scholarly journals For the Greater (Bacterial) Good: Heterogeneous Expression of Energetically Costly Virulence Factors

2020 ◽  
Vol 88 (7) ◽  
Author(s):  
Kimberly M. Davis
Keyword(s):  
Virulence Factor ◽  
Host Response ◽  
Bacterial Cells ◽  
Bacterial Populations ◽  
Factor Expression ◽  
Heterogeneous Expression ◽  
Host Cell Interactions ◽  
Slow Growing ◽  
Kinetics Of ◽  
Host Tissues

ABSTRACT Bacterial populations are phenotypically heterogeneous, which allows subsets of cells to survive and thrive following changes in environmental conditions. For bacterial pathogens, changes within the host environment occur over the course of the immune response to infection and can result in exposure to host-derived, secreted antimicrobials or force direct interactions with immune cells. Many recent studies have shown host cell interactions promote virulence factor expression, forcing subsets of bacterial cells to battle the host response, while other bacteria reap the benefits of this pacification. It still remains unclear whether virulence factor expression is truly energetically costly within host tissues and whether expression is sufficient to impact the growth kinetics of virulence factor-expressing cells. However, it is clear that slow-growing subsets of bacteria emerge during infection and that these subsets are particularly difficult to eliminate with antibiotics. This minireview will focus on our current understanding of heterogenous virulence factor expression and discuss the evidence that supports or refutes the hypothesis that virulence factor expression is linked to slowed growth and antibiotic tolerance.

scholarly journals Modifying TIMER to generate a slow-folding DsRed derivative for optimal use in quickly-dividing bacteria

2021 ◽  
Vol 17 (7) ◽  
pp. e1009284
Author(s):  
Pavan Patel ◽  
Brendan J. O’Hara ◽  
Emily Aunins ◽  
Kimberly M. Davis
Keyword(s):  
Stationary Phase ◽  
Yersinia Pseudotuberculosis ◽  
Fluorescent Protein ◽  
Bacterial Species ◽  
Bacterial Cells ◽  
Deep Tissue ◽  
Bacterial Populations ◽  
Novel Variant ◽  
Slow Growing ◽  
Host Tissues

It is now well appreciated that members of pathogenic bacterial populations exhibit heterogeneity in growth rates and metabolic activity, and it is known this can impact the ability to eliminate all members of the bacterial population during antibiotic treatment. It remains unclear which pathways promote slowed bacterial growth within host tissues, primarily because it has been difficult to identify and isolate slow growing bacteria from host tissues for downstream analyses. To overcome this limitation, we have developed a novel variant of TIMER, a slow-folding fluorescent protein, named DsRed42, to identify subsets of slowly dividing bacteria within host tissues. The original TIMER folds too slowly for fluorescence accumulation in quickly replicating bacterial species (Escherichia coli, Yersinia pseudotuberculosis), however DsRed42 accumulates red fluorescence in late stationary phase cultures of E. coli and Y. pseudotuberculosis. We show DsRed42 signal also accumulates during exposure to sources of nitric oxide (NO), suggesting DsRed42 signal detects growth-arrested bacterial cells. In a mouse model of Y. pseudotuberculosis deep tissue infection, DsRed42 signal was detected, and primarily accumulates in bacteria expressing markers of stationary phase growth. There was no significant overlap between DsRed42 signal and NO-exposed subpopulations of bacteria within host tissues, suggesting NO stress was transient, allowing bacteria to recover from this stress and resume replication. This novel DsRed42 variant represents a tool that will enable additional studies of slow-growing subpopulations of bacteria, specifically within bacterial species that quickly divide.

scholarly journals Modifying TIMER, a slow-folding DsRed derivative, for optimal use in quickly-dividing bacteria

