scholarly journals The effect of penicillin on Chlamydia trachomatis DNA replication

Microbiology ◽  
2006 ◽  
Vol 152 (9) ◽  
pp. 2573-2578 ◽  
Author(s):  
Paul R. Lambden ◽  
Mark A. Pickett ◽  
Ian N. Clarke
Keyword(s):  
Dna Replication ◽  
Chlamydia Trachomatis ◽  
Plasmid Dna ◽  
Host Cells ◽  
Developmental Cycle ◽  
Chromosomal Dna ◽  
Rapid Phase ◽  
Reticulate Body ◽  
Transmission Electron ◽  
Infected Cells

Chlamydia trachomatis L2 was used to infect BGMK cells at an m.o.i. of 1.0, and the developmental cycle was followed by transmission electron microscopy and quantitative PCR (QPCR) for both chromosomal and plasmid DNA. Samples were taken at sequential 6 h time points. Subsequent analysis by QPCR showed that there was an initial slow replication period (0–18 h), followed by a rapid phase (18–36 h) coinciding with exponential division when the DNA doubling time was 4.6 h. Chromosomal DNA was amplified 100–200-fold corresponding to 7–8 generations for the complete developmental cycle. Penicillin (10 and 100 units ml−1) was added to cultures at 20 h post-infection (p.i.). This blocked binary fission and also prevented reticulate body (RB) to elementary body transition. However, exposure to penicillin did not prevent chromosomal or plasmid DNA replication. After a short lag period, following the addition of penicillin, chlamydial chromosomal DNA replication resumed at the same rate as in control C. trachomatis-infected cells. C. trachomatis-infected host cells exposed to penicillin did not lyse, but instead harboured large, aberrant RBs in massive inclusions that completely filled the cell cytoplasm. In these RBs, the DNA continued to replicate well beyond the end of the normal developmental cycle. At 60 h p.i. each aberrant RB contained a minimum of 16 chromosomal copies.

scholarly journals Effects ofMentha suaveolensEssential Oil onChlamydia trachomatis

2015 ◽  
Vol 2015 ◽  
pp. 1-7 ◽  
Author(s):  
Rosa Sessa ◽  
Marisa Di Pietro ◽  
Fiorenzo De Santis ◽  
Simone Filardo ◽  
Rino Ragno ◽  
...  
Keyword(s):  
Chlamydia Trachomatis ◽  
Substantial Reduction ◽  
Developmental Cycle ◽  
Elementary Body ◽  
Promising Candidate ◽  
Common Cause ◽  
Sexually Transmitted ◽  
Reticulate Body ◽  
Preventative Strategy

Chlamydia trachomatis, the most common cause of sexually transmitted bacterial infection worldwide, has a unique biphasic developmental cycle alternating between the infectious elementary body and the replicative reticulate body.C. trachomatisis responsible for severe reproductive complications including pelvic inflammatory disease, ectopic pregnancy, and obstructive infertility. The aim of our study was to evaluate whetherMentha suaveolensessential oil (EOMS) can be considered as a promising candidate for preventingC. trachomatisinfection. Specifically, we investigated thein vitroeffects of EOMS towardsC. trachomatisanalysing the different phases of chlamydial developmental cycle. Our results demonstrated that EOMS was effective towardsC. trachomatis, whereby it not only inactivated infectious elementary bodies but also inhibited chlamydial replication. Our study also revealed the effectiveness of EOMS, in combination with erythromycin, towardsC. trachomatiswith a substantial reduction in the minimum effect dose of antibiotic. In conclusion, EOMS treatment may represent a preventative strategy since it may reduceC. trachomatistransmission in the population and, thereby, reduce the number of new chlamydial infections and risk of developing of severe sequelae.

scholarly journals Chlamydial infection and disease in the koala

2005 ◽  
Vol 26 (2) ◽  
pp. 65 ◽  
Author(s):  
Peter Timms
Keyword(s):  
Chlamydia Trachomatis ◽  
Chlamydial Infection ◽  
Developmental Cycle ◽  
Elementary Body ◽  
Obligate Intracellular ◽  
Molecular Approaches ◽  
Reticulate Body ◽  
Wide Range ◽  
The Family ◽  
Serious Disease

