Significance of circulating toxin and antitoxin in unimmunized tetanus cases of neonates and infants.

The level of circulating tetanus toxin, antitoxin and their individual influence on the outcome of tetanus cases were determined in unimmunized 125 neonatal and 39 infant cases of tetanus. PHA (passive haemagglutination) test showed 40% positive cases for toxin while its absence in the remaining cases indicated of either toxin fixation to the central nervous system (CNS) or it got neutralized by antitoxin. TN (toxin neutralization) and PHA test carried out in 46 sera samples revealed a strong positive correlation (r = 0.9) showing that 35/46 (76%) and 38/46 (82.6%) samples were positive for antitoxin, respectively. 25.4% of the neonate and infant cases and 34% of the control group had a protective serum tetanus antitoxin level. 42.5% of the paired sera from unimmunized mothers and their neonates showing nonprotective antitoxin levels suggested that a high level of antitoxin is needed for transplacental transfer, although transfer may not play a decisive role in the resistance against the disease. The presence of toxin or antitoxin in the clinical cases did not affect the outcome of the disease, although in neonates, presence of toxin was found to be a bad prognostic sign. This study explicitly advocates for the need to improve the vaccination coverage strategy.

Genetic analysis of precore/core and partial pol genes in an unprecedented outbreak of fulminant hepatitis B in India.

We investigated an unprecedented outbreak of fulminant hepatitis B virus (HBV) that occurred in Modasa, Gujarat (India) in 2009. Genomic analysis of all fulminant hepatic failure cases confirmed exclusive predominance of subgenotype D1. A1762T, G1764A basal core promoter (BCP) mutations, insertion of isoleucine after nt 1843, stop codon mutation G1896A, G1862T transversion plus seven other mutations in the core gene caused inhibition of HBeAg expression implicating them as circulating precore/BCP mutant virus. Two rare mutations at amino acids 89 (Ile→Ala) and 119 (Leu→Ser) in addition to other mutations in the polymerase (pol) gene may have caused some alteration in either of four pol gene domains to affect encapsidation of pregenomic RNA to enhance pathogenicity. Sequence similarity among patients' sequences suggested an involvement of a single hepatitis B mutant strain/source to corroborate the finding of gross and continued usage of HBV mutant-contaminated syringes/needles by a physician which resulted in this unprecedented outbreak of fulminant hepatitis B. The fulminant exacerbation of the disease might be attributed to mutations in the BCP/precore/core and pol genes that may have occurred due to selection pressure during rapid spread/mutation of the virus.

Correlation of partial env gene sequences with disease progression parameters in HIV-positive pregnant women from India.

Ever since the beginning of the epidemic of HIV, one of the poignant aspects of HIV infection is transmission of the virus from mother to child. It is not known whether pregnancy accelerates the progression of HIV infection from a clinically asymptomatic stage to a progressive clinical phase. Present study was carried out to understand disease progression in pregnant women from India. We studied co-receptor utilization (the major determinant of HIV disease progression), N-glycosylation sites, and sequence variability. Blood samples were collected from 25 HIV sero-positive patients, eleven from the antenatal risk group (experimental group), nine from heterosexual male, and five from heterosexual female risk group (control group). Partial env gene was amplified by PCR and sequenced. BLAST search and phylogenetic analysis were used to determine the subtype. The deduced amino acid sequence of the V3 region was used to predict co-receptor, determine sequence variability and N-glycosylation site. The experimental group comprising the antenatal risk group did not exhibit any difference in terms of co-receptor, N-glycosylation, and sequence variability when compared with the control, non-pregnant group. Pregnancy does not seem to accelerate the clinical course of HIV infection. The female body during the gestation phase possibly acquires certain strategies to impede or at least alleviate the disease progression during the crucial immune-compromised pregnancy phase, which would otherwise adversely affect the mother as well as the fetus during the infection.

Characterization of RPO B gene for detection of rifampicin drug resistance by SSCP and sequence analysis.

