Rootstock genotype and orchard replant position rather than soil fumigation or compost amendment determine tree growth and rhizosphere bacterial community composition in an apple replant soil

Apple replant disease (ARD) is a complex soilborne disease syndrome that often causes problems when renovating old orchard sites. Soil fumigants sometimes control ARD, but biological and cultural alternatives are needed. In this study the growth of two widely used clonal apple (Malus domestica) rootstocks (M7 and M26) were compared to three new rootstocks from the Cornell-Geneva series (CG16, CG30 and CG210 (a.k.a. CG6210)) in an orchard site with a history of ARD, in Ithaca, NY. Trees were planted in two distinguishable positions – the previous tree rows versus the old inter-row grass lanes. Additionally, we compared the effects of compost amendment and fumigation with dichloropropene plus chloropicrin on tree growth on each replant rootstock. Rhizosphere bacteria and actinobacteria were assessed using PCR-DGGE for the rootstocks M7, M26, CG30 and CG210. Tree growth on the rootstocks M7, M26 and CG16 was suppressed in the old tree rows relative to grass lanes, while trees on CG30 and CG210 rootstocks grew equally well in both positions. The species composition of rhizosphere bacteria and actinobacteria differed significantly between the planting positions and between the rootstocks M7 and M26 compared to CG30 and CG210. In contrast, the preplant compost or fumigation soil treatments had no effect on tree growth and little impact on rhizosphere bacterial community composition. Abbreviations: ARD – apple replant disease; DGGE – denaturing gradient gel electrophoresis; PCR – polymerase chain reaction.     

Growth rate of fine root systems influences rootstock tolerance to replant disease

Aims

Plant tolerance to herbivory has often been linked to plant growth rate, with faster growing plants that present high tissue turnover rates expected to be more tolerant than slower-growing plants. We tested whether this relationship also holds for rootstock growth rate and tolerance to apple replant disease (ARD).

Methods

An ARD susceptible rootstock, M.26 and ARD tolerant rootstock, CG.6210 were grown in soil from an apple replant site (FS) and in pasteurized soil (PS) from the same site. Total below ground biomass production was determined by harvesting a subset of plants per soil treatment and rootstock at 11, 17, and 23 weeks after planting. Root samples were collected prior to each harvesting date to determine root respiration and total carbon (C) and nitrogen (N) content. Root dynamics were tracked during the growing season by digitally photographing root observation windows.

Results

Total root biomass, first and second order roots, and second-to-first order root ratio were higher in CG.6210 than in M.26 in both soil treatments. Roots of CG.6210 were thinner and had lower N concentration than those of M.26. Roots of M.26 had longer lifespans than those of CG.6210, and the mortality risk of M.26 roots was 56 % that of CG.6210 roots.

Conclusion

Our study indicates that rootstocks with faster growing root systems can tolerate ARD infection by investing fewer resources in individual root construction that can be shed more readily.     

Linking Soil Biotic and Abiotic Factors to Apple Replant Disease: a Greenhouse Approach

Apple replant disease (ARD) is a frequently occurring plant disease, which causes retarded growth and mortality of young apple trees in replanted orchards. The aetiology is not well understood, but soil‐borne micro‐organisms are often discussed as primary causal agents of the replant problem. A greenhouse study was conducted in Laimburg, Italy, with orchard soils from the region, with the aim of obtaining information about the influence of soil biotic and abiotic factors on the aetiology of the disease. Apple rootstocks (M9) were planted into soils cultivated with apple trees that were either fumigated with chloropicrin or not fumigated, as well as mixtures of fumigated and non‐fumigated soils. In addition, uncultivated soils (from the inter‐row, from a fallow plot and from a meadow) were taken as controls. Various parameters were measured after 62 days in a controlled pot assay. Soils fumigated with chloropicrin resulted in higher apple shoot growth and lower microbial biomass carbon than non‐fumigated soils. Uncultivated soils had generally the highest microbial biomass carbon and the highest ergosterol contents. No considerable differences between basal respiration, ergosterol content, pH, electrical conductivity, and most nutrient and metal contents were observed between fumigated and non‐fumigated soils. Denaturing gradient gel electrophoresis gels of DNA extracted from the soils revealed differences in the fungal, bacterial and actinobacterial communities of the different soils, indicating significant shifts in microbial community composition after chloropicrin treatment. This study indicates biotic factors in soil to be a causal agent of apple replant disease.     

