Molecular and phylogenetic evidence of parallel expansion of anion channels in plants.

Aluminum-activated malate transporters (ALMTs) and slow anion channels (SLACs) are important in various physiological processes in plants, including stomatal regulation, nutrient uptake, and in response to abiotic stress such as aluminum toxicity. To understand their evolutionary history and functional divergence, we conducted phylogenetic and expression analyses of ALMTs and SLACs in green plants. Our findings from phylogenetic studies indicate that ALMTs and SLACs may have originated from green algae and red algae, respectively. The ALMTs of early land plants and charophytes formed a monophyletic clade consisting of three subgroups. A single duplication event of ALMTs was identified in vascular plants and subsequent duplications into six clades occurred in angiosperms, including an identified clade, 1-1. The ALMTs experienced gene number losses in clades 1-1 and 2-1 and expansions in clades 1-2 and 2-2b. Interestingly, the expansion of clade 1-2 was also associated with higher expression levels compared to genes in clades that experienced apparent loss. SLACs first diversified in bryophytes, followed by duplication in vascular plants, giving rise to three distinct clades (I, II, and III), and clade II potentially associated with stomatal control in seed plants. SLACs show losses in clades II and III without substantial expansion in clade I. Additionally, ALMT clade 2-2 and SLAC clade III contain genes specifically expressed in reproductive organs and roots in angiosperms, lycophytes, and mosses, indicating neofunctionalization. In summary, our study demonstrates the evolutionary complexity of ALMTs and SLACs, highlighting their crucial role in the adaptation and diversification of vascular plants.

Infection with 'Candidatus Liberibacter asiaticus' improves the fecundity of Diaphorina citri aiding its proliferation: A win-win strategy.

The evolution of insect vector-pathogen relationships has long been of interest in the field of molecular ecology. One system of special relevance, due to its economic impacts, is that between Diaphorina citri and 'Candidatus Liberibacter asiaticus' (CLas), the cause of the severe Asian form of huanglongbing. CLas-positive D. citri are more fecund than their CLas-negative counterparts, boosting opportunities for pathogens to acquire new vector hosts. The molecular mechanism behind this life-history shift remains unclear. Here, we found that CLas promoted ovarian development and increased the expression of the vitellogenin receptor (DcVgR) in ovaries. DcVgR RNAi significantly decreased fecundity and CLas titer in ovaries, extended the preoviposition period, shortened the oviposition period and blocked ovarian development. Given their importance in gene regulation, we explored the role of miRNAs in shaping these phenotypes and their molecular triggers. Our results showed that one miRNA, miR-275, suppressed DcVgR expression by binding to its 3' UTR. Overexpression of miR-275 knocked down DcVgR expression and CLas titer in ovaries, causing reproductive defects that mimicked DcVgR knockdown phenotypes. We focused, further, on roles of the Juvenile Hormone (JH) pathway in shaping the observed fecundity phenotype, given its known impacts on ovarian development. After CLas infection, this pathway was upregulated, thereby increasing DcVgR expression. From these combined results, we conclude that CLas hijacks the JH signalling pathway and miR-275, thereby targeting DcVgR to increase D. citri fecundity. These changes simultaneously increase CLas replication, suggesting a pathogen-vector host mutualism, or a seemingly helpful, but cryptically costly life-history manipulation.

Natural Infection of Murraya paniculata and Murraya sumatrana with 'Candidatus Liberibacter asiaticus' in Java.

The phloem-limited bacterium 'Candidatus Liberibacter asiaticus' (CLas) is the putative causal pathogen of the severe Asiatic form of huanglongbing (citrus greening) and is most commonly transmitted by the Asiatic citrus psyllid Diaphorina citri. CLas severely affects many Citrus species and hybrids and has been recorded in the Citrus relative, orange jasmine, Murraya paniculata (L.) Jack (syn. M. exotica L.). In this study, 13 accessions of three Murraya species (M. paniculata, M. sumatrana Roxb., and M. lucida [G.Forst.] Mabb.) and the Papuan form of a putative hybrid (M. omphalocarpa Hayata) were identified morphologically and molecularly based on sequence identity of the matK-5'trnK region of the chloroplast genome, and infection on these plants under field conditions was determined by PCR and quantitative real-time PCR (qPCR) on two to four occasions over 14 months. CLas was repeatedly detected in leaflet midribs by PCR and qPCR on four and three accessions of M. paniculata and M. sumatrana, respectively. It was not detected in leaflet midribs of single accessions of M. lucida and M. omphalocarpa. The species identification of the CLas-positive accessions was further confirmed using all the molecular taxonomic markers consisting of the six fragments of the maternally inherited chloroplast genome and part of the nuclear-encoded internal transcribed spacer (ITS) region. The results indicated that natural infection of M. paniculata and M. sumatrana with CLas can occur in Java. To our knowledge, this is the first demonstration of the natural infection of M. sumatrana with CLas. Further studies are required to determine whether infections persist in the absence of D. citri.

