Production and Characterization of Motile and Chemotactic Bacterial Minicells.
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ABSTRACT:
SUBMITTER: Ni B
PROVIDER: S-EPMC8218304 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Marine heterotrophic microorganisms remineralize about half of the annual primary production, with the microbiomes on and around algae and particles having a major contribution. These microbiomes specifically include free-living chemotactic and particle-attached bacteria, which are often difficult to analyze individually, as the standard method of size-selective filtration only gives access to particle-attached bacteria. In this study, we demonstrated that particle collection in Imhoff sedimentation cones enriches microbiomes that included free-living chemotactic bacteria and were distinct from particle microbiomes obtained by filtration or centrifugation. Coastal seawater was collected during North Sea phytoplankton spring blooms, and the microbiomes were investigated using 16S rRNA amplicon sequencing and fluorescence microscopy. Enrichment factors of individual operational taxonomic units (OTUs) were calculated for comparison of fractionated communities after separation with unfractionated seawater communities. Filtration resulted in a loss of cells and yielded particle fractions including bacterial aggregates, filaments, and large cells. Centrifugation had the lowest separation capacity. Particles with a sinking rate of >2.4 m day-1 were collected in sedimentation cones as a bottom fraction and enriched in free-living chemotactic bacteria, i.e., Sulfitobacter, Pseudoalteromonas, and Vibrio. Subfractions of these bottom fractions, obtained by centrifugation, showed enrichment of either free-living or particle-attached bacteria. We identified five distinct enrichment patterns across all separation techniques: mechano-sensitive and mechano-stable free-living bacteria and three groups of particle-attached bacteria. Simultaneous enrichment of particle-attached and chemotactic free-living bacteria in Imhoff sedimentation cones is a novel experimental access to these groups providing more insights into the diversity, structure, and function of particle-associated microbiomes, including members of the phycosphere.
| S-EPMC8047139 | biostudies-literature
Project description:Type III protein secretion systems are being considered for vaccine development as virtually any protein antigen can be engineered for delivery by these nanomachines into the class I antigen presentation pathway to stimulate antigen-specific CD8(+) T cells. A limitation in the use of this system is that it requires live virulence-attenuated bacteria, which may preclude its use in certain populations such as children and the immunocompromised. Here we report the engineering of the Salmonella Typhimurium type III secretion system in achromosomal, non-replicating nanoparticles derived from bacterial minicells. The engineered system is shown to be functional and capable of delivering heterologous antigens to the class I antigen presentation pathway stimulating immune responses both in vitro and in vivo. This antigen delivery platform offers a novel approach for vaccine development and cellular immunotherapy.
| S-EPMC3693737 | biostudies-literature
Project description:Bacterial chemotaxis is a canonical system for the study of signal transduction. One of the hallmarks of this system is its robust adaptive behavior. However, how fast the system adapts remains controversial. The adaptation time measured at the level of the kinase activity was tens of seconds, whereas that measured at the level of the flagellar motor was <10 s. The flagellar motor was recently shown to exhibit adaptive remodeling, its main physiological function being to provide a robust match between the chemoreceptor output and the motor input, whereas its adaptation timescale was thought to be too slow to contribute much to the overall adaptation timescale of the chemotaxis system. Here, through theoretical modeling of the motor adaptive remodeling and experimental tests, we show that this motor adaptation contributes significantly to speeding up the overall chemotactic adaptation, thereby resolving the previous inconsistency.
