Escherichia coli biofilms formed under low-shear modeled microgravity in a ground-based system.
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ABSTRACT:
SUBMITTER: Lynch SV
PROVIDER: S-EPMC1694224 | biostudies-literature | 2006 Dec
REPOSITORIES: biostudies-literature
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Escherichia coli biofilms formed under low-shear modeled microgravity in a ground-based system.
Lynch S V SV Mukundakrishnan K K Benoit M R MR Ayyaswamy P S PS Matin A A
Applied and environmental microbiology 20061006 12
Bacterial biofilms cause chronic diseases that are difficult to control. Since biofilm formation in space is well documented and planktonic cells become more resistant and virulent under modeled microgravity, it is important to determine the effect of this gravity condition on biofilms. Inclusion of glass microcarrier beads of appropriate dimensions and density with medium and inoculum, in vessels specially designed to permit ground-based investigations into aspects of low-shear modeled microgra ...[more]
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Project description:BackgroundExtra-cellular shear force is an important environmental parameter that is significant both medically and in the space environment. Escherichia coli cells grown in a low-shear modeled microgravity (LSMMG) environment produced in a high aspect rotating vessel (HARV) were subjected to transcriptional and physiological analysis.ResultsAerobic LSMMG cultures were grown in rich (LB) and minimal (MOPS + glucose) medium with a normal gravity vector HARV control. Reproducible changes in transcription were seen, but no specific LSMMG responsive genes were identified. Instead, absence of shear and a randomized gravity vector appears to cause local extra-cellular environmental changes, which elicit reproducible cellular responses. In minimal media, the majority of the significantly up- or down-regulated genes of known function were associated with the cell envelope. In rich medium, most LSMMG down-regulated genes were involved in translation. No observable changes in post-culture stress responses and antibiotic sensitivity were seen in cells immediately after exposure to LSMMG. Comparison with earlier studies of Salmonella enterica serovar Typhimurium conducted under similar growth conditions, revealed essentially no similarity in the genes that were significantly up- or down-regulated.ConclusionComparison of these results to previous studies suggests that different organisms may dramatically differ in their responses to medically significant low-shear and space environments. Depending on their specific response, some organisms, such as Salmonella, may become preadapted in a manner that predisposes them to increased virulence.
| S-EPMC1852313 | biostudies-literature
Project description:The long-term response of microbial communities to the microgravity environment of space is not yet fully understood. Of special interest is the possibility that members of these communities may acquire antibiotic resistance. In this study, Escherichia coli cells were grown under low-shear modeled microgravity (LSMMG) conditions for over 1,000 generations (1000G) using chloramphenicol treatment between cycles to prevent contamination. The results were compared with data from an earlier control study done under identical conditions using steam sterilization between cycles rather than chloramphenicol. The sensitivity of the final 1000G-adapted strain to a variety of antibiotics was determined using Vitek analysis. In addition to resistance to chloramphenicol, the adapted strain acquired resistance to cefalotin, cefuroxime, cefuroxime axetil, cefoxitin, and tetracycline. In fact, the resistance to chloramphenicol and cefalotin persisted for over 110 generations despite the removal of both LSMMG conditions and trace antibiotic exposure. Genome sequencing of the adapted strain revealed 22 major changes, including 3 transposon-mediated rearrangements (TMRs). Two TMRs disrupted coding genes (involved in bacterial adhesion), while the third resulted in the deletion of an entire segment (14,314?bp) of the genome, which includes 14 genes involved with motility and chemotaxis. These results are in stark contrast with data from our earlier control study in which cells grown under the identical conditions without antibiotic exposure never acquired antibiotic resistance. Overall, LSMMG does not appear to alter the antibiotic stress resistance seen in microbial ecosystems not exposed to microgravity.IMPORTANCE Stress factors experienced during space include microgravity, sleep deprivation, radiation, isolation, and microbial contamination, all of which can promote immune suppression (1, 2). Under these conditions, the risk of infection from opportunistic pathogens increases significantly, particularly during long-term missions (3). If infection occurs, it is important that the infectious agent should not be antibiotic resistant. Minimizing the occurrence of antibiotic resistance is, therefore, highly desirable. To facilitate this, it is important to better understand the long-term response of bacteria to the microgravity environment. This study demonstrated that the use of antibiotics as a preventive measure could be counterproductive and would likely result in persistent resistance to that antibiotic. In addition, unintended resistance to other antimicrobials might also occur as well as permanent genome changes that might have other unanticipated and undesirable consequences.
