To investigate the inflammatory cytokine production of human epithelial cell lines stimulated with uropathogenic E. coli strains showing 3 different adherence patterns, differential expression of inflammatory cytokine (IL-1a, IL-lB, IL-8, TNFa, and TGFB) mRNA were detected by RT-PCR. IL-1a, IL-1B, IL-8, and TGFB mRNAs constitutively expressed in epithelial cell lines, but not TNFa. The expression of IL-1a and IL-1B mRNA was increased in J-82 cells stimulated with E. coli strains showing DA, LA, or AggA pattern. The expression of IL-8 mRNA was increased, whereas TGFj3 mRNA was decreased in J-82 cells stimulated with E. coli strain showing AggA.pattern. Treatment with crude bacterial adhesins (CBA) isolated from E. coli strains showing DA or LA pattern increased IL-la, IL-lB, IL-S, and TGFj3 mRNA expressions in J-82 cells and HeLa cells. IL-la, IL-lB, and TGFB mRNA expressions were decreased in epitheUal cells stimulated with CBA from E. coli strain showing AggA pattern, whereas IL-8 mRNA expression was significantly increased. The expressions of cytokine mRNAs showed little differences between epithelial ceRs used, but great differences between CBA from DA or LA and AggA strain. LPS stimulation was little changed cytokine mRNA expressions in epithelial cells. This study suggests that cytokine gene expression of epithelial cells by the bacterial stimulation mainly depends on the bacterial adhesins recognized by the respective receptors of epithelial cells.
Adhesins, Bacterial ; Epithelial Cells* ; Gene Expression ; HeLa Cells ; Humans ; Interleukin-8 ; RNA, Messenger ; Uropathogenic Escherichia coli*

In order to understand the role of mec regulator genes in the evolution of methicillin-resistant S. aureus (MRSA), the distribution of the mec regulator genes among the 66 clinical isolates of MRSA was analysed. And also the correlation between gene mutation and degree of phenotypic expression of resistance was studied. Fifty strains carried whole mec regulator region, while the mecI gene and nearly half of the 3'-end of the mecR#l gene were deleted in fifteen strains. The mecRl MS gene was detected among all of the mecA carried strains, but the mecRl PB gene was carried by 77% of the MRSA strains. At least a portion of the 5'-end region of the mecRl gene was carried by all MRSA strains tested. Forty-seven strains were finally confirmed to have mecI gene and each mecI gene of above strains was sequenced for identification of the relationship between repressor function of mecI gene on mecA transcription and MIC level of methicillin. Point mutations were detected in 11 strains of 47 strains. In 8 strains, there was one nucleotide substitution (C to T at position 202) that produced a new termination codon at position 201. In 3 strains, one nucleotide substitution from G to T at position 43 caused an amino acid substitution from Val to Phe. The MIC of methicillin of strains carrying mutated mecI genes ranged 256 ug/ ml to 1024 ug/ml. Transcription level of amplified cDNA corresponding to mecA was determined by the method of RT-PCR of extracted RNA. Total RNA was extracted from two strains with mutated mecI gene and a strain with intact mecI gene. Deletional loss or the mutational inactivation of the mecI gene did not affect the level of mecA transcription. Role of mecI gene as a strong repressor function on mecA gene seemed to be skeptical.
Amino Acid Substitution ; Codon, Terminator ; DNA, Complementary ; Genes, Regulator* ; Methicillin Resistance ; Methicillin* ; Methicillin-Resistant Staphylococcus aureus ; Point Mutation ; RNA

No abstract available.

Mobility shifts in non-denatured gel electrophoresis of PCR-amplified Rif' region in each of fifteen different mutants of M. tuberculosis were discerned by single strand conformation polymorphism (SSCP) analysis. The findings of mobility differences between rifampin-resistant and susceptible strains showed an excellent agreement with data obtained by traditional susceptibility test. SSCP-PCR seemed to replace the cultivation method of susceptibility test that was known to be time-consuming, labor wasting, and skeptical in quality control. After screening of rpoB gene mutation by SSCP-PCR, detection of specific sequence changes in the region of rpoB gene was attempted through the procedures of PCR-amplification, cloning of PCR-products using pGEM-T vector and DNA thermocycling sequencing. Fifteen different types of mutations were identified among fifty strains of rifampin-resistant strains while five rifampin-susceptible control strains showed no sequence changes of rpoB gene as well as reference strain H37rv. Most mutation appeared to be a point mutation due to substitution or deletion except seven mutants showing somewhat complex mutation. Each of mut#ated loci inclined to clustering within a region of eighteen amino acids involving eight codons. The most common mutation of Ser425 shared among twenty-nine mutants and followed by eleven mutants of His420. Several mutants alleles identified in this study appeared to be dissimilar to those of previous reports.
Alleles ; Amino Acid Substitution* ; Amino Acids ; Clone Cells ; Cloning, Organism ; Codon ; DNA ; Electrophoresis ; Mass Screening ; Mycobacterium tuberculosis* ; Mycobacterium* ; Point Mutation ; Quality Control ; Rifampin* ; Tuberculosis

