Streamlined spatial and environmental expression signatures characterize the minimalist duckweed Wolffia australiana.

Single-cell genomics permits a new resolution in the examination of molecular and cellular dynamics, allowing global, parallel assessments of cell types and cellular behaviors through development and in response to environmental circumstances, such as interaction with water and the light-dark cycle of the Earth. Here, we leverage the smallest, and possibly most structurally reduced, plant, the semiaquatic Wolffia australiana, to understand dynamics of cell expression in these contexts at the whole-plant level. We examined single-cell-resolution RNA-sequencing data and found Wolffia cells divide into four principal clusters representing the above- and below-water-situated parenchyma and epidermis. Although these tissues share transcriptomic similarity with model plants, they display distinct adaptations that Wolffia has made for the aquatic environment. Within this broad classification, discrete subspecializations are evident, with select cells showing unique transcriptomic signatures associated with developmental maturation and specialized physiologies. Assessing this simplified biological system temporally at two key time-of-day (TOD) transitions, we identify additional TOD-responsive genes previously overlooked in whole-plant transcriptomic approaches and demonstrate that the core circadian clock machinery and its downstream responses can vary in cell-specific manners, even in this simplified system. Distinctions between cell types and their responses to submergence and/or TOD are driven by expression changes of unexpectedly few genes, characterizing Wolffia as a highly streamlined organism with the majority of genes dedicated to fundamental cellular processes. Wolffia provides a unique opportunity to apply reductionist biology to elucidate signaling functions at the organismal level, for which this work provides a powerful resource.

Phytohormonal regulation determines the organization pattern of shoot aerenchyma in greater duckweed (Spirodela polyrhiza).

Airspace or aerenchyma is crucial for plant development and acclimation to stresses such as hypoxia, drought, and nutritional deficiency. Although ethylene-mediated signaling cascades are known to regulate aerenchyma formation in stems and roots under hypoxic conditions, the precise mechanisms remain unclear. Moreover, the cellular dynamics underlying airspace formation in shoots are poorly understood. We investigated the stage-dependent structural dynamics of shoot aerenchyma in greater duckweed (Spirodela polyrhiza), a fast-growing aquatic herb with well-developed aerenchyma in its floating fronds. Using X-ray micro-computed tomography and histological analysis, we showed that the spatial framework of aerenchyma is established before frond volume increases, driven by cell division and expansion. The substomatal cavity connecting aerenchyma to stomata formed via programmed cell death (PCD) and was closely associated with guard cell development. Additionally, transcriptome analysis and pharmacological studies revealed that the organization of aerenchyma in greater duckweed is determined by the interplay between PCD and proliferation. This balance is governed by spatiotemporal regulation of phytohormone signaling involving ethylene, abscisic acid, and salicylic acid. Overall, our study reveals the structural dynamics and phytohormonal regulation underlying aerenchyma development in duckweed, improving our understanding of how plants establish distinct architectural arrangements. These insights hold the potential for wide-ranging application, not only in comprehending aerenchyma formation across various plant species but also in understanding how airspaces are formed within the leaves of terrestrial plants.

Gene expression and metabolite levels converge in the thermogenic spadix of skunk cabbage.

The inflorescence (spadix) of skunk cabbage (Symplocarpus renifolius) is strongly thermogenic and can regulate its temperature at around 23 °C even when the ambient temperature drops below freezing. To elucidate the mechanisms underlying developmentally controlled thermogenesis and thermoregulation in skunk cabbage, we conducted a comprehensive transcriptome and metabolome analysis across 3 developmental stages of spadix development. Our RNA-seq analysis revealed distinct groups of expressed genes, with selenium-binding protein 1/methanethiol oxidase (SBP1/MTO) exhibiting the highest levels in thermogenic florets. Notably, the expression of alternative oxidase (AOX) was consistently high from the prethermogenic stage through the thermogenic stage in the florets. Metabolome analysis showed that alterations in nucleotide levels correspond with the developmentally controlled and tissue-specific thermogenesis of skunk cabbage, evident by a substantial increase in AMP levels in thermogenic florets. Our study also reveals that hydrogen sulfide, a product of SBP1/MTO, inhibits cytochrome c oxidase (COX)-mediated mitochondrial respiration, while AOX-mediated respiration remains relatively unaffected. Specifically, at lower temperatures, the inhibitory effect of hydrogen sulfide on COX-mediated respiration increases, promoting a shift toward the dominance of AOX-mediated respiration. Finally, despite the differential regulation of genes and metabolites throughout spadix development, we observed a convergence of gene expression and metabolite accumulation patterns during thermogenesis. This synchrony may play a key role in developmentally regulated thermogenesis. Moreover, such convergence during the thermogenic stage in the spadix may provide a solid molecular basis for thermoregulation in skunk cabbage.

