Studies on diagnostic potential of secretory antigens of Mycobacterium bovis.

Culture filtrate proteins (CFPs) of M. bovis were produced by culturing the organism for a period of 3 weeks. These CFPs elicited a good delayed hypersensitivity (DTH) reaction in cattle and guinea pig. However, CFPs showed least cross-reactivity when tested in guinea pigs sensitized with M. bovis BCG and M. phlei. CFPs were found to be well recognized by T-cells of single intradermal test (SIDT) positive cattle and were also sero-reactive. Lymphocyte transformation assay revealed the culture filtrate antigens as major T-cell antigens. CFPs induced a good interferon gamma release as compared to conventional antigen, PPD, thereby highlighting their diagnostic capabilities.

Comparison of the diagnostic value of local Widal slide test with imported Widal slide test.

A laboratory study comparing the Widal slide agglutination test using local antigens produced by Mekar Jaya Diagnostica (SAT-MJD) with imported antigens (Murex, Abbott) was carried out on 55 sera of typhoid fever patients with positive blood culture and 56 sera of non-typhoid febrile patients. The SAT-MJD antigens consisted of a mixture of 5 different phage-types of S. typhi dominantly found in Indonesia. This study revealed the following results: the diagnostic sensitivity of local and imported antigens was 83.93%, the diagnostic specificity of local antigens was higher than the imported antigens ie 82.14% compared with 64.28%, the diagnostic efficiency of local antigens was 82.88% compared with 73.87% of the imported antigens. The diagnostic positive and negative predictive values of the local antigens were 80.70% and 83.63%, respectively. The imported antigen revealed diagnostic positive and negative predictive values of 69.69% and 80%, respectively. The diagnostic specificity and efficiency of local antigens were significantly different (p < 0.02 and p < 0.05) from the imported antigens. The local antigens have some advantages. There was no variation in within-run and between-day test, compared with a 6.6% variation shown by the imported antigen. The test results obtained 5 minutes after mixing the serum with antigens reduced the possibility of false-positive and false-negative results. The cost of local antigens is lower than the imported antigens. Based on these data, the Widal SAT-MJD has a reliable diagnostic value and can be used in small laboratories, such as primary health centers (Puskesmas).

Serodiagnosis of extra pulmonary tuberculosis using A-60 antigen.

The diagnosis of extrapulmonary tuberculosis including neurotuberculosis is difficult because of the low yield of culture positivity for Mycobacterium tuberculosis (M. tb). Serodiagnosis has emerged as a useful aid to the diagnosis of extrapulmonary tuberculosis. The utility and efficacy of detection of antimycobacterial antibodies to A-60 antigen in serum and/or cerebrospinal fluid (CSF) was analysed in 100 patients-neurotuberculosis-72, abdominal tuberculosis-12 and others-16. The overall positivity rate for the test was 75%. The positivity rate of the test in serum and/or CSF was 79.2% (57 of 72) in neurotuberculosis and 62.5% (10 of 16) for other forms of extrapulmonary tuberculosis. The positivity rate for antimycobacterial antibodies was higher for patients with tubercular meningitis -94.7%. To conclude, testing for antimycobacterial antibodies to A-60 antigen is a useful adjunct in the diagnosis of extrapulmonary tuberculosis especially neurotuberculosis.

DTH responsiveness of HIV-infected Thai adults.

A study was carried out in Thailand to determine the frequency of reactivity to delayed-type hypersensitivity (DTH) skin tests used for the staging of HIV patients in the United States. A four-antigen panel which included tetanus toxoid (1:10), Candida (1:10), mumps and Trichophyton antigens was assessed in 221 adult subjects from across the full immunological spectrum of HIV disease. Complete anergy was found in 38 per cent of 73 subjects with CD4 counts of 0-200 cells/ml and in 6 per cent of 78 subjects with 201-400 cells/ml. Partial anergy (response to 1 of 4 antigens) was found in 26 per cent of the 0-200 cell/ml group and decreased progressively with increasing CD4 cell count. Results suggested that a 3-member recall antigen panel would provide nearly all the clinically useful information gained by the more standard 4-member panel. In conclusion, DTH skin testing was confirmed to provide a method of assessing the integrity of cellular immune function of HIV-infected Thai adults which correlated with disease progression.

Diagnostic utility of estimation of mycobacterial antigen A60 specific immunoglobulins in serum and CSF in adult neurotuberculosis.

