Indu Verma | PGIMER,CHD,India - Academia.edu (original) (raw)

Papers by Indu Verma

Bone tuberculosis is widely characterized by irreversible bone destruction caused by Mycobacteriu... more Bone tuberculosis is widely characterized by irreversible bone destruction caused by Mycobacterium tuberculosis. Mycobacterium has the ability to adapt to various environmental stresses by altering its transcriptome in order to establish infection in the host. Thus, it is of critical importance to understand the transcriptional profile of M. tuberculosis during infection in the bone environment compared to axenic cultures of exponentially growing M.tb. In the current study, we characterized the in vivo transcriptome of M. tuberculosis within abscesses or necrotic specimens obtained from patients with bone TB using whole genome microarrays in order to gain insight into the M. tuberculosis adaptive response within this host microenvironment. A total of 914 mycobacterial genes were found to be significantly over-expressed and 1688 were repressed (fold change>2; p-value ≤0.05) in human bone TB specimens. Overall, the mycobacteria displayed a hypo-metabolic state with significant (p≤0...

Future Microbiology

Background: Disseminated Mycobacterium avium complex (MAC) and Mycobacterium tuberculosis infecti... more Background: Disseminated Mycobacterium avium complex (MAC) and Mycobacterium tuberculosis infections have almost similar clinical presentations but require different therapeutic management. Materials & methods: A duplex PCR was designed based on the sequence variation between the genes encoding catalase-peroxidase (KatG) of M. avium complex and M. tuberculosis, so as to discriminate MAC, M. tuberculosis and mixed mycobacterial (MAC + M. tuberculosis) infections in HIV patients. Results: An accurate, single-step differential diagnosis of disseminated mycobacterial infections in HIV patients was achieved with specific detection of a single band each for M. avium (120 bp) and M. tuberculosis (90 bp) and two bands for the mixed (120 and 90 bp) infections. Conclusion: katG gene-based duplex PCR can facilitate quick differential diagnosis of disseminated MAC and M. tuberculosis infections in HIV patients.

The Indian Journal of Medical Research, 2012

Sir, Sarcoidosis is a granulomatous disorder of unknown aetiology characterized by the presence o... more Sir, Sarcoidosis is a granulomatous disorder of unknown aetiology characterized by the presence of noncaseating granulomas in multiple organs1. Mycobacterial nucleic acids have been demonstrated in sarcoidosis lesions2, and we have reported 50 per cent prevalence of mycobacterial DNA in sarcoidosis samples using PCR for 65 kDa protein gene3. Genes located on the region of difference 1 (RD1) of Mycobacterium tuberculosis such as the 6-kDa early secreted antigenic target (ESAT-6) and the 10-kDa culture filtrate protein (CFP-10) are not shared by BCG strains and most non-tuberculous mycobacteria, and these antigens are specific indicators of M. tuberculosis complex infection4. We hypothesized that if mycobacteria are aetiologically linked to sarcoidosis, the humoral responses against ESAT-6 and CFP-10 should be demonstrable in the serum samples of sarcoidosis patients. Newly diagnosed glucocorticoid-naive cases of pulmonary sarcoidosis [18 patients (8 females); mean (95% CI) age, 34.9 ...

Future Microbiology

Background: Disseminated Mycobacterium avium complex (MAC) and Mycobacterium tuberculosis infecti... more Background: Disseminated Mycobacterium avium complex (MAC) and Mycobacterium tuberculosis infections have almost similar clinical presentations but require different therapeutic management. Materials & methods: A duplex PCR was designed based on the sequence variation between the genes encoding catalase-peroxidase (KatG) of M. avium complex and M. tuberculosis, so as to discriminate MAC, M. tuberculosis and mixed mycobacterial (MAC + M. tuberculosis) infections in HIV patients. Results: An accurate, single-step differential diagnosis of disseminated mycobacterial infections in HIV patients was achieved with specific detection of a single band each for M. avium (120 bp) and M. tuberculosis (90 bp) and two bands for the mixed (120 and 90 bp) infections. Conclusion: katG gene-based duplex PCR can facilitate quick differential diagnosis of disseminated MAC and M. tuberculosis infections in HIV patients.

BackgroundDiabetes is an important risk factor for developing tuberculosis. This association lead... more BackgroundDiabetes is an important risk factor for developing tuberculosis. This association leads to exacerbation of tuberculosis symptoms and delayed treatment of both the diseases. Molecular mechanism and biomarkers/drug targets related to copathogenesis of tuberculosis and diabetes, however, still remains to be poorly understood. In this study, proteomics based 2D-MALDI/MS approach was employed to identify host signature proteins which are altered during copathogenesis of tuberculosis and diabetes.MethodsComparative proteome of human peripheral blood mononuclear cells (PBMCs) from healthy controls, tuberculosis and diabetes patients in comparison to comorbid diabetes and tuberculosis patients was analyzed. Gel based proteomics approach followed by in gel trypsin digestion and peptide identification by mass spectrometry was used for signature protein identification.ResultsTotal of 18 protein spots with differential expression in TBDM patients in comparison to other groups were id...