Caloric restriction extends lifespan in a clonal plant.

When subjected to dietary caloric restriction (CR), individual animals often outlive well-fed conspecifics. Here, we address whether CR also extends lifespan in plants. Whereas caloric intake in animals comes from ingestion, in plants it derives from photosynthesis. Thus, factors that reduce photosynthesis, such as reduced light intensity, can induce CR. In two lab experiments investigating the aquatic macrophyte Lemna minor, we tracked hundreds of individuals longitudinally, with light intensity-and hence, CR-manipulated using neutral-density filters. In both experiments, CR dramatically increased lifespan through a process of temporal scaling. Moreover, the magnitude of lifespan extension accorded with the assumptions that (a) light intensity positively relates to photosynthesis following Michaelis-Menten kinetics, and (b) photosynthesis negatively relates to lifespan via a power law. Our results emphasize that CR-mediated lifespan extension applies to autotrophs as well as heterotrophs, and suggest that variation in light intensity has quantitatively predictable effects on plant aging trajectories.

Spatio-Temporal Study of Galactolipid Biosynthesis in Duckweed Using Mass Spectrometry Imaging and in vivo Isotope Labeling.

Isotope labeling coupled with mass spectrometry imaging (MSI) presents a potent strategy for elucidating the dynamics of metabolism at cellular resolution, yet its application to plant systems is scarce. It has the potential to reveal the spatio-temporal dynamics of lipid biosynthesis during plant development. In this study, we explore its application to galactolipid biosynthesis of an aquatic plant, Lemna minor, with D2O labeling. Specifically, matrix-assisted laser desorption/ionization-MSI data of two major galactolipids in L. minor, monogalactosyldiacylglycerol and digalactosyldiacylglycerol, were studied after growing in 50% D2O media over a 15-day time period. When they were partially labeled after 5 d, three distinct binomial isotopologue distributions were observed corresponding to the labeling of partial structural moieties: galactose only, galactose and a fatty acyl chain and the entire molecule. The temporal change in the relative abundance of these distributions follows the expected linear pathway of galactolipid biosynthesis. Notably, their mass spectrometry images revealed the localization of each isotopologue group to the old parent frond, the intermediate tissues and the newly grown daughter fronds. Besides, two additional labeling experiments, (i) 13CO2 labeling and (ii) backward labeling of completely 50% D2O-labeled L. minor in H2O media, confirm the observations in forward labeling. Furthermore, these experiments unveiled hidden isotopologue distributions indicative of membrane lipid restructuring. This study suggests the potential of isotope labeling using MSI to provide spatio-temporal details in lipid biosynthesis in plant development.

Auxin and carbohydrate control flower bud development in Anthurium andraeanum during early stage of sexual reproduction.

