scholarly journals Multi-Omics of Pine Wood Nematode Pathogenicity Associated With Culturable Associated Microbiota Through an Artificial Assembly Approach

2022 ◽  
Vol 12 ◽  
Author(s):  
Shouping Cai ◽  
Jiayu Jia ◽  
Chenyang He ◽  
Liqiong Zeng ◽  
Yu Fang ◽  
...  
Keyword(s):  
Pine Wilt Disease ◽  
Pinus Massoniana ◽  
Tree Seedlings ◽  
Post Inoculation ◽  
Pinewood Nematode ◽  
Masson Pine ◽  
Pine Tree ◽  
Pine Trees ◽  
Bacteria And Fungi ◽  
The Relationship

Pinewood nematode (PWN), the causal agent of pine wilt disease (PWD), causes massive global losses of Pinus species each year. Bacteria and fungi existing in symbiosis with PWN are closely linked with the pathogenesis of PWD, but the relationship between PWN pathogenicity and the associated microbiota is still ambiguous. This study explored the relationship between microbes and the pathogenicity of PWN by establishing a PWN-associated microbe library, and used this library to generate five artificial PWN–microbe symbiont (APMS) assemblies with gnotobiotic PWNs. The fungal and bacterial communities of different APMSs (the microbiome) were explored by next-generation sequencing. Furthermore, different APMSs were used to inoculate the same Masson pine (Pinus massoniana) cultivar, and multi-omics (metabolome, phenomics, and transcriptome) data were obtained to represent the pathogenicity of different APMSs at 14 days post-inoculation (dpi). Significant positive correlations were observed between microbiome and transcriptome or metabolome data, but microbiome data were negatively correlated with the reactive oxygen species (ROS) level in the host. Five response genes, four fungal genera, four bacterial genera, and nineteen induced metabolites were positively correlated with the ROS level, while seven induced metabolites were negatively correlated. To further explore the function of PWN-associated microbes, single genera of functional microbes (Mb1–Mb8) were reloaded onto gnotobiotic PWNs and used to inoculate pine tree seedlings. Three of the genera (Cladophialophora, Ochroconis, and Flavobacterium) decreased the ROS level of the host pine trees, while only one genus (Penicillium) significantly increased the ROS level of the host pine tree seedlings. These results demonstrate a clear relationship between associated microbes and the pathogenicity of PWN, and expand the knowledge on the interaction between PWD-induced forest decline and the PWN-associated microbiome.

scholarly journals The specialization of Esteya vermicola hyphae in infection to Bursaphelenchus xylophilus and its colonization of pine tree

2020 ◽  
Author(s):  
Hai-Hua Wang ◽  
Can Yin ◽  
Jie Gao ◽  
Ran Tao ◽  
Piao-Piao Dai ◽  
...  
Keyword(s):  
Pine Wilt Disease ◽  
Bursaphelenchus Xylophilus ◽  
Wilt Disease ◽  
Fungal Colonization ◽  
Pinewood Nematode ◽  
Pine Tree ◽  
Pine Wilt ◽  
Pine Trees ◽  
Colonization Patterns ◽  
Esteya Vermicola

