<Emphasis Type="Italic">Diaporthe</Emphasis> from walnut tree (<Emphasis Type="Italic">Juglans regia</Emphasis>) in China,with insight of the <Emphasis Type="Italic">Diaporthe eres</Emphasis> complex

Species of Diaporthe are important plant pathogenic fungi that commonly occur on a wide range of hosts. They are relatively difficult to identify due to their extreme similarity in morphology and confusing multigene phylogeny, especially in the Diaporthe eres complex. In the present study, isolates were collected from diseased branches of Juglans regia in China. Most strains were clustered into the D. eres species complex based on the combined internal transcribed spacer (ITS) region, partial calmodulin (CAL), histone H3 (HIS), translation elongation factor 1-alpha (TEF1-α) and beta-tubulin (TUB) genes. To focus on this complex, CAL, TEF1-α and TUB were selected in further phylogenetic analyses that showed a better topology compared with combined five-gene phylogeny. Results revealed that all strains which clustered in the Diaporthe eres complex from Juglans regia in China were Diaporthe eres. Results suggested a revised species criterion in the Diaporthe eres complex. The current study uncovered a new species here described as Diaporthe. tibetensis.     

Genetic diversity and population structure of common walnut (<Emphasis Type="Italic">Juglans regia</Emphasis>) in China based on EST-SSRs and the nuclear gene phenylalanine ammonia-lyase (<Emphasis Type="Italic">PAL</Emphasis>)

Common walnut (Juglans regia L.) is an economically important temperate tree species valued for both its nut and wood. We investigated the genetic diversity and population structure of J. regia germplasm from 13 locations in China using 10 markers derived from expressed sequences (EST-SSR) and sequence polymorphisms within the phenylalanine ammonia-lyase (PAL) gene. Analysis of the population genetic structure based on EST-SSRs showed distinct populations in northern versus southern China that were not reflected in the spatial distribution of PAL haplotypes. High levels of population differentiation were probably the result of reproductive isolation and in southern China, hybridization with Juglans sigillata. The results indicate the possible presence of distinct evolutionary lineages of J. regia in the Qinling and Daba Mountains of China and in Yunnan province that may require ecological management if they are to be retained as in situ resources.     

Genetic diversity and population structure in the narrow endemic Chinese walnut <Emphasis Type="Italic">Juglans hopeiensis</Emphasis> Hu: implications for conservation

The conservation of narrow endemic species relies on accurate information regarding their population structure. Juglans hopeiensis Hu (Ma walnut), found only in Hebei province, Beijing, and Tianjin, China, is a threatened tree species valued commercially for its nut and wood. Sequences of two maternally inherited mitochondrial markers and two maternally inherited chloroplast intergenic spacers, three nuclear DNA sequences, and allele sizes from 11 microsatellites were obtained from 108 individuals of J. hopeiensis, Juglans regia, and Juglans mandshurica. Haplotype networks were constructed using NETWORK. Genetic diversity, population differentiation, and analysis of molecular variance (AMOVA) were used to determine genetic structure. MEGA was used to construct phylogenetic trees. Genetic diversity of J. hopeiensis was moderate based on nuclear DNA, but low based on uniparentally inherited mitochondrial DNA and chloroplast DNA. Haplotype networks showed that J. hopeiensis haplotypes were different than haplotypes found in J. regia and J. mandshurica. Allelic variants in nuclear genes that were shared among J. hopeiensis populations were not found in J. regia or J. mandshurica. Sampled populations of J. hopeiensis showed clear genetic structure. The maximum parsimony (MP) tree showed J. hopeiensis to be distinct from J. mandshurica but threatened by hybridization with J. regia and J. mandshurica. J. hopeiensis populations are strongly differentiated from sympatric Juglans species, but they are threatened by small population sizes and hybridization.     

