The Importance of Serial Time Point Quantitative Assessment of Cardiac Troponin I in the Diagnosis of Acute Myocardial Damage.

OBJECTIVE: The present study was aimed to establish a threshold value for cardiac troponin I (cTnI) for nonacute coronary syndrome (ACS) participants from the local population and also to determine the importance of serial time point estimation of cTnI in acute myocardial infarction (AMI), non-ST-elevated MI (NSTEMI), and unstable angina cases. METHODS: The present study included 194 cases, admitted in ICCU with the complaint of anginal pain; 31 were diagnosed with AMI with typical electrocardiography (ECG) changes; whereas, 48 cases were diagnosed with NSTEMI. The latter group of cases was selected for the time point study of cTnI release at 0-4 h, 6-12 h, 72 h, and 144 h of admission. cTnI levels were assessed using the Abbott ARCHITECT i1000SR system. RESULTS: ACS was clinically ruled out in 98 cases, and cTnI level for them was used to decide cTnI threshold for the non-ACS group. cTnI level was checked in 17 cases of unstable angina. The threshold value of cTnI for non-ACS participants was 0.1 ng/ml and can be considered as cut-off value for the regional population. The data suggested that the peak of cTnI levels in most of the AMI cases reached during 6-12 h. The cTnI levels were lower than 0.1 ng/ml, and no significant change in ECG was noticed in 17 cases of unstable angina. CONCLUSION: The present study suggested that the repeat of cTnI assay after 4-6 h of admission is required if the initial value is <3 ng/ml.

Cardiac troponin I: A potent biomarker for myocardial damage assessment following high voltage electric burn.

Myocardial infarction (MI) following high voltage electric burn is very rare, and its pathogenesis remains controversial. Electrical burns represent only 4% of all burns. Hence, clinical managements have taken a slow pace in developing. The recent guidelines laid down by the cardiology societies include cardiac troponin I (cTnI) as the gold standard marker for the assessment of myocardial damage assessment. Two patients were admitted to our hospital at the different time with the same kind of high voltage electric burn. Both patients had complained with chest discomfort during admission, and cardiac parameter assessment was done for both the patients. cTnI was also measured for both patients, and marked increase in the values was seen within 5 h of onset of myocardial damage and got into normal range within 72 h. Myocardial damage following electric burn needs to be suspected and assessed as early as possible. Hence, cTnI should be the valuable tool to detect the severity of myocardial damage incurred in the electric burn cases.

In vitro activity of daptomycin & linezolid against methicillin resistant Staphylococcus aureus & vancomycin resistant enterococci isolated from hospitalized cases in Central India.

BACKGROUND & OBJECTIVES: Growing incidence of methicillin resistant Staphylococcus aureus (MRSA) and vancomycin resistant enteroccoci (VRE) is posing a therapeutic problem due to limited drug options. Therefore, the present study was undertaken to check susceptibility of MRSA and VRE isolates against new antimicrobials such as daptomycin and linezolid. METHODS: A total of 586 Gram-positive isolates comprising 442 S. aureus and 144 enterococci isolated from hospitalized cases included in the study, were subjected to in vitro antimicrobial susceptibility testing by disc diffusion method. One hundred twenty four enterococci obtained from rectal swabs of neonates were also included. Minimum inhibitory concentration (MIC) was determined for daptomycin, linezolid, vancomycin and teicoplanin against 50 each isolates of MRSA and VRE by E strip. RESULTS: Among the staphylococci, 326 (73.85%) isolates were MRSA. MIC for vancomycin and teicoplanin among MRSA was ≤ 3 µg/ml. MIC for daptomycin among MRSA was found to be in the range of 0.064-1.5 µg/ml. Percentage of VRE among clinical samples was 14.29 per cent while it was 47.06 per cent among enterococci from rectal swabs of neonates. MIC was >256 µg/ml for vancomycin among VRE and was associated with van A genotype. MIC range for daptomycin among VRE was 0.38-3 µg/ml. MIC for linezolid among MRSA and VRE was in the range of 0.25 to 1 and 0.38 -1.5 µg/ml, respectively. INTERPRETATION & CONCLUSIONS: The present study showed a rise in MIC to vancomycin for sizable number of MRSA and growing percentage of VRE at our centre. Daptomycin and linezolid showed 100 per cent activity against MRSA and VRE.

