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Chlamydia trachomatis elementary body (EB) and reticulate body (RB) developmental stages have polymorphic plasmid DNA. Several plasmid forms separated by gel electrophoresis were identified as topoisomers by treatment with topoisomerase I. Among these topoisomers was one form unique to EBs and one form unique to RBs. The unique EB plasmid topoisomer was characterized as highly supercoiled, on the basis of band migrations by gel electrophoresis and its appearance by electron microscopy. The unusual physical state of this topoisomer was probably mediated, in part, by DNA-specific structural proteins. The unique RB plasmid topoisomer was a supercoiled form of lower superhelical density than the other identified topoisomers. Developmental-stage-specific differences in super-helical density of plasmid DNA suggest cause-and-effect relationships between DNA topology and metabolic activity in RBs and metabolic quiescence in EBs.
Mol Microbiol 1990 Sep
PMID:Developmental-stage-specific plasmid supercoiling in Chlamydia trachomatis. 198 Oct 86

A 27kDa Chlamydia trachomatis L2 protein was characterized by the use of monoclonal antibodies and by two-dimensional gel electrophoresis. The protein was shown to be located in the membrane of reticulate bodies as well as elementary bodies. Its synthesis could be detected from 10 hours post-infection. Cloning and sequence analysis of the distal part of the gene revealed an open reading frame of 175 amino acids. Comparison of the deduced amino acid sequence with the NBRF data base revealed significant homology between the 27 kDa chlamydial membrane protein and the product of the macrophage infectivity potentiator (mip) gene of Legionella pneumophila.
Mol Microbiol 1991 Jan
PMID:Chlamydia trachomatis contains a protein similar to the Legionella pneumophila mip gene product. 201 97

Low copy-number bacterial plasmids F (the classical Escherichia coli sex factor) and prophage P1 encode partitioning functions which may provide fundamental insights into the active processes which ensure that bacterial genomes are segregated to both daughter cells prior to cell division. These partitioning systems involve two proteins: ParA and ParB. We report that incC from the broad host-range plasmid RK2 is a member of the family of ParA partitioning proteins and that these proteins (as well as related proteins encoded by plasmids from Agrobacterium tumefaciens and Chlamydia trachomatis) contain type I nucleotide-binding motifs. Also, we show that the cell division inhibitor MinD is homologous to members of the ParA family. Sequence comparisons of ParB proteins suggest that they may contain sites for phosphorylation. We propose that ATP hydrolysis by the ParA protein may result in phosphorylation of the ParB protein, thereby causing a conformational shift necessary to separate paired plasmid molecules at the cell division plane.
Mol Microbiol 1990 Sep
PMID:A family of ATPases involved in active partitioning of diverse bacterial plasmids. 214 83

A study was conducted to compare results between culture methods and the Gen-Probe (Gen-Probe Inc. San Diego, California) chemiluminescent technique of nucleic acid hybridization to identify Chlamydia trachomatis from genital specimens from 117 females and 70 males. Specimens collected from sexually transmitted diseases (STD) and infertility clinics were randomized as to whether probe or culture swabs were collected first. The Gen-Probe demonstrated a sensitivity of 83% and a specificity of 75% in the female population and a sensitivity of 68% and a specificity of 75% in the male population when compared to the reference culture method using cycloheximide-treated McCoy cells. Gen-Probe had an overall sensitivity of 74% and specificity of 75% when the two groups were combined. Chlamydiazyme (Abbott Labs) results were obtained on 135 specimens; 90 of which correlated with probe and culture. The remaining 45 specimens had varying combinations of probe, culture and Chlamydiazyme results. MicroTrak (Syva) was done on 49 specimens; 35 of which correlated with probe and culture. The remaining 14 specimens had varying combinations of probe, culture and MicroTrak results. The apparent lack of sensitivity of the DNA probe is a major drawback of this system.
Mol Cell Probes 1990 Feb
PMID:Comparison of a DNA probe with culture for detecting Chlamydia trachomatis directly from genital specimens. 217 11

Two cysteine-rich proteins of Chlamydia trachomatis are essential structural components of the unique outer membrane of the infectious elementary body. These 58,000 (outer membrane protein 2; OMP2) and 15,000 (OMP3) proteins also differ structurally and chemically between biovariants that differ in invasive capability. We have identified the gene for OMP3 and sequenced both trachoma and lymphogranuloma venereum (LGV) omp3 genes. We have previously sequenced omp2 from the LGV biovar and now describe the omp2 sequence for a trachoma biovariant. Amino acid sequence differences between biovariants were few but, significantly, these changes have altered the charge of both OMP2 and OMP3 such that the net charge of each protein differs between biovariants. These compensatory charge alterations have implications for the outer membrane organization of these proteins. In addition, examination of the OMP3 sequence suggests that OMP3 may be a lipoprotein.
Mol Microbiol 1990 Sep
PMID:Cysteine-rich outer membrane proteins of Chlamydia trachomatis display compensatory sequence changes between biovariants. 228 77

