Defense responses of arbuscular mycorrhizal fungus-colonized poplar seedlings against gypsy moth larvae: a multiomics study

Arbuscular mycorrhizal (AM) fungi may help protect plants against herbivores; however, their use for the pest control of woody plants requires further study. Here, we investigated the effect of Glomus mosseae colonization on the interactions between gypsy moth larvae and Populus alba × P. berolinensis seedlings and deciphered the regulatory mechanisms underlying the mycorrhizal-induced resistance in the leaves of mycorrhizal poplar using RNA-seq and nontargeted metabolomics. The resistance assay showed that AM fungus inoculation protected poplar seedlings against gypsy moth larvae, as evidenced by the decreased larval growth and reduced larval survival. A transcriptome analysis revealed that differentially expressed genes (DEGs) were involved in jasmonic acid biosynthesis (lipoxygenase, hydroperoxide dehydratase, and allene oxide cyclase) and signal transduction (jasmonate-ZIM domain and transcription factor MYC2) and identified the genes that were upregulated in mycorrhizal seedlings. Except for chalcone synthase and anthocyanidin synthase, which were downregulated in mycorrhizal seedlings, all DEGs related to flavonoid biosynthesis were upregulated, including 4-coumarate-CoA ligase, chalcone isomerase, flavanone 3-hydroxylase, flavonol synthase, and leucoanthocyanidin reductase. The metabolome analysis showed that several metabolites with insecticidal properties, including coumarin, stachydrine, artocarpin, norizalpinin, abietic acid, 6-formylumbelliferone, and vanillic acid, were significantly accumulated in the mycorrhizal seedlings. These findings suggest the potential of mycorrhiza-induced resistance for use in pest management of woody plants and demonstrate that the priming of JA-dependent responses in poplar seedlings contributes to mycorrhiza-induced resistance to insect pests.

Genome-Wide Analysis of the Homeobox Gene Family and Identification of Drought-Responsive Members in Populus trichocarpa

Homeobox (HB) genes play critical roles in the regulation of plant morphogenesis, growth and development. Here, we identified a total of 156 PtrHB genes from the Populus trichocarpa genome. According to the topologies and taxonomy of the phylogenetic tree constructed by Arabidopsis thaliana HB members, all PtrHB proteins were divided into six subgroups, namely HD-ZIP, ZF-HD, HB-PHD, TALE, WOX and HB-OTHERS. Multiple alignments of conserved homeodomains (HDs) revealed the conserved loci of each subgroup, while gene structure analysis showed similar exon–intron gene structures, and motif analysis indicated the similarity of motif number and pattern in the same subgroup. Promoter analysis indicated that the promoters of PtrHB genes contain a series of cis-acting regulatory elements involved in responding to various abiotic stresses, indicating that PtrHBs had potential functions in these processes. Collinearity analysis revealed that there are 96 pairs of 127 PtrHB genes mainly distributing on Chromosomes 1, 2, and 5. We analyzed the spatio-temporal expression patterns of PtrHB genes, and the virus-induced gene silencing (VIGS) of PtrHB3 gene resulted in the compromised tolerance of poplar seedlings to mannitol treatment. The bioinformatics on PtrHB family and preliminary exploration of drought-responsive genes can provide support for further study of the family in woody plants, especially in drought-related biological processes. It also provides a direction for developing new varieties of poplar with drought resistance. Overall, our results provided significant information for further functional analysis of PtrHB genes in poplar and demonstrated that PtrHB3 is a dominant gene regulating tolerance to water stress treatment in poplar seedlings.

