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Biomedical subjects

S Ratnam

Publications and source records attributed to S Ratnam.

At least 91 records · Page 5Linked to original sources
PubMed1988

Establishment of human endometrial cell cultures.

Epithelial and stromal endometrial cells from 19 patients at different phases of the menstrual cycle were enzymatically separated, isolated by successive centrifugation and primary cultures established for in-vitro studies on implantation. The behaviour of cells in vitro was evaluated using Nomarski's inverted optics, May-Grunwald-Giemsa stained coverslips and scanning electron microscopy. Epithelial and stromal cells from all patients grew successfully in Chang's medium and formed a mixed confluent monolayer of epithelioid and fibroblastic cells in 3-7 days and such monolayers could be maintained alive up to 3-4 weeks. Epithelioid cells were polyhedral and grew as islands in a whorl-like wavy pattern around glandular fragments. Fibroblasts were spindle-shaped, more long-lived and grew rapidly to form parallel bundles of cells. Significant differences were observed in the number of multinucleated cells and cells with intracytoplasmic vacuoles between endometrium from proliferative, postovulatory and secretory phases (P less than 0.01). Scanning electron micrographs showed cells with cilia with varying densities of microvilli and apical protrusions. Endometrial cells in culture showed structural features remarkably similar to those described for cells in situ. The method described allows the propagation in vitro of separate endometrium cell types which can be used to study implantation mechanisms in unstimulated and stimulated cycles.

Cells, Cultured↗
PubMed1988

Comparison of API ZYM system with API AN-Ident, API 20A, Minitek Anaerobe II, and RapID-ANA systems for identification of Clostridium difficile.

The API ZYM system was compared with four anaerobe identification systems for the definitive identification of Clostridium difficile by using 88 cultures of C. difficile grown on Mueller-Hinton blood agar medium. The API ZYM system yielded a distinct and consistent enzyme profile for all test strains, whereas the sensitivities of the other systems in identifying C. difficile ranged from 78 to 96% (AN-Ident, 77.9%; RapID-ANA, 88.6%; Minitek Anaerobe II, 90.9%; and API 20A, 95.5%). The API ZYM system is highly reliable in identifying C. difficile accurately, is rapid, and is relatively simple to use.

Bacteriological Techniques↗
PubMed1988

Characterization of Escherichia coli serotype O157:H7.

A total of 174 strains of Escherichia coli serotype O157:H7 representing human isolates obtained from outbreaks and sporadic cases of hemorrhagic colitis, hemolytic-uremic syndrome, and nonbloody diarrheal illnesses as well as from asymptomatic carriers across Canada and the United States were examined. E. coli serotype O157:H7 possessed distinct biochemical markers, a 100% negative reaction for beta-glucuronidase and sorbitol, and a 100% positive reaction for raffinose and dulcitol; all strains otherwise were biochemically typical of E. coli. The vast majority (97%) of the strains were susceptible to commonly used antimicrobial agents. All strains produced readily detectable levels of Verotoxin; however, with polymyxin extraction, nearly 50% of the strains showed up to a 10-fold increase in the toxin level. None were found to mediate hemagglutination of human group A erythrocytes with or without D-mannose. The majority (approximately 70%) of the strains showed localized and diffuse adherence to HEp-2 cells and Henle 407 cells, and the adherence patterns were not very different from those observed among other E. coli strains. Twenty phage types were recognized, with phage types 1 and 2 accounting for 65% of the test strains. Plasmid analysis indicated three basic plasmid profiles: profile I was characterized by 68.7- and 4.2-megadalton (MDa) plasmids (62% of strains), profile II was characterized by 66.2- and 1.8-MDa plasmids (20% of strains), and profile III was characterized by a 62.5-MDa plasmid (18% of strains). A small number (19%) of the strains carried at least one additional plasmid over the basic complements, and these could be considered to constitute a miscellaneous category. None of the above-described characteristics of E. coli serotype O157:H7 could be directly correlated with one another, with the nature of infection, or with the geographical distribution of strains.

Bacterial Adhesion↗
PubMed1987

Purification and characterization of dihydrofolate reductase from soybean seedlings.

Dihydrofolate reductase (DHFR; EC 1.5.1.3) was purified to homogeneity from soybean seedlings by affinity chromatography on methotrexate-aminohexyl Sepharose, gel filtration on Ultrogel AcA-54, and Blue Sepharose chromatography. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the enzyme gave a single protein band corresponding to a molecular weight of 22,000. The enzyme is not a 140,000 Da heteropolymer as reported by others. Amino acid sequence-specific antibodies to intact human DHFR and also antibodies to CNBr-generated fragments of human DHFR bound to the plant enzyme on Western blots and cross-reacted significantly in immunoassays, indicating the presence of sequence homology between the two enzymes. The plant and human enzymes migrated similarly on nondenaturing polyacrylamide electrophoretic gels as monitored by activity staining with a tetrazolium dye. The specific activity of the plant enzyme was 15 units/mg protein, with a pH optimum of 7.4. Km values of the enzyme for dihydrofolate and NADPH were 17 and 30 microM, respectively. Unlike other eukaryotic enzymes, the plant enzyme showed no activation with organic mercurials and was inhibited by urea and KCl. The affinity of the enzyme for folate was relatively low (I50 = 130 microM) while methotrexate bound very tightly (KD less than 10(-10) M). Binding of pyrimethamine to the plant enzyme was weaker, while trimethoprim binding was stronger than to vertebrate DHFR. Trimetrexate, a very potent inhibitor of the human and bacterial enzymes showed weak binding to the plant enzyme. However, certain 2,4-diaminoquinazoline derivatives were very potent inhibitors of the plant DHFR. Thus, the plant DHFR, while showing similarity to the vertebrate and bacterial enzymes in terms of molecular weight and immunological cross-reactivity, can be distinguished from them by its kinetic properties and interaction with organic mercurials, urea, KCl and several antifolates.

