Detection of phytoplasma infections in declining Populus nigra ‘Italica’ trees and molecular differentiation of the aster yellows phytoplasmas identified in various Populus species

A disease of Populus nigra‘Italica’ associated with foliar yellowing, sparse foliage, stunting, dieback, and decline was observed in south-western Germany; a witches’ broom disease of Populus alba that is known in other countries was also detected in Hungary and Germany. The aetiology of the diseases was studied by fluorescence microscopy and polymerase chain reaction (PCR) amplification. Using fluorescence microscopy, phytoplasmas could be detected only in P. alba. However, most diseased trees of P. nigra‘Italica’ tested phytoplasma-positive by PCR. In some of the trees the phytoplasma numbers were so low that nested PCR was required to detect the infection. Very low phytoplasma numbers were also observed in diseased Populus tremula. The identity of phytoplasmas from P. nigra‘Italica’ sampled in Germany and France, P. alba and also P. tremula was examined by restriction fragment length polymorphism (RFLP) analysis of PCR-amplified ribosomal DNA. In all poplars, phytoplasmas of the aster yellows group were detected. However, three different RFLP groups were identified that consisted of (1) French strains from P. nigra‘Italica’, (2) German strains from P. nigra‘Italica’ and (3) strains from P. alba and P. tremula. The profile observed in the last group was probably the result of sequence heterogeneity in the two 16S RNA genes.     

Fungi that destroyed a Populus tremula stand in Lagow Lubuski

The decay of a 20-year-old stand of Populus tremula growing on old agricultural waste land was observed from 1975 to 1981. There were 16 species of pathogenic and saprophytic fungi found on dying and dead trees, of which Heterobasidion annosum (Fr.) Bref. and Cytospora chrysosperma Fr. were the most frequent. The circumstances of combined action of both fungi in the disease process are discussed.     

Genotype differencies in direct plant regeneration from stem explants ofPopulus tremula in Turkey

Plant regeneration from 9 clones ofPopulus tremula representing diverse genotypes originated from Turkey, has been studied in different media, such as Murashige and Skoog’s Medium (MS), Aspen Culture Medium (ACM) and Woody Plant Medium (WPM) with appropriate supplements of growth regulators. Regeneration efficiency on WPM containing 1 mg/l zeatin using stem explantsin vitro, was higher than on MS and ACM. In this system, the time required for shoot regeneration was 2 weeks, which was shorter by two weeks than that of previous reports. A strong genotype dependence was observed among the tested nine clones. The regeneration system, we describe here, did not involve the callus phase and thus was less prone to somaclonal variation. When adventitious microshoots were cultured on WPM supplemented with 0.5 mg/l or 2 mg/l indole-3-butyric acid (IBA), root formation was obtained after 1 week of culture. Adaptation of regenerated plantletsin vivo was 100%. This system for micropropagation ofPopulus tremula is expected to be suitable for studies on transformation of genes involved in lignin biosynthesis via modification of lignin content. This study was a part of the Project No. 62-02-001 TUBITAK-MAN-GMBAE named “Investigations on Biogenetically Develop Poplar Clones as a Raw Material for Pulp and Paper Industry” and financially supported by General Management of Turkish Pulp and Paper Mills.     

Observations on some Italian provenances of Melampsora populnea

In some Italian provenances of Melampsora populnea on Populus tremula from mountain zones, the presence of Melampsora pinitorqua was tested through teliospore germinability tests and sub-sequent basidiospore inoculations. In this study the risk of twist rust epidemics in new planta-tions is discussed.     

木麻黄青枯病菌的分离及强致病菌株的筛选

[目的]筛选出强致病菌株用于木麻黄抗病育种研究工作。[方法]在广东沿海木麻黄青枯病发病区采集病根,开展病原菌两种不同分离方法的比较研究,对分离出的31个病株进行16s rRNA测序鉴定及致病性测定。[结果]采用稀释分离法及根系溢出法在TTC培养基上共分离出了31个病原菌株,根系溢出法操作简便,杂菌含量低,分离率在60%左右,可作为常规稀释分离法的补充。31个菌株进行分子鉴定,只有22个菌株扩增出了特异性条带,经测序比对确定这22个菌株为青枯菌。青枯菌株致病性测定结果显示菌株致病性在无性系间、菌株间及菌株与无性系间的交互作用均具有极显著差异(P0.01),不同接种方法间菌株致病性相关系数值较小,介于0.496 6~0.731 0之间,即室内水培接种与小苗盆栽接种不存在密切的直线相关关系。[结论]综合选择在不同无性系及不同接种方法中均具有较强致病性的GL-2、H、M、TC-1、F、Q菌株作为下一步木麻黄种质资源抗性鉴定及抗病育种研究试验菌株。     