BACKGROUND: Flower buds of Anthurium andraeanum frequently cease to grow and abort during the early flowering stage, resulting in prolonged planting times and increased commercialization costs. Nevertheless, limited knowledge exists of the mechanism of flower development after initiation in A. andraeanum. RESULTS: In this study, the measurement of carbohydrate flow and intensity between leaves and flowers during different growth stages showed that tender leaves are strong sinks and their concomitant flowers are weak ones. This suggested that the tender leaves compete with their concomitant flower buds for carbohydrates during the early growth stages, potentially causing the abortion of the flower buds. The analysis of transcriptomic differentially expressed genes suggested that genes related to sucrose metabolism and auxin response play an important role during flower bud development. Particularly, co-expression network analysis found that AaSPL12 is a hub gene engaged in flower development by collaborating carbohydrate and auxin signals. Yeast Two Hybrid assays revealed that AaSPL12 can interact with AaARP, a protein that serves as an indicator of dormancy. Additionally, the application of exogenous IAA and sucrose can suppress the expression of AaARP, augment the transcriptional abundance of AaSPL12, and consequently expedite flower development in Anthurium andraeanum. CONCLUSIONS: Collectively, our findings indicated that the combination of auxin and sugar signals could potentially suppress the repression of AaARP protein to AaSPL12, thus advancing the development of flower buds in Anthurium andraeanum.

Optimizing biochar, vermicompost, and duckweed amendments to mitigate arsenic uptake and accumulation in rice (Oryza sativa L.) cultivated on arsenic-contaminated soil.

The accumulation of arsenic (As) in rice (Oryza sativa L.) grain poses a significant health concern in Bangladesh. To address this, we investigated the efficacy of various organic amendments and phytoremediation techniques in reducing As buildup in O. sativa. We evaluated the impact of five doses of biochar (BC; BC0.1: 0.1%, BC0.28: 0.28%, BC0.55: 0.55%, BC0.82: 0.82% and BC1.0: 1.0%, w/w), vermicompost (VC; VC1.0: 1.0%, VC1.8: 1.8%, VC3.0: 3.0%, VC4.2: 4.2% and VC5.0: 5.0%, w/w), and floating duckweed (DW; DW100: 100, DW160: 160, DW250: 250, DW340: 340 and DW400: 400 g m- 2) on O. sativa cultivated in As-contaminated soil. Employing a three-factor five-level central composite design and response surface methodology (RSM), we optimized the application rates of BC-VC-DW. Our findings revealed that As contamination in the soil negatively impacted O. sativa growth. However, the addition of BC, VC, and DW significantly enhanced plant morphological parameters, SPAD value, and grain yield per pot. Notably, a combination of moderate BC-DW and high VC (BC0.55VC5DW250) increased grain yield by 44.4% compared to the control (BC0VC0DW0). As contamination increased root, straw, and grain As levels, and oxidative stress in O. sativa leaves. However, treatment BC0.82VC4.2DW340 significantly reduced grain As (G-As) by 56%, leaf hydrogen peroxide by 71%, and malondialdehyde by 50% compared to the control. Lower doses of BC-VC-DW (BC0.28VC1.8DW160) increased antioxidant enzyme activities, while moderate to high doses resulted in a decline in these activities. Bioconcentration and translocation factors below 1 indicated limited As uptake and translocation in plant tissues. Through RSM optimization, we determined that optimal doses of BC (0.76%), VC (4.62%), and DW (290.0 g m- 2) could maximize grain yield (32.96 g pot- 1, 44% higher than control) and minimize G-As content (0.189 mg kg- 1, 54% lower than control). These findings underscore effective strategies for enhancing yield and reducing As accumulation in grains from contaminated areas, thereby ensuring agricultural productivity, human health, and long-term sustainability. Overall, our study contributes to safer food production and improved public health in As-affected regions.

Trophic interactions in microbiomes influence plant host population size and ecosystem function.

Plant microbiomes that comprise diverse microorganisms, including prokaryotes, eukaryotes and viruses, are the key determinants of plant population dynamics and ecosystem function. Despite their importance, little is known about how species interactions (especially trophic interactions) between microbes from different domains modify the importance of microbiomes for plant hosts and ecosystems. Using the common duckweed Lemna minor, we experimentally examined the effects of predation (by bacterivorous protists) and parasitism (by bacteriophages) within microbiomes on plant population size and ecosystem phosphorus removal. Our results revealed that the addition of predators increased plant population size and phosphorus removal, whereas the addition of parasites showed the opposite pattern. The structural equation modelling further pointed out that predation and parasitism affected plant population size and ecosystem function via distinct mechanisms that were both mediated by microbiomes. Our results highlight the importance of understanding microbial trophic interactions for predicting the outcomes and ecosystem impacts of plant-microbiome symbiosis.