First Report of Leaf Blight Caused by Nigrospora oryzae on Poplar in China

Populus alba L. × P. berolinensis Dipp. (a hybrid poplar, ‘PaPb poplar’) exhibits fast growth and beautiful tree shape with high drought and cold tolerance, and is widely planted in the cities of Northeast China because of its ornamental and ecological value (Wang et al. 2008). In October 2020, an unknown leaf blight symptom was observed on the seedlings of ‘PaPb poplar’ at Shenyang Agricultural University (41°49′N, 123°34′E) located in Shenyang City, Liaoning Province, China. The disease incidence was 50% in a survey of 200 seedlings on the campus. The typical symptoms were brown-to-black, irregular-shaped lesions (Fig. 1A). To investigate the disease, five symptomatic leaves were collected, and pieces were cut at the margin of diseased and healthy tissue. These pieces were surface sterilized with 2% sodium hypochlorite for 2 min, rinsed three times with sterile distilled water, air dried, placed on potato dextrose agar (PDA) and incubated at 28°C. After 5 days of incubation, three isolates with similar morphological characteristics were observed. Isolate N03 was chosen and used for pathogen identification. The fungal colonies were initially white in color, and later turned gray to black (Fig. 1D). Conidia were single-celled, black, spherical or oblate in shape measuring 9.19 to 12.78 μm × 12.61 to 14.81 μm in diameter (n=40) (Fig. 1E). These were borne on hyaline vesicles at the tip of a conidiophore. Morphologically, the isolate N03 was identified as Nigrospora oryzae (Berk. and Broome) Petch (Wang et al. 2017). The genomic DNA was extracted with a SP Fungal DNA Kit (D5542-01, OMEGA). The internal transcribed spacer (ITS) region, translation elongation factor 1-alpha (TEF1-α), and partial beta-tubulin (TUB) genes were amplified using the primers ITS5/ITS4 (White et al. 1990), EF1-728F/EF-2R (Carbone and Kohn, 1999; O’Donnell et al. 1998), and Bt-2a/Bt-2b (Glass and Donaldson, 1995) primer sets, respectively. The PCR products of ITS, TEF1-α, and TUB were amplified, sequenced, and deposited in GenBank with the following accession numbers MZ148528, MZ182080, and MZ182079, respectively. BLASTn analysis of the ITS, TEF1-α, and TUB sequences had 99.3%, 99.8%, and 99.27% nucleotide identities to MK131325, KY019328, and KY019559, respectively. A phylogenetic tree based on combined ITS, TEF1-α, and TUB sequences was constructed using a Maximum Likelihood method with 1000 bootstraps showing that N03 was grouped with other N. oryzae isolates (Fig. 2). The fungus was identified as N. oryzae based on morphological characteristics and molecular analyses. Koch’s postulates were completed to confirm the pathogenity of N. oryzae on ‘PaPb poplar’. The N03 spore suspension (105 spores/mL) was used to inoculate detached leaves and field leaves in two experiments. The two experiments were repeated three times, respectively. In the detached leaf test, 10 healthy leaves collected from 1-year-old ‘PaPb poplar’ seedlings were inoculated with N03 by spraying with the spore suspension followed by incubation at 28°C on wet filter papers in a petri dish for 7 days. 10 leaves were sprayed with sterile water to save as the controls. For field leaf test, leaves of 5 plants were spray-inoculated with the spore suspension at the 4-week-old growth stage, and an additional 5 plants were sprayed with sterile water. Seven days after inoculation, brown-to-black, irregular-shaped lesions on the margin of leaves were observed on inoculated leaves but not on the controls (Fig. 1B and C). All detached leaves inoculated with N03 were symptomatic. In the field tests, symptom appeared on 20 of the 30 inoculated leaves. N. oryzae was re-isolated from all the inoculated detached leaves and inoculated plants, but not from the controls. N. oryzae is a known pathogen of several hosts, such as Costus speciosus (Koen.) Sm. and Mentha spicata L., but has not been reported on any species of Populus. To our knowledge, this is the first report of leaf blight of ‘PaPb poplar’ caused by N. oryzae in China and the world. This disease could affect growth and development of ‘PaPb poplar’ seedlings, and may cause economic losses in the future. Appropriate strategies should be developed to manage this disease.

CREATION OF FOREST CARBON (CARBON) LANDSCAPES: ECONOMIC COMPONENT

The article discusses the economic components of the creation and maintenance of forest carbon (carbonic) landfills by poplar seedlings with a closed root system on the territory of the Voronezh region. A general description of the consequences of climate change, an increase in the concentration of carbon dioxide in the atmosphere is given, with an emphasis on considering such consequences in the forest industry. The technology of creating forest carbonic polygons has been determined, taking into account the influence of the main factors (growing conditions, species composition, soil group, number and diameter of stumps, rutting length, planting density, average distance to crossings). When determining the costs of creating and maintaining a forest carbon (carbonic) landfill with poplar seedlings with a closed root system, a costly and normative approach was used. In the course of the study, it was determined that the creation and maintenance of forest carbonic landfills is a costly process, but the activity of carbon sequestration implies economic and environmental benefits. The cost of creating and maintaining one hectare of carboniferous forest landfill, created by seedlings of poplar with a closed root system, amounted to 303.9 thousand rubles. The creation of carbon polygons will allow controlling the carbon balance in the country's territories, helping to improve the environmental situation and increase the economic benefits of business by producing more competitive products.

