Biology subjects

Cronenberg, T.

Publications and source records attributed to Cronenberg, T..

5 recordsLinked to original sources

bioRxiv2022

Collective polarization dynamics in gonococcal colonies

Membrane potential in bacterial systems has been shown to be dynamic and tightly related to survivability at the single cell level. However, little is known about spatio-temporal patterns of membrane potential in bacterial colonies and biofilms. Here, we discovered a switch from uncorrelated to collective dynamics within colonies formed by the human pathogen Neisseria gonorrhoeae. In freshly assembled colonies, polarization is heterogeneous with instances of transient and uncorrelated hyper- or depolarization of individual cells. As colonies reach a critical size, the polarization behaviour switches to collective dynamics: A hyperpolarized shell forms at the centre, travels radially outward, and halts several micrometres from the colony periphery. Once the shell has passed, we detect an influx of potassium correlated with depolarisation. Transient hyperpolarization also demarks the transition from volume to surface growth. By combining simulations and the use of an alternative electron acceptor for the respiratory chain, we provide strong evidence that local oxygen gradients shape the collective polarization dynamics. Finally, we show that within the hyperpolarized shell, tolerance against aminoglycoside antibiotics but not against {beta}-lactam antibiotics is increased, suggesting that depolarization instantaneously protects cells, while the protective effect of growth arrest does not set in immediately. These findings highlight that the polarization pattern can demark the differentiation into distinct subpopulations with different growth rates and antibiotic tolerance. Significance statementAt the level of single bacteria, membrane potential is surprisingly dynamic and transient hyperpolarization has been associated with increased death rate. Yet, little is known about the spatiotemporal dynamics of membrane polarization during biofilm development. Here, we reveal a discrete transition from uncorrelated to collective polarization dynamics within spherical colonies. Suddenly, a shell of hyperpolarized cells forms at the colony centre and hyperpolarization travels radially outward. In the wake of this shell, bacteria depolarize, reduce their growth rate, and become tolerant against antibiotics, indicating the onset of habitat diversity. Single cell live imaging and modelling link hyperpolarization to an oxygen gradient formed within the colonies. We anticipate that dynamical polarization patterns are tightly connected to biofilm differentiation in various bacterial species.

biophysics↗

bioRxiv2021

Spatio-temporal dynamics of growth and death within spherical bacterial colonies

Bacterial growth within colonies and biofilms is heterogeneous. Local reduction of growth rates has been associated with tolerance against various antibiotics. However, spatial gradients of growth rates are poorly characterized in three-dimensional bacterial colonies. Here, we report two spatially resolved methods for measuring growth rates in bacterial colonies. As bacteria grow and divide, they generate a velocity field that is directly related to the growth rates. We derive profiles of growth rates from the velocity field and show that they are consistent with the profiles obtained by single cell counting. Using these methods, we reveal that even small colonies comprising a few thousand cells of the human pathogen Neisseria gonorrhoeae develop a steep gradient of growth rates within two generations. Furthermore, we show that stringent response decelerates growth inhibition at the colony centre. Based on our results, we suggest that aggregation-related growth inhibition can protect gonococci from external stresses even at early biofilm stages.

biophysics↗

bioRxiv2020

Antibiotics modulate attractive interactions in bacterial colonies affecting survivability under combined treatment