A non-cell-autonomous circadian rhythm of bioluminescence reporter activities in individual duckweed cells.

The circadian clock is responsible for the temporal regulation of various physiological processes in plants. Individual cells contain a circadian oscillator consisting of a clock gene circuit that coordinates physiological rhythms within the plant body in an orderly manner. The coordination of time information has been studied from the perspective of cell-cell local coupling and long-distance communication between tissues based on the view that the behavior of circadian oscillators represents physiological rhythms. Here, we report the cellular circadian rhythm of bioluminescence reporters that are not governed by the clock gene circuit in expressing cells. We detected cellular bioluminescence rhythms with different free-running periods in the same cells using a dual-color bioluminescence monitoring system in duckweed (Lemna minor) transfected with Arabidopsis CIRCADIAN CLOCK ASSOCIATED 1::luciferace+ (AtCCA1::LUC+) and Cauliflower mosaic virus 35S::modified click-beetle red-color luciferase (CaMV35S::PtRLUC) reporters. Co-transfection experiments with the two reporters and a clock gene-overexpressing effector revealed that the AtCCA1::LUC+ rhythm, but not the CaMV35S::PtRLUC rhythm, was altered in cells with a dysfunctional clock gene circuit. This indicated that the AtCCA1::LUC+ rhythm is a direct output of the cellular circadian oscillator, whereas the CaMV35S::PtRLUC rhythm is not. After plasmolysis, the CaMV35S::PtRLUC rhythm disappeared, whereas the AtCCA1::LUC+ rhythm persisted. This suggests that the CaMV35S::PtRLUC bioluminescence has a symplast/apoplast-mediated circadian rhythm generated at the organismal level. The CaMV35S::PtRLUC-type bioluminescence rhythm was also observed when other bioluminescence reporters were expressed. These results reveal that the plant circadian system consists of both cell-autonomous and noncell-autonomous rhythms that are unaffected by cellular oscillators.

Characterization of the cryptic interspecific hybrid Lemna×mediterranea by an integrated approach provides new insights into duckweed diversity.

Lemnaceae taxonomy is challenged by the particular morphology of these tiny free-floating angiosperms. Although molecular taxonomy has helped clarify the phylogenetic history of this family, some inconsistency with morphological data leads to frequent misclassifications in the genus Lemna. Recently, the finding that Lemna japonica is an interspecific hybrid between Lemna minor and Lemna turionifera provided a clear explanation for one such taxonomic question. Here we demonstrated that L. minor is also capable of hybridizing with Lemna gibba, generating a cryptic but widespread taxon in the Mediterranean area. The nothotaxon Lemna ×mediterranea is described and compared with clones of the putative parental species L. minor and L. gibba. Genetic analysis by nuclear and plastid markers, as well as genome size measurement, revealed that two different cytotypes, diploid and triploid, originated by at least two independent hybridization events. Despite high overall similarity, morphometrical, physiological, and biochemical analyses showed an intermediate position of L. ×mediterranea between its parental species in most qualitative and quantitative characters, and also separation of the two hybrid cytotypes by some criteria. These data provide evidence that hybridization and polyploidization, driving forces of terrestrial plant evolution, contribute to duckweed genetic diversity and may have shaped the phylogenetic history of these mainly asexual, aquatic plants.

Return of the Lemnaceae: duckweed as a model plant system in the genomics and postgenomics era.