Reintroduced Native Populus nigra in Restored Floodplain Reduces Spread of Exotic Poplar Species

Exotic Populus taxa pose a threat to the success of riparian forest restoration in floodplain areas. We evaluated the impact of exotic Populus taxa on softwood riparian forest development along the river Common Meuse after introducing native Populus nigra and after the re-establishment of the natural river dynamics. We sampled 154 poplar seedlings that spontaneously colonized restored habitat and assessed their taxonomy based on diagnostic chloroplast and nuclear microsatellite markers. Furthermore, by using a paternity analysis on 72 seedlings resulting from six open pollinated P. nigra females, we investigated natural hybridization between frequently planted cultivated poplars and native P. nigra. The majority of the poplar seedlings from the gravel banks analyzed where identified as P. nigra; only 2% of the sampled seedlings exhibited genes of exotic poplar species. Similarly, the majority of the seedlings from the open pollinated progenies were identified as P. nigra. For three seedlings (4%), paternity was assigned to a cultivar of P. × canadensis. Almost two decades after reintroducing P. nigra, the constitution of the seed and pollen pools changed in the study area in favor of reproduction of the native species and at the expense of the exotic poplar species. This study indicates that, although significant gene flow form exotic poplars is observed in European floodplains, restoration programs of the native P. nigra can vigorously outcompete the exotic gene flows and strongly reduce the impact of exotic Populus taxa on the softwood riparian forest development.

Enhancing the expression of ARK1 genes in poplar leads to multiple branches and transcriptomic changes

The ARBORKNOX1 ( ARK1 ) gene is an important gene for regulating plant growth and development; however, transcriptomic responses of enhancing expression of ARK1 gene in poplar are poorly investigated. To provide insight into the gene function of the ARK1 gene in poplar, the ARK1 transgenic poplar ‘717' and ‘84 K' lines were obtained, the morphology of transgenic plants was observed, and transcriptome profiles were compared. The results showed that there were multiple branches in ARK1 transgenic seedlings compared with non-transgenic seedlings. The results of transcriptome analysis showed that there were significant differences in transcriptome profiles between the transgenic lines of ‘717' and ‘84 K', and between non-transgenic lines (CK) and transgenic plants. The real-time quantitative polymerase chain reaction (RT-qPCR) analysis confirmed the expression levels of the genes involved in the pathway of zeatin biosynthesis and brassinosteroid biosynthesis. The increase in expression levels of AHP and CYCD3 was related to multiple branches. Enhancing the expression of the ARK1 gene in poplar seedlings leads to multiple branches and transcriptomic changes.

Intake of Radionuclides in the Trees of Fukushima Forests 4. Binding of Radioiodine to Xyloglucan

The 1,4-linked glucans such as xyloglucan and amylose are known to form a complex with iodine/iodide ions and to also be precipitated with CaCl2 in the presence of iodine. Here, we show that iodine gas could be specifically incorporated into xyloglucan. Furthermore, we show that [125I]I2 gas is, over time, incorporated at high levels into the entire outer surface of poplar seedlings but that spraying seedlings with abscisic acid to close stomata decreases the incorporation of the gas. There was less incorporation of the gas in a transgenic poplar overexpressing xyloglucanase at the early stages when compared with a wild type. This shows that xyloglucan serves as a key absorber of iodine gas into a plant body. After individual leaves of cultured seedlings were exposed to the gas for 30 min, no radioiodine was emitted from those leaves over the following two weeks, indicating that no turnover occurs in radioiodine once it is bound to the polysaccharides in plant tissues. We conclude that forest trees could serve as one of the largest enormous capture systems for the radioiodine fallout following the nuclear power plant accident in Fukushima.