The major outer membrane protein (MOMP) of Chlamydia trachomatis carries serovar-, subspecies-, species- and genus immunodomains, antibodies to which may be protective. We have compared the inferred amino acid sequences for MOMP from different serovars of C. trachomatis and from Chlamydia psittaci to identify the likely locations of these sero-taxonomic epitopes. Overlapping peptides corresponding to each of these regions were synthesized on a solid phase and probed with monoclonal antibodies (MAbs) of appropriate specificities. We describe the primary structures of the binding sites of MAb to each of these four epitopes on C. trachomatis serovar L1 MOMP.
Mol Microbiol 1988 Sep
PMID:Epitope mapping with solid-phase peptides: identification of type-, subspecies-, species- and genus-reactive antibody binding domains on the major outer membrane protein of Chlamydia trachomatis. 246 Jul 19

Fragments of the gene encoding the major outer membrane porin protein (MOMP) of Chlamydia trachomatis serovar L1 were ligated into the pUC plasmid vectors to give a series of overlapping recombinants expressing MOMP from the lac promoter. Induction of this promoter with IPTG leads to high-level expression of the recombinant porin protein. Electron microscopy shows the presence of insoluble inclusions within the Escherichia coli host cells. Probing the expressed MOMP fragments with a set of monoclonal antibodies permitted localization of the four binding sites (epitopes) of primary-sequence-dependent monoclonal antibodies that exhibit genus-, species-, subspecies- and type (serovar)-specific reactivities.
Mol Microbiol 1988 Sep
PMID:High-level expression and epitope localization of the major outer membrane protein of Chlamydia trachomatis serovar L1. 246 Jul 20

The polymerase chain reaction is an in vitro procedure for primer-directed enzymatic amplification of specific template nucleic acid sequences. This technique was used to detect and differentiate Chlamydia trachomatis and Chlamydia psittaci in laboratory samples of infected McCoy cells. The polymerase chain reaction was shown to be both sensitive, detecting in the order of one chlamydial DNA molecule in 10(5) cells, and specific. No cross reaction (amplified product) was detected when a variety of mammalian cell and bacterial DNAs were used as template with the Chlamydia-specific oligonucleotide primers.
Mol Cell Probes 1989 Dec
PMID:A polymerase chain reaction (PCR) protocol for the specific detection of Chlamydia spp. 261 67

Sequence analysis of a 7.5 kb DNA plasmid isolated from Chlamydia trachomatis shows 8 open reading frames (ORFs) regularly spaced along most of the sequence. One of these ORFs encodes a 451-amino-acid polypeptide highly homologous to the DnaB protein of Escherichia coli. A region between ORFs 6 and 7 contains a cluster of alternating ATs and a 22 bp sequence tandemly repeated 4 times, suggesting a replication control region. Several ORFs correspond to plasmid-specific polypeptides that have been described. Codons ending with A or T are more frequent, as might be expected from the high A/T content (64%) of the plasmid, and codon usage is similar to that of the C. trachomatis chromosomal gene, omp1L2.
Mol Microbiol 1988 Jul
PMID:The structure of a plasmid of Chlamydia trachomatis believed to be required for growth within mammalian cells. 284 28

The detection of Chlamydia trachomatis by in situ DNA hybridization in urogenital smears was investigated using a commercially available biotinylated DNA probe. Intracellular staining of inclusion bodies was used as the criterion for positivity. Of 35 patients with a culture proven chlamydial infection 19 had smears in which C. trachomatis was detected by in situ DNA hybridization, indicating a sensitivity of 54%. Of 57 patients with a negative culture, two had positive smears by in situ DNA hybridization. To compare whether the criterion for positivity had influenced the sensitivity obtained with in situ DNA hybridization, 14 duplicate smears from culture positive patients were analysed with in situ DNA hybridization and immunofluorescence. Both methods detected intracellular inclusion bodies in seven of these smears, suggesting that the presence of infected cells mainly determines the sensitivity. The DNA probe did not cross-react with micro-organisms commonly found in the urogenital tract.
Mol Cell Probes 1988 Dec
PMID:Detection of Chlamydia trachomatis in culture and urogenital smears by in situ DNA hybridization using a biotinylated DNA probe. 307 11


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