The aquatic Lemnaceae family, commonly called duckweed, comprises some of the smallest and fastest growing angiosperms known on Earth. Their tiny size, rapid growth by clonal propagation, and facile uptake of labeled compounds from the media were attractive features that made them a well-known model for plant biology from 1950 to 1990. Interest in duckweed has steadily regained momentum over the past decade, driven in part by the growing need to identify alternative plants from traditional agricultural crops that can help tackle urgent societal challenges, such as climate change and rapid population expansion. Propelled by rapid advances in genomic technologies, recent studies with duckweed again highlight the potential of these small plants to enable discoveries in diverse fields from ecology to chronobiology. Building on established community resources, duckweed is reemerging as a platform to study plant processes at the systems level and to translate knowledge gained for field deployment to address some of society's pressing needs. This review details the anatomy, development, physiology, and molecular characteristics of the Lemnaceae to introduce them to the broader plant research community. We highlight recent research enabled by Lemnaceae to demonstrate how these plants can be used for quantitative studies of complex processes and for revealing potentially novel strategies in plant defense and genome maintenance.

The evolution of the duckweed ionome mirrors losses in structural complexity.

BACKGROUND AND AIMS: The duckweeds (Lemnaceae) consist of 36 species exhibiting impressive phenotypic variation, including the progressive evolutionary loss of a fundamental plant organ, the root. Loss of roots and reduction of vascular tissues in recently derived taxa occur in concert with genome expansions of ≤14-fold. Given the paired loss of roots and reduction in structural complexity in derived taxa, we focus on the evolution of the ionome (whole-plant elemental contents) in the context of these fundamental changes in body plan. We expect that progressive vestigiality and eventual loss of roots might have both adaptive and maladaptive consequences that are hitherto unknown. METHODS: We quantified the ionomes of 34 accessions in 21 species across all duckweed genera, spanning 70 Myr in this rapidly cycling plant (doubling times are as rapid as ~24 h). We related both micro- and macroevolutionary ionome contrasts to body plan remodelling and showed nimble microevolutionary shifts in elemental accumulation and exclusion in novel accessions. KEY RESULTS: We observed a robust directional trend in calcium and magnesium levels, decreasing from the ancestral representative Spirodela genus towards the derived rootless Wolffia, with the latter also accumulating cadmium. We also identified abundant within-species variation and hyperaccumulators of specific elements, with this extensive variation at the fine (as opposed to broad) scale. CONCLUSIONS: These data underscore the impact of root loss and reveal the very fine scale of microevolutionary variation in hyperaccumulation and exclusion of a wide range of elements. Broadly, they might point to trade-offs not well recognized in ionomes.

The immediate metabolomic effects of whole-genome duplication in the greater duckweed, Spirodela polyrhiza.

PREMISE: In plants, whole-genome duplication (WGD) is a common mutation with profound evolutionary potential. Given the costs associated with a superfluous genome copy, polyploid establishment is enigmatic. However, in the right environment, immediate phenotypic changes following WGD can facilitate establishment. Metabolite abundances are the direct output of the cell's regulatory network and determine much of the impact of environmental and genetic change on the phenotype. While it is well known that an increase in the bulk amount of genetic material can increase cell size, the impact of gene dosage multiplication on the metabolome remains largely unknown. METHODS: We used untargeted metabolomics on four genetically distinct diploid-neoautotetraploid pairs of the greater duckweed, Spirodela polyrhiza, to investigate how WGD affects metabolite abundances per cell and per biomass. RESULTS: Autopolyploidy increased metabolite levels per cell, but the response of individual metabolites varied considerably. However, the impact on metabolite level per biomass was restricted because the increased cell size reduced the metabolite concentration per cell. Nevertheless, we detected both quantitative and qualitative effects of WGD on the metabolome. Many effects were strain-specific, but some were shared by all four strains. CONCLUSIONS: The nature and impact of metabolic changes after WGD depended strongly on the genotype. Dosage effects have the potential to alter the plant metabolome qualitatively and quantitatively, but were largely balanced out by the reduction in metabolite concentration due to an increase in cell size in this species.

Neopolyploidy-induced changes in giant duckweed (Spirodela polyrhiza) alter herbivore preference and performance and plant population performance.

