Biology subjects

Khare, A.

Publications and source records attributed to Khare, A..

8 recordsLinked to original sources

bioRxiv2025

Stenotrophomonas maltophilia exhibits defensive multicellularity in response to a Pseudomonas aeruginosa quorum sensing molecule

Microorganisms commonly exist in polymicrobial communities, where they can respond to interspecies secreted molecules by altering behaviors and physiology, however, the underlying mechanisms remain underexplored. Here we investigated interactions between Stenotrophomonas maltophilia and Pseudomonas aeruginosa, co-infecting opportunistic pathogens found in pneumonia and chronic lung infections, including in cystic fibrosis. We found that S. maltophilia forms robust protective multicellular aggregates upon exposure to P. aeruginosa secreted factors. Experimental evolution for lack of aggregation selected for fimbrial mutations and we found that fimbriae are required on both interacting S. maltophilia cells for aggregation. Untargeted metabolomics and targeted validations revealed that the quorum sensing molecule Pseudomonas quinolone signal (PQS) directly induced S. maltophilia aggregation, and co-localized with the aggregates. Further, in co-culture with P. aeruginosa, wild-type S. maltophilia formed aggregates, resulting in up to 75-fold increased survival from P. aeruginosa competition compared to fimbrial mutants. Finally, multiple other bacterial species similarly aggregated upon exposure to P. aeruginosa exoproducts, indicating a more general response. Collectively, our work identifies a novel multispecies interaction where a quorum sensing molecule from a co-infecting pathogen is sensed as a danger signal, thereby inducing a protective multicellular response.

microbiology↗

bioRxiv2025

Genetic determinants of gene amplifications alter frequency and evolutionary trajectory of antibiotic resistance in Staphylococcus aureus

Gene amplifications are thought to be common in bacterial populations, providing a rapid reversible mode of adaptation to diverse stresses, including the acquisition of antibiotic resistance. We previously showed that the opportunistic pathogen Staphylococcus aureus evolves resistance to the dual-targeting fluoroquinolone delafloxacin (DLX) that inhibits both the DNA gyrase and DNA topoisomerase IV via gene amplifications of an efflux pump encoding gene sdrM. However, the pathways that control the formation or selection of gene amplifications, and consequently adaptive trajectories, remain understudied, especially in gram-positive bacteria like S. aureus. Here, we show that specific DNA repair and chromosomal separation pathways alter the frequency of formation and selection of gene amplifications in S. aureus. Through a screen of 36 mutants deficient in various DNA processes, we found that while sdrM amplification was still the almost universal path to DLX resistance, other mutations that increased sdrM expression reduced the selection frequency of sdrM amplifications, demonstrating the critical role of sdrM in DLX resistance. We found that similar to other bacteria, the formation and loss of sdrM amplifications required a functional RecA recombinase, but multiple other mutants in pathways required for amplifications in other species still exhibited frequent sdrM amplifications, suggesting that S. aureus may have alternate routes of amplification formation. Finally, mutants in the tyrosine recombinase XerC that is involved in chromosomal separation were deficient for sdrM amplifications, indicating that XerC is a novel modulator of amplification formation, maintenance, or selection. Thus, our work sheds light on genetic factors that alter gene amplification-mediated evolutionary trajectories to antibiotic resistance in S. aureus and can potentially unlock mechanisms by which such evolution of resistance can be inhibited.

microbiology↗

bioRxiv2025

Temperature robustness of the timing network within songbird premotor nucleus HVC

Many neuronal processes are temperature-sensitive. Cooling by 10 {degrees}C typically slows ion channel dynamics by more than a factor of two (Q10 > 2). Nevertheless, behaviors can remain robust despite variations in brain temperature. For instance, cooling the premotor nucleus HVC in zebra finches by 10 {degrees}C slows song production by only a factor of Q10 [~] 1.3. Here we examine the temperature robustness of the synaptic chain network within HVC. Burst spike propagation along such a chain network is postulated to control the tempo of the song. We show that the dynamics of this network are resilient to cooling and that the slowing of burst propagation exhibits a Q10 similar to that observed for the song. We identify two key factors underlying this robustness: the reliance on axonal delays, which are more resistant to temperature changes than ion channels, and enhanced synaptic efficacy at lower temperatures. We propose that these mechanisms represent general principles by which neural circuits maintain functional stability despite temperature fluctuations in the brain. Significance StatementMany animal behaviors remain robust despite temperature fluctuations in the brain. By studying timing circuits in songbirds, we identify key circuit elements that contribute to this resilience, including axonal delays and synaptic integration. Our work highlights how these mechanisms interact to maintain stable neuronal dynamics in response to temperature changes.