新疆额尔齐斯河流域杨属植物种间关系的SSR分析

[目的]为探讨额河杨和银灰杨天然杂种的起源机制,[方法]应用18对SSR标记,从分子水平上对新疆额尔齐斯河流域杨属植物的种间关系进行分析研究。[结果]表明:(1)SSR系统发育树将整个流域天然杨属植物分为两大类群,即黑杨派和青杨派为一类,白杨派为一类;(2)白杨派派内系统聚类图显示,银白杨、欧洲山杨、银灰杨三个树种均有较大的遗传分化,特别是杂种银灰杨似乎更大;(3)黑杨派和青杨派的UPGMA分类图显示,青杨派和黑杨派分属于2个分支,其中,青杨派内部分化相对简单,分为2支,均为典型的苦杨;黑杨派内部的分化较为复杂,可分为4类,包括典型的欧洲黑杨、额河杨和回交子代。[结论]杂种额河杨具有更多的欧洲黑杨的遗传成分,因此,将额河杨放到黑杨派是正确的。     

An inoculation experiment of Japanese <Emphasis Type="Italic">Bursaphelenchus</Emphasis> nematodes on Japanese black and red pine, <Emphasis Type="Italic">Pinus thunbergii</Emphasis> and <Emphasis Type="Italic">P. densiflora</Emphasis>

The pinewood nematode (PWN) Bursaphelenchus xylophilus is an invasive pathogen that was introduced from North America to Asian countries and Portugal and is devastating native pine forests. Some native European and Asian Bursaphelenchus nematodes also have weak to moderate pathogenicity to native pine species. To evaluate the potential risk of native Bursaphelenchus species, we inoculated ten Japanese Bursaphelenchus species into native pine species (the dominant forest species) in Japan, and evaluated their pathogenicity using mortality and tracheal tissue damage as indices. Inoculation was conducted on August 3, 2007, and the symptoms were observed every 2 weeks until February 1, 2008. None of the inoculated trees, excluding the pathogenic PWN inoculated control, showed external disease symptoms; however, four species [a less pathogenic PWN isolate, B. luxuriosae, Bursaphelenchus sp. NK215 (undescribed), and NK224 (undescribed)] caused tracheal tissue damage in inoculated seedlings and showed weak pathogenicity. Therefore, we conclude that there are some potentially pathogenic native species of nematodes distributed in Japan. Interestingly, two of these weakly pathogenic species, B. luxuriosae and NK215, are not associated with Pinaceae trees, suggesting that nematode pathogenicity may be a pre-adaptive character. More experimental studies under different conditions are necessary to accurately evaluate the potential risk of these pathogens.     

Pathogenicity to alder of Phytophthora species from riparian ecosystems in western Oregon

Described as one of the most destructive pathogens of agricultural crops and forest trees, Phytophthora is a genus of microorganisms containing over 100 known species. Phytophthora alni has caused collar and root disease in alders throughout Europe, and a subspecies has recently been isolated in North America. Reports of canopy dieback in red alder, Alnus rubra, prompted a survey of their overall health and to determine whether P. alni was present in western Oregon riparian ecosystems. Over 1100 Phytophthora isolates were recovered, representing 20 species and 2 taxa. Phytophthora‐type cankers were observed in many trees, and their incidence was positively correlated with canopy dieback. High levels of mortality for red alder were not observed, which suggests these Phytophthora species are not aggressive pathogens. To test this hypothesis, three stem wound inoculations and one root dip were conducted on red alder seedlings using 13 Phytophthora species recovered from the riparian survey. Ten of the 13 Phytophthora species produced significant lesions in at least one pathogenicity test. Phytophthora siskiyouensis produced the largest lesions on red alder from the two stem wound inoculation tests conducted under summer conditions, while P. taxon Pgchlamydo caused the largest lesions during the winter stem wound inoculation test. Phytophthora gonapodyides, P. taxon Pgchlamydo and P. siskiyouensis have previously been found associated with necrotic alder roots and bole cankers in the field, and with the pathogenicity results reported here, we have established these species as causes of Phytophthora root disease and Phytophthora bole canker of alder in Oregon. While none of the Phytophthora species were especially aggressive towards red alder in the pathogenicity tests, they did cause localized disease symptoms. By weakening the root systems or boles of alders, the Phytophthoras could be leaving alders more susceptible to other insects and pathogens.     