Amino Acids↗
PubMed1987

Comparative evaluation of commercial enzyme immunoassay kits for detection of hepatitis B seromarkers.

The commercial hepatitis B enzyme immunoassay kits of Abbott Laboratories and Organon Teknika were compared for their sensitivity, specificity, and reproducibility in detecting the hepatitis B seromarkers hepatitis B surface and e antigens and antibodies to hepatitis B core, e, and surface antigens. With the exception of the Organon kit for antibodies to hepatitis B surface antigen, the specificity and reproducibility were about the same for both products, but the level of sensitivity was generally lower for the Organon kits; this, however, may not be critical in routine clinical application. The Organon kits have a longer shelf life and are cheaper.

Costs and Cost Analysis↗
PubMed1987

Simplified acetylcysteine-alkali digestion-decontamination procedure for isolation of mycobacteria from clinical specimens.

During our attempts to reevaluate and improve mycobacteriology laboratory procedures, we found that isolation of mycobacteria from undecontaminated specimens through the use of selective media was unrewarding and that specimen decontamination was an essential step. A reevaluation of the N-acetyl-L-cysteine-sodium hydroxide (NALC-NaOH) decontamination procedure (G. P. Kubica, W. E. Dye, M. L. Cohn, and G. Middlebrook, Am. Rev. Respir. Dis. 87:775-779, 1963) indicated that this method could be simplified and shortened without affecting the isolation rate or growth and without increasing the contamination rate. The modified NALC-NaOH procedure effectively combines the decontamination step with the concentration step: specimens are mixed with the NALC-NaOH solution on a Vortex mixer and, without any waiting or addition of buffer or water to the digested specimens, centrifuged at 3,000 X g for 15 min, and the sediment is suspended in phosphate buffer (pH 5.3). The modified method is simpler, faster, and safer than the original procedure, and the reduced manipulation is also likely to minimize the chance for cross-contamination in sequential specimen processing. The total alkali exposure time in the modified NALC-NaOH method should be kept to a minimum; if necessary or feasible, the concentration of NaOH in the digestion solution may be reduced from the usual strength of 2% to about 1.5% to compensate for the longer alkali exposure time in the modified method to maintain the isolation rate or to improve it.

Acetylcysteine↗
PubMed1987

Characterization of serum, liver, and intestinal sialyltransferases from rats treated with colchicine.

A modified high pressure liquid chromatographic method using lactose (Gal beta 1----4Glc) as an exogenous acceptor has been used to characterize the sialyltransferases known to increase in the serum of colchicine-treated rats. The results show a 10-fold increase of Gal beta 1----4GlcNAc alpha 2----6 sialyltransferase (alpha 2----6 ST), whereas the Gal beta 1----3GlcNAc alpha 2----3 sialyltransferase showed only 1.6-fold increase in the serum after 17 h of colchicine treatment. The sialyltransferase activity in serum using exogenous desialylated, alpha 1-acid glycoprotein as acceptor also showed an eightfold increase. In liver homogenate and Golgi membrane, the sialyltransferase activity when assayed with desialylated alpha 1-acid glycoprotein as acceptor showed a slight decrease after 4 h, but returned to normal level after 17 h. A similar trend was seen when the two transferases were assayed with lactose as acceptor. The antiserum to rat alpha 2----6 ST inhibited the sialyltransferase activity in serum, liver, and jejunal incubation medium. Jejunal sections from rats treated with colchicine for 4 h in presence of heated serum showed a decrease of sialyltransferase, with consequent increase of the alpha 2----6 ST enzyme activity in the medium. This result suggests that intestinal tissue could be a source of increased serum enzyme activity in colchicine treatment.

Animals↗
PubMed1986

Lack of evidence of hepatitis D (delta) infection in Newfoundland and Labrador.

Epidemiologic knowledge of hepatitis D virus (HDV) infection is limited. A seroepidemiologic study was undertaken to determine the prevalence of the infection in Newfoundland and Labrador. Between October 1983 and October 1985 over 200 people were recognized through routine serodiagnosis and screening as having hepatitis B seromarkers. A total of 223 serum samples from 186 of these people were tested for anti-HDV. The subjects were mainly asymptomatic carriers of hepatitis B surface antigen or patients with acute or chronic hepatitis B from the native Indian and Inuit and the non-native populations. None of the serum samples were positive for anti-HDV. The absence of anti-HDV in a substantial number of people in the province who are infected with hepatitis B virus is strong evidence that HDV infection is not prevalent in the local population, including native people.

