Pinus Thunbergii Parl Research Articles

Thinning shelter forest reduced the soil priming effect and soil microbial respiration by altering the contribution of soil organic carbon from different sources

Thinning affects the understorey microclimate and the material input into the shelter forest soil, and it alters the contribution of the plant-derived and microbial-derived carbon (C). This difference effects the utilization of C by soil microbes and ultimately soil C emission. There is still no adequate characterization for the contribution of plant-derived and microbial-derived C to soil organic C (SOC) under thinning. Moreover, how this difference affects microbially driven C emissions (soil microbial respiration, SMR) and the contribution to the soil priming effect in this process are still unknown. The contribution of plant-derived and microbial-derived C to SOC were quantified by comparing lignin phenols and amino sugars under thinning and control treatments. Meanwhile, a 30-day indoor cultivation experiment was conducted. Soil priming effect and SMR were tested by adding 13C-labelled glucose to the soil of the thinning and control treatments. The purpose was to investigate the contribution of C from different sources to SOC, as well as its influence mechanism on soil priming effect and SMR.The results indicate that thinning significantly increased the fungal-derived (10.19 and 7.34 g kg−1) and microbial-derived C (11.05 and 8.19 g kg−1) of Quercus acutissima Carruth (QAC) and Pinus thunbergii Parl (PTP). However, the bacterial-derived C (0.86 and 0.85 g kg−1) decreased. Compared to the control treatment, the plant-derived, fungal-derived and microbial-derived C increased by 6.6 % and 1.1 %, 17.9 % and 12.2 %, and 16.6 % and 10.1 %, respectively, while bacterial-derived C decreased by 1.3 % and 2.1 %. Thinning reduced the soil priming effect of QAC by lowering the pH and increasing F/B and fungal-derived C. At the same time, this phenomenon was not evident in PTP, mainly attributed to the content of SOC and vanillyl and syringyl monomers. Thinning reduced the SMR, which was mainly related to the suppression of soil priming effect. In summary, this study has demonstrated that thinning reduced the soil priming effect and SMR of shelter forests by altering the contribution of plant-derived and microbial-derived C to SOC.

Pinus thunbergii Parl. Somatic Plants' Resistance to Bursaphelenchus xylophilus Depends on Pathogen-Induced Differential Transcriptomic Responses.

Pinus thunbergii Parl. is an economically and medicinally important plant, as well as a world-renowned horticultural species of the Pinus genus. Pine wilt disease is a dangerous condition that affects P. thunbergii. However, understanding of the genetics underlying resistance to this disease is poor. Our findings reveal that P. thunbergii's resistance mechanism is based on differential transcriptome responses generated by the early presence of the pathogen Bursaphelenchus xylophilus, also known as the pine wood nematode. A transcriptome analysis (RNA-seq) was performed to examine gene expression in shoot tissues from resistant and susceptible P. thunbergii trees. RNA samples were collected from the shoots of inoculated pines throughout the infection phases by the virulent Bursaphelenchus xylophilus AMA3 strain. The photosynthesis and plant-pathogen interaction pathways were significantly enriched in the first and third days after infection. Flavonoid biosynthesis was induced in response to late infestation (7 and 14 days post-infestation). Calmodulin, RBOH, HLC protein, RPS, PR1, and genes implicated in phytohormone crosstalk (e.g., SGT1, MYC2, PP2C, and ERF1) showed significant alterations between resistant and susceptible trees. Furthermore, salicylic acid was found to aid pine wood nematodes tolerate adverse conditions and boost reproduction, which may be significant for pine wood nematode colonization within pines. These findings provide new insights into how host defenses overcame pine wood nematode infection in the early stage, which could potentially contribute to the development of novel strategies for the control of pine wilt disease.

Antifungal and Plant-Growth Promotion Effects of Bacillus velezensis When Applied to Coastal to Pine (Pinus thunbergii Parl.) Seedlings

Fungal diseases such as root rot and leaf blight cause substantial losses in coastal pine (Pinus thunbergii Parl.) seedling production, which hinders afforestation/forest restoration programs. We isolated and identified Fusarium oxysporum and Alternaria alternata as the causal agents of root rot and needle blight diseases and investigated the biocontrol efficacy against the fungal pathogens and growth promotion of coastal pine seedlings using Bacillus velezensis CE 100. The bacterium produced the hydrolytic enzymes chitinase, β-1,3-glucanase, and protease enzymes, and the crude enzyme fraction of the biocontrol strain caused the deformation of the fungal cell wall and antagonized F. oxysporum and A. alternata, causing respective inhibition of spore germination by 91.0% and 85.9% and mycelial growth by 58.3% and 54.3%, at a concentration of 1000 µL/mL. Consequently, the bacterial treatment improved the survival rate of seedlings 1.9 times relative to the control group. The bacterium secreted indole-acetic acid (IAA) phytohormone and enhanced root growth and absorption of nutrients, which notably enhanced the biomass production of coastal pine seedlings. Therefore, these results provide evidence that B. velezensis CE 100 is an effective antifungal and growth-promoting bacterium that can facilitate the production of high-quality coastal pine seedlings for the restoration and establishment of coastal forests.

