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1. A modified aeroponic system for growing small-seeded legumes and other plants to study root systems

   https://doi.org/10.1186/s13007-023-01000-6  

   Cai, Jingya ; Veerappan, Vijaykumar ; Arildsen, Kate ; Sullivan, Catrina ; Piechowicz, Megan ; Frugoli, Julia ; Dickstein, Rebecca ( March 2023 , Plant Methods)

   Various growth systems are available for studying plant root growth and plant–microbe interactions including hydroponics and aeroponics. Although some of these systems work well withArabidopsis thalianaand smaller cereal model plants, they may not scale up as well for use with hundreds of plants at a time from a larger plant species. The aim of this study is to present step-by-step instructions for fabricating an aeroponic system, also called a “caisson,” that has been in use in several legume research labs studying the development of symbiotic nitrogen fixing nodules, but for which detailed directions are not currently available. The aeroponic system is reusable and is adaptable for many other types of investigations besides root nodulation.

   An aeroponic system that is affordable and reusable was adapted from a design invented by French engineer René Odorico. It consists of two main components: a modified trash can with a lid of holes and a commercially available industrial humidifier that is waterproofed with silicon sealant. The humidifier generates a mist in which plant roots grow, suspended from holes in trash can lid. Results from use of the aeroponic system have been available in the scientific community for decades; it has a record as a workhorse in the lab.

   Aeroponic systems present a convenient way for researchers to grow plants for studying root systems and plant–microbe interactions in root systems. They are particularly attractive for phenotyping roots and following the progress of nodule development in legumes. Advantages include the ability to precisely control the growth medium in which the plants grow and easy observations of roots during growth. In this system, mechanical shear potentially killing microbes found in some other types of aeroponic devices is not an issue. Disadvantages of aeroponic systems include the likelihood of altered root physiology compared to root growth on soil and other solid substrates and the need to have separate aeroponic systems for comparing plant responses to different microbial strains.

    

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2. Specific Root Exudate Compounds Sensed by Dedicated Chemoreceptors Shape Azospirillum brasilense Chemotaxis in the Rhizosphere

   https://doi.org/10.1128/AEM.01026-20  

   O’Neal, Lindsey ; Vo, Lam ; Alexandre, Gladys ; Parales, Rebecca E. ( July 2020 , Applied and Environmental Microbiology)

   ABSTRACT Plant roots shape the rhizosphere community by secreting compounds that recruit diverse bacteria. Colonization of various plant roots by the motile alphaproteobacterium Azospirillum brasilens e causes increased plant growth, root volume, and crop yield. Bacterial chemotaxis in this and other motile soil bacteria is critical for competitive colonization of the root surfaces. The role of chemotaxis in root surface colonization has previously been established by endpoint analyses of bacterial colonization levels detected a few hours to days after inoculation. More recently, microfluidic devices have been used to study plant-microbe interactions, but these devices are size limited. Here, we use a novel slide-in chamber that allows real-time monitoring of plant-microbe interactions using agriculturally relevant seedlings to characterize how bacterial chemotaxis mediates plant root surface colonization during the association of A. brasilens e with Triticum aestivum (wheat) and Medicago sativa (alfalfa) seedlings. We track A. brasilense accumulation in the rhizosphere and on the root surfaces of wheat and alfalfa. A. brasilense motile cells display distinct chemotaxis behaviors in different regions of the roots, including attractant and repellent responses that ultimately drive surface colonization patterns. We also combine these observations with real-time analyses of behaviors of wild-type and mutant strains to link chemotaxis responses to distinct chemicals identified in root exudates to specific chemoreceptors that together explain the chemotactic response of motile cells in different regions of the roots. Furthermore, the bacterial second messenger c-di-GMP modulates these chemotaxis responses. Together, these findings illustrate dynamic bacterial chemotaxis responses to rhizosphere gradients that guide root surface colonization. IMPORTANCE Plant root exudates play critical roles in shaping rhizosphere microbial communities, and the ability of motile bacteria to respond to these gradients mediates competitive colonization of root surfaces. Root exudates are complex chemical mixtures that are spatially and temporally dynamic. Identifying the exact chemical(s) that mediates the recruitment of soil bacteria to specific regions of the roots is thus challenging. Here, we connect patterns of bacterial chemotaxis responses and sensing by chemoreceptors to chemicals found in root exudate gradients and identify key chemical signals that shape root surface colonization in different plants and regions of the roots. 