2021 ◽  
Author(s):  
Pavan Patel ◽  
Brendan J. O’Hara ◽  
Emily Aunins ◽  
Kimberly M. Davis
Keyword(s):  
Nitric Oxide ◽  
Cell Division ◽  
Stationary Phase ◽  
Yersinia Pseudotuberculosis ◽  
Fluorescent Protein ◽  
Bacterial Species ◽  
Bacterial Cells ◽  
E Coli ◽  
Slow Growing ◽  
Host Tissues

AbstractIt is now well appreciated that members of pathogenic bacterial populations exhibit heterogeneity in growth rates and metabolic activity, and it is known this can impact the ability to eliminate all members of the bacterial population during antibiotic treatment. It remains unclear which pathways promote slowed bacterial growth within host tissues, primarily because it has been difficult to identify and isolate slow growing bacteria from host tissues for downstream analyses. To overcome this limitation, we have developed a novel variant of TIMER, a slow-folding fluorescent protein, to identify subsets of slowly dividing bacteria within host tissues. The original TIMER folds too slowly for fluorescence accumulation in quickly replicating bacterial species (Escherichia coli, Yersinia pseudotuberculosis), however this TIMER42 variant accumulates signal in late stationary phase cultures of E. coli and Y. pseudotuberculosis. We show TIMER42 signal also accumulates during exposure to sources of nitric oxide (NO), suggesting TIMER42 signal detects growth-arrested bacterial cells. In a mouse model of Y. pseudotuberculosis deep tissue infection, TIMER42 signal is clearly detected, and primarily accumulates in bacteria expressing markers of stationary phase growth. There was not significant overlap between TIMER42 signal and NO-exposed subpopulations of bacteria within host tissues, suggesting NO stress was transient, allowing bacteria to recover from this stress and resume replication. This novel TIMER42 variant represents a new faster folding TIMER that will enable additional studies of slow-growing subpopulations of bacteria, specifically within bacterial species that quickly divide.Author SummaryWe have generated a variant of TIMER that can be used to mark slow-growing subsets of Yersinia pseudotuberculosis, which has a relatively short division time, similar to E. coli. We used a combination of site-directed and random mutagenesis to generate the TIMER42 variant, which has red fluorescent signal accumulation in post-exponential or stationary phase cells. We found that nitric oxide (NO) stress is sufficient to promote TIMER42 signal accumulation in culture, however within host tissues, TIMER42 signal correlates with a stationary phase reporter (dps). These results suggest NO may cause an immediate arrest in bacterial cell division, but during growth in host tissues exposure to NO is transient, allowing bacteria to recover from this stress and resume cell division. Thus instead of indicating a response to host stressors, TIMER42 signal accumulation within host tissues appears to identify slow-growing cells that are experiencing nutrient limitation.

scholarly journals Host Response and Bacterial Virulence Factor Expression in Pseudomonas aeruginosa and Streptococcus pneumoniae Corneal Ulcers

PLoS ONE ◽  
2013 ◽  
Vol 8 (6) ◽  
pp. e64867 ◽  
Author(s):  
Rajapandian SivaGanesa Karthikeyan ◽  
Jeganathan Lakshmi Priya ◽  
Sixto M. Leal ◽  
Jonida Toska ◽  
Arne Rietsch ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Streptococcus Pneumoniae ◽  
Virulence Factor ◽  
Host Response ◽  
Bacterial Virulence ◽  
Corneal Ulcers ◽  
Factor Expression ◽  
Bacterial Virulence Factor

scholarly journals Stochasticity in Colonial Growth Dynamics of Individual Bacterial Cells

2013 ◽  
Vol 79 (7) ◽  
pp. 2294-2301 ◽  
Author(s):  
Konstantinos P. Koutsoumanis ◽  
Alexandra Lianou
Keyword(s):  
Kinetic Parameters ◽  
Single Cells ◽  
Growth Curves ◽  
Growth Dynamics ◽  
Time Lapse ◽  
Initial Population ◽  
Bacterial Cells ◽  
Stochastic Growth ◽  
Bacterial Populations ◽  
Content Type

