Statistics as Part of Scientific Reasoning in Plant Sciences:Overlooked Issues and Recommended Solutions

New research tools for modern life sciences are emerging every few years and being implemented to reveal the underlying mechanisms of scientific questions of interest.However,statistical practice in modern life sciences has barely moved forward and probably has regressed.In many cases,the decisions from biological experiments are predominantly made by using significant P values involving inappropriate statistical analyses and standards,and positive,favored results are preferentially reported (Fanelli,2012;Head et al.,2015;Demidenko,2016;Yong, 2017).

Calmodulins and calmodulin-like proteins-mediated plant organellar calcium signaling networks under abiotic stress

Plant calmodulins(CaMs)and calmodulin-like proteins(CMLs)mediate Ca~(2+)signaling in response to abiotic stresses.Manipulation of this signaling in crops could increase stress tolerance.We review methods for detecting Ca~(2+)signals,regulatory roles of Ca Ms and CMLs,binding targets,and Ca~(2+)networks under abiotic stress in organelles.

Characteristics of long-distance mobile mRNAs from shoot to root in grafted plant species

Thousands of long-distance mobile mRNAs were identified from different grafting systems,based on high-throughput sequencing technology.Moreover,the long-distance delivery of RNAs was proved to involve multiple mechanisms.Here,we analyzed the homology,motif,and tRNA-like structure(TLS)of long-distance mobile mRNAs identified by RNA-seq as well as the RNA-binding protein(RBP)in nine grafting combinations including Arabidopsis thaliana,Vitis vinifera,Cucumis sativus,Citrullus lanatus,Nicotiana benthamiana,Malus domestica,Pyrus spp.,Glycine max and Phaseolus vulgaris.Although several mRNAs were found to be shared in herbaceous,woody,and related species,the vast majority of long-distance mobile mRNAs were species-specific.Four non-specific movement-related motifs were identified,while the TLS was not necessary for mRNA long distance mobility.In addition,we found that RBPs were conserved among herbaceous and woody plants as well as related species.This paper reports a further in-depth analysis of the endogenous mechanisms by which the species-specific transportable m RNAs were selected by bioinformatics,in order to provide insights for future research on long-distance mobile mRNAs.

SPATULA as a Versatile Tool in Plant:The Progress and Perspectives of SPATULA(SPT)Transcriptional Factor

With the rapid development of modern molecular biology and bioinformatics,many studies have proved that transcription factors play an important role in regulating the growth and development of plants.SPATULA(SPT)belongs to the bHLH transcription family and participates in many processes of regulating plant growth and development.This review systemically summarizes the multiple roles of SPT in plant growth,development,and stress response,including seed germination,flowering,leaf size,carpel development,and root elongation,which is helpful for us to better understand the functions of SPT.

Strategies and prospects for melatonin to alleviate abiotic stress in horticultural plants

Melatonin is a conserved pleiotropic molecule in animals and plants.Melatonin is involved in many development processes and stress responses;thus,exploring its function in plants,particularly in horticultural plants,has become a rapidly developing field.Many studies have revealed that phytomelatonin acts as a plant biostimulant and increase its tolerance to various abiotic stressors,including extreme temperature,drought,osmotic disturbance,heavy metals,and ultraviolet(UV).Melatonin appears to have roles in the scavenging of reactive oxygen species(ROS)and other free radicals,affecting the primary and secondary metabolism of plants,regulating the transcripts of stress-related enzymes and transcription factors,and crosstalk with other hormones under different environmental conditions.This pleiotropy makes phytomelatonin an attractive regulator to improve resistance to abiotic stress in plants.The recent discovery of the potential phytomelatonin receptor CAND2/PMTR1 and the proposition of putative models related to the phytomelatonin signaling pathways makes phytomelatonin a new plant hormone.Based on relevant studies from our laboratory,this review summarizes the phytomelatonin biosynthetic and metabolic pathways in plants and the latest research progress on phytomelatonin in abiotic stress of horticultural plants.This study will provide a reference for elucidating the regulatory mechanism of phytomelatonin affecting the resistance to abiotic stress in plants.