Evolutionary Significance of NHX Family and NHX1 in Salinity Stress Adaptation in the Genus Oryza.

Rice (Oryza sativa), a staple crop for a substantial part of the world's population, is highly sensitive to soil salinity; however, some wild Oryza relatives can survive in highly saline environments. Sodium/hydrogen antiporter (NHX) family members contribute to Na+ homeostasis in plants and play a major role in conferring salinity tolerance. In this study, we analyzed the evolution of NHX family members using phylogeny, conserved domains, tertiary structures, expression patterns, and physiology of cultivated and wild Oryza species to decipher the role of NHXs in salt tolerance in Oryza. Phylogenetic analysis showed that the NHX family can be classified into three subfamilies directly related to their subcellular localization: endomembrane, plasma membrane, and tonoplast (vacuolar subfamily, vNHX1). Phylogenetic and structural analysis showed that vNHX1s have evolved from streptophyte algae (e.g., Klebsormidium nitens) and are abundant and highly conserved in all major land plant lineages, including Oryza. Moreover, we showed that tissue tolerance is a crucial trait conferring tolerance to salinity in wild rice species. Higher Na+ accumulation and reduced Na+ effluxes in leaf mesophyll were observed in the salt-tolerant wild rice species O. alta, O. latifolia, and O. coarctata. Among the key genes affecting tissue tolerance, expression of NHX1 and SOS1/NHX7 exhibited significant correlation with salt tolerance among the rice species and cultivars. This study provides insights into the evolutionary origin of plant NHXs and their role in tissue tolerance of Oryza species and facilitates the inclusion of this trait during the development of salinity-tolerant rice cultivars.

A ß-ketoacyl carrier protein reductase confers heat tolerance via the regulation of fatty acid biosynthesis and stress signaling in rice.

Heat stress is a major environmental threat affecting crop growth and productivity. However, the molecular mechanisms associated with plant responses to heat stress are poorly understood. Here, we identified a heat stress-sensitive mutant, hts1, in rice. HTS1 encodes a thylakoid membrane-localized ß-ketoacyl carrier protein reductase (KAR) involved in de novo fatty acid biosynthesis. Phylogenetic and bioinformatic analysis showed that HTS1 probably originated from streptophyte algae and is evolutionarily conserved in land plants. Thermostable HTS1 is predominantly expressed in green tissues and strongly induced by heat stress, but is less responsive to salinity, cold and drought treatments. An amino acid substitution at A254T in HTS1 causes a significant decrease in KAR enzymatic activity and, consequently, impairs fatty acid synthesis and lipid metabolism in the hts1 mutant, especially under heat stress. Compared to the wild-type, the hts1 mutant exhibited heat-induced higher H2 O2 accumulation, a larger Ca2+ influx to mesophyll cells, and more damage to membranes and chloroplasts. Also, disrupted heat stress signaling in the hts1 mutant depresses the transcriptional activation of HsfA2s and the downstream target genes. We suggest that HTS1 is critical for underpinning membrane stability, chloroplast integrity and stress signaling for heat tolerance in rice.

Sodium sequestration confers salinity tolerance in an ancestral wild rice.