Biofilm formation protects bacteria from antibiotics. Very little is known about the response of biofilm-dwelling bacteria to antibiotics at the single cell level. Here, we developed a cell-tracking approach to investigate how antibiotics affect structure and dynamics of colonies formed by the human pathogen Neisseria gonorrhoeae. Antibiotics targeting different cellular functions enlarge the cell volumes and modulate within-colony motility. Focusing on azithromycin and ceftriaxone, we identify changes in type 4 pilus (T4P) mediated cell-to-cell attraction as the molecular mechanism for different effects on motility. By using strongly attractive mutant strains, we reveal that the survivability under ceftriaxone treatment depends on motility. Combining our results, we find that sequential treatment with azithromycin and ceftriaxone is synergistic. Taken together, we demonstrate that antibiotics modulate T4P-mediated attractions and hence cell motility and colony fluidity. Author SummaryAggregation into colonies and biofilms can enhance bacterial survivability under antibiotic treatment. Aggregation requires modulation of attractive forces between neighboring bacteria, yet the link between attraction and survivability is poorly characterized. Here, we quantify these attractive interactions and show that different antibiotics enhance or inhibit them. Live-cell tracking of single cells in spherical colonies enables us to correlate attractive interactions with single cell motility and colony fluidity. Even moderate changes in cell-to-cell attraction caused by antibiotics strongly impact on within-colony motility. Vice versa, we reveal that motility correlates with survivability under antibiotic treatment. In summary, we demonstrate a link between cellular attraction, colony fluidity, and survivability with the potential to optimize the treatment strategy of commonly used drug combinations.

microbiology↗

bioRxiv2018

Motor properties of PilT-independent type 4 pilus retraction in gonococci

Bacterial type 4 pili (T4P) belong to the strongest molecular machines. The gonococcal T4P retraction ATPase PilT supports forces exceeding 100 pN during T4P retraction. Here, we address the question whether gonococcal T4P retract in the absence of PilT. We show that pilT deletion strains indeed retract their T4P but the maximum force is reduced to 5 pN. Similarly, the speed of T4P retraction is lower by orders of magnitude compared to T4P retraction driven by PilT. Deleting the pilT paralogues pilU and pilT2 in the{Delta} pilT background did not inhibit T4P retraction, indicating that the PilT-like proteins do not compensate for PilT. Furthermore, we show that depletion of proton motive force did not inhibit pilT-independent T4P retraction. We conclude that the retraction ATPase is not essential for gonococcal T4P retraction. However, the force generated in the absence of PilT is too low to support important functions of T4P including twitching motility, fluidization of colonies, or induction of host cell response.\n\nIMPORTANCEBacterial type 4 pili (T4P) have been termed the swiss army knive of bacteria because they perform numerous functions including host cell interaction, twitching motility, colony formation, DNA uptake, protein secretion, and surface sensing. The pilus fibre continuously elongates or retracts and these dynamics are functionally important. Curiously, only a subset of T4P systems employs T4P retraction ATPases to power T4P retraction. Here we show that one of the strongest T4P machines, the gonococcal T4P, retracts without a retraction ATPase. Biophysical characterization reveal strongly reduced force and speed compared to retraction with ATPase. We propose that bacteria encode for retraction ATPases when T4P have to generate high force supporting functions like twitching motility, triggering host cell response, or fluidizing colonies.

biophysics↗

bioRxiv2018

Type IV pilin post-translational modifications modulate materials properties of bacterial colonies

Bacterial type 4 pili (T4P) are extracellular polymers that initiate the formation of microcolonies and biofilms. T4P continuously elongate and retract. These pilus dynamics crucially affects the local order, shape, and fluidity of microcolonies. The major pilin subunit of the T4P bears multiple post-translational modifications. By interfering with different steps of the pilin glycosylation and phosphoform modification pathways, we investigated the effect of pilin post-translational modification on the shape and dynamics of microcolonies formed by Neisseria gonorrhoeae. Deleting the phosphotransferase responsible for phosphoethanolamine modification at residue serine 68 (S68) inhibits shape relaxations of microcolonies after pertubation and causes bacteria carrying the phosphoform modification to segregate to the surface of mixed colonies. We relate these mesoscopic phenotypes to increased attractive forces generated by T4P between cells. Moreover, by deleting genes responsible for the pilin glycan structure, we show that the number of saccharides attached at residue serine 63 (S63) affect the ratio between surface tension and viscosity and cause sorting between bacteria carrying different pilin glycoforms. We conclude that different pilin post-translational modifications moderately affect the attractive forces between bacteria but have severe effects on the materials properties of microcolonies.

biophysics↗