Comparison of procedures to evaluate the pathogenicity of Ceratocystisfimbriata sensu lato isolates from Eucalyptus in South Africa

Ceratocystis fimbriata sensu lato (s.l.) is an important pathogen of Eucalyptus. Pathogenicity of isolates has typically been evaluated by inoculating seedlings under greenhouse conditions. It is, however, not clear how accurately this reflects pathogenicity under field conditions. In this study, five techniques to potentially screen C. fimbriata isolates for their relative pathogenicity to Eucalyptus were compared. These included: in vitro growth comparisons on artificial media; inoculations on apples; inoculation on Eucalyptus seedlings in a greenhouse; inoculations on Eucalyptus bolts freshly cut from stems of young trees; and field inoculations on young trees. Eight isolates of C. fimbriata s.l. collected from various areas in South Africa were used. There was considerable variation in growth in culture and aggressiveness of the eight isolates. Field inoculations on young trees were best correlated with inoculations of bolts (r = 0.76). Lower correlation coefficients were obtained with seedlings (r = 0.59), apple inoculations (r = 0.56), and in vitro colony growth (r = 0.42). Inoculation of bolts provides a rapid and reliable method to screen isolates of C. fimbriata s.l. for pathogenicity to Eucalyptus.     

Two pathogenic races of Melampsora larici-populina in north-western Europe

Three collections of Melampsora larici-populina Kleb. originating from Belgium, France and the Netherlands were compared for their pathogenicity on eleven poplar clones. Qualitative type reactions suggest the existence of two pathogenic races designated E1 and E2. E2 is able to infect some clones previously considered to be resistant.     

Pathogenic variation of Seiridium spp. isolated from cankered Cuppressaceae hosts in New Zealand

Field and glasshouse inoculation tests with isolates of S. cardinale and S. unicorne (Cke & Ell.) Sutton on C. macrocarpa and C lusitanien showed symptoms caused by the two pathogens to be indistinguishable. Isolates of S. cardinale showed an overall higher pathogenicity and less variability than those of S. unicorne. Neither pathogen seemed capable of infecting hosts outside the Cupressaceae, An isolate of Boesewinkel originating from Crvptowcria and supposed to be a distinct species from the anamorph of Lepteutypa cupressi, failea to infect Cryptomeria; neither could L. cupressi and S. cardinale, though all of them readily infected C. macrocarpa and C. lusitanica. Isolates of S. unicoene showed vast differences in pathogenicity in any one test, but many of the Isolates also showed extremi pathogenicity variation between repeat tests, hence pathogenicity determination became uncertain or probalistic. There is no evidence that the decline of S cardinale in this country is due to reduced pathogenicity or to possible interaction with S. unicorne in host infection.     

Clonal variation in root suckering ability of hybrid aspen (Populus tremula L. × P. tremuloides Michx.)

Current information on the root sucker ability of hybrid aspen (Populus tremula L. × P. tremuloides Michx.) is in most cases based on clone mixtures. In this study, we could separate the performance among clones by using two experimental sites with monoclonal plots of the crossing. The natural regeneration was followed for 2 years after harvest of the formerly planted stands, clear-cut at 22 and 25 years of age, respectively. We found that clonal differences were present in the number of root suckers produced per area unit and their biomass production. However, all included clones produced sufficiently many root suckers for a successful regeneration. To receive a more complete evaluation of the root sucker dynamics for future selection on the clonal level, further studies are needed where vitality, self-thinning and effects of thinning measures are coupled to the goal of the aspen forestry.     