Wild rice Oryza rufipogon, a progenitor of cultivated rice Oryza sativa L., possesses superior salinity tolerance and is a potential donor for breeding salinity tolerance traits in rice. However, a mechanistic basis of salinity tolerance in this donor species has not been established. Here, we examined salinity tolerance from the early vegetative stage to maturity in O. rufipogon in comparison with a salt-susceptible (Koshihikari) and a salt-tolerant (Reiziq) variety of O. sativa. We assessed their phylogeny and agronomical traits, photosynthetic performance, ion contents, as well as gene expression in response to salinity stress. Salt-tolerant O. rufipogon exhibited efficient leaf photosynthesis and less damage to leaf tissues during the course of salinity treatment. In addition, O. rufipogon showed a significantly higher tissue Na+ accumulation that is achieved by vacuolar sequestration compared to the salt tolerant O. sativa indica subspecies. These findings are further supported by the upregulation of genes involved with ion transport and sequestration (e.g. high affinity K+ transporter 1;4 [HKT1;4], Na+ /H+ exchanger 1 [NHX1] and vacuolar H+ -ATPase c [VHA-c]) in salt-tolerant O. rufipogon as well as by the close phylogenetic relationship of key salt-responsive genes in O. rufipogon to these in salt-tolerant wild rice species such as O. coarctata. Thus, the high accumulation of Na+ in the leaves of O. rufipogon acts as a cheap osmoticum to minimize the high energy cost of osmolyte biosynthesis and excessive reactive oxygen species production. These mechanisms demonstrated that O. rufipogon has important traits that can be used for improving salinity tolerance in cultivated rice.

Genomic adaptation to drought in wild barley is driven by edaphic natural selection at the Tabigha Evolution Slope.

Ecological divergence at a microsite suggests adaptive evolution, and this study examined two abutting wild barley populations, each 100 m across, differentially adapted to drought tolerance on two contrasting soil types, Terra Rossa and basalt at the Tabigha Evolution Slope, Israel. We resequenced the genomes of seven and six wild barley genotypes inhabiting the Terra Rossa and basalt soils, respectively, and identified a total of 69,192,653 single-nucleotide variants (SNVs) and insertions/deletions in comparison with a reference barley genome. Comparative genomic analysis between these abutting wild barley populations involved 19,615,087 high-quality SNVs. The results revealed dramatically different selection sweep regions relevant to drought tolerance driven by edaphic natural selection within 2,577 selected genes in these regions, including key drought-responsive genes associated with ABA synthesis and degradation (such as Cytochrome P450 protein) and ABA receptor complex (such as PYL2, SNF1-related kinase). The genetic diversity of the wild barley population inhabiting Terra Rossa soil is much higher than that from the basalt soil. Additionally, we identified different sets of genes for drought adaptation in the wild barley populations from Terra Rossa soil and from wild barley populations from Evolution Canyon I at Mount Carmel. These genes are associated with abscisic acid signaling, signaling and metabolism of reactive oxygen species, detoxification and antioxidative systems, rapid osmotic adjustment, and deep root morphology. The unique mechanisms for drought adaptation of the wild barley from the Tabigha Evolution Slope may be useful for crop improvement, particularly for breeding of barley cultivars with high drought tolerance.

Genome Sequence Resource of 'Candidatus Liberibacter asiaticus' from Diaphorina citri Kuwayama (Hemiptera: Liviidae) from La Réunion.

'Candidatus Liberibacter asiaticus' is an insect-transmitted, phloem-restricted α-proteobacterium associated with huanglongbing. Here, we provide the whole genome sequence of 'Ca. L. asiaticus' strain ReuSP1 from its insect vector Diaphorina citri (Hemiptera: Liviidae) collected in La Réunion. The genome is composed of 1,230,064 bp and has a 36.5% G+C content. This study reports the first 'Ca. L. asiaticus' genome sequence from La Réunion, which will add to 'Ca. L. asiaticus' genome resources and help elucidate our understanding of the introduction pathway into La Réunion.

Mechanistic Insights into Potassium-Conferred Drought Stress Tolerance in Cultivated and Tibetan Wild Barley: Differential Osmoregulation, Nutrient Retention, Secondary Metabolism and Antioxidative Defense Capacity.