| S-EPMC6336426 | biostudies-literature
Project description:The goal of this study was to assess whether low shear-modeled microgravity (LSMMG) effects yeast ,genomic expression patterns using the powerful tool of whole genome microarray hybridization. We determined ,changes in the yeast model organism, Saccharomyces cerevisisae, when grown in LSMMG using the rotating High ,Aspect Ratio Vessel (HARV). A significant number of genes were up- or down-regulated by at least two fold in cells ,that were grown for 5 generations or 25 generations in HARVs. We identified genes in cell wall integrity signaling ,pathways containing MAP kinase cascades that may provide clues to novel physiological responses of eukaryotic ,cells to the external stress of a low-shear modeled microgravity environment. A comparison of the microgravity ,response to other environmental stress response (ESR) genes showed that 26% of the genes that respond ,significantly to LSMMG are involved in a general environmental stress response, while 74% of the genes may ,represent a unique transcriptional response to microgravity. In addition, we found changes in genes involved in ,budding, cell polarity establishment, and cell separation that confirm our hypothesis that exposure to LSMMG ,causes changes in gene transcription resulting in a phenotypic response. The results of the study provide interesting ,clues to potential mechanisms involved in the response to, adaptation to, and survival of eukaryotic cells in a ,microgravity environment and our findings may have important health implications for human spaceflight. Experiment Overall Design: Four conditions are compared with three replicates each: yeast grown in low-shear modeled microgravity (HARV bioreactor) for 5 and 25 generations; yeast grown in a horizontal (non-LSMMG) HARV bioreactor for 5 and 25 generations.
2007-07-07 | E-GEOD-4136 | biostudies-arrayexpress
Project description:UnlabelledThe low-shear microgravity environment, modeled by rotating suspension culture bioreactors called high aspect ratio vessels (HARVs), allows investigation in ground-based studies of the effects of microgravity on eukaryotic cells and provides insights into the impact of space flight on cellular physiology. We have previously demonstrated that low-shear modeled microgravity (LSMMG) causes significant phenotypic changes of a select group of Saccharomyces cerevisiae genes associated with the establishment of cell polarity, bipolar budding, and cell separation. However, the mechanisms cells utilize to sense and respond to microgravity and the fundamental gene expression changes that occur are largely unknown. In this study, we examined the global transcriptional response of yeast cells grown under LSMMG conditions using DNA microarray analysis in order to determine if exposure to LSMMG results in changes in gene expression.ResultsLSMMG differentially changed the expression of a significant number of genes (1372) when yeast cells were cultured for either five generations or twenty-five generations in HARVs, as compared to cells grown under identical conditions in normal gravity. We identified genes in cell wall integrity signaling pathways containing MAP kinase cascades that may provide clues to novel physiological responses of eukaryotic cells to the external stress of a low-shear modeled microgravity environment. A comparison of the microgravity response to other environmental stress response (ESR) genes showed that 26% of the genes that respond significantly to LSMMG are involved in a general environmental stress response, while 74% of the genes may represent a unique transcriptional response to microgravity. In addition, we found changes in genes involved in budding, cell polarity establishment, and cell separation that validate our hypothesis that phenotypic changes observed in cells grown in microgravity are reflected in genome-wide changes. This study documents a considerable response to yeast cell growth in low-shear modeled microgravity that is evident, at least in part, by changes in gene expression. Notably, we identified genes that are involved in cell signaling pathways that allow cells to detect environmental changes, to respond within the cell, and to change accordingly, as well as genes of unknown function that may have a unique transcriptional response to microgravity. We also uncovered significant changes in the expression of many genes involved in cell polarization and bud formation that correlate well with the phenotypic effects observed in yeast cells when grown under similar conditions. These results are noteworthy as they have implications for human space flight.
| S-EPMC1774566 | biostudies-literature
Project description:Anticipating the risk for infectious disease during space exploration and habitation is a critical factor to ensure safety, health and performance of the crewmembers. As a ubiquitous environmental organism that is occasionally part of the human flora, Pseudomonas aeruginosa could pose a health hazard for the immuno-compromised astronauts. In order to gain insights in the behavior of P. aeruginosa in spaceflight conditions, two spaceflight-analogue culture systems, i.e. the rotating wall vessel (RWV) and the random position machine (RPM), were used. Microarray analysis of P. aeruginosa PAO1 grown in the low shear modeled microgravity (LSMMG) environment of the RWV compared to the normal gravity control (NG), revealed a regulatory role for AlgU (RpoE). Specifically, P. aeruginosa cultured in LSMMG exhibited increased alginate production and up-regulation of AlgU-controlled transcripts, including those encoding stress-related proteins. This study also shows the involvement of Hfq in the LSMMG response, consistent with its previously identified role in the Salmonella LSMMG- and spaceflight response. Furthermore, cultivation in LSMMG increased heat- and oxidative stress resistance and caused a decrease in the culture oxygen transfer rate. Interestingly, the global transcriptional response of P. aeruginosa grown in the RPM was similar to that in NG. The possible role of differences in fluid mixing between the RWV and RPM is discussed, with the overall collective data favoring the RWV as the optimal model to study the LSMMG-response of suspended cells. This study represents a first step towards the identification of specific virulence mechanisms of P. aeruginosa activated in response to spaceflight-analogue conditions, and could direct future research regarding the risk assessment and prevention of Pseudomonas infections for the crew in flight and the general public.
