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ABSTRACT Microorganisms often inhabit environments that are suboptimal for growth and reproduction. To survive when challenged by such conditions, individuals engage in dormancy, where they enter a metabolically inactive state. For this persistence strategy to confer an evolutionary advantage, microorganisms must be able to resuscitate and reproduce when conditions improve. Among bacteria in the phylum Actinomycetota, dormancy can be terminated by resuscitation-promoting factor (Rpf), an exoenzyme that hydrolyzes glycosidic bonds in the peptidoglycan of cell walls. We characterized Rpf fromMicrococcusKBS0714, a bacterium isolated from agricultural soil. The protein exhibited high substrate affinityin vitro, even though resuscitation was maximized in live-cell assays at micromolar concentrations. Site-directed mutations at conserved catalytic sites significantly reduced or eliminated resuscitation, as did the deletion of repeating motifs in a lectin-encoding linker region. We then tested the effects of recombinant Rpf fromMicrococcusKBS0714 on a diverse set of dormant soil bacteria. Patterns of resuscitation mapped onto strain phylogeny, which reflected core features of the cell envelope. Additionally, the direction and magnitude of the Rpf effect were associated with functional traits, in particular, aspects of the moisture niche and biofilm production, which are critical for understanding dormancy and the persistence of microbial populations in soils. These findings expand our understanding of how Rpf may affect seed bank dynamics with implications for the diversity and functioning of microorganisms in terrestrial ecosystems. IMPORTANCEDormancy is a process whereby individuals enter a reversible state of reduced metabolic activity. In fluctuating environments, dormancy protects individuals from unfavorable conditions, enhancing fitness and buffering populations against extinction. However, waking up from dormancy is a critical yet risky decision. Some bacteria resuscitate stochastically, while others rely on environmental cues or signals from neighboring cells to transition back to active growth. Resuscitation-promoting factor (Rpf) is an exoenzyme that cleaves bonds in the peptidoglycan of bacterial cell walls, facilitating dormancy termination and enabling regrowth. Although this family of proteins has been well characterized in model organisms and clinically relevant strains, our study characterizes Rpf from a soil bacterium and examines its effects on resuscitation across a diverse collection of bacteria, linking it to functional traits that may influence dormancy dynamics in both natural and managed ecosystems. 

Abstract Microorganisms often inhabit environments that are suboptimal for growth and reproduction. To survive when challenged by such conditions, individuals may engage in dormancy where they enter a metabolically inactive state. For this persistence strategy to confer an evolutionary advantage, microorganisms must be able to resuscitate and reproduce when conditions improve. Among bacteria in the phylum Actinomycetota, dormancy can be terminated by resuscitation-promoting factor (Rpf), an exoenzyme that hydrolyzes glycosidic bonds in the peptidoglycan of the cell wall. We characterized Rpf fromMicrococcusKBS0714, a bacterium isolated from agricultural soil. Compared to previous studies, the Rpf elicited activity at relatively high concentrations, yet demonstrated high substrate affinity. Site-directed mutations at conserved catalytic sites significantly reduced or abolished resuscitation, as did the deletion of repeating motifs in a lectin-encoding linker region. We then tested the effects of recombinant Rpf fromMicrococcusKBS0714 on a diverse set of dormant soil bacteria. Patterns of resuscitation mapped onto strain phylogeny, which reflected core features of the cell envelope. Additionally, the direction and magnitude of the Rpf effect were associated with functional traits, in particular, aspects of the moisture niche and biofilm production, which are critical for understanding persistence and resuscitation during dormancy. These findings expand our understanding of how Rpf may affect seed-bank dynamics and have implications for the diversity and functioning of soil ecosystems. 

Abstract While microorganisms are recognized for driving belowground processes that influence the productivity and fitness of plant populations, the vast majority of bacteria and fungi in soil belong to a seed bank consisting of dormant individuals. However, plant performance may be affected by microbial dormancy through its effects on the activity, abundance, and diversity of soil microorganisms. To test how microbial seed banks influence plant‐soil interactions, we purified recombinant resuscitation promoting factor (Rpf), a bacterial protein that terminates dormancy. In a factorially designed experiment, we then applied the Rpf to soil containing field mustard (Brassicarapa), an agronomically important plant species. Plant biomass was ~33% lower in the Rpf treatment compared to plants grown with an unmanipulated microbial seed bank. In addition, Rpf reduced soil respiration, decreased bacterial abundance, and increased fungal abundance. These effects of Rpf on plant performance were accompanied by shifts in bacterial community composition, which may have diluted mutualists or resuscitated pathogens. Our findings suggest that changes in microbial seed banks may influence the magnitude and direction of plant‐soil feedbacks in ways that affect above‐ and belowground biodiversity and function. 

Abstract Pioneer trees require high‐light environments for successful seedling establishment. Consequently, seeds of these species often persist in the soil seed bank (SSB) for periods ranging from several weeks to decades. How they survive despite extensive pressure from seed predators and soil‐borne pathogens remains an intriguing question.This study aims to test the hypotheses that decades‐old seeds collected from the SSB in a lowland tropical forest remain viable by (i) escaping infection by fungi, which are major drivers of seed mortality in tropical soils, and/or (ii) maintaining high levels of seed dormancy and seed coat integrity when compared to inviable seeds.We collected seeds ofTrema micranthaandZanthoxylum ekmaniiat Barro Colorado Island, Panama, from sites where adult trees previously occurred in the past 30 years. We used carbon dating to measure seed age and characterized seed coat integrity, seed dormancy and fungal communities.Viable seeds from the SSB ranged in age from 9 to 30 years forT. micrantha, and 5 to 33 years forZ. ekmanii. We found no evidence that decades‐old seeds maintain high levels of seed dormancy or seed coat integrity. Fungi were rarely detected in fresh seeds (no soil contact), but phylogenetically diverse fungi were detected often in seeds from the SSB. Although fungal infections were more commonly detected in inviable seeds than in viable seeds, a lack of differences in fungal diversity and community composition between viable and inviable seeds suggested that viable seeds are not simply excluding fungal species to survive long periods in the SSB.Synthesis.Our findings reveal the importance of a previously understudied aspect of seed survival, where the impact of seed–microbial interactions may be critical to understand long‐term persistence in the SSB. Read the freePlain Language Summaryfor this article on the Journal blog.