Scientific Reports

The upsurge of drug resistant tuberculosis is major health threat globally. To counteract, antimi... more The upsurge of drug resistant tuberculosis is major health threat globally. To counteract, antimicrobial peptides are being explored as possible alternatives. However, certain limitations of peptidebased drugs such as potential toxicity, high cost and relatively low stability need to be addressed to enhance their clinical applicability. Use of computer predicted short active motifs of AMps along with nanotechnology could not only overcome the limitations of AMps but also potentiate their antimicrobial activity. Therefore, present study was proposed to in silico identify short antimicrobial motif (Pep-H) of human neutrophil peptide-1 (HNP-1) and explore its antimycobacterial activity in free form and using nanoparticles-based delivery systems. Based on colony forming unit analysis, motif Pep-H led to killing of more than 90% M. tb in vitro at 10 μg/ml, whereas, similar activity against intracellularly growing M. tb was observed at 5 μg/ml only. Thereafter, chitosan (244 nm) and gold nanoparticles (20 nm) were prepared for Pep-H with both the formulations showing minimal effects on the viability of human monocyte derived macrophages (MDMs) and RBC integrity. The antimycobacterial activity of pep-H against intracellular mycobacteria was enhanced in both the nanoformulations as evident by significant reduction in CFU (>90%) at 5-10 times lower concentrations than that observed for free Pep-H. Thus, Pep-H is an effective antimycobacterial motif of HNP-1 and its activity is further enhanced by chitosan and gold nanoformulations. Antimicrobial peptides (AMPs) have great potential to be explored as efficient drug candidates against tuberculosis (TB), the leading cause of deaths amongst infectious diseases across the globe 1. The complex survival strategies employed by mycobacteria to reside successfully inside the host led to usage of lengthy multidrug cocktail regime for more than forty years. Lack of addition of new drugs to the age old therapy and patient incompliance has provided favourable conditions for mycobacteria to evolve into various drug resistant strains and posing great challenges for TB control. Thus, AMPs, essential part of our first line innate immune defense, having ability to not only form cytotoxic pores in bacterial membranes, but also inhibit cell wall, nucleic acid, and protein biosynthesis are appropriate contenders to enhance efficacy of current chemotherapy. Previously our laboratory reported Human neutrophil peptide-1 (HNP-1), a human α-defensin (AMP), as a potential adjunct to existing chemotherapy and also established its specific mode of action against mycobacteria 2-4. However, there are various drawbacks of a peptide drug to be used as a pharmacological agent. The problems that hamper the progress of AMPs as clinical candidates include lack of stability, short in vivo half-life, proteolytic cleavage, toxicity and high cost of manufacturing. Recently, studies have focused on exploring the role of shorter peptides of AMPs as ideal therapeutic candidates against range of infectious agents 5. These shorter

Tuberculosis

Diabetes affects the presentation of tuberculosis including delayed clearance of the bacteria fro... more Diabetes affects the presentation of tuberculosis including delayed clearance of the bacteria from host cells, however, the molecular changes which help survival of phagocytosed mycobacterium in the diabetic host are still not clear. The effect of in vitro high glucose concentrations on the proteome of the phagocytosed mycobacterium isolated from the human monocytic THP1 cell line derived macrophages has been investigated in the present study. Concurrent tuberculosis and hyperglycemia conditions were mimicked by growing M. tuberculosis infected THP1 cells under high glucose conditions. Phagocytosed bacilli were isolated 5 days post infection. Proteomics analysis of the isolated bacilli was done by two-dimensional gel electrophoresis followed by mass spectrometry. A total of 224 ± 18 protein spots were obtained out of which 10 were found to be differentially expressed under high glucose concentrations in comparison to normal glucose concentration. Further, identity of all the ten proteins namely Tgs3, Rv0547c, AcrA1, EsxU, Rv2219, Mpa, Rv2308, ORN, LucA, and Rv1414 was elucidated by peptide mass finger printing using Matrix-assisted laser desorption and ionization-mass spectrometry (MALDI/MS) assisted with MASCOT software. Though Tgs3, Rv0547c, AcrA1 and Mpa proteins have been demonstrated to play a major role in lipid metabolism under nitric oxide stress conditions, the functional role of rest of the differentially expressed proteins remains to be elucidated. Under hyperglycemic conditions in the host cells, differential expression of these proteins might help in the better survival of mycobacteria and can further act as suitable targets to design novel drugs for more effective therapy for comorbid tuberculosis and diabetes.

Allergy & rhinology (Providence, R.I.)

Ever since its characterization in the 1970s, allergic fungal rhinosinusitis (AFRS) has been the ... more Ever since its characterization in the 1970s, allergic fungal rhinosinusitis (AFRS) has been the subject of much controversy, especially regarding its pathogenesis. In this study, we analyzed the differential expression of genes that encode protease-activated receptors (PAR) in patients with AFRS and patients with chronic rhinosinusitis, and tried to understand the pathogenic basis of this disease. To analyze the differential expression of PAR genes in patients with AFRS and in patients with chronic rhinosinusitis. Mucosa from ethmoid sinuses of 51 patients (tests and controls) was biopsied and evaluated for messenger RNA expression of PAR genes by using reverse transcriptase-polymerase chain reaction. Each of the four PAR genes, i.e., par1, par2, par3 and par4 was amplified, the final gene products were run on 1.8% agarose gel and analyzed by densitometry to calculate differential expression. The significance level was determined as p ≤ 0.05. It was observed that the expressions of...