Long-term orchard groundcover management systems affect soil microbial communities and apple replant disease severity

Apple replant disease (ARD) is a soil-disease syndrome of complex etiology that affects apple tree roots in replanted orchards, resulting in stunted tree growth and reduced yields. To investigate whether different groundcover management systems (GMSs) influence subsequent ARD severity, we grew apple seedlings in an outdoor nursery in pots containing orchard soil from field plots where four GMSs had been maintained for 14 years in an orchard near Ithaca, NY, USA. The GMS treatments were: (1) pre-emergence herbicide (Pre-H), bare soil strips maintained by applying tank-mixed glyphosate, norflurazon and diuron herbicides annually; (2) post-emergence herbicide (Post-H), sparse weed cover maintained by applying glyphosate in May and July each year; (3) mowed sod grass (Mowed Sod); and (4) bark mulch (Mulch). Soils were also sampled from the grass drive lane maintained between the trees in the orchard (Grass Lane). Sampled soils (Orchard soil) were either pasteurized or left untreated, placed into 4-L pots, and planted with one apple seedling per pot. After 3 months of growth, soil (Bioassay soil) and apple tree roots (Bioassay roots) were sampled from each pot and microbial populations colonizing samples were characterized. Seedling growth was reduced in soils sampled from all four GMS treatments compared to the Grass Lane soils. Among the GMS treatments, seedling biomass was greater in Pre-H than in the Post-H soil. Soil microbial communities and nutrient availability differed among all four GMS treatments and the Grass Lane. Root-lesion (Pratylenchus sp.) nematode populations were higher in the Mowed Sod than in the other GMS treatments. Soil bacterial and fungal community composition was assessed in Orchard and Bioassay soils and Bioassay roots with a DNA fingerprinting method (T-RFLP). Redundancy analysis indicated that soils sampled from the different GMS treatments differentially influenced seedling biomass. A clone library of 267 soil bacteria was developed from sampled Orchard soils and Bioassay roots. These communities were dominated by Acidobacteria (25% of sequences), Actinobacteria (19%), δ-Proteobacteria (12%), β-Proteobacteria (10%), and these ratios differed among the GMS soils. Members of the family Comamonadaceae were detected only in tree-row soil, not in the Grass Lanes. The dominant sequences among 145 cloned fungi associated with apple seedling roots were Fusarium oxysporum (16% of sequences), an uncultured soil fungus submitted under DQ420986 (12%), and Rhodotorula mucilaginosa (9%). In a redundancy analysis, factors including fungal and oomycete community compositions, soil respiration rates, population sizes of culturable bacteria and fungi, soil organic matter content, and nutrient availability, were not significant predictors of apple seedling biomass in these soils. Different GMS treatments used by apple growers may influence subsequent ARD severity in replanted trees, but edaphic factors commonly associated with soil fertility may not reliably predict tree-root health and successful establishment of replanted orchards.     

Effect of rootstock on apple (Malus domestica) tree water relations

The effects of rootstock on mid-season water relations, under orchard conditions of non-limiting soil moisture, were determined for bearing 'Empire' apple trees ( Malus domestica Borkh.) on the clonal rootstocks M9, M26, M7, MM106, and MM104 (most to least dwarfing) in their sixth and seventh growing seasons. Stem water potentials (ψ^stem) of trees on M9 and M26 were more negative at midday, under warm, sunny conditions, than were the trees on the other three rootstocks. However, change in ψ^stem per change in stem distance through the canopy (water potential gradient) did not vary among rootstocks at midday. There was no rootstock effect on diurnal variation in transpiration or stomatal conductance. Differences in water storage capacitance, relative to tree size, were determined in a separate study but did not account for the differences observed in ψ^stem. Calculated hydraulic conductivities of xylem water transport suggest that rootstocks differ in their ability to conduct water to the scion, but hydraulic conductivity of the scion was not affected by rootstock. Root-stock differences in hydraulic conductivity were not accounted for by differences in tree size.     

