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Biomedical subjects

Y J Tang

Publications and source records attributed to Y J Tang.

13 recordsLinked to original sources

Genomic fingerprinting of epidemic and endemic strains of Stenotrophomonas maltophilia (formerly Xanthomonas maltophilia) by arbitrarily primed PCR.

Arbitrarily primed PCR (AP-PCR) was used to type 64 clinical isolates of Stenotrophomonas maltophilia from 60 patients and the hands of one nurse. Forty-seven different patterns were observed, most patients having isolates with unique genomic fingerprints. A single pattern, however, was obtained from six of eight patients involved in an intensive care nursery outbreak, confirming the suspected nosocomial transmission of this microorganism. This strain was also found in four other patients hospitalized at the same time but in different units. AP-PCR typing results had a good correlation with the 49 patterns obtained when the isolates were typed by contour-clamped homogeneous electric field gel electrophoresis. Although AP-PCR is slightly less discriminatory than contour-clamped homogeneous electric field gel electrophoresis, it offers several advantages and should be considered as a practical option for molecular typing during investigations of outbreaks.

Bacterial Typing Techniques

Apparent outbreaks of Clostridium difficile-associated diarrhea in horses in a veterinary medical teaching hospital.

Intestinal colonization with toxigenic strains of Clostridium difficile was documented in 9 of 10 horses with acute onset diarrhea in a veterinary medical teaching hospital, whereas a similar isolate was detected in only 1 of 23 other horses without diarrhea in the hospital. One horse with diarrhea was infected simultaneously with both C. difficile and Salmonella krefeld. Clostridium difficile was detected by fecal culture on selective medium, confirmed with a latex particle agglutination test, and identified as toxigenic by polymerase chain reaction amplification of toxin A and toxin B gene sequences. Using an arbitrarily-primed polymerase chain reaction, 6 distinct C. difficile isolates were detected in the feces of the 9 affected horses at the time of the outbreak of diarrhea.

Animals

Specific detection of Clostridium difficile toxin A gene sequences in clinical isolates.

The polymerase chain reaction (PCR) was used to specifically detect toxin A gene sequences of Clostridium difficile in DNA isolated from human faeces. A set of oligonucleotide primers derived from the non-repetitive region of the toxin A gene was developed to amplify a 634-bp DNA fragment. All 28 cytotoxic strains of C. difficile, previously characterized by a toxin B-PCR assay, were positive for the presence of toxin A gene sequences. No amplification products were obtained from DNAs extracted from non-toxigenic strains, strains of C. sordellii, or C. bifermentans. In addition, amplification of DNA extracted from C. difficile 8864, a strain which does not produce toxin A, resulted in multiple bands which probed negative for toxin A gene sequences. DNAs extracted from nine stool specimens which were positive for toxin B by the cytotoxicity assay and by the toxin B-PCR assay were also positive in this assay. Toxin A gene sequences were detected in DNAs obtained from 4/11 stool specimens which were negative by the toxin B cytotoxicity assay. These four specimens were from patients who had a history of relapses due to C. difficile-associated colitis, and whose stools had previously been found to be positive by the toxin B-PCR test despite no detectable toxin B in the specimens. These data indicate a comparable degree of clinical sensitivity between these two toxin-gene PCR-based assays. This rapid, sensitive and specific assay may be useful not only in the diagnosis of C. difficile infections, but also in molecular studies of the toxin A gene in C. difficile strains.

Bacterial Proteins

Genotyping of Clostridium difficile isolates.

Arbitrarily primed polymerase chain reaction (AP-PCR) was used to genotype Clostridium difficile isolates from various sources. Four major molecular types were identified among strains from the American Type Culture Collection previously typed by serogroup and from isolates from patients at the University of California, Davis Medical Center, from a patient at a Utah institution, and from the environment. These groups contained subgroups that displayed, in addition to the common group bands, at least one unique band. Two strains isolated from patients at our institution had the same DNA banding patterns. These patients were hospitalized during the same period, raising the possibility of cross-infection through hospital contact or another common source. These results suggest that this AP-PCR approach will be useful in epidemiologic studies of C. difficile infections.