Asian cardiovascular & thoracic annals, 2017

Background Renal dysfunction is a well-recognized major complication after coronary artery bypass... more Background Renal dysfunction is a well-recognized major complication after coronary artery bypass grafting. Off-pump coronary artery bypass theoretically appears to have less impact on renal function. We estimated preoperative and postoperative creatinine clearance as a marker of renal dysfunction in patients undergoing off-pump and on-pump coronary artery bypass. Methods Thirty patients undergoing coronary artery bypass were randomly allocated to undergo either on-pump ( n = 15) or off-pump surgery ( n = 15). The two groups had similar preoperative demographic characteristics. Serum creatinine and creatinine clearance were measured for 4 days postoperatively and the results were compared with preoperative levels. Results The rise in serum creatinine on postoperative day 1 was 0.28 mgċdL(-1) in the on-pump group and 0.22 mgċdL(-1) in the off-pump group ( p = 0.27); on postoperative day 4 it was 0.15 mgċdL(-1) and 0.10 mgċdL(-1), respectively, ( p = 0.28). Similarly, the fall in crea...

PLOS ONE, 2017

Pulmonary tuberculosis, the disease caused by Mycobacterium tuberculosis, still retains a top ran... more Pulmonary tuberculosis, the disease caused by Mycobacterium tuberculosis, still retains a top rank among the deadliest communicable diseases. Sputum expectorated during the disease continues to be a primary diagnostic specimen and also serves as a reservoir of bacteria. The expression pattern of mycobacteria in sputum will lead to an insight into bacterial adaptation at the most highly transmissible stage of infection and can also help in identifying newer diagnostic as well as drug targets. Thus, in the present study, a whole genome microarray of Mycobacterium tuberculosis was used to elucidate the transcriptional profile of mycobacteria in the sputum samples of smear positive pulmonary tuberculosis patients. Overall, the mycobacteria in sputum appeared to be in a low energy and low replicative state as compared to in vitro grown log phase M. tb with downregulation of genes involved in ATP synthesis, aerobic respiration and translational machinery. Simultaneously, downregulation was also seen in the genes involved in secretion machinery of mycobacteria along with the downregulation of genes involved in the synthesis of phthiocerol dimycocerosate and phenol glycolipids. In contrast, the majority of the genes which showed an upregulation in sputum mycobacteria were of unknown function. Further identification of these genes may provide new insights into the mycobacterial behavior during this phase of infection and may help in deciphering candidates for development of better diagnostic and drug candidates.

Indian journal of experimental biology, 2010

Tuberculin skin test (TST), an age old method is based on measuring delayed-type hypersensitivity... more Tuberculin skin test (TST), an age old method is based on measuring delayed-type hypersensitivity (DTH) response to purified protein derivative (PPD). However, inspite of simplicity, ease and cost effectiveness, the usefulness of PPD test is limited due to its inability to distinguish among a protective immune response, latent infection and active tuberculosis disease. On the other hand, a skin test based on RD antigens would add advantages of a high specificity of antigens with the logistics of a skin test. However, except few reports, in vivo data of intradermal use of RD antigens for skin testing is limited. Therefore, in the present study, four M. tuberculosis (Mtb) specific antigens (ESAT6, CFP10, CFP21 and MPT64) were evaluated for their diagnostic utility based on DTH response. These antigens alone and their multiple combinations induced strong DTH response in Mtb infected guinea pigs and the response was negligible in BCG vaccinated and sham immunized animals.

Scientific Reports

The upsurge of drug resistant tuberculosis is major health threat globally. To counteract, antimi... more The upsurge of drug resistant tuberculosis is major health threat globally. To counteract, antimicrobial peptides are being explored as possible alternatives. However, certain limitations of peptidebased drugs such as potential toxicity, high cost and relatively low stability need to be addressed to enhance their clinical applicability. Use of computer predicted short active motifs of AMps along with nanotechnology could not only overcome the limitations of AMps but also potentiate their antimicrobial activity. Therefore, present study was proposed to in silico identify short antimicrobial motif (Pep-H) of human neutrophil peptide-1 (HNP-1) and explore its antimycobacterial activity in free form and using nanoparticles-based delivery systems. Based on colony forming unit analysis, motif Pep-H led to killing of more than 90% M. tb in vitro at 10 μg/ml, whereas, similar activity against intracellularly growing M. tb was observed at 5 μg/ml only. Thereafter, chitosan (244 nm) and gold nanoparticles (20 nm) were prepared for Pep-H with both the formulations showing minimal effects on the viability of human monocyte derived macrophages (MDMs) and RBC integrity. The antimycobacterial activity of pep-H against intracellular mycobacteria was enhanced in both the nanoformulations as evident by significant reduction in CFU (>90%) at 5-10 times lower concentrations than that observed for free Pep-H. Thus, Pep-H is an effective antimycobacterial motif of HNP-1 and its activity is further enhanced by chitosan and gold nanoformulations. Antimicrobial peptides (AMPs) have great potential to be explored as efficient drug candidates against tuberculosis (TB), the leading cause of deaths amongst infectious diseases across the globe 1. The complex survival strategies employed by mycobacteria to reside successfully inside the host led to usage of lengthy multidrug cocktail regime for more than forty years. Lack of addition of new drugs to the age old therapy and patient incompliance has provided favourable conditions for mycobacteria to evolve into various drug resistant strains and posing great challenges for TB control. Thus, AMPs, essential part of our first line innate immune defense, having ability to not only form cytotoxic pores in bacterial membranes, but also inhibit cell wall, nucleic acid, and protein biosynthesis are appropriate contenders to enhance efficacy of current chemotherapy. Previously our laboratory reported Human neutrophil peptide-1 (HNP-1), a human α-defensin (AMP), as a potential adjunct to existing chemotherapy and also established its specific mode of action against mycobacteria 2-4. However, there are various drawbacks of a peptide drug to be used as a pharmacological agent. The problems that hamper the progress of AMPs as clinical candidates include lack of stability, short in vivo half-life, proteolytic cleavage, toxicity and high cost of manufacturing. Recently, studies have focused on exploring the role of shorter peptides of AMPs as ideal therapeutic candidates against range of infectious agents 5. These shorter

Oxidative Stress in Applied Basic Research and Clinical Practice, 2014

The Indian Journal of Medical Research, Mar 1, 2015

Pleural effusion is a common occurrence in patients with late-stage chronic kidney disease (CKD).... more Pleural effusion is a common occurrence in patients with late-stage chronic kidney disease (CKD). In developing countries, many effusions remain undiagnosed after pleural fluid analysis (PFA) and patients are empirically treated with antitubercular therapy. The aim of this study was to evaluate the role of adenosine deaminase (ADA), nucleic acid amplification tests (NAAT) and medical thoracoscopy in distinguishing tubercular and non-tubercular aetiologies in exudative pleural effusions complicating CKD.

Vaccine, 2005

The cytotoxic T-lymphocyte (CTL) responses to culture filtrate antigens of Mycobacterium tubercul... more The cytotoxic T-lymphocyte (CTL) responses to culture filtrate antigens of Mycobacterium tuberculosis H 37 Rv (RvCFP) and purified protein derivative (PPD) were investigated in active pulmonary tuberculosis patients, healthy tuberculosis contacts and non-contacts. Healthy tuberculin skin test (Mantoux) positive tuberculosis contacts demonstrated strong CTL response against RvCFP and Mantoux reactivity was found to correlate with CTL response. The specificity of CTL response in healthy Mantoux positive contacts was further assessed using different molecular weight fractions of RvCFP. Peripheral blood mononuclear cells (PBMCs) derived CTLs recognized multiple antigenic targets and demonstrated predominant cytotoxicity against low molecular weight (below 15 kDa) protein fractions as well as those migrated in the region of 30 kDa. Subsequently, evaluation of CTL responses against selected purified prominent T-cell antigens indicated maximum CTL response directed against Ag85 complex proteins; most notably Ag85A. From this study, it is suggested that identification of more mycobacterial antigens activating various CTL subsets could be an important step for the rational designing of future antituberculous vaccine.

Vaccine, 1997

Six diJg^erent secretory proteins of molecular weights (15,26,30,41, 55 and 70 kDa) were isolated... more Six diJg^erent secretory proteins of molecular weights (15,26,30,41, 55 and 70 kDa) were isolated from 8-day-old culture Jiltrate of Mycobacterium tuberculosis H,,Ra using d@erent column chromatography techniques. These proteins were further examined for their ability to induce cell mediated (T-cell proliferation assay) and humoral immune response (ELISA) in mice immunized with total culture jiltrate proteins. Out of six proteins,

Tropical and Geographical Medicine, 1992

Muria gond tribals (n = 473) from district Bastar, central India, an area known to be hyperendemi... more Muria gond tribals (n = 473) from district Bastar, central India, an area known to be hyperendemic for malaria, were investigated for malarial infection and glucose-6-phosphate dehydrogenase deficiency. The frequency of G-6-PD deficiency was 21.3% among male subjects and 3.7% among females. Assay of malarial antibodies showed that seropositivity as well as the level of antibodies was significantly higher in male subjects with normal enzyme levels as compared with males G-6-PD deficiency. Females with normal G-6-PD enzyme levels too had higher seropositivity as well as level of antibodies against malaria as compared with females having G-6-PD deficiency. This suggests that G-6-PD deficiency correlates with a higher degree of resistance.

Eur Resp J, 2006

The differences in specificity of human lung and peripheral lymphocytes for mycobacterial antigen... more The differences in specificity of human lung and peripheral lymphocytes for mycobacterial antigens (Ag) need to be evaluated in order to identify vaccine candidates against pulmonary tuberculosis (TB).

Indian J Med Microbiol, 2010

Biochemical or nucleic acid based diagnostic techniques for MAC infection are unsatisfactory. Thi... more Biochemical or nucleic acid based diagnostic techniques for MAC infection are unsatisfactory. This study aims to identify and evaluate M. avium secretory protein(s) of diagnostic potential, so as to develop a rapid and simple method for diagnosis of MAC infection. Initially, a specific protein band of approximately 80-85 kDa was recognised by differential immunoblotting; which was subjected to anion exchange column chromatography for purification of proteins. After fractionisation using SDS-PAGE and electroelution, blast search was carried out. Further immunoreactivity studies were done with M. avium and Mtb infected mice sera. Clinical utilisation of separated protein was evaluated by conducting indirect ELISA with serum samples from mycobacterial infected patients. A specific 81.6 kDa protein, shown to be catalase-peroxidase protein (KatG) by blast search was separated. Immunoreactivity studies of purified KatG proteins with mice sera confirmed it to be specific for M. avium infection. Indirect ELISA with patient samples further confirmed it to be M. avium infection specific. KatG protein is specifically recognised by MAC patients and can be used as a marker for simple and rapid ELISA based tests for differential diagnosis of M. avium infection.