Fine-root system development and susceptibility to pathogen colonization

Root development may exert control on plant–pathogen interactions with soil-borne pathogens by shaping the spatial and temporal availability of susceptible tissues and in turn the impact of pathogen colonization on root function. To evaluate the relationship between root development and resistance to apple replant disease (ARD) pathogens, pathogen abundance was compared across root branching orders in a bioassay with two rootstock genotypes, M.26 (highly susceptible) and CG.210 (less susceptible). Root growth, anatomical development and secondary metabolite production were evaluated as tissue resistance mechanisms. ARD pathogens primarily colonized first and second order roots, which corresponded with cortical tissue senescence and loss in second and third order roots. Defense compounds were differentially allocated across root branching orders, while defense induction or stress response was only detected in first order and pioneer roots. Our results suggest disease development is based largely on fine-root tip attrition. In accordance, the less susceptible rootstock supported lower ARD pathogen abundance and altered defense compound production in first order and pioneer roots and maintained higher rates of root growth in both the ARD soil and pasteurized control compared to the more susceptible. Thus, this rootstock’s ability to maintain shoot growth in replant soil may be attributable to relative replant pathogen resistance in distal root branches as well as tolerance of infection based on rates of root growth.     

Factors promoting infestation of newly planted, nonbearing apple orchards by dogwood borer (Lepidoptera: Sesiidae)

The initiation and level of infestation by dogwood borer, Synanthedon scitula (Harris), was tracked over three consecutive years in two nonbearing apple (Malus spp.) orchards in West Virginia and Virginia. The orchards were planted on a number of rootstock-variety (approximately cultivar) combinations and grown using different cultural practices. Infestations were detected during the first season after planting and continued to increase annually. The amount of burr knot tissue had the greatest impact on dogwood borer populations, because increasing amounts of burr knot tissue resulted in higher infestation rates. The use of plastic spiral wrap tree guards seemed to increase the development of burr knot tissue, resulting in significantly greater infestation compared with trees without tree guards in the West Virginia orchard. Variety also had a significant effect, because 'Idared' trees on M.26 had significantly greater levels of infestation compared with 'Buckeye Gala' on M.26, with or without tree guards, in the Virginia orchard. Mounding soil around the rootstock to a height just above the graft union prevented or tremendously curtailed infestation by dogwood borer, but it led to scion rooting that seemed to have an impact on size-controlling features of dwarfing rootstocks. Removal of the mounds at the beginning of the third growing season resulted in infestation of the rooted tissue during the same season. As long as apple cultivars continue to be planted on size-controlling rootstocks, dogwood borer will likely remain a serious pest, requiring either chemical treatments or a behavioral control strategy, such as mating disruption, to protect trees from infestation and damage.     

环渤海湾苹果产区老果园与连作果园土壤线虫群落特征

应用类群多样性、类群丰富度、个体密度和功能类群指数等群落参数,于2009年5-10月分3次对环渤海湾苹果栽植区老果园不同位置和连作果园取样研究了土壤线虫群落特征。结果表明:苹果连作对果园土壤生物环境具有明显的恶化作用,连作果园土壤环境条件恶劣;对功能类群指数的统计表明,苹果连作明显改变了果园土壤中植物寄生性线虫r-选择和k-选择的比例,与自由生活线虫比较,连作对植物寄生线虫影响更明显;连作提高了果园的土壤线虫PPI/MI,说明连作对果园土壤健康的扰动最大;土壤不同食性线虫数量统计结果显示,环渤海湾苹果栽植区植食性线虫数量未达线虫伤害阈值,线虫不是引起环渤海湾地区苹果连作障碍的主要原因。     

复合微生物菌剂对重茬苹果园土壤细菌群落的影响

【目的】了解抗重茬复合微生物菌剂对重茬苹果园土壤细菌数量及多样性的影响。【方法】对重茬苹果园土壤进行复合微生物菌剂处理后,采用自动核糖体内转录间隔区序列分析方法(Automated ribosomal intergenic spacer analysis,ARISA)对新茬苹果园、重茬苹果园土壤细菌群落多样性进行分析。【结果】经复合微生物菌剂处理2年后,重茬果园土壤OTUs总数和Shannon-Weiner指数分别为250和4.44,新茬果园OTUs总数和Shannon-Weiner指数分别为234和3.81,经One-Way ANOSIM法分析得到二者差异系数P分别为0.108 3和0.084 3。另外,重茬园和新茬园共享核心OTUs有89个,二者的Jaccard群落相似性系数为0.47,相似程度中等。【结论】复合微生物菌剂具有提高重茬果园土壤微生物多样性,促进重茬苹果园土壤微生物生态系统恢复的作用。     

Tissue age,orchard location and disease management influence the composition of fungal and bacterial communities present on the bark of apple trees

Plants host microbial communities that can be affected by environmental conditions and agronomic practices. Despite the role of bark as a reservoir of plant pathogens and beneficial microorganisms, no information is available on the effects of disease management on the taxonomic composition of the bark-associated communities of apple trees. We assessed the impact of disease management strategies on fungal and bacterial communities on the bark of a scab-resistant apple cultivar in two orchard locations and for two consecutive seasons. The amplicon sequencing revealed that bark age and orchard location strongly affected fungal and bacterial diversity. Microbiota dissimilarity between orchards evolved during the growing season and showed specific temporal series for fungal and bacterial populations in old and young bark. Disease management did not induce global changes in the microbial populations across locations and seasons, but specifically affected the abundance of some taxa according to bark age, orchard location and sampling time. Therefore, the disease management applied to scab-resistant cultivars, which is based on a limited use of fungicides, partially changed the taxonomic composition of bark-associated fungal and bacterial communities, suggesting the need for a more accurate risk assessment regarding possible pathogen outbreaks.     

Phytophthora collar rot of apple: influence of the rootstock on scion variety resistance

In field experiments with young trees great differences were found in the resistance to Phytophthora cactorum of Cox's Orange Pippin apple scions grafted on different clonal rootstocks. The rootstock effect on scion resistance was inversely related to the effect on tree vigour: the rootstocks inducing high resistance were dwarfing (M. 9) or semi-dwarfing (M. 7, M. 26, MM. 106), and those inducing low resistance were vigorous or very vigorous (M. 25, MM. 104, MM. 109). Mean lesion lengths in Cox on MM. 104 were five to eight times greater than those in Cox on M. 9. The rootstock influence on scion resistance was associated primarily with effects on the rate of lesion extension: during the early stages only of host colonization there appeared to be threshold extension rates below which host resistance factors effectively suppressed a large proportion of infections. The influence of the root-stock on scion resistance was apparently unrelated to inherent rootstock resistance. On all rootstocks Cox showed diminished resistance to infection during the period from the swelling of buds to the early stages of shoot growth. Although most susceptible during the ‘mouse-ear’ and ‘pink bud’ stages of development, suscpetibility was not associated with flowering per se. Rootstock type did not affect the resistance of Cox scions to P. syringae, for which the period of susceptibility to infection occurred in the dormant season.     

Global RNA association with the transcriptionally active chromosome of chloroplasts

Key message

Gene expression studies in roots of apple replant disease affected plants suggested defense reactions towards biotic stress to occur which did not lead to adequate responses to the biotic stressors.

Abstract

Apple replant disease (ARD) leads to growth inhibition and fruit yield reduction in replanted populations and results in economic losses for tree nurseries and fruit producers. The etiology is not well understood on a molecular level and causal agents show a great diversity indicating that no definitive cause, which applies to the majority of cases, has been found out yet. Hence, it is pivotal to gain a better understanding of the molecular and physiological reactions of the plant when affected by ARD and later to overcome the disease, for example by developing tolerant rootstocks. For the first time, gene expression was investigated in roots of ARD affected plants employing massive analysis of cDNA ends (MACE) and RT-qPCR. In reaction to ARD, genes in secondary metabolite production as well as plant defense, regulatory and signaling genes were upregulated whereas for several genes involved in primary metabolism lower expression was detected. For internal verification of MACE data, candidate genes were tested via RT-qPCR and a strong positive correlation between both datasets was observed. Comparison of apple ‘M26’ roots cultivated in ARD soil or γ-irradiated ARD soil suggests that typical defense reactions towards biotic stress take place in ARD affected plants but they did not allow responding to the biotic stressors attack adequately, leading to the observed growth depressions in ARD variants.

