Zhonghua Minguo wei sheng wu ji mian yi xue za zhi = Chinese journal of microbiology and immunology最新文献

The VIDAS rubella IgG(RBG) is a new, automated, enzyme-linkad fluorescent assay(ELFA) for detecting IgG antibodies to rubella virus in serum. The purpose of this study was to evaluate the usefulness of the qualitative and quantitative VIDAS RBG as laboratory tests in the rapid detecting anti-rubella IgG antibodies. Simultaneous parallel testing was performed by using the qualitative and quantitative RBG along with RUBAZYME(ABBOTT LABORATORIES) and the standard hemagglutination-inhibition(HI) test(R-HI kit, SEIKEN) on 200 blood samples submitted for anti-rubella IgG antibodies testing from patients at the VGH-Kaohsiung(Mar.-Sep., 1993). The qualitative and quantitative VIDAS RBG and RUBAZYME assay were compared to the standard R-HI test. Significant differences among the sensitivities of these three methods were found (100% sensitivity for the qualitative VIDAS RBG, 88.89% for the quantitative VIDAS RBG and 97.67% for the RUBAZYME). However, the specificities of these three methods were all the same, 100%. With these results we can conclude that the qualitative VIDAS RBG will provide a very good, precise and reliable method to determine serum specific IgG antibodies against rubella virus. Furthermore, the VIDAS RBG is fully automated and time-saving procedures (for every batch of test, it takes 25 minutes with VIDAS RBG, 2.5 hours with RUBAZYME and 4.5 hours with R-HI) in clinical laboratories.

Restriction fragment length polymorphism analysis of rRNA genes (ribotyping) was used to differentiate Vibrio vulnificus isolates. Among the 10 restriction enzymes tested, HindIII was shown to provide the most discriminatory patterns. Stul was used for further analysis of strains that were indistinguishable with HindIII. Thirteen clinical V. vulnificus strains were analyzed for their ribotypes with HindIII, as well as Stul when necessary. Four of the clinical strains were isolated from different samples collected from the same patient, and were shown to have identical ribotypes. All the others gave unique ribotypes, indicating the large genetic divergence in V. vulnificus clinical isolates. The ribotype of V. vulnificus by HindIII remained unchanged after successive in vitro and in vivo passages. HindIII gave rise to five bands which were shown in every V. vulnificus strain but not in other vibrio species tested, suggesting that ribotyping with this restriction enzyme may be useful for confirming the identification of this bacterium.

To type the Vibrio cholerae strains isolated from sporadic and epidemic cases in Taiwan, 28 toxigenic isolates were studied by sequencing polymerase chain reaction-amplified cholera toxin gene (ctx) fragments. Based on specific base substitutions on positions 115 and 203 of ctxB and comparison with previously published typing system from Centers for Disease Control and Prevention (Olsvik theta et al., J Clin Microbiol 1993; 31:22-5, Ref.1), two genotypes were identified. Cholera strains from imported seafood and sporadic cases in Taiwan had ctxB polymorphism of genotype 1; strains from patients in the 1962 Taiwan epidemic and Taiwan raised soft-shell turtles had ctxB polymorphism of genotype 3. Moreover, one toxigenic non-O1 strain was found to have other 11 different nucleotides in ctxB compared with those of the O1 and O139 strains. Therefore, DNA sequencing is a useful method for obtaining more complete genetic information. The approach could be improved by applying it to other more polymorphic regions of bacterial genome to obtain better epidemiological information among infected cases.

The virulence and some related factors of Vibrio parahaemolyticus are regulated by the level of iron. In this study, five Mn-resistant mutants were selected after N-methyl-N'-nitrosoguanidine treatment and two transfers in medium containing high levels of manganese chloride. Production of siderophores and the 77-kDa iron-regulated outer-membrane protein and the bacterial growth in these Mn-resistants were deregulated, as compared with the wild-type strain. In addition, the regulation of these phenomena was partially or completely restored by the introduction of Escherichia coli fur gene. Also, the total cellular protein profiles of the wild-type and mutants showed that production of some proteins were positively or negatively regulated by iron, and expression of some of these proteins remained unaffected in these mutants. These results suggested the presence of a complicated iron regulation system, similar to the Fur system of E. coli, in this pathogen.

It has become clear that the genotypes of HCV vary with respect to pathogenicity, infectivity, response to antiviral therapy and geographic clustering. The prevalence of genotype distribution of HCV infection in Taiwan was investigated by typing with type-specific DNA primers in HCV core region. Using a design by Okamoto et al., it was found that in 280 serum samples examined, 3.3% (4/122) of the virus detected were mixed type. The implication of mixed type infection remains to be clarified: whether it is a single infection with a new variant, or infection with two HCV virions at different times or confusion with type-specific DNA primers themselves. The nucleotide sequences of the recombinant plasmid DNAs and the PCR products recovered from gel electrophoresis were analyzed by autosequencer. Gene sequences of HCV cDNAs of the two blood donors were used as control. To double check the results, we have also analyzed the DNA sequences of the cloned plasmids in the NS5 region with the primer system designed by Chayama et al. Results indicated that the hemodialysis patient was doubly infected with HCV, rather than by a HCV variant.