Resistant and Susceptible Pinus thunbergii ParL. Show Highly Divergent Patterns of Differentially Expressed Genes during the Process of Infection by Bursaphelenchus xylophilus.

Pine wilt disease (PWD) is a devastating disease that threatens pine forests worldwide, and breeding resistant pines is an important management strategy used to reduce its impact. A batch of resistant seeds of P. thunbergii was introduced from Japan. Based on the resistant materials, we obtained somatic plants through somatic embryogenesis. In this study, we performed transcriptome analysis to further understand the defense response of resistant somatic plants of P. thunbergii to PWD. The results showed that, after pine wood nematode (PWN) infection, resistant P. thunbergii stimulated more differential expression genes (DEGs) and involved more regulatory pathways than did susceptible P. thunbergii. For the first time, the alpha-linolenic acid metabolism and linoleic acid metabolism were intensively observed in pines resisting PWN infection. The related genes disease resistance protein RPS2 (SUMM2) and pathogenesis-related genes (PR1), as well as reactive oxygen species (ROS)-related genes were significantly up-expressed in order to contribute to protection against PWN inoculation in P. thunbergii. In addition, the diterpenoid biosynthesis pathway was significantly enriched only in resistant P. thunbergii. These findings provided valuable genetic information for future breeding of resistant conifers, and could contribute to the development of new diagnostic tools for early screening of resistant pine seedlings based on specific PWN-tolerance-related markers.

Effects of Nitrogen Deposition on Leaf Litter Decomposition and Soil Organic Carbon Density in Arid and Barren Rocky Mountainous Regions: A Case Study of Yimeng Mountain

The ecological impact of nitrogen (N) deposition has gained significance since the advent of the industrial revolution. Although numerous studies have examined the impact of N deposition on soil organic carbon (SOC), certain arid and barren rocky mountainous regions, which experience more pronounced N limitations, have been overlooked. This study was conducted in the Yimeng Mountains, examining eight treatments created by four N addition levels (0 kg N ha−1 yr−1, 50 kg N ha−1 yr−1, 100 kg N ha−1 yr−1 and 200 kg N ha−1 yr−1) and two tree species (Quercus acutissima Carruth. and Pinus thunbergii Parl.). The research revealed variations in the effect of N addition on leaf litter decomposition and SOC density (SOCD) between different tree species. Notably, N addition stimulated the decomposition of leaf litter from Quercus acutissima Carruth. However, the decomposition of Pinus thunbergii Parl. leaf litter was enhanced at N addition levels below 100 kg N ha−1 yr−1, while it was hindered at levels exceeding 100 kg N ha−1 yr−1. In the Quercus acutissima Carruth. forest, the N addition levels of 50 kg N ha−1 yr−1, 100 kg N ha−1 yr−1 and 200 kg N ha−1 yr−1 resulted in decreases in SOCD by 10.57%, 22.22% and 13.66%, respectively, compared to 0 kg N ha−1 yr−1. In the Pinus thunbergii Parl. forest, the N addition levels of 50 kg N ha−1 yr−1, 100 kg N hm−2 ha−1 and 200 kg N ha−1 yr−1 led to increases in SOCD by 49.53%, 43.36% and 60.87%, respectively, compared to 0 kg N ha−1 yr−1. Overall, N addition decreases the SOCD of Quercus acutissima Carruth., but it increases the SOCD of Pinus thunbergii Parl., attributed to the alteration in soil enzyme stoichiometry and nutrient cycling by N addition. This study fills a theoretical gap concerning leaf litter decomposition and SOC sequestration in arid and barren rocky mountainous regions under global climate change.