ABSTRACTConventional bacterial growth studies rely on large bacterial populations without considering the individual cells. Individual cells, however, can exhibit marked behavioral heterogeneity. Here, we present experimental observations on the colonial growth of 220 individual cells ofSalmonella entericaserotype Typhimurium using time-lapse microscopy videos. We found a highly heterogeneous behavior. Some cells did not grow, showing filamentation or lysis before division. Cells that were able to grow and form microcolonies showed highly diverse growth dynamics. The quality of the videos allowed for counting the cells over time and estimating the kinetic parameters lag time (λ) and maximum specific growth rate (μmax) for each microcolony originating from a single cell. To interpret the observations, the variability of the kinetic parameters was characterized using appropriate probability distributions and introduced to a stochastic model that allows for taking into account heterogeneity using Monte Carlo simulation. The model provides stochastic growth curves demonstrating that growth of single cells or small microbial populations is a pool of events each one of which has its own probability to occur. Simulations of the model illustrated how the apparent variability in population growth gradually decreases with increasing initial population size (N0). For bacterial populations withN0of >100 cells, the variability is almost eliminated and the system seems to behave deterministically, even though the underlying law is stochastic. We also used the model to demonstrate the effect of the presence and extent of a nongrowing population fraction on the stochastic growth of bacterial populations.

scholarly journals Characterization and Molecular Epidemiology of a Fungal Infection of Edible Crabs (Cancer pagurus) and Interaction of the Fungus with the Dinoflagellate Parasite Hematodinium

2012 ◽  
Vol 79 (3) ◽  
pp. 783-793 ◽  
Author(s):  
Amanda L. Smith ◽  
Kristina M. Hamilton ◽  
Lucy Hirschle ◽  
Emma C. Wootton ◽  
Claire L. Vogan ◽  
...  
Keyword(s):  
Ribosomal Dna ◽  
Host Response ◽  
Small Subunit ◽  
Fungal Isolate ◽  
Content Type ◽  
Cancer Pagurus ◽  
South Wales ◽  
Disease Condition ◽  
Fungal Sepsis ◽  
Host Tissues

ABSTRACTThis study reports on an emerging fungal disease of the edible crab,Cancer pagurus. Juvenile (prerecruit) crabs were found to be subject to this disease condition during the months of May to September at two intertidal sites in South Wales, United Kingdom. Histopathology revealed that the fungi overwhelm the host response in the tissues, leading to progressive septicemia. The causative agent of this infection was isolated and grown in pure culture and was identified as a member of theOphiocordycepsclade by sequencing of the small subunit of the fungal ribosomal DNA (rDNA). Of the crabs naturally infected with the fungus, 94% had a coinfection with the parasitic dinoflagellateHematodiniumspecies. To determine if there was any interaction between the two disease-causing agents, apparently fungus-free crabs, both with and without naturalHematodiniuminfections, were challenged with the fungal isolate. The presence ofHematodiniumcaused a significant reduction in fungal multiplication in the hemocoel of the crabs in comparison to that inHematodinium-free individuals. Histopathology of coinfected crabs showed a systemic multiplication ofHematodiniumwithin host tissues, leading to a rapid death, whileHematodinium-free crabs experimentally infected with the fungal isolate died due to fungal sepsis (septicemia) with the same characteristic pathology as seen in natural infections.

scholarly journals Prokaryotic Single-Cell RNA Sequencing by In Situ Combinatorial Indexing

2019 ◽  
Author(s):  
Sydney B. Blattman ◽  
Wenyan Jiang ◽  
Panos Oikonomou ◽  
Saeed Tavazoie
Keyword(s):  
Single Cell ◽  
Rna Sequencing ◽  
Low Cost ◽  
Bacterial Cells ◽  
Single Experiment ◽  
Bacterial Populations ◽  
Single Cell Rna Sequencing ◽  
Gene Expression Heterogeneity ◽  
Mrna Polyadenylation