2010-07-05 | GSE16970 | GEO
Project description:Anticipating the risk for infectious disease during space exploration and habitation is a critical factor to ensure safety, health and performance of the crewmembers. As a ubiquitous environmental organism that is occasionally part of the human flora, Pseudomonas aeruginosa could pose a health hazard for the immuno-compromised astronauts. In order to gain insights in the behavior of P. aeruginosa in spaceflight conditions, two spaceflight-analogue culture systems, i.e. the rotating wall vessel (RWV) and the random position machine (RPM), were used. Microarray analysis of P. aeruginosa PAO1 grown in the low shear modeled microgravity (LSMMG) environment of the RWV compared to the normal gravity control (NG), revealed a regulatory role for AlgU (RpoE). Specifically, P. aeruginosa cultured in LSMMG exhibited increased alginate production and up-regulation of AlgU-controlled transcripts, including those encoding stress-related proteins. This study also shows the involvement of Hfq in the LSMMG response, consistent with its previously identified role in the Salmonella LSMMG- and spaceflight response. Furthermore, cultivation in LSMMG increased heat- and oxidative stress resistance and caused a decrease in the culture oxygen transfer rate. Interestingly, the global transcriptional response of P. aeruginosa grown in the RPM was similar to that in NG. The possible role of differences in fluid mixing between the RWV and RPM is discussed, with the overall collective data favoring the RWV as the optimal model to study the LSMMG-response of suspended cells. This study represents a first step towards the identification of specific virulence mechanisms of P. aeruginosa activated in response to spaceflight-analogue conditions, and could direct future research regarding the risk assessment and prevention of Pseudomonas infections for the crew in flight and the general public. The wild type P. aeruginosa PAO1 strain (ATCC 15692) was used in this study and all cultures were grown in Lennox L Broth Base (LB) (Life Technologies) at 28 °C. An overnight shaking culture (125 r.p.m.) of P. aeruginosa in LB was washed and diluted in 0.85% NaCl solution to an OD600 of 1. This bacterial suspension was used to inoculate fresh LB medium at a final concentration of 10-4 CFU/ml. Synthecon Rotating Wall Vessel bioreactors (RWV) (50 ml or 10 ml) were filled with inoculated medium so that no headspace (i.e. no bubbles) was present. Other than for stress resistance assays, for which 10 ml capacity bioreactors were used, RWV bioreactors with a capacity of 50 ml were adopted for all experiments. Identical bioreactors were mounted in triplicate on (i) a RWV device in vertical position (LSMMG) (Cellon), (ii) a RWV device in horizontal position (NG) and (iii) the center of the inner Random Positioning Machine (RPM) frame (RG) (Fokker Space), and placed in a large humidified (70%-80% relative humidity) culture chamber, to avoid evaporation of culture medium through the gas-permeable membrane at the back of each vessel (Figure 1). A 25 r.p.m. rotation speed was adopted for the RWV cultures, while RPM-cultures were randomly rotated at 10 r.p.m. (60°/s). Bacteria were grown in the three described test conditions for 24 hours. After 24 hours of cultivation, the contents of every bioreactor was gently mixed by pipetting and divided into several aliquots. Ten millilitres of culture from each growth condition was immediately fixed with RNA Protect Reagent (Qiagen), following the manufacturer's instructions, and fixed cell pellets were frozen at -20 °C until RNA extraction. Samples were immediately exposed to different stresses.
2010-07-29 | E-GEOD-16970 | biostudies-arrayexpress
Project description:The low-shear environment of optimized rotation suspension culture allows both eukaryotic and prokaryotic cells to assume physiologically relevant phenotypes that have led to significant advances in fundamental investigations of medical and biological importance. This culture environment has also been used to model microgravity for ground-based studies regarding the impact of space flight on eukaryotic and prokaryotic physiology. We have previously demonstrated that low-shear modeled microgravity (LSMMG) under optimized rotation suspension culture is a novel environmental signal that regulates the virulence, stress resistance, and protein expression levels of Salmonella enterica serovar Typhimurium. However, the mechanisms used by the cells of any species, including Salmonella, to sense and respond to LSMMG and identities of the genes involved are unknown. In this study, we used DNA microarrays to elucidate the global transcriptional response of Salmonella to LSMMG. When compared with identical growth conditions under normal gravity (1 x g), LSMMG differentially regulated the expression of 163 genes distributed throughout the chromosome, representing functionally diverse groups including transcriptional regulators, virulence factors, lipopolysaccharide biosynthetic enzymes, iron-utilization enzymes, and proteins of unknown function. Many of the LSMMG-regulated genes were organized in clusters or operons. The microarray results were further validated by RT-PCR and phenotypic analyses, and they indicate that the ferric uptake regulator is involved in the LSMMG response. The results provide important insight about the Salmonella LSMMG response and could provide clues for the functioning of known Salmonella virulence systems or the identification of uncharacterized bacterial virulence strategies.