AbstractPine wilt disease (PWD) caused by the nematode Bursaphelenchus xylophilus is a serious problem on pines, and there is currently no effective control strategy for this disease. Although the endoparasitic fungus Esteya vermicola showed great effectiveness in controlling pine wilt disease, the colonization patterns of the host pine tree xylem by this fungus are unknown. To investigate the colonization patterns of pine xylem by this fungus, the species Pinus koraiensis grown in a greenhouse was used as an experimental host tree. The fungal colonization of healthy and wilting pine trees by E. vermicola was quantified using PCR with a TaqMan probe, and a green fluorescence protein (GFP) transformant was used for visualization. The results reported a specific infection approach used by E. vermicola to infect B. xylophilus and specialized fungal parasitic cells in PWN infection. In addition, the inoculated blastospores of E. vermicola germinated and grew inside of healthy pine xylem, although the growth rate was slow. Moreover, E. vermicola extended into the pine xylem following spray inoculation of wounded pine seedling stems, and a significant increase in fungal quantity was observed in response to B. xylophilus invasion. An accelerated extension of E. vermicola colonization was shown in PWN-infected wilting pine trees, due to the immigration of fungal-infected PWNs. Our results provide helpful knowledge about the extension rate of this fungus in healthy and wilting PWN-susceptible pine trees in the biological control of PWD and will contribute to the development of a management method for PWD control in the field.Author summaryPine wilt disease, caused by Bursaphelenchus xylophilus, has infected most pine forests in Asian and European forests and led to enormous losses of forest ecosystem and economy. Esteya vermicola is a bio-control fungus against pinewood nematode, showed excellent control efficient to pine wilt disease in both of greenhouse experiments and field tests. Although this bio-control agent was well known for the management of pine wilt disease, the infection mechanism of fungal infection and colonization of host pine tree are less understand. Here, we use GFP-tagged mutant to investigate the fungal infection to pinewood nematode; additionally, the temporal and spatial dynamics of E. vermicola colonize to pine tree were determined by the TaqMan real-time PCR quantification, as well as the response to pinewood nematode invasion. We found a specific infection approach used by E. vermicola to infect B. xylophilus and specialized fungal parasitic cells in PWN infection. In addition, the fungal germination and extension inside of pine tree xylem after inoculation were revealed. In addition, the quantity of E. vermicola increased as response to pinewood nematode invasion was reported. Our study provides two novel technologies for the visualization and detection of E. vermicola for the future investigations of fungal colonization and its parasitism against pinewood nematode, and the mechanisms of the bio-control process.

scholarly journals Optimization of factors affecting the rooting of pine wilt disease resistant Masson pine (Pinus massoniana) stem cuttings

PLoS ONE ◽  
2021 ◽  
Vol 16 (9) ◽  
pp. e0251937
Author(s):  
Ting Pan ◽  
Xue-lian Chen ◽  
Yan-ping Hao ◽  
Chun-wu Jiang ◽  
Song Wang ◽  
...  
Keyword(s):  
Pine Wilt Disease ◽  
Pinus Massoniana ◽  
Wilt Disease ◽  
Stem Cuttings ◽  
Simple Method ◽  
Masson Pine ◽  
Factors Affecting ◽  
Pine Wilt ◽  
Pine Trees ◽  
Root Quality

Pine wilt disease (PWD) is a devastating disease affecting trees belonging to the genus Pinus. To control the spread of PWD in the Masson pine forest in China, PWD resistant Masson pine clones have been selected by the Anhui Academy of Forestry. However, because Masson pine is a difficult-to-root species, producing seedlings is challenging, especially from trees older than 5 years of age, which impedes the application of PWD resistant clones. In this study, we investigated the factors affecting rooting of PWD resistant clones and established a cheap, reliable, and simple method that promotes rooting. We tested the effects of three management methods, four substrates, two cutting materials, two cutting treatments, and three collection times on the rooting of cuttings obtained from 9-year-old PWD resistant clones. Rooting was observed only in stem cuttings treated with the full-light automatic spray management method. Additionally, stem cuttings showed a significantly higher rooting rate and root quality than needles cuttings. Compared with other substrates, stem cuttings planted in perlite produced the longest adventitious root and the highest total root length and lateral root number. Moreover, stem cuttings of PWD resistant clones collected in May showed a significantly higher rooting rate and root quality than those collected in June and July. Moreover, stem cuttings prepared with a horizontal cut while retaining the needles showed significantly higher rooting rate and root quality than those prepared with a diagonal cut while partly removing the needles. This study promotes the reproduction of seedlings of PWD-resistant Masson pine clones which helps control the spread of PWD, meanwhile, provides a technical reference for the propagation of mature pine trees via cuttings.

scholarly journals Early detection of pine wilt disease in Pinus tabuliformis in North China using a field portable spectrometer and UAV-based hyperspectral imagery