An ELISA assay based on mycobacterial antigen A60 (Anda, Biologicals France) was used to detect specific immunoglobulins (IgM, IgA and IgG) in 48 cases of adult neurotuberculosis (24 TBM; 24 Tuberculoma) and in 48 controls (24 diseased controls; 24 healthy controls). Serum was analysed in all the subjects whereas CSF was assayed only in TBM cases and diseased controls. The cut off values used for IgM, IgG and IgA in this study were 1.500 ODI (optical density index) at 1:100 dil, 250 units/ml and 150units/ml respectively in serum; and 1.500 ODI at 1:10 dil, 10 units/ml and 10 units/ml respectively in CSF. The mean titres of all three antibodies were found to be significantly higher in cases as compared to controls. In cases of TBM, in serum, the percentage positivity for IgM, IgG, IgA and combination of IgG or IgA were 41.67, 87.50 87.50 and 95.83 respectively. The corresponding figures in CSF were 62.50, 75.0, 66.67 and 79.16 for IgM, IgG, IgA and 'IgA or IgM' respectively. In tuberculoma cases, in serum, the figures were 37.50, 75.0, 75.0 and 83.33 respectively. Overall, a high sensitivity and specificity were obtained in cases of TBM (Serum: ST = 95.83%: SP = 87.50%; CSF ST = 79.16%. SP = 100%) and Tuberculoma cases (serum: ST = 83.33% SP = 87.50%) employing the combined antibody estimations.

Diagnostic utility of ELISA test using antigen A60 in suspected cases of tuberculous meningitis in paediatric age group.

The aim of the study was evaluation of the utility of ELISA test using antigen A60 for rapid diagnosis of tuberculous menigitis (TBM) in paediatric age group. ELISA test based on mycrobacterial antigen A60 (Anda biological, France) was used to estimate specific IgM and IgG antibodies in the sera and CSF of 20 suspected cases of TBM which were selected on the basis of numerous parameters and were smear negative on concentrated smear of CSF. Sera of 20 Montoux negative healthy children was taken as control by detecting IgM and IgG antibodies to A60 antigen. Response to anti-tubercular treatment was observed in all the suspected cases of TBM. This study showed that specificity for diagnosis of TBM by detecting IgM and IgG antibodies in sera was 90% and 80% respectively. Sensitivity of the test by detecting IgM and IgG antibodies in sera was 85% and 80% respectively with positive predictive value of 89.47% for IgM antibody and 80% for IgG antibody. In CSF IgM and IgG antibodies were found in 75% and 60% cases respectively. Both were positive only in 60% of cases. It is concluded from this study that 80-85% cases of TBM in paediatric age group have eigher IgM or IgG antibodies in sera whereas 60-75% have antibodies in CSF.

Sensitivity of EIA in the diagnosis of tuberculosis using 38-kDa antigen.

Patients, who reported to Government Medical College Hospital, Chandigarh with suspected signs and symptoms of pulmonary tuberculosis, were examined. Their sputum was investigated for the presence of acid-fast bacilli, using the routine Ziehl-Neelsen staining, for 3 consecutive days. The positive cases were referred for treatment and the rest were referred for supplementary tests like x-ray, Mantoux test, FNAC and the presence of IgG antibodies in their serum against recombinant 38-kDa antigen derived from Mycobacterium tuberculosis, using micro-ELISA plates. It was found that 37.8% of the cases who were negative for acid-fast bacilli found to be positive for the presence of antimycobacterial IgG antibodies with an antibody titre greater than the cut off value in addition to one of the above mentioned positive supplementary finding. Hence, EIA for tuberculosis is more sensitive as compared to the routine acid-fast bacilli staining.

Post-vaccination sensitization with ICRC vaccine.

ICRC vaccine is one of the candidate anti-leprosy vaccines under test in a large scale comparative vaccine in trial. The objectives of the present study was to study the sensitization potential, as measured by Rees' MLSA and lepromin, and reactogenicity of this vaccine preparation in the local population. The study included 368 'healthy' individuals aged 1-70 years. Each individual received either ICRC vaccine or normal saline (control) by random allocation. They were also tested with Rees' MLSA and lepromin-A, 12 weeks after vaccination. Reactions to Rees' MLSA were measured after 48 hours and those to lepromin-A after 48 hours and three weeks. Character and size of local response, at the vaccination site, were recorded at 3rd, 8th and 15th week after vaccination. The results of the study showed that healing of vaccination lesion was uneventful, the mean size of the lesion being 10.3 mm. The mean sizes of post-vaccination reactions, to Rees' MLSA and lepromin (both early and late reactions), were significantly higher in the vaccine group compared to that in the normal saline group; the sensitizing effect attributable to the vaccine was of the order of 3.5 mm, 1.7 mm and 2.2 mm respectively. In conclusion, the study has demonstrated that ICRC vaccine was 'safe' and produced significant sensitizing effect as measured by post-vaccination sensitization to Rees' MLSA and lepromin, in the local population.

ELISA for diagnosis of tuberculosis.

Antigen A 60 of Mycobacterium bovis is a cell wall antigen with almost complete cross reactivity with antigen A 60 of Mycobacterium tuberculosis. This antigen was used for the ELISA technique for 100 cases suspected of childhood tuberculosis to evaluate the usefulness of the method in the diagnosis. The positivity rate in pulmonary tuberculosis was 65.1%, in central nervous system tuberculosis 50.0% and in the miscellaneous group 76.4%. The overall positivity was 61.0%, proving the utility of the method for diagnosis.