Biology subjects

Shuppara, A. M.

Publications and source records attributed to Shuppara, A. M..

3 recordsLinked to original sources

bioRxiv2024

Combining multiple stressors unexpectedly blocks bacterial migration and growth

In nature, organisms experience combinations of stressors. However, laboratory studies typically simplify reality and focus on the effects of an individual stressor. Here, we use a microfluidic approach to simultaneously provide a physical stressor (shear flow) and a chemical stressor (H2O2) to the human pathogen Pseudomonas aeruginosa. By treating cells with levels of flow and H2O2 that commonly co-occur in nature, we discover that previous reports significantly overestimate the H2O2 levels required to block bacterial growth. Specifically, we establish that flow increases H2O2 effectiveness 50-fold, explaining why previous studies lacking flow required much higher concentrations. Using natural H2O2 levels, we identify the core H2O2 regulon, characterize OxyR-mediated dynamic regulation, and dissect the redundant roles of multiple H2O2 scavenging systems. By examining single-cell behavior, we serendipitously discover that the combined effects of H2O2 and flow block pilus-driven surface migration. Thus, our results counter previous studies and reveal that natural levels of H2O2 and flow synergize to restrict bacterial colonization and survival. By studying two stressors at once, our research highlights the limitations of oversimplifying nature and demonstrates that physical and chemical stress can combine to yield unpredictable effects.

microbiology↗

bioRxiv2024

Fluid flow overcomes antimicrobial resistance by boosting delivery

Antimicrobial resistance is an emerging global threat to humanity. As resistance outpaces development, new perspectives are required. For decades, scientists have prioritized chemical optimization, while largely ignoring the physical process of delivery. Here, we used biophysical simulations and microfluidic experiments to explore how fluid flow delivers antimicrobials into communities of the highly resistant pathogen Pseudomonas aeruginosa. We discover that increasing flow overcomes bacterial resistance towards three chemically distinct antimicrobials: hydrogen peroxide, gentamicin, and carbenicillin. Without flow, resistant P. aeruginosa cells generate local zones of depletion by neutralizing all three antimicrobials through degradation or chemical modification. As flow increases, delivery overwhelms neutralization, allowing antimicrobials to regain effectiveness against resistant bacteria. Additionally, we discover that cells on the edge of a community shield internal cells, and cell-cell shielding is abolished in higher flow regimes. Collectively, our quantitative experiments reveal the unexpected result that physical flow and chemical dosage are equally important to antimicrobial effectiveness. Thus, our results should inspire the incorporation of flow into the discovery, development, and implementation of antimicrobials, and could represent a new strategy to combat antimicrobial resistance.

microbiology↗

bioRxiv2022

Fluid flow sensitizes bacterial pathogens to chemical stress

Cells regularly experience fluid flow in natural systems. However, most experimental systems rely on batch cell culture and fail to consider the effect of flow-driven dynamics on cell physiology. Using microfluidics and single-cell imaging, we discover that the interplay of physical shear rate (a measure of fluid flow) and chemical stress trigger a transcriptional response in the human pathogen Pseudomonas aeruginosa. In batch cell culture, cells protect themselves by quickly scavenging the ubiquitous chemical stressor hydrogen peroxide (H2O2) from the media. In microfluidic conditions, we observe that cell scavenging generates spatial gradients of H2O2. High shear rates replenish H2O2, abolish gradients, and generate a stress response. Combining mathematical simulations and biophysical experiments, we find that cells in flow are sensitive to a H2O2 concentration that is 100-1000 times lower than traditionally studied in batch cell culture. Surprisingly, the shear rate and H2O2 concentration required to trigger a transcriptional response closely match their respective values in the human bloodstream. Thus, our results explain a long-standing discrepancy between H2O2 levels in experimental and natural systems. Finally, we demonstrate that the shear rate and H2O2 concentration found in the human bloodstream trigger gene expression in the blood-relevant human pathogen Staphylococcus aureus, suggesting that flow sensitizes bacteria to chemical stress in natural environments.

microbiology↗