The effect of thinning intensity on the soil carbon pool mediated by soil microbial communities and necromass carbon in coastal zone protected forests

Thinning is a common forest management measure that can effectively maintain the ecological service function of protected forests. However, the effect of thinning on the soil carbon (C) pool remains uncertain. In particular, we lack an understanding of the complete link between thinning and microbial communities, microbial necromass C, and consequently, soil C pools in coastal zone protected forests. In this study, three thinning intensities, i.e., a control treatment (CT, i.e., no thinning), light thinning (LT) and heavy thinning (HT), were established in three types of forests (Quercus acutissima Carruth, Pinus thunbergii Parl and mixed Quercus acutissima Carruth and Pinus thunbergii Parl, i.e., QAC, PTP and QP, respectively). Two years after the completion of thinning, we investigated the changes in the soil organic carbon (SOC) fractions, soil microbial community and soil microbial necromass C in the surface layer (0–20 cm) and thoroughly evaluated the relationship between the potential change in SOC and the microbial community. Compared with CT, there was no change in the SOC content under LT and HT, but thinning conducted in QAC increased the proportion of mineral-associated organic C (MAOC) in SOC. Moreover, both LT and HT reduced the soil carbon lability (CL) in the QAC and QP forests. Different thinning intensities changed the soil microbial community structure, and most of the variation was explained by thinning and the soil physicochemical properties. The proportion of soil bacterial and fungal necromass C to SOC increased with increasing thinning intensity. The content of soil bacterial and fungal necromass C was mainly controlled by the relative abundance of the core phylum (relative abundance>10 %). Thinning affected the soil C pool by affecting the content of soil bacterial and fungal necromass C, but their accumulation pathways was different. The results showed that thinning was beneficial to the stability of SOC. The microbial C pool, total organic C pool and even bacterial and fungal C pools should be distinguished when studying the soil C pool, which can effectively deepen our understanding of the mechanism by which soil microorganisms affect the soil C pool.

Promoting the application of Pinus thunbergii Parl. to enhance the growth and survival rates of post-germination somatic plantlets

ObjectiveThere is a growing need for nematode resistant Pinaceae species plantlets to cope with the global scale degradation of coniferous forests, due to the prevalence of pine wilt disease. One of the bottlenecks that limits the commercialization of Pinaceae species plantlets is regeneration following their transfer from controlled sterile environments to the field while maintaining high survival rates.MethodsThe growth factors of somatic plantlets (SPs), such as sucrose, media, culture substrate, brassinolide and spectrum were investigated to promote the application of somatic nematode-resistant P. thunbergii plants in afforestation.ResultsThe 1/2 WPM liquid medium, culture substrate (perlite and vermiculite =1:1), and carbohydrate (20 g/L sucrose) were effective in stimulating the growth of rooted SPs. While for unrooted SPs, 1 ug/L of brassinolide enhanced plantlet growth and rooting. And blue light (B) significantly promoted the longitudinal growth of shoots, while red light (R) was beneficial for root growth during the laboratory domestication stage. High quality SPs were obtained at a R/B ratio of 8:2. Following this acclimatization protocol, the P. thunbergii SPs could be directly transplanted to the field with a higher survival rate (85.20 %) in a forcing house.Conclusionthis acclimatization protocol extremely improved the survival rate of P. thunbergii SPs. Moreover, this work will contribute to enhancing the possibilities for somatic plant afforestation with Pinus species.

Maximum Rooting Depth of Pinus thunbergii Parl. Estimated with Depth at the Center Point of Rotation in a Tree-Pulling Experiment in a Coastal Forest in Japan

Tree resistance to uprooting can be estimated as the critical turning moment in tree-pulling experiments. The depth at the center point of rotation (Dcp) in tree-pulling experiments is measured as an indicator of below-ground traits and is related to this critical turning moment. However, few researchers have investigated the relationship between the Dcp and maximum root depth. Our objective in this study was to clarify whether the Dcp in tree-pulling experiments can be estimated as the maximum root depth of Pinus thunbergii Parl. in sandy soils. We also estimated which position of displacement of the center of rotation (Cp) can be applied as the Dcp. We conducted tree-pulling experiments, and compared the Dcp obtained from images with the measured maximum root depth. We found significant positive correlations between the Dcp and maximum root depth. The Cp displacement concentrated immediately below the stem when the maximum critical turning moment was reached. This position should be measured as the Dcp, which is related to the maximum root depth. We found that the Dcp can serve as a parameter, preventing the need for uprooting, when tree-pulling experiments are performed to obtain an important below-ground trait for understanding the critical turning moment.

Effects of vegetation and terrain changes on spatial heterogeneity of soil C-N-P in the coastal zone protected forests at northern China.