The Indian Journal of Medical Research, Jun 1, 2012

Bone tuberculosis is widely characterized by irreversible bone destruction caused by Mycobacteriu... more Bone tuberculosis is widely characterized by irreversible bone destruction caused by Mycobacterium tuberculosis. Mycobacterium has the ability to adapt to various environmental stresses by altering its transcriptome in order to establish infection in the host. Thus, it is of critical importance to understand the transcriptional profile of M. tuberculosis during infection in the bone environment compared to axenic cultures of exponentially growing M.tb. In the current study, we characterized the in vivo transcriptome of M. tuberculosis within abscesses or necrotic specimens obtained from patients with bone TB using whole genome microarrays in order to gain insight into the M. tuberculosis adaptive response within this host microenvironment. A total of 914 mycobacterial genes were found to be significantly over-expressed and 1688 were repressed (fold change>2; p-value ≤0.05) in human bone TB specimens. Overall, the mycobacteria displayed a hypo-metabolic state with significant (p≤0...

Future Microbiology

Background: Disseminated Mycobacterium avium complex (MAC) and Mycobacterium tuberculosis infecti... more Background: Disseminated Mycobacterium avium complex (MAC) and Mycobacterium tuberculosis infections have almost similar clinical presentations but require different therapeutic management. Materials & methods: A duplex PCR was designed based on the sequence variation between the genes encoding catalase-peroxidase (KatG) of M. avium complex and M. tuberculosis, so as to discriminate MAC, M. tuberculosis and mixed mycobacterial (MAC + M. tuberculosis) infections in HIV patients. Results: An accurate, single-step differential diagnosis of disseminated mycobacterial infections in HIV patients was achieved with specific detection of a single band each for M. avium (120 bp) and M. tuberculosis (90 bp) and two bands for the mixed (120 and 90 bp) infections. Conclusion: katG gene-based duplex PCR can facilitate quick differential diagnosis of disseminated MAC and M. tuberculosis infections in HIV patients.

The Indian Journal of Medical Research, 2012

Sir, Sarcoidosis is a granulomatous disorder of unknown aetiology characterized by the presence o... more Sir, Sarcoidosis is a granulomatous disorder of unknown aetiology characterized by the presence of noncaseating granulomas in multiple organs1. Mycobacterial nucleic acids have been demonstrated in sarcoidosis lesions2, and we have reported 50 per cent prevalence of mycobacterial DNA in sarcoidosis samples using PCR for 65 kDa protein gene3. Genes located on the region of difference 1 (RD1) of Mycobacterium tuberculosis such as the 6-kDa early secreted antigenic target (ESAT-6) and the 10-kDa culture filtrate protein (CFP-10) are not shared by BCG strains and most non-tuberculous mycobacteria, and these antigens are specific indicators of M. tuberculosis complex infection4. We hypothesized that if mycobacteria are aetiologically linked to sarcoidosis, the humoral responses against ESAT-6 and CFP-10 should be demonstrable in the serum samples of sarcoidosis patients. Newly diagnosed glucocorticoid-naive cases of pulmonary sarcoidosis [18 patients (8 females); mean (95% CI) age, 34.9 ...

Future Microbiology

Background: Disseminated Mycobacterium avium complex (MAC) and Mycobacterium tuberculosis infecti... more Background: Disseminated Mycobacterium avium complex (MAC) and Mycobacterium tuberculosis infections have almost similar clinical presentations but require different therapeutic management. Materials & methods: A duplex PCR was designed based on the sequence variation between the genes encoding catalase-peroxidase (KatG) of M. avium complex and M. tuberculosis, so as to discriminate MAC, M. tuberculosis and mixed mycobacterial (MAC + M. tuberculosis) infections in HIV patients. Results: An accurate, single-step differential diagnosis of disseminated mycobacterial infections in HIV patients was achieved with specific detection of a single band each for M. avium (120 bp) and M. tuberculosis (90 bp) and two bands for the mixed (120 and 90 bp) infections. Conclusion: katG gene-based duplex PCR can facilitate quick differential diagnosis of disseminated MAC and M. tuberculosis infections in HIV patients.

BackgroundDiabetes is an important risk factor for developing tuberculosis. This association lead... more BackgroundDiabetes is an important risk factor for developing tuberculosis. This association leads to exacerbation of tuberculosis symptoms and delayed treatment of both the diseases. Molecular mechanism and biomarkers/drug targets related to copathogenesis of tuberculosis and diabetes, however, still remains to be poorly understood. In this study, proteomics based 2D-MALDI/MS approach was employed to identify host signature proteins which are altered during copathogenesis of tuberculosis and diabetes.MethodsComparative proteome of human peripheral blood mononuclear cells (PBMCs) from healthy controls, tuberculosis and diabetes patients in comparison to comorbid diabetes and tuberculosis patients was analyzed. Gel based proteomics approach followed by in gel trypsin digestion and peptide identification by mass spectrometry was used for signature protein identification.ResultsTotal of 18 protein spots with differential expression in TBDM patients in comparison to other groups were id...