PREMISE: Polyploidy is a widespread mutational process in angiosperms that may alter population performance of not only plants but also their interacting species. Yet, knowledge of whether polyploidy affects plant-herbivore dynamics is scarce. Here, we tested whether aphid herbivores exhibit preference for diploid or neopolyploid plants, whether polyploidy impacts plant and herbivore performance, and whether these interactions depend on the plant genetic background. METHODS: Using independently synthesized neotetraploid strains paired with their diploid progenitors of greater duckweed (Spirodela polyrhiza), we evaluated the effect of neopolyploidy on duckweed's interaction with the water-lily aphid (Rhopalosiphum nymphaeae). Using paired-choice experiments, we evaluated feeding preference of the herbivore. We then evaluated the consequences of polyploidy on aphid and plant performance by measuring population growth over multiple generations. RESULTS: Aphids preferred neopolyploids when plants were provided at equal abundances but not at equal surface areas, suggesting the role of plant population surface area in driving this preference. Additionally, neopolyploidy increased aphid population performance, but this result was dependent on the plant's genetic lineage. Lastly, the impact of herbivory on neopolyploid vs. diploid duckweed varied greatly with genetic lineage, where neopolyploids appeared to be variably tolerant compared to diploids, sometimes mirroring the effect on herbivore performance. CONCLUSIONS: By experimentally testing the impacts of polyploidy on trophic species interactions, we showed that polyploidization can impact the preference and performance of herbivores on their plant hosts. These results have significant implications for the establishment and persistence of plants and herbivores in the face of plant polyploidy.

Exploring the ecotoxicological impact of meropenem on Lemna minor: Growth, photosynthetic activity, and oxidative stress.

Meropenem is a potent carbapenem antibiotic frequently used in medical settings. Several studies have confirmed the pervasive presence of these antibiotics in wastewater treatment plants and aquatic environments. However, the effects of these substances on non-target organisms, such as plants, have not been adequately monitored. Thus, this study aimed to assess the short-term impact of meropenem on the growth, photosynthesis, chlorophyll content, and enzyme activity of the macrophyte plant Lemna minor. The methods involved exposing the plant to meropenem under controlled conditions and assessing physiological and biochemical parameters to determine the impact on photosynthetic activity and oxidative stress. These analyses included growth rate, antioxidant enzyme activity, and photosynthetic capacity. The findings suggest that the growth rate of Lemna minor remained unaffected by meropenem at concentrations <200000 µgL-1. However, plants exposed to concentrations >20 µgL-1showed physiological alterations, such as decreased net photosynthesis rate (17%) and chlorophyll concentration (57%), compared to the control group. For acute toxicity assays, the calculated EC50 7-day and EC20 7-day were 1135 µgL-1and 33 µgL-1, respectively. In addition, in most treatments tested, meropenem caused an increase in the superoxide dismutase (SOD), catalase (CAT) and ascorbate peroxidase (APX) activity as a defense mechanism against oxidative stress. Our results suggest that meropenem affects photosynthetic processes and induces oxidative stress in the macrophyte plant Lemna minor. Further studies are needed to assess the physiological and metabolic interactions between antibiotics and primary producers at different long-term trophic levels.

Xanthomonas Strains Isolated from Hosts in the Family Araceae Reveal Diverse Phylogenetic Relationships and Origins.