Soil organic carbon (SOC), total nitrogen (TN) and total phosphorus (TP) are important indicators reflecting soil quality, and they can be used to effectively evaluate the effect of soil remediation. Many studies have evaluated the content of SOC, TN and TP in different ecosystems. However, after constructing protected forests for ecological restoration in the ecologically fragile coastal zone, the spatial distribution and influencing mechanism of SOC, TN and TP content is still uncertain. In this study, the spatial heterogeneity and influencing factors of SOC, TN and TP in surface (0-20cm) soil were analyzed by traditional analysis and geostatistics. A total of 39 soil samples were collected under the coastal zone protected forest types including Quercus acutissima Carruth (QAC), Pinus thunbergii Parl (PTP), mixed PTP and QAC (QP) and Castanea mollissima BL (CMB) in the coastal zone protected forests in northern China. The results show that SOC, TN and TP content were defined as moderate variation, and they also show significant changes under different protected forest types (P<0.05). The semivariance results indicate that SOC, TN and TP all exhibited strong spatial dependence class, with Range of 224m, 229m and 282m respectively, which were more than the sampling scale of 200m. The spatial prediction results showed that SOC, TN and TP content all appear in large areas of extremely low value in CMB, and its cross validation results showed that using vegetation and terrain factors as covariates in the spatial prediction of SOC, TN and TP can improve the prediction accuracy. The results of correlation analysis showed that the influencing factor for SOC and TN, and TP were NDVI and topographical changes, respectively. In general, vegetation and terrain factors as auxiliary factors can improved the accuracy of soil C-N-P spatial distribution prediction after afforestation in coastal zone.

Terpene Production Varies in Pinus thunbergii Parl. with Different Levels of Resistance, with Potential Effects on Pinewood Nematode Behavior

Determining the mechanisms of pine wilt disease (PWD) resistance in Pinus is a popular research topic, but information on volatile organic substances (VOS) and their role in PWD is lacking. Whether the difference in VOS among Pinus thunbergii parl. that have different levels of resistance with pine wood nematodes (PWNs) is the reason for the differing resistance needs to be studied. In this study, resistant P. thunbergii introduced from Japan and susceptible P. thunbergii native to China were used to investigate the effects of different lines inoculated with PWN. We determined the expression levels of the terpene synthesis-related genes geranylgeranyl diphosphate synthase (GGPPS), 3-hydroxy-3-methylglutaryl-coenzyme A reductase 1 (HMDH1), two kinds of alpha-farnesene synthase (PT) genes. The types and the relative percentage content of terpenoids in the pine needles were also determined by gas chromatography coupled with mass spectrometry (GC-MS). Results show that the growth, population size and migration of PWNs were significantly inhibited. The expression of terpene synthesis genes in the resistant P. thunbergii was higher than that in the susceptible one. The analysis of terpenoids revealed a total of 41 terpenoids, of which resistant P. thunbergii contained 39 and susceptible P. thunbergii only 28; 14 terpenoids were specific to resistant P. thunbergii, in which 8 of the terpenoids were constitutive terpenes and 6 were inducible terpenes. There were 3 terpenes unique to the susceptible P. thunbergii, and only 1 inducible terpene. Our results showed that the reduction in the expression of disease symptom and suppression of PWNs in resistant P. thunbergii was likely related to differences in the types and content of resistance-related substances in the trees. This study does not specifically connect elevated compounds in resistant P. thunbergii to resistance to PWN and assays should be conducted to establish direct effects of terpenoids on pinewood nematode activity and reproduction.

First Report of Needle Blight on Pinus thunbergii Parl. Caused by Fusarium proliferatum in China.

