Biology subjects

Maroti, G.

Publications and source records attributed to Maroti, G..

At least 19 recordsLinked to original sources

bioRxiv2026

Most metagenomic vanT calls are not vancomycin resistance genes

Metagenomic resistome surveys frequently report vanTG as the dominant glycopeptide resistance gene, but these calls rely on homology without verifying catalytic residues or operon context. By analyzing 107,008 metagenome-assembled genomes, we demonstrate that only 3.7% of predicted vanTG genes possess the required active-site substitution, with the vast majority being misannotated alanine racemases. Incorporating structural and genomic context corrections reveals a novel split-architecture lineage in gut-associated Erysipelotrichales missed by current profiling tools.

microbiology↗

bioRxiv2024

Metagenomic peek into a corn mummy

Numerous studies have shown that metagenomics has opened a dimension in reading the contents of archaeological remains as time capsules. Corn mummies are ritual objects from ancient Egypt, created by forming human-shaped figures from cereal grains grown in a mixture of water and earth. The aim of our study was to determine whether ancient DNA could be preserved in the mummy, and if so, which organisms it might have originated from. To find answers, we performed metagenomic analyses on samples taken from a corn mummy dating to the second half of the third century BC. Alongside a number of clearly modern contaminants, we identified organisms that cannot be excluded as being of historical origin. Besides considerable amounts of bacterial sequences belonging to the genus Bacillus, Mesobacillus, Metabacillus, Neobacillus, Niallia, Peribacillus and Paenibacillus, we also found traces of plants, animals, and humans. Sequences assigned to the genus Triticum showed the highest similarity to ancient T. turgidum ssp. dicoccum specimens from Egypt and the southern Levant. The fragments identified as of Lepidopteran origin showed the greatest similarity to Sphingidae genomes. Analysis of the human-derived sequences revealed L3 (mtDNA), E, and J (Y chromosome) haplotypes, which are common lineages in Africa today.

bioinformatics↗

bioRxiv2023

Investigation of singlet oxygen sensitive genes in the cyanobacterium Synechocystis PCC 6803

Singlet oxygen (1O2) is an important reactive oxygen species whose formation by the type-II, light-dependent, photodynamic reaction is inevitable during photosynthetic processes. In the last decades, the recognition that 1O2 is not only a damaging agent, but can also affect gene expression and participates in signal transduction pathways has received increasing attention. However, contrary to several other taxa, 1O2-specific genes have not been identified in the important cyanobacterial model organism Synechocystis PCC 6803. By using global transcript analysis we have identified a large set of 1O2-responsive Synechocystis genes, whose transcript levels were either enhanced or repressed in the presence of 1O2. Characteristic 1O2 responses were observed in several light-inducible genes of Synechocystis, especially in the hli (or scp) family encoding HLIP/SCP proteins involved in photoprotection. Other important 1O2-induced genes include components of the Photosystem II repair machinery (psbA2 and ftsH2, ftsH3), iron homeostasis genes isiA and idiA, the Group-2 sigma factor sigD, some components of the transcriptomes induced by salt-, hyperosmotic and cold-stress, as well as several genes of unknown function. One of the most pronounced 1O2-induced upregulation was observed for the hliB gene, whose deletion provided tolerance against 1O2-mediated light damage. A bioreporter Synechocystis strain was created by fusing the hliB promoter to the bacterial luciferase (lux), which showed its utility for continuous monitoring of 1O2 concentrations inside the cell.

plant biology↗

bioRxiv2023

Taxonomy of Pseudomonas spp determines interactions with Bacillus subtilis

Bacilli and pseudomonads are among the most well-studied microorganisms commonly found in soil, and frequently co-isolated. Despite this, no systematic approach has been employed to assess the pairwise compatibility of members from these genera. Here, we screened 720 fluorescent soil isolates for their effects on Bacillus subtilis pellicle formation in two types of media and found a predictor for interaction outcome in Pseudomonas taxonomy. Interactions were context-dependent and both medium composition and culture conditions strongly influenced interactions. Negative interactions were associated with Pseudomonas capeferrum, Pseudomonas entomophila and Pseudomonas protegens, and 2,4-diacetylphloroglucinol was confirmed as a strong (but not exclusive) inhibitor of B. subtilis. Non-inhibiting strains were closely related to Pseudomonas trivialis and Pseudomonas lini, but in this case, cocultures with increased B. subtilis pellicle formation were spatially segregated. Our study is the first to propose an overall negative outcome from pairwise interactions between B. subtilis and fluorescent pseudomonads, hence cocultures comprising members from these groups are likely to require additional microorganisms for coexistence.