Clones No 1333 ofPopulus alba × P. davidiana and No 1132 ofP. davidiana × P. alba × P. daviana

The hybridization experiment was initiated in 1975, in which the parents ofP. davidiana were collected from Dailing, Heilongjiang Province,P. suaveolens were from Baicheng,P. simonii from Zhaodong of Heilongjiang Province, andP. tremula from Shanxi Province. Clones No 1333 ofP. Alba × P. davidiana and No 1132 ofP. davidiana × (P. alba × P. davidiana F1) had greater genetic variation and heritability in clones tested. (Responsible Editor: Dai Fangtian)     

Pathogenicity of seed‐associated fungi to Podocarpus falcatus in vitro

Twenty‐nine fungi that were isolated from seeds and female cones of Podocarpus falcatus from natural forests in Ethiopia, were assessed for their impact on seeds and seedlings of the same host. Based on the results from in vitro seed inoculation tests, we could group the fungi into five categories as: (i) isolates that were pathogenic only to seeds and had no obvious impacts on the germlings; (ii) isolates that were pathogenic only to the germlings; (iii) isolates that were pathogenic both to seeds and the emerging germlings; (iv) isolates that were more or less harmless; and (v) isolates that were germination promoters. Inoculation tests were also performed on 4–7‐day‐old aseptically grown seedlings. Fusarium oxysporum and Polyporus sp., were strongly pathogenic to both seeds and seedlings, while Nectria gliocladioides, Peniophora cinerea and Pestalotiopsis neglecta also demonstrated pathogenicity but to a lesser extent. Other isolates, e.g. Diaporthe spp. resulted in increased germination of P. falcatus seeds and no pathogenicity to seedlings. However, further investigations are required in order to find out how these fungi behave under nursery or field conditions.     

Discula quercina as a possible causal agent of dieback on oak trees in Iran

During spring and summer of 2016, dieback symptoms including blights of leaves, twigs, and acorns were observed on current year shoots of Quercus infectoria in the Zagros oak forests of Iran. The fungus isolated from damaged tissues was identified as Discula quercina based on both morphological characteristics and ITS sequencing. To fulfil the Koch's postulates, a representative isolate was inoculated onto shoots of Q. infectoria and Quercus libani in both laboratory and forest conditions. Responses of the two oak species to inoculation with D. quercina were examined under laboratory and forest conditions in a completely randomized experiment. Discula quercina was clearly involved in oak dieback, and Q. infectoria was more susceptible than Q. libani to damage by the pathogen. This is the first record of the occurrence and pathogenicity of the fungus Discula quercina on Quercus infectoria. The fungus is considered as an emerging pathogen on oak trees in Zagros forests in Iran. Furthermore, the pathogenicity of the Discula quercina on Q. libani under laboratory and forest conditions increases the potential importance of this pathogen in Zagros forests.     

Vegetative compatibility and pathogenicity of Diplodia mutila isolates on oak

Forty isolates of Diplodia mutila, a dieback agent on oaks were characterized by their capacity to produce heterokaryons, by their vegetative compatibility, and by their pathogenicity tests on Quercus cerris. The forty isolates were collected from Quercus cerris, Quercus frainetto, Quercus ilex, Quercus pubescens, Quercus robur and Quercus suber in different regions in Italy. Three nit mutants (nit_t, nit and nit_M) were identified with different nitrogen sources: sodium nitrite, sodium nitrate, uric acid, ammonium tartrate and hypoxanthine. On the two nutrient media employed, a potato dextrose agar (PDA) medium with chlorate (PDC), and a minimal agar medium with chlorate (MMC), isolates obtained from Q. cerris yielded the greatest number of mutants: 254 on PDC (out of a total of 583) and 190 on MMC (out of a total of 440). Nine vegetative compatibility groups (VCGs) were identified. There was a strong association between these VCGs and host species. All isolates tested except one from Q. cerris were pathogenic.     