Pinus thunbergii Parl. (Japanese black pine), an evergreen species, is distributed along the seacoasts of China. In addition, this species has been planted along seacoasts as a windbreak to prevent soil erosion due to its resistance to salt and various environmental stresses. It can also be found in public parks and gardens due to its exquisite appearance and toughness. In August 2020, needle blight symptoms were found on several black pine trees in Sichuan Province, China. Further surveys showed that these symptoms are common. The disease incidence is less than 30% while severity of the disease is high. The tips of old needles first turn grayish green that developed into brown bands ranging from 1 to 2 mm. To determine the pathogen, small needle pieces (3-4 mm2 long) from the margin of fresh lesions were surface-sterilized for 30 s in 75% ethanol, follow by 1% NaOCl for 90 s, then washed three times with sterile water, and then were placed on potato dextrose agar (PDA) with 0.1 mg/mL ampicillin and incubated at 25°C. Pure cultures of 8 isolates were obtained by monosporic isolation, and a representative isolate (SC03) was deposited in the Co-Innovation Center for Sustainable Forestry in Southern China, Nanjing Forestry University. When cultured on Spezieller Nährstoffarmer Agar medium (Leslie and Summerell, 2006), the SC03 colony was beige-white, cottony from top view and pale orange near the center on the reverse side. The daily growth rate was 11.8 mm/day at 25°C in the dark. Microscopic observations showed hyaline and septate hyphae, slightly curved macroconidia with two to three septa measuring 17.5 - 30 × 3.7 - 7.5 μm (23.2 × 5.7 on average), and aseptate microconidia measuring 7.5 - 12.5 × 2.5 - 5.0 μm, (9.7 × 4.3 on average). The morphological characteristics of conidia and other structures of SC03 matched those of the Fusarium fujikuroi species complex (FFSC) (Abdalla et al. 2000). For accurate identification, translation elongation factor 1-alpha（TEF-1α） , and the second largest RNA polymerase subunit (RPB2) were amplified and sequenced using the primer pairs EF1 and EF2, RPB5f2 and RPB7cr. The sequences were deposited in GenBank [Accession TEF-1α: ON049647, RBP2: ON049648]. A Blast search of GenBank showed that TEF-1α and RPB2 sequences of SC03 matched Fusarium proliferatum (Matsush) Nirenberg at a high level (>99%). Phylogenetic analysis using neighbor joining and concatenated sequences (TEF-1α and RPB2) with MEGA X placed SC03 in F. proliferatum. For the pathogenicity test, a conidial suspension was prepared with a concentration of 2.0 × 107 conidia/ml. The suspension was sprayed onto 3 annual seedlings' needles, and the control was sprayed with sterile water. Inoculated and uninoculated plants were kept in humid chambers in a glasshouse where the average humidity was 60% and the average temperature was 27℃. After 10 days, typical symptoms appeared on inoculated needles, whereas control needles remained symptomless. These symptoms were similar to those observed in field. The fungus, F. proliferatum, was reisolated from those lesions, confirming Koch's postulates. No symptoms were observed on control plants. Fusarium proliferatum is a ubiquitous saprophytic fungus on cankers and very rarely reported to cause disease on pine needles. It has been reported to cause leaf blight of Polygonatum cyrtonema (Zhou et al. 2021) and Majesty palm (Polizzi and Vitale 2003). To our knowledge, this is the first report of needle blight on P. thunbergii caused by F. proliferatum in China. The disease represents a threat to producers and more research on the biology and management is needed.

Short Communication: Thromboplerous hyphae of the ectomycorrhizal mushroom Rhizopogon roseolus with and without a host tree

Abstract. Putra IP, Aimi, T, Shimomura N. 2022. Short Communication: Thromboplerous hyphae of the ectomycorrhizal mushroom Rhizopogon roseolus with and without a host tree. Nusantara Bioscience 14: 47-52. Thromboplerous hyphae are modified hyphae found in the basidiocarp of many agarics. However, data on their development and function as well as on their formation in pure cultures of ectomycorrhizal mushrooms are scarce. The aim of the present study was to characterize the cytological descriptions of thromboplerous hyphae from pure cultures of Rhizopogon roseolus (Corda) Th.M. Fr. with or without the host pine tree Pinus thunbergii Parl. Thromboplerous hyphae formed on the mycelium and mycelial cords in all experimental settings. Results revealed that thromboplerous hyphae were extremely melanized, smooth, cracked on the surface, and swollen at both hyphal termini. Thromboplerous hyphae produced with a host were mainly unbranched in shape in contrast to those formed without a host that possessed additional twisted, semi-twisted, and branched forms. Some thromboplerous hyphae that grew without a host had large diameters, and few showed notable septa and clamp connections. The present study also provides evidence of the initial development of thromboplerous hyphae via vegetative hyphae and tubular hyphae. This study contributes to understanding the cytology of thromboplerous hyphae grown with and without a host tree. Further investigations on the specific functions of thromboplerous hyphae are needed to deepen the current knowledge of fungal cytology.

Genetic evidence of stable northward extension of Pinus thunbergii Parl. forests in the Democratic People’s Republic of Korea

Genetic evidence of stable northward extension of Pinus thunbergii Parl. forests in the Democratic People’s Republic of Korea

