Native ectomycorrhizal fungi from the endangered pine rocklands are superior symbionts to commercial inoculum for slash pine seedlings.

The south Florida pine rocklands is a critically endangered, fire-dependent ecosystem dominated by the overstory tree Pinus densa (South Florida slash pine). Because pine recruitment in this ecosystem has proven problematic, restoration efforts need to include replanting slash pine trees. Even though ectomycorrhizal fungi are known to be critical symbionts of young pines and are necessary for the development of healthy pines, virtually nothing is known about these mutualists and their role in pine establishment and survival in the pine rocklands. One approach to improve pine establishment is to grow seedlings in a nursery before outplanting, facilitating early associations with ectomycorrhizae, and therefore improving seedling health. In this study, we compared health metrics (height, stem diameter, final needle length, root length, root colonization, needle greenness, root volume, and root:shoot ratio) of seedlings grown in soil amended with five commercially available mycorrhizal inocula versus field soil collected from three pine rockland fragments. Seedlings grown with native field soil from the pine rocklands generally performed better than those grown with commercial inoculum in all metrics except root length. According to their labels, each commercial inoculum contained between 4 and 10 ectomycorrhizal fungi species. However, no ectomycorrhizal fungi were recovered from two of the inoculum products and only three ectomycorrhizal fungi in total were recovered from the other three products. In contrast, seedlings grown with field soil are associated with ten ectomycorrhizal species. Our results highlight the importance of incorporating native ectomycorrhizal fungi into pine seedling replanting as part of restoration efforts in the pine rocklands.

Responses of soil bacterial and fungal communities to the long-term monoculture of grapevine.

Soil microorganisms are essential for the long-term sustainability of agricultural ecosystems. However, continuous grapevine replanting can disrupt the stability of soil microbial communities. We investigated the bacterial and fungal abundance, diversity, and community composition in rhizosphere soils with continuous grapevine replanting for 5, 6, 7 (Y5, Y6, and Y7; short-term), and 20 (Y20; long-term) years with high-throughput sequencing. Results showed that diversities and abundances of bacterial and fungal communities in Y20 were significantly lower than in other samples. The bacterial and fungal community compositions were markedly affected by the replanting time and planting year. After short-term grapevine replanting, relative abundances of potential beneficial bacteria and harmful fungi in rhizosphere soils were higher compared to long-term planting. Bacterial and fungal communities were significantly correlated with available nitrogen (AN), available phosphorus, available potassium (AK), and pH. AK and AN were the primary soil factors related to the shift of bacterial and fungal communities. Bacterial and fungal co-occurrence patterns were remarkably affected by replanting time, showing that fallow land harbored co-occurrence networks more complex than those in other groups, with the Y20 group showing the lowest complexity. Then, we isolated the dominant fungi in grapevine rhizosphere soil after continuous replanting and verified the harmful effects of three candidate strains through pot experiments. The results showed that 12 days post-treating the soil with fungal spore suspensions significantly inhibited grapevine seedlings' growth, whereas Fusarium solani inhibited plant growth. Overall, we showed that F. solani might be a potentially harmful fungus related to grapevine replant diseases. KEY POINTS: • Continuous grapevine planting reduced soil microbe diversities/abundances. • Beneficial bacteria and harmful fungi increased after short-term replanting. • F. solani may be a harmful fungus related to grapevine replant diseases.

First Report of Mucor spp. Causing Root Rot of Radix pseudstellariae in Guizhou Province of China.

Radix pseudostellariae L. is one of the most common and highly-prized Chinese medicinal plants with various pharmacological effects, and mainly produced in acid soils in the Guizhou and Fujian provinces of southwestern and southeastern China, respectively (Wu et al. 2020). However, consecutive monoculture of R. pseudostellariae results in severe root rot and decline in biomass and quality of underground tubers. Root tubers of R. pseudostellariae are typically planted in December and harvested in next June. Root rot commonly starts developing in May. The disease incidence of root rot was ranging from 37 to 46% in root portions and basal stem of R. pseudostellariae under the consecutive monoculture fields in Shibing County, Guizhou Province, China (108°12'E, 27°03'N) (Li et al. 2017). Severe root rot was observed in Shibing County in May 2018. Infected plants displayed curly, withered, and yellow leaves, blight, retarded growth, root rot, and yield losses. Abundant whitish mycelia were observed on roots and surrounding soil. Two fungal isolates, designated GZ20190123 and GZ20190124, were obtained from symptomatic roots cultured on potato dextrose agar (PDA). The optimum temperature range for growth of the two isolates was 25 to 30°C. The optimum pH range for the growth of GZ20190123 was 5 to 5.5, whereas GZ20190124 grew better between pH 5 to 8.5. The mean mycelial growth rates of GZ20190123 and GZ20190124 at 30°C were 2.1 and 1.5 cm/day, respectively. Conidia of the two isolates were ovoid or obclavate and were produced in single or branched chains. The internal transcribed spacer (ITS) region was amplified with primers ITS1 and ITS4 (White et al. 1990). The sequences were deposited in GenBank as accession No. MN726736 for GZ20190123 and MN726738 for GZ20190124. Sequence comparison revealed 99% (GZ20190123) and 97% (GZ20190124) identity with previously reported isolate xsd08071 of Mucor racemosus Bull. (accession No. FJ582639.1) and isolate BM3 of Mucor fragilis Bainier (accession No. MK910058.1), respectively, which was confirmed by phylogenetic analysis. The two isolates were tested for pathogenicity on R. pseudostellariae. Six roots of R. pseudostellariae were surface-sterilized with 75% ethanol and stab inoculated with mycelia using a sterile toothpick for each isolate. Sterile distilled water was stab inoculated to twelve roots to serve as the control. Treated roots were incubated in a greenhouse with 16 h day length [light intensity 146.5 µmol/(m2·s)] and day/night temperature 26°C/18°C. The inoculated roots showed the expected symptoms on roots and sprouts 7 days after inoculation, whereas the control roots with sprouts did not show any symptom. The fungi were re-isolated from the diseased roots and confirmed as expected M. racemosus or M. fragilis based on the ITS sequences, which satisfied Koch's postulates. Thus, isolate GZ20190123 was identified as M. racemosus and GZ20190124 as M. fragilis. Previously, M. racemosus and M. fragilis have been reported as a pathogen on tomato (Kwon and Hong 2005) and grape (Ghuffar et al. 2018), respectively. To our knowledge, this is the first report of M. racemosus and M. fragilis causing root rot of R. pseudostellariae in southwestern China, where the disease could cause a significant loss to production of this important medicinal plant.

Spectral Data Collection by Dual Field-Of-View System Under Changing Atmospheric Conditions-A Case Study of Estimating Early Season Soybean Populations.

There is an increasing interest in using hyperspectral data for phenotyping and crop management while overcoming the challenge of changing atmospheric conditions. The Piccolo dual field-of-view system collects up- and downwelling radiation nearly simultaneously with one spectrometer. Such systems offer great promise for crop monitoring under highly variable atmospheric conditions. Here, the system's utility from a tractor-mounted boom was demonstrated for a case study of estimating soybean plant populations in early vegetative stages. The Piccolo system is described and its performance under changing sky conditions are assessed for two replicates of the same experiment. Plant population assessment was estimated by partial least squares regression (PLSR) resulting in stable estimations by models calibrated and validated under sunny and cloudy or cloudy and sunny conditions, respectively. We conclude that the Piccolo system is effective for data collection under variable atmospheric conditions, and we show its feasibility of operation for precision agriculture research and potential commercial applications.

Managing declining yields from ageing tea plantations.

Strong growth in the demand for tea requires further increases in the productivity of plantations. Declining or stagnant yields are commonly observed in older plantations. Possible controlling factors for yield decline are reviewed including ageing of plants, chronic disease and sub-optimal soil conditions such as excess soil acidity and low soil organic matter. Management options for addressing these factors are evaluated, including replanting. A systematic approach to decision-making about replanting is presented. Practice for replanting is reviewed and it is concluded that evidence to support a general case for replanting is limited, unless based on the introduction of more productive clones and/or better plant spacing.