2021 ◽  
Vol 8 (1) ◽  
Author(s):  
Run Yu ◽  
Lili Ren ◽  
Youqing Luo
Keyword(s):  
Early Stage ◽  
Vegetation Indices ◽  
Pine Wilt Disease ◽  
Wilt Disease ◽  
Pinus Tabulaeformis ◽  
Pine Tree ◽  
Preventative Measures ◽  
Pine Wilt ◽  
Pine Trees ◽  
Airborne Vehicle

Abstract Background Pine wilt disease (PWD) is a major ecological concern in China that has caused severe damage to millions of Chinese pines (Pinus tabulaeformis). To control the spread of PWD, it is necessary to develop an effective approach to detect its presence in the early stage of infection. One potential solution is the use of Unmanned Airborne Vehicle (UAV) based hyperspectral images (HIs). UAV-based HIs have high spatial and spectral resolution and can gather data rapidly, potentially enabling the effective monitoring of large forests. Despite this, few studies examine the feasibility of HI data use in assessing the stage and severity of PWD infection in Chinese pine. Method To fill this gap, we used a Random Forest (RF) algorithm to estimate the stage of PWD infection of trees sampled using UAV-based HI data and ground-based data (data directly collected from trees in the field). We compared relative accuracy of each of these data collection methods. We built our RF model using vegetation indices (VIs), red edge parameters (REPs), moisture indices (MIs), and their combination. Results We report several key results. For ground data, the model that combined all parameters (OA: 80.17%, Kappa: 0.73) performed better than VIs (OA: 75.21%, Kappa: 0.66), REPs (OA: 79.34%, Kappa: 0.67), and MIs (OA: 74.38%, Kappa: 0.65) in predicting the PWD stage of individual pine tree infection. REPs had the highest accuracy (OA: 80.33%, Kappa: 0.58) in distinguishing trees at the early stage of PWD from healthy trees. UAV-based HI data yielded similar results: the model combined VIs, REPs and MIs (OA: 74.38%, Kappa: 0.66) exhibited the highest accuracy in estimating the PWD stage of sampled trees, and REPs performed best in distinguishing healthy trees from trees at early stage of PWD (OA: 71.67%, Kappa: 0.40). Conclusion Overall, our results confirm the validity of using HI data to identify pine trees infected with PWD in its early stage, although its accuracy must be improved before widespread use is practical. We also show UAV-based data PWD classifications are less accurate but comparable to those of ground-based data. We believe that these results can be used to improve preventative measures in the control of PWD.

scholarly journals Chitosan increases Pinus pinaster tolerance to the pinewood nematode (Bursaphelenchus xylophilus) by promoting plant antioxidative metabolism

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Marta Nunes da Silva ◽  
Carla S. Santos ◽  
Ana Cruz ◽  
Adrián López-Villamor ◽  
Marta W. Vasconcelos
Keyword(s):  
Phenolic Compounds ◽  
Pinus Pinaster ◽  
Plant Defence ◽  
Pine Wilt Disease ◽  
Bursaphelenchus Xylophilus ◽  
Nematode Population ◽  
Post Inoculation ◽  
Pinewood Nematode ◽  
Plant Tolerance ◽  
The Moment

AbstractThe pine wilt disease (PWD), for which no effective treatment is available at the moment, is a constant threat to Pinus spp. plantations worldwide, being responsible for significant economic and environmental losses every year. It has been demonstrated that elicitation with chitosan increases plant tolerance to the pinewood nematode (PWN) Bursaphelenchus xylophilus, the causal agent of the PWD, but the biochemical and genetic aspects underlying this response have not been explored. To understand the influence of chitosan in Pinus pinaster tolerance against PWN, a low-molecular-weight (327 kDa) chitosan was applied to mock- and PWN-inoculated plants. Nematode population, malondialdehyde (MDA), catalase, carotenoids, anthocyanins, phenolic compounds, lignin and gene expression related to oxidative stress (thioredoxin 1, TRX) and plant defence (defensin, DEF, and a-farnesene synthase, AFS), were analysed at 1, 7, 14, 21 and 28 days post-inoculation (dpi). At 28 dpi, PWN-infected plants elicited with chitosan showed a sixfold lower nematode population when compared to non-elicited plants. Higher levels of MDA, catalase, carotenoids, anthocyanins, phenolic compounds, and lignin were detected in chitosan-elicited plants following infection. The expression levels of DEF gene were higher in elicited plants, while TRX and AFS expression was lower, possibly due to the disease containment-effect of chitosan. Combined, we conclude that chitosan induces pine defences against PWD via modulation of metabolic and transcriptomic mechanisms related with plant antioxidant system.

