‘Le Crete 1’ and ‘Le Crete 2’: two newly patented Seiridium cardinale canker‐resistant cultivars of Cupressus sempervirens

In the Mediterranean area, common cypress (Cupressus sempervirens) has traditionally been used as a multipurpose tree, for its symbolic and ornamental role, for its valuable timber, as well as for windbreaks and soil protection. The epidemic spread of the Seiridium cardinale canker has limited the use of this tree since the 1970s, inducing researchers to develop a breeding programme of cypress aimed at selecting canker‐resistant lines for different uses and to support a flourishing trade of cypress plants. ‘Le Crete 1’ and ‘Le Crete 2’ described here are two new canker‐resistant C. sempervirens varieties patented in 2010, selected through a 13‐year assessment of their response to artificial inoculations and growth traits. Both are characterized by a rapid growth and by a columnar and fastigiated habit that confers them a notable ornamental effect. Preliminary observations showed also that both tend to produce yearly few microsporophylls and little pollen. ‘Le Crete 2’ was also selected for the high growth rate it maintained on heavy, clayey soils.     

Defence responses in plantation‐grown Eucalyptus globulus and Eucalyptus nitens after artificial fungal inoculation

The formation of reaction and barrier zones was studied in the xylem of Eucalyptus globulus and Eucalyptus nitens tree stems after wounding and artificial inoculation with two white rot fungi. The study had two objectives: to describe host responses in Eucalyptus spp. by light microscopy and to determine whether they would differ in a fungal treatment (wounding and inoculation by one of two fungal isolates) when compared to a control treatment (wounding only). Eucalyptus globulus and E. nitens developed similar reaction and barrier zones. The E. globulus barrier zone was characterized by kino vein formation. In both hosts, the reaction zone was primarily influenced by content and distribution of living tracheids and parenchyma cells within the sapwood. By contrast, the anatomy of the barrier zone showed similarities to the basic xylem structure of each host, except for some cell types that were newly formed (sclereids, kino veins) or increased in number (parenchyma cells, tracheids). Other cell types were reduced in number or completely absent. Host response in terms of barrier zone width appeared to be greater in the fungal than control treatment. Both wood decay fungi appeared to induce a wider barrier zone in both species than that associated with non‐specific damage caused exclusively by wounding. However, the small number of replicates available for this study was possibly insufficient to provide statistical evidence for different barrier zone width between fungal and control treatments.     

Occurrence of ‘Candidatus Phytoplasma ziziphi’ in apple trees in China

Young apple trees exhibiting symptoms of little leaf, margin involute and yellows were observed in an open‐air nursery in Yangling, Shaanxi, China. Transmission electron microscopy showed typical phytoplasma bodies in the sieve tube elements of symptomatic leaf samples. In two nested polymerase chain reaction (PCR) assays, products of expected size of 1.1 and 1.2 kb were separately generated from the total DNAs of symptomatic samples. A restriction fragment length polymorphism (RFLP) analysis of the purified 1.2 kb PCR products indicated that the disease associated with apple trees was ‘Ca. Phytoplasma ziziphi’. To our knowledge, this is the first report of ‘Ca. Phytoplasma ziziphi’ infecting apple trees in China.     

“Candidatus Phytoplasma solani” associated with Eucalyptus witches’ broom in Iran

During summer of 2015, Eucalyptus camaldulensis plants showing witches’ broom, little leaf and general yellowing of the foliage were observed in west of Fars and Khozestan province of Iran. DNA from samples of 22 symptomatic and two asymptomatic trees was extracted and subjected to molecular analyses. Nested‐PCR test using R16F2n/R16R2 primers confirmed phytoplasma presence in 63% of symptomatic Eucalyptus plants. Sequence analysis along with virtual RFLP of the 16S ribosomal DNA allowed to classify three Eucalyptus witches’ broom strains into the “stolbur” (“Candidatus phytoplasma solani”) 16SrXII‐A subgroup. Comparison of the secA and secY gene sequences with sequences deposited in GenBank confirmed the phytoplasma identity. Real and virtual RFLPs of the amplified secY gene using HaeIII, MseI and RsaI restriction enzymes showed profiles indistinguishable from each other. This is the first study reporting E. camaldulensis as a new host species for “Ca. P. solani.”     