As a part of investigations to characterize trypanosome infections in Taiwan, sera collected from patients admitted to Veterans General Hospital, Taipei were tested for antitrypanosome antibodies. A Trypanosoma cruzi extract-based enzyme-linked immunosorbent assay (ELISA) was used to screen and titrate 1,297 patient sera. High antitrypanosome titers were detected in 166 (12.8%) of these sera. Retitration of random samples of the high titer (HT) sera indicated a 5.4% false positive. Thirteen donors with high antitrypanosome ELISA titers were followed up. Twelve of then remained high serum titers also showed high ELISA titers against an extract of Trypanosoma conorhini. Hemocultures conducted on freshly drawn blood specimens of the 13 subjects did not provide any evidence of trypanosome infections. Electrophoretic analyses of sera from HT and low titer (LT) patients suggested differences between serum proteins of the subjects in each of the groups. Atypical reactions were observed in immunodiffusion tests performed with HT and LT sera and trypanosome extracts, while western blot analyses revealed a complex pattern of binding by both sera. The qualitative and quantitative differences in these tests suggested interactions of T. cruzi antigens with donor antibodies against unrelated antigens and/or with autoantibodies. Subsequent analyses did not indicate any association between rheumatoid factor and the reactivities of the HT sera with the parasites. However, antinuclear antibodies were detected with an indirect fluorescent antibody test (IFAT) in 50% of the HT sera and 22% of the LT sera. No differences were found between the levels of antilaminin activity of the two groups. The IFAT employing T. cruzi epimastigotes was positive for 100% of the HT sera and 22% of the LT sera. The data indicate that the high seropositivity recognized in this study is due in part to the activities of cross-reacting antibodies and/or autoantibodies in the sample population.

Streptococcus mutans constitutively expresses three glucosyltransferases, i.e., GtfB, GtfC, and GtfD; which synthesize glucan polymers from sucrose. To obtain individual GTF without complexing with one another, a purification strategy was developed to recover recombinant GTF expressed from Escherichia coli. The recombinant GtfC was aggregated and associated with the insoluble fraction in E. coli homogenates. GtfC was solublized with the 8M urea, renatured to its biologically active form by serial dialysis against sodium phosphate buffer, and subsequently purified to homogeneity by DEAE-Sephacel and hydroxylapatite column chromatography. The GtfC enzyme preparation was purified 16.3-fold and the molecular weight was estimated to be 140 kDa. GtfC synthesized water insoluble glucan in a primer independent manner and its enzymatic activities could be enhanced by dextran. Purified GtfC had a pH optimum of 6.5, a K(m) of 9.26 mM for sucrose and a pI of 5.5. Distinct from the previous reports, results from this study offers an alternative for the purification of the recombinant GTFs free from any detergent contamination to make it more suitable for utilization in vivo.

The distribution pattern of HLA antigens varies to a great extent among different ethnic groups. Availability of HLA antigen distribution information is very important for disease association study, paternity testing and recipient/donor matching. Analysis of 11,383 blood donors from the Taipei Blood Center, Chinese Blood Services Foundation (CBSF), gave evidence that the distribution pattern of HLA-A,B antigens was unique, yet closer to Southern Chinese with higher A11, A33, B16, B40 frequencies, and B46, B48, B54 as unique antigens when compared with Caucasians. Pairwise linkage disequilibrium analysis between HLA-A, -B and -DR antigens from 238 blood donors revealed unique linkages of A33-B17, A2-B46, A11-B15, A24-B40; A11-DR5, A2-DR9, B17-DR3, B40-DR8, B13-DR2. In addition, A33-B58-DR3 were the most frequent 3-loci haplotypes. Knowing linkage disequilibrium between HLA loci and preferential association of DR specificity among various HLA-A,B haplotypes may provide a more efficient strategy to obtain an HLA-DR or HLA-D region compatible unrelated bone marrow donor from an existing HLA data bank.

The susceptibility of 46 pneumococcal isolates collected during October 1989 to May 1995 from National Taiwan University Hospital and Taipei Municipal Yang Ming Hospital was studied. Among these isolates, the resistant rate of penicillin G was 21.7%; the penicillin G-resistant strains were more frequently resistant than the penicillin-sensitive strains to other beta-lactam antimicrobial drugs. The minimum bactericidal concentrations (MBCs) of penicillin G for all isolates were equal to, or one dilution higher than, minimum inhibitory concentrations (MICs). Three strains were false positive for penicillin resistance among isolates of Streptococcus pneumoniae screened with oxacillin. On the other hand, resistance to penicillin G was often independent of resistance to erythromycin. Vancomycin was the most active agent tested.

A cell line, J5, derived directly from the human hepatocellular carcinoma (HCC) by in vitro culture, and three other lines, J2-23, J3-27 and J3-28, which were established in culture only after passages in nude mice were examined. Since nude mice as well as various strains of normal mice often produce infectious murine retroviruses, the HCC cells passaged as a heterotransplant in nude mice may be infected with these viruses. The supernatant of the J2-23, J3-27 and J3-28 cell co-cultured with a wild mouse cell line, SC-1, which supports the murine ecotropic retroviruses, showed the presence of a murine N-tropic virus. No murine xenotropic retrovirus was detected in these cells using S+L- mink cell assay method. A clone (J3-28 Clone 1) was derived from the above mentioned J3-28 cells, since the latter contained a mixture of human and murine cells. The J3-28 Clone 1 was found to be entirely of human karyotype. This clone as well as the J5 and the PLC/PRF/5 cel lines which have never been passaged in nude mice, showed none of these murine retroviruses. By Southern-blot hybridization, no change was detectable for c-abl, c-ras, c-mos, and c-myc protooncogenes in the chromosomal DNA of J2-23, J3-27, J3-28 Clone 1, and J5 cell lines. The karyotypes of all the HCC cell lines were aneuploid, and those of the J2-23 and J3-27 cell lines were of mouse, while the J3-28 Clone 1 and J5 cell lines were of human. The chromosomes of the J2-23 and J3-27 cells showed, except for the presence of a few marker chromosomes in the former, no apparent changes in the length were observed. In contrast, those of the J3-28 Clone 1 and J5 cells exhibited various changes, including 6-8 marker chromosomes, and an increase or decrease in the length of p and/or q arms of various chromosomes.