海岸黑松和紫穗槐对间歇强净风和风沙流吹袭的生理响应机制

以海岸防风固沙优势树种紫穗槐（Amorpha fruticosa Linn）和黑松（Pinus thunbergii Parl）为研究对象，利用野外便携式沙风洞用间歇风吹模拟自然阵风，通过分析间歇强净风（18m/s）和强风沙流（172.93g cm^-1 min^-1）吹袭过程中和风后恢复中，两树种叶片膜脂过氧化产物含量、抗氧化酶活力、渗透调节物含量的变化，以探讨其对自然阵风吹袭响应机制及自愈修复生理机制。结果表明，自然状况下，紫穗槐和黑松叶片相对含水量（RWC）相近，但抗氧化酶活力及种类和渗透调节物含量及种类上存在差异。紫穗槐叶片丙二醛含量（MDA）、脯氨酸含量及过氧化氢酶（CAT）和过氧化物酶（POD）活力分别较黑松高93.3%、78.6%、118.8%、6.5倍。而黑松可溶糖含量和超氧化物歧化酶（SOD）活力较紫穗槐高111.5%和28.2%。在间歇净风和风沙流处理中，随着风吹袭次数增多，黑松叶片RWC趋于小幅降低，可溶性糖含量及POD、SOD、CAT活力呈小幅波动式变化；紫穗槐叶片RWC大幅下降，伴随着脯氨酸含量，POD、CAT、SOD活力大幅增高和MDA含量下降。在风吹袭中，叶片抗氧化酶活力和渗透调节含量的变化与叶片含水量变化密切相关。在风吹袭处理后的恢复期，黑松叶片MDA、可溶性糖、脯氨酸含量和CAT活力下降，但叶片一直维持较高可溶性糖含量和SOD活力；而紫穗槐伴随叶片RWC回升，而MDA、可溶性糖、脯氨酸含量下降和POD、CAT、SOD活力的增高。研究表明，风吹袭引发叶片水分亏缺成为启动植物对缺水补偿机制的诱导因子（自愈修复）。在风吹袭中增高的渗透调节物通过提高细胞吸水力为自愈修复提供物质基础，而抗氧化酶通过抑制膜脂过氧化、维护膜的完整性和弹性为自愈修复提供生理保护。因此，渗透调节和抗氧化酶通过维护水分平衡和氧自由基代谢平衡而在抗风生长和自愈修复中起重要生理保护作用。;In this study, the coastal area wind and sand blowing resistant Amorpha fruticosa Linn and Pinus thunbergii Parl were used as materials to learn how they responded physiologically to intervals of strong wind and wind-drift blowing, and what mechanisms of physiological adaptation and self-repairing were involved in their resistance to wind-drift blowing and self-repairing after wind blowing. The plants were subjected to 18 m/s wind speed and wind-drift blowing (172.93g cm^-1 min^-1) for 20 min and allowed to repair for 24h after the blowing treatment (repair period); these treatments were repeated three times using an outdoor portable wind tunnel. Relative water content (RWC), malondialdehyde (MDA), proline, soluble sugar, and the activities of superoxide dismutase (SOD), catalase (CAT), and peroxidase (POD) were measured in the leaves of the plants. The results showed that in the natural environment, there were no difference in RWC in the two species, but there were differences in the kinds of antioxidant enzyme and osmotic regulators between A. fruticosa and P. thunbergii, with the MDA content, proline content, and the activities of CAT and POD in leaves of A. fruticosa higher by 93.3%, 78.6%, 118.8%, and 650% than those in leaves of P. thunbergii. The soluble sugar content and SOD activity in leaves of P. thunbergii were higher by 111.5% and 28.2% than those in leaves of A. fruticosa. With increasing repetitions of blowing, RWC tended to decrease to a lower level, while the soluble sugar content and the activities of POD, SOD, CAT exhibited slight fluctuations in the leaves of P. thunbergii, but in A. fruticosa leaves, RWC dramatically dropped, and proline content and the activities of POD, CAT, and SOD significantly increased. Furthermore, during the repairing period, the contents of MDA, soluble sugar, and proline and CAT activity decreased in the leaves of P. thunbergii but the contents of MDA, soluble sugar, and proline decreased, and the activities of POD, CAT, and SOD increased, accompanied by an increase of RWC, in the leaves of A. fruticosa. These results indicate that water shortage induced by wind blowing was a direct factor causing physiological regulation and activating the compensation mechanism for water shortage (self-healing repair) in both trees. Although the two tree species belonged to different families and used different kinds of osmotic regulator and antioxidant enzymes in the physiological regulation of wind resistance, they both had the same strong ability to regulate physiological mechanisms during strong wind blows and post-wind repairing periods. It suggests that water shortage might activate the self-healing physiological mechanism. Under wind blowing, the increased content of soluble sugar and proline might provide materials for self-healing by improving cell water absorption, while antioxidases played an important physiological protective role in inhibition of membrane lipid peroxidation, thus maintaining membrane integrity and elasticity for self-healing. Therefore, self-healing is the key physiological adaptative mechanism for the two trees to adjust to the intervals of strong sea wind blowing on the coast.

Morphological and Phylogenetic Resolution of Diplodia neojuniperi Emerging Diplodia Top Dieback of Pinus thunbergii Parl. in China

In Bazhong City, Sichuan Province, China, top dieback symptoms were found on many pine trees (Pinus thunbergii Parl). The tips of old needles first turned grayish-green and then developed into brown bands in the field. Phylogenetic analysis of concatenated ITS and EF1-α indicated the pathogen of this dieback disease as Diplodia neojuniperi. Additionally, effects of temperature, pH and medium on the mycelial growth were also characterized. The most favorable temperature and pH level for mycelial growth are 25°C and 8, respectively. The optimal medium for mycelial growth is PDA medium. To our knowledge, this is the first report of D. neojuniperi causes Diplodia top dieback on Pinus thunbergii. Our results provide fundamental information for monitoring and preventing such disease in the future.