No abstract available.
Pseudomonas aeruginosa* ; Pseudomonas* ; Pyocins*

No abstract available.
Molecular Biology* ; Shigella*

In order to evaluate the efficiency of serological typing of A. baumannii in practical application, a total of 63 strains of A. baumannii and 234 strains of Gram-negative, lactose non-fermenting bacteria were tested with polyclonal rabbit immunized sera (RIS) against heat-killed A. baumannii strains by slide agglutination tests. Six typing sera of RIS were finally obtained after the checkerboard agglutination test and reciprocal cross-absorption. Species identification of sixty-three strains of A. baumannii were confirmed by ribotyping. Forty-seven (74.6%) of the 63 strains of A. baumannii showed strong positive reaction by slide agglutination tests. Thirty-nine strains could be serotypable and thus classified into 6 distinct serovars of A. baumannii, but 8 strains were unable to classify into specific serovar. Serovar 4 was the most frequent arbitrary serovar and included 17 strains among the 39. When slide agglutination tests were performed with 50-fold diluted pooled polyclonal RIS, there was no cross-reactions except one of E. coli strain among 234 strains of various Gram-negative lactose non-fermenting. Although each profile of LPS-gel electrophoresis of A. baumannii appeared to be unrelated with serovar, the patterns of western-blot of LPS after immunostaining with homologous RIS showed serovar-specificity. Several fractions of low molecular weight LPS showed cross-reaction with antisera of other serovars. In conclusion, the sensitivity and specificity of serological identification of A. baumannii strains were 74.6% and 99.6%, respectively. This result suggests that serotyping is a useful method for the identification of A. baumannii strains as well as is the epidemiological tool to trace back the source of the nosocomial outbreaks.
Acinetobacter baumannii* ; Acinetobacter* ; Agglutination Tests ; Bacteria ; Disease Outbreaks ; Electrophoresis ; Immune Sera ; Lactose ; Molecular Weight ; Ribotyping ; Sensitivity and Specificity ; Serotyping

No abstract available.
Gene Expression* ; Virulence*

No abstract available.
Gene Expression* ; Microscopy, Electron, Scanning* ; Virulence*

One hundred and thirty trimethoprim-resistant R plasmids derived from of Escherichia coli isolated from clinical specimens and feces of healthy collegians were examined for incompatibility, EcoRI endonuclease restriction fragment pattern, and Southern hybridization with DHFR I, II, III, V, and VII probe. 1. Most trimethoprim-resistant R plasmids were resistant to ampicillin, tetracycline, chloramphenicol, gentamicin, and kanamycin, and showed multiple drug resistance and various antimicrobial resistance patterns. 2. Trimethoprim-resistant R plasmids ranged from 90 to 50 kilobase and 42.3% of R plasmids tested were classified to incompatibilty group Inc FI, Inc FII or Inc FIV, 3. Among 48 random selected R plasmids from various origin, 14 R plasmids (including 9 of 14 Inc FII plasmids and 3 of 14 Inc FI plasmids) hybridized with DHFR VII oligonucleotide probe but others did not respond to any of DHFR probes used. 4. Most R plasmids showed various EcoRI endonuclease fragments and different reaction sites by Southern hybridization. Six plasmids showed identical or nearly identical molecular weight, EcoRI endonuclease fragment patterns and different sites of Southern hybridization. But 2 Inc FII plasmids derived from urine and feces showed identical pattern. These findings, if confirmed by further studies, suggest that normal flora E. coli can act as reservoir of resistant genes and, consequently, as a factor in the dissemination of these genes among enteric pathogens and need to be examined further.
Ampicillin ; Chloramphenicol ; Deoxyribonuclease EcoRI ; Drug Resistance, Multiple ; Escherichia coli* ; Escherichia* ; Feces ; Gentamicins ; Immunodeficiency Virus, Feline ; Kanamycin ; Molecular Biology* ; Molecular Weight ; Plasmids ; R Factors ; Tetracycline ; Trimethoprim Resistance* ; Trimethoprim*