microbiology↗

bioRxiv2023

Apis mellifera filamentous virus from a honey bee gut microbiome survey in Hungary

In Hungary, as part of a nationwide, climatically balanced survey for a next-generation sequencing-based study of the honey bee (Apis mellifera) gut microbiome, repeated sampling was carried out during the honey production season (March and May 2019). Among other findings, the presence of Apis mellifera filamentous virus (AmFV) was detected in all samples, some at very high levels. AmFV-derived reads were more abundant in the March samples than in the May samples. In March, a higher abundance of AmFV-originated reads was identified in samples collected from warmer areas compared to those collected from cooler areas. A lower proportion of AmFV-derived reads were identified in samples collected in March from the wetter areas than those collected from the drier areas. AmFV-read abundance in samples collected in May showed no significant differences between groups based on either environmental temperature or precipitation. The AmFV abundance correlated negatively with Bartonella apihabitans, Bartonella choladocola, and positively with Frischella perrara, Gilliamella apicola, Gilliamella sp. ESL0443, Lactobacillus apis, Lactobacillus kullabergensis, Lactobacillus sp. IBH004. De novo metagenome assembly of four samples resulted in almost the complete AmFV genome. According to phylogenetic analysis based on DNA polymerase, the Hungarian strains are closest to the strain CH-05 isolated in Switzerland.

bioinformatics↗

bioRxiv2023

Shotgun metagenomic analysis of the skin mucus bacteriome of the common carp (Cyprinus carpio)

The skin mucus bacteriome of fish plays an important role in the health of their hosts. Despite the economic importance of the common carp (Cyprinus carpio), research on its skin bacteriome composition is still missing. To date, most studies on the composition of fish skin bacteriome have used amplicon sequencing, despite the limitations associated with this method. In our study, a shotgun metagenomic approach was applied to characterize the external mucus bacteriome of 8 carp specimens from two different ponds on a fish farm in Hungary. Besides the carp samples, water was also sequenced from the two corresponding ponds. Each carp skin sample was dominated by the phylum Proteobacteria, followed by Actinobacteria, Bacteroidota, Firmicutes, Cyanobacteria and Planctomycetota. Additionally, we have found strong concordance between the water and carp skin mucus samples, despite most studies describing an opposite relationship. Furthermore, shotgun metagenomics allowed us to apply functional annotation to the metagenomes, which revealed several metabolic functions. We present, to our knowledge, the first description of the common carp (Cyprinus carpio) skin mucus bacteriome. Even though our results showed a high level of host genome contamination, we could still provide valuable insight into the external bacterial community of this species. The presented data can provide a basis for future metagenome studies of carp or other fish species.

microbiology↗

bioRxiv2023

Enhanced specificity of Bacillus metataxonomics using a tuf-targeted amplicon sequencing approach

Bacillus species are ubiquitous in nature and have tremendous application potential in agriculture, medicine, and industry. However, the individual species of this genus vary widely in both ecological niches and functional phenotypes, which, hence, requires accurate classification of these bacteria when selecting them for specific purposes. Although analysis of the 16S gene has been widely used to disseminate the taxonomy of most bacterial species, this gene fails proper classification of Bacillus species. To circumvent this restriction, we designed novel primers and optimized them to allow exact species resolution of Bacillus species in both synthetic and natural communities using high-throughput amplicon sequencing. The primers designed for the tuf gene were not only specific for the Bacillus genus but also sufficiently discriminated species both in silico and in vitro in a mixture of 11 distinct Bacillus species. Investigating the primers using a natural soil sample, 13 dominant species were detected including Bacillus badius, Bacillus velezensis, and Bacillus mycoides as primary members, neither of which could be distinguished with 16S sequencing. In conclusion, a set of high-throughput primers were developed which allows unprecedented species-level identification of Bacillus species, including agriculturally important species.

microbiology↗

bioRxiv2023

Enhanced niche colonisation and competition during bacterial adaptation to a fungus