Chlamydiae are obligate intracellular bacterial pathogens able to infect and cause serious disease in humans, birds and a remarkably wide range of warm and cold-blooded animals. The family Chlamydiaciae have traditionally been defined by their unique biphasic developmental cycle, involving the interconversion between an extracellular survival form, the elementary body and an intracellular replicative form, the reticulate body. However, as with many other bacteria, molecular approaches including 16SrRNA sequence are becoming the standard of choice. As a consequence, the chlamydiae are in a taxonomic state of flux. Prior to 1999, the family Chlamydiaceae consisted of one genus, Chlamydia, and four species, Chlamydia trachomatis, C. psittaci, C. pecorum and C. pneumoniae. In 1999, Everett et al proposed a reclassification of Chlamydia into two genera (Chlamydia and Chlamydophila) and nine species (Chlamydia trachomatis, C. suis, and C. muridarum and Chlamydophila psittaci, C. pneumoniae, C. felis, C. pecorum, C. abortus, and C. caviae). While some of these species are thought to be host specific (C. suis ? pigs, C. muridarum ? mice, C. felis ? cats, C. caviae ? guinea pigs) many are known to infect and cause disease in a wide range of hosts.

Persistent Infection of Mouse Fibroblasts (L Cells) with Chlamydia psittaci : Evidence for a Cryptic Chlamydial Form

1980 ◽  
Vol 30 (3) ◽  
pp. 874-883
Author(s):  
James W. Moulder ◽  
Nancy J. Levy ◽  
Laura P. Schulman
Keyword(s):  
Host Cell ◽  
Chlamydia Psittaci ◽  
Host Cells ◽  
Developmental Cycle ◽  
Persistently Infected ◽  
Mouse Fibroblasts ◽  
Persistent Infections ◽  
L Cells ◽  
Infected Cells ◽  
Parasite Multiplication

When monolayers of mouse fibroblasts (L cells) were infected with enough Chlamydia psittaci (strain 6BC) to destroy most of the host cells, 1 in every 10 5 to 10 6 originally infected cells gave rise to a colony of L cells persistently infected with strain 6BC. In these populations, the density of L cells and 6BC fluctuated periodically and reciprocally as periods of host cell increase were followed by periods of parasite multiplication. Successive cycles of L-cell and 6BC reproduction were sustained indefinitely by periodic transfer to fresh medium. Isolation of L cells and 6BC from persistent infections provided no evidence that there had been any selection of variants better suited for coexistence. Persistently infected populations consisting mainly of inclusion-free L cells yielded only persistently infected clones, grew more slowly, and cloned less efficiently. They were also almost completely resistant to superinfection with high multiplicities of either 6BC or the lymphogranuloma venereum strain 440L of Chlamydia trachomatis . These properties of persistently infected L cells may be accounted for by assuming that all of the individuals in these populations are cryptically infected with 6BC and that cryptic infection slows the growth of the host cell and makes it immune to infection with exogenous chlamydiae. According to this hypothesis, the fluctuations in host and parasite density occur because some factor periodically sets off the conversion of cryptic chlamydial forms into reticulate bodies that multiply and differentiate into infectious elementary bodies in a conventional chlamydial developmental cycle.

Activation of caspase-3 during Chlamydia trachomatis-induced apoptosis at a late stage

2019 ◽  
Vol 65 (2) ◽  
pp. 135-143 ◽  
Author(s):  
Junji Matsuo ◽  
Sanae Haga ◽  
Kent Hashimoto ◽  
Torahiko Okubo ◽  
Takeaki Ozawa ◽  
...  
Keyword(s):  
Chlamydia Trachomatis ◽  
Late Stage ◽  
Caspase 3 ◽  
Luciferase Activity ◽  
Firefly Luciferase ◽  
Host Cells ◽  
Chlamydia Infection ◽  
Developmental Cycle ◽  
Apoptosis Pathway ◽  
Induced Apoptosis

The obligate intracellular bacterium Chlamydia trachomatis activates the host cell apoptosis pathway at a late stage of its developmental cycle. However, whether caspase-3, which is a key enzyme of apoptosis, is activated in Chlamydia-infected cells remains unknown. Here, we established HEp-2 cells stably expressing cFluc-DEVD, which is a caspase-3 substrate sequence inserted into cyclic firefly luciferase, and then monitored the dynamics of caspase-3 activity in cells infected with Chlamydia. Transfected cells without infection showed a significant increase in luciferase activity due to stimulation with staurosporine, an inducer of apoptosis. Activation was significantly blocked by addition of caspase inhibitor z-VAD-fmk. Furthermore, as expected, Chlamydia infection caused a significant increase in luciferase activation at 36–48 h postinfection with a contrastive decrease at 24 h postinfection, which is already well known. Such activation caused by the infection was much stronger when the amount of bacteria was increased. Thus, caspase-3 activation was accurately monitored by the luciferase activity in HEp-2 cells constitutively expressing the cFluc-DEVD probe. Furthermore, our data showed that C. trachomatis activates caspase-3 in host cells at a late stage of infection.

