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Biomedical subjects

T Victor

Publications and source records attributed to T Victor.

At least 19 recordsLinked to original sources
PubMed1996

Polymerase chain reaction in the diagnosis of urinary tract tuberculosis.

The polymerase chain reaction (PCR) is a technique that can be used to amplify a specific DNA genomic sequence, whereby the presence of an extremely small number of bacteria can be detected. The high sensitivity of PCR is particularly useful in paucibacillary situations such as non-pulmonary tuberculosis (TB). The aims of the present study were to establish a PCR assay for the rapid detection of Mycobacterium tuberculosis (MTb) in urine, to compare the sensitivity of PCR with routine culture technique (Bactec) and to determine the optimal type of urine specimen for PCR detection of MTb. In the first phase of the study, a total of 92 urine specimens were collected from 83 patients with suspected urinary tract TB. Two urine specimens in 2 patients were positive for TB by both PCR and Bactec, while 90 specimens from 81 patients were negative by both methods. Inhibition of PCR was present in nine urine specimens (10%). In the second phase of the study, a further seven patients were selected for intensive investigation to determine the optimal urine sampling for PCR detection of MTb. The conclusions of the study are that PCR can provide much faster confirmation of urinary TB (within 24-48 h) than Bactec urine culture (which may take several weeks). About 10% of urine specimens could not be evaluated by PCR due to the presence of inhibitory substances of unknown nature. MTb organisms were found to be excreted intermittently in the urine of infected patients, and single specimens were more likely to be false negative than a 24-h sample. The best method appeared to be the concentration of a large volume of urine, for instance 11 concentrated to 2 ml.

Adult↗
PubMed1995

Erythema induratum of Bazin. A clinicopathological study of 20 cases and detection of Mycobacterium tuberculosis DNA in skin lesions by polymerase chain reaction.

Erythema induratum of Bazin (EIB) is a chronic, recurring panniculitis that is found predominantly on the legs of women with tuberculin hypersensitivity. A causal relationship between EIB and Mycobacterium tuberculosis remains elusive because of the absence of demonstrable organisms in skin lesions. We reviewed the clinicopathological features of 20 patients (all women) with positive Mantoux tests (1:10,000 dilution) and characteristic skin lesions of EIB that cleared up with combined antituberculous treatment. Histological examination of skin lesions confirmed panniculitis with varying combinations of granulomatous inflammation, primary vasculitis, and necrosis. Sections from 20 formalin-fixed, paraffin-embedded skin biopsies obtained from the 20 patients were submitted for polymerase chain reaction (PCR) using oligonucleotide primers for the detection of a 123-bp DNA fragment specific for the M. tuberculosis complex. M. tuberculosis DNA was identified in five of the 20 biopsies. Meticulous care was taken to prevent contamination as a source of false-positive results. Mycobacterial DNA was absent in all negative controls and in normal skin biopsies from purified protein derivative-positive patients with and without EIB. These results provide direct evidence that mycobacterial components are present in EIB lesions and strongly suggest that M. tuberculosis is involved in the pathogenesis of EIB.

Adolescent↗
PubMed1994

The detection of Mycobacterium tuberculosis in bone marrow aspirate using the polymerase chain reaction.

SETTING: Tygerberg Hospital, South Africa. OBJECTIVE: Bone marrow aspirate and biopsy were obtained from 37 patients who were in-patients at the Tygerberg hospital. The specificity and sensitivity of the polymerase chain reaction (PCR) in the detection of Mycobacterium tuberculosis in bone marrow aspirate was evaluated. DESIGN: The PCR was compared to standard culture as well as to clinical and bone marrow biopsy data in 24 patients with suspected tuberculosis (TB). RESULTS: 12 of the 24 patients eventually had definite or probable TB and in these 12 patients the detection incidence was 42% for PCR and 25% for culture. CONCLUSION: This study confirms that it is possible to use PCR to detect M. tuberculosis in bone marrow aspirate material and that this technique is more sensitive than culture methods. The PCR technique has the added advantage of being a rapid test yielding results within 2 days of sampling. Overall sensitivity for the detection of M. tuberculosis in bone marrow aspirate may be improved to 58% [corrected] by using both culture and PCR techniques.