Bacterial-fungal interactions (BFIs) influence microbial community performance of most ecosystems and elicit specific microbial behaviours, including stimulating specialised metabolite production. Using a simple BFI system encompassing the Gram-positive bacterium Bacillus subtilis and the black mould fungus Aspergillus niger, we established a co-culture experimental evolution method to investigate bacterial adaptation to the presence of a fungus. In the evolving populations, B. subtilis was rapidly selected for enhanced production of the lipopeptide surfactin and accelerated surface spreading ability, leading to inhibition of fungal expansion and acidification of the environment. These phenotypes were explained by specific mutations in the DegS-DegU two-component system. In the presence of surfactin, fungal hyphae exhibited bulging cells with delocalised secretory vesicles and RlmA-dependent cell wall stress induction. Increased surfactin production typically enhances the competitive success of bacteria against fungi, which likely explains the primary adaption path in the presence of A. niger. Significance statementExperimental evolution and co-cultivation of different microbes are important and useful techniques for discovering new traits and unravelling cryptic regulatory connections. We combined these methods by evolving the Gram-positive bacterium Bacillus subtilis in the presence of the black mould fungus Aspergillus niger that were previously shown to engage in an intricate and physical interaction. Both are ubiquitous, environmentally and industrially relevant model microbes in the colonisation of rhizo- and endosphere and in the production of enzymes. Our results demonstrate how laboratory adaptation can be exploited to improve biocontrol properties of bacteria.

microbiology↗

bioRxiv2023

Inter-kingdom microbial interactions revealed by a comparative machine-learning guided multi-omics analysis of industrial-scale biogas plants

Multi-omics analysis is a powerful tool for the detection and study of inter-kingdom interactions, such as those between bacterial and archaeal members of complex biogas-producing microbial communities. In the present study, the microbiomes of three industrial-scale biogas digesters, each fed with different substrates, were analysed using a machine-learning guided genome-centric metagenomics framework complemented with metatranscriptome data. This data permitted us to elucidate the relationship between abundant core methanogenic communities and their syntrophic bacterial partners. In total, we detected 297 high-quality, non-redundant metagenome-assembled genomes (nrMAGs). Moreover, the assembled 16S rRNA gene profiles of these nrMAGs showed that the phylum Firmicutes possessed the highest copy number, while the representatives of the Archaeal domain had the lowest. Further investigation of the three anaerobic microbial communities showed characteristic alterations over time but remained specific to each industrial-scale biogas plant. The relative abundance of various microbes as revealed by metagenome data were independent from corresponding metatranscriptome activity data. Interestingly, Archaea showed considerably higher activity than was expected from their abundance. We detected 53 nrMAGs that were present in all three biogas plant microbiomes with different abundances. The core microbiome correlated with the main chemical fermentation parameters and no individual parameter emerged as a predominant shaper of community composition. Various interspecies H2/electron transfer mechanisms were assigned to hydrogenotrophic methanogens in the biogas plants that ran on agricultural biomass and wastewater. Analysis of metatranscriptome data revealed that methanogenesis pathways were the most active of all main metabolic pathways. These findings highlight the importance of a combinatorial omics data framework to identify and characterise the activity of specific microbes in complex environments.

microbiology↗

bioRxiv2022

Dual RNA-Seq profiling unveils mycoparasitic activities of Trichoderma atroviride against haploid Armillaria ostoyae in antagonistic interaction assays

Armillaria ostoyae, a species among the destructive forest pathogens from the genus Armillaria, causes root rot disease on woody plants worldwide. Efficient control measures to limit the growth and impact of this severe underground pathogen are currently under investigation. In a previous study, a new soilborne fungal isolate, Trichoderma atroviride SZMC 24276, exhibited high antagonistic efficacy, which suggested that it could be utilized as a biocontrol agent. The dual culture assay results indicated that the haploid A. ostoyae derivative SZMC 23085 (C18/9) is highly susceptible to the mycelial invasion of T. atroviride SZMC 24276. In the present study we analyzed the transcriptome of A. ostoyae SZMC 23085 (AO) and that of T. atroviride SZMC 24276 (TA) in in vitro dual culture assays to test the molecular arsenal of Trichoderma antagonism and the defense mechanisms of Armillaria. We conducted time-course analysis, functional annotation, analyzed enriched pathways, and differentially expressed genes (DEGs) including biocontrol-related candidate genes from TA and defense-related candidate genes from AO. The results indicated that TA deployed several biocontrol mechanisms when confronted with AO. In response, AO initiated multiple defense mechanisms to protect against the fungal attack. To our knowledge, the present study offers the first transcriptome analysis of a biocontrol fungus attacking A. ostoyae. Overall, this study provides insights that aid the further exploration of plant pathogen - biocontrol agent interaction mechanisms. IMPORTANCEArmillaria species can survive for decades in the soil on dead woody debris, develop rapidly under favourable conditions, and harmfully infect newly planted forests. Our previous study found Trichoderma atroviride to be highly effective in controlling Armillaria growth; therefore, our current work explored the molecular mechanisms that might play a key role in Trichoderma-Armillaria interactions. Direct confrontation assays combined with time course-based dual transcriptome analysis provided a reliable system for uncovering the interactive molecular dynamics between the fungal plant pathogen and its mycoparasitic partner. Furthermore, using a haploid Armillaria isolate allowed us to survey the deadly prey-invading activities of the mycoparasite and the ultimate defensive strategies of its prey. Our current study provides detailed insights into the essential genes and mechanisms involved in Armillaria defense against Trichoderma and the genes potentially involved in the efficiency of Trichoderma to control Armillaria. In addition, using a sensitive haploid Armillaria strain (C18/9), with its complete genome data already available, also offers the opportunity to test possible variable molecular responses of Armillaria ostoyae towards diverse Trichoderma isolates with varying biocontrol abilities. Initial molecular tests of the dual interactions may soon help to develop a targeted biocontrol intervention with mycoparasites against plant pathogens.