Base Sequence

Fecal shedding of Clostridium difficile in dogs: a period prevalence survey in a veterinary medical teaching hospital.

The goal of this study was to determine the fecal prevalence of Clostridium difficile in dogs who were patients at a veterinary medical teaching hospital. Stool specimens collected from 152 dogs (in- and outpatients) were analyzed for the presence of C. difficile. An additional 42 stool specimens were collected and examined from dogs recently housed at local animal shelters. Following culture on selective medium, C. difficile was identified by a latex agglutination test, and the presence of the toxin A and B genes was determined individually by polymerase chain reaction. Clostridium difficile was isolated from the feces of 28 of the veterinary hospital patients (18.4%); isolates from 14 of these patients (50.0%) were toxigenic. Diarrhea was a clinical finding in 5 (35.7%) of the dogs carrying toxigenic isolates of C. difficile, whereas diarrhea was noted in only 2 of 14 dogs (14.3%) shedding nontoxigenic isolates. Three of 14 dogs (21.4%) shedding toxigenic isolates of C. difficile were receiving antibiotics at the time of stool collection, whereas 5 of 14 dogs (37.5%) shedding nontoxigenic strains of C. difficile were receiving antibiotics. The carriage rate of C. difficile was significantly higher for animals categorized as inpatients of the veterinary hospital. The carriage rate also provided evidence for an increased risk for fecal shedding with increasing age. Clostridium difficile was not isolated from any of the 42 dogs recently housed at local animal shelters. This study confirms the presence of toxigenic C. difficile in dogs at a veterinary teaching hospital.(ABSTRACT TRUNCATED AT 250 WORDS)

Age Factors

Diagnosis and monitoring of Clostridium difficile infections with the polymerase chain reaction.

Toxigenic Clostridium difficile is the etiologic agent of pseudomembranous colitis. We have developed an assay system for the rapid direct detection of toxigenic C. difficile in human stool samples. After DNA extraction, polymerase chain reaction (PCR) amplification is undertaken with primers targeting specific sequences in the C. difficile 16S rRNA gene. Next, toxigenic strains of C. difficile are distinguished from nontoxigenic strains by PCR amplification of toxin A and/or B gene sequences. This study included 12 patients with C. difficile colitis, seven of whom had clinical relapses after discontinuation of vancomycin therapy. We detected toxigenic C. difficile in stools from four (57%) of these seven patients before relapse--at a time when no toxin B was detectable in stools and results of anaerobic culture were negative. The PCR assay is 100-fold more sensitive than anaerobic culture methods. The course of the infection in one patient (both during and after therapy) was monitored by the PCR technique. The multigene analysis approach permitted the detection of colonization with a nontoxigenic strain when this patient's relapses had cleared. This clinically applicable assay allows earlier detection of infection with toxigenic C. difficile. The result is more timely therapeutic intervention.

Aged

Study on sperm function in men long after cessation of gossypol treatment.

Sperm function was assessed in 19 men 3-10 years after cessation of gossypol treatment and 2-9 years after recovery of normal sperm density. Nineteen normal fertile men of similar age served as the controls. The zona-free hamster egg-sperm penetration assay (SPA) revealed a highly significant difference (P less than 0.01) in the penetration rates between the treated and the control groups. Hormone assays indicated that there were no significant differences in circulating levels of luteinizing hormone (LH) and testosterone (T) between the two groups, but the follicle stimulating hormone (FSH) level of the treated group was significantly higher than that of the controls (P less than 0.05). The results showed that sperm function in the treated group was lower than that in the control, which may be a result of persistent gossypol-mediated damage to the testes.

Adult

[Experimental study on anti-acute leukemia with Chinese traditional drugs].

This paper deals with the experimental study on L7212 leukemic model of mice with Chinese medicine Liushenwan, Zhijinding and Xihuangwan for the treatment. This study proved that these drugs possessed the effect of inhibiting and killing L7212 leukemic cells of the experimental leukemic mice (P less than 0.05-0.001). They affected the S stage of the cell multiplication cycle time mainly (P less than 0.01). They could relieve the infiltration of leukemic cells in the liver and spleen of L7212 mice (P less than 0.001) and obviously prolong the survival time of the mice (P less than 0.01-0.001).