Biopsy↗
PubMed1994

Incidence of heat-labile enterotoxin-producing Escherichia coli detected by means of polymerase chain reaction amplification.

Diarrhoea can be caused by many different organisms, some of which are notoriously difficult to identify. One of these is enterotoxin-producing Escherichia coli. Recently a new diagnostic technique that uses polymerase chain reaction DNA amplification was developed for detection of the 'A' subunit of the labile enterotoxin-producing E. coli gene. This technique was used to evaluate the incidence of heat-labile (LT+) enterotoxin-producing E. coli in the causation of diarrhoea. The results from this study showed that LT+ E. coli is a cause of diarrhoea in the western Cape and that 5.3% of non-diagnosed diarrhoea patients in Tygerberg Hospital were infected with this pathogen. This represented less than 1% of the total number of cases of diarrhoea investigated in this hospital. The peak coincides with the wetter months in this locality and the infection rate is lower than that reported in most other countries. Given the low incidence of occurrence of this organism we do not recommend routine implementation of the diagnostic procedure. However, this test may be useful at times, e.g. to ascertain the source of a diarrhoea epidemic.

Adult↗
PubMed1993

Laboratory experience and guidelines for avoiding false positive polymerase chain reaction results.

Despite the widespread use of polymerase chain reaction (PCR) for diagnosis of infectious diseases, the technology has not been generally introduced into routine diagnostic laboratories. One of the most serious problems which has influenced the acceptance of this technology is the occurrence of false positive PCR results. This study describes the experience, in a hospital laboratory setting, of using PCR for the diagnosis of heat-labile enterotoxin-producing E. coli, M. tuberculosis, M. paratuberculosis and human papillomavirus. Results indicate that a build-up of amplicons, generated during the amplification process in the laboratory, is the main source of PCR-contamination. Protocols are described that include both physical and chemical procedures to prevent contamination. The use of photo-induced psoralen is recommended for those laboratories already involved in PCR work where amplicons are likely to be present. An enzymatic system (uracil-N-glycosylase) was evaluated and is recommended for workers intending to start diagnostic PCR. Attention was given to simple control measures which are easily implemented in a routine diagnostic laboratory. Protocols such as these are likely to have a major impact on the introduction of PCR-based methods into routine laboratories.

Bacteria↗
PubMed1992

Purification of sputum samples through sucrose improves detection of Mycobacterium tuberculosis by polymerase chain reaction.

A method is described for the routine preparation of sputum samples for detection of Mycobacterium tuberculosis by using polymerase chain reaction amplification. Liquefaction of sputum samples with NaOH and subsequent removal of inhibitors of the polymerase reaction with a 50% sucrose centrifugation step (5 min) in a desktop centrifuge allow direct amplification of a 123-bp repetitive region of the M. tuberculosis genome. We have evaluated the sensitivity and specificity of the sucrose method with 155 sputum specimens from patients suspected of having tuberculosis and from normal healthy volunteers. This method, in our opinion, is reproducible, sensitive, and reliable. Multiple samples can be handled simultaneously, and results can be obtained in one working day.

Centrifugation, Density Gradient↗
PubMed1992

Papulonecrotic tuberculid. Identification of Mycobacterium tuberculosis DNA by polymerase chain reaction.

Sections from 22 formalin-fixed, paraffin-embedded skin biopsies from 12 patients with papulonecrotic tuberculid (PNT) were examined for the presence of Mycobacterium tuberculosis DNA with use of the polymerase chain reaction. All patients had a positive tuberculin skin test and a compatible clinical picture and responded to antituberculous therapy. Histological examination showed the typical morphology of PNT lesions with dermal necrosis surrounded by an ill-formed granulomatous infiltrate. Mycobacterial DNA was detected in 11 of the 22 biopsies. None of the negative controls showed positive DNA identification by amplification. Great care was taken in avoiding false-positive results due to contamination. After reviewing the literature, we believe this is the first time that PNT lesions have been investigated by PCR for the presence of mycobacterial DNA. These findings provide direct proof that mycobacterial products are present in PNT lesions and support the theory that this organism is in some way responsible for the development of PNT.