In-house, simple & economical phage technique for rapid detection of rifampicin, isoniazid, ethambutol, streptomycin & ciprofloxacin drug resistance using Mycobacterium tuberculosis isolates.

BACKGROUND & OBJECTIVES: Multiple drug resistance (MDR) among Mycobacterium tuberculosis poses a serious therapeutic problem. Early detection of MDR can be valuable but the conventional drug susceptibility tests take 4-6 wk time after the laboratory isolation of M. tuberculosis. The bacterial phage assay has been reported as a rapid tool for rifampicin susceptibility testing of tubercle bacilli using the suspension of isolated cultures. The present study was aimed to set up a phage assay for testing drug susceptibility to isoniazid (INH), rifampicin, ethambutol, streptomycin and ciprofloxacin in M. tuberculosis isolates. METHODS: Mueller-Hinton broth instead of Middle Brook 7H9 broth was used to make it more economical. The phage assay was compared with the proportion method using 100 M. tuberculosis isolates from pulmonery TB cases. Phage assay results were available in 48 h for rifampicin and streptomycin while 72 h required for INH, ethambutol and ciprofloxacin. The assay was compared with gold standard proportion method. Interpretation of the results was easy and clear. RESULTS: In the present study, sensitivity and specificity of the phage assay when compared to proportion method were in the range of 97 to 100 per cent for all the drugs except for ciprofloxacin for which it was 93 and 96 per cent, respectively. INTERPRETATION & CONCLUSIONS: The phage assay was economic, easy to perform and rapid for the detection of drug resistance in M. tuberculosis isolates with no requirement of expensive equipment. It is within the reach of microbiology laboratories in developing countries having high loads of tuberculosis.

Evaluation of whole blood IFNgamma test using PPD and recombinant antigen challenge for diagnosis of pulmonary and extra-pulmonary tuberculosis.

Quantiferon TB gold (QFT-G) with recombinant antigen cocktail is well evaluated for diagnosis of pulmonary tuberculosis (PTB). However, diagnosis of extra-pulmonary tuberculosis (EPTB) is more difficult due to limitations of conventional techniques. This study compares recombinant antigens based QFT-G and low cost PPD based interferon test for the diagnosis of PTB and EPTB. IFNgamma release, with recombinant antigens and PPD, was assayed by ELISA from 140 cases of EPTB, 100 cases of PTB along with acid fast bacillus (AFB) detection, AFB culture on LJ and MGIT BACTEC. Sensitivity and specificity for QFT-G recombinant antigens was 84.29% and 96%, while for PPD based interferon was 70% and 84% for EPTB group. The sensitivity was far superior to AFB smear and culture for both the antigens. Nine samples were identified as non-tubercular mycobacteria (NTM) in the EPTB group and all were negative for QFT-G, but six of them were positive for PPD based test. Results of the study show that QFT-G using recombinant antigen is sensitive and specific for both PTB and EPTB diagnosis. The PPD based test is economic and offers comparable performance for PTB and EPTB diagnosis and also useful for diagnosis of NTM.

Insights to the diphtheria toxin encoding prophages amongst clinical isolates of Corynebacterium diphtheriae from India.

Diphtheria is a dreadful disease caused by Corynebacterium diphtheriae. Lysogenised bacteriophages carrying toxin gene in C. diphtheriae can make the strain toxigenic. However, such phage disseminates the toxin genes to other strains when it undergoes lytic phase. As little is known about the phage diversity in C. diphtheriae in India, the present study was undertaken to investigate the prophages integrated into the genome of 29 clinical isolates of C. diphtheriae using whole-genome shotgun sequencing. Amongst these isolates, 27 were toxigenic, while 2 were non-toxigenic strains. Of the 27 toxigenic strains, all harbored known phages carrying toxin gene and two other phages with unknown function. However, the two non-toxin strains did not harbour any of the phages in the genome. It is imperative to devise prevention strategies that hinder the dissemination of toxin by prophages, as it may increase the complications of diphtheria post-immunisation.

Role of cancer antigen-125 from pleural & ascitic fluid samples in non malignant conditions.