Over-expression of <Emphasis Type="Italic">JrsHSP17.3</Emphasis> gene from <Emphasis Type="Italic">Juglans regia</Emphasis> confer the tolerance to abnormal temperature and NaCl stresses

Small heat shock proteins (sHSPs) are an HSP subgroup and involved in environmental stress response. In the current study, to understand the role of sHSP protein in a widely distribution nut woody tree, a sHSP gene was cloned from Juglans regia (JrsHSP17.3, GeneBank No.: KT277704). Compared with control condition, the expression of JrsHSP17.3 was induced to 58.1-fold (6 h) in the roots, 86.8-fold in the stems (9 h), 50.9-fold in the leaves (6 h) under 10°C; and was up-regulated to 2.9- ～ 79.9-fold response to 40°C for 3～9 h; meanwhile, it was transcribed to 5.9 - ～39.7-fold under 9 h NaCl treatment, suggesting the potential role of JrsHSP17.3 to cold, heat and NaCl stimulus. Further, JrsHSP17.3 transgenic yeasts showed improved tolerance to freezing, heat and salt stresses compared with control yeast. JrsHSP17.3 was transient over-expressed in J. regia leaves. The leaves non-transgenic (NT) and vector prokII transgenic (empty, PT) were used as control. The expression of JrsHSP17.3 was 81.6-, 125.4-, and 54.2-fold of the control lines under normal conditions, indicating the success over-expression of JrsHSP17.3. Cell damage staining and physiological index determination showed that JrsHSP17.3 transformed lines, NT and PT displayed no obvious difference under control conditions, however, after treated with 16°C, 40°C and NaCl, JrsHSP17.3 transformed lines displayed weaker cell damage, lower level of electrolyte leakages (EL) rate, malondialdehyde (MDA) and H₂O₂ content, and higher activities of catalase (CAT), glutathione transferase (GST), superoxide dismutase (SOD) and peroxidase (POD) as well as more accumulation of proline than NT and PT. Meanwhile, NT and PT were similar and showed no significant difference under all conditions. All of these results indicated that JrsHSP17.3 can improve plant tolerance to abnormal temperatures and NaCl stresses, it represents a potential candidate gene for molecular breeding to enhance stress tolerance in plants.     

In planta characterization of a tau class glutathione S-transferase gene from <Emphasis Type="Italic">Juglans regia</Emphasis> (<Emphasis Type="Italic">JrGSTTau1</Emphasis>) involved in chilling tolerance

Key message

JrGSTTau1 is an important candidate gene for plant chilling tolerance regulation.

Abstract

A tau subfamily glutathione S-transferase (GST) gene from Juglans regia (JrGSTTau1, GeneBank No.: KT351091) was cloned and functionally characterized. JrGSTTau1 was induced by 16, 12, 10, 8, and 6 °C stresses. The transiently transformed J. regia showed much greater GST, glutathione peroxidase (GPX), superoxide dismutase (SOD), and peroxidase (POD) activities and lower H₂O₂, malondialdehyde (MDA), reactive oxygen species (ROS), and electrolyte leakage (EL) rate than prokII (empty vector control) and RNAi::JrGSTTau1 under cold stress, indicating that JrGSTTau1 may be involved in chilling tolerance. To further confirm the role of JrGSTTau1, JrGSTTau1 was heterologously expressed in tobacco, transgenic Line5, Line9, and Line12 were chosen for analysis. The germinations of WT, Line5, Line9, and Line12 were similar, but the fresh weight, primary root length, and total chlorophyll content (tcc) of the transgenic lines were significantly higher than those of WT under cold stress. When cultivated in soil, the GST and SOD activities of transgenic tobacco were significantly higher than those of WT; however, the MDA and H₂O₂ contents of WT were on average 1.47- and 1.96-fold higher than those of Line5, Line9, and Line12 under 16 °C. The DAB, Evans blue, and PI staining further confirmed these results. Furthermore, the abundances of NtGST, MnSOD, NtMAPK9, and CDPK15 were elevated in 35S::JrGSTTau1 tobacco compared with WT. These results suggested that JrGSTTau1 improves the plant chilling tolerance involved in protecting enzymes, ROS scavenging, and stress-related genes, indicating that JrGSTTau1 is a candidate gene for the potential application in molecular breeding to enhance plant abiotic stress tolerance.

