Patterns of bacterial motility in microfluidics-confining environments

Understanding the motility behavior of bacteria in confining microenvironments, in which they search for available physical space and move in response to stimuli, is important for environmental, food industry, and biomedical applications. We studied the motility of five bacterial species with various sizes and flagellar architectures (Vibrio natriegens, Magnetococcus marinus, Pseudomonas putida, Vibrio fischeri, and Escherichia coli) in microfluidic environments presenting various levels of confinement and geometrical complexity, in the absence of external flow and concentration gradients. When the confinement is moderate, such as in quasi-open spaces with only one limiting wall, and in wide channels, the motility behavior of bacteria with complex flagellar architectures approximately follows the hydrodynamics-based predictions developed for simple monotrichous bacteria. Specifically, V. natriegens and V. fischeri moved parallel to the wall and P. putida and E. coli presented a stable movement parallel to the wall but with incidental wall escape events, while M. marinus exhibited frequent flipping between wall accumulator and wall escaper regimes. Conversely, in tighter confining environments, the motility is governed by the steric interactions between bacteria and the surrounding walls. In mesoscale regions, where the impacts of hydrodynamics and steric interactions overlap, these mechanisms can either push bacteria in the same directions in linear channels, leading to smooth bacterial movement, or they could be oppositional (e.g., in mesoscale-sized meandered channels), leading to chaotic movement and subsequent bacterial trapping. The study provides a methodological template for the design of microfluidic devices for single-cell genomic screening, bacterial entrapment for diagnostics, or biocomputation.

Machine Learning Algorithms Applied to Identify Microbial Species by Their Motility

(1) Background: Future missions to potentially habitable places in the Solar System require biochemistry-independent methods for detecting potential alien life forms. The technology was not advanced enough for onboard machine analysis of microscopic observations to be performed in past missions, but recent increases in computational power make the use of automated in-situ analyses feasible. (2) Methods: Here, we present a semi-automated experimental setup, capable of distinguishing the movement of abiotic particles due to Brownian motion from the motility behavior of the bacteria Pseudoalteromonas haloplanktis, Planococcus halocryophilus, Bacillus subtilis, and Escherichia coli. Supervised machine learning algorithms were also used to specifically identify these species based on their characteristic motility behavior. (3) Results: While we were able to distinguish microbial motility from the abiotic movements due to Brownian motion with an accuracy exceeding 99%, the accuracy of the automated identification rates for the selected species does not exceed 82%. (4) Conclusions: Motility is an excellent biosignature, which can be used as a tool for upcoming life-detection missions. This study serves as the basis for the further development of a microscopic life recognition system for upcoming missions to Mars or the ocean worlds of the outer Solar System.

Rhamnolipids and their 3-(3-hydroxyalkanoyloxy)alkanoic acid precursors activate Arabidopsis innate immunity through two independent mechanisms

Plant innate immunity is activated upon perception of invasion pattern molecules by plant cell-surface immune receptors. Several bacteria of the genera Pseudomonas and Burkholderia produce rhamnolipids (RLs) from L-rhamnose and (R)-3-hydroxyalkanoate precursors (HAAs). RL and HAA secretion is required to modulate bacterial swarming motility behavior. The bulb-type lectin receptor kinase LIPOOLIGOSACCHARIDE-SPECIFIC REDUCED ELICITATION/S-DOMAIN-1-29 (LORE/SD1-29) mediates medium-chain 3-hydroxy fatty acid (mc-3-OH-FA) sensing in the plant Arabidopsis thaliana. Here, we show that the lipidic secretome from Pseudomonas aeruginosa comprising RLs, HAAs and mc-3-OH-FAs stimulates Arabidopsis immunity. HAAs, like mc-3-O-FAs, are sensed by LORE and induce canonical immune signaling and local resistance to plant pathogenic Pseudomonas infection. By contrast, RLs trigger an atypical immune response and resistance to Pseudomonas infection independent of LORE. Thus, the glycosyl moieties of RLs, albeit abolishing sensing by LORE, do not impair their ability to trigger plant defense. In addition, our results show that RL-triggered immune response is affected by the sphingolipid composition of the plasma membrane. In conclusion, RLs and their precursors released by bacteria can both be perceived by plants but through distinct mechanisms.

A multiscale 3D chemotaxis assay reveals bacterial navigation mechanisms

How bacteria navigate environmental chemical gradients has implications ranging from health to climate science. The underlying behavioral mechanisms are unknown for most species due to a lack in techniques bridging scales from individual 3D motility behavior to the statistical power required to assess the resulting performance. We present the first demonstration of such a multiscale 3D chemotaxis assay and reveal that Caulobacter crescentus chemotaxis breaks with the Escherichia coli paradigm.

Antigen and G-Protein Coupled Receptor signaling differentially control CD8 T cell motility immediately before and after virus clearance in a primary infection

In mice, experimental influenza virus infection stimulates CD8 T cell infiltration of the airways. Virus is cleared by day 9, and between days 8 and 9 there is an abrupt change in CD8 T cell motility behavior transitioning from low velocity and high confinement on day 8, to high velocity with continued confinement on day 9. We hypothesized that it is loss of virus and/or antigen signals in the context of high chemokine levels that drives the T cells into a rapid surveillance mode. Virus infection induces chemokine production, which may change when the virus is cleared. We therefore sought to examine this period of rapid changes to the T cell environment in the tissue and seek evidence on the roles of peptide-MHC and chemokine receptor interactions. Experiments were performed to block G protein coupled receptor (GPCR) signaling with Pertussis toxin (Ptx). Ptx treatment generally reduced cell velocities and mildly increased confinement, except on day 8 when velocity increased and confinement was relieved, suggesting chemokine mediated arrest. Blocking specific peptide-MHC with monoclonal antibody unexpectedly decreased velocities on days 7 through 9, suggesting TCR/peptide-MHC interactions promote cell mobility in the tissue. Together, these results suggest the T cells are engaged with antigen bearing and chemokine producing cells that affect motility in ways that vary with the day after infection. The increase in velocities on day 9 were reversed by addition of specific peptide, consistent with the idea that antigen signals become limiting on day 9 compared to earlier time points. Thus, antigen and chemokine signals act to alternately promote and restrict CD8 T cell motility until the point of virus clearance, suggesting the switch in motility behavior on day 9 may be due to a combination of limiting antigen in the presence of high chemokine signals as the virus is cleared.