PURPOSE: Because of the emergence of multidrug-resistant tuberculosis in recent times, the rapid detection of resistance to the first-line anti-tuberculosis drug rifampicin was felt worldwide. Accordingly, this study was conducted to evaluate the diagnostic potential of polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) for checking its utility as a rapid screening test for determination of rifampicin drug resistance. MATERIALS AND METHODS: A total of 34 isolates of Mycobacterium tuberculosis (M. tuberculosis) (22 rifampicin resistant, 11 rifampicin sensitive and one control H37Rv) strains were analysed by PCR-SSCP and DNA sequencing within the 157-bp region of the rpo B gene (Ala 500-Val 550). RESULTS: Rifampicin resistance was detected successfully by PCR-SSCP in 20/22(90.90%) of rifampicin-resistant strains showing a total of nine different mutations in seven codon positions: codon 513 (CAA-->CCA), 516 (GAC-->GTC), 507 (GGC-->GAC), 526 (CAC-->GAC, TAC), 531 (TCG-->TTG, TGG), 522 (TCG-->TGG) and 533 (GTG-->CCG). Two rifampicin-resistant strains showed an identical PCR-SSCP pattern with the wild type H37Rv; 77.27% rifampicin-resistant strains showed a single point mutation and 9.09% had no mutation. Three rifampicin-resistant strains showed characteristic double mutations at codon positions 526 and 531. Sensitivity and specificity were calculated as 90.90% and 100%. CONCLUSIONS: Rifampicin-resistant genotypes were mainly found in codon positions 516, 526 and 531. PCR-SSCP seems to be an efficacious method of predicting rifampicin resistance and substantially reduces the time required for susceptibility testing from 4 to 6 weeks to a few weeks.

Diagnostic potential of IS6110, 38kDa, 65kDa and 85B sequence-based polymerase chain reaction in the diagnosis of Mycobacterium tuberculosis in clinical samples.

PURPOSE: The correlation between the presence of specific gene sequence of M. tuberculosis and specific diagnosis of clinical tuberculosis is not known. This study compared the results of polymerase chain reaction (PCR) amplification of M. tuberculosis specific DNA sequences (IS6110, 65kDa, 38kDa and mRNA coding for 85B protein) from different clinical samples of pulmonary and extrapulmonary tuberculosis. METHODS: One hundred and seventy-two clinical samples from suspected tuberculosis patients were tested for smear examination, culture (LJ and rapid BACTEC 460 TB system) and PCR. PCR was performed with specific primers for the targets: IS6110, 65 kDa, 38 kDa and 85 B. RESULTS: Each PCR test was found to have a much higher positivity than conventional test and BACTEC culture (P < 0.05). Smear positive samples (56) and the samples (36) showing positive results by conventional methods (smear and LJ medium culture) and BACTEC were found to be positive by all PCR protocols. No significant difference was found between the four PCR protocols (P> 0.05). The primer specific for amplifying the 123bp IS6110 fragment gave the highest positivity (83%), followed by 65kDa, 38kDa and 85B RT-PCR in descending order. CONCLUSIONS: These data suggest that the presence of IS6110 correlates more closely with the diagnosis of clinical tuberculosis than that of 65kDa, 38kDa and 85B proteins.

Protein antigen b (Pab) based PCR test in diagnosis of pulmonary and extra-pulmonary tuberculosis.

BACKGROUND AND OBJECTIVES: Diagnosis of tuberculosis (TB) is largely based on microscopy and culture examination which are either less sensitive, or time consuming. In the present study a PCR (polymerase chain reaction) test based on DNA sequence coding for a 38-kilodalton protein antigen b (Pab) ,specific for Mycobacterium tuberculosis was compared with Ziehl-Neelsen (ZN) stained AFB (acid fast bacilli) smear examination, culture based on conventional Lowenstein-Jensen (LJ) medium and radiometric BACTEC 460 system for the diagnosis of TB using clinical samples obtained from pulmonary and extra-pulmonary cases of TB. METHODS: Clinical samples obtained from 168 patients of suspected TB (pulmonary and extrapulmonary) were subjected to ZN smear examination, LJ culture, radiometric BACTEC culture and a PCR test by amplifying 419 bp sequence coding for Pab, a glycoprotein of molecular weight 38 kDa. RESULTS: A significant difference was seen in the sensitivity of different tests, the figures being 74.2 per cent for PCR test, 53.4 per cent for BACTEC culture, 47.1 per cent for LJ medium based culture and 35.2 per cent for ZN smear examination (P<0.05). However, there was no significant difference between different tests as far as specificity was concerned. PCR test sensitivity in pulmonary and extra-pulmonary clinical samples were 74.3 and 71.5 per cent respectively, being significantly higher (P<0.05) when compared with sensitivity of other tests. The mean detection time for M. tuberculosis was 24.0 days by LJ media culture, 12.8 days by BACTEC culture and less than 1 day by smear examination and PCR test. INTERPRETATION AND CONCLUSION: PCR test is more sensitive than ZN smear examination, LJ medium culture and BACTEC culture for diagnosing TB in pulmonary and extra-pulmonary clinical samples.

Comparison of various microbiological tests including polymerase chain reaction for the diagnosis of osteoarticular tuberculosis.