BACKGROUND & OBJECTIVE: CA-125, an ovarian tumor marker is known to increase in non malignant conditions such as tubercular and non tubercular pleuritis and ascites. We undertook this study to evaluate non-specific rise in CA-125 levels in conditions associated with pleural effusion and ascites and also to understand the mechanism of its secretion. METHODS: CA-125 levels in 38 pleural and 46 ascitic fluid samples from non malignant cases and 10 blood samples from pulmonary tuberculosis cases were estimated by ELISA. The ascitic fluid samples were collected from cases of bacterial peritonitis, tuberculosis, hepatitis, cirrhosis of other aetiology and pleural fluid samples were from cases of tubercular, pyogenic, cardiomegaly and other conditions. RESULTS: Both ascitic and pleural fluid samples (transudative and exudative) showed elevated CA- 125 levels. The CA-125 levels were significantly higher in ascitic fluid samples than in pleural fluid samples. INTERPRETATION & CONCLUSION: Our findings showed that elevated levels of CA-125 in pleural and ascitic fluid could be because of varied aetiologies which need to be ruled out before considering malignancy. Peritoneum has a greater capacity to secrete CA-125 than the pleural epithelium and the secretion occurs following inflammation or mechanical distress. Pulmonary tuberculosis as a closed lesion without involvement of pleural epithelium does not evoke high CA-125 release.

Chronic knee monoarthritis caused by Mycobacterium chelonae.

Extrapulmonary tuberculosis occurs in 20% of all patients with tuberculosis and tubercular arthritis occurs in 10% of those with extrapulmonary tuberculosis. Arthritis caused by Mycobacterium tuberculosis is not uncommon in India. However, arthritis caused by Mycobacterium chelonae has not been reported to the best of our knowledge. We report a patient with arthritis caused by Mycobacterium chelonae in whom the diagnosis was confirmed by smear and culture of acid-fast bacilli. Polymerase chain reaction of the synovial fluid using IS6110 was negative.

Helium-neon and nitrogen laser irradiation accelerates the phagocytic activity of human monocytes.

OBJECTIVE: Intracellular survival of mycobacteria within monocytes is a crucial stage in the pathogenesis of tuberculosis. The aim was to check intracellular survival of Mycobacterium fortuitum within the human monocytes exposed to He-Ne and nitrogen laser irradiation. BACKGROUND DATA: Tuberculosis remains one of the most important infectious diseases for developing countries. Low-level laser therapy (LLLT) has been tried to treat tubercular cavitory lung disease with encouraging results. The in vitro photobiological effect of low level laser radiation on the intracellular mycobacteria needs to be evaluated before we could go for large clinical trials. METHODS: The aliquots of human monocytes from peripheral blood of healthy volunteers and tuberculosis cases were exposed to He-Ne or nitrogen laser beam. The non-irradiated monocytes from the same source served as controls. The monocytes were then challenged with M. fortuitum, and surviving mycobacteria within monocytes were subjected to viable counts. RESULTS: Enhanced killing of mycobacterial cells was seen among monocytes exposed to He-Ne and nitrogen laser irradiation. CONCLUSIONS: He-Ne and nitrogen laser irradiation activates the monocytes to increase intracellular killing of mycobacteria.

In vitro effect of nitrogen and He-Ne laser on the apoptosis of human polymorphonuclear cells from burn cases and healthy volunteers.

OBJECTIVE: The aim of this study was to examine the effect of He-Ne and nitrogen lasers on the apoptosis of PMN in normal versus burn patients. BACKGROUND DATA: Nitrogen and He-Ne laser exposure increases the apoptotic death rate for human macrophages. Inflammation is a major consequence of thermal injury, and polymorphonuclear cell (PMN) infiltration exacerbates inflammatory process through the release of proinflammatory cytokines. The apoptotic death instead of necrotic death of PMN under the situation may help to resolve inflammation. METHODS: Ten healthy volunteers and 10 burn cases (30-50% burn surface) were included in the study. The PMN was separated by dextran sedimentation and density gradient centrifugation before suspending in RPMI-1640 medium supplemented with autologus serum. The cell suspension aliquoted in microwells was exposed to nitrogen (wavelength of 337 nm with power output of 3 mW) and He-Ne (LGN model no. 111, Russia, wavelength of 632.8 nm with power output of 3 mW) lasers for 10 and 5 min. The wells not exposed to laser were used as controls. After 24-36 h of incubation, the apoptotic rates were measured as percentage by morphological studies on acridine orange-ethidium bromide stained preparation using fluorescent microscope. RESULTS: Percentage of apoptotic death increases from 32.9% (SD +/- 4.14) in control PMN to 41.97% (SD +/- 14) in PMN exposed to nitrogen laser for 5 min and further increased to 62.7% (SD +/- 15.11) with nitrogen laser exposure for 10 min. He-Ne laser exposure for 10 min increased apoptotic cell percentage to 41.9%. Increased apoptosis in PMN exposed to nitrogen laser was statistically significant (p < 0.03) both for PMN from healthy subjects and burn cases. It was significantly elevated (p = 0.005) only for PMN from healthy volunteers exposed to He-Ne laser for 10 min but not among He-Ne exposed PMN from burn cases. CONCLUSIONS: These observations support the therapeutic application of nitrogen laser to reduce inflammation and improve wound healing for burn cases.