Adolescent↗
PubMed1991

The use of the polymerase chain reaction test in the diagnosis of tuberculosis.

Current techniques for laboratory diagnosis of tuberculosis have some serious limitations. These include the high cost and time required for the current assays. The development of a rapid, sensitive, specific and low-cost assay is therefore of considerable importance. We report here the development and laboratory testing of a polymerase chain reaction DNA-based diagnostic test for the presence of Mycobacterium tuberculosis in sputum. The assay shows a high level of sensitivity and specificity and requires considerably less capital, consumables and time inputs than existing laboratory tests. We believe this technology is ready for large-scale evaluation and use, particularly in hospital-based laboratories.

Humans↗
PubMed1991

Improved method for the routine identification of toxigenic Escherichia coli by DNA amplification of a conserved region of the heat-labile toxin A subunit.

This report describes a DNA amplification procedure for routine identification of heat-labile-toxin-producing Escherichia coli. Two oligonucleotide primers were used in a polymerase chain reaction procedure to amplify a highly conserved region of the A subunit of the heat-labile enterotoxin gene. Amplifications were done directly on E. coli colonies from plates when Salmonella, Shigella, or parasite infections were excluded as agents of the severe diarrhea in the patients. The conditions for the polymerase chain reaction method were empirically determined, and the procedure is inexpensive, sensitive, and specific. Positive results can be obtained over a wide variation in bacterial numbers, with no inhibition of Thermus aquaticus DNA polymerase. Detection of the amplified product can be done by agarose gel electrophoresis, which is specific and sensitive enough for routine diagnosis of this pathogen in clinical isolates. If greater sensitivity and specificity are required, hybridization with 32P- or alkaline phosphatase-labeled oligonucleotide probes can be used. Our results suggest that heat-labile-toxin-producing E. coli is responsible for about 9% of nondiagnosed diarrhea cases in Tygerberg Hospital, Tygerberg, Republic of South Africa.

Alkaline Phosphatase↗
PubMed1990

No evidence for point mutations in codons 12, 13, and 61 of the ras gene in a high-incidence area for esophageal and gastric cancers.

The molecular mechanisms underlying the induction of esophageal and gastric cancer are not yet understood. It is possible that different etiological factors from geographically distinct areas play a role in the onset of these cancers. Twenty-seven primary esophageal and 11 gastric cancers originating from the high-incidence areas of South Africa were analyzed for the presence of ras protooncogene mutations. We found no evidence for mutations in codons 12, 13, or 61 or the H-ras, K-ras, and N-ras genes in these primary cancers. Our results indicate that etiological factors such as fungal contamination of basic foodstuffs in a high-incidence area for these cancers do not play a role in the activation of ras genes and that mutations in these genes are not directly involved in the development of primary esophageal and gastric cancers in the South African population.

Base Sequence↗
PubMed1989

Estradiol induction of accelerated energy metabolism in prepuberal rat uteri in vitro: mRNA hybridization and [13C]NMR studies.

In vitro treatment with 30 nM 17 beta-estradiol stimulated the induction of mRNA for the brain type isozyme of creatine kinase BB (CKBB) and stimulated glucose metabolism in perifused uteri from 27-29-day-old rats. The perifusion conditions maintained the normal NMR spectrum of high energy phosphates for at least 24 h. This technique permitted the demonstration that perifused rat uteri stimulated by 17 beta-estradiol show increased mRNA for creatine kinase BB, 1 h after estrogen addition. The time-course of increase, measured by Northern blot hybridization, parallels that seen in mRNA extracted from uteri after in vivo induction by i.p. injection of 5 micrograms 17 beta-estradiol; the maximal increase is seen at 2-4 h. Experiments utilizing actinomycin D (4 micrograms/ml) for inhibition of RNA synthesis showed that CKB mRNA from both untreated and estradiol stimulated uteri had a similar half-life, of approximately 2 h, indicating that CKB mRNA is transcriptionally regulated. In the same system, the rate of glycolysis was measured by NMR spectroscopy using [1-13C]glucose. Following in vitro stimulation with 30 nM estradiol, glycolysis increased within 3 h, in parallel to increases previously found in uteri from in vivo stimulated rats.