‘Morfeo’ Elm: a new variety resistant to Dutch elm disease

Dutch elm disease (DED) has spread through Europe and North America since the beginning of the twentieth century. In response, several independent genetic improvement programmes for breeding DED‐resistant elms have been established on both sides of the Atlantic. Assimilating a wide range of resistant clones of different parentage should help ensure survival of the elm in the event of mutation of the pathogen. The Italian elm breeding programme began in the late 1970s and has recently raised a new variety by crossing a specimen of U. chenmoui with the Dutch hybrid clone ‘405’. This new release, named ‘Morfeo’, is extremely resistant to DED and has an attractive form and foliage. It is also fast‐growing, tolerant of drought and soils waterlogged in winter, therefore proving well adapted to the climates of both north‐western Europe and the Mediterranean. Following trials in England, ‘Morfeo’ is considered of potential importance in the conservation of several invertebrates endangered by the consequences of DED.     

Retention-tree groups in clear-cuts: Do they constitute ‘life-boats’ for spiders and carabids?

Green-tree retention aims at maintaining biodiversity while harvesting timber, but knowledge of its efficiency is still limited. Green-tree retention may function as a ‘life-boat’ for organisms, maintain structural features, or create stepping-stones in the harvested landscape. The aim of this study is to determine whether larger retention-tree groups established in clear-cuts maintain forest species better than smaller ones. We posed hypotheses for different habitat affinity groups of spiders and carabids, and tested them in Eastern Finland with 11 retention-tree groups, 0.09–0.55 ha in size (47–385 trees per tree group). Although some of the analysed species responded as expected, we did not find unambiguous support for the hypotheses that (1) species requiring forest and/or (medium-) moist habitat should be more abundant in larger retention-tree groups, (2) species requiring (semi-) open and/or (medium-) dry habitat should be more common in smaller retention-tree groups, and (3) species indifferent with respect to light intensity and moisture should be caught in equal numbers in all retention-tree groups irrespective of their size. However, the mean numbers of individuals of many forest species and/or species requiring (medium-) moist habitat decreased over the 3 years after logging in all retention-tree groups, whereas the mean numbers of many species of (semi-) open and/or (medium-) dry habitat increased over the 3 years after logging. Furthermore, species composition changed from the pre-treatment year for spiders. We conclude that green-tree retention is justified because (1) retention-tree groups tend to maintain some forest species, thereby mitigating the effects of clear-cutting, at least in the short term, (2) they provide dead wood, large old trees, and other structural features, and (3) there are gaps in our ecological knowledge that warrant the adoption of the precautionary principle. However, as there are economic costs associated with the establishment of retention-tree groups, their ecological benefits as compared to other approaches aimed at maintaining biodiversity should be assessed carefully.     

‘Pink rot’: infection of Castanea sativa fruits by Colletotrichum acutatum

Pink‐coloured tissues were observed in rotting chestnut nuts collected from soil in different orchards in Italy (Emilia Romagna, Tuscany and Trentino). Morphological and molecular analysis confirmed the presence of Colletotrichum acutatum (J.H. Simmons) in affected fruits, and inoculation tests corroborated the ability of the pathogen to colonize nuts.     