Keeping the significance of potassium (K) nutrition in focus, this study explores the genotypic responses of two wild Tibetan barley genotypes (drought tolerant XZ5 and drought sensitive XZ54) and one drought tolerant barley cv. Tadmor, under the exposure of polyethylene glycol-induced drought stress. The results revealed that drought and K deprivation attenuated overall plant growth in all the tested genotypes; however, XZ5 was least affected due to its ability to retain K in its tissues which could be attributed to the smallest reductions of photosynthetic parameters, relative chlorophyll contents and the lowest Na+/K+ ratios in all treatments. Our results also indicate that higher H+/K+-ATPase activity (enhancement of 1.6 and 1.3-fold for shoot; 1.4 and 2.5-fold for root), higher shoot K+ (2 and 2.3-fold) and Ca2+ content (1.5 and 1.7-fold), better maintenance of turgor pressure by osmolyte accumulation and enhanced antioxidative performance to scavenge ROS, ultimately suppress lipid peroxidation (in shoots: 4% and 35%; in roots 4% and 20% less) and bestow higher tolerance to XZ5 against drought stress in comparison with Tadmor and XZ54, respectively. Conclusively, this study adds further evidence to support the concept that Tibetan wild barley genotypes that utilize K efficiently could serve as a valuable genetic resource for the provision of genes for improved K metabolism in addition to those for combating drought stress, thereby enabling the development of elite barley lines better tolerant of abiotic stresses.

Characterization of the bacterial communities of psyllids associated with Rutaceae in Bhutan by high throughput sequencing.

BACKGROUND: Several plant-pathogenic bacteria are transmitted by insect vector species that often also act as hosts. In this interface, these bacteria encounter plant endophytic, insect endosymbiotic and other microbes. Here, we used high throughput sequencing to examine the bacterial communities of five different psyllids associated with citrus and related plants of Rutaceae in Bhutan: Diaphorina citri, Diaphorina communis, Cornopsylla rotundiconis, Cacopsylla heterogena and an unidentified Cacopsylla sp. RESULTS: The microbiomes of the psyllids largely comprised their obligate P-endosymbiont 'Candidatus Carsonella ruddii', and one or two S-endosymbionts that are fixed and specific to each lineage. In addition, all contained Wolbachia strains; the Bhutanese accessions of D. citri were dominated by a Wolbachia strain first found in American isolates of D. citri, while D. communis accessions were dominated by the Wolbachia strain, wDi, first detected in D. citri from China. The S-endosymbionts from the five psyllids grouped with those from other psyllid taxa; all D. citri and D. communis individuals contained sequences matching 'Candidatus Profftella armatura' that has previously only been reported from other Diaphorina species, and the remaining psyllid species contained OTUs related to unclassified Enterobacteriaceae. The plant pathogenic 'Candidatus Liberibacter asiaticus' was found in D. citri but not in D. communis. Furthermore, an unidentified 'Candidatus Liberibacter sp.' occurred at low abundance in both Co. rotundiconis and the unidentified Cacopsylla sp. sampled from Zanthoxylum sp.; the status of this new liberibacter as a plant pathogen and its potential plant hosts are currently unknown. The bacterial communities of Co. rotundiconis also contained a range of OTUs with similarities to bacteria previously found in samples taken from various environmental sources. CONCLUSIONS: The bacterial microbiota detected in these Bhutanese psyllids support the trends that have been seen in previous studies: psyllids have microbiomes largely comprising their obligate P-endosymbiont and one or two S-endosymbionts. In addition, the association with plant pathogens has been demonstrated, with the detection of liberibacters in a known host, D. citri, and identification of a putative new species of liberibacter in Co. rotundiconis and Cacopsylla sp.

Molecular differentiation of the Murraya paniculata Complex (Rutaceae: Aurantioideae: Aurantieae).