Animals

A NosA-specific bacteriophage can be used to select denitrification-defective mutants of Pseudomonas stutzeri.

phi PS5, a double-stranded DNA bacteriophage of Pseudomonas stutzeri JM604 that adsorbs specifically to the outer-membrane protein NosA, was isolated from stagnant irrigation ditch water. Mutant strains that do not produce NosA are resistant to phi PS5 and cannot grow anaerobically with N2O as the sole electron acceptor. phi PS5 did not adsorb to nosA mutants and adsorption to the wild-type strain was reduced when cells were grown with a high concentration of copper, a condition that represses the synthesis of NosA. The isolation of spontaneous phi PS5-resistant mutants yielded strains that were clearly defective in growth on N2O at about a 10% incidence. About half of these strains could respire N2O when supplied with a high concentration of exogenous copper.

Adsorption

A Pseudomonas stutzeri outer membrane protein inserts copper into N2O reductase.

Among a set of frameshift mutagen (ICR-191; Polysciences, Inc.)-induced mutations that confer inability to grow anaerobically with N2O as the sole electron acceptor, one class was found that produced an inactive N2O reductase which lacked copper. All of these mutant strains failed to produce a 61,000-Mr protein located in the outer membrane. This protein, termed NosA, seems not to be responsible for bringing copper into the cell because the mutant strains and their parent were similarly sensitive to the copper content of the growth medium and no intermediate copper concentration in the medium permitted the mutant strains (nosA) to grow anaerobically with N2O as the sole electron acceptor. We conclude that NosA is necessary to insert copper into N2O reductase or to maintain it there.

Bacterial Outer Membrane Proteins

Immediate hypersensitivity to Cryptococcus neoformans.

The role of the capsular polysaccharide in anaphylactic reactions to Cryptococcus neoformans was investigated. Groups of mice were sensitized with viable cells of either a moderately encapsulated strain of C. neoformans or a non-encapsulated variant. Anaphylactic reactions were observed in both groups of mice to a similar extent when challenged with whole cells. Mice sensitized with the encapsulated strain and challenged with homologous polysaccharide showed only mild hypersensitivity symptoms. Mice sensitized with either the encapsulated or the nonencapsulated strain showed cross-reactivity when challenged with killed cells of the heterologous strain. These data indicate that the capsular polysaccharide plays a minor role in anaphylactic reactions to C. neoformans and that the sensitizing antigen is probably located in the cell wall of the yeast.

Anaphylaxis

Use of the polymerase chain reaction for the specific and direct detection of Clostridium difficile in human feces.

The polymerase chain reaction was used for the detection of Clostridium difficile, the etiologic agent of antibiotic-associated colitis. An upstream primer identical to a coding region (segment I) of the C. difficile 16S rRNA gene and a downstream primer complementary to a highly conserved region of eubacterial 16S rRNA served to amplify a targeted 270-base-pair fragment of genomic DNA. This technique allowed the detection of as few as 10 C. difficile organisms among 10(6) Escherichia coli bacteria. This level of sensitivity represents a 100-fold increase over that of conventional anaerobic culture. C. difficile was detected in DNA extracted directly from the stools of 23 patients with antibiotic-associated colitis and from those of four patients with diarrhea whose stools had been negative for C. difficile when assessed in a cytotoxicity assay. No amplification products were found in the stools of asymptomatic patients. When detected in stools of symptomatic patients, amplification products of C. difficile were confirmed by Southern blotting with a nonradioactive, horseradish peroxidase-catalyzed, chemiluminescent probing system in which biotin-labeled oligonucleotides were used. This system discriminates between C. difficile and similar organisms, such as Clostridium sordellii and Clostridium bifermentans. The combination of the polymerase chain reaction with enzyme-linked probing results in a faster and more sensitive assay for C. difficile than standard culture.

Base Sequence