     

The crucial role of roots in increased cadmium-tolerance and Cd-accumulation in the pea mutant <Emphasis Type="Italic">SGECd</Emphasis><Superscript><Emphasis Type="Italic">t</Emphasis></Superscript>

Elucidation of mechanisms underlying plant tolerance to cadmium, a widespread toxic soil pollutant, and accumulation of Cd in plants are urgent tasks. For this purposes, the pea (Pisum sativum L.) mutant SGECd^t (obtained by treatment of the laboratory pea line SGE with ethylmethane sulfonate) was reciprocally grafted with the parental line SGE, and four scion/rootstock combinations were obtained: SGE/SGE, SGECd^t/SGECd^t, SGE/SGECd^t, and SGECd^t/SGE. They were grown in hydroponics in the presence of 1 μM CdCl₂ for 30 d. The SGE and SGECd^t scions on the SGECd^t rootstock had a higher root and shoot biomass and an elevated root and shoot Cd content compared with the grafts having SGE rootstock. Only the grafts with the SGE rootstock showed chlorosis and roots demonstrating symptoms of Cd toxicity. The content of nutrient elements in roots (Fe, K, Mg, Mn, Na, P, and Zn) was higher in the grafts having the SGECd^t rootstock, and three elements, namely Ca, Fe, and Mn, were efficiently transported by the SGECd^t root to the shoot of these grafts. The content of other measured elements (K, Mg, Na, P, and Zn) was similar in the root and shoot in all the grafts. Then, the non-grafted plants were grown in the presence of Cd and subjected to deficit or excess concentrations of Ca, Fe, or Mn. Exclusion of these elements from the nutrient solution retained or increased differences between SGE and SGECd^t in growth response to Cd toxicity, whereas excess of Ca, Fe, or Mn decreased or eliminated such differences. The obtained results assign a principal role of roots to realizing the increased Cd-tolerance and Cdaccumulation in the SGECd^t mutant. Efficient translocation of Ca, Fe, and Mn from roots to shoots appeared to counteract Cd toxicity, although Cd was actively taken up by roots and accumulated in shoots.     

Concentration of some metals in soil and plant organs and their biochemical profiles in <Emphasis Type="Italic">Tulipa luanica</Emphasis>, <Emphasis Type="Italic">T. kosovarica</Emphasis> and <Emphasis Type="Italic">T. albanica</Emphasis> native plant species

The purpose of this study was to determine the concentration of some metals (Al, Cd, Co, Cr, Cu, Fe, Mn, Ni, Pb, Zn, Ca and Mg) in soil of serpentine and limestone sites, their bioaccumulation and impact on some biochemical parameters in T. luanica, T. kosovarica and T. albanica plants. T. kosovarica and T. albanica grows in serpentine soil, while T. luanica grow in limestone soil. The research showed that concentrations of Cd, Co, Cr, Fe, Mn and Ni were significantly higher at serpentine soil sites in comparison with limestone sites, while concentrations of Pb, Cd, Co and Cr in bulbs, leaves and seeds were under the limit of detection. The concentration of Ni in plant samples of T. kosovarica was significantly higher in comparison with its concentration in T. albanica, but it was under the limit of detection in T. luanica. Moreover, concentrations of Al and Fe in leaves of T. kosovarica and T. albanica were higher in comparison with T. luanica. The concentration of Mg was significantly higher in T. kosovarica and T. albanica than in T. luanica. The δ-aminolevulinic acid dehydratase activity, malondialdehyde and glutathione contents in leaves of T. luanica were higher in comparison with T. kosovarica and T. albanica. In addition, the amounts of total chlorophyll and δ-aminolevulinic acid (ALA) in leaves of T. albanica were higher in comparison with T. kosovarica and T. luanica. Our findings show that target organs of metal accumulation in three Tulip species appears to be leaves?>?seeds?>?bulbs, while the biochemical parameters show that limestone sites represent a less stressful habitat for growing these plant species in comparison with serpentine sites.     