Modified blood supplemented Lowenstein Jensen medium improves recovery of mycobacteria from sputum samples.

Lowenstein Jensen medium containing 3% human blood in CPDA anticoagulant was compared with plain LJ medium for mycobacterial growth using 565 sputum samples. Mycobacterial growth appeared on both the media in case of 148 samples. However, growth was faster by one week and colony size larger over blood supplemented LJ in 53 of the 145 culture positives. Additional 12 samples which showed no growth on plain LJ could grow only on LJ supplemented with blood. While 3 samples revealed scanty growth on plain LJ alone. The experience suggests that two LJ slants; one plain and the other supplemented with blood be in inoculated routinely to increase speed of growth and recovery of mycobacteria from clinical samples.

A simple and economical in-house phage technique for the rapid detection of rifampin, isoniazid, ethambutol, streptomycin, and ciprofloxacin drug resistance in Mycobacterium tuberculosis, directly on decontaminated sputum samples.

OBJECTIVES: The early detection of drug resistance would be a boon for TB control programs. The aim of the present study was to set up a rapid phage assay for the testing of drug susceptibility of Mycobacterium tuberculosis to rifampin, isoniazid, ethambutol, streptomycin, and ciprofloxacin, directly on decontaminated sputum samples. METHODS: Mueller-Hinton broth was used instead of 7H9 broth to make the method more economical. Vancomycin and polymyxin B were added to the concentrated sputum samples to reduce the bacterial contamination. The phage assay on decontaminated sputum samples was compared with the proportion method using M. tuberculosis isolates from the same sputum samples. RESULTS: Phage assay results were available within 48h for rifampin and streptomycin and within 72h for all the other drugs. In contrast the proportion method required 4-6 weeks from the primary cultures. The sensitivity of the phage assay was in the range of 93% to 100% and specificity in the range of 96% to 100% for all the drugs tested. The interpretation of results was possible for 334 of the 370 (90.3%) acid-fast bacillus (AFB) smear-positive sputum samples by the phage assay. CONCLUSIONS: The phage assay for the detection of drug resistance on direct decontaminated sputum samples is economical, easy to perform, and rapid.

Augmentation in zone of inhibition of cefoperazone/cefoperazone+ sulbactum compares well with the clinical laboratory standard institute standard extended spectrum beta-lactamase detection method as well as the polymerase chain reaction method.

BACKGROUND: With increased prevalence of extended spectrum beta-lactamase (ESBL) in hospital practice globally, reporting of extended spectrum beta-lactamase along with drug susceptibility test is expected from clinical microbiology laboratory. The aim was to evaluate cefoperazone and cefoperazone+sulbactum disc for phenotypic detection of extended spectrum beta-lactamase among E. coli and Klebsiella spp. isolates. METHODOLOGY: A total of 948 clinical specimens were analysed which included 496 E. coli and 392 Klebsiella pneumoniae. For confirmation of extended spectrum beta-lactamase ceftazidime/ceftazidime+clavulanidc acid and cefotaxime/ cefotaxime+clavulanic acid discs were used as recommended by Clinical Laboratory Standard Institute (CLSI). Simultaneously randomly selected 100 isolates, each of E. coli and Klebsiella spp. were identified for extended spectrum beta-lactamase genes coding for the TEM, SHV and CTX by polymerase chain reaction (PCR). The results were compared with cefoperazone/cefoperazone+sulbactum disc diffusion method. RESULT: Phenotypic characterization identified a high extended spectrum beta-lactamase rate. Four hundred out of 496 (80.64%) E. coli and 392 out of 452 Klebsiella spp.(86.7%) were positive for extended spectrum beta-lactamase by the Clinical Laboratory Standard Institute method. The increase in zone size of cefoprazone/cefoperazone+sulbactum (≥ 5 mm) was seen for all the isolates of E. coli and Klebsiella spp. which were confirmed as extended spectrum beta-lactamase by Clinical Laboratory Standard Institute as well as polymerase chain reaction (PCR) method. CONCLUSION: cefoperazone/cefoperazone+sulbactum disc diffusion showed 100% concordance with extended spectrum beta-lactamase detection by ceftazidime/ ceftazidime+clavulanidc cefotaxime/cefotaxime+clavulanic acid disc diffusion method and polymerase chain reaction.