新疆额尔齐斯河流域白杨派植物居群遗传多样性分析

应用SSR标记技术,对新疆齐斯河流域河谷分布的白杨派树种银白杨、银灰杨和欧洲山杨天然居群的克隆结构、克隆多样性和遗传多样性进行研究。结果表明:欧洲山杨、银白杨和银灰杨均有很强的克隆繁殖特性。欧洲山杨、银白杨居群的克隆多样性均比较高,Simpson指数分别为0.987和0.983。与欧洲山杨相比较,银白杨居群具有较低的遗传多样性,Shannon信息指数分别为1.068 9和0.324 9,Nei多样性指数平均为0.505 6和0.211 2。欧洲山杨和银白杨居群间的遗传一致度均较高,变幅分别为0.778 1 0.954 4和0.975 1 0.994 6,反映出其超长距离的基因流特性,超强的基因流阻止了银白杨和欧洲山杨居群的遗传分化。研究发现,银白杨和欧洲山杨分别有95%和89.98%的遗传变异存在于居群内。     

Mechanical characterization of juvenile European aspen (Populus tremula) and hybrid aspen (Populus tremula × Populus tremuloides) using full-field strain measurements

Functional analysis of genes and proteins involved in wood formation and fiber properties often involves phenotyping saplings of transgenic trees. The objective of the present study was to develop a tensile test method for small green samples from saplings, and to compare mechanical properties of juvenile European aspen (Populus tremula) and hybrid aspen (Populus tremula × tremuloides). Small microtomed sections were manufactured and successfully tested in tension parallel to fiber orientation. Strain was determined by digital speckle photography. Results showed significantly lower values for juvenile hybrid aspen in both Young’s modulus and tensile strength parallel to the grain. Average Young’s moduli spanned the ranges of 5.9–6.6 and 4.8–6.0 GPa for European aspen and hybrid aspen, respectively. Tensile strength was in the range of 45–49 MPa for European aspen and 32–45 MPa for hybrid aspen. The average density (oven-dry) was 284 kg/m³ for European aspen and 221 kg/m³ for hybrid aspen. Differences in mechanical properties correlated with differences in density. Part of this article was presented at the 3rd International Symposium on Wood Machining, May 21–23, 2007, Lausanne, Switzerland     

Pathogenicity of Phytophthora species to Pacific Northwest Conifers

Phytophthora root rot is described for the first time killing sugar pine (Pinus lambertiana) in a seed orchard and four species of true fir (Abies spp.) in a forest nursery. P. cactorum was recovered from true firs and P. megasperma was recovered from sugar pine. P. cryptogea was recovered from sugar pine and true fir but isolates from the two locations differed from each other in pathogenicity and colony appearance. Isolates recovered from these hosts and isolates of 6 Phytophthora species previously recovered from Douglas-fir (Pseudotsuga menziesii) were then tested for pathogenicity on seedlings of 9 Northwest conifers. P. megasperma Group 1, P. cryptogea, and P. cinnamomi were pathogenic to all tree species except western redcedar (Thujaplicata). Western hemlock (Tsuga heterophylla) and true firs were susceptible to most species tested; ponderosa (P. ponderosa) and sugar pines were damaged only by P. cryptogea and P. cinnamomi; western redcedar was resistant to all isolates.     

Fungi in leaves,twigs and stem bark of Populus tremula from northern Spain

Fungal species were recorded from Populus tremula leaves, twigs and bark (trunk) at eight sites in the northern region of Spain, with the aim of evaluating the effect of sampling site, sampling tissue and isolation method on the frequency and species distribution of the fungi recovered from necrotic and healthy tissues. Two different isolation methods were used: the first consisted of finding fruitbodies on plant tissues after incubating them in wet and warm conditions, and the second included growing mycelia from plant fragments plated on potato dextrose agar (PDA). A total of 48 fungal species were isolated from 960 plant fragments. Cytospora chrysosperma, previously reported to be a pathogen of poplar, was recovered very frequently and was recorded as much in healthy as in dead or dying tissues. Cladosporium maculicola, Elsinoe veneta, Alternaria alternata, Aureobasidium pullulans and Pollaccia radiosa were also frequently isolated. The isolation frequencies from necrotic and healthy plant tissues were significantly different between sites and isolation methods. The incubation in moist chambers revealed significantly more fungal isolates and a higher species richness (38 species) than isolations on PDA. However, 16% of the fungal species were exclusively recorded from cultures. Therefore, a combination of several isolation methods is recommended for surveys of fungal communities associated with trees.