| S-EPMC129779 | biostudies-literature
Project description:The goal of this study was to assess whether low shear-modeled microgravity (LSMMG) effects yeast genomic expression patterns using the powerful tool of whole genome microarray hybridization. We determined changes in the yeast model organism, Saccharomyces cerevisisae, when grown in LSMMG using the rotating High Aspect Ratio Vessel (HARV). A significant number of genes were up- or down-regulated by at least two fold in cells that were grown for 5 generations or 25 generations in HARVs. We identified genes in cell wall integrity signaling pathways containing MAP kinase cascades that may provide clues to novel physiological responses of eukaryotic cells to the external stress of a low-shear modeled microgravity environment. A comparison of the microgravity response to other environmental stress response (ESR) genes showed that 26% of the genes that respond significantly to LSMMG are involved in a general environmental stress response, while 74% of the genes may represent a unique transcriptional response to microgravity. In addition, we found changes in genes involved in budding, cell polarity establishment, and cell separation that confirm our hypothesis that exposure to LSMMG causes changes in gene transcription resulting in a phenotypic response. The results of the study provide interesting clues to potential mechanisms involved in the response to, adaptation to, and survival of eukaryotic cells in a microgravity environment and our findings may have important health implications for human spaceflight. Keywords: time course, stress response, budding, microgravity
2006-03-01 | GSE4136 | GEO
Project description:Waterborne pathogenic mycobacteria can form biofilms, and certain species can cause hard-to-treat human lung infections. Astronaut health could therefore be compromised if the spacecraft environment or water becomes contaminated with pathogenic mycobacteria. This work uses Mycobacterium marinum to determine the physiological changes in a pathogenic mycobacteria grown under low-shear modeled microgravity (LSMMG). M. marinum were grown in high aspect ratio vessels (HARVs) using a rotary cell culture system subjected to LSMMG or the control orientation (normal gravity, NG) and the cultures used to determine bacterial growth, bacterium size, transcriptome changes, and resistance to stress. Two exposure times to LSMMG and NG were examined: bacteria were grown for ~40 h (short), or 4 days followed by re-dilution and growth for ~35 h (long). M. marinum exposed to LSMMG transitioned from exponential phase earlier than the NG culture. They were more sensitive to hydrogen peroxide but showed no change in resistance to gamma radiation or pH 3.5. RNA-Seq detected significantly altered transcript levels for 562 and 328 genes under LSMMG after short and long exposure times, respectively. Results suggest that LSMMG induced a reduction in translation, a downregulation of metabolism, an increase in lipid degradation, and increased chaperone and mycobactin expression. Sigma factor H (sigH) was the only sigma factor transcript induced by LSMMG after both short and long exposure times. In summary, transcriptome studies suggest that LSMMG may simulate a nutrient-deprived environment similar to that found within macrophage during infection. SigH is also implicated in the M. marinum LSMMG transcriptome response.
| S-EPMC5515531 | biostudies-literature
Project description:Spaceflight imposes numerous adaptive challenges for terrestrial life. The reduction in gravity, or microgravity, represents a novel environment that can disrupt homeostasis of many physiological processes. Additionally, it is becoming increasingly clear that an organism's microbiome is critical for host health and examining its resiliency in microgravity represents a new frontier for space biology research. In this study, we examine the impact of microgravity on the interactions between the squid Euprymna scolopes and its beneficial symbiont Vibrio fischeri, which form a highly specific binary mutualism. First, animals inoculated with V. fischeri aboard the space shuttle showed effective colonization of the host light organ, the site of the symbiosis, during space flight. Second, RNA-Seq analysis of squid exposed to modeled microgravity conditions exhibited extensive differential gene expression in the presence and absence of the symbiotic partner. Transcriptomic analyses revealed in the absence of the symbiont during modeled microgravity there was an enrichment of genes and pathways associated with the innate immune and oxidative stress response. The results suggest that V. fischeri may help modulate the host stress responses under modeled microgravity. This study provides a window into the adaptive responses that the host animal and its symbiont use during modeled microgravity.
| S-EPMC5385879 | biostudies-literature