First Report of Brown Spot Needle Blight on Pinus thunbergii Parl. Caused by Aureobasidium pullulans in China.

Pinus thunbergii Parl., known as black pine, is widely distributed all over China. This pine variety can prevent soil desertification and promote soil conservation and is excellent for constructing fast-growing forests and shelter belts. The timber of this species can be used for infrastructure construction and furniture production. In August 2020, needle blight symptoms were found on several trees of black pine in Sichuan Province, China. Further surveys showed that these symptoms are common while the disease incidence is less than 30% which indicated the severity of the disease is mild. The tips of old needles first turn grayish green and developed into brown bands ranging from 1 to 2 mm. To determine the pathogen, 20 needle samples with typical symptoms were disinfected with 75% alcohol, and sections of the tissue were cut from joints of diseased and healthy tissues (visually healthy) with a sterilized scalpel, surface sterilized for 45 seconds in 75% alcohol, soaked for 90 seconds in 1.5% NaCIO, rinsed in sterilized water and dried. Small cut tissues were placed on potato dextrose agar (PDA) at 25℃ for 10 days. Pure cultures were obtained by monosporic isolation. The colonies initially appeared white to cream, yeast-like, and later turned to pink and remained at least 10 days. Conidia were hyaline, smooth-walled, single-celled, and ellipsoidal with variable shape and size, 7.5 to 16 × 3.5 to 7 µm (Zalar et al. 2008). DNA was extracted from the mycelium of the isolate by the cetyltriethylammonium bromide (CTAB) method and amplified through polymerase chain reaction (PCR) with the internal transcribed spacer (ITS) region of rDNA and partial β-tubulin genes of a representative isolate (SC05) were amplified using the ITS1/ITS4 and Bt2a/Bt2b primer pairs, respectively(Wu et al. 2017). The sequences submitted to GenBank (Accession Nos. MW228368 for ITS and MW256762 for β-tubulin) showed high similarity with BLAST sequences of Aureobasidium pullulans (ITS, KR704881 [100%]; β-tubulin, MT671934 [99.49%]). For the pathogenicity test, a conidial suspension was prepared with a concentration of 2.0 × 107 conidia/ml. The suspension was sprayed onto 3 annual seedlings' needles, and the control was sprayed with sterile water. Inoculated and non-inoculated plants were kept in humid chambers in a glasshouse. After 10 days, typical symptoms appeared on inoculated needles, whereas control needles remained symptomless. The fungus, A. pullulans, was reisolated from those lesions, confirming Koch's postulates. No symptoms were observed on control plants. Aureobasidium pullulans, a ubiquitous saprophytic fungus on many fruits and very rarely reported to cause disease on pine needles. Only reported invasion of Ozone-injured needles in P. strobus (Costonis and Sinclair 1972) and needles damaged by acid rain in P. sylvestris (Ranta 1990). To our knowledge, this is the first report of brown spot needle blight on P. thunbergii caused by A. pullulans in China. The disease represents a threat to pine manufactures and more research on the pathogenesis and management is needed. .

Diversity and Distribution of Xylophagous Beetles from Pinus thunbergii Parl. and Pinus massoniana Lamb. Infected by Pine Wood Nematode

The vectors of pinewood nematode of Bursaphelenchus xylophilus (Steiner &amp; Bührer, 1934) are mainly known as xylophagous beetles. Understanding the composition and distribution of these xylophagous beetles in host pine trees infected by PWN is critical to control the spread of PWN. In this study, we investigated the community structures of the xylophagous beetles in two main host trees in Fujian and Shandong, Pinus massoniana Lamb. and Pinus thunbergia Parl., in different stages of infection. All beetles were collected by dissecting the whole pine trees and then identified by their morphological characteristics and COI genes. The results showed that the diversity of xylophagous beetles was different not only between the two host pine trees but also among the different infection stages. The diversity of P. massoniana xylophagous beetles was significantly higher than that of P. thunbergii, and there were also significant differences in the different stages of PWN infection. In total, Scolytinae was the most common (53.70%), followed by Curculionidae (18.26%), Cerambycidae (16.31%), and Cleridae (6.04%). Monochamus alternatus, the most important vector of PWN, occupied a large niche and showed different aggregation positions during the three infection stages in both host trees. This result might be related to the resistance of bark beetles to host trees and competition with other xylophagous beetles. The community diversity of xylophagous beetles was jointly affected by both the infection stages of PWN and the spatial niche of xylophagous beetles. Knowledge of the diversity and competitive relationships among xylophagous beetles might help regulate the population dynamics of these beetles.