A new record of Rhizoctonia butinii associated with Picea glauca ‘Conica’ in Slovenia

In 2013, the death of needles of dwarf Alberta spruce (Picea glauca ‘Conica’) seedlings was observed in an arboretum in Slovenia. Needles on infected P. glauca ‘Conica’ twigs were covered with thick, yellowish brown hyphal cushions. The outgrowing mycelium was white and had completely overgrown some needles and connected them to each other. To definitively identify the fungus affecting P. glauca ‘Conica’, morphological examinations of isolated pure cultures were performed, and the internal transcribed spacer (ITS) region of the rDNA was sequenced. Based on morphological and molecular phylogenetic analyses together with the performed pathogenicity trial, we identified the causal agent of the observed symptoms as Rhizoctonia butinii – a recently described species. This report connects this fungus to a new locality and a new host plant.     

Salix tetradenia Hand.‐Mazz: a new natural plant host of ‘Candidatus phytoplasma’

This article reports Salix tetradenia Hand.‐Mazz as a new host of Candidatus phytoplasma and demonstrates its association with witches' broom disease on S. tetradenia plants. Plants exhibited typical visual symptoms of phytoplasma with virescence, abnormality of flowers and witches' broom, and phytoplasma bodies were observed by transmission electron microscopy. Products of 1.2 kb were amplified by nested PCR using phytoplasma universal primer pairs R16F2n/R16R2, but no amplification products were obtained from symptomless plants. The sequence analysis of three 16S rDNA isolates showed 99.84%, 99.68% and 99.76% identify, respectively, with the homologous gene (nc_005303) of member of ‘Candidatus phytoplasma asteris’ (16SrI) group. Phylogenetic and virtual computer‐simulated restriction fragment length polymorphism analysis of the 16S rRNA, tuf and rp gene sequences confirmed that this phytoplasma clustered in the 16SrI‐B subgroup. These results indicated that the diseased S. tetradenia plants were infected by a phytoplasma of the 16SrI group. This is the first report on the occurrence of phytoplasma disease on S. tetradenia worldwide.     

Ethnobotanical, pharmacological and chemical studies of plants used in the treatment of ‘Mwandza’ dermatites

An ethnobotanical investigation in the Bokouélé country, in the Cuvette region of Congo, was undertaken in order to record some of the plants used by local traditional therapists for the treatment of a disease locally known as ‘Mwandza’. Out of 60 plants recorded, a statistical study of those used in the presumably most efficient protocols, narrowed down the number to 20. Subsequent pharmacological tests have shown that some of these plants exhibit antibacterial and/or antitumoral activities.     

Genetic variability of the ‘bark canker resistance’ character in several natural provenances of Cupressus sempervirens

Several Aegean (Greece) and Anatolian (Turkey) cypress provenances were studied for resistance variability to bark canker, a disease caused by the fungal pathogen Seiridium cardinale. The investigation also examined whether the low disease rate within the natural area of cypress was due to genetic or geographic‐climatic reasons. Results demonstrated strong variability for the ‘bark canker resistance’ character, in particular for trees within families. As trees from the provenances studied were not found to have genetic superiority for bark canker resistance, the above‐mentioned low disease rate could be due to geographic‐climatic barriers that inhibit the development of the fungus or its ability to infect the host. Several half‐sib progenies exhibited high resistance, suggesting that this character is totally inherited through the maternal line. Should this finding be confirmed by further research, it would facilitate the task of genetic improvement for resistance, allowing progenies of resistant trees to be obtained.     

Identification of ‘Candidatus phytoplasma ziziphi’ associated with persimmon (Diospyros kaki Thunb.) fasciation in China

Persimmon (Diospyros kaki Thunb.) trees, with fasciation symptoms (PF), were observed in an orchard located in suburban Tai'an, Shandong Province, China. A specific fragment of the phytoplasma 16S rRNA gene, approximately 1.2 kb in length, was amplified from two symptomatic plants via nested polymerase chain reaction, while no fragment was obtained from healthy controls. The two samples (PF1 and PF2) resulted with 99.5% nucleotide sequence identity. Phylogenetic and restriction fragment length polymorphism (RFLP) analysis of 16S rRNA gene sequences revealed that PF1 was a member of ribosomal subgroup B of the elm yellows group (16SrV), and PF2 may represent a novel subgroup within the 16SrV group, designed as 16SrV‐I. This is the first report of phytoplasmas of the 16SrV group associated with persimmon fasciation disease.     

