IgG response to Mycobacterium tuberculosis non-polar lipids and sonicated extracts among tuberculous meningitis patients.

INTRODUCTION: The diagnosis of tuberculous meningitis (TBM) is a major global health concern due to its protean nature. There is a need to identify better biomarkers for the rapid and definitive diagnosis of TBM. Lipids have been poorly explored as diagnostic markers in TBM. AIM: Non-polar lipids (NPL) and mycobacterial sonicate extract (MTSE) antigens were assessed for diagnosis of Mycobacterium tuberculosis . METHODOLOGY: A total of 110 cerebrospinal fluid samples were categorized as confirmed, suspected and non-TBM cases according to clinical presentation and laboratory investigations, which were further analysed by NPL and MTSE ELISA. RESULTS: The sensitivity and specificity of the NPL ELISA were 39.6 and 96 %, respectively, whereas the MTSE ELISA was 17 % sensitive and 92 % specific. The combination of the NPL and MTSE ELISA test was superior to these tests alone, with sensitivity and specificity of 43 and 88 %, respectively. CONCLUSION: This combination may be useful as an adjunct in the laboratory diagnosis of TBM. However, future studies in different settings among different populations, such as those with human immunodeficiency virus co-infection, are desirable to explore the full potential of biomarkers.

Ultrastructural analysis of cell wall of drug resistant and sensitive Mycobacterium tuberculosis isolated from cerebrospinal fluid by transmission electron microscope.

Drug-resistant tuberculosis is being increasingly recognized and is one among the leading cause of death worldwide. Remarkable impermeability of cell wall to antituberculous drugs protects the mycobacteria from drug action. The present study analyzed the cell wall thickness among first-line drug resistant and sensitive Mycobacterium tuberculosis (Mtb) isolated from cerebrospinal fluid by transmission electron microscopy (TEM). The average thickness of the cell wall of sensitive isolates was 13.60 ± 0.98 nm. The maximum difference (26.48%) in the cell wall thickness was seen among multi-drug resistant (18.50 ± 1.71 nm) isolates and the least difference (4.14%) was shown by streptomycin-resistant (14.18 ± 1.38 nm) isolates. The ultrastructural study showed evident differences in the cell wall thickness among sensitive and resistant isolates. Preliminary TEM examination of cells indicates that morphological changes occur in the cell wall which might be attributed to the drug resistance. The thickened wall of Mtb appears to help the bacilli to overcome the action of antituberculous drugs.

A rapid and simple resazurin assay to detect minimum inhibitory concentrations of first-line drugs for Mycobacterium tuberculosis isolated from cerebrospinal fluid.

OBJECTIVES: Central nervous system tuberculosis (CNS-TB) is a devastating manifestation of TB. The most common form of CNS-TB is tuberculous meningitis. Drug-resistant TB poses a major threat to the control of TB worldwide. Timely treatment dramatically improves the outcome. Colorimetric techniques for drug susceptibility testing based on the oxidation-reduction principle give results quick and are less expensive. The objectives of this study were to compare the susceptibility of Mycobacterium tuberculosis isolated from cerebrospinal fluid to four first-line drugs using the MGIT automated mycobacterial detection system and the resazurin assay (RA) as well as to estimate the minimum inhibitory concentrations (MICs) by RA. METHODS: A total of 42 M. tuberculosis isolates were analysed for their susceptibilities by MGIT and RA. RESULTS: Of the 42 isolates, 35 gave concordant results with both methods. Agreement between the two tests for streptomycin and rifampicin was 100% with a Fleiss' kappa (κ) value of 1, whereas for isoniazid and ethambutol agreement was 92.86% and 90.48%, respectively, with κ values of 0.853 and 0.738. CONCLUSION: The RA appears to be a good alternative to the automated MGIT technique in resource-limited settings.

Nitric oxide in cerebrospinal fluid of central nervous system tuberculosis: correlations with culture, antibody response, and cell count.

BACKGROUND/AIM: The role of nitric oxide (NO) has been established in infection over the years. NO functions by inhibiting the growth of intracellular pathogens. The present study was undertaken to ascertain the role of NO in central nervous system (CNS) infection by Mycobacterium tuberculosis. MATERIALS AND METHODS: A total of 781 chronic meningitis cerebrospinal fluid (CSF) samples suspected of CNS tuberculosis (TB) were categorized based on M. tuberculosis culture positivity, anti-TB antibody response, and CSF cell count and were analyzed for NO. RESULTS: We found that NO levels were positive in 10.88% of the CSF samples. Positivity for NO was 18%, 11.67%, 13.68%, 9.32%, and 9.66% in the cases with mycobacterial culture positivity, anti-TB antibody positivity, high cell count, low cell count, and zero cell count, respectively. Among the above cell count categories, NO levels were noticed to be elevated in high cell count samples with mononuclear cell predominance. CONCLUSION: This study suggests that NO might play some role in the later stages of tuberculous meningitis. This is the first study to our knowledge in which NO was evaluated in CSF in relation to immune response and the presence of a pathogen with such a large number of subjects.

Significance of immune response to Mycobacterium tuberculosis culture filtrate protein antigens in cerebrospinal fluid of tuberculous meningitis patients: A search for diagnostic marker.

Mycobacterium tuberculosis (H37Ra) culture filtrate proteins (CFP) are explored as a diagnostic marker for tuberculous meningitis (TBM). Cerebrospinal fluid (CSF) samples from patients were categorized as confirmed (n = 47), suspected (n = 20), and non-TBM (n = 25) cases. Immune response by Western blot revealed TBM CSF samples are having heterogeneous response to CFP. CFP ELISA was 92% sensitive and 38.30% specific. ODs of confirmed TBM and non-TBM cases were significantly different (P < 0.0001) and also the suspected TBM and non-TBM cases (P = 0.0001). No significant difference noticed in TBM and suspected TBM (P = 0.90). Thus, CFP can be a better biomarker for the diagnosis of TBM.

Correlation of clinical, laboratory and drug susceptibility profiles in 176 patients with culture positive TBM in a tertiary neurocare centre.

Drug resistance has increased the difficulties in control of tuberculosis infection. The present study evaluated the clinical and laboratory features among tuberculous meningitis (TBM) patients and the drug susceptibility of Mycobacterium tuberculosis (M.tb) isolated from CSF. Out of 698 CSF samples, 176 (25.21%) were M.tb culture positive. Among the clinical signs and symptoms, fever, headache and altered sensorium were found to be statistically significant (P<0.05). ELISA was a better predictor of disease and found to be statistically significant (P<0.001) in culture-proven TBM cases. Totally, 57 (32.4%) isolates were resistant to one or more drugs that include 5 (2.8%) multidrug-resistant isolates. In conclusion, the search for antibody in CSF and also CSF chloride can represent as an adjunct in the diagnosis of TBM. Screening of drug susceptibility is a very important factor and would help in better management of the disease.

Immunoconfirmation of central nervous system tuberculosis by blotting: A study of 300 cases.

Tuberculous meningitis (TBM) is a serious form of disease of the central nervous system. Early and accurate diagnosis of the disease and effective treatment are key important factors to contain the disease. The disease presents as chronic meningitis where other partners such as fungal meningitis, neurosyphilis, cysticercal meningitis, carcinomatous meningitis and partially treated pyogenic meningitis share a similar clinical picture making the diagnosis complicated. Culturing of the pathogen Mycobacterium tuberculosis (MTB) from the cerebrospinal fluid (CSF) sample has shown a poor response. The main immunological method for the immunodiagnosis of TBM is the detection of an antibody response in the CSF. In the present study, total MTB sonicated extract antigen was used for ELISA and Western blot. ELISA shows overall immune response of the test sample, whereas Western blotting reveals the specific reactivity to a particular molecular weight antigen. This would also reveal the immunodominant antigen. A total of 300 CSF samples were analyzed by both ELISA and Western blotting. Of the 240 clinically suspected TBM cases, 111 samples were positive by ELISA and 81 samples by Western blot. A total of 76 CSF samples were positive by both ELISA and Western blot. None of the control samples showed positivity either by ELISA or by Western blot. TBM patients revealed major antibody reactivity to 30-40 kD region, followed by 14 kD region. ELISA is sensitive with mild non-specific binding, but Western blot is specific in detecting the immune response. The findings will be useful in definitive immunodiagnosis of TBM.

Probiotic lactic acid bacterium from kanjika as a potential source of vitamin B12: evidence from LC-MS, immunological and microbiological techniques.

Vitamin B(12) was produced by probiotic Lactobacillus plantarum in submerged fermentation (96 h) with successive anaerobic and aerobic phases of 48 h each to give 13 ng vitamin B(12)/g dry biomass. Sodium cyanide-mediated cell lysis, followed by benzyl alcohol/chloroform/water extraction, improved the release of intracellular vitamin B(12) for analysis. The presence of the K(+) adduct of cyanocobalamin (m/z of 1394) was established using electron spray ionization-mass spectra; growth of a mutant of Escherichia coli in the presence of cyanocobalamin ascertained its bioavailability.