AbstractDespite longstanding appreciation of gene expression heterogeneity in isogenic bacterial populations, affordable and scalable technologies for studying single bacterial cells have been limited. While single-cell RNA sequencing (scRNA-seq) has revolutionized studies of transcriptional heterogeneity in diverse eukaryotic systems, application of scRNA-seq to prokaryotes has been hindered by their extremely low mRNA abundance, lack of mRNA polyadenylation, and thick cell walls. Here, we present Prokaryotic Expression-profiling by Tagging RNA In Situ and sequencing (PETRI-seq), a low-cost, high-throughput, prokaryotic scRNA-seq pipeline that overcomes these technical obstacles. PETRI-seq uses in situ combinatorial indexing to barcode transcripts from tens of thousands of cells in a single experiment. PETRI-seq captures single cell transcriptomes of Gram-negative and Gram-positive bacteria with high purity and low bias, with median capture rates >200 mRNAs/cell for exponentially growing E. coli. These characteristics enable robust discrimination of cell-states corresponding to different phases of growth. When applied to wild-type S. aureus, PETRI-seq revealed a rare sub-population of cells undergoing prophage induction. We anticipate broad utility of PETRI-seq in defining single-cell states and their dynamics in complex microbial communities.

Modulation of Virulence Factor Expression by Pathogen Target Cell Contact

Science ◽  
1996 ◽  
Vol 273 (5279) ◽  
pp. 1231-1233 ◽  
Author(s):  
J. Pettersson ◽  
R. Nordfelth ◽  
E. Dubinina ◽  
T. Bergman ◽  
M. Gustafsson ◽  
...  
Keyword(s):  
Virulence Factor ◽  
Target Cell ◽  
Cell Contact ◽  
Factor Expression

scholarly journals EnterotoxigenicE. colivirulence gene regulation in human infections

2018 ◽  
Vol 115 (38) ◽  
pp. E8968-E8976 ◽  
Author(s):  
Alexander A. Crofts ◽  
Simone M. Giovanetti ◽  
Erica J. Rubin ◽  
Frédéric M. Poly ◽  
Ramiro L. Gutiérrez ◽  
...  
Keyword(s):  
Gene Expression ◽  
Virulence Factor ◽  
Virulence Gene ◽  
Human Infection ◽  
Anaerobic Growth ◽  
Infection Model ◽  
Low Oxygen ◽  
Virulence Gene Expression ◽  
Factor Expression

EnterotoxigenicEscherichia coli(ETEC) is a global diarrheal pathogen that utilizes adhesins and secreted enterotoxins to cause disease in mammalian hosts. Decades of research on virulence factor regulation in ETEC has revealed a variety of environmental factors that influence gene expression, including bile, pH, bicarbonate, osmolarity, and glucose. However, other hallmarks of the intestinal tract, such as low oxygen availability, have not been examined. Further, determining how ETEC integrates these signals in the complex host environment is challenging. To address this, we characterized ETEC’s response to the human host using samples from a controlled human infection model. We found ETEC senses environmental oxygen to globally influence virulence factor expression via the oxygen-sensitive transcriptional regulator fumarate and nitrate reduction (FNR) regulator. In vitro anaerobic growth replicates the in vivo virulence factor expression profile, and deletion offnrin ETEC strain H10407 results in a significant increase in expression of all classical virulence factors, including the colonization factor antigen I (CFA/I) adhesin operon and both heat-stable and heat-labile enterotoxins. These data depict a model of ETEC infection where FNR activity can globally influence virulence gene expression, and therefore proximity to the oxygenated zone bordering intestinal epithelial cells likely influences ETEC virulence gene expression in vivo. Outside of the host, ETEC biofilms are associated with seasonal ETEC epidemics, and we find FNR is a regulator of biofilm production. Together these data suggest FNR-dependent oxygen sensing in ETEC has implications for human infection inside and outside of the host.

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