Rapid detection and identification of ‘Candidatus Phytoplasma pini’‐related strains based on genomic markers present in 16S rRNA and tuf genes

In order to devise a method for rapid detection of ‘Candidatus (Ca.) Phytoplasma pini’ and for distinguishing it rapidly from other phytoplasmas, we carried out preliminary sequencing of Lithuanian ‘Ca. Phytoplasma pini’ strain PineBL2 using Illumina (NGS) technology and targeted sequencing employing universal phytoplasma primers. We focused on two resulting chromosomal segments that contained a 16S rRNA gene and a translation elongation factor EF‐TU gene (tuf), respectively. Based on alignments of the ‘Ca. Phytoplasma pini’ gene sequences with the corresponding sequences of other phytoplasmas, we designed new primer pairs for PCR‐based detection of ‘Ca. Phytoplasma pini’. Because ‘Ca. Phytoplasma pini’ strains are expected to reside in the pine phloem in a very low titre, one might expect that they could be detected only by nested PCR. By contrast, the primers and PCR protocols designed in the current work enabled rapid direct PCR detection and identification of ‘Ca. Phytoplasma pini ’ by amplifying a 484 bp 16S rDNA segment and a 513 bp tuf gene fragment that contain regions unique to this phytoplasma .     

Behaviour of an Armillaria in some Eucalyptus obliqua – Eucalyptus regnans forests in Tasmania and its role in their decline

An undescribed but native species of Armillaria is a secondary pathogen in declining stands of Eucalyptus obliqua and Eucalyptus regnans in southern Tasmania. The fungus is almost ubiquitous in healthy and diseased forest and occurred epiphytically or parasitically in local lesions on the roots of 74 % of eucalypts partially excavated in 15 forest stands, 25–75 years of age. The fungus infected 85 % of logging stumps and 64 % of fire killed trees. Although the fungus only infected the sapwood of the eucalypts, it survived in infected stumps for at least 70 years. Rhizomorphs initiated most infections but did not grow freely through the soil. Pathogenicity tests confirmed that the Armillaria is a weak pathogen.     

Isolation of Xanthomonas campestris pv. populi from stem lesions on Salix matsudana X alba ‘Aokautere’ in New Zealand

A yellow-pigmented, Gram-negative, aerobic, salicin utilising bacterium-identified as Xanthomonas campestris pv. populi was isolated from one year old stems of S. matsudana X alba‘Aokautere’ willows exhibiting stem necrosis. Although the bacterium was isolated from necrotic stem tissue almost in pure culture it was not pathogenic to vigorously growing material of S. matsudana X alba‘Aokautere’ in field inoculations. Stress and predisposing climatic conditions during winter (frost, rain, wind, mild temperatures) are thought to have favoured the initial infections of this opportunistic pathogen. This disease is not considered serious.     

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.     

The response of Populus ‘NE 42’ (P. maximowiczii × P. trichocarpa) to infection by Ceratocystis fimbriata isolated from cacao-tree and plane

Shoots of a poplar clone susceptible to a Ceratocystis fimbriata strain from poplar were inoculated with strains of the fungus isolated from cacao-tree and plane. Disease symptoms and pathological changes in shoots as well as the development of the fungus in tissues are reported.     

The role of free proline in the sensitivity of poplar (Populus ‘Robusta’) plants to the action of SO11

The action of SO₂ caused an increase in the content of free proline in the leaves of Populus ‘Robusta’ plants. A protective role of proline pretreatment was demonstrated against injury, changes in water content and chlorophyll loss in the leaves of plants exposed to the gas.     

A TaqMan real‐time PCR assay for the detection of Phytophthora ