Biomedical subjects
Publications and source records attributed to A Rattan.
Salmonella typhimurium isolated from an infant and the powder milk as well as from three different epidemiologically unrelated patients had the same biochemical reactions, antibiogram, serotype and all were non-typable by phage typing. The clonal nature of epidemiologically related isolates was established by SDS PAGE analysis.
HBsAg was screened by Reverse Passive Haemagglutination Test (RPHA) and was confirmed by ELISA test in 157 pregnant females and their newborns. Anti-HBc and IgM anti-HBc was done in these cases by enzyme immuno-assay. The overall prevalence of HBsAg in mothers was 16 out of 157 (10 per cent) and in cord blood of newborns 5 per cent. The transplacental transmission was found in eight of 16 (50 per cent) HBsAg positive mothers. Anti-HBc was present in 12 out of 16 (75 per cent) HBsAg positive mothers and, of these, seven (58 per cent) neonates acquired HBsAg infection. IgM anti-HBc was present in seven out of eight (88 per cent) HBsAg positive neonates, suggesting active in utero infection. Fourteen out of 16 (88 per cent) neonates born to HBsAg positive mothers were alive and healthy, one was stillborn and one had a congenital anomaly.
It has been reported recently that isoniazid resistant strains of Mycobacterium tuberculosis have lost the katG gene which encodes the catalase-peroxidase enzyme. A 35 mer oligonucleotide probe specific for the katG gene of M tuberculosis, 3' end-labelled with digoxigenin, was constructed and hybridised with DNA extracted from 26 clinical isolates of M tuberculosis under high stringency conditions. Twenty two of these isolates were resistant to 0.2 microgram/ml isoniazid and 20 to 1.0 microgram/ml isoniazid. Semiquantitative detection of catalase did not show any discrimination between isoniazid sensitive and resistant strains. The katG gene was present in all clinical strains of M tuberculosis. Therefore, complete deletion of the katG gene does not seem to be the mechanism of isoniazid resistance in M tuberculosis strains isolated from patients in India.
Two groups of 4-5 week old DBA/2J Nii mice were put on either a yogurt-based (n = 33) or a milk-based (n = 32) diet for a period of 4 weeks. At the end of the feeding trial one sub group of mice each from the two dietary groups was sacrificed for assessment of immune response. The remaining mice were challenged intragastrically with 2 x 10(10) live Salmonella typhimurium organisms and continued on their respective diets for 8 days after which they were also sacrificed. The immune response was measured by tritiated thymidine uptake by splenic or intestinal lymphocytes in response to the mitogens concanavalin A (Con A), Phytohaemaggutinin (PHA), and Lipopolysaccharide from Escherichia coli (LPS). Serum Immunoglobulin A levels were also estimated. Feed efficiency, measured as weight gain per unit energy intake, was significantly higher for the yogurt diet than for the milk diet. The mitogenic response of splenic and intestinal lymphocytes in the two groups of unchallenged mice was not different. In the Salmonella-challenged mice the stimulation index (SI) of splenic lymphocytes from yogurt-fed mice (mean +/- SD) was significantly higher (P = 0.001) in response to Con A (24.71 +/- 3.40) than that of milk-fed mice (15.85 +/- 2.09). Further, in these mice the SI of intestinal lymphocytes from yogurt-fed mice was higher than that of milk-fed mice in response to Con A (7.35 +/- 0.61 vs 5.65 +/- 0.78, P = 0.016) and LPS (9.04 +/- 0.93 vs 6.15 +/- 1.32, P = 0.016). Serum IgA levels in Salmonella-challenged mice were significantly higher 8 days after the challenge in the yogurt-fed group than in the milk-fed group (P < 0.001). The experiments indicate an improvement in local gastrointestinal as well as systemic immunity on a yogurt diet as compared to a milk diet.
Forty two strains of Pseudomonas aeruginosa isolated from bronchoalveolar lavage fluid from intubated patients admitted to the Intensive care unit in AIIMS between December 1993 to June 1994 were included in the study. After obtaining typical biochemical profile, antimicrobial susceptibility was performed against ceftazidime, amikacin, gentamicin, ampicillin, cefotaxime and ciprofloxacin. Pyocin typing of these 42 strains was performed by scrape and streak method using 22 indicator strains. Forty strains could be typed showing excellent discrimination but on repeated testing the group designation changed indicating that the system had low reproducibility. SDS-PAGE of whole cell protein profile indicated the presence of 45 protein bands of different molecular weights, individual isolates had 37 to 42 protein band ranging in molecular weight from 340 kDa to 14.3 kDa. On the basis of Dice index of similarity the strains could be grouped into 20 types. Since all strains could be typed, the system has adequate typability. Similar results were obtained on repeated testing indicating good reproducibility.