scholarly journals Chlamydial Effector Proteins Localized to the Host Cell Cytoplasmic Compartment

2008 ◽  
Vol 76 (11) ◽  
pp. 4842-4850 ◽  
Author(s):  
Betsy Kleba ◽  
Richard S. Stephens
Keyword(s):  
Bacterial Pathogens ◽  
Eukaryotic Cell ◽  
Effector Proteins ◽  
Host Cells ◽  
Developmental Cycle ◽  
Secretion Systems ◽  
Bacterial Proteins ◽  
Cell Functions ◽  
Infected Cells ◽  
Cytoplasmic Compartment

ABSTRACT Disease-causing microbes utilize various strategies to modify their environment in order to create a favorable location for growth and survival. Gram-negative bacterial pathogens often use specialized secretion systems to translocate effector proteins directly into the cytosol of the eukaryotic cells they infect. These bacterial proteins are responsible for modulating eukaryotic cell functions. Identification of the bacterial effectors has been a critical step toward understanding the molecular basis for the pathogenesis of the bacteria that use them. Chlamydiae are obligate intracellular bacterial pathogens that have a type III secretion system believed to translocate virulence effector proteins into the cytosol of their host cells. Selective permeabilization of the eukaryotic cell membrane was used in conjunction with metabolic labeling of bacterial proteins to identify chlamydial proteins that localize within the cytosol of infected cells. More than 20 Chlamydia trachomatis and C. pneumoniae proteins were detected within the cytoplasmic compartment of infected cells. While a number of cytosolic proteins were shared, others were unique to each species, suggesting that variation among cytosolic chlamydial proteins contributes to the differences in the pathogenesis of the chlamydial species. The spectrum of chlamydial proteins exported differed concomitant with the progress of the developmental cycle. These data confirm that a dynamic relationship exists between Chlamydia and its host and that translocation of bacterial proteins into the cytosol is developmentally dependent.

scholarly journals Role of High-Mobility Group Box 1 Protein and Poly(ADP-Ribose) Polymerase 1 Degradation in Chlamydia trachomatis-Induced Cytopathicity

2010 ◽  
Vol 78 (7) ◽  
pp. 3288-3297 ◽  
Author(s):  
Hangxing Yu ◽  
Katja Schwarzer ◽  
Martin Förster ◽  
Olaf Kniemeyer ◽  
Vera Forsbach-Birk ◽  
...  
Keyword(s):  
Chlamydia Trachomatis ◽  
Cell Membrane ◽  
Hela Cells ◽  
High Mobility ◽  
Host Cells ◽  
High Mobility Group ◽  
Cell Membrane Permeability ◽  
Dependent Manner ◽  
Infected Cells ◽  
Parp 1

ABSTRACT As intracellular bacteria, chlamydiae block the apoptotic pathways of their host cells. However, the infection of epithelial cells causes the loss of cell membrane integrity and can result in nonapoptotic death. Normally, cells undergoing necrosis release high-mobility group box 1 protein (HMGB1) that acts as an important proinflammatory mediator. Here, we show that in Chlamydia trachomatis-infected HeLa cells HMGB1 is not translocated from the nucleus to the cytosol and not released from injured cells in increased amounts. At 48 h after infection, degradation of HMGB1 was observed. In infected cells, poly(ADP-ribose) polymerase 1 (PARP-1), a DNA repair enzyme that also regulates HMGB1 translocation, was found to be cleaved into fragments that correspond to a necrosislike pattern of PARP-1 degradation. Cell-free cleavage assays and immunoprecipitation using purified proteolytic fractions from infected cells demonstrated that the chlamydial-protease-like activity factor (CPAF) is responsible for the cleavage of both HMGB1 and PARP-1. Proteolytic cleavage of PARP-1 was accompanied by a significant decrease in the enzymatic activity in a time-dependent manner. The loss of PARP-1 function obviously affects the viability of Chlamydia-infected cells because silencing of PARP-1 in uninfected HeLa cells with specific small interfering RNA results in increased cell membrane permeability. Our findings suggest that the Chlamydia-specific protease CPAF interferes with necrotic cell death pathways. By the degradation of HMGB1 and PARP-1, the pathogen may have evolved a strategy to reduce the inflammatory response to membrane-damaged cells in vivo.