BACKGROUND: Orange jasmine has a complex nomenclatural history and is now known as Murraya paniculata (L.) Jack. Our interest in this common ornamental stemmed from the need to resolve its identity and the identities of closely related taxa as hosts of the pathogen 'Candidatus Liberibacter asiaticus' and its vector Diaphorina citri. Understanding these microbe-vector-plant relationships has been hampered by taxonomic confusion surrounding Murraya at both the generic and specific levels. RESULTS: To resolve the taxonomic uncertainty, six regions of the maternally-inherited chloroplastal genome and part of the nuclear-encoded ITS region were amplified from 85 accessions of Murraya and Merrillia using the polymerase chain reaction (PCR). Clustering used maximum parsimony (MP), maximum likelihood (ML) and Bayesian inference (BI). Chronograms were produced for molecular dating, and to test the monophyly of Murraya rigorously, using selected accessions of Murraya and 26 accessions of the Rutaceae and Simarubaceae. Sequence data from the ITS and chloroplastal regions suggest that Murraya paniculata (sensu (Swingle WT and Reece CR, The Citrus Industry, p. 190-430, 1967)) can be separated into four distinct but morphologically somewhat cryptic taxa: Murraya paniculata (sensu (Mabberley DJ, Taxon 65:366-371, 2016)), M. elongata, M. sumatrana and M. lucida. In addition, Murraya omphalocarpa was identified as a putative hybrid of M. paniculata and M. lucida with two geographically isolated nothovarieties representing reciprocal crosses. Murraya is monophyletic, and molecular dating suggests that it diverged from Merrillia during the Miocene (23-5 Ma) with this Murraya group speciating and dispersing during the Middle Miocene onwards. CONCLUSIONS: The accessions from Asia and Australasia used in this study grouped into biogeographical regions that match herbarium specimen records for the taxa that suggest natural allopatric distributions with limited overlap and hybridity. Murraya paniculata has been distributed around the world as an ornamental plant. The division of the Murraya paniculata complex into four species with a rare hybrid also confirms morphological studies.

Field study reveals core plant microbiota and relative importance of their drivers.

Harnessing plant microbiota can assist in sustainably increasing primary productivity to meet growing global demands for food and biofuel. However, development of rational microbiome-based approaches for improving crop yield and productivity is currently hindered by a lack of understanding of the major biotic and abiotic factors shaping the crop microbiome under relevant field conditions. We examined bacterial and fungal communities associated with both aerial (leaves, stalks) and belowground (roots, soil) compartments of four commercial sugarcane varieties (Saccharum spp.) grown in several growing regions in Australia. We identified drivers of the sugarcane microbiome under field conditions and evaluated whether the plants shared a core microbiome. Sugarcane-associated microbial assemblages were primarily determined by plant compartment, followed by growing region, crop age, variety and Yellow Canopy Syndrome (YCS). We detected a core set of microbiota and identified members of the core microbiome that were influenced by YCS incidence. Our study revealed key hub microorganisms in the core microbiome networks of sugarcane leaves, stalks, roots and rhizosphere soil despite location and time-associated shifts in the community assemblages. Elucidating their functional roles and identification of the keystone core microbiota that sustain plant health could provide a technological breakthrough for a sustainable increase in crop productivity.

Roles of Chloroplast Retrograde Signals and Ion Transport in Plant Drought Tolerance.

Worldwide, drought affects crop yields; therefore, understanding plants' strategies to adapt to drought is critical. Chloroplasts are key regulators of plant responses, and signals from chloroplasts also regulate nuclear gene expression during drought. However, the interactions between chloroplast-initiated retrograde signals and ion channels under stress are still not clear. In this review, we summarise the retrograde signals that participate in regulating plant stress tolerance. We compare chloroplastic transporters that modulate retrograde signalling through retrograde biosynthesis or as critical components in retrograde signalling. We also discuss the roles of important plasma membrane and tonoplast ion transporters that are involved in regulating stomatal movement. We propose how retrograde signals interact with ion transporters under stress.

Tolerance to Drought, Low pH and Al Combined Stress in Tibetan Wild Barley Is Associated with Improvement of ATPase and Modulation of Antioxidant Defense System.

