Extracellular vesicles as indicators of environmental stress response in Lactiplantibacillus plantarum: a multi-platform study

Razim, A.; Laimer-Digruber, A.; Schmid, A. M.; Edelbacher, T. V.; Paschal, M. E.; Weinmayer, T.; Thaler, M.; Skalska, M. E.; Migdal, P.; Morandi, M. I.; Daniel, C.; Srutkova, D.; Schwarzer, M.; Hrdy, J.; Ehling-Schulz, M.; Gorska, S.; Inic-Kanada, A.; Wiedermann, U.; Schabussova, I.

doi:10.1101/2025.11.21.689665

bioRxiv · 10.1101/2025.11.21.689665

Extracellular vesicles as indicators of environmental stress response in Lactiplantibacillus plantarum: a multi-platform study

Razim, A.·Laimer-Digruber, A.·Schmid, A. M.·Edelbacher, T. V.·Paschal, M. E.·Weinmayer, T.·Thaler, M.·Skalska, M. E.·Migdal, P.·Morandi, M. I.·Daniel, C.·Srutkova, D.·Schwarzer, M.·Hrdy, J.·Ehling-Schulz, M.·Gorska, S.·Inic-Kanada, A.·Wiedermann, U.·Schabussova, I.

View original source ↗Export metadata ↓

Abstract

Extracellular vesicles (EVs) are key mediators of bacterial communication and adaptation to environmental stress. Their size, cargo, and surface charge are influenced by several factors, including environmental conditions, bacterial physiology, and isolation methods, and are highly strain-specific. Herein, we investigated how exposure to bile, a physiological component of the gut environment, affects the production and properties of EVs released by the probiotic strain Lactiplantibacillus plantarum NCIMB 8826. Through ultracentrifugation followed by size-exclusion chromatography (SEC), we isolated highly purified L. plantarum EVs (LpEVs) and characterized them according to the Minimal Information for Studies of Extracellular Vesicles guidelines. SEC purification significantly reduced the contents of contaminating proteins and peptidoglycans, improving the compositional purity of the isolated EVs. Compared with the parent bacteria, purified LpEVs exhibited distinct surface lipid profiles and zeta potential as well as remarkable stability across varying pH levels, elevated NaCl concentrations, and increasing detergent challenges. Under bile stress, the bacteria released larger LpEVs enriched in bile metabolism-related proteins, suggesting vesicle-mediated adaptation. Fourier-transform infrared spectroscopy further revealed bile-induced molecular alterations in LpEVs that differed from those in the parent bacteria. These findings highlight bacterial EVs as dynamic environmental communicators that respond to stress and may modulate host-microbe interactions before detectable changes occur in the bacterial cells.

Explore related subjects

microbiology Preprint

Keep this discovery

Explore connections, maps & timelines

BibTeX RIS

Razim, A., Laimer-Digruber, A., Schmid, A. M., Edelbacher, T. V., Paschal, M. E., Weinmayer, T., Thaler, M., Skalska, M. E., Migdal, P., Morandi, M. I., Daniel, C., Srutkova, D., Schwarzer, M., Hrdy, J., Ehling-Schulz, M., Gorska, S., Inic-Kanada, A., Wiedermann, U., Schabussova, I.. 2025-11-21. Extracellular vesicles as indicators of environmental stress response in Lactiplantibacillus plantarum: a multi-platform study. https://doi.org/10.1101/2025.11.21.689665

Cite the original work for its findings. Save a collection to share your selection of sources.