Management of Strawberry Grey Mold Disease Using Biocontrol Agents and Plant Extracts

Strawberry (Fragaria × ananassa Duch.) is a significant global soft fruit crop, prized for its nutrient content and pleasant flavor. However, diseases, particularly grey mold caused by Botrytis cinerea Pers. Fr. poses major constraints to strawberry production and productivity. Grey mold severely impacts fruit quality and quantity, diminishing market value. This study evaluated five B. cinerea isolates from various locations in the Ri-Bhoi district of Meghalaya. All isolates were pathogenic, with isolate SGM 2 identified as highly virulent. Host range studies showed the pathogen-producing symptoms in the fava bean pods, marigold, gerbera, and chrysanthemum flowers and in the fava bean, gerbera, and lettuce leaves. In vitro tests revealed that neem extract (15% w/v) achieved the highest mycelial growth inhibition at 76.66%, while black turmeric extract (5% w/v) had the lowest inhibition at 9.62%. Dual culture methods with bio-control agents indicated that Bacillus subtilis recorded the highest mean inhibition at 77.03%, while Pseudomonas fluorescens had the lowest at 20.36% against the two virulent isolates. Pot evaluations demonstrated that B. subtilis resulted in the lowest percent disease index at 20.59%, followed by neem extract at 23.31%, with the highest disease index in the control group at 42.51%. Additionally, B. subtilis significantly improved plant growth, yielding an average of 0.32 kg compared to 0.14 kg in the control. The promising results of B. subtilis and neem leaf extract from this study suggest their potential for eco-friendly managing grey mold in strawberries under field conditions.

Wild soybean(Glycine soja)transcription factor GsWRKY40 plays positive roles in plant salt tolerance

Wild soybean(Glycine soja),a relative of cultivated soybean,shows high adaptability to adverse environmental conditions.We identified and characterized a wild soybean transcription factor gene,GsWRKY40,that promotes plant salt stress.GsWRKY40 was highly expressed in wild soybean roots and was up-regulated by salt treatment.GsWRKY40 was localized in nucleus and demonstrated DNA-binding activities but without transcriptional activation.Mutation and overexpression of GsWRKY40 altered salt tolerance of Arabidopsis plants.To understand the molecular mechanism of GsWRKY40 in regulating plant salt resistance,we screened a cDNA library and identified a GsWRKY40 interacting protein GsbHLH92 by using yeast two-hybrid approach.The physical interaction of GsWRKY40 and GsbHLH92 was confirmed by co-immunoprecipitation(co-IP),GST pull-down,and bimolecular fluorescence complementation(BiFC)techniques.Intriguingly,co-overexpression of GsWRKY40 and GsbHLH92 resulted in higher salt tolerance and lower ROS levels than overexpression of GsWRKY40 or GsbHLH92 in composite soybean plants,suggesting that GsWRKY40 and GsbHLH92 may synergistically regulate plant salt resistance through inhibiting ROS production.qRT-PCR data indicated that the expression level of GmSPOD1 gene encoding peroxidase was cooperatively regulated by GsWRKY40 and GsbHLH92,which was confirmed by using a dual luciferase report system and yeast one-hybrid experiment.Our study reveals a pathway that GsWRKY40 and GsbHLH92 collaboratively up-regulate plant salt resistance through impeding GmSPOD1 expression and reducing ROS levels,providing a novel perspective on the regulatory mechanisms underlying plant tolerance to abiotic stresses.

Ca^(2+)-homeostasis Differs Between Plant Species with Different Cold-tolerance at 4 ℃ Chilling