microbiology↗

bioRxiv2022

A climatically representative analysis of Hungarian Ixodes ricinus tick bacteriome

BackgroundThe microbial communities of disease vectors may represent a key feature in several biological functions and thus deserve special attention in light of climate change and the consequent need to develop novel control strategies. Nevertheless, vector-borne microbial networks are still poorly understood. Assessing vectors microbial interactions and climatic dependencies may contribute to better-estimating pathogen transmission characteristics, public health risks and the urgency for control steps to be taken. MethodsIn a climatically representative country-wide survey, Ixodes ricinus ticks were collected from 17 locations in Hungary. Using shotgun metagenomic sequencing, the bacteriome composition was analyzed by investigating the relationship between the abundances of nymphs and females in various climatic environments. ResultsBacterial composition on the genus level revealed a significant difference between the samples from females and nymphs. Within the core bacteriome, females and nymphs showed significant variation in the following genera: Arsenophonus, Bacillus, Candidatus Midichloria, Rhodococcus, Sphingomonas, Staphylococcus, Wolbachia. Among females, according to temperature strata, the following were found differentiating: Curtobacterium, Pseudomonas, Sphingomonas. There was no genus with a significant difference in precipitation categories for females. In the nymphs, Curtobacterium showed significant variation between temperature and Bacillus and Curtobacterium for various precipitation levels. Based on the full sample set, Arsenophonus and Wolbachia correlated positively which we assumed to have occurred due to the presence of I. hookeri. ConclusionsThe composition of vector-borne bacteriome members showed significant alterations at sampling points with different climatic conditions and development stages of the tick hosts. Our findings not only pave the way towards understanding tick-borne bacterial networks and interdependencies but also shed light on the high potential for the presence of a possible biological tick control species, the tick parasitoid, Ixodiphagus hookeri based on related bacteriome patterns. The results of conscious tick microbiome assessment studies may contribute to precision tick control strategies of the future.

genomics↗

bioRxiv2021

Adaptation and phenotypic diversification of Bacillus thuringiensis 407 biofilm are accompanied by a fuzzy spreader morphotype

Bacillus cereus group (Bacillus cereus sensu lato) has a diverse ecology, including various species that produce biofilms on abiotic and biotic surfaces. While genetic and morphological diversification enable the adaptation of multicellular communities, this area remains largely unknown in the Bacillus cereus group. In this work, we dissected the experimental evolution of Bacillus thuringiensis 407 Cry-during continuous recolonization of plastic beads. We observed the evolution of a distinct colony morphotype that we named fuzzy spreader (FS) variant. Most multicellular traits of the FS variant displayed higher competitive ability versus the ancestral strain, suggesting an important role for diversification in the adaptation of B. thuringiensis to the biofilm lifestyle. Further genetic characterization of FS variant revealed the disruption of a guanylyltransferase gene by an insertion sequence (IS) element, which could be similarly observed in the genome of a natural isolate. The evolved FS and the deletion mutant in the guanylyltransferase gene (Bt407{Delta}rfbM) displayed similarly altered aggregation and hydrophobicity compared to the ancestor strain, suggesting that adaptation process highly depends on the physical adhesive forces.