Autotoxin affects the rhizosphere microbial community structure by influencing the secretory characteristics of grapevine roots.

Autotoxins secreted by roots into the soil can trigger rhizosphere microecological imbalances and affect root secretory properties resulting in conditions such as replanting disease. However, information on the effect of autotoxins on root secretion characteristics and regulation of the composition of rhizosphere microorganisms by altered root exudates is limited. In this study, autotoxin ρ-hydroxybenzoic acid (4-HBA) was added to the soil of potted grapevine seedlings, CO2 pulse-labeling, and DNA stable isotope probing were used to track the rhizosphere microbiome that assimilates root exudates. Bacterial and fungal microbiomes that assimilated plant-derived carbon were identified by high-throughput sequencing. Results showed that 4-HBA treatment altered bacterial and fungal communities in 13C-labeled organisms, with a lower abundance of beneficial bacteria (e.g., Gemmatimonas, Streptomyces, and Bacillus) and a higher abundance of potential pathogen fungi (e.g., Fusarium, Neocosmospora, Gibberella, and Fusicolla) by changing the composition of root exudates. The exogenous addition of upregulated compound mixtures of root exudates reduced the abundance of beneficial bacterial Bacillus and increased the abundance of potential pathogen fungi Gibberella. These results suggest that 4-HBA can alter root secretion properties and altered root exudates may enrich certain potential pathogens and reduce certain beneficial bacteria, thereby unbalancing the structure of the rhizosphere microbial community.

Geneva® Series Rootstocks for Apple Trees Under Extreme Replanting Conditions in Southern Brazil.

Geneva® rootstocks in Brazil are known to be efficient in controlling vigor, and are precocious and resistant to diseases. The objective of this study was to evaluate the performance of apple tree cultivars grafted on the Geneva® rootstocks in severe replant disease areas, by planting 60 days after the eradication. The experiments were implemented in 2017, in São Joaquim and Vacaria. The Gala Select and Fuji Suprema cultivars were grafted onto 'G.202', 'G.814', 'G.210', and 'G.213' rootstocks in the Tall Spindle training system. In 2018/2019, total thinning was carried out to promote plant growth. In São Joaquim, partial thinning was carried out in 2019/2020 harvest of 'Gala Select'. The rootstocks were divided into two groups based on vigor, for both areas and cultivars. 'G.202' and 'G.213' were 40% less vigorous than 'G.210' and 'G.814'. For 'Gala Select', the extreme non-fallow condition mainly affected the vigor and productivity of 'G.213' in both areas. At the end of two harvests, 'G.213' was 17% less productive than 'G.210', contrary to what is observed in areas where the fallow period is respected. However, 'G.213' confirmed a greater yield efficiency, which was 27% higher than 'G.210'. This suggests that a perspective of forecasting production for the third crop is higher for 'G.213' than for 'G.210'. In the case of 'Fuji Suprema', the G.210 rootstock was the most productive in both areas. In São Joaquim, 'G.202' matched 'G.210' in productivity and efficiency as it sprouts better in colder regions. Considering the fruit quality, 'G.213' anticipated the maturation with fruits of larger size and higher total soluble solids (TSS) in both areas and cultivars, making it possible to anticipate the harvest. It was concluded that the non-fallow condition does not alter the relative differences in vigor and fruit quality among the rootstocks. However, notwithstanding the overall replant tolerance of these rootstocks, it does reduce productivity by mainly affecting less vigorous rootstocks that need about three crops to overcome the allelopathic effects of the soil and start growing normally. The G.210 semi-dwarfing rootstock is an alternative for the immediate conversion of apple orchards of Gala Select and Fuji Suprema cultivars in southern Brazil.

Overexpression of RgPAL family genes involved in phenolic biosynthesis promotes the replanting disease development in Rehmannia glutinosa.