A comparative study was carried out on the EM_cytochemical localization of calcium and Ca 2+ _ATPase activity in the suspension_cultured cells between the chilling_sensitive maize ( Zea mays L. cv. Black Mexican Sweet) and chilling_insensitive Trititrigia ( Triticum sect. Trititrigia mackey) at 4 ℃ chilling. When maize and Tyititrigia cells were cultured at 26 ℃, electron microscopic observations revealed that the electron_dense calcium antimonate deposits, an indication of the calcium localization, were localized mainly in the vacuoles, and few was found in the cytosol and nuclei. The electron_dense cerium phosphate deposits, an indication of Ca 2+ _ATPase activity, were abundantly distributed on the plasma membrane (PM). When the cells from both species were cultured at 4 ℃ for 1 and 3 h, an elevation of Ca 2+ level in the cytosol and nuclei was observed, whereas the cerium phosphate deposits on the PM showed no quantitative difference from those of the 26 ℃_cultured cells, indicating that the enzymatic activities were not altered during these chilling periods. However, there was a distinct difference in the dynamics of the Ca 2+ distribution and the PM Ca 2+ _ATPase activity between maize and Trititrigia when chilled at 4 ℃ for 12, 24 and 72 h. In maize cells, a large number of Ca 2+ deposits still existed in the cytosol and nuclei, and the PM Ca 2+ _ATPase became less and less active, and even inactive at all. In Trititrigia cells, the increased cytosolic and nuclear Ca 2+ ions decreased after 12 h chilling. By chilling up to 24 and 72 h, the intracellular Ca 2+ concentration had been restored to a similar low level as those of the warm temperature_cultured cells, while the activity of the PM Ca 2+ _ATPase maintained high. The transient cytosolic and nuclear Ca 2+ increase and the activities of PM Ca 2+ _ATPase during chilling are discussed in relation to plant cold hardiness.

Effects of Plant Density on Yield and Canopy Micro Environment in Hybrid Cotton

A rational plant population is an important attribute to high yield of cotton, because it can provide a beneficial micro environment within the canopy for plant growth and development as well as yield formation. A 2-yr field experiment was conducted to determine the optimal plant density based on cotton yield in relation to the canopy micro environment （canopy temperature, relative humidity and light transmittance）. Six plant densities （1.2-5.7 plants m^-2） were arranged with a completely randomized block design. The highest cotton yield （1 507 kg ha^-1） was obtained at 3.0 plants m^-2 due to more bolls per unit ground area （79 bolls m2）, while the lowest yield （1 091 kg ha1） was obtained at 1.2 plants m^-2. Under the moderate plant density （3.0 plants m^-2）, there was a lower mean daily temperature （MDT, 27. 1℃） attributing to medium daily minimum temperature （Train, 21.9℃） and the lowest daily maximum temperature （Tmax, 35.8℃）, a moderate mean canopy light transmittance of 0.51, and lower mean daily relative humidity （MRH） of 79.7% from June to October. The results suggest that 3.0 plants m^-2 would be the optimal plant density because it provides a better canopy micro environment.

Plant salt tolerance and Na^+ sensing and transport

Salinity is a global challenge to agricultural production. Understanding Na^+ sensing and transport in plants under salt stress will be of benefit for breeding robustly salt-tolerant crop species. In this review, first, possible salt stress sensor candidates and the root meristem zone as a tissue harboring salt stress-sensing components are proposed. Then,the importance of Na^+ exclusion and vacuolar Na^+ sequestration in plant overall salt tolerance is highlighted. Other Na^+ regulation processes, including xylem Na^+ loading and unloading, phloem Na^+ recirculation, and Na^+ secretion, are discussed and summarized.Along with a summary of Na^+ transporters and channels, the molecular regulation of Na^+ transporters and channels in response to salt stress is discussed. Finally, some largely neglected issues in plant salt stress tolerance, including Na^+ concentration in cytosol and the role of Na^+ as a nutrient, are reviewed and discussed.

Fertilizer ^(15)N Accumulation, Recovery and Distribution in Cotton Plant as Affected by N Rate and Split

N fertilization of 300 kg N ha-1 is normally applied to cotton crops in three splits： pre-plant application （PPA, 30%）, first bloom application （FBA, 40%） and peak bloom application （PBA, 30%） in the Yangtze River Valley China. However, low fertilizer N plant recovery （NPR） （30-35%） causes problems such as cotton yield stagnation even in higher N rate, low profit margin of cotton production and fertilizer release to the environment. Therefore, it is questioned： Are these three splits the same significance to cotton N uptake and distribution？ An outdoor pot trial was conducted with five N rates and 15 N labeled urea to determine the recovery and distribution of 15N from different splits in cotton （Gossypium hirsutum L. cv. Huazamian H318） plant. The results showed that, cotton plant absorbed fertilizer 15N during the whole growing period, the majority during flowering for 18-20 d regardless of N rates （150-600 kg ha-1）. Fertilizer 15N proportion to the total N accumulated in cotton plant increased with N rates, and it was the highest in reproductive organs （88% averaged across N rates） among all the plant parts. FBA had the highest NPR （70%）, the lowest fertilizer N lose （FNL, 19%）, and the highest contribution to the fertilizer 15N proportion to the total N （46%） in cotton plant, whereas PPA had the reverse effect. It suggests that FBA should be the most important split for N absorption and yield formation comparatively and allocating more fertilizer N for late application from PPA should improve the benefit from fertilizer.