scholarly journals Study of symptoms and gene expression in four Pinus species after pinewood nematode infection

2011 ◽  
Vol 9 (2) ◽  
pp. 272-275 ◽  
Author(s):  
Albina R. Franco ◽  
Carla Santos ◽  
Mariana Roriz ◽  
Rui Rodrigues ◽  
Marta R. M. Lima ◽  
...  
Keyword(s):  
Virulent Strain ◽  
Pine Wilt Disease ◽  
Bursaphelenchus Xylophilus ◽  
Wilt Disease ◽  
Pinewood Nematode ◽  
Pine Trees ◽  
Severe Infections ◽  
Genomic Results ◽  
Avirulent Isolate

Pine wilt disease, caused by the pinewood nematode Bursaphelenchus xylophilus (Steiner and Buhrer) Nickle, is originating severe infections in pine trees. The disease is detected when external symptoms appear (e.g. needle chlorosis), but trees could remain asymptomatic for long periods and serve as a long-term host. The primary goal of this study was to assess the effect of inoculation with an avirulent isolate of B. xylophilus (C14-5) on different Pinus spp. seedlings (P. sylvestris, P. nigra, P. pinea and P. pinaster). At the same time, seedlings were also inoculated with a virulent strain, HF, in order to compare the phenotypic and genomic results of the two types of inoculations. The effect of inoculation was determined in terms of expression of various Pinus genes potentially involved in the response to the disease.The results suggest that P. pinea and P. nigra are more resistant to infection by the nematode than P. sylvestris and P. pinaster. The phenotypic and genetic differences were more marked among P. pinea and P. pinaster.

scholarly journals Comparative Transcriptome Analysis of the Pinewood Nematode Bursaphelenchus xylophilus Reveals the Molecular Mechanism Underlying Its Defense Response to Host-Derived α-pinene

2019 ◽  
Vol 20 (4) ◽  
pp. 911 ◽  
Author(s):  
Yongxia Li ◽  
Fanli Meng ◽  
Xun Deng ◽  
Xuan Wang ◽  
Yuqian Feng ◽  
...  
Keyword(s):  
Molecular Mechanisms ◽  
Bursaphelenchus Xylophilus ◽  
Stress Reaction ◽  
Reproduction Rate ◽  
Molecular Response ◽  
Pinewood Nematode ◽  
Comparative Transcriptome ◽  
Pine Tree ◽  
Pine Trees

Bursaphelenchus xylophilus is fatal to the pine trees around the world. The production of the pine tree secondary metabolite gradually increases in response to a B. xylophilus infestation, via a stress reaction mechanism(s). α-pinene is needed to combat the early stages of B. xylophilus infection and colonization, and to counter its pathogenesis. Therefore, research is needed to characterize the underlying molecular response(s) of B. xylophilus to resist α-pinene. We examined the effects of different concentrations of α-pinene on the mortality and reproduction rate of B. xylophilus in vitro. The molecular response by which B. xylophilus resists α-pinene was examined via comparative transcriptomics of the nematode. Notably, B. xylophilus genes involved in detoxification, transport, and receptor activities were differentially expressed in response to two different concentrations of α-pinene compared with control. Our results contribute to our understanding of the molecular mechanisms by which B. xylophilus responds to monoterpenes in general, and the pathogenesis of B. xylophilus.

