Biology subjects

Nijjer, J.

Publications and source records attributed to Nijjer, J..

4 recordsLinked to original sources

bioRxiv2025

Single-cell imaging reveals spontaneous phenotypic sorting and bet-hedging in developing biofilms

How phenotypic heterogeneity shapes biofilm architecture and development remains poorly understood. Motivated by this, we developed imaging tools to track intracellular levels of c-di-GMP, a key second messenger that controls the motile-to-sessile transition, in developing Vibrio cholerae biofilms at single-cell resolution. We show that c-di-GMP levels spontaneously bifurcate into a bimodal distribution that forms a spatially sorted pattern: High-c-di-GMP cells dominate the biofilm core while low-c-di-GMP cells localize to the periphery. Combining single-lineage tracing, mutant analysis, and agent-based modeling, we reveal that this pattern arises from differential viscosity and surface friction mediated by matrix-dependent interactions between cells and their microenvironments. We demonstrate that this heterogeneity and phenotypic sorting enable continuous emergence and shedding of planktonic cells, enhancing fitness in fluctuating environments. Our findings uncover a differential drag mechanism for pattern formation in multicellular systems, and expand the classical picture of biofilm lifecycle by highlighting the functional significance of phenotypic heterogeneity.

biophysics↗

bioRxiv2023

Lipoarabinomannan regulates septation in Mycobacterium smegmatis

The growth and division of mycobacteria, which include several clinically relevant pathogens, deviate significantly from that of canonical bacterial models. Despite their Gram-positive ancestry, mycobacteria synthesize and elongate a diderm envelope asymmetrically from the poles, with the old pole elongating more robustly than the new pole. In addition to being structurally distinct, the molecular components of the mycobacterial envelope are also evolutionarily unique, including the phosphatidylinositol-anchored lipoglycans lipomannan (LM) and lipoarabinomannan (LAM). LM and LAM modulate host immunity during infection, but their role outside of intracellular survival remains poorly understood, despite their widespread conservation among non-pathogenic and opportunistically pathogenic mycobacteria. Previously, Mycobacterium smegmatis and Mycobacterium tuberculosis mutants producing structurally altered LM and LAM were shown to grow slowly under certain conditions and to be more sensitive to antibiotics, suggesting that mycobacterial lipoglycans may support cellular integrity or growth. To test this, we constructed multiple biosynthetic lipoglycan mutants of M. smegmatis and determined the effect of each mutation on cell wall biosynthesis, envelope integrity, and division. We found that mutants deficient in LAM, but not LM, fail to maintain cell wall integrity in a medium-dependent manner, with envelope deformations specifically associated with septa and new poles. Conversely, a mutant producing abnormally large LAM formed multiseptated cells in way distinct from that observed in a septal hydrolase mutant. These results show that LAM plays critical and distinct roles at subcellular locations associated with division in mycobacteria, including maintenance of local cell envelope integrity and septal placement. SignificanceMycobacteria cause many diseases including tuberculosis (TB). Lipoarabinomannan (LAM) is a lipoglycan of mycobacteria and related bacteria, playing important roles as a surface-exposed pathogen-associated molecular pattern during host-pathogen interactions. Its importance is highlighted by the facts that anti-LAM antibody appears to be protective against TB disease progression, and urine LAM serves as a diagnostic marker for active TB. Given the clinical and immunological relevance of the molecule, it was a striking gap in knowledge that we did not know the cellular function of this lipoglycan in mycobacteria. In this study, we demonstrated that LAM regulates septation, a principle potentially generalizable to other lipoglycans widely found in a group of Gram-positive bacteria that lack lipoteichoic acids.

microbiology↗

bioRxiv2021

Biofilm self-patterning: mechanical force landscaping drives a collective reorientation cascade

In growing active matter systems, a large collection of engineered or living autonomous units metabolize free energy and create order at different length scales as they proliferate and migrate collectively. One such example is bacterial biofilms, which are surface-attached aggregates of bacterial cells embedded in an extracellular matrix. However, how bacterial growth coordinates with cell-surface interactions to create distinctive, long-range order in biofilms remains elusive. Here we report a collective cell reorientation cascade in growing Vibrio cholerae biofilms, leading to a differentially ordered, spatiotemporally coupled core-rim structure reminiscent of a blooming aster. Cell verticalization in the core generates differential growth that drives radial alignment of the cells in the rim, while the radially aligned rim in turn generates compressive stresses that expand the verticalized core. Such self-patterning disappears in adhesion-less mutants but can be restored through opto-manipulation of growth. Agent-based simulations and two-phase active nematic modeling reveal the strong interdependence of the driving forces for the differential ordering. Our findings provide insight into the collective cell patterning in bacterial communities and engineering of phenotypes and functions of living active matter.

biophysics↗

bioRxiv2021

Mechanical stress determines morphogenesis and cell ordering in confined bacterial biofilms

Biofilms are aggregates of bacterial cells surrounded by an extracellular matrix. Much progress has been made in studying biofilm growth on solid substrates; however, little is known about the biophysical mechanisms underlying biofilm development in three-dimensional confined environments, in which the biofilm-dwelling cells must push against and even damage the surrounding environment to proliferate. Here, combining single-cell imaging, mutagenesis, and rheological measurement, we reveal the key morphogenesis steps of Vibrio cholerae biofilms embedded in hydrogels as they grow by four orders of magnitude from their initial size. We show that the morphodynamics and cell ordering in embedded biofilms are fundamentally different from those of biofilms on flat surfaces. Treating embedded biofilms as inclusions growing in an elastic medium, we quantitatively show that the stiffness contrast between the biofilm and its environment determines biofilm morphology and internal architecture, selecting between spherical biofilms with no cell ordering and oblate ellipsoidal biofilms with high cell ordering. When embedded in stiff gels, cells self-organize into a bipolar structure that resembles the molecular ordering in nematic liquid crystal droplets. In vitro biomechanical analysis shows that cell ordering arises from stress transmission across the biofilm-environment interface, mediated by specific matrix components. Our imaging technique and theoretical approach are generalizable to other biofilm-forming species, and potentially to biofilms embedded in mucus or host tissues as during infection. Our results open an avenue to understand how confined cell communities grow by means of a compromise between their inherent developmental program and the mechanical constraints imposed by the environment. Significance StatementBiofilms are microbial cities in which bacterial cells reside in a polymeric matrix. They are commonly found inside soft confining environments such as food matrices and host tissues, against which bacteria must push to proliferate. Here, by combining single-cell live imaging and mechanical characterization, we show that the confining environment determines the dynamics of biofilm shape and internal structure. This self-organized evolution of biofilm architecture is caused by force transmission between the environment and the biofilm, mediated by the extracellular matrix secreted by the cells. Our findings lead to new ways to understand how bacterial communities develop under mechanical constraints, and potentially to new strategies for preventing and controlling biofilm growth in three-dimensional environments.

microbiology↗