neuroscience↗

bioRxiv2024

Mutations in the Staphylococcus aureus Global Regulator CodY Confer Tolerance to an Interspecies Redox-Active Antimicrobial

Bacteria often exist in multispecies communities where interactions among different species can modify individual fitness and behavior. Although many competitive interactions have been characterized, molecular adaptations that can counter this antagonism and preserve or increase fitness remain underexplored. Here, we characterize the adaptation of Staphylococcus aureus to pyocyanin, a redox-active interspecies antimicrobial produced by Pseudomonas aeruginosa, a co-infecting pathogen frequently isolated from wound and chronic lung infections with S. aureus. Using experimental evolution, we identified mutations in a conserved global transcriptional regulator, CodY, that confer tolerance to pyocyanin and thereby enhance survival of S. aureus. The transcriptional response of a pyocyanin tolerant CodY mutant to pyocyanin indicated a two-pronged defensive response compared to the wild type. Firstly, the CodY mutant strongly suppressed metabolism, by downregulating pathways associated with core metabolism, especially translation-associated genes, upon exposure to pyocyanin. Metabolic suppression via ATP depletion was sufficient to provide comparable protection against pyocyanin to the wild-type strain. Secondly, while both the wild-type and CodY mutant strains upregulated oxidative stress response pathways, the CodY mutant overexpressed multiple stress response genes compared to the wild type. We determined that catalase overexpression was critical to pyocyanin tolerance as its absence eliminated tolerance in the CodY mutant and overexpression of catalase was sufficient to impart tolerance to the wild-type strain. Together, these results suggest that both transcriptional responses likely contribute to pyocyanin tolerance in the CodY mutant. Our data thus provide new mechanistic insight into adaptation toward interbacterial antagonism via altered regulation that facilitates multifaceted protective cellular responses.

microbiology↗

bioRxiv2024

Interspecies secreted surfactants induce emergent motility in Pseudomonas aeruginosa

In most natural environments, bacteria live in polymicrobial communities where secreted molecules from neighboring species alter bacterial behaviors including motility, but such interactions are understudied. Pseudomonas aeruginosa is a motile opportunistic pathogen that exists in diverse multispecies environments such as the soil and is frequently found in human wound and respiratory tract co-infections with other bacteria including Staphylococcus aureus. Here we show that P. aeruginosa can co-opt secreted surfactants from other species for flagellar-based surface motility. We found that exogenous surfactants from S. aureus, other bacteria, and interkingdom species enabled P. aeruginosa to switch from swarming to an alternative surface spreading motility on semi-solid surfaces and allowed for the emergence of surface motility on hard agar where P. aeruginosa was otherwise unable to move. This motility was distinct from the response of other motile bacteria in the presence of exogenous surfactants. Mutant analysis indicated that this P. aeruginosa motility was similar to a previously described mucin-based motility, surfing, albeit with divergent regulation. Thus, our study demonstrates that secreted surfactants from the host as well as neighboring bacterial and interkingdom species act as public goods facilitating P. aeruginosa flagella-mediated surfing-like surface motility, thereby allowing it to access different environmental niches.

microbiology↗

bioRxiv2022

Efflux pump gene amplifications bypass necessity of multiple target mutations for resistance against dual-targeting antibiotic