Pine chemical volatiles promote dauer recovery of a pine parasitic nematode, Bursaphelenchus xylophilus

Parasitology ◽  
2019 ◽  
Vol 147 (1) ◽  
pp. 50-57 ◽  
Author(s):  
Wei Zhang ◽  
Yongxia Li ◽  
Long Pan ◽  
Xuan Wang ◽  
Yuqian Feng ◽  
...  
Keyword(s):  
Parasitic Nematode ◽  
Pine Wilt Disease ◽  
Bursaphelenchus Xylophilus ◽  
Wilt Disease ◽  
Pine Forests ◽  
Pinewood Nematode ◽  
Dispersal Stage ◽  
Pine Trees ◽  
The Common ◽  
Gradient Effects

AbstractPinewood nematode, Bursaphelenchus xylophilus, a pine parasitic nematode, poses a serious threat to its host pine forests globally. When dispersal-stage larvae 4 (dauer, DL4) of B. xylophilus enters the new pine, it moults into propagative adult (dauer recovery) and reproduces quickly to kill the host pine. Here, we found pine chemical volatiles, rather than the common dauer recovery factors of nematodes (e.g. suitable temperatures, nutrient availability or density), promote B. xylophilus dauer recovery. The results showed that volatilization of chemicals in host pines could attract DL4 and promote DL4 recovery. To identify which chemicals promote this process, we determined the stimulated activity of the main volatiles of pines including six monoterpenes and two sesquiterpenes. Results showed that all the six monoterpenes promoted dauer recovery, especially β-pinene and β-myrcene, but the two sesquiterpenes have no effect on the transformation. Furthermore, β-pinene performed gradient effects on dauer recovery. We hypothesized that when DL4 infect pine trees, the pine volatiles released from the feeding wounds are used as chemical signals for DL4 transformation to adult to reproduce and rapidly kill the pines. Our study identified the B. xylophilus dauer recovery chemical signal and may contribute to preventing pine wilt disease.

Resistance of an indigenous biological system against expansion of the invasive nematode, Bursaphelenchus xylophilus, in cool areas of Japan

Nematology ◽  
2008 ◽  
Vol 10 (5) ◽  
pp. 679-687 ◽  
Author(s):  
Shota Jikumaru ◽  
Katsumi Togashi
Keyword(s):  
Air Temperature ◽  
Pinus Densiflora ◽  
Pine Wilt Disease ◽  
Bursaphelenchus Xylophilus ◽  
Vector System ◽  
Insect Vector ◽  
Disease Development ◽  
Biotic Factors ◽  
Pine Tree ◽  
Pine Trees

Abstract Bursaphelenchus xylophilus is the pathogen associated with pine wilt disease (PWD), an infectious disease of pine trees transmitted by cerambycid beetles of the genus Monochamus. Bursaphelenchus xylophilus is an invasive species, whilst B. mucronatus is a native congener and non-pathogenic to pine trees in Japan. To provide experiment evidence of the biotic and/or abiotic resistance to PWD expansion in a cool area of Japan, we inoculated B. xylophilus into healthy pine trees in a Japanese red pine (Pinus densiflora) stand with the indigenous pine tree-B. mucronatus-insect vector system before invasion of B. xylophilus in 1993 and 1994. Extremely cool air temperature with high precipitation and extremely high air temperature with low precipitation were observed in the summers of the 2 years of inoculation, respectively. The cool summer induced a low incidence of PWD and delayed disease development, resulting in the replacement of B. xylophilus by B. mucronatus within diseased trees and the emergence of Monochamus saltuarius beetles carrying B. mucronatus from the trees 2 years after the inoculation. The hot summer induced disease development in trees in the year of inoculation, but such diseased trees did not become infection sources because of the lack of M. alternatus, whose oviposition was synchronised with the period of disease development in pine trees. This study indicated that biotic factors were important in the inhibitory mechanism in a pine forest against the spread of PWD in the stand. Relevant biotic factors were the lack of M. alternatus and the presence of M. saltuarius carrying B. mucronatus.

