Detecting the Different Responses of Roots and Shoots to Gravity in Salix matsudana (Koidz)

The study of the gravity response of roots and shoots is of great significance when exploring the polarity of plants and the development of the forest industry. In our study, normal and inverted cuts of Salix matsudana (Koidz) were cultured. The total RNAs of roots and shoots were extracted. Based on the comparative transcriptome, 412 and 668 genes were differentially expressed. The plasma membrane-, cell wall-, and extracellular region-related genes were up-regulated in the shoots, while the carbon metabolism and the nitrogen metabolism were up-regulated in the roots. Combining the alternative splicing genes, we found a potential gravity response network: in the shoots, LecRLKs were highly sensitive to gravity and further affected the alternative splicing of SNARE, as well as inducing an inhomogeneous distribution of auxin and a negative geotropism curve. In the roots, AP2/ERFs and STRKs were highly sensitive to gravity and regulated the expression level of STPKs and WAKs, finally resulting in a geotropism curve. Moreover, cell division was suppressed in both the roots and the shoots under inverted conditions with different mechanisms. Cell division inhibitors (KRPs) were up-regulated in the roots, while DNA helicase MCMs were down-regulated in the shoots. These results provide an important foundation for further studies of the molecular mechanisms and genetic regulation of plant responses to gravity and the plant polarity of forest trees.

Comparative study on windbreak effects of two different configuration shelterbelts

Pinus sylvestris, Artemisia ordosica, Tamarix chinensis Lour, Elaeagnus angustifolia and Salix matsudana Koidz were selected to design the models of forest belts. Through wind tunnel experiments, we analyzed the wind speed frequency, wind speed flow field and wind protection efficiency of two different configuration shelterbelts. The results showed the wind speed frequency of mixed shelterbelts was generally lower than that of single shelterbelts with the same configuration. The similarity of the average wind speed between No.1 A3 and No.2 C3 indicates that the windbreak effect of low-density arbor-shrub mixed shelterbelt was similar to that of high-density single shelterbelt. The average wind protection efficiency of No.1 A1 was similar to that of No.1 C2, indicating that the influence of density of pinus sylvestrison of wind protection efficiency is greater than the height of forest belt. the average wind protection efficiency of No. 2 A2 was 57.64%, which was similar to that of No. 2 B3, indicating that the height of the forest belt increased, and the influence of density on the protection effect of the forest belt decreased.Therefore, the wind protection ability of multi-species and highly hierarchical mixed forests was higher than that of single shelterbelt.

Insights into the isotopic mismatch between bulk soil water and <i>Salix matsudana</i> Koidz trunk water from root water stable isotope measurements

Increasing numbers of field studies have detected isotopic mismatches between plant trunk water and its potential sources. However, the cause of these isotopic offsets is not clear, and it is uncertain whether they occur during root water uptake or during water transmission from root to trunk. Thus, we measured the specific isotopic composition (δ2H and δ18O) of each component (e.g. bulk soil water, mobile water, groundwater, trunk water and root water of Salix matsudana Koidz trees) in the soil–root–trunk continuum with a resolution of about 3 days. We report three main findings. First, we detected a clear separation between the isotopic compositions of mobile water and bulk soil water, but the distinction between mobile water and bulk soil water gradually decreased with increasing soil depth. Second, root water composition deviated from bulk soil water isotopic composition but overlapped with the composition derived for less mobile water. The maximum differences in δ2H and δ18O between bulk soil water and root water were −8.6 ‰ and −1.8 ‰, respectively. Third, trunk water was only isotopically similar to root water at 100–160 cm depths, and it remained stable during the experimental period, suggesting that the trees consistently used the stable deep water source. In conclusion, the isotopic offset between bulk soil water and trunk water of S. matsudana reflected an isotopic mismatch between root water and bulk soil water associated with the heterogeneity of the soil water. Our results illuminate relationships between the isotopic compositions of soil waters of various mobilities, root water and trunk water that may be useful for advancing our understanding of root water uptake and transport.

Impacto de la aplicación de una metodología de marcación del cambium en la evaluación de vasos formados exclusivamente en el xilema bajo tratamiento de estrés hídrico

La formación de xilema es un proceso cíclico a partir del cambium. La actividad estacional del cambium puede ser estudiada con diferentes metodologías (lesión mecánica en este trabajo). Identificar con exactitud el xilema formado durante un período de tratamiento de estrés abiótico permitiría realizar interpretaciones correctas en cuanto al efecto específico del estrés en la producción de xilema. Los objetivos fueron: 1- Evaluar si las mediciones de vasos realizadas sólo en el xilema formado durante el período de estrés hídrico tienen mayor sensibilidad que cuando se realizan en la totalidad del xilema; 2- Explicar los puntos más importantes de la técnica de marcación y los pasos metodológicos posteriores en el análisis morfométrico. Se trabajó sobre diez plantas de Salix matsudana ´NZ692´, a 5 plantas se las inundó por 35 días y las otras 5 fueron controles. Posteriormente crecieron por 28 días de recuperación. Las plantas fueron marcadas con una incisión de bisturí a nivel del cambium al inicio y al final del tratamiento. El análisis morfométrico de los vasos se realizó sobre 3 áreas en el xilema marcado: 1- sólo el área de tratamiento, 2- sólo el área de recuperación y 3- todo el crecimiento (suma de pre-tratamiento, tratamiento y recuperación). Los vasos producidos exclusivamente durante el período de estrés se diferenciaron de los vasos formados posteriormente y de los medidos en todo el tallo. Asimismo, la técnica de marcación del cambium permitió identificar el área específica de formación de xilema bajo estrés hídrico. La utilización de esta técnica relativamente sencilla, puede mejorar sustancialmente la interpretación de cambios anatómicos ocurridos en el xilema bajo estrés.

Messenger RNAs Mobile in Salix Matsudana Grafts Were in Association With Plant Rooting

Messenger RNAs exchanged between scions and rootstocks of grafted plants seriously affect their traits performance. The study goals were to identify the long-distance mRNA transmission events in grafted willows using a transcriptome analysis and to reveal the possible effects on rooting traits. The results showed that the Salix matsudana variety 9901 has better rooting ability than YJ, which reasonably improved the rooting performance of the heterologous grafts 9901 (scion) / YJ (rootstock). A transcriptome analysis showed that 2,948 differentially expressed genes (DEGs) were present in the rootstock of 9901/YJ grafted plants in comparison with YJ/YJ. Among them, 692 were identified as mRNAs moved from 9901 scion based on a SNP analysis of two parents. They were mostly 1,001–1,500 bp, had 40–45% GC contents, or had expression abundance values less than 10. However, mRNAs over 4,001 bp, having 50–55% GC contents, or having expression abundance values of 10–20 were preferentially transferred. Eight mRNAs subjected to long-distance trafficking were involved in the plant hormone pathways and may significantly promote the root growth of grafted plants. Thus, heterologous grafts of Salix matsudana could efficiently influence plant rooting since of the mRNAs transport from scion to rootstock. Thus, the grafting parents and grafting patterns would be much concerned in the breeding process to gain the expected results in future.

First report of leaf spot caused by Colletotrichum siamense on Salix matsudana in China

Salix matsudana Koidz. (Chinese willow) is an important landscaping tree species widely grown in China (Zhang et al. 2017). In October 2019, a characteristic leaf spot disease of S. matsudana was found on the campus of Nanjing Forestry University. Most 25-year-old S. matsudana trees (13 out of 21, approximately 62%) on campus showed the leaf spot disease. On average, 70% of the leaves per individual tree were affected by this disease. Foliar symptoms began as dark brown, irregular spots and the centers were gray-white, gradually enlarging with time. Leaf spot symptomatic leaves were collected from three infected S. matsudana trees (10 leaves/tree), and small infected tissues (3–4 mm2) were surface-sterilized in 75% ethanol for 30 s, 1% NaClO for 90 s, rinsed in ddH2O, dried on sterilized filter paper, and plated on potato dextrose agar (PDA), and then incubated at 25°C. Three isolates (NHY1-1, NHY1-2, and NHY1-3) of the same fungus were obtained in 85% of the samples and deposited in China's Forestry Culture Collection Center (NHY1-1: cfcc55354, NHY1-2: cfcc55355, NHY1-3: cfcc55359). The colonies of three isolates were white, but the reverse side was grayish-white. The conidia of NHY1-1 were one-celled, straight, subcylindrical, hyaline, 14.4 ± 0.9 × 5.4 ± 0.4 µm (n = 50), with a rounded end. Conidiophores were hyaline to pale brown, septate, and branched. Appressoria were one-celled, ellipsoidal, brown or dark brown, thick-walled, 8.0 ± 0.9 × 5.9 ± 0.5 µm (n = 50). The conidia and appressoria of the other two isolates weralmost identical to NHY1-1. The morphological characters of the three isolates were matched with those of the Colletotrichum gloeosporioides complex (Weir et al. 2012). For accurate identification, the DNA of the three isolates was extracted. The internal transcribed spacer region (ITS), actin (ACT), calmodulin (CAL), chitin synthase (CHS-1), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), superoxide dismutase (SOD2), and β-tubulin 2 (TUB2) genes were amplified using the primer pairs ITS1/ITS4, ACT-512F/ACT-783R, CL1C/CL2C, CHS-79F/CHS-345R, GDF1/GDR1, SODglo2-F/SODglo2-R, and Bt2a/Bt2b, respectively (Weir et al. 2012). The sequences were deposited in GenBank [Accession Nos. MW784679 and MW808959 to MW808964 for NHY1-1; MW784726 and MW808965 to MW808970 for NHY1-2; MW784729 and MW808971 to MW808976 for NHY1-3]. A BLAST search of GenBank showed that ITS, ACT, CAL, GAPDH, SOD2, and TUB2 sequences of the three isolates were identical to Colletotrichum siamense at a high level (>99%), and CHS-1 sequences of three isolates were consistent with Colletotrichum fructicola at a high level (>99%). A maximum likelihood and Bayesian posterior probability analyses using IQtree v. 1.6.8 and Mr. Bayes v. 3.2.6 with the concatenated sequences (ITS, ACT, CAL, CHS-1, GAPDH, SOD2, and TUB2) placed NHY1-1, NHY1-2, and NHY1-3 in the clade of C. siamense with high bootstrap support values (ML/BI = 93/1). The pathogenicity of three isolates were tested on potted 2-yr-old seedlings (50-cm tall) of S. matsudana, which were grown in a greenhouse. Healthy leaves were wounded with a sterile needle and then inoculated with 10 µL of conidial suspension (106 conidia/mL). Controls were treated with ddH2O (Zhu et al. 2019). In total, 12 seedlings were inoculated including controls. Three seedlings/isolate and 10 leaves/seedling were used for each treatment. The plants were covered with plastic bags after inoculation and sterilized H2O was sprayed into the bags twice/day to maintain humidity and kept in a greenhouse at the day/night temperatures at 25 ± 2 / 16 ± 2°C. Within 7 days, all the inoculated points showed lesions similar to those observed in field, whereas controls were asymptomatic. The infection rate of each of the three isolates is 100%. C. siamense was re-isolated from the lesions, whereas no fungus was isolated from control leaves. The diseases caused by C. siamense often occur in tropical and subtropical regions of China, with a wide range of hosts, such as Hevea brasiliensis and Coffea arabica, etc. (Cao et al. 2019; Liu et al. 2018). This is the first report of C. siamense causing leaf spot of S. matsudana in China and the world. These data will help to develop effective strategies for managing this newly emerging disease.

Genome-wide investigation of the AP2/ERF superfamily and their expression under salt stress in Chinese willow (Salix matsudana)

AP2/ERF transcription factors (TFs) play indispensable roles in plant growth, development, and especially in various abiotic stresses responses. The AP2/ERF TF family has been discovered and classified in more than 50 species. However, little is known about the AP2/ERF gene family of Chinese willow (Salix matsudana), which is a tetraploid ornamental tree species that is widely planted and is also considered as a species that can improve the soil salinity of coastal beaches. In this study, 364 AP2/ERF genes of Salix matsudana (SmAP2/ERF) were identified depending on the recently produced whole genome sequencing data of Salix matsudana. These genes were renamed according to the chromosomal location of the SmAP2/ERF genes. The SmAP2/ERF genes included three major subfamilies: AP2 (55 members), ERF (301 members), and RAV (six members) and two Soloist genes. Genes’ structure and conserved motifs were analyzed in SmAP2/ERF family members, and introns were not found in most genes of the ERF subfamily, some unique motifs were found to be important for the function of SmAP2/ERF genes. Syntenic relationships between the SmAP2/ERF genes and AP2/ERF genes from Populus trichocarpa and Salix purpurea showed that Salix matsudana is genetically more closely related to Populus trichocarpa than to Salix purpurea. Evolution analysis on paralog gene pairs suggested that progenitor of S. matsudana originated from hybridization between two different diploid salix germplasms and underwent genome duplication not more than 10 Mya. RNA sequencing results demonstrated the differential expression patterns of some SmAP2/ERF genes under salt stress and this information can help reveal the mechanism of salt tolerance regulation in Salix matsudana.

Affinibrenneria salicis gen. nov. sp. nov. isolated from Salix matsudana bark canker

L3-3HAT, a Gram-negative-staining, facultatively anaerobic, motile bacterial strain, was isolated from the symptomatic bark of Salix matsudana canker in China. 16S rRNA gene analysis revealed that the novel strain shares the highest sequence similarity with Brenneria goodwinii FRB141T (95.5 %). In phylogenetic trees based on four housekeeping genes (gyrB, rpoB, atpD and infB) and the 16S rRNA gene sequence, the novel strain formed a separate branch from the five genera of the family Pectobacteriaceae (Lonsdalea, Brenneria, Dickeya, Pectobacterium and Sodalis), suggesting that the novel strain should belong to a novel species of a novel genus within the family Pectobacteriaceae. The result was also supported by phylogenomics, amino acid identity and average nucleotide identity. The major fatty acids were C14:0, C16:0, C17:0 cyclo, and C19:0 cyclo ɷ8c. Genome analysis showed that the novel strain has a large genome (5.89 Mb) with 5,052 coding genes, including 181 virulence genes by searching the pathogen-host interactions database (PHI-base), indicating that the novel strain is a potential pathogen of plants and animals. Based on phenotypic and genotypic characteristics, the L3-3HAT strain represents a novel species of a novel genus in the Pectobacteriaceae family, for which the name Affinibrenneria salicis gen nov. sp. nov. is proposed. The strain type is L3-3HAT (= CFCC 15588T = LMG 31209T).