Gene expression profiling of murine macrophages in response to infection with Cryptococcus neoformans yeast cells at distinct physiological states, including the dormant state known as viable but non-culturable (VBNC)
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PROVIDER: PRJNA1018842 | ENA |
REPOSITORIES: ENA
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Project description:Macrophages play a pivotal role in determining the fate of invading Cryptococcus neoformans cells, and this outcome is highly dependent on how the pathogen modulates phagocyte activation and function. A common consequence of the C. neoformans-host interaction is the establishment of a latent infection, in which the yeast enters a dormant state. In this study, we investigated whether dormant cells could modulate murine bone marrow-derived macrophages (BMDMs) differently from non-dormant cells, thereby providing mechanistic insights into latent infection. To accomplish this, we examined the transcriptional responses of BMDMs upon in vitro infection with distinct physiological states of C. neoformans – exponential phase, stationary phase, or viable but non-culturable (VBNC; equivalent of dormant) cells – over a period of 6 hours. Our findings unveiled a relationship between the physiological state of the yeast and its impact on the transcriptome of BMDMs. Compared to non-dormant cells, particularly those in the logarithm phase, VBNC cells trigger a modest disruption in the host transcriptome, regulating a reduced number of genes, which clustered in far fewer Gene Ontology terms, many of which were unique to this specific condition. Nevertheless, VBNC cells retained the ability to regulate genes that are important for immune response, such as NLRP3 inflammasome-related genes. Our results suggest that VBNC cells retain the low immunostimulatory profile that is critical for the persistence of this fungal pathogen within the host.
2023-11-23 | GSE243519 | GEO
Project description:The phenomenon of viable but non-culturable (VBNC) referred as a dormant state of non-sporulating bacteria enhancing the survival in adverse environments. To our knowledge, only few studies have been performed on whole genomic expression of V. parahaemolyticus in VBNC state compared with cells in exponential and early stationary phases. Since many VBNC state studies found DNA, RNA and protein degradation, we hypothesise that gene regulation of VBNC cells is highly reduced, down-regulation of gene expression is dominant and only metabolic functions crucial for survival are kept on a sustained basis. In the VBNC state we found 509 significantly induced genes and 309 significantly repressed by more than twofold compared with unstressed phases among 4820 investigated genes (adjusted P-value < 0.05). Furthermore, up-regulation was dominant in most of the non-metabolism functional categories, while five metabolism-related functional categories revealed down-regulation in the VBNC state. To our knowledge, this is the first study of comprehensive transcriptomic analyses of three phases of V. parahaemolyticus RIMD2210633. Although the mechanism of VBNC state is not yet clear, massive regulation of gene expression occurs in the VBNC state compared with expression in unstressed phases and thus, VBNC cells are active cells. VBNC state gene expression was detected in total bacterial RNA of V. parahaemolyticus. Three phases (exponential phase, early stationary phase, and VBNC state) were used in 8 biological replicates. Gene expression in exponential phase and early stationary phase was used for normalization, respectively.
2015-01-28 | E-GEOD-65340 | biostudies-arrayexpress
Project description:The phenomenon of viable but non-culturable (VBNC) referred as a dormant state of non-sporulating bacteria enhancing the survival in adverse environments. To our knowledge, only few studies have been performed on whole genomic expression of V. parahaemolyticus in VBNC state compared with cells in exponential and early stationary phases. Since many VBNC state studies found DNA, RNA and protein degradation, we hypothesise that gene regulation of VBNC cells is highly reduced, down-regulation of gene expression is dominant and only metabolic functions crucial for survival are kept on a sustained basis. In the VBNC state we found 509 significantly induced genes and 309 significantly repressed by more than twofold compared with unstressed phases among 4820 investigated genes (adjusted P-value < 0.05). Furthermore, up-regulation was dominant in most of the non-metabolism functional categories, while five metabolism-related functional categories revealed down-regulation in the VBNC state. To our knowledge, this is the first study of comprehensive transcriptomic analyses of three phases of V. parahaemolyticus RIMD2210633. Although the mechanism of VBNC state is not yet clear, massive regulation of gene expression occurs in the VBNC state compared with expression in unstressed phases and thus, VBNC cells are active cells.