Carbon isotopic measurements from coastal zone protected forests in northern China: Soil carbon decomposition assessment and its influencing factors

Panting protected forests to increase soil carbon sequestration is an effective means of reducing carbon emissions. Soil organic carbon (SOC) decomposition is one of the main indicators of soil carbon sequestration. However, SOC decomposition and its influencing factors in protected forests have not been fully characterized, especially in coastal zones. In this paper, coastal zone protected forest stands composed of Quercus acutissima Carruth (QAC), Pinus thunbergii Parl (PTP) and mixed PTP and QAC (MF) were selected as the research objects. The trends of the SOC decomposition rate were characterized by the beta (β) value, and the influencing factors were further explored with structural equation models. The results were as follows: The SOC content decreased from leaf to litter and then to the soil profile at all sites, while the δ13C value increased. The β value ranged from −3.12 to −5.76, with an average of −3.81. The β value was positively correlated with the diversity and richness of soil bacteria, supporting the hypothesis that the increase in δ13C with depth was mainly caused by isotope fractionation in the process of microbial SOC decomposition. The structural equation model showed that nitrogen and the availability of nitrogen have a strong ability to explain the value of β, which indicates that nitrogen-based edaphic variables play an important role in affecting SOC decomposition. The SOC decomposition rate in PTP was higher than that in QAC and MF. The results of this study indicate that the prediction of SOC decomposition based on the β value is suitable for coastal zone protected forests. The incorporation of edaphic variables into global carbon cycle models may enhance the predictions of SOC dynamics in coastal zone protected forests.

Reconstruction of Conifer Root Systems Mapped with Point Cloud Data Obtained by 3D Laser Scanning Compared with Manual Measurement

Three-dimensional (3D) root system architecture (RSA) is a predominant factor in anchorage failure in trees. Only a few studies have used 3D laser scanners to evaluate RSA, but they do not check the accuracy of measurements. 3D laser scanners can quickly obtain RSA data, but the data are collected as a point cloud with a large number of points representing surfaces. The point cloud data must be converted into a set of interconnected axes and segments to compute the root system traits. The purposes of this study were: (i) to propose a new method for easily obtaining root point data as 3D coordinates and root diameters from point cloud data acquired by 3D laser scanner measurement; and (ii) to compare the accuracy of the data from main roots with intensive manual measurement. We scanned the excavated root systems of two Pinus thunbergii Parl. trees using a 3D laser scanner and neuTube software, which was developed for reconstructing the neuronal structure, to convert the point cloud data into root point data for reconstructing RSA. The reconstruction and traits of the RSA calculated from point cloud data were similar in accuracy to intensive manual measurements. Roots larger than 7 mm in diameter were accurately measured by the 3D laser scanner measurement. In the proposed method, the root point data were connected as a frustum of cones, so the reconstructed RSAs were simpler than the 3D root surfaces. However, the frustum of cones still showed the main coarse root segments correctly. We concluded that the proposed method could be applied to reconstruct the RSA and calculate traits using point cloud data of the root system, on the condition that it was possible to model both the stump and ovality of root sections.

Evaluation of somatic embryo production during embryogenic tissue proliferation stage using morphology, maternal genotype, proliferation rate and tissue age of Pinus thunbergii Parl

To determine the optimal embryogenic capacity (somatic embryo production) of the selected elite nematode-resistant genotypes of Pinus thunbergii, variables such as embryogenic tissue (ET) morphology, maternal genotype, proliferation rate and tissue age were analyzed. ET morphology and histological evaluation of the proliferation stage showed a decrease in filamentous clump and protuberant surfaces and a decline in the acetocarmine-staining area, which indicates a decrease in somatic embryo production (SEP). Variations in cell physiology during the proliferation stage showed that SEP was positively correlated with soluble sugars and proteins, but negatively correlated with starch, peroxidase, and superoxidase. In addition, SEP was significantly (p < 0.001) affected by maternal genotype, tissue age and proliferation rate. Moreover, SEP was positively correlated with proliferation rate (r = 0.98, p < 0.001), but negatively correlated with tissue age (r = − 0.95, p < 0.001). In general, the results suggest that SEP could be assessed in ET proliferation stages by the apparent cell morphology, histology, proliferation rate and tissue age, which provides novel insights for evaluating the ET maturation capacity (number of somatic embryos) during the proliferation stage of P. thunbergii somatic embryogenesis.