The rise of antimicrobial resistance has motivated the development of antibiotics that have multiple cellular targets, to theoretically reduce the frequency of resistance evolution, but adaptive trajectories and genetic determinants of resistance against such antibiotics are understudied. Here we investigate these in methicillin resistant Staphylococcus aureus (MRSA) using experimental evolution of ten independent populations in the presence of delafloxacin (DLX), a novel fluoroquinolone that targets both DNA gyrase and topoisomerase IV. We show that coding sequence mutations and genomic amplifications of the gene encoding a poorly characterized efflux pump, SdrM, lead to the evolution of high DLX resistance, circumventing the requirement for mutations in the target enzymes. Almost all of our evolved populations had one of two SdrM coding sequence mutations, which led to moderate DLX resistance. Additionally, these populations had 13 distinct genomic amplifications, each containing sdrM and two adjacent genes encoding efflux pumps, which resulted in up to 100-fold higher DLX resistance. While increased sdrM expression provided the selective advantage of the amplification in the DLX evolution, the adjacent efflux pumps hitchhiking in the genomic amplification contributed to cross-resistance against the aminoglycoside streptomycin. Finally, lack of sdrM necessitated mutations in both DNA gyrase and topoisomerase IV to evolve DLX resistance, and the presence of sdrM thus increased the frequency of resistance evolution. Our study highlights that instead of reduced rates of resistance, evolution of resistance to antibiotics with multiple cellular targets can involve alternate high-frequency evolutionary paths such as genomic amplifications of efflux pumps, that may cause unexpected alterations of the fitness landscape, including antibiotic cross-resistance.

microbiology↗

bioRxiv2021

Systematic identification of molecular mediators underlying sensing of Staphylococcus aureus by Pseudomonas aeruginosa

Bacteria typically exist in dynamic, multispecies communities where polymicrobial interactions influence fitness. Elucidating the molecular mechanisms underlying these interactions is critical for understanding and modulating bacterial behavior in natural environments. While bacterial responses to foreign species are frequently characterized at the molecular and phenotypic level, the exogenous molecules that elicit these responses are understudied. Here we outline a systematic strategy based on transcriptomics combined with genetic and biochemical screens of promoter-reporters to identify the molecules from one species that are sensed by another. We utilized this method to study interactions between the pathogens Pseudomonas aeruginosa and Staphylococcus aureus that are frequently found in co-infections. We discovered that P. aeruginosa senses diverse staphylococcal exoproducts including the metallophore staphylopine, intermediate metabolites citrate and acetoin, and multiple molecules that modulate its iron starvation response. Further, we show that staphylopine inhibits biofilm formation and that P. aeruginosa can utilize citrate and acetoin for growth, revealing that these interactions have both antagonistic and beneficial effects. Our screening approach thus identified multiple S. aureus secreted molecules that are sensed by P. aeruginosa and affect its physiology, demonstrating the efficacy of this approach, and yielding new insight into the molecular basis of interactions between these two species.

microbiology↗

bioRxiv2021

A literature-derived knowledge graph augments the interpretation of single cell RNA-seq datasets

Technology to generate single cell RNA-sequencing (scRNA-seq) datasets and tools to annotate them have rapidly advanced in the past several years. Such tools generally rely on existing transcriptomic datasets or curated databases of cell type defining genes, while the application of scalable natural language processing (NLP) methods to enhance analysis workflows has not been adequately explored. Here we deployed an NLP framework to objectively quantify associations between a comprehensive set of over 20,000 human protein-coding genes and over 500 cell type terms across over 26 million biomedical documents. The resultant gene-cell type associations (GCAs) are significantly stronger between a curated set of matched cell type-marker pairs than the complementary set of mismatched pairs (Mann Whitney p < 6.15x10-76, r = 0.24; cohens D = 2.6). Building on this, we developed an augmented annotation algorithm that leverages GCAs to categorize cell clusters identified in scRNA-seq datasets, and we tested its ability to predict the cellular identity of 185 clusters in 13 datasets from human blood, pancreas, lung, liver, kidney, retina, and placenta. With the optimized settings, the true cellular identity matched the top prediction in 66% of tested clusters and was present among the top five predictions for 94% of clusters. Further, contextualization of differential expression analyses with these GCAs highlights poorly characterized markers of established cell types, such as CLIC6 and DNASE1L3 in retinal pigment epithelial cells and endothelial cells, respectively. Taken together, this study illustrates for the first time how the systematic application of a literature derived knowledge graph can expedite and enhance the annotation and interpretation of scRNA-seq data.

genomics↗