     

L'epoca Della Differenziazione Delle Gemme Fiorali Nell'olivo

Abstract

Calcium, as in general for all plants, is very important for fruit trees and a tight correlation between leaf content and cropping efficiency has been found.

The annual removal of calcium oxide of an orchard has been estimated in kg per hectare: pear 200, apple 180, peach 150, grape 60–130, olive 35–70, kiwi 55–60.

The rootstock affects the calcium uptake from the soil and content of the scion; frequently a higher calcium content is found in trees grafted an dwarfing rootstocks as pear on quince, apple on M9 and M26, peach on Damas.

By the horticultural point of view, calcium is responsible of two main problems: chlorosis due to high active Ca content in the soil and bitter pit, on the apposite, due to a low Ca level in the fruits.

From soil and leaf studies it seems clear that lime-induced Fe chlorosis results from two conditions: a) slow availability of Fe in the soil, and b) immobilization of Fe in the tree in forms that are not available for chlorophyll formation.

Breeding tolerant rootstocks has been the practical solution of chlorosis for most of the susceptible species.

Bitter pit is a physiological disorder of apple fruits, sometime already evident before picking, more frequently after harvesting, during the storage.

The prevention of the disorder is, essentially, a good horticultural practice (pruning, fertilization, irrigation, fruit thinning). Very effective are also Ca sprays as chloride or nitrate, or citrate, or phosphate, starting after setting, 4–5 times every 10–12 days.     

Effects of Rootstocks and Interstocks on the Xylem Sap Composition in Apple Trees: Effects on Nitrogen, Phosphorus, and Potassium Content

This paper records the concentration of N, P, and K in sap whichexuded from the cut stumps of composite orchard apple trees.The trees were either single-worked on different rootstocksor incorporated various interstocks. While the volume of sap exuded per unit area of cut surfacedid not appear to vary with rootstock or interstock, exudatefrom the dwarfing rootstocks or interstocks contained depletedlevels of nutrients. A possible mechanism for rootstock and interstock effects isdiscussed in the light of these results.     

连作苹果园土壤真菌的T-RFLP分析

为探讨连作苹果园不同土壤空间真菌群落结构,应用T-RFLP(Terminal Restriction Fragment Length Polymorphism)技术,比较了3个连作园不同取样位置(行间、原穴、株间)和不同土层(0—30 cm、30—60 cm)的土壤真菌多样性,并结合不同样品TRFLP图谱的差异,采用多样性指数分析、聚类分析和主成分分析(PCA),分析了3个连作园土壤真菌群落结构特征。结果表明,磁窑、道朗和金城连作园的土壤真菌多样性存在差异,各采样地区的Shannon多样性指数在0.43—2.47之间,Pielou均匀度指数在0.17—0.85之间,Simpson优势度指数在0.12—0.81之间,Margalef丰富度指数最高的是金城树穴0—30 cm土层(R=4.55),最低的是磁窑行间30—60 cm土层(R=0.77)。在调查的不同取样位置、不同土层中,原树穴具有最高的多样性指数、均匀度指数、丰富度指数和最低的优势度指数;0—30 cm土层的土壤真菌多样性指数、均匀度指数、丰富度指数均高于30—60cm土层,而优势度指数的趋势正好相反;PCA和聚类分析结果显示磁窑、道朗和金城连作园的土壤真菌群落结构均有明显差异,3个连作园的土壤真菌各自构成一个独立的群落结构,这些群落能够适应各自的土壤环境并成为环境的优势群落。     

Apple Replant Disorder of Pingyitiancha Rootstock is Closely Associated with Rhizosphere Fungal Community Development