Rehmannia glutinosa production is affected by the replanting disease, which involves autotoxic harm mediated by specific endogenous allelochemicals in root exudates. Many phenolics that act as allelochemical agents are mostly phenylpropanoid products of secondary metabolism in plants. Phenylalanine ammonia-lyase (PAL) is the first enzyme that catalyses the deamination of l-phenylalanine for entrance into the phenylpropanoid pathway. PAL family genes have been isolated and functionally characterized in many plant species. However, PAL family genes involved in phenolic biosynthesis remain largely uncharacterized in R. glutinosa. Here, we identified and characterized four PAL family genes (RgPAL2 to RgPAL5) in the species whose sequences exhibited highly conserved domains of PALs according to in silico analysis, implying their potential function in phenolic biosynthesis. Overexpression of RgPALs in R. glutinosa enhanced phenolic production, verifying that RgPAL family genes participate in phenolic biosynthesis pathways. Moreover, we found that the release of several allelopathic phenolics from the roots of RgPAL-overexpressing transgenic R. glutinosa increased, implying that the RgPALs positively promote their release. Importantly, under continuous monoculture stress, we found that the RgPAL transgenic plants exhibited more significant autotoxic harm than did non-transgenic (WT) plants by activating the phenolics/phenylpropanoid pathway, indicating that RgPAL family genes function as positive regulators of the replanting disease development in R. glutinosa. This study revealed that RgPAL family genes are involved in the biosynthesis and release of several phenolics and positively control the replanting disease development in R. glutinosa, laying a foundation for further clarification of the molecular mechanisms underlying the disease formation.

Influence of the level of arterial resection on the replanting and revascularization results in hand surgery: prospective study over 22 months.

The objective of this work was to assess whether the injury mechanisms are responsible for histological arterial lesions. This prospective single-center study included adults with wrist or hand arterial injury. Arterial resection of at least 2 mm from the proximal and distal stumps was performed before the arterial anastomosis. Histological analysis of the arterial stumps was performed. An ultrasound was performed 1 month postoperatively to check arterial patency. A clinical and functional evaluation was done at 1 month postoperative, then every 3 months. From 2018 to 2020, 46 patients were included with a maximum follow-up of 13 months. There were 35 cuts, 2 crush injuries, 8 amputation and 1 blast injury. Macroscopically, 37% of the margins were considered damaged. Histological analysis showed significant damage in 59% of the sections (27 out of 46 patients) with 50% for crush injury, 55% for cuts by mechanical tool, 62% for cuts by power tool, 62% for amputations and 100% for blasts. The failure rate was 9%: 2 replantations and 2 asymptomatic thromboses diagnosed by ultrasound. Postoperative pain on VAS was 1.75/10, range of motion was 87%, Quick DASH was 8%, SF36 PCS was 69% and SF36 MCS was 70%. Factors influencing the success or failure of anastomosis were the mechanism of injury (p = 0.02), associated nerve damage (p = 0.014) and length of proximal arterial cut (p = 0.046). Histological arterial lesions seem to correlate with the injury mechanism. Cuts caused by glass or crush injuries do not seem to require arterial resections of more than 2 mm. A continuation of the study with a larger number of subjects may generate statistically significant results.

Recent Advances in Saffron Soil Remediation: Activated Carbon and Zeolites Effects on Allelopathic Potential.

Saffron (Crocus sativus L.) is a highly valuable plant. Iran provides nearly 90% of the world's total saffron and is the biggest global producer. The allelopathic effects of saffron corm (SC) and saffron field soil (SFS) have been hypothesized to play an important role in replanting. Recently, adsorbent materials have been used to neutralize the effects of allelochemicals. These materials, including activated carbon and zeolite, have large surface areas, pore volumes, as well as tremendous adsorptive capacity and complex chemical and physical properties. In this study, three independent experiments were conducted. In the first test, the allelopathic effects of aqueous and methanolic extracts of SC remnant and 9-year-old SFS as well as filtered aqueous extract of soil were investigated. In the second assay, the effects of SC remnants and SFS with different ages (i.e., 4, 6, and 9 years old) in combination with adsorbents were examined on the germination and growth of lettuce (Lactuca sativa L.) seedlings by the sandwich method. In the third experiment, we examined the effects of SC remnants combined with adsorbents on lettuce growth parameters. Our results showed that the allelopathic effects of aqueous and methanolic extracts of SC remnant were significantly superior to those of 9-year-old SFS. The aqueous extract of SC remnant reduced the root length of lettuce by 50%. The use of activated carbon and zeolites significantly decreased the observed allelopathic effect. Moreover, lettuce growth in rhizosphere soil was significantly inhibited by SC remnant and SFS extracts. The allelopathic effects of SC remnants caused a growth imbalance between the shoot and roots. Based on biochemical analyses, using the adsorbents increased the carotenoid content and chlorophyll index of lettuce by 23.33% and 5.25%, respectively. Adsorbents may play a role in treating soils contaminated by allelochemicals.

