors were analysed. Methods and Material: Anti-CMV antibodies were assayed by ELISA on the sera of 26 CMV PCR positive and 21 PCR negative patients and 35 normal healthy blood donors. Statistical analysis: Chi square and Fischer exact test were used for statistical analysis. Results: Anti-CMV antibodies (IgG or IgG and IgM) were present in 20 (76.9%) of 26 PCR positive and 13 (61.9%) of 21 PCR negative patients. ELISA was negative in six (23.1%) of 26 PCR positive patients. Of the 28 paediatric patients, ELISA was positive in 14 (73.7%) of 19 PCR positive and three (33.3%) of nine PCR negative patients showing a statistically significant difference (Chi square test, P value 0.038). Among the 19 patients having complications after organ transplant, ELISA showed anti-CMV antibodies in six (85.7%) of seven PCR positive and 11 (91.7%) of 12 PCR negative patients showing no significant difference. CMV-DNA was not detected in the buffy coat of 35 sero-positive blood donors. Conclusion: ELISA has no diagnostic value in the detection of CMV activation although it may help in the differential diagnosis of CMV infection in the paediatric age group. (J Postgrad Med 2002;48:176-178

Development and application of multiplex polymerase chain reaction for the etiological diagnosis of infectious endophthalmitis

Background: Uniplex polymerase chain reaction (PCR) for detection of bacterial and panfungal genome has been applied onto a large number of intraocular fluids facilitating management of infective endophthalmitis. Aim: To develop and apply a novel, rapid multiplex polymerase chain reaction (mPCR) to detect the presence of eubacterial, Propionibacterium acnes and panfungal genomes in intraocular fluids from patients clinically diagnosed to have infective endophthalmitis. Settings and Design: Prospective study. Materials and Methods: Conventional methods of direct microscopy by KOH/calcofluor mount, Gram\u2032s staining and culture were done on 30 (19 Aqueous humor-AH and 11 Vitreous fluid-VF) intraocular specimens and mPCR done for simultaneous detection of eubacterial, P. acnes and panfungal genomes. Results: mPCR detected an infectious etiology in 18 (60%) of 30 intraocular specimens. Eubacterial genome was detected in 12 (40%) specimens, P. acnes genome in 4 (13.3%) specimens and panfungal genome in 2 (6.6%) specimens. mPCR results correlated with those of uniplex PCR. mPCR results were available within 5-6 hours after receipt of specimen, as against 8 hours required for each uniplex PCR with three separate thermalcyclers for their completion. Consumption of Taq polymerase was reduced considerably for mPCR. Conclusion: mPCR is a cost effective, single tube method for the simultaneous detection of eubacterial, P. acnes and panfungal genomes in intraocular specimens from patients with infective endophthalmitis. It is a more rapid procedure than uniplex PCRs and requires only a single thermalcycler