Animals↗
PubMed1985

Mitochondrial phospholipid composition and microviscosity in myocardial ischaemia.

Normothermic ischaemic arrest of the isolated perfused rat heart causes profound changes in mitochondrial ultrastructure. Since the mitochondrial membranes contain a high percentage of phospholipids, an evaluation of the effect of different periods of ischaemia on mitochondrial phospholipid content and fatty acid composition was made. The results showed that ischaemia had no effect on the content of the different phospholipid classes and no correlation was observed between ultrastructural changes and mitochondrial phospholipid content. However, the phospholipid fatty acid composition of several phospholipids showed marked changes. For example, with lysophosphatidylcholine a progressive increase in the percentage saturated fatty acids was observed with increasing periods of ischaemia, while a reduction occurred in lysophosphatidylethanolamine. To determine whether the ischaemia-induced changes in mitochondrial phospholipid fatty acid composition had an effect on the physical properties of the membrane, the microviscosity of mitochondrial preparations was studied, using the lipophilic probe, 1,6-diphenyl-1,3,5-hexatrine. Mitochondria isolated from ischaemic hearts showed a progressive increase in fluorescence polarization with longer periods of ischaemia, indicating an overall increase in microviscosity. This phenomenon may be responsible for the increased mitochondrial fragility which is characteristic of ischaemic damage.

Animals↗
PubMed1985

Physiological and biochemical measurements during a 4-day surf-ski marathon.

We studied competitors in the 1983 Texan Challenge surf-ski paddle marathon to determine the effects of 4 days prolonged paddling on sweat rates, rectal temperatures, renal function, serum glucose, free fatty acid, porphyrin and C-reactive protein levels and serum creatine kinase activity. Sweat rates during the race varied from 0.5 to 1.0 l/h, and peak rectal temperatures did not exceed 38 degrees C, even in the most dehydrated subjects. Renal function was unchanged during the race. Plasma renin activity remained low during the race but C-reactive protein levels and serum creatine kinase activity were elevated. Seventy per cent of the subjects had immediate post-exercise blood glucose levels below 3.9 mmol/l after the 1st and 4th days, and 27% had values below 3.0 mmol/l, one competitor requiring intravenous glucose therapy on the beach. Surf-ski paddling is therefore associated with low sweat rates, low levels of dehydration, low body temperature and unchanged renal function. The low post-exercise blood glucose levels indicate that competitors must eat high-carbohydrate diets for the duration of the event and must either eat carbohydrate-containing foods or drink concentrated carbohydrate solutions while paddling.

Blood Glucose↗
PubMed1984

Myocardial tissue free fatty acids.

Myocardial ischaemia, whether produced by coronary artery ligation or by hypoxic low-flow perfusion of the isolated rat heart, has been shown to be associated with a significant depression in mitochondrial function, as well as an increase in tissue free fatty acid (FFA) levels. Although the effects of FFA on mitochondrial oxidative phosphorylation function in vitro are well established, it has not yet been shown that increased tissue FFA levels are causing the depression in mitochondrial function in ischaemia. Using the isolated perfused rat heart, several experiments were performed to gain more information regarding (i) the validity of the Dole FFA extraction technique; (ii) controlling factors; (iii) the relationship between tissue FFA and mitochondrial function and (iv) sources of tissue FFA in ischaemia. Significant elevation of tissue FFA was achieved by perfusion with (i) substrate-free Ringer and (ii) phosphatidylcholine. Elevation of tissue FFA obtained by perfusing with long-chain fatty acids was due to extracellular accumulation. Reduction of tissue FFA levels was observed by perfusing with (i) albumin, (ii) glucose, insulin and propranolol. Our results also suggest that lysosomal involvement could cause the increase in tissue FFA levels in myocardial ischaemia.

Animals↗