Aluminum (Al) toxicity and drought are two major constraints on plant growth in acidic soils, negatively affecting crop performance and yield. Genotypic differences in the effects of Al/low pH and polyethyleneglycol (PEG) induced drought stress, applied either individually or in combination, were studied in Tibetan wild (XZ5, drought-tolerant; XZ29, Al-tolerant) and cultivated barley (Al-tolerant Dayton; drought-tolerant Tadmor). Tibetan wild barley XZ5 and XZ29 had significantly higher H⁺-ATPase, Ca2+Mg2+-ATPase, and Na⁺K⁺-ATPase activities at pH 4.0+Al+PEG than Dayton and Tadmor. Moreover, XZ5 and XZ29 possessed increased levels in reduced ascorbate and glutathione under these conditions, and antioxidant enzyme activities were largely stimulated by exposure to pH 4.0+PEG, pH 4.0+Al, and pH 4.0+Al+PEG, compared to a control and to Dayton and Tadmor. The activity of methylglyoxal (MG) was negatively correlated with increased levels of glyoxalase (Gly) I and Gly II in wild barley. Microscopic imaging of each genotype revealed DNA damage and obvious ultrastructural alterations in leaf cells treated with drought or Al alone, and combined pH 4.0+Al+PEG stress; however, XZ29 and XZ5 were less affected than Dayton and Tadmor. Collectively, the authors findings indicated that the higher tolerance of the wild barley to combined pH 4.0+Al+PEG stress is associated with improved ATPase activities, increased glyoxalase activities, reduced MG, and lower reactive oxygen species levels (like O2- and H2O2) due to increased antioxidant enzyme activities. These results offer a broad comprehension of the mechanisms implicated in barley's tolerance to the combined stress of Al/low pH and drought, and may provide novel insights into the potential utilization of genetic resources, thereby facilitating the development of barley varieties tolerant to drought and Al/low pH stress.

QTLs for stomatal and photosynthetic traits related to salinity tolerance in barley.

BACKGROUND: Stomata regulate photosynthesis and transpiration, and these processes are critical for plant responses to abiotic stresses such as salinity. A barley double haploid population with 108 lines derived from a cross between CM72 (salt-tolerant) and Gairdner (salt-sensitive) was used to detect quantitative trait loci (QTLs) associated with stomatal and photosynthetic traits related to salinity tolerance. RESULTS: A total of 11 significant QTLs (LOD > 3.0) and 11 tentative QTLs (2.5 < LOD < 3.0) were identified. These QTLs are distributed on all the seven chromosomes, except 5H and explain 9.5-17.3% of the phenotypic variation. QTLs for biomass, intercellular CO2 concentration, transpiration rate and stomatal conductance under control conditions co-localised together. A QTL for biomass also co-located with one for transpiration rate under salinity stress. A linkage was found between stomatal pore area and gas exchange. A QTL for salinity tolerance also co-localised with QTLs for grain yield and biomass on chromosome 3H. Based on the draft barley genome, the candidate genes for salinity tolerance at this locus are proposed. CONCLUSIONS: The lack of major QTLs for gas exchange and stomatal traits under control and saline conditions indicates a complex relationship between salinity and leaf gas exchange due to the fact that these complex quantitative traits are under the control of multiple genes.

Genotypic Differences in Morphological, Physiological and Agronomic Traits in Wheat (Triticum aestivum L.) in Response to Drought.

Drought is one of the main environmental factors affecting crop growth, and breeding drought-tolerant cultivars is one of the most economic and effective ways of increasing yields and ensuring sustainable agricultural production under drought stress. To facilitate the breeding of drought-tolerant wheat, this study was conducted to evaluate genotypic differences in the drought tolerance of 334 wheat genotypes collected from China and Australia with the aim of screening for drought-tolerant and -sensitive genotypes and to elucidate the corresponding physiological mechanisms. A hydroponic-air experiment (roots exposed to air for 7 h/d and continued for 6 d) showed significant genotypic differences in shoot and root dry weights among the genotypes. The relative shoot and root dry weights, expressed as the percentage of the control, showed a normal distribution, with variation ranges of 20.2-79.7% and 32.8-135.2%, respectively. The coefficients of variation were in the range of 18.2-22.7%, and the diversity index was between 5.71 and 5.73, indicating a rich genetic diversity among the wheat genotypes for drought tolerance. Using phenotypic differences in relative dry weights in responses to drought stress, 20 of each of the most drought-tolerant and drought-sensitive genotypes were selected; these were further evaluated in pot experiments (watering withheld until the soil moisture content reached four percent). The results showed that the trends in drought tolerance were consistent with the hydroponic-air experiment, with genotypes W147 and W235 being the most drought-tolerant and W201 and W282 the most sensitive. Significant genotypic differences in water use efficiency in response to drought were observed in the pot experiment, with the drought-tolerant genotypes being markedly higher and the two sensitive genotypes being no different from the control. A marked increase in bound water content in the drought stress plants was observed in the two drought-tolerant genotypes, while a decrease occurred in the free water. The reductions in photochemical efficiencies of PSII, transpiration rates, net photosynthesis rates, chlorophyll contents and stomatal conduction in the drought-sensitive genotypes W201 and W282 under drought stress were higher than the two tolerant genotypes. This study provides a theoretical guide and germplasm for the further genetic improvement of drought tolerance in wheat.