The Influence of Transgenic cry1Ab/cry1Ac,cry1C and cry2A Rice on Non-Target Planthoppers and Their Main Predators Under Field Conditions

Transgenic Bt rice has been shown to be an effective means of controlling Lepidoptera pests of rice. However, the potential roles of transgenic rice on planthoppers and their predators need to be investigated before its commercialization. Population density, species dominance and population dynamics are important parameters of arthropods populations in field. So the impacts of three transgenic Bt rice strains expressing crylAb/crylAc, crylC and cry2A on population density, species dominance and population dynamics of three species of planthoppers （Nilaparvata lugens, Sogatella furcifera and Laodelphax striatellus） and their three main predators （ Cyrtorhinus lividipennis, Pirata subpiraticus and Theridium octomaculatum） were evaluated at three sites in Hubei Province, China, in the current study. The results showed that among three species of planthoppers, both in transgenic and non-transgenic rice field, the predominant species ofplanthoppers within phytophagous guild was S. furcifera at any site either growing season （46-50%）. Significantly higher population density ofN. lugens was observed in T2A-1 field relative to Minghui 63 field at Wuxue in 2010. The species dominance of predator, P. subpiraticus, in TT51 field was significantly higher than that in T 1 C-19 and T2A-1 fields in 2009 at Xiaogan site. Sampling date significantly influenced six arthropods except for P. subpiraticus in 2010. The interaction between rice strain^sampling date had no significant adverse effects on the population dynamics of three species of planthoppers and their predators, except for several individual species in 2009. The interaction among rice strain^sampling date^sampling site also had no significant effect on six arthropods except for S. furcifera in 2009. The results indicated that transgenic Bt rice expressing crylAb/crylAc, cry2A and crylC had no significant adverse effects on the population dynamics of three planthoppers and their predators in most investigated data and sampling site.

The role of calcium sensor-interacting protein kinases in plant adaptation to potassium-deficiency: new answers to old questions

Potassium （K^＋） is an essential macronutrient for all living organisms and large amounts are required for plant growth and development. In many regions of Asia K＋-fertilization has been neglected and soils have become K＋-depleted. K＋- deficiency in the field diminishes not only crop production but also leads to environmental problems due to inefficient usage and leaching of nitrate. Consequences of K^＋-deficiency on crop production range from decreased biomass, nutritional quality and taste of the crops to inferior harvest and storage properties, as well as increased susceptibility to disease. Effects of K^＋-deficiency on plant physiology include decreased photosynthetic rate, impaired tissue allocation of sugars and amino acids, decreased protein synthesis.

Twenty years of Chinese vascular plant novelties,2000 through 2019

From 2000 to 2019,11,895 new names or new additions to the Chinese vascular flora were proposed by 4226 individuals(4086 articles and 140 books),as documented in the Chinese Plant Names Index(CPNI).During those 20 years,4407 new taxa of vascular plants were described from China,including 7 new families,132 new genera,3543 new species,68 new subspecies,497 new varieties and 160 new forms.Additionally,3562 new combinations and names at new rank and 306 new replacement names were also proposed.Among these various new names were 150 invalid names and 108 illegitimate names,including some that have not been resolved.Six hundred and forty three vascular plants were reported as new to China,while 2349 names were reduced to synonyms of 1406 taxa.The data show that the Chinese flora increased in size at the rate of about 200 taxa annually during those years.Despite the increased attention given to biodiversity in recent years,the evidence indicates that a large number of species in China have yet to be discovered.Further basic investigation of the Chinese flora is needed.Additionally,in the past two decades only 8.5%of the newly published species have been based on molecular evidence,but in the past five years such data have increased significantly,reaching about 20%.Molecular data will undoubtedly become increasingly significant in the discovery of new species in the coming years.Yunnan,Guangxi,Sichuan,Xizang and Taiwan were important sources of new discoveries,with more than 3300 new taxa and records from these five provinces.By area,Taiwan and Hainan,two islands in southern China,have the highest density of newly discovered species.Regional plant surveys are still needed,especially in areas in the southwest and on the southern islands.