KEEP EXPLORING

Related preprints

bioRxiv2026

Cryo-EM Structure of Mycobacteriophage D29 Reveals a Specialized Portal-Capsid Junction and a Unique 13-Subunit Baseplate Collar

Mycobacteriophages are promising agents against drug-resistant mycobacterial infections, yet their structural organization and host-recognition mechanisms remain poorly understood. Here, we determined cryo-EM structures of the capsid, connector, tail tube, and baseplate of the lytic, broad-host-range mycobacteriophage D29, revealing a near-complete virion architecture. At the capsid-portal interface, five copies of the previously uncharacterized protein gp19.1 bridge adjacent capsid subunits around the dodecameric portal, potentially reinforcing the symmetry-mismatched vertex. At the baseplate, 13 copies of gp6 assemble into a collar surrounding the sixfold-symmetric gp23-gp27 core. Remarkably, this 6:13 symmetry mismatch is accommodated through six distinct gp6- gp27 interfaces involving 12 gp6 subunits, leaving one subunit largely disengaged from the core. This asymmetric organization produces a tilted, outward-flaring collar that presents an extensive surface with the potential to engage the host cell. Whole-cell binding and plaque-competition assays further implicate the gp6 collar and the flexible trimeric central fiber gp31 in host recognition. Together, these findings reveal distinct structural adaptations at symmetry-mismatched interfaces and provide a structural framework for understanding mycobacteriophage assembly and host engagement.

microbiology↗

bioRxiv2026

A Luciferase Reporter of Ester Prodrug Delivery to the Cytosol of Intracellular Mycobacteria

The bacterial cell envelope restricts access to cytoplasmic targets and is a principal reason that otherwise potent antibacterial compounds fail in cells. Masking an anionic carboxylic acid as an ester is a common way to cross that barrier, but the strategy succeeds only if the target organism also hydrolyzes the ester once the compound arrives. These two requirements, permeation and intracellular unmasking, are rarely measured together in live bacterial cells, and existing reporters cannot specify where unmasking occurred. Here we describe a bioluminescent platform that resolves both steps in real time in live mycobacteria using carboxylate esters of D-luciferin. Luciferase requires that an ester both reach the bacterial cytoplasm and be hydrolyzed there, which makes light a compartment-specific readout of productive delivery. Across a series of alkyl esters in Mycobacterium smegmatis, chain length governed permeation while branching adjacent to the carbonyl governed esterase processing, indicating that the two parameters can be tuned independently. The n-butyl ester gave the strongest signal in broth, in infected macrophages, and in the pathogen Mycobacterium avium, whereas the tert-butyl ester was inert in Mycobacterium avium and unmasking of the whole series was attenuated at phagosomal pH. Ester activation is therefore both species- and compartment-dependent, and masking groups are best selected in the organism and environment in which they must act.

microbiology↗

bioRxiv2026

Flexible phosphorus allocation to the phosphoproteome underpins diatom regulatory adaptations to fluctuating phosphate supply

Diatoms are major drivers of marine phosphorus (P) cycling and are highly adapted to cope with variable P supply. P-starved diatoms can sense increases in P availability within seconds, accumulating this limiting resource through luxury polyphosphate (polyP) storage. Yet, how diatoms coordinate such responses is unknown. Protein phosphorylation is a core mechanism for environmental signal transduction in eukaryotes, but how cells balance investment in protein phosphorylation against competing physiological demands during P limitation is poorly understood. Here, we characterise system-wide remodelling of the Phaeodactylum tricornutum phosphoproteome in response to P deficiency and subsequent resupply. We identify widespread proteome dephosphorylation across 44% of phosphosites, revealing a previously unrecognised P-sparing strategy in marine phytoplankton. Following P resupply, diatoms orchestrate extensive phosphoproteome reprogramming within minutes. We characterise previously unrecognized components for early recovery from P limitation, including a novel environmentally prevalent intracellular H+/phosphate transporter strongly conserved in diverse diatoms. We also identify a vacuolar Ca2+-dependent protein kinase (CDPK) associated with polyP-storage granules. Mutant and inhibitor analyses demonstrate a role for Ca2+- and CDPK-signalling in the regulation of luxury polyP accumulation. Our findings reveal that recovery from P starvation is mediated by rapid phosphoregulatory control of intracellular P trafficking, organellar ATP generation coupled with luxury P storage, enabling diatoms to exploit newly available P and thrive in fluctuating P regimes. Collectively, our study uncovers the phosphoproteome as a flexible intracellular P reservoir and establishes mechanistic links between intracellular P budgeting, signalling and the marine P cycle.

microbiology↗