Involvement of annexin II in DNA replication: evidence from cell-free extracts of Xenopus eggs

1993 ◽  
Vol 105 (2) ◽  
pp. 533-540 ◽  
Author(s):  
J.K. Vishwanatha ◽  
S. Kumble
Keyword(s):  
Cell Cycle ◽  
Dna Replication ◽  
Plasmid Dna ◽  
Annexin Ii ◽  
Chromosomal Dna ◽  
Stringent Requirement ◽  
Exogenous Dna ◽  
Nuclear Assembly ◽  
Egg Extracts ◽  
Higher Eukaryotes

Cell-free extracts of Xenopus eggs efficiently initiate and complete semiconservative DNA replication of exogenously added plasmid DNA. DNA replication in such extracts can be neutralized by a monoclonal antibody (D1/274.5) against human annexin II. Specific immunodepletion of Xenopus annexin II from the egg extracts results in loss of DNA replicative ability. Immunodepletion of annexin II does not prevent nuclear assembly, a stringent requirement for DNA synthesis on exogenous DNA in this system. Replicative ability can be restored to the immunodepleted extracts by the addition of purified human annexin II. These results demonstrate that annexin II is involved in chromosomal DNA replication and has a role in the cell cycle of higher eukaryotes.

scholarly journals Chlamydia trachomatis Inhibits Homologous Recombination Repair of DNA Breaks by Interfering with PP2A Signaling

mBio ◽  
2018 ◽  
Vol 9 (6) ◽  
Author(s):  
Yang Mi ◽  
Rajendra Kumar Gurumurthy ◽  
Piotr K. Zadora ◽  
Thomas F. Meyer ◽  
Cindrilla Chumduri
Keyword(s):  
Cell Cycle ◽  
Chlamydia Trachomatis ◽  
Homologous Recombination ◽  
Double Strand Breaks ◽  
Host Cells ◽  
Dna Double Strand Breaks ◽  
Content Type ◽  
Strand Breaks ◽  
Recombination Repair ◽  
Infected Cells

ABSTRACT Cervical and ovarian cancers exhibit characteristic mutational signatures that are reminiscent of mutational processes, including defective homologous recombination (HR) repair. How these mutational processes are initiated during carcinogenesis is largely unclear. Chlamydia trachomatis infections are epidemiologically associated with cervical and ovarian cancers. Previously, we showed that C. trachomatis induces DNA double-strand breaks (DSBs) but suppresses Ataxia-telangiectasia mutated (ATM) activation and cell cycle checkpoints. The mechanisms by which ATM regulation is modulated and its consequences for the repair pathway in C. trachomatis-infected cells remain unknown. Here, we found that Chlamydia bacteria interfere with the usual response of PP2A to DSBs. As a result, PP2A activity remains high, as the level of inhibitory phosphorylation at Y307 remains unchanged following C. trachomatis-induced DSBs. Protein-protein interaction analysis revealed that C. trachomatis facilitates persistent interactions of PP2A with ATM, thus suppressing ATM activation. This correlated with a remarkable lack of homologous recombination (HR) repair in C. trachomatis-infected cells. Chemical inhibition of PP2A activity in infected cells released ATM from PP2A, resulting in ATM phosphorylation. Activated ATM was then recruited to DSBs and initiated downstream signaling, including phosphorylation of MRE11 and NBS1 and checkpoint kinase 2 (Chk2)-mediated activation of the G2/M cell cycle checkpoint in C. trachomatis-infected cells. Further, PP2A inhibition led to the restoration of C. trachomatis-suppressed HR DNA repair function. Taking the data together, this study revealed that C. trachomatis modulates PP2A signaling to suppress ATM activation to prevent cell cycle arrest, thus contributing to a deficient high-fidelity HR pathway and a conducive environment for mutagenesis. IMPORTANCE Chlamydia trachomatis induces DNA double-strand breaks in host cells but simultaneously inhibits proper DNA damage response and repair mechanisms. This may render host cells prone to loss of genetic integrity and transformation. Here we show that C. trachomatis prevents activation of the key DNA damage response mediator ATM by preventing the release from PP2A, leading to a complete absence of homologous recombination repair in host cells.