Multi-Location Investigation of Optimum Planting Density and Boll Distribution of High-Yielding Cotton (G.hirsutum L.) in Hubei Province,China

Cotton yield per unit ground area has stagnated for a dozen years in Hubei Province, China, although a series of new high- yielding varieties have been commercialized. A multi-location investigation was carried out in 2008 and 2009 in 13 counties to determine if increased planting population density （PPD） would break the stagnant yield. The results showed that significant differences among the fields existed in theoretical yield, PPD, and bolls per square meter （BPM）. The lowest yield of 1 641.1 kg ha-I was resulted from the lowest PPD of 1.7 plants m-2 and the lowest BPM of 71.8 bolls m-2, while the highest yield of 2 779.7 kg ha-~ was resulted from the highest PPD of 2.5 plants m-2, and the highest BPM of 129.4 bolls m-z. Plant mapping revealed that boll retention rate （BRR） was maintained over 30 or 40% for the first 17-18 fruiting branches （FBs） and decreased dramatically thereafter, rotten boll rate （RBR） decreased, but open boll rate （OBR） rose first and dropped later with rising FB from the bottom to the top. But BRR, RBR, and OBR were all dropped with the fruiting positions （FPs） extending outwards. The optimum range of plant density would be 2-3 plants m-2 and the proper individual plant structure would be 16-19 FBs with 5-7 FPs for cotton production in Hubei Province.

Metabolic analyses reveal different mechanisms of leaf color change in two purple-leaf tea plant (Camellia sinensis L.) cultivars

Purple-leaf tea plants,as anthocyanin-rich cultivars,are valuable materials for manufacturing teas with unique colors or flavors.In this study,a new purple-leaf cultivar“Zixin”(“ZX”)was examined,and its biochemical variation and mechanism of leaf color change were elucidated.The metabolomes of leaves of“ZX”at completely purple,intermediately purple,and completely green stages were analyzed using ultra-performance liquid chromatography quadrupole time of flight mass spectrometry(UPLC-QTOF-MS).Metabolites in the flavonoid biosynthetic pathway remained at high levels in purple leaves,whereas intermediates of porphyrin and chlorophyll metabolism and carotenoid biosynthesis exhibited high levels in green leaves.In addition,fatty acid metabolism was more active in purple leaves,and steroids maintained higher levels in green leaves.Saponin,alcohol,organic acid,and terpenoid-related metabolites also changed significantly during the leaf color change process.Furthermore,the substance changes between“ZX”and“Zijuan”(a thoroughly studied purple-leaf cultivar)were also compared.The leaf color change in“Zijuan”was mainly caused by a decrease in flavonoids/anthocyanins.However,a decrease in flavonoids/anthocyanins,an enhancement of porphyrin,chlorophyll metabolism,carotenoid biosynthesis,and steroids,and a decrease in fatty acids synergistically caused the leaf color change in“ZX”.These findings will facilitate comprehensive research on the regulatory mechanisms of leaf color change in purple-leaf tea cultivars.

Genetic variations in plant architecture traits in cotton(Gossypium hirsutum) revealed by a genomewide association study

Plant architecture traits influence crop yield. An understanding of the genetic basis of cotton plant architecture traits is beneficial for identifying favorable alleles and functional genes and breeding elite cultivars. We collected 121 cotton accessions including 100 brownfiber and 21 white-fiber accessions, genotyped them by whole-genome resequencing, and phenotyped them in multiple environments. This genome-wide association study(GWAS)identified 11 quantitative trait loci(QTL) for two plant architecture traits: plant height and fruit spur branch number. Negative-effect alleles were enriched in the elite cultivars. Based on these QTL, gene annotation information, and published QTL, candidate genes and natural genetic variations in four QTL were identified. Ghir_D02 G017510 and Ghir_D02 G017600 were identified as candidate genes for qD02-FSBN-1, and a premature start codon gain variation was found in Ghir_D02 G017510. Ghir_A12 G026570, the candidate gene of qA12-FSBN-2, belongs to the pectin lyase-like superfamily, and a significantly associated SNP, A12_105366045(T/C), in this gene represents an amino acid change. The QTL, candidate genes, and associated natural variations in this study are expected to lay a foundation for studying functional genes and developing breeding programs for desirable architecture in brown-fiber cotton.