Evaluation of real-time polymerase chain reaction, interferon-gamma, adenosine deaminase, and immunoglobulin A for the efficient diagnosis of pleural tuberculosis.

OBJECTIVE: Pleural tuberculosis (TB) is a diagnostic challenge because of its non-specific clinical presentation and paucibacillary nature. Conventional diagnosis methods have limitations. We evaluated the real-time polymerase chain reaction (PCR), interferon-gamma (IFN-γ), adenosine deaminase (ADA), and immunoglobulin A (IgA). METHODS: We assessed 204 cases: 50 were confirmed pleural TB, 104 were probable pleural TB, and 50 formed the non-TB group. IFN-γ and IgA were measured by enzyme-linked immunosorbent assay and ADA was measured by colorimetric assay. Real-time PCR was carried out using the 16S rRNA sequence, pleural biopsy specimens were submitted to histopathologic examination, pleural fluid culture was undertaken using Lowenstein-Jensen and MGIT-BACTEC, and pleural fluid smears were stained with auramine O. RESULTS: For confirmed and probable pleural TB cases, the area under the curve (AUC) of the receiver operating characteristic (ROC) curve was highest for IFN-γ (0.994 and 0.963, respectively), followed by ADA (0.989 and 0.945, respectively), real-time PCR (0.898 and 0.784, respectively), and IgA (0.817 and 0.784, respectively). For confirmed and probable pleural TB cases, IFN-γ showed the highest sensitivity (98% and 76.9%, respectively), followed by ADA (92% and 73%, respectively), real-time PCR (80% and 57.7%, respectively), and IgA (70% and 57.7%, respectively). With regard to combined positivity, the combination of 'either real-time PCR or IFN-γ' showed the highest sensitivity: 100% in confirmed pleural TB and 96.2% in probable pleural TB. CONCLUSIONS: IFN-γ showed the highest sensitivity as an individual diagnostic test. When a combination of tests was used, positivity of 'either IFN-γ or real-time PCR' appeared valuable for the diagnosis of pleural TB.

Ciprofloxacin therapy for typhoid fever needs reconsideration.

Outbreaks of multidrug-resistant Salmonella enterica serotype Typhi in the Indian subcontinent in the late 1980s resulted in the failure of conventional drugs, and ciprofloxacin became the firstline drug to treat enteric fever. However, reduced susceptibility to ciprofloxacin, reported widely since 1994, has posed a therapeutic difficulty. The aim of the present work was to review the situation of drug resistance among S. enterica serotype Typhi in central India from 1988 to 2005. A minimum inhibitory concentration (MIC) study for ciprofloxacin was carried out by the agar dilution method on 314 stock cultures preserved since 1988. The MIC for ciprofloxacin was < or = 0.125 mg/l for the 50 isolates isolated during 1989-1994, but during 1998-1999, 60% of the 50 isolates showed MIC > 0.125 mg/l, while in 2002-2003, 82.5 % of the 97 isolates had MIC > 0.125 mg/l and 35% had MIC > 1 mg/l (high-level resistance). In 2004-2005, 88.2% of the 77 isolates had MIC > 0.125 mg/l and 15% had MIC > 1 mg/l (high-level resistance). Sixty-four isolates showing MIC > 1 mg/l with the agar dilution method were also checked by Epsilometer test (E-test, AB Biodisk, Solna, Sweden). Based on the data, it is suggested to withdraw ciprofloxacin as a therapeutic agent for enteric fever. Fortunately, multiple drug resistance, with concurrent resistance to chloramphenicol, cotrimoxazole, and ampicillin, which had reached more than 90% in 1990-1991, started declining over the years and was as low as 5.6% in 2004-2005. According to these observations, older drugs such as chloramphenicol, cotrimoxazole, and ampicillin could be recalled to treat enteric fever.