2015-01-28 | GSE65340 | GEO
Project description:Metabolically quiescent pathogens can persist in a viable non-replicating state for months or even years. In specific infectious diseases, such as tuberculosis, cryptococcosis, histoplasmosis, latent infection is a corollary of this dormant state, with at some point a phase of reactivation, leading to the symptoms of the infection. During murine cryptococcosis and macrophage uptake, stress and host immunity induce C. neoformans heterogeneity with generation of a sub-population of yeasts that has been shown to harbor a phenotype compatible with dormancy (low stress response, latency of growth). In this subpopulation, mitochondrial transcriptional activity has been shown to be up regulated and this phenotype has been considered as a hallmark of quiescence in stem cells. Based on these findings, we worked to reproduce this phenotype in vitro model and standardized the experimental conditions to generate dormancy in Cryptococcus neoformans. We found that incubation of stationary phase yeasts (STAT) in nutriment limited conditions and hypoxia during 8 days (8D-HYPOx) were able to mimic the phenotype obtained in vivo. In these conditions, mortality and/or apoptosis occurred in less than 5% of the yeasts compared to 30-40% of apoptotic or dead yeasts upon incubation in normoxia (8D-NORMOx). Yeasts in 8D-HYPOx harbored a lower stress response, delayed growth and less that 1% of culturability on agar plates, suggesting that these yeasts are viable but non culturable cells (VBNC). These VBNC were able to reactivate in the presence of pantothenic acid, a vitamin that is known to be involved in quorum sensing and precursor of acetyl-CoA. Global metabolism of 8D-HYPOx cells showed some specific requirements and was globally shut down compared to 8D-NORMOx and STAT conditions. Mitochondrial analyses showed that the mitochondrial masse increased with mitochondria mostly depolarized in 8D-HYPOx compared to 8D-NORMox with an increased expression of mitochondrial genes. Proteomic and transcriptomic analyses of 8D-HYPOx revealed that the number of secreted proteins and transcripts detected also decreased compared to 8D-NORMOx and STAT, but proteome, secretome and transcriptome harbored specific profiles that are engaged as soon as four days of incubation. Importantly, acetyl-CoA and fatty acid pathway involving mitochondria is required for the generation and viability maintenance of VBNC. All together, these data show that we were able to generate for the first time VBNC phenotype in Cryptococcocus neoformans. This VBNC state is associated with a reduced and specific metabolism that should be further studied to understand dormancy/quiescence in this organism.
2019-08-08 | PXD012570 | Pride
Project description:Viable but nonculturable (VBNC) organisms have been underestimated and neglected when studying dormant phenotypes. In clinical settings, VBNC cells may contribute to non-apparent infections capable of being reactivated after months or even years, as for the case of Mycobacterium tuberculosis. The lack of specific and reliable methodology prevents the proper characterization of the VBNC state. Ultimately, these organisms pose a public health risk with potential implications in several industries ranging from pharmaceuticals to food industry. Research regarding their induction and resuscitation is of major importance. Bacteria are able to respond to several environmental and physiological oscillations in part via two-component systems (TCSs). BtsS/BtsR and YpdA/YpdB are two TCSs of Escherichia coli that form a pyruvate sensing network. Their role in the VBNC state is explored in this study.
2018-11-23 | PXD009463 | Pride
Project description:"Viable but non-culturable” (VBNC) states pose challenges for environmental and clinical microbiology, but their biological mechanisms remain obscure. Mycobacterium tuberculosis (Mtb), the leading cause of death from infection until COVID-19, affords a striking example. Mtb can enter into a “differentially detectable” (DD) state associated with phenotypic antimicrobial resistance in which Mtb cells are viable but undetectable as colony-forming units. We found that Mtb cells enter the DD state when they undergo sublethal oxidative stress that damages their DNA, proteins, and lipids, and in addition, their replication is delayed, allowing repair. Mycobacterium bovis and BCG fail to enter the DD state under similar conditions. These findings have implications for TB latency, detection, relapse, treatment monitoring, and development of regimens that overcome phenotypic antimicrobial resistance.