PURPOSE: To evaluate the utility of the polymerase chain reaction (PCR) test for diagnosing osteoarticular tuberculosis (TB). METHODS: Clinical samples (synovial tissue and synovial fluid) obtained from 23 cases of suspected osteoarticular tuberculosis were subjected to Ziehl Neelsen (ZN) smear examination, radiometric BACTEC culture and PCR test for tuberculosis by amplifying 65 kDa antigen coding region of Mycobacterium tuberculosis (M.tb) genome. RESULTS: PCR test was found to be much sensitive than the ZN smear examination and BACTEC culture (p<0.05) in the diagnosis of osteoarticular TB. In synovial fluid samples, PCR was positive in 73.9%, ZN smear examination in 17.39% and BACTEC culture in 39.13% of cases. The positivities were relatively lower with synovial tissue samples, the corresponding figures being 60.8, 8.6 and 26.08% respectively. Moreover, on combining the results of synovial fluid and tissues, the corresponding figures further increased to 78.2, 21.7 and 43.3% respectively. Further, sensitivity and specificity for PCR employing BACTEC culture as the "gold standard" was 100% respectively. Using BACTEC culture, the earliest positivity was seen in three days using synovial tissue specimen and 13 days with synovial fluid, the average detection times being 23.2 days and 32.6 days respectively. On the other hand, PCR test gave a positive result within 24 hours. CONCLUSIONS: PCR test was shown to be much more sensitive than ZN smear examination and BACTEC culture test for diagnosing osteoarticular tuberculosis.

Comparison of the conventional diagnostic modalities, bactec culture and polymerase chain reaction test for diagnosis of tuberculosis.

PURPOSE: To evaluate the performance of 65 kDa antigen based PCR assay in clinical samples obtained from pulmonary and extrapulmonary cases of tuberculosis. METHODS: One hundred and fifty six samples were processed for detection of Mycobacterium tuberculosis by ZN smear examination, LJ medium culture, BACTEC radiometric culture and PCR tests. RESULTS: A significant difference was seen in the sensitivities of different tests, the figures being 74.4% for PCR test, 33.79% for ZN smear examination, 48.9% for LJ culture and 55.8% for BACTEC culture (P< 0.05). However, there was no significant difference (P>0.05) as far as specificity of different tests was concerned. PCR test sensitivity in pulmonary and extrapulmonary clinical samples were 72.7% and 75.9% respectively and found to be significantly higher (P< 0.05) when compared with those of other tests. The mean detection time for M.tuberculosis was 24.03 days by LJ medium culture, 12.89 days by BACTEC culture and less than one day by PCR test. CONCLUSIONS: PCR is a rapid and sensitive method for the early diagnosis of pulmonary and extrapulmonary tuberculosis.

Comparison of magnetic resonance cholangiography and percutaneous transhepatic cholangiography in the evaluation of bile duct strictures after cholecystectomy.

BACKGROUND: Percutaneous transhepatic cholangiography (PTC) has been the preferred investigation to delineate the anatomy of the biliary tract in a patient with a bile duct stricture after cholecystectomy. Recently magnetic resonance cholangiography (MRC) has been described to evaluate the obstructed biliary tract. This paper reports a comparison of MRC with PTC in evaluating patients with an iatrogenic bile duct stricture. METHODS: This was a prospective study of 26 patients who had surgery for a bile duct stricture after cholecystectomy. Before operation all patients underwent both MRC and PTC, the results of which were compared with the intraoperative findings. RESULTS: Both PTC and MRC were comparable with regard to image quality, detection of intrahepatic bile duct dilatation, assessment of the level of injury and detection of abnormalities such as intraduct calculi, cholangitic liver abscesses and atrophy of liver lobes. MRC provided additional information in four patients, including detection of associated fluid collections (n = 3) and portal hypertension (n = 1). In eight patients more than one puncture had to be performed during PTC to delineate the complete anatomy. CONCLUSION: MRC is an accurate and non-invasive imaging procedure for preoperative evaluation of patients with a bile duct injury after cholecystectomy, and is capable of providing additional information which may not be available with PTC.

Experience of surgical management of pseudo-aneurysms of branches of the coeliac axis in a North Indian Hospital.

BACKGROUND: Bleeding splanchnic artery pseudo-aneurysm is a rare but frequently fatal complication that can be successfully managed by angiographic embolization. However, certain patients because of hemodynamic instability, non-availability of technique or angiographic failure may require primary surgical intervention. METHOD: Retrospective review of 13 patients presenting with exsanguinating hemorrhage from ruptured pseudo-aneurysm arising from branches of coeliac axis, managed surgically in absence of angiographic embolization. RESULTS: Splenic artery was most commonly involved (n = 7) followed by hepatic (n = 3), gastroduodenal (n = 2) and left gastric artery (n = 1). The most common underlying aetiology was pancreatitis (n = 8, acute = 2; chronic = 6) followed by iatrogenic (n = 3), liver abscess (n = 1) and gastric ulcer (n = 1). Seven patients presented with upper gastrointestinal (GI) bleed, while 2 each with lower GI bleed, haemobilia and bleeding through tube drains. CT-scan accurately demonstrated the pseudo-aneurysm in 11 (84.6%) patients and additionally demonstrated the underlying pathology. The surgical management chiefly consisted of ligation of offending vessel and additional procedures directed at primary pathology. Overall, 77% patients had a favourable outcome while 23% died consequent to ongoing hemorrhage. CONCLUSION: Pseudo-aneurysm involving the branches of coeliac axis most commonly arises as a result of pancreatitis and affects splenic artery. CT-scan accurately demonstrates pseudo-aneurysm and associated pathology in majority of cases. Primary surgical management in the presence of hemodynamic instability and non-availability of angiographic embolization is a viable alternative.