The application of molecular biology methods has contributed significant insight into the epidemiology of infections caused by Mycobacterium tuberculosis. However, most of the techniques remain out of reach of laboratories in the tropics. We describe here a simple method for extraction of DNA from the pathogen, a first step in the application of molecular biology tools.
Circulating immune complexes (CIC) were estimated in 31 cases of head and neck malignancies by polyethylene glycol (PEG) precipitation and latex agglutination inhibition (LAI) techniques. Results were compared with 25 age and sex-matched control volunteers. Seropositivity for CIC by PEG precipitation test was 54.83% compared to 61.29% by LAI test. No positive case was detected in control group. Seropositivity for CIC by combination of both test results was 67.74%. In stage I cancer seropositivity for CIC by both techniques was 33.33%. In stage II it was 36.36% by PEG precipitation test and 45.45% by LAI test. In stage III it was 64.28% by PEG precipitation test and 71.42% by LAI test. In stage IV all cases were seropositive by both techniques. LAI test was more sensitive in CIC detection in cancer stages II and III. PEG precipitation test had the advantage of detecting quantitative CIC levels by PEG index. Follow-up in 10 postoperative cases revealed significant decline in CIC levels by both tests (p < 0.01) after 3 months of surgery; 90% patients became seronegative for CIC by PEG precipitation technique while 80% became seronegative by LAI technique. However, 15 patients were followed up after 3 1/2 years of surgery; 75% patients remained seronegative while 25% patients became seropositive by both tests and manifested clinical recurrences.
We assessed the utility of bronchoalveolar lavage (BAL) in the diagnosis of pulmonary tuberculosis (PTB) in 50 consecutive HIV-negative patients with clinical and radiographic findings suggestive of PTB, but with negative microscopy for acid-fast bacilli (AFB) on sputum smear. Patients were grouped, using a scoring system, into relative likelihoods of having PTB (I-IV, in descending probability). Patients were started on anti-tuberculosis treatment according to the BAL results. Bacteriological diagnosis of PTB was confirmed in 22/50 BAL; 11 (91.6%), seven (37%) and four (40%) of groups I-III, respectively. In 13 cases, an early diagnosis of PTB was made by positive microscopy for AFB on BAL; an alternative diagnosis was made in six cases (bacterial pneumonia 4, carcinoma 2). A decision analysis model was created to assess the overall utility of BAL. This suggested that in a region of high PTB prevalence, and when the clinical diagnosis of PTB is likely, empirical treatment is the best course of action, with BAL being reserved for further investigation of non-responders. Early BAL should be considered when the diagnosis of PTB is uncertain.
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AIMS: To evaluate a safe sputum processing method for detection of tuberculosis in developing countries. METHODS: A sample processing method was developed in which acid fast bacilli were killed with 1% sodium hypochlorite and concentrated by flotation on a layer of xylene before staining by the Ziehl Neelsen or auramine O methods. RESULTS: Best results were obtained by auramine O staining after flotation. Staining by the Ziehl Neelsen method after flotation gave better results than direct Ziehl Neelsen staining without flotation. CONCLUSIONS: The flotation method with Ziehl Neelsen staining offers advantages for smear preparation in the tuberculosis control programmes of developing countries.
The first case of infection with Group B meningococcus in India is reported. The patient was a 4-month-old boy who presented with meningitis and died within 6 h of admission. Gram stain of CSF showed meningococci and latex particle agglutination test on CSF was strongly positive for Neisseria meningitidis serogroup B. The CSF was also positive for meningococcus by polymerase chain reaction using primers NM1 and NM6, which amplify a 650 bp region of the dihydropteroate synthase (dhps) gene of N. meningitidis.
We have compared the in vitro activity of commonly used aminoglycosides--Gentamicin and Amikacin with Ciprofloxacin to determine whether the latter has any role in the empiric treatment of serious bacterial infections. Our results indicate that ciproloxacin was comparable or had a better activity than gentamicin or Amikacin against clinical isolates. As it can be administered orally, hospitalization can be avoided in a large number of patients.