2021-11-24 | GSE180505 | GEO
Project description:“Viable but non-culturable” (VBNC) states pose challenges for environmental and clinical microbiology, but their biological mechanisms remain obscure. Mycobacterium tuberculosis (Mtb), the leading cause of death from infection until COVID-19, affords a striking example. Mtb can enter into a “differentially detectable” (DD) state associated with phenotypic antimicrobial resistance in which Mtb cells are viable but undetectable as colony-forming units. We found that Mtb cells enter the DD state when they undergo sublethal oxidative stress that damages their DNA, proteins, and lipids, and in addition, their replication is delayed, allowing repair. Mycobacterium bovis and BCG fail to enter the DD state under similar conditions. These findings have implications for TB latency, detection, relapse, treatment monitoring, and development of regimens that overcome phenotypic antimicrobial resistance.
2021-11-24 | GSE180792 | GEO
Project description:“Viable but non-culturable” (VBNC) states pose challenges for environmental and clinical microbiology, but their biological mechanisms remain obscure. Mycobacterium tuberculosis (Mtb), the leading cause of death from infection until COVID-19, affords a striking example. Mtb can enter into a “differentially detectable” (DD) state associated with phenotypic antimicrobial resistance in which Mtb cells are viable but undetectable as colony-forming units. We found that Mtb cells enter the DD state when they undergo sublethal oxidative stress that damages their DNA, proteins, and lipids, and in addition, their replication is delayed, allowing repair. Mycobacterium bovis and BCG fail to enter the DD state under similar conditions. These findings have implications for TB latency, detection, relapse, treatment monitoring, and development of regimens that overcome phenotypic antimicrobial resistance.
2021-11-24 | GSE180658 | GEO
Project description:Dormant cells of Mycobacterium tuberculosis, in addition to low metabolic activity and a high level of drug resistance, are characterized by ‘non-culturability’ – a specific reversible state of the inability of the cells to grow on solid media. The biochemical characterization of this physiological state of the pathogen is only superficial, pending clarification of the metabolic processes that may exist in such cells. In this study, applying LC-MS proteomic profiling, we report the analysis of proteins accumulated in dormant, ‘non-culturable’ M. tuberculosis cells in an in vitro model of self-acidification of mycobacteria in the post-stationary phase, simulating the in vivo persistence conditions. This approach revealed the preservation of 1379 proteins in cells after 5 months of storage in dormancy; among them, 468 proteins were statistically different from those in the actively growing cells and bore a positive fold change (FC). Differential analysis revealed the proteins of the pH-dependent regulatory system PhoP and allowed the reconstruction of the reactions of central carbon/glycerol metabolism, as well as revealing the salvaged pathways of mycothiol and UMP biosynthesis, establishing the cohort of survival enzymes of dormancy. The annotated pathways mirror the adaptation of the mycobacterial metabolic machinery to life within lipid-rich macrophages: especially the involvement of the methyl citrate and glyoxylate pathways. Thus, the current in vitro model of M. tuberculosis self-acidification reflects the biochemical adaptation of these bacteria to persistence in vivo. Comparative analysis with published proteins displaying antigenic properties makes it possible to distinguish immunoreactive proteins among the proteins bearing a positive FC in dormancy, which may include specific antigens of latent tuberculosis. Additionally, the biotransformatory enzymes (oxidoreductases and hydrolases) capable of prodrug activation and stored up in the dormant state were annotated. These findings may potentially lead to the discovery of immunodiagnostic tests for early latent tuberculosis and trigger the discovery of efficient drugs/prodrugs with potency against non-replicating, dormant populations of mycobacteria.
2022-07-07 | PXD028849 | Pride
Project description:Characterization of the putative genetic determinants of the VBNC state in a known spore-forming Gram-positive organism Bacillus subtilis 168. The VBNC state was induced under osmotic stress and aminoglycoside treatment. The transcriptome landscape of VBNC cells was compared to the viable, antibiotic sensitive B. subtilis cells and to the viable cells with no antibiotic treatment.
2022-07-02 | GSE207180 | GEO