| S-EPMC5883565 | biostudies-literature
Project description:BackgroundWe have harnessed a novel biological system, the bacterial minicell, to deliver cancer therapeutics to cancer cells. Preclinical studies showed that epidermal growth factor receptor (EGFR)-targeted, paclitaxel-loaded minicells (EGFRminicellsPac) have antitumor effects in xenograft models. To examine the safety of the minicell delivery system, we initiated a first-time-in-human, open-label, phase I clinical study of EGFRminicellsPac in patients with advanced solid tumors.MethodologyPatients received 5 weekly infusions followed by a treatment free week. Seven dose levels (1x108, 1x109, 3x109, 1x1010, 1.5x1010, 2x1010, 5x1010) were evaluated using a 3+3 dose-escalation design. Primary objectives were safety, tolerability and determination of the maximum tolerated dose. Secondary objectives were assessment of immune/inflammatory responses and antitumor activity.Principal findingsTwenty eight patients were enrolled, 22 patients completed at least one cycle of EGFRminicellsPac; 6 patients did not complete a cycle due to rapidly progressive disease. A total of 236 doses was delivered over 42 cycles, with a maximum of 45 doses administered to a single patient. Most common treatment-related adverse events were rigors and pyrexia. No deaths resulted from treatment-related adverse events and the maximum tolerated dose was defined as 1x1010 EGFRminicellsPac. Surprisingly, only a mild self-limiting elevation in the inflammatory cytokines IL-6, IL-8 and TNF? and anti-inflammatory IL-10 was observed. Anti-LPS antibody titers peaked by dose 3 and were maintained at that level despite repeat dosing with the bacterially derived minicells. Ten patients (45%; n = 22) achieved stable disease as their best response.Conclusions/significanceThis is the first study in humans of a novel biological system that can provide targeted delivery of a range of chemotherapeutic drugs to solid tumor cells. Bispecific antibody-targeted minicells, packaged with the chemotherapeutic paclitaxel, were shown to be safe in patients with advanced solid tumors with modest clinical efficacy observed. Further study in Phase II trials is planned.Trial registrationAustralian New Zealand Clinical Trials Registry ACTRN12609000672257.
| S-EPMC4699457 | biostudies-literature
Project description:Exploring the complexity of host-pathogen communication is vital to understand why microbes persist within a host, while others are cleared. Here, we employed a Dual-sequencing approach to unravel conversational turn-taking of dynamic host-pathogen communications. We demonstrate that upon hitting a host cell, motile Pseudomonas aeruginosa induce a specific gene expression program. This results in the expression of spermidine on the surface, which specifically activates the PIP3-pathway to induce phagocytic uptake into primary or immortalized murine cells. Non-motile bacteria are more immunogenic due to a lower expression of arnT upon host cell contact, but do not produce spermidine and are phagocytosed less. We demonstrate that not only the presence of pathogen inherent molecular patterns induces immune responses, but that bacterial motility is linked to a host-cell induced expression of additional immune modulators. Our results emphasize on the value of integrating microbiological and immunological findings to unravel complex and dynamic host-pathogen interactions.
2020-09-25 | GSE141757 | GEO
Project description:The quality of sensing and response to external stimuli constitutes a basic element in the selective performance of living organisms. Here we consider the response of Escherichia coli to chemical stimuli. For moderate amplitudes, the bacterial response to generic profiles of sensed chemicals is reconstructed from its response function to an impulse, which then controls the efficiency of bacterial motility. We introduce a method for measuring the impulse response function based on coupling microfluidic experiments and inference methods: The response function is inferred using Bayesian methods from the observed trajectories of bacteria swimming in microfluidically controlled chemical fields. The notable advantages are that the method is based on the bacterial swimming response, it is noninvasive, without any genetic and/or mechanical preparation, and assays the behavior of the whole flagella bundle. We exploit the inference method to measure responses to aspartate and ?-methylaspartate--measured previously by other methods--as well as glucose, leucine, and serine. The response to the attractant glucose is shown to be biphasic and perfectly adapted, as for aspartate. The response to the attractant serine is shown to be biphasic yet imperfectly adapted, that is, the response function has a nonzero (positive) integral. The adaptation of the response to the repellent leucine is also imperfect, with the sign of the two phases inverted with respect to serine. The diversity in the bacterial population of the response function and its dependency upon the background concentration are quantified.