scholarly journals Inhibition of Fusion of Chlamydia trachomatis Inclusions at 32°C Correlates with Restricted Export of IncA

2002 ◽  
Vol 70 (7) ◽  
pp. 3816-3823 ◽  
Author(s):  
K. A. Fields ◽  
E. Fischer ◽  
T. Hackstadt
Keyword(s):  
Chlamydia Trachomatis ◽  
Hela Cells ◽  
Immunofluorescence Microscopy ◽  
Parasitophorous Vacuole ◽  
Obligate Intracellular Bacterium ◽  
Inclusion Membrane ◽  
Obligate Intracellular ◽  
Transmission Electron ◽  
Infected Cells ◽  
Exocytic Pathway

ABSTRACT Chlamydia trachomatis is an obligate intracellular bacterium that develops within a parasitophorous vacuole termed an inclusion. The inclusion is nonfusogenic with lysosomes but intercepts lipids from a host cell exocytic pathway. Initiation of chlamydial development is concurrent with modification of the inclusion membrane by a set of C. trachomatis-encoded proteins collectively designated Incs. One of these Incs, IncA, is functionally associated with the homotypic fusion of inclusions. Inclusions also do not fuse when cultures are multiply infected with C. trachomatis and cultivated at 32°C. We obtained evidence linking these experimental observations by characterizing IncA localization in 32°C cultures. Analysis of inclusions by light and transmission electron microscopy confirmed that HeLa cells infected with multiple C. trachomatis elementary bodies and cultivated at 32°C for 24 h contained multiple, independent inclusions. Reverse transcriptase PCR and immunoblot analyses of C. trachomatis-infected HeLa cells demonstrated the presence of IncA at 24 h in 32°C cultures. When parallel cultures were probed with IncA-specific antibodies in indirect immunofluorescence assays, IncA was detectable in intracellular chlamydiae but not within the inclusion membrane. In addition, analysis of purified reticulate bodies from 37 and 32°C cultures showed that bacterium-associated pools of IncA are enriched in cultures grown at 32°C. Microscopic observation of infected cells revealed that some vacuoles had fused by 48 h postinfection, and this finding was correlated with the detection of IncA in inclusion membranes by immunofluorescence microscopy. The data are consistent with a requirement for IncA in fusions of C. trachomatis inclusions and suggest that the effect of incubation at 32°C is manifested by restricted export of IncA to the inclusion membrane.

Expression of recombinant DNA introduced into Chlamydia trachomatis by electroporation

1994 ◽  
Vol 40 (7) ◽  
pp. 583-591 ◽  
Author(s):  
Jeffrey E. Tam ◽  
Carolyn H. Davis ◽  
Priscilla B. Wyrick
Keyword(s):  
Chlamydia Trachomatis ◽  
Plasmid Dna ◽  
Southern Hybridization ◽  
Recombinant Dna ◽  
Chloramphenicol Acetyltransferase ◽  
Developmentally Regulated ◽  
Reticulate Body ◽  
Body Development ◽  
Coli Plasmid

Electroporation was used to introduce DNA into the elementary bodies of the obligate parasitic bacterium Chlamydia trachomatis. The source of DNA for these experiments was the chimeric plasmid pPBW100, which was constructed from the well-characterized 7.5-kb plasmid of C. trachomatis and the Escherichia coli plasmid pBGS9. To select directly for C. trachomatis carrying pPBW100, an in-frame gene fusion between the chlamydial promoter P7248 and a promoterless chloramphenicol acetyltransferase (cat) cassette was incorporated into the plasmid. After infection of McCoy cells with electroporated elementary bodies containing pPBW100, the following were observed: (i) the plasmid DNA was detected inside the chloramphenicol-resistant chlamydial inclusions by in situ and Southern hybridization analyses; (ii) both physical and biochemical evidence showed that chloramphenicol acetyltransferase was synthesized by the electroporated C. trachomatis; (iii) expression of P7248::cat was developmentally regulated and occurred during the early stages of chlamydial reticulate body development; and (iv) although the expression from P7248::cat was mainly transient, there were rare instances where chloramphenicol-resistant C. trachomatis were observed after four passages.Key words: chlamydia, electroporation, chimeric plasmid, expression.

Sign in / Sign up

Export Citation Format

Share Document