microbiology↗

bioRxiv2021

Experimental evolution of Bacillus subtilis on Arabidopsis thaliana roots reveals fast adaptation and improved root colonization in the presence of soil microbes

The soil ubiquitous Bacillus subtilis is known to promote plant growth and protect plants against disease. These characteristics make B. subtilis highly relevant in an agricultural perspective, fueling the interest in studying B. subtilis-plant interactions. Here, we employ an experimental evolution approach to explore adaptation of B. subtilis to Arabidopsis thaliana roots. We found that B. subtilis rapidly adapted to the plant root environment, as evidenced by improved root colonizers observed already after 12 consecutive transfers between seedlings in a hydroponic setup. In addition, two selected evolved isolates from independent populations from transfer 30 outcompeted the ancestor during root colonization. Re-sequencing of single evolved isolates and endpoint populations revealed mutations in genes related to different bacterial traits. Further, phenotypic characterization of evolved isolates from transfer 30 showed that increased root colonization was associated with robust biofilm formation in response to the plant polysaccharide xylan. Additionally, several evolved isolates across independent populations were impaired in motility, a redundant trait in the selective environment. Interestingly, two evolved isolates suffered a fitness disadvantage in a non-selective environment, demonstrating an evolutionary cost of adaptation to the plant root. Finally, increased root colonization by a selected evolved isolate was also demonstrated in the presence of resident soil microbes. Our findings provide novel insights into how a well-known plant growth-promoting rhizobacterium rapidly adapts to an ecologically relevant environment and reveal evolutionary consequences that are fundamental to consider when evolving strains for biocontrol purposes.

microbiology↗

bioRxiv2021

Mobile antimicrobial resistance genes in probiotics

Even though people around the world tend to consume probiotic products for their beneficial health effects on a daily basis, recently, concerns were outlined regarding the uptake and potential intestinal colonisation of the bacteria that they transfer. These bacteria are capable of executing horizontal gene transfer (HGT) which facilitates the movement of various genes, including antimicrobial resistance genes (ARGs), among the donor and recipient bacterial populations. Within our study, 47 shotgun sequencing datasets deriving from various probiotic samples (isolated strains and metagenomes) were bioinformatically analysed. We detected more than 70 ARGs, out of which rpoB mutants conferring resistance to rifampicin, tet(W/N/W) and potentially extended-spectrum beta-lactamase (ESBL) coding TEM-116 were the most common. Numerous ARGs were associated with integrated mobile genetic elements, plasmids or phages promoting the HGT. Our findings raise clinical and public health concerns as the consumption of probiotic products may lead to the transfer of ARGs to human gut bacteria.

genomics↗

bioRxiv2021

Diversification of B. subtilis during experimental evolution on A. thaliana leads to synergism in root colonization of evolved subpopulations

The soil bacterium Bacillus subtilis is known to suppress pathogens as well as promote plant growth. However, in order to fully exploit the potential as natural fertilizer, we need a better understanding of the interactions between B. subtilis and plants. Here, B. subtilis was examined for root colonization through experimental evolution on Arabidopsis thaliana. The populations evolved rapidly, improved in root colonization and diversified into three distinct morphotypes. In order to better understand the adaptation that had taken place, single evolved isolates from the final transfer were randomly selected for further characterization, revealing changes in growth and pellicle formation in medium supplemented with plant polysaccharides. Intriguingly, certain evolved isolates showed improved root colonization only on the plant species they evolved on, but not on another plant species, namely tomato, suggesting A. thaliana specific adaption paths. Finally, the mix performed better than the sum of its constituents in monoculture, which was demonstrated to be caused by complementarity effects. Our results suggest, that genetic diversification occurs in an ecological relevant setting on plant roots and proves to be a stable strategy for root colonization. Significance StatementUnderstanding how plant-growth-promoting rhizobacteria (PGPR) colonize plant roots is crucial to fully utilize their potential for agricultural applications. Here, we employ experimental evolution of the PGPR Bacillus subtilis on Arabidopsis thaliana to study root colonization. We revealed that evolving populations rapidly improve in root colonization and diversify into distinct morphotypes. Notably, improved root colonization by evolved isolates was observed on A. thaliana, not on tomato. Moreover, isolates of distinct morphotypes interacted during root colonization and the mixture of morphotypes showed higher productivity than predicted. These findings suggest that genetic diversification might be a stable strategy to maximize root colonization.