The partial nucleotide sequence of a recombinant plasmid containing the 23S rRNA gene of Mycobacterium tuberculosis was determined and an assay was developed for amplifying 23S rRNA gene sequences of mycobacteria. The PCR-based non-radioactive test enabled us to distinguish Mycobacterium from other closely related genera and was sensitive enough to detect 2 bacterial genome equivalents. The assay was extended to the detection of mycobacterial DNA in uncultured clinical specimens; 23S rRNA sequences were detected in thirty four of forty eight (70.8%) sputum and cerebrospinal fluid (CSF) specimens by the PCR assay, whereas direct smear examination and culture methods demonstrated a positivity rate of 29.2% and 16.7% respectively for the same specimens. A RNA-based PCR assay with a detection limit of 1 genome equivalent was also developed. These PCR assays should prove useful for the early and rapid detection of mycobacterial infection in uncultured clinical specimens.
A sandwich ELISA to detect specific protein antigens of Mycobacterium tuberculosis was developed by using polyclonal anti-BCG rabbit antibodies as the primary capture antibodies. The mycobacterial antigens were detected with horseradish peroxidase conjugated monoclonal antibodies (P 6) as secondary antibodies. The enzyme was detected by using 1,2 Phenylenediamine dihydrochloride (OPD) and Hydrogen peroxide as substrate. The antigen could be quantitated through linear regression analysis with lower detection limit of 1.25 micrograms/ml. 50 consecutive cases of infertility were examined by laparoscopy and tested for the presence of the antigen in the serum. Mycobacterial antigen could be detected in 9 of the 12 cases with definitive diagnosis of tuberculosis, 5 of the 23 where the diagnosis of tuberculosis was probable and in only 1 of 15 patients who had no laparoscopic abnormalities indicative of tuberculosis.
Clinicobacteriological profile of omphalitis neonatorum was analysed in this prospective study which comprised 4776 neonates (4410 hospital-born, 366 deliveries at home). The incidence of omphalitis in the hospital-born babies was 2.3%. About 21.3% babies delivered at home were admitted for neonatal sepsis, meningitis, birth asphyxia, etc. They were found to be concomitantly suffering from omphalitis. Improper severing of the umbilical cord, application of oily substances on the umbilical stump and unhygienic rearing practices during neonatal period were some of the important predisposing factors. The fall of the umbilical stump and the diagnosis of omphalitis neonatorum was made significantly earlier (p < 0.001) in the hospital-born babies and none of them developed sepsis. The institution of therapy for umbilical sepsis was considerably delayed in the babies delivered at home and the omphalitis was the probable cause of sepsis in 46.6% cases. The Gram-negative organisms were responsible for omphalitis in 57.1% cases. Klebsiella was the commonest Gram-negative organism. Its incidence was more among the babies delivered at home signifying a potentially infective environment in the community. Gram's stain was a reliable and easy method for grossly identifying the organism in the umbilical smear.
This is the first report of Pneumocystis carinii infection seen in patients with AIDS in India. Two of the three patients were of Indian origin while the third was an American tourist. This report indicates that the technical problem of demonstrating P carinii may be an important factor underlying the diagnosis of PCP.
Seven clones of murine monoclonal antibodies specific for group-A streptococci were generated. All of them were of IgM isotypes and recognized trypsinized as well as nontrypsinized group-A streptococci and polysaccharide. These were devoid of reactivity with streptococci-B, -C, -G and Staphylococcus aureus. Polyclonal antibodies against group-A polysaccharide (APS) were also raised in rabbits by linking APS to bovine serum albumin, and rendered monospecific by adsorption. Latex agglutination assays were developed employing both types of antibodies. The assay employing monospecific polyvalent antibodies had a sensitivity of 12.5 ng APS/ml as compared to 1 microgram APS/ml for latex sensitized with monoclonal antibodies. Both assays were specific, as no agglutination was observed with polysaccharides obtained from streptococci-B, -C, -G, Staph. aureus, Cornybacterium diptheriae, Candida albicans, Candida spp., Morexella catarrhalis, Klebsiella pneumoniae, Pseudomonas aeruginosa, Escherichia coli, Streptococcus agalactiae, Strep. pneumoniae. Salmonella typhi and S. paratyphi A and B. Throat swabs from children obtained in duplicate, when tested for the presence of streptococci-A, showed a good correlation of the results obtained by latex agglutination assay with the microbial culture test and serogrouping.
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