The conserved role of miR-2 and novel miR-109 in the increase in fecundity of Diaphorina citri induced by symbiotic bacteria and pathogenic fungi.

Infection with pathogens can increase the fecundity and other fitness-related traits of insect vectors for their own advantage. Our previous research has reported the pivotal role of DcKr-h1 in the fecundity improvement of Diaphorina citri induced by the bacterium, "Candidatus Liberibacter asiaticus" (CLas), and the fungus, Cordyceps fumosorosea (Cf). However, the posttranscriptional regulation of this process remains poorly understood. Given the significance of miRNAs in gene regulation, we delved into their roles in shaping phenotypes and their underlying molecular mechanisms. Our results indicated that two miRNAs, miR-2 and novel-miR-109, jointly inhibited DcKr-h1 expression by binding to its 3' untranslated region (UTR). In the D. citri-CLas interaction, the expression levels of miR-2 and novel-miR-109 in the ovaries of CLas-positive psyllids were lower compared to CLas-negative individuals. Overexpression of miR-2 or novel-miR-109 significantly decreased fecundity and CLas titer in ovaries and caused reproductive defects reminiscent of DcKr-h1 knockdown. Similarly, in the D. citri-Cf interaction, the levels of miR-2 and novel-miR-109 markedly decreased in the ovaries. Upregulation of miR-2 or novel-miR-109 also resulted in reduced fecundity and ovary defects similar to those caused by DcKr-h1 silencing. Moreover, feeding antagomir-2 or antagomir-109 partially rescued the defective phenotypes caused by DcKr-h1 silencing in both model systems, and miR-2 and novel-miR-109 were repressed by juvenile hormone (JH) and regulated the genes associated with egg development. This study shows a conserved regulatory mechanism, whereby JH suppresses the expression of miR-2 and novel-miR-109 which, together with JH-induced transcription of DcKr-h1, increases female fecundity induced by both symbiotic bacteria and pathogenic fungi. IMPORTANCE: Infection with pathogens can increase the fecundity and other fitness-related traits of insect vectors for their own advantage. Our previous research has reported that DcKr-h1 plays a critical role in the increase in fecundity of Diaphorina citri induced by the bacterium, "Candidatus Liberibacter asiaticus" (CLas) and the fungus, Cordyceps fumosorosea (Cf). However, the posttranscriptional regulation of this process remains poorly understood. Given the significance of miRNAs in gene regulation, we delved into their roles in shaping phenotypes and their underlying molecular mechanisms. Our results indicated that two miRNAs, miR-2 and novel-miR-109, jointly inhibited DcKr-h1 expression by binding to its 3' untranslated region (UTR). In both D. citri-CLas and D. citri-Cf interactions, the increased juvenile hormone (JH) titer and reduced abundance of miR-2 and novel-miR-109 ensure high levels of DcKr-h1 expression, consequently stimulating ovarian development and enhancing fecundity. These observations provide evidence that miR-2 and miR-109 are crucial players in the JH-dependent increase in fecundity in psyllids induced by infection with different pathogens.

Adipokinetic hormone signaling mediates the enhanced fecundity of Diaphorina citri infected by 'Candidatus Liberibacter asiaticus'.