scholarly journals Virulence of Bursaphelenchus xylophilus Isolated from Naturally Infested Pine Forests to Five Resistant Families of Pinus thunbergii

Plant Disease ◽  
2012 ◽  
Vol 96 (2) ◽  
pp. 249-252 ◽  
Author(s):  
Mitsuteru Akiba ◽  
Makoto Ishihara ◽  
Norio Sahashi ◽  
Katsunori Nakamura ◽  
Mineko Ohira ◽  
...  
Keyword(s):  
Disease Incidence ◽  
Pine Wilt Disease ◽  
Bursaphelenchus Xylophilus ◽  
Pinus Thunbergii ◽  
Breeding Program ◽  
Pinewood Nematode ◽  
Differential Host ◽  
Pine Trees ◽  
Efficient Breeding ◽  
Highly Virulent

Pine wilt disease is one of the most serious epidemic tree diseases in Japan, and resistant pine trees have been developed through a breeding program. To evaluate resistance of resistant families of Japanese black pine, Pinus thunbergii, to the pinewood nematode, Bursaphelenchus xylophilus, isolated from the field, and to determine whether differentiation of pathogenicity to resistant pine families appears in the nematode isolates, seedlings of five resistant pine families were inoculated with 25 nematode isolates. Disease incidence 18 weeks after inoculation was significantly different among nematode isolates and among pine families but there was no interaction effect between nematode isolate and pine family. This indicates that nematode isolates did not have differential host specificity to resistant families of P. thunbergii. Isolate Shimabara, a test isolate of the breeding program, showed the same degree of virulence as the highly virulent isolates frequently used in experiments. However, more virulent isolates than Shimabara were found among the isolates collected from natural pine forest. This indicated that B. xylophilus populations with higher virulence than Shimabara exist in the natural population. These findings are important in development of more efficient breeding procedures for resistant pine trees.

scholarly journals Transformation of Multi-Antibiotic Resistant Stenotrophomonas maltophilia with GFP Gene to Enable Tracking its Survival on Pine Trees

Forests ◽  
2019 ◽  
Vol 10 (3) ◽  
pp. 231 ◽  
Author(s):  
Yu-Long Li ◽  
Cui-Yun Zheng ◽  
Kan-Cheng Liu ◽  
Yang Wu ◽  
Ben Fan ◽  
...  
Keyword(s):  
Stenotrophomonas Maltophilia ◽  
Fluorescent Protein ◽  
Ecological Impact ◽  
Pine Wilt Disease ◽  
Multidrug Resistant ◽  
The Body ◽  
Resistant Bacteria ◽  
Planted Forests ◽  
Masson Pine ◽  
Pine Trees

Pinus massoniana Lamb., commonly known as Masson Pine, is one of the most important tree species for planted forests in China. This species is, however, threatened by pine wilt disease caused by Bursaphelenchus xylophilus. Stenotrophomonas maltophilia (Palleroni & Bradbury 1993) Smal-007, a bacterium isolated from the body surface of native B. xylophilus, was evidenced to possess the ability to prevent and control this disease. In this study, we focus on exploring effective transformation and green fluorescent protein (GFP)-labeling of Smal-007, in order to facilitate its later investigation. The results indicated that the recombination of antibiotic Tp (trimethoprim), and the uncoupling reagent, CCCP (carbonyl cyanide m-chlorophenyl hydrazine), was effective for the transformation of the multidrug-resistant bacterium. An optimal transformation procedure, including electroporation, was established. To the best of our knowledge, this is the first report where such a method was used for S. maltophilia transformation. Furthermore, Smal-007 was labeled by GFP, allowing the monitoring of its survival ability in pine trees. The labeling was robust and recognizable in isolates recovered from pine needles and bark. In summary, our study indicated that combining uncoupling reagents could be a useful approach to finding operative antibiotic markers for the transformation of multidrug-resistant bacteria. In addition, our successful labeling of Smal-007 with GFP could improve the understanding of its ecological impact, when used as a biocontrol agent.

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