Maintenance of mesophyll potassium and regulation of plasma membrane H^+-ATPase are associated with physiological responses of tea plants to drought and subsequent rehydration

Drought stress is one of the main factors limiting yield in tea plants. The plant cell's ability to preserve K^+homeostasis is an important strategy for coping with drought stress. Plasma membrane H^+-ATPase in the mesophyll cell is important for maintaining membrane potential to regulate K^+transmembrane transport. However, no research to date has investigated the possible relationship between plasma membrane H^+-ATPase and mesophyll K^+retention in tea plants under drought and subsequent rehydration conditions. In our experiment, drought stress inhibited plasma membrane H^+-ATPase activities and induced net H^+influx, leading to membrane potential depolarization and inducing a massive K^+efflux in tea plant mesophyll cells. Subsequent rehydration increased plasma membrane H^+-ATPase activity and induced net H^+efflux, leading to membrane potential hyperpolarization and thus lowering K^+loss. A first downregulated and then upregulated plasma membrane H^+-ATPase protein expression level was also observed under drought and subsequent rehydration treatment, a finding in agreement with the change of measured plasma membrane H^+-ATPase activities. Taken together, our results suggest that maintenance of mesophyll K^+in tea plants under drought and rehydration is associated with regulation of plasma membrane H^+-ATPase activity.

A RHOse by any other name:a comparative analysis of animal and plant Rho GTPases

Rho GTPases are molecular switches that act as key regulators of a many cellular processes, including cell movement, morphogenesis, host defense, cell division and gene expression. Rho GTPases are found in all eukaryotic kingdoms. Plants lack clear homologs to conventional Rho GTPases found in yeast and animals; instead, they have over time developed a unique subfamily, ROPs, also known as RAC. The origin of ROP-like proteins appears to precede the appearance of land plants. This review aims to discuss the evolution of ROP/RAC and to compare plant ROP and animal Rho GTPases, focusing on similarities and differences in regulation of the GTPases and their downstream effectors.

Comparison of yield performance between direct-seeded and transplanted double-season rice using ultrashort-duration varieties in central China

Labor scarcity requires double-season rice to be planted by direct seeding as an alternative to transplanting. Only ultrashort-duration varieties can be used in direct-seeded, double-season rice(DSD) in central China where thermal time is limited. Whether ultrashort-duration varieties grown in DSD can be as productive and efficient in nitrogen(N) use as transplanted double-season rice(TPD) remains unclear. Field experiments were conducted in Hubei province, central China with two establishment methods(DSD,TPD) and three N rates in the early and late seasons of 2017 and 2018. Nitrogen treatments included zero-N control(N0), total N rate of 60 kg N ha;with equal splits at basal, midtillering, and panicle initiation(N1), and weekly N application at 15 kg ha;from seeding/transplanting to heading(N2). Both early-and late-season rice under DSD matured within 95 days, on average 9 days shorter than rice under TPD. The grain yield of DSD was comparable to or higher than that of TDP in both seasons, although the daily yield was significantly higher under DSD than under TDP. Before heading, DSD had higher leaf area,stem number, intercepted radiation, and radiation use efficiency than TPD, which compensated for the negative effect of short growth duration on biomass production. Total dry weight and harvest index under DSD were comparable to or higher than those under TDP. In general, the recovery efficiency of fertilizer-N under DSD was higher than that under TPD, but the reverse was true for physiological N use efficiency. Thus, there was no significant difference in agronomic N use efficiency between DSD and TPD. These results suggested that DSD with ultrashort-duration varieties is a promising alternative to TPD in central China for maintaining high grain yield and N fertilizer use efficiency with less labor input.