Scientific Reports

The upsurge of drug resistant tuberculosis is major health threat globally. To counteract, antimi... more The upsurge of drug resistant tuberculosis is major health threat globally. To counteract, antimicrobial peptides are being explored as possible alternatives. However, certain limitations of peptidebased drugs such as potential toxicity, high cost and relatively low stability need to be addressed to enhance their clinical applicability. Use of computer predicted short active motifs of AMps along with nanotechnology could not only overcome the limitations of AMps but also potentiate their antimicrobial activity. Therefore, present study was proposed to in silico identify short antimicrobial motif (Pep-H) of human neutrophil peptide-1 (HNP-1) and explore its antimycobacterial activity in free form and using nanoparticles-based delivery systems. Based on colony forming unit analysis, motif Pep-H led to killing of more than 90% M. tb in vitro at 10 μg/ml, whereas, similar activity against intracellularly growing M. tb was observed at 5 μg/ml only. Thereafter, chitosan (244 nm) and gold nanoparticles (20 nm) were prepared for Pep-H with both the formulations showing minimal effects on the viability of human monocyte derived macrophages (MDMs) and RBC integrity. The antimycobacterial activity of pep-H against intracellular mycobacteria was enhanced in both the nanoformulations as evident by significant reduction in CFU (>90%) at 5-10 times lower concentrations than that observed for free Pep-H. Thus, Pep-H is an effective antimycobacterial motif of HNP-1 and its activity is further enhanced by chitosan and gold nanoformulations. Antimicrobial peptides (AMPs) have great potential to be explored as efficient drug candidates against tuberculosis (TB), the leading cause of deaths amongst infectious diseases across the globe 1. The complex survival strategies employed by mycobacteria to reside successfully inside the host led to usage of lengthy multidrug cocktail regime for more than forty years. Lack of addition of new drugs to the age old therapy and patient incompliance has provided favourable conditions for mycobacteria to evolve into various drug resistant strains and posing great challenges for TB control. Thus, AMPs, essential part of our first line innate immune defense, having ability to not only form cytotoxic pores in bacterial membranes, but also inhibit cell wall, nucleic acid, and protein biosynthesis are appropriate contenders to enhance efficacy of current chemotherapy. Previously our laboratory reported Human neutrophil peptide-1 (HNP-1), a human α-defensin (AMP), as a potential adjunct to existing chemotherapy and also established its specific mode of action against mycobacteria 2-4. However, there are various drawbacks of a peptide drug to be used as a pharmacological agent. The problems that hamper the progress of AMPs as clinical candidates include lack of stability, short in vivo half-life, proteolytic cleavage, toxicity and high cost of manufacturing. Recently, studies have focused on exploring the role of shorter peptides of AMPs as ideal therapeutic candidates against range of infectious agents 5. These shorter

Tuberculosis

Diabetes affects the presentation of tuberculosis including delayed clearance of the bacteria fro... more Diabetes affects the presentation of tuberculosis including delayed clearance of the bacteria from host cells, however, the molecular changes which help survival of phagocytosed mycobacterium in the diabetic host are still not clear. The effect of in vitro high glucose concentrations on the proteome of the phagocytosed mycobacterium isolated from the human monocytic THP1 cell line derived macrophages has been investigated in the present study. Concurrent tuberculosis and hyperglycemia conditions were mimicked by growing M. tuberculosis infected THP1 cells under high glucose conditions. Phagocytosed bacilli were isolated 5 days post infection. Proteomics analysis of the isolated bacilli was done by two-dimensional gel electrophoresis followed by mass spectrometry. A total of 224 ± 18 protein spots were obtained out of which 10 were found to be differentially expressed under high glucose concentrations in comparison to normal glucose concentration. Further, identity of all the ten proteins namely Tgs3, Rv0547c, AcrA1, EsxU, Rv2219, Mpa, Rv2308, ORN, LucA, and Rv1414 was elucidated by peptide mass finger printing using Matrix-assisted laser desorption and ionization-mass spectrometry (MALDI/MS) assisted with MASCOT software. Though Tgs3, Rv0547c, AcrA1 and Mpa proteins have been demonstrated to play a major role in lipid metabolism under nitric oxide stress conditions, the functional role of rest of the differentially expressed proteins remains to be elucidated. Under hyperglycemic conditions in the host cells, differential expression of these proteins might help in the better survival of mycobacteria and can further act as suitable targets to design novel drugs for more effective therapy for comorbid tuberculosis and diabetes.

Allergy & rhinology (Providence, R.I.)

Ever since its characterization in the 1970s, allergic fungal rhinosinusitis (AFRS) has been the ... more Ever since its characterization in the 1970s, allergic fungal rhinosinusitis (AFRS) has been the subject of much controversy, especially regarding its pathogenesis. In this study, we analyzed the differential expression of genes that encode protease-activated receptors (PAR) in patients with AFRS and patients with chronic rhinosinusitis, and tried to understand the pathogenic basis of this disease. To analyze the differential expression of PAR genes in patients with AFRS and in patients with chronic rhinosinusitis. Mucosa from ethmoid sinuses of 51 patients (tests and controls) was biopsied and evaluated for messenger RNA expression of PAR genes by using reverse transcriptase-polymerase chain reaction. Each of the four PAR genes, i.e., par1, par2, par3 and par4 was amplified, the final gene products were run on 1.8% agarose gel and analyzed by densitometry to calculate differential expression. The significance level was determined as p ≤ 0.05. It was observed that the expressions of...