Acute Disease↗
PubMed1986

Sporadic occurrence of hemorrhagic colitis associated with Escherichia coli O157:H7 in Newfoundland.

During a 9-month period in 1984, 113 fecal samples obtained from 92 patients with diarrheal illness were cultured for Escherichia coli serotype O157:H7 to determine the occurrence of this agent in diarrheal illness in Newfoundland. E. coli O157:H7 was isolated in almost pure culture from 12 stool specimens obtained from 7 (15%) of the 47 patients who had grossly bloody diarrhea; none of the 12 yielded any of the usual enteric pathogens. The agent was not isolated from the stool specimens obtained from the remaining 45 patients, who did not have bloody diarrhea. All seven patients whose specimens were positive for E. coli O157:H7 had clinical manifestations typical of hemorrhagic colitis, but the syndrome was clinically suspected and a specific test requested in only two cases. The seven cases were not clustered geographically or temporally, and no common exposure was identified. To determine whether hamburger meat was the source of E. coli O157:H7, 66 samples obtained from various retail outlets were tested; none were found to be positive. Hemorrhagic colitis may be a common disease, and E. coli O157:H7 should be considered as an agent in bloody diarrheal illness.

Adolescent↗
PubMed1986

Laboratory studies on salmonella-contaminated cheese involved in a major outbreak of gastroenteritis.

A major outbreak of gastroenteritis was traced to Cheddar cheese contaminated with Salmonella typhimurium. There were no significant differences in pH values of the contaminated (mean pH 5.31) and non-contaminated (mean pH 5.39) cheese. The isolation rates of Salm. typhimurium were about the same when cheese samples were homogenized in lactose broth, lactose broth containing 1% Tween 80, or in aqueous 2% sodium citrate. Salmonella typhimurium was isolated regardless of preenrichment in lactose broth, but required selective enrichment in selenite cystine or tetrathionate brilliant green broth. There were no marked differences in the isolation rates obtained with different selective enrichment media, or after incubation at 36 degrees and 43 degrees C for 24 or 48 h. Contaminated samples of cheese failed to yield Salm. typhimurium consistently despite large and multiple samplings; samples from the interior of cheese blocks yielded positive results more frequently than the samples from the exterior. The number of Salm. typhimurium in factory sealed blocks as well as in samples obtained from the homes of known cases of salmonellosis was found to range from less than 3/100 g to 9/100 g of cheese. The infective dose of Salm. typhimurium in contaminated cheese was probably no greater than 10(4) organisms, and a rapid decline in numbers of Salm. typhimurium must have occurred subsequent to the outbreak.

Animals↗
PubMed1986

Effect of relative centrifugal force and centrifugation time on sedimentation of mycobacteria in clinical specimens.

Optimum relative centrifugal force (RCF) and centrifugation time to concentrate mycobacteria in clinical specimens were determined by processing split samples of sputa and urines containing mycobacteria with combinations of different RCFs and centrifugation times. Although individual test results showed considerable variation in the recovery rates of mycobacteria in the sediment, the data indicated that higher recovery rates occurred as centrifugation speed and time were increased. With a 15- to 20-min centrifugation time, on the average, 67 to 71% of mycobacteria were recovered at an RCF of 2,074 X g, and 76 to 80% were recovered at 3,005 or 3,895 X g at maximum radius. The remainder of mycobacteria was mostly recovered from the supernatant, but culturing of supernatant was not profitable. Increasing RCF had a negligible effect on acid-fast bacillus smear sensitivity. The smear sensitivity for about 25,000 clinical specimens processed with an RCF of 3,800 X g for 20 min was 71% compared with 69% as determined for over 30,000 specimens processed in a similar manner but an RCF of 2,000 X g. An RCF of 3,000 X g applied for 15 min, or an RCF of about 2,000 to 2,500 X g applied for 20 min, is considered adequate to concentrate mycobacteria in clinical specimens.

Bacteriuria↗
PubMed1986

Whirlpool-associated folliculitis caused by Pseudomonas aeruginosa: report of an outbreak and review.

An outbreak of folliculitis caused by Pseudomonas aeruginosa serotype O:7 occurred among the guests of a hotel in St. John's, Newfoundland, Canada, and the source of the infection was traced to the hotel whirlpool. Of 36 persons who used the whirlpool, 26 (72%) developed folliculitis within 1 to 5 days after exposure; the attack rate was significantly higher for children (90%) than for adults (50%). The rash characteristics were consistent with those of Pseudomonas folliculitis previously described (T. L. Gustafson, J. D. Band, R. H. Hutcheson, Jr., and W. Schaffner, Rev. Infect. Dis. 5:1-8, 1983). This is considered to be the first outbreak in which P. aeruginosa serotype O:7 has been incriminated. Published reports to date of outbreaks of Pseudomonas folliculitis associated with the use of whirlpools, hot tubs, swimming pools, etc., were reviewed.

Adolescent↗