Molecular phylogenetic analysis of key <Emphasis Type="Italic">Jatropha</Emphasis> species inferred from nrDNA ITS and chloroplast (<Emphasis Type="Italic">trn</Emphasis>L-F and <Emphasis Type="Italic">rbc</Emphasis>L) sequences

The genus Jatropha (Euphorbiaceae) contains species that are of significant economic and ornamental value. However, Jatropha breeding material is rather limited due to incomplete information regarding phylogenetic relationships among germplasm resources. Phylogenetic analyses were performed based on the internal transcribed spacer of nuclear ribosomal DNA (nrDNA ITS), two chloroplast regions (trnL-F and rbcL), and the combined (ITS+trnL-F+rbcL) dataset among twenty-five specimens representing six key Jatropha species. Phylogenetic relationships of Jatropha were well resolved between subgenus Curcas and subgenus Jatropha, and demonstrated the intermediate position of section Polymorphae among sections of both subgenera. Jatropha curcas and J. integerrima demonstrated a close phylogenetic relationship. The molecular data agreed with the morphological classification that recognized J. multifida and J. podagrica in sec. Peltatae. The distinct intraspecific divergence that occurred in J. curcas could be attributed to restricted gene flow caused by geographical isolation and different ecological conditions. Phylograms produced with trnL-F and rbcL sequence data suggested slow rates of sequence divergence among Jatropha spp., while the ITS gene tree had good resolution suggesting high genetic variation of ITS among Jatropha species.     

Bioinformatics identification of the methylerythritol phosphate pathway associated genes in <Emphasis Type="Italic">Arabidopsis thaliana</Emphasis> with <Emphasis Type="Italic">ceh1</Emphasis> mutant

The methylerythritol phosphate (MEP) pathway for the production of isoprenoids is recently discovered. The current study aimed to identify MEP pathway disorder-related molecular mechanisms and potential genes in Arabidopsis thaliana. Microarray data (GSE61675) obtained from ceh1 mutant plants and corresponding parental lines were retrieved from Gene Expression Omnibus (GEO) database and were applied for differentially expressed genes (DEGs) screening. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis of DEGs were performed. Protein-protein interaction (PPI) network was then constructed and displayed by Cytoscape software. Total 762 DEGs including 620 up-regulated and 142 down-regulated genes were screened. In addition, a great many of DEGs were mainly involved in biosynthesis and metabolism-related pathways, such as stilbenoid, diarylheptanoid, and gingerol biosynthesis, and biosynthesis of terpenoids and steroids. Moreover, a PPI network contained 90 down-regulated genes and 497 up-regulated genes were obtained. Up-regulated DEGs including glutaredoxin (GRX480, cytochrome BC1 synthase (BCS1, syntaxin of plants 121 (SYP121) and A. thaliana MAP kinase 11 (ATMPK11) with higher degree in this network were hub nodes. Pathways including stilbenoid, diarylheptanoid, and gingerol biosynthesis obtained in our study were consistent with previous studies. Importantly, GRX480, BCS1 and ATMPK11 could have close interactions with the MEP pathway and may play important roles in the biosynthesis of isoprenoids.     

<Emphasis Type="Italic">Diplazium</Emphasis> hybrids involving <Emphasis Type="Italic">D. plantaginifolium</Emphasis> and <Emphasis Type="Italic">D</Emphasis>. <Emphasis Type="Italic">ternatum</Emphasis> from Mexico and Central America

Here we evaluate the origins and relationships of Mexican and Central American Diplazium hybrids derived from crosses involving either D. plantaginifolium or D. ternatum. Based on study of live plants and herbarium specimens, we distinguish D. ×verapax from the similar D. riedelianum and present evidence that the former is a sterile hybrid derived from a cross between D. plantaginifolium and D. werckleanum. We also describe new hybrids, D. ×torresianum and D. ×subternatum from Mexico and northern Central America. Both involve D. ternatum as one parent. Diplazium. cristatum is the other putative parent of D. ×torresianum, and D. plantaginifolium is the second parent of D. ×subternatum. We also designate lectotypes for D. cordovense and D. dissimile.