Asian cardiovascular & thoracic annals, 2017

Background Renal dysfunction is a well-recognized major complication after coronary artery bypass... more Background Renal dysfunction is a well-recognized major complication after coronary artery bypass grafting. Off-pump coronary artery bypass theoretically appears to have less impact on renal function. We estimated preoperative and postoperative creatinine clearance as a marker of renal dysfunction in patients undergoing off-pump and on-pump coronary artery bypass. Methods Thirty patients undergoing coronary artery bypass were randomly allocated to undergo either on-pump ( n = 15) or off-pump surgery ( n = 15). The two groups had similar preoperative demographic characteristics. Serum creatinine and creatinine clearance were measured for 4 days postoperatively and the results were compared with preoperative levels. Results The rise in serum creatinine on postoperative day 1 was 0.28 mgċdL(-1) in the on-pump group and 0.22 mgċdL(-1) in the off-pump group ( p = 0.27); on postoperative day 4 it was 0.15 mgċdL(-1) and 0.10 mgċdL(-1), respectively, ( p = 0.28). Similarly, the fall in crea...

PLOS ONE, 2017

Pulmonary tuberculosis, the disease caused by Mycobacterium tuberculosis, still retains a top ran... more Pulmonary tuberculosis, the disease caused by Mycobacterium tuberculosis, still retains a top rank among the deadliest communicable diseases. Sputum expectorated during the disease continues to be a primary diagnostic specimen and also serves as a reservoir of bacteria. The expression pattern of mycobacteria in sputum will lead to an insight into bacterial adaptation at the most highly transmissible stage of infection and can also help in identifying newer diagnostic as well as drug targets. Thus, in the present study, a whole genome microarray of Mycobacterium tuberculosis was used to elucidate the transcriptional profile of mycobacteria in the sputum samples of smear positive pulmonary tuberculosis patients. Overall, the mycobacteria in sputum appeared to be in a low energy and low replicative state as compared to in vitro grown log phase M. tb with downregulation of genes involved in ATP synthesis, aerobic respiration and translational machinery. Simultaneously, downregulation was also seen in the genes involved in secretion machinery of mycobacteria along with the downregulation of genes involved in the synthesis of phthiocerol dimycocerosate and phenol glycolipids. In contrast, the majority of the genes which showed an upregulation in sputum mycobacteria were of unknown function. Further identification of these genes may provide new insights into the mycobacterial behavior during this phase of infection and may help in deciphering candidates for development of better diagnostic and drug candidates.

Indian journal of experimental biology, 2010

Tuberculin skin test (TST), an age old method is based on measuring delayed-type hypersensitivity... more Tuberculin skin test (TST), an age old method is based on measuring delayed-type hypersensitivity (DTH) response to purified protein derivative (PPD). However, inspite of simplicity, ease and cost effectiveness, the usefulness of PPD test is limited due to its inability to distinguish among a protective immune response, latent infection and active tuberculosis disease. On the other hand, a skin test based on RD antigens would add advantages of a high specificity of antigens with the logistics of a skin test. However, except few reports, in vivo data of intradermal use of RD antigens for skin testing is limited. Therefore, in the present study, four M. tuberculosis (Mtb) specific antigens (ESAT6, CFP10, CFP21 and MPT64) were evaluated for their diagnostic utility based on DTH response. These antigens alone and their multiple combinations induced strong DTH response in Mtb infected guinea pigs and the response was negligible in BCG vaccinated and sham immunized animals.

Scientific Reports

The upsurge of drug resistant tuberculosis is major health threat globally. To counteract, antimi... more The upsurge of drug resistant tuberculosis is major health threat globally. To counteract, antimicrobial peptides are being explored as possible alternatives. However, certain limitations of peptidebased drugs such as potential toxicity, high cost and relatively low stability need to be addressed to enhance their clinical applicability. Use of computer predicted short active motifs of AMps along with nanotechnology could not only overcome the limitations of AMps but also potentiate their antimicrobial activity. Therefore, present study was proposed to in silico identify short antimicrobial motif (Pep-H) of human neutrophil peptide-1 (HNP-1) and explore its antimycobacterial activity in free form and using nanoparticles-based delivery systems. Based on colony forming unit analysis, motif Pep-H led to killing of more than 90% M. tb in vitro at 10 μg/ml, whereas, similar activity against intracellularly growing M. tb was observed at 5 μg/ml only. Thereafter, chitosan (244 nm) and gold nanoparticles (20 nm) were prepared for Pep-H with both the formulations showing minimal effects on the viability of human monocyte derived macrophages (MDMs) and RBC integrity. The antimycobacterial activity of pep-H against intracellular mycobacteria was enhanced in both the nanoformulations as evident by significant reduction in CFU (>90%) at 5-10 times lower concentrations than that observed for free Pep-H. Thus, Pep-H is an effective antimycobacterial motif of HNP-1 and its activity is further enhanced by chitosan and gold nanoformulations. Antimicrobial peptides (AMPs) have great potential to be explored as efficient drug candidates against tuberculosis (TB), the leading cause of deaths amongst infectious diseases across the globe 1. The complex survival strategies employed by mycobacteria to reside successfully inside the host led to usage of lengthy multidrug cocktail regime for more than forty years. Lack of addition of new drugs to the age old therapy and patient incompliance has provided favourable conditions for mycobacteria to evolve into various drug resistant strains and posing great challenges for TB control. Thus, AMPs, essential part of our first line innate immune defense, having ability to not only form cytotoxic pores in bacterial membranes, but also inhibit cell wall, nucleic acid, and protein biosynthesis are appropriate contenders to enhance efficacy of current chemotherapy. Previously our laboratory reported Human neutrophil peptide-1 (HNP-1), a human α-defensin (AMP), as a potential adjunct to existing chemotherapy and also established its specific mode of action against mycobacteria 2-4. However, there are various drawbacks of a peptide drug to be used as a pharmacological agent. The problems that hamper the progress of AMPs as clinical candidates include lack of stability, short in vivo half-life, proteolytic cleavage, toxicity and high cost of manufacturing. Recently, studies have focused on exploring the role of shorter peptides of AMPs as ideal therapeutic candidates against range of infectious agents 5. These shorter