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Project description:Exploring the complexity of host-pathogen communication is vital to understand why microbes persist within a host, while others are cleared. Here, we employed a dual-sequencing approach to unravel conversational turn-taking of dynamic host-pathogen communications. We demonstrate that upon hitting a host cell, motile Pseudomonas aeruginosa induce a specific gene expression program. This results in the expression of spermidine on the surface, which specifically activates the PIP3-pathway to induce phagocytic uptake into primary or immortalized murine cells. Non-motile bacteria are more immunogenic due to a lower expression of arnT upon host-cell contact, but do not produce spermidine and are phagocytosed less. We demonstrate that not only the presence of pathogen inherent molecular patterns induces immune responses, but that bacterial motility is linked to a host-cell-induced expression of additional immune modulators. Our results emphasize on the value of integrating microbiological and immunological findings to unravel complex and dynamic host-pathogen interactions.
| S-EPMC7538158 | biostudies-literature
Project description:Bacterial chemotaxis is one of the most extensively studied adaptive responses in cells. Many bacteria are able to bias their apparently random motion to produce a drift in the direction of the increasing chemoattractant concentration. It has been recognized that the particular motility pattern employed by moving bacteria has a direct impact on the efficiency of chemotaxis. The linear theory of chemotaxis pioneered by de Gennes allows for calculation of the drift velocity in small gradients for bacteria with basic motility patterns. However, recent experimental data on several bacterial species highlighted the motility pattern where the almost straight runs of cells are interspersed with turning events leading to the reorientation of the cell swimming directions with two distinct angles following in strictly alternating order. In this manuscript we generalize the linear theory of chemotaxis to calculate the chemotactic drift speed for the motility pattern of bacteria with two turning angles. By using the experimental data on motility parameters of V. alginolyticus bacteria we can use our theory to relate the efficiency of chemotaxis and the size of bacterial cell body. The results of this work can have a straightforward extension to address most general motility patterns with alternating angles, speeds and durations of runs.
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Project description:While mucosal inflammation is a major source of stress during enteropathogen infection, it remains to be fully elucidated how the host benefits from this environment to clear the pathogen. Here, we show that host stress induced by different stimuli mimicking inflammatory conditions strongly reduces the binding of Shigella flexneri to epithelial cells. Mechanistically, stress activates acid sphingomyelinase leading to host membrane remodeling. Consequently, knockdown or pharmacological inhibition of the acid sphingomyelinase blunts the stress-dependent inhibition of Shigella binding to host cells. Interestingly, stress caused by intracellular Shigella replication also results in remodeling of the host cell membrane, in vitro and in vivo, which precludes re-infection by this and other non-motile pathogens. In contrast, Salmonella Typhimurium overcomes the shortage of permissive entry sites by gathering effectively at the remaining platforms through its flagellar motility. Overall, our findings reveal host membrane remodeling as a novel stress-responsive cell-autonomous defense mechanism that protects epithelial cells from infection by non-motile bacterial pathogens.
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Project description:Bacterial cellulose (BC) is characterized for its high water holding capacity, high crystallinity, an ultrafine fiber network and high tensile strength. This work demonstrates the production of a new interpenetrated polymer network nanocomposite obtained through the incorporation of poly(vinyl alcohol) (PVA) on the BC matrix and evaluates the effect of oven drying on the morphological, mechanical and mass transfer properties of the composite membranes. Both the addition of PVA and oven drying induce the appearance of larger pores (circa 1-3 µm in average diameter) in dried BC/PVA membranes. Both types of treatments also affect the permeability of the composite, as assessed by the diffusion coefficients of polyethylene glycol (PEG) molecules (900, 8,000, 35,000 and 100,000 Da) across the membranes. Finally, the Young's modulus of dry pristine BC decreases following PVA incorporation, resulting in a change from 3.5 GPa to 1 GPa and a five-fold loss in tensile strength.
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