microbiology↗

bioRxiv2020

Adaptation of Bacillus thuringiensis to plant colonisation affects differentiation and toxicity

Bacillus cereus group (Bacillus cereus sensu lato) has a diverse ecology, including various species that are vertebrate or invertebrate pathogens. Few isolates from the B. cereus group have however been demonstrated to benefit plant growth. Therefore, it is crucial to explore how bacterial development and toxicity evolves during plant colonization. Herein, we investigated Bacillus thuringiensis (Cry-) adaptation to the colonization of Arabidopsis thaliana roots, and monitored changes in cellular differentiation in experimentally evolved isolates. Isolates from two populations displayed improved iterative ecesis on roots and increased toxicity against insect larvae. Molecular dissection and recreation of a causative mutation revealed the importance of a non-sense mutation in the rho transcription terminator gene. Transcriptome analysis revealed how Rho impacts various B. thuringiensis genes involved in carbohydrate metabolism and virulence. Our work suggests that evolved multicellular aggregates have a fitness advantage over single cells when colonizing plants, creating a trade-off between swimming and multicellularity in evolved lineages, in addition to unrelated alterations in pathogenicity. ImportanceBiologicals-based plant protection relies on the use of safe microbial strains. During application of biologicals to the rhizosphere, microbes adapt to the niche, including genetic mutations shaping the physiology of the cells. Here, the experimental evolution of Bacillus thuringiensis lacking the insecticide crystal toxins was examined on the plant root to reveal how adaptation shapes the differentiation of this bacterium. Interestingly, evolution of certain linages led to increased emetic toxin production, hemolysis and insect larvae pathogenesis in B. thuringiensis driven by transcriptional rewiring. Further, our detailed study reveals how inactivation of the transcription termination protein Rho promotes aggregation on the plant root in addition to altered differentiation and pathogenesis in B. thuringiensis.

microbiology↗

bioRxiv2020

A glimpse of antimicrobial resistance gene diversity in kefir and yoghurt

Antimicrobial resistance (AMR) is a global threat gaining more and more practical significance every year. The protection of bacteria against antimicrobials based on antimicrobial resistance genes (ARGs) developed in evolution. One of the essential clinical questions is the origin of ARGs of pathogen bacteria. Since the bacteria can share genetic components by horizontal gene transfer (HGT), all even non-pathogen bacteria may provide ARG to any pathogens when they became close physically. The bacteria of the human gut may make contact with bacteria entered into the body by food. The fermented food contains bacteria in high amount by its nature. Here we studied the diversity of ARG content by a unified metagenomic approach in various kefir and yoghurt products, in grain and isolated bacterial strains. We found numerous ARGs of commonly used fermenting bacteria with diversity characteristics in kefir and yoghurt samples. Even with the strictest filter restrictions we identified ARGs undermining the efficacy of aminocoumarin, aminoglycoside, carbapenem, cephalosporin, cephamycin, diaminopyrimidine, elfamycin, fluoroquinolone, fosfomycin, glycylcycline, lincosamides, macrolide, monobactam, nitrofuran, nitroimidazole, penam, penem, peptide, phenicol, rifamycin, tetracycline and triclosan. In the case of gene lmrD, we detected genetic environment providing mobility of this ARG. Our findings support that theory during the fermentation process the food ARG content can grow by the bacteria multiplication. Results presented suggest that starting culture strains of fermented food should be monitored and selected to decrease the ARG amount intake by nutrition.

genomics↗

bioRxiv2020

Impact of Rap-Phr system abundance on adaptation of Bacillus subtilis

Microbes commonly display great genetic plasticity, which has allowed them to colonize all ecological niches on Earth. Bacillus subtilis is a soil-dwelling organism that can be isolated from a wide variety of environments. An interesting characteristic of this bacterium is its ability to form biofilms that display complex heterogeneity: individual, clonal cells develop diverse phenotypes in response to different environmental conditions within the biofilm. Here, we scrutinized the impact that the number and variety of the Rap-Phr family of regulators and cell-cell communication modules of B. subtilis has on genetic adaptation and evolution. We examine how the Rap family of phosphatase regulators impacts sporulation in diverse niches using a library of single and double rap-phr mutants in competition under 4 distinct growth conditions. Using specific DNA barcodes and whole-genome sequencing, population dynamics were followed, revealing the impact of individual Rap phosphatases and arising mutations on the adaptability of B. subtilis.

microbiology↗