Critical states and fractal attractors in fractal tongues: localization in the Harper map.

Localized states of Harper's equation correspond to strange nonchaotic attractors in the related Harper mapping. In parameter space, these fractal attractors with nonpositive Lyapunov exponents occur in fractally organized tongue-like regions which emanate from the Cantor set of eigenvalues on the critical line epsilon=1. A topological invariant characterizes wave functions corresponding to energies in the gaps in the spectrum. This permits a unique integer labeling of the gaps and also determines their scaling properties as a function of potential strength.

Benign signet ring cell change with multilayering in the gallbladder mucosa--a case report.

We describe a case of benign signet ring cell change in the gallbladder mucosa. On histopathological examination of H&E-stained sections, the gallbladder epithelium showed multilayering. The epithelial cells were large, columnar to polygonal with a small round basal or eccentric nucleus and vacuolated cytoplasm, giving them a signet ring appearance. There was no nuclear atypia, hyperchromatism or mitotic activity. The cells showed uniform positivity with mucicarmine, PAS and Alcian blue stains. The cytoplasmic vacuolations were negative for fat stains (Oil red O and Sudan IV). On immunohistochemistry, the cells showed positivity with antibodies for pancytokeratin (PCK) and epithelial membrane antigen (EMA). A diagnosis of benign signet ring cell change with multilayering in the gall bladder mucosa was made. Thoroughly reviewing the literature, we found only one case of benign signet ring cell aggregates in the gallbladder mucosa documented earlier. The lesion is hereby reported because of the unique histomorphology and the diagnostic dilemma which can occur as a malignant change in situ has to be excluded.

Paraganglionoma of extrahepatic biliary tract causing obstructive jaundice.

We report a young woman with paraganglionoma arising from the extrahepatic bile duct presenting with acute obstructive jaundice. The patient underwent excision of the gall bladder and extrahepatic bile duct with the tumor, and Roux-en-Y hepaticojejunostomy. She is asymptomatic 9 months later, with normal biochemical investigations and imaging.

Mirage formation in the thermal region.

Formation of a mirage in the 8-12-µm band has been observed with a high-resolution thermal camera designed and developed by us. Thermal imagery recorded under actual field conditions is presented to illustrate the mirage phenomenon. Further analysis is in progress to estimate the effect of mirages in this waveband for civil and military applications.

Simple determination of microbial protein in rumen liquor.

A method for microbial protein determination based on Lowry's assay was devised. Differential centrifugation was used to separate bacterial cells from rumen liquor. Proteins from cells were released by suspending in .25 N sodium hydroxide and heating in boiling water bath for 10 min. Protein was determined by the Folin phenol method. The method is simple, accurate, reproducible, sensitive, specific, brief, and can be performed with small amount of rumen liquor (5 ml). A large number of samples can be handled conveniently.

Changes in blood constituents of guinea pigs in lantana toxicity.

Lantana toxicity of guinea pigs elicited an increase in hematocrit, erythrocyte and leukocyte number, hemoglobin, urea-nitrogen and bilirubin contents in the blood of the affected animals. Most of the bilirubin was present in the conjugated form. Enzyme activities of glutamic oxaloacetic transaminase, acid phosphatase, lactate dehydrogenase, glutamate dehydrogenase, sorbitol dehydrogenase in the blood plasma of affect animals exhibited a marked increase. Acid phosphatase activity was inhibited by tartrate. Enzyme activity of alkaline phosphatase remained unchanged while that of glutamic pyruvic transaminase showed a marginal decrease.

Chronic aflatoxicosis in cattle.

Pathological, hematological and plasma enzymatic studies were made on milk cattle affected by chronic aflatoxicosis caused by the prolonged feeding of concentrate feed mixtures containing contaminated groundnut cake having aflatoxin B1 (110 ug/kg groundnut cake at the time of sampling), B2, G1 and G2. Clinical and necropsy observations on liver included proliferation of connective tissue along portal triads leaving small group of hepatocytes intact. Liver function tests showed liver damage in three of the four affected animals studied.