The aim of this study was to examine rhizosphere fungal community development in apple replant soil and a new planting soil in Beijing through a potted experiment with the apomictic triploid Pingyitiancha rootstock using the Illumina Miseq method. Steam sterilization significantly enhanced seedling growth in replant soil, while root segment addition did not enhance replant disease symptoms. The bulk soil of the replant site (ReSoil) and the new soil of the planting site (NewSoil) were sampled prior to nursery planting, and rhizosphere soils were r Qinping esampled after one growing season as RePlant and NewPlant, respectively. Approximately 48.3, 47.9, 59.8 and 68.8% of the operational taxonomic units (OTUs) were classified as Ascomycota and Basidiomycota in the community of ReSoil, New Soil, RePlant and NewPlant, respectively. Shannon indices suggested that planting apple nurseries increased the fungal diversity of rhizosphere soil in both soils. Apple replant disorder (ARD) was highly associated with an unbalanced microbial ecosystem with more saprophytic and pathogenic fungi, but less species known to produce antimicrobial metabolites, while Rhizopus, Pyrenochaeta and Eurotium were identified as the key factors in ARD.     

Effects of soil fumigation on the growth of apple and cherry rootstocks on land previously cropped with apples

Apple and cherry rootstocks were planted into land on which apple trees had previously been grown and which pot tests had shown to be subject to apple replant disease. Prior to planting the land was either left unfumigated or zones of progressively increasing area (0.052 m², 0.839 m³, 3.356 m²) centred on the proposed planting holes were fumigated with chloropicrin. Fumigation greatly increased the growth of the apple rootstocks but not of the cherry rootstocks. In the first year after planting the effects of the two larger fumigated areas were identical but in later years apple rootstock growth was directly related to the size of the fumigated area. This indicated that the replant disease does not simply affect tree establishment as was previously thought to be the case. The implications of this are discussed.     

Streptomycin Application Has No Detectable Effect on Bacterial Community Structure in Apple Orchard Soil

Streptomycin is commonly used to control fire blight disease on apple trees. Although the practice has incited controversy, little is known about its nontarget effects in the environment. We investigated the impact of aerial application of streptomycin on nontarget bacterial communities in soil beneath streptomycin-treated and untreated trees in a commercial apple orchard. Soil samples were collected in two consecutive years at 4 or 10 days before spraying streptomycin and 8 or 9 days after the final spray. Three sources of microbial DNA were profiled using tag-pyrosequencing of 16S rRNA genes: uncultured bacteria from the soil (culture independent) and bacteria cultured on unamended or streptomycin-amended (15 μg/ml) media. Multivariate tests for differences in community structure, Shannon diversity, and Pielou''s evenness test results showed no evidence of community response to streptomycin. The results indicate that use of streptomycin for disease management has minimal, if any, immediate effect on apple orchard soil bacterial communities. This study contributes to the profile of an agroecosystem in which antibiotic use for disease prevention appears to have minimal consequences for nontarget bacteria.     

Effects of transgenic rootstocks on growth and development of non-transgenic scion cultivars in apple

Although cultivation of genetic modified (GM) annual crops has been steadily increasing in the recent 10 years, the commercial cultivation of GM fruit tree is still very limited and reports of field trials on GM fruit trees are rare. This is probably because development and evaluation of GM fruit trees require a long period of time due to long life cycles of trees. In this study, we report results from a field trial on three rolB transgenic dwarfing apple rootstocks of M26 and M9 together with non-transgenic controls grafted with five non-transgenic scion cultivars. We intended to investigate the effects of transgenic rootstock on non-transgenic scion cultivars under natural conditions as well as to evaluate the potential value of using the rolB gene to modify difficult-to-root rootstocks of fruit trees. The results showed that all rolB transgenic rootstocks significantly reduced vegetative growth including tree height regardless of scion cultivar, compared with the non-transgenic rootstocks. Flowering and fruiting were also decreased for cultivars grown on the transgenic rootstocks in most cases, but the fruit quality was not clearly affected by the transgenic rootstocks. Cutting experiment and RT-PCR analysis showed that the rolB gene was stably expressed under field conditions. PCR and RT-PCR analyses displayed that the rolB gene or its mRNA were not detectable in the scion cultivars, indicating no translocation of the transgene or its mRNA from rootstock to scion. Our results suggest that rolB modified rootstocks should be used in combination with vigorous scion cultivars in order to obtain sufficient vegetative growth and good yield. Alternatively, the rolB gene could be used to dwarf vigorous rootstocks of fruit trees or produce bonzai plants as it can significantly reduce the vegetative growth of plants.