Diaphorina citri serves as the primary vector for 'Candidatus Liberibacter asiaticus (CLas),' the bacterium associated with the severe Asian form of huanglongbing. CLas-positive D. citri are more fecund than their CLas-negative counterparts and require extra energy expenditure. Therefore, understanding the molecular mechanisms linking metabolism and reproduction is of particular importance. In this study, we found adipokinetic hormone (DcAKH) and its receptor (DcAKHR) were essential for increasing lipid metabolism and fecundity in response to CLas infection in D. citri. Knockdown of DcAKH and DcAKHR not only resulted in the accumulation of triacylglycerol and a decline of glycogen, but also significantly decreased fecundity and CLas titer in ovaries. Combined in vivo and in vitro experiments showed that miR-34 suppresses DcAKHR expression by binding to its 3' untranslated region, whilst overexpression of miR-34 resulted in a decline of DcAKHR expression and CLas titer in ovaries and caused defects that mimicked DcAKHR knockdown phenotypes. Additionally, knockdown of DcAKH and DcAKHR significantly reduced juvenile hormone (JH) titer and JH signaling pathway genes in fat bodies and ovaries, including the JH receptor, methoprene-tolerant (DcMet), and the transcription factor, Krüppel homolog 1 (DcKr-h1), that acts downstream of it, as well as the egg development related genes vitellogenin 1-like (DcVg-1-like), vitellogenin A1-like (DcVg-A1-like) and the vitellogenin receptor (DcVgR). As a result, CLas hijacks AKH/AKHR-miR-34-JH signaling to improve D. citri lipid metabolism and fecundity, while simultaneously increasing the replication of CLas, suggesting a mutualistic interaction between CLas and D. citri ovaries.

Insights into the ameliorative effect of ZnONPs on arsenic toxicity in soybean mediated by hormonal regulation, transporter modulation, and stress responsive genes.

Arsenic (As) contamination of agricultural soils poses a serious threat to crop productivity and food safety. Zinc oxide nanoparticles (ZnONPs) have emerged as a potential amendment for mitigating the adverse effects of As stress in plants. Soybean crop is mostly grown on marginalized land and is known for high accumulation of As in roots than others tissue. Therefore, this study aimed to elucidate the underlying mechanisms of ZnONPs in ameliorating arsenic toxicity in soybean. Our results demonstrated that ZnOB significantly improved the growth performance of soybean plants exposed to arsenic. This improvement was accompanied by a decrease (55%) in As accumulation and an increase in photosynthetic efficiency. ZnOB also modulated hormonal balance, with a significant increase in auxin (149%), abscisic acid (118%), gibberellin (160%) and jasmonic acid content (92%) under As(V) stress assuring that ZnONPs may enhance root growth and development by regulating hormonal signaling. We then conducted a transcriptomic analysis to understand further the molecular mechanisms underlying the NPs-induced As(V) tolerance. This analysis identified genes differentially expressed in response to ZnONPs supplementation, including those involved in auxin, abscisic acid, gibberellin, and jasmonic acid biosynthesis and signaling pathways. Weighted gene co-expression network analysis identified 37 potential hub genes encoding stress responders, transporters, and signal transducers across six modules potentially facilitated the efflux of arsenic from cells, reducing its toxicity. Our study provides valuable insights into the molecular mechanisms associated with metalloid tolerance in soybean and offers new avenues for improving As tolerance in contaminated soils.

Luxury Zinc Supply Prevents the Depression of Grain Nitrogen Concentrations in Rice (Oryza sativa L.) Typically Induced by Elevated CO2.

Rice (Oryza sativa L.) has inherently low concentrations of nitrogen (N) and zinc (Zn), and those concentrations are falling as the atmospheric concentration of carbon dioxide ([CO2]) increases, threatening the quality of human diets. We investigated the effect of two levels of Zn supply (marginal and luxury), on Zn and N concentrations in whole grain of two indica rice cvv. Differing in Zn-efficiency (IR26 (inefficient) and IR36 (efficient)), grown in sand culture at ambient (400 µL CO2 L-1 (a[CO2])) and elevated (700 µL CO2 L-1 (e[CO2])) CO2 concentrations. For both cvv., luxury Zn-supply increased vegetative growth, and the foliar and grain Zn concentrations; the increases in grain yield were greater at e[CO2]. The e[CO2] decreased grain Zn concentrations ([Zn]), as is consistently observed in other studies. However, unique to our study, luxury Zn-supply maintained grain N concentrations at e[CO2]. Our data also show that enhanced Zn uptake is the basis of the greater Zn-efficiency of IR36. Lastly, luxury Zn-supply and e[CO2] appreciably decreased the time to panicle emergence and, consequently, to maturity in both cvv. Since Zn-supply can be manipulated by both soil and foliar applications, these findings are potentially important for the quality and quantity of the global rice supply. That is, further investigation of our findings is justified. Key message: Luxury zinc supply maintains grain N concentration at 700 µL CO2 L-1.