Rehmannia glutinosa Replant Issues: Root Exudate-Rhizobiome Interactions Clearly Influence Replant Success.

Production of medicinal tubers of Rehmannia glutinosa is severely hindered by replanting issues. However, a mechanistic understanding of the plant-soil factors associated with replant problems is currently limited. Thus, we aimed to identify the R. glutinosa root exudates, evaluate their potential phytotoxicity and profile the interactions between the plant and its associated rhizobiome. Stereomicroscopy and liquid chromatography coupled to a quadrupole/time of flight mass spectrometer were used to monitor and identify secreted metabolites, respectively. Seedling bioassays were used to evaluate the phytotoxicity of R. glutinosa root exudates. Two complimentary experiments were performed to investigate allelochemical fate in rhizosphere soil and profile the associated microbiota. Root specific microbes were further isolated from R. glutinosa rhizosphere. Impacts of isolated strains were evaluated by co-cultivation on plate and on seedlings in tissue culture, with a focus on their pathogenicity. Interactions between key R. glutinosa root exudates and isolated rhizobiomes were investigated to understand the potential for plant-soil feedbacks. Quantification and phytotoxic analysis of metabolites released from R. glutinosa indicated catalpol was the most abundant and bioactive metabolite in root exudates. Subsequent microbial profiling in soil containing accumulated and ecologically significant levels of catalpol identified several taxa (e.g., Agromyces, Lysobacter, Pseudomonas, Fusarium) that were specifically shifted. Isolation of R. glutinosa rhizobiomes obtained several root specific strains. A significant antagonistic effect between strain Rh7 (Pseudomonas aeruginosa) and two pathogenic strains Rf1 (Fusarium oxysporum) and Rf2 (Fusarium solani) was observed. Notably, the growth of strain Rh7 and catalpol concentration showed a hormesis-like effect. Field investigation further indicated catalpol was increasingly accumulated in the rhizosphere of replanted R. glutinosa, suggesting that interactions of biocontrol agents and pathogens are likely regulated by the presence of bioactive root exudates and in turn impact the rhizo-ecological process. In summary, this research successfully monitored the release of R. glutinosa root exudates, identified several abundant bioactive R. glutinosa secreted metabolites, profiled associated root specific microbes, and investigated the plant-soil feedbacks potentially regulated by catalpol and associated rhizobiomes. Our findings provide new perspectives toward an enhanced understanding R. glutinosa replant problems.

Biochar mediates microbial communities and their metabolic characteristics under continuous monoculture.

Biochar amendment has been extensively used to improve plant performance and suppress disease in monoculture systems; however, few studies have focused on the underlying control mechanisms of replanting disease. In this study, we assessed the effects of biochar application on Radix pseudostellariae plant growth, rhizosphere soil microbial communities, and the physiological properties of microorganisms in a consecutive monoculture system. We found that biochar addition had little impact on the physiological parameters of tissue cultures of R. pseudostellaria but did significantly mediate microbial abundance in the rhizosphere soil of different consecutive monoculture years, leading to decreases in the abundance of pathogenic Fusarium oxysporum, Talaromyces helicus, and Kosakonia sacchari. Furthermore, biochar amendment had negative effects on the growth of beneficial bacteria, such as Burkholderia ambifaria, Pseudomonas chlororaphis, and Bacillus pumilus. Metabolomic analysis indicated that biochar significantly influenced the metabolic processes of F. oxysporum while inhibiting the mycelial growth and abating the virulence on plants. In summary, this study details the potential mechanisms responsible for the biochar-stimulated changes in the abundances and metabolism of rhizosphere bacteria and fungi, decreases in the contents of pathogens, and therefore improvements in the environmental conditions for plants growth. Further research is needed to evaluate the effects of biochar in long-term field trials.