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3. Intraspecific variation mediates density dependence in a genetically diverse plant species

   https://doi.org/10.1002/ecy.3502  

   Zaiats, Andrii ; Germino, Matthew J. ; Serpe, Marcelo D. ; Richardson, Bryce A. ; Caughlin, T. Trevor ( September 2021 , Ecology)

   Interactions between neighboring plants are critical for biodiversity maintenance in plant populations and communities. Intraspecific trait variation and genome duplication are common in plant species and can drive eco‐evolutionary dynamics through genotype‐mediated plant–plant interactions. However, few studies have examined how species‐wide intraspecific variation may alter interactions between neighboring plants. We investigate how subspecies and ploidy variation in a genetically diverse species, big sagebrush (Artemisia tridentata), can alter the demographic outcomes of plant interactions. Using a replicated, long‐term common garden experiment that represents range‐wide diversity ofA. tridentata, we ask how intraspecific variation, environment, and stand age mediate neighbor effects on plant growth and survival. Spatially explicit models revealed that ploidy variation and subspecies identity can mediate plant–plant interactions but that the effect size varied in time and across experimental sites. We found that demographic impacts of neighbor effects were strongest during early stages of stand development and in sites with greater growth rates. Within subspecies, tetraploid populations showed greater tolerance to neighbor crowding compared to their diploid variants. Our findings provide evidence that intraspecific variation related to genome size and subspecies identity impacts spatial demography in a genetically diverse plant species. Accounting for intraspecific variation in studies of conspecific density dependence will improve our understanding of how local populations will respond to novel genotypes and biotic interaction regimes. As introduction of novel genotypes into local populations becomes more common, quantifying demographic processes in genetically diverse populations will help predict long‐term consequences of plant–plant interactions.

    

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4. Autopolyploidy alters nodule‐level interactions in the legume – rhizobium mutualism

   https://doi.org/10.1002/ajb2.1375  

   Forrester, Nicole J. ; Ashman, Tia‐Lynn ( November 2019 , American Journal of Botany)

   Polyploidy is a major genetic driver of ecological and evolutionary processes in plants, yet its effects on plant interactions with mutualistic microbes remain unresolved. The legume–rhizobium symbiosis regulates global nutrient cycles and plays a role in the diversification of legume species. In this mutualism, rhizobia bacteria fix nitrogen in exchange for carbon provided by legume hosts. This exchange occurs inside root nodules, which house bacterial cells and represent the interface of legume–rhizobium interactions. Although polyploidy may directly impact the legume–rhizobium mutualism, no studies have explored how it alters the internal structure of nodules.

   We created synthetic autotetraploids usingMedicago sativasubsp.caerulea. Neotetraploid plants and their diploid progenitors were singly inoculated with two strains of rhizobia,Sinorhizobium melilotiandS. medicae. Confocal microscopy was used to quantify internal traits of nodules produced by diploid and neotetraploid plants.

   Autotetraploid plants produced larger nodules with larger nitrogen fixation zones than diploids for both strains of rhizobia, although the significance of these differences was limited by power. NeotetraploidM. sativasubsp.caeruleaplants also produced symbiosomes that were significantly larger, nearly twice the size, than those present in diploids.

   This study sheds light on how polyploidy directly affects a plant–bacterium mutualism and uncovers novel mechanisms. Changes in plant–microbe interactions that directly result from polyploidy likely contribute to the increased ability of polyploid legumes to establish in diverse environments.

    

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5. Interactions among plants, bacteria, and fungi reduce extracellular enzyme activities under long‐term N fertilization

   https://doi.org/10.1111/gcb.14081  

   Carrara, Joseph E. ; Walter, Christopher A. ; Hawkins, Jennifer S. ; Peterjohn, William T. ; Averill, Colin ; Brzostek, Edward R. ( February 2018 , Global Change Biology)

   Atmospheric nitrogen (N) deposition has enhanced soil carbon (C) stocks in temperate forests. Most research has posited that these soil C gains are driven primarily by shifts in fungal community composition with elevated N leading to declines in lignin degradingBasidiomycetes. Recent research, however, suggests that plants and soil microbes are dynamically intertwined, whereby plants send C subsidies to rhizosphere microbes to enhance enzyme production and the mobilization of N. Thus, under elevated N, trees may reduce belowground C allocation leading to cascading impacts on the ability of microbes to degrade soil organic matter through a shift in microbial species and/or a change in plant–microbe interactions. The objective of this study was to determine the extent to which couplings among plant, fungal, and bacterial responses to N fertilization alter the activity of enzymes that are the primary agents of soil decomposition. We measured fungal and bacterial community composition, root–microbial interactions, and extracellular enzyme activity in the rhizosphere, bulk, and organic horizon of soils sampled from a long‐term (>25 years), whole‐watershed, N fertilization experiment at the Fernow Experimental Forest in West Virginia, USA. We observed significant declines in plant C investment to fine root biomass (24.7%), root morphology, and arbuscular mycorrhizal (AM) colonization (55.9%). Moreover, we found that declines in extracellular enzyme activity were significantly correlated with a shift in bacterial community composition, but not fungal community composition. This bacterial community shift was also correlated with reduced AM fungal colonization indicating that declines in plant investment belowground drive the response of bacterial community structure and function to N fertilization. Collectively, we find that enzyme activity responses to N fertilization are not solely driven by fungi, but instead reflect a whole ecosystem response, whereby declines in the strength of belowground C investment to gain N cascade through the soil environment.

    

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