Oxidative Stress in Applied Basic Research and Clinical Practice, 2014

The Indian Journal of Medical Research, Mar 1, 2015

Pleural effusion is a common occurrence in patients with late-stage chronic kidney disease (CKD).... more Pleural effusion is a common occurrence in patients with late-stage chronic kidney disease (CKD). In developing countries, many effusions remain undiagnosed after pleural fluid analysis (PFA) and patients are empirically treated with antitubercular therapy. The aim of this study was to evaluate the role of adenosine deaminase (ADA), nucleic acid amplification tests (NAAT) and medical thoracoscopy in distinguishing tubercular and non-tubercular aetiologies in exudative pleural effusions complicating CKD.

Vaccine, 2005

The cytotoxic T-lymphocyte (CTL) responses to culture filtrate antigens of Mycobacterium tubercul... more The cytotoxic T-lymphocyte (CTL) responses to culture filtrate antigens of Mycobacterium tuberculosis H 37 Rv (RvCFP) and purified protein derivative (PPD) were investigated in active pulmonary tuberculosis patients, healthy tuberculosis contacts and non-contacts. Healthy tuberculin skin test (Mantoux) positive tuberculosis contacts demonstrated strong CTL response against RvCFP and Mantoux reactivity was found to correlate with CTL response. The specificity of CTL response in healthy Mantoux positive contacts was further assessed using different molecular weight fractions of RvCFP. Peripheral blood mononuclear cells (PBMCs) derived CTLs recognized multiple antigenic targets and demonstrated predominant cytotoxicity against low molecular weight (below 15 kDa) protein fractions as well as those migrated in the region of 30 kDa. Subsequently, evaluation of CTL responses against selected purified prominent T-cell antigens indicated maximum CTL response directed against Ag85 complex proteins; most notably Ag85A. From this study, it is suggested that identification of more mycobacterial antigens activating various CTL subsets could be an important step for the rational designing of future antituberculous vaccine.

Vaccine, 1997

Six diJg^erent secretory proteins of molecular weights (15,26,30,41, 55 and 70 kDa) were isolated... more Six diJg^erent secretory proteins of molecular weights (15,26,30,41, 55 and 70 kDa) were isolated from 8-day-old culture Jiltrate of Mycobacterium tuberculosis H,,Ra using d@erent column chromatography techniques. These proteins were further examined for their ability to induce cell mediated (T-cell proliferation assay) and humoral immune response (ELISA) in mice immunized with total culture jiltrate proteins. Out of six proteins,

Tropical and Geographical Medicine, 1992

Muria gond tribals (n = 473) from district Bastar, central India, an area known to be hyperendemi... more Muria gond tribals (n = 473) from district Bastar, central India, an area known to be hyperendemic for malaria, were investigated for malarial infection and glucose-6-phosphate dehydrogenase deficiency. The frequency of G-6-PD deficiency was 21.3% among male subjects and 3.7% among females. Assay of malarial antibodies showed that seropositivity as well as the level of antibodies was significantly higher in male subjects with normal enzyme levels as compared with males G-6-PD deficiency. Females with normal G-6-PD enzyme levels too had higher seropositivity as well as level of antibodies against malaria as compared with females having G-6-PD deficiency. This suggests that G-6-PD deficiency correlates with a higher degree of resistance.

Eur Resp J, 2006

The differences in specificity of human lung and peripheral lymphocytes for mycobacterial antigen... more The differences in specificity of human lung and peripheral lymphocytes for mycobacterial antigens (Ag) need to be evaluated in order to identify vaccine candidates against pulmonary tuberculosis (TB).

Indian J Med Microbiol, 2010

Biochemical or nucleic acid based diagnostic techniques for MAC infection are unsatisfactory. Thi... more Biochemical or nucleic acid based diagnostic techniques for MAC infection are unsatisfactory. This study aims to identify and evaluate M. avium secretory protein(s) of diagnostic potential, so as to develop a rapid and simple method for diagnosis of MAC infection. Initially, a specific protein band of approximately 80-85 kDa was recognised by differential immunoblotting; which was subjected to anion exchange column chromatography for purification of proteins. After fractionisation using SDS-PAGE and electroelution, blast search was carried out. Further immunoreactivity studies were done with M. avium and Mtb infected mice sera. Clinical utilisation of separated protein was evaluated by conducting indirect ELISA with serum samples from mycobacterial infected patients. A specific 81.6 kDa protein, shown to be catalase-peroxidase protein (KatG) by blast search was separated. Immunoreactivity studies of purified KatG proteins with mice sera confirmed it to be specific for M. avium infection. Indirect ELISA with patient samples further confirmed it to be M. avium infection specific. KatG protein is specifically recognised by MAC patients and can be used as a marker for simple and rapid ELISA based tests for differential diagnosis of M. avium infection.

The Indian Journal of Medical Research, Jun 1, 2012