Optimal control of a vectored plant disease model for a crop with continuous replanting.

Vector-transmitted diseases of plants have had devastating effects on agricultural production worldwide, resulting in drastic reductions in yield for crops such as cotton, soybean, tomato, and cassava. Plant-vector-virus models with continuous replanting are investigated in terms of the effects of selection of cuttings, roguing, and insecticide use on disease prevalence in plants. Previous models are extended to include two replanting strategies: frequencyreplanting and abundance-replanting. In frequency-replanting, replanting of infected cuttings depends on the selection frequency parameter ε, whereas in abundance-replanting, replanting depends on plant abundance via a selection rate parameter also denoted as ε. The two models are analysed and new thresholds for disease elimination are defined for each model. Parameter values for cassava, whiteflies, and African cassava mosaic virus serve as a case study. A numerical sensitivity analysis illustrates how the equilibrium densities of healthy and infected plants vary with parameter values. Optimal control theory is used to investigate the effects of roguing and insecticide use with a goal of maximizing the healthy plants that are harvested. Differences in the control strategies in the two models are seen for large values of ε. Also, the combined strategy of roguing and insecticide use performs better than a single control.

Protocol for the Micropropagation of Coconut from Plumule Explants.

Coconut is a crop that is economically important in several countries throughout the world. Unfortunately, production is decreasing because palms are affected by very serious phytoplasma diseases, such as lethal yellowing, and also most of coconuts are already very old. On the other hand, markets for coconut products have been rapidly growing in recent years. Hence, replanting of most cultivation surface worldwide, as well as establishing new surface, is urgently needed. This is an immense task, requiring at least a billion coconut palms that cannot be accomplished by traditional propagation through seed. Therefore the biotechnological alternative of micropropagation by somatic embryogenesis is needed. Research has been carried out on this subject in laboratories in several countries studying different approaches, testing different types of explants. The most responsive tissue has been plumule from zygotic embryos. A protocol for micropropagation of coconut based on plumule explants is described here. It involves the use of different media that are based on Y3 medium complemented with activated charcoal, gelling agent, sucrose, and growth regulators. These media allow the formation of embryogenic callus and somatic embryos, growth of shoots, and development of plantlets.

Rhizospheric microbial communities are driven by Panax ginseng at different growth stages and biocontrol bacteria alleviates replanting mortality.

The cultivation of Panax plants is hindered by replanting problems, which may be caused by plant-driven changes in the soil microbial community. Inoculation with microbial antagonists may efficiently alleviate replanting issues. Through high-throughput sequencing, this study revealed that bacterial diversity decreased, whereas fungal diversity increased, in the rhizosphere soils of adult ginseng plants at the root growth stage under different ages. Few microbial community, such as Luteolibacter, Cytophagaceae, Luteibacter, Sphingomonas, Sphingomonadaceae, and Zygomycota, were observed; the relative abundance of microorganisms, namely, Brevundimonas, Enterobacteriaceae, Pandoraea, Cantharellales, Dendryphion, Fusarium, and Chytridiomycota, increased in the soils of adult ginseng plants compared with those in the soils of 2-year-old seedlings. Bacillus subtilis 50-1, a microbial antagonist against the pathogenic Fusarium oxysporum, was isolated through a dual culture technique. These bacteria acted with a biocontrol efficacy of 67.8%. The ginseng death rate and Fusarium abundance decreased by 63.3% and 46.1%, respectively, after inoculation with B. subtilis 50-1. Data revealed that microecological degradation could result from ginseng-driven changes in rhizospheric microbial communities; these changes are associated with the different ages and developmental stages of ginseng plants. Biocontrol using microbial antagonists alleviated the replanting problem.