The family Araceae, comprising ornamentals including Anthurium, Dieffenbachia, Philodendron, Colocasia, and Zantedeschia, is susceptible to Xanthomonas pathogens. Previous analyses have established heterogeneity in aroid strains, yet unresolved taxonomic positions and dynamics between Xanthomonas and frequently associated Stenotrophomonas in aroids necessitate in-depth genetic investigation to resolve these complex relationships. This study utilized multilocus sequence analysis of housekeeping genes atpD, dnaA, dnaK, gltA, and gyrB to investigate 59 aroid strains, selected based on hosts, time, and geographical origins. After adding sequences from additional strains from NCBI GenBank, analysis of 161 concatenated sequences indicated that all aroid strains fell within Xanthomonas and Stenotrophomonas. Thirty-six strains isolated from Anthurium grouped under X. phaseoli, with outliers including one strain each in X. arboricola and X. sacchari and two in Stenotrophomonas. Six strains from Caladium, Dieffenbachia, and Philodendron formed host-specific subgroups within X. euvesicatoria. One strain from Dieffenbachia aligned with X. campestris, whereas strains from Colocasia, Aglaonema, and Spathiphyllum clustered with X. sacchari. Apart from the zantedeschia strain described as X. arboricola pv. zantedeschiae, two colocasia, one epipremnum, and one anthurium strain joined the X. arboricola group. Overall, this study revealed significant heterogeneity among aroid strains, with anthurium strains clustering closely despite distant geographical origins. The analysis underscores the complexity of host-pathogen specificity within Xanthomonas and emphasizes the need for further taxonomic clarification through whole-genome analysis of representative strains. The findings of this research will facilitate strain selection for inclusivity and exclusivity panels in developing diagnostic assays for X. phaseoli and xanthomonads affecting aroids.

Actinomycetospora lemnae sp. nov., A Novel Actinobacterium Isolated from Lemna aequinoctialis Able to Enhance Duckweed Growth.

Duckweed-associated actinobacteria are co-existing microbes that affect duckweed growth and adaptation. In this study, we aimed to report a novel actinobacterium species and explore its ability to enhance duckweed growth. Strain DW7H6T was isolated from duckweed, Lemna aequinoctialis. Phylogenetic analysis based on its 16S rRNA gene sequence revealed that the strain was most closely related to Actinomycetospora straminea IY07-55T (99.0%), Actinomycetospora chibensis TT04-21T (98.9%), Actinomycetospora lutea TT00-04T (98.8%) and Actinomycetospora callitridis CAP 335T (98.4%). Chemotaxonomic and morphological characteristics of strain DW7H6T were consistent with members of the genus Actinomycetospora, while average nucleotide identity (ANI) and digital DNA-DNA hybridization (dDDH) between the draft genomes of this strain and its closely related type strains were below the proposed threshold values used for species discrimination. Based on chemotaxonomic, phylogenetic, phenotypic, and genomic evidence obtained, we describe a novel Actinomycetospora species, for which the name Actinomycetospora lemnae sp. nov. is proposed. The type strain is DW7H6T (TBRC 15165T, NBRC 115294T). Additionally, the duckweed-associated actinobacterium strain DW7H6T was able to enhance duckweed growth when compared to the control, in which the number of fronds and biomass dry weight were increased by up to 1.4 and 1.3 fold, respectively. Moreover, several plant-associated gene features in the genome of strain DW7H6T potentially involved in plant-microbe interactions were identified.

Two lineages of Lemna aequinoctialis (Araceae, Lemnoideae) based on physiology, morphology, and phylogeny.

Lemna aequinoctialis Welw. is a widely spread species that has diverse physiological and molecular properties. Flower characteristics are important factors in deducing taxonomical status; however, owing to the rarity of flowering observations in Lemna, studying them has been a prolonged challenge. In this study, physiological and morphological analyses were conducted by inducing flowering, and molecular analysis was done based on the two chloroplast DNA loci (matK, atpF-atpH intergeneric spacer) of L. aequinoctialis sensu Landolt (1986) from 70 strains found in 70 localities in Japan, Korea, Thailand, and the US. In total, 752 flowering fronds from 13 strains were observed based on axenic conditions. Two different trends in flower organ development-protogyny and adichogamy-were detected in these strains. Their physiological traits were divided into two groups, showing different morphological features based on frond thickness, root cap, and anther sizes. Molecular analysis showed two lineages corresponding to two physiological groups. These were identified as L. aequinoctialis sensu Beppu et al. (1985) and L. aoukikusa Beppu et Murata based on the description of the nomenclature of L. aoukikusa. These were concluded as independent taxa and can be treated as different species. Furthermore, the distribution of L. aoukikusa is not only limited to Japan.

Responses of Vallisneria natans and Pistia stratiotes to Cu2+ and Mn2+ stress: Occurrence of caffeic acid and its degradation kinetics during chlorination.

Macrophytes are crucial in maintaining the equilibrium of aquatic ecosystems. However, the pattern of macrophyte-derived caffeic acid (CA) release under heavy metal stress is yet to be fully understood. More importantly, due to its functional groups, CA may be a precursor to the formation of disinfection by-products, posing threats to water ecology and even safety of human drinking water. This study analyzed the responses of CA released by Vallisneria natans (V. natans) and Pistia stratiotes (P. Stratiotes) when exposed to Cu2+ and Mn2+ stress. Additionally, the CA levels in two constructed wetland ponds were detected and the degradation kinetics of CA during chlorination were investigated. Results indicated that CA occurred in two constructed wetland ponds with the concentrations of 44.727 µg/L (planted with V. natans) and 61.607 µg/L (planted with P. Stratiotes). Notably, heavy metal stress could significantly affect CA release from V. natans and P. Stratiotes. In general, under Cu2+ stress, V. natans secreted far more CA than under Mn2+ stress, the level could reach up to 435.303 µg/L. However, compared to V. natans, P. Stratiotes was less affected by Cu2+ and Mn2+ stress, releasing a maximum CA content of 55.582 µg/L under 5 mg/L Mn2+ stress. Aquatic macrophytes secreted more CA in response to heavy metal stresses and protected macrophytes from harmful heavy metals. CA degradation followed the pseudo first-order kinetics model, and the chlorination of CA conformed to a second-order reaction. The reaction rate significantly accelerated as NaClO, pH, temperature and Br- concentration increased. A new pathway for CA degradation and a new DBP 2, 2, 3, 3-tetrachloropropanal were observed. These findings pointed at a new direction into the adverse effect of CA, potentially paving the way for new strategies to solve drinking water safety problems.

Effects of humic acid on nutrient removal efficiency of aquatic duckweed (Lemna minor) and both growth performance, and hemato-biochemical parameters of Nile tilapia (Oreochromis niloticus) cultured in water recirculating system.

This study was carried out to evaluate the effects of humic acid (HA) on the nutrient removal efficiencies of aquatic duckweed plant (Lemna minor) from a water recirculating system used to culture Nile tilapia (Oreochromis niloticus) fish for 30 days. The HA was added to water at three concentrations of 0 (Control), 1.5, and 3 mg/L in triplicate. Water quality parameters, growth performance, and some hemato-biochemical parameters of the fish in variable HA concentrations were compared. The total ammonia nitrogen (TAN) and total phosphorous (TP) removal efficiency of L. minor increased with increasing the HA concentration from 0 mg/L to 3 mg/L (p < 0.05). The concentration of nitrate (NO3-) in the HA-3 mg/L was higher than that in the other groups on days 20 and 30 of the fish cultivation period (p < 0.05). The growth performance of fish improved in the HA-3 mg/L compared to the other groups. The addition of different concentrations of HA to water had no adverse effect on the hematological properties of the Nile tilapia. The plasma aspartate aminotransferase (AST), alanine aminotransferase (ALT), and alkaline phosphatase (ALP) levels in the HA-0 mg/L and HA-1.5 mg/L groups were higher than in the HA-3 mg/L (p < 0.05). No significant differences in the plasma glucose and cholesterol levels were observed between the HA-groups (p > 0.05), while the triglyceride level increased in the HA-3 mg/L compared to the control (p < 0.05). These results indicated that adding HA to water could be an effective method to enhance the bioremediation performance of the aquatic duckweed plants as biofilter and thus improve water quality, subsequently, fish growth performance in RASs.

The current study applied aquatic duckweed plant (Lemna minor) as a new biofilter in a water recirculating system used to culture Nile tilapia (Oreochromis niloticus) fish. The effects of three concentrations of humic acid (HA) as water additive on the nutrient removal efficiency of L. minor from water were investigated. HA improved bioremediation performance of the aquatic duckweed plant.