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.

scholarly journals Paclitaxel Arrests Growth of IntracellularToxoplasma gondii

1998 ◽  
Vol 42 (8) ◽  
pp. 2036-2040 ◽  
Author(s):  
Randee Estes ◽  
Nicolas Vogel ◽  
Douglas Mack ◽  
Rima McLeod
Keyword(s):  
Toxoplasma Gondii ◽  
Host Cell ◽  
Fresh Medium ◽  
Host Cells ◽  
Normal Morphology ◽  
Intracellular Form ◽  
Human Foreskin Fibroblasts ◽  
The Future ◽  
Infected Cells ◽  
Parasite Multiplication

ABSTRACT Addition of paclitaxel (Taxol) at a concentration of 1 μM toToxoplasma gondii-infected human foreskin fibroblasts arrested parasite multiplication. Division of theT. gondii tachyzoite nucleus was inhibited, leading to syncytium-like parasite structures within the fibroblasts by 24 h after infection and treatment of the cultures. By 4 days after infection and treatment of the cultures with paclitaxel, this inhibition was irreversible, since the arrested intracellular form was incapable of leaving the host cell, infecting new cells, and initiating the growth of tachyzoites with normal morphology. Specifically, when paclitaxel was added to infected cells for 4 days and then removed by washing and the infected, paclitaxel-treated cells were cultured for 4 more days, there were no remaining T. gondii organisms with normal morphology. Syncytium-like structures in the cultures that were infected and treated with paclitaxel for 8 days were similar in appearance to those in preparations of infected paclitaxel-treated fibroblasts that had been cultured for 24 to 48 h. Pretreatment of the tachyzoites for 1 h with paclitaxel followed by the removal of the paclitaxel by repeatedly centrifuging and resuspending the parasites in fresh medium without paclitaxel and then adding fresh medium prior to culture of the parasites with fibroblasts did not prevent their invasion of fibroblasts but did affect their subsequent ability to replicate within fibroblasts. Pretreatment of the fibroblasts with paclitaxel also diminished subsequent replication ofT. gondii in such host cells after 8 days. Thus, paclitaxel alters the ability of T. gondii to replicate in host cells. Inhibition of parasite microtubules by such compounds at concentrations which do not interfere with the function of host cell microtubules may be useful for development of novel medicines to treat T. gondii infections in the future.

Respiration of L cells infected with Chlamydia psittaci

1970 ◽  
Vol 16 (11) ◽  
pp. 1033-1039 ◽  
Author(s):  
Shawn D'arcy Gill ◽  
Robert B. Stewart
Keyword(s):  
Tissue Culture ◽  
Oxidative Phosphorylation ◽  
Host Cell ◽  
Respiratory Quotient ◽  
Culture Medium ◽  
Mouse Fibroblast ◽  
Chlamydia Psittaci ◽  
Concomitant Increase ◽  
L Cells ◽  
Infected Cells

Strain 929 mouse fibroblast L cells demonstrated an increased rate of endogenous and exogenous respiration when infected by the 6BC strain of the psittacosis organism. The respiratory quotient (R.Q.) values for infected cultures were about 1.0. Studies involving the use of 2,4-DNPand reduced phosphate suggested that the accelerated respiration was a result of increased oxidative phosphorylation. Infection was accompanied by a decline in host cell ATP levels with a concomitant increase in the ADP levels. A reduction in tissue culture medium phosphate resulted in a reduced yield of psittacosis organisms from infected cells and lower cellular levels of ATP. These results are discussed in relation to an energy parasite hypothesis.

scholarly journals Attachment defect in mouse fibroblasts (L cells) persistently infected with Chlamydia psittaci.

1981 ◽  
Vol 34 (1) ◽  
pp. 285-291 ◽  
Author(s):  
J W Moulder ◽  
N J Levy ◽  
S L Zeichner ◽  
C K Lee
Keyword(s):  
Chlamydia Psittaci ◽  
Persistently Infected ◽  
Mouse Fibroblasts ◽  
L Cells

scholarly journals Revisiting Ehrlichia ruminantium Replication Cycle Using Proteomics: The Host and the Bacterium Perspectives

2021 ◽  
Vol 9 (6) ◽  
pp. 1144
Author(s):  
Isabel Marcelino ◽  
Philippe Holzmuller ◽  
Ana Coelho ◽  
Gabriel Mazzucchelli ◽  
Bernard Fernandez ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Host Cell ◽  
Cell Metabolism ◽  
Replication Cycle ◽  
Host Cells ◽  
Ehrlichia Ruminantium ◽  
Host Interaction ◽  
Infected Cells ◽  
Related Proteins

The Rickettsiales Ehrlichia ruminantium, the causal agent of the fatal tick-borne disease Heartwater, induces severe damage to the vascular endothelium in ruminants. Nevertheless, E. ruminantium-induced pathobiology remains largely unknown. Our work paves the way for understanding this phenomenon by using quantitative proteomic analyses (2D-DIGE-MS/MS, 1DE-nanoLC-MS/MS and biotin-nanoUPLC-MS/MS) of host bovine aorta endothelial cells (BAE) during the in vitro bacterium intracellular replication cycle. We detect 265 bacterial proteins (including virulence factors), at all time-points of the E. ruminantium replication cycle, highlighting a dynamic bacterium–host interaction. We show that E. ruminantium infection modulates the expression of 433 host proteins: 98 being over-expressed, 161 under-expressed, 140 detected only in infected BAE cells and 34 exclusively detected in non-infected cells. Cystoscape integrated data analysis shows that these proteins lead to major changes in host cell immune responses, host cell metabolism and vesicle trafficking, with a clear involvement of inflammation-related proteins in this process. Our findings led to the first model of E. ruminantium infection in host cells in vitro, and we highlight potential biomarkers of E. ruminantium infection in endothelial cells (such as ROCK1, TMEM16K, Albumin and PTPN1), which may be important to further combat Heartwater, namely by developing non-antibiotic-based strategies.

scholarly journals Type I Interferon acts as a major barrier to the establishment of infectious bursal disease virus (IBDV) persistent infections

2020 ◽  
pp. JVI.02017-20
Author(s):  
Laura Broto ◽  
Nicolás Romero ◽  
Fernando Méndez ◽  
Elisabet Diaz-Beneitez ◽  
Oscar Candelas-Rivera ◽  
...  
Keyword(s):  
Disease Virus ◽  
Cell Lines ◽  
Infectious Bursal Disease Virus ◽  
Infectious Bursal Disease ◽  
Type I ◽  
Persistently Infected ◽  
Persistent Infections ◽  
Avian Pathogen ◽  
Bursal Disease ◽  
Infected Cells

Infectious bursal disease virus (IBDV), the best characterized member of the Birnaviridae family, is a highly relevant avian pathogen causing both acute and persistent infections in different avian hosts. Here, we describe the establishment of clonal, long-term, productive persistent IBDV infections in DF-1 chicken embryonic fibroblasts. Although virus yields in persistently-infected cells are exceedingly lower than those detected in acutely infected cells, the replication fitness of viruses isolated from persistently-infected cells is higher than that of the parental virus. Persistently-infected DF-1 and IBDV-cured cell lines derived from them do not respond to type I interferon (IFN). High-throughput genome sequencing revealed that this defect is due to mutations affecting the IFNα/β receptor subunit 2 (IFNAR2) gene resulting in the expression of IFNAR2 polypeptides harbouring large C-terminal deletions that abolish the signalling capacity of IFNα/β receptor complex. Ectopic expression of a recombinant chicken IFNAR2 gene efficiently rescues IFNα responsiveness. IBDV-cured cell lines derived from persistently infected cells exhibit a drastically enhanced susceptibility to establishing new persistent IBDV infections. Additionally, experiments carried out with human HeLa cells lacking the IFNAR2 gene fully recapitulate results obtained with DF-1 cells, exhibiting a highly enhanced capacity to both survive the acute IBDV infection phase and to support the establishment of persistent IBDV infections. Results presented here show that the inactivation of the JAK-STAT signalling pathway significantly reduces the apoptotic response induced by the infection, hence facilitating the establishment and maintenance of IBDV persistent infections.IMPORTANCE Members of the Birnaviridae family, including infectious bursal disease virus (IBDV), exhibit a dual behaviour, causing acute infections that are often followed by the establishment of life-long persistent asymptomatic infections. Indeed, persistently infected specimens might act as efficient virus reservoirs, hence potentially contributing to virus dissemination. Despite the key importance of this biological trait, information about mechanisms triggering IBDV persistency is negligible. Our report evidences the capacity of IBDV, a highly relevant avian pathogen, to establishing long-term, productive, persistent infections in both avian and human cell lines. Data presented here provide novel and direct evidence about the crucial role of type I IFNs on the fate of IBDV-infected cells and their contribution to controlling the establishment of IBDV persistent infections. The use of cell lines unable to respond to type I IFNs opens a promising venue to unveiling additional factors contributing to IBDV persistency.

scholarly journals Role for nsP2 Proteins in the Cessation of Alphavirus Minus-Strand Synthesis by Host Cells

2006 ◽  
Vol 80 (1) ◽  
pp. 360-371 ◽  
Author(s):  
Dorothea L. Sawicki ◽  
Silvia Perri ◽  
John M. Polo ◽  
Stanley G. Sawicki
Keyword(s):  
Host Response ◽  
Rna Synthesis ◽  
Late Phase ◽  
Host Cells ◽  
Minus Strand ◽  
Wild Type ◽  
Persistent Infections ◽  
Infected Cells ◽  
Replicative Intermediates ◽  
Transcription Complexes

ABSTRACT In order to establish nonlytic persistent infections (PI) of BHK cells, replicons derived from Sindbis (SIN) and Semliki Forest (SFV) viruses have mutations in nsP2. Five different nsP2 PI replicons were compared to wild-type (wt) SIN, SFV, and wt nsPs SIN replicons. Replicon PI BHK21 cells had viral RNA synthesis rates that were less than 5% of those of the wt virus and ∼10% or less of those of SIN wt replicon-infected cells, and, in contrast to wt virus and replicons containing wt nsP2, all showed a phenotype of continuous minus-strand synthesis and of unstable, mature replication/transcription complexes (RC+) that are active in plus-strand synthesis. Minus-strand synthesis and incorporation of [3H]uridine into replicative intermediates differed among PI replicons, depending on the location of the mutation in nsP2. Minus-strand synthesis by PI cells appeared normal; it was dependent on continuous P123 and P1234 polyprotein synthesis and ceased when protein synthesis was inhibited. The failure by the PI replicons to shut off minus-strand synthesis was not due to some defect in the PI cells but rather was due to the loss of some function in the mutated nsP2. This was demonstrated by showing that superinfection of PI cells with wt SFV triggered the shutdown of minus-strand synthesis, which we believe is a host response to infection with alphaviruses. Together, the results indicate alphavirus nsP2 functions to engage the host response to infection and activate a switch from the early-to-late phase. The loss of this function leads to continuous viral minus-strand synthesis and the production of unstable RC+.

scholarly journals Characterization of a Chlamydia psittaciDNA Binding Protein (EUO) Synthesized during the Early and Middle Phases of the Developmental Cycle

1998 ◽  
Vol 66 (3) ◽  
pp. 1167-1173 ◽  
Author(s):  
Li Zhang ◽  
Annemarie L. Douglas ◽  
Thomas P. Hatch
Keyword(s):  
Promoter Region ◽  
Chlamydia Psittaci ◽  
Host Cells ◽  
Upstream Open Reading Frame ◽  
Developmental Cycle ◽  
Binding Property ◽  
Logarithmic Growth Phase ◽  
Reading Frame

ABSTRACT The EUO gene (for early upstream open reading frame) ofChlamydia psittaci was previously found to be transcribed better at 1 than at 24 h postinfection. We found that the EUO gene encodes a minor protein that is expressed within 1 h of infection of host cells with C. psittaci 6BC but that protein quantity peaks during the logarithmic growth phase of reticulate bodies (RBs), declines late in the infection (after 20 h) when RBs reorganize into elementary bodies (EBs), and is absent in infectious EBs. EUO protein lacks homology to known proteins but does contain a putative helix-turn-helix motif. We found that recombinant EUO binds to DNA in vitro with a relatively broad specificity. Using the bp −200 to +67 promoter region of the cysteine-rich envelope protein (crp) operon as a model, we show that EUO protein preferentially binds to AT-rich sequences and protects crpDNA from DNase I from approximately bp −60 to −9. We also found that native EUO protein in extracts of RBs binds to the promoter region of the crp operon, demonstrating that the DNA binding property of EUO protein is not an artifact of recombinant methods. Although EUO protein appears to bind to the crp operon with high affinity in vitro (Kd of about 15 nM), it is not known whether the protein binds the crp DNA in vivo.

scholarly journals Type I Interferon acts as a major barrier to the establishment of infectious bursal disease virus (IBDV) persistent infections

2020 ◽  
Author(s):  
Laura Broto ◽  
Nicolás Romero ◽  
Fernando Méndez ◽  
Elisabet Diaz-Beneitez ◽  
Oscar Candelas-Rivera ◽  
...  
Keyword(s):  
Disease Virus ◽  
Cell Lines ◽  
Infectious Bursal Disease Virus ◽  
Infectious Bursal Disease ◽  
Type I ◽  
Persistently Infected ◽  
Persistent Infections ◽  
Avian Pathogen ◽  
Bursal Disease ◽  
Infected Cells

ABSTRACTInfectious bursal disease virus (IBDV), the best characterized member of the Birnaviridae family, is a highly relevant avian pathogen causing both acute and persistent infections in different avian hosts. Here, we describe the establishment of clonal, long-term, productive persistent IBDV infections in DF-1 chicken embryonic fibroblasts. Although virus yields in persistently-infected cells are exceedingly lower than those detected in acutely infected cells, the replication fitness of viruses isolated from persistently-infected cells is higher than that of the parental virus. Persistently-infected DF-1 and IBDV-cured cell lines derived from them do not respond to type I interferon (IFN). High-throughput genome sequencing revealed that this defect is due to mutations affecting the IFNα/β receptor subunit 2 (IFNAR2) gene resulting in the expression of IFNAR2 polypeptides harbouring large C-terminal deletions that abolish the signalling capacity of IFNα/β receptor complex. Ectopic expression of a recombinant chicken IFNAR2 gene efficiently rescues IFNα responsiveness. IBDV-cured cell lines derived from persistently infected cells exhibit a drastically enhanced proneness to establishing new persistent IBDV infections. Additionally, experiments carried out with human HeLa cells lacking the IFNAR2 gene fully recapitulate results obtained with DF-1 cells, exhibiting a highly enhanced capacity to both survive the acute IBDV infection phase and to support the establishment of persistent IBDV infections. Results presented here show that the inactivation of the JAK-STAT signalling pathway significantly reduces the apoptotic response induced by the infection, hence facilitating the establishment and maintenance of IBDV persistent infections.IMPORTANCEMembers of the Birnaviridae family, including infectious bursal disease virus (IBDV), exhibit a dual behaviour, causing acute infections that are often followed by the establishment of life-long persistent asymptomatic infections. Indeed, persistently infected specimens might act as efficient virus reservoirs, hence potentially contributing to virus dissemination. Despite the key importance of this biological trait, information about mechanisms triggering IBDV persistency is negligible. Our report evidences the capacity of IBDV, a highly relevant avian pathogen, to establishing long-term, productive, persistent infections in both avian and human cell lines. Data presented here provide novel and direct evidence about the crucial role of type I IFNs on the fate of IBDV-infected cells and their contribution to controlling the establishment of IBDV persistent infections. The use of cell lines unable to respond to type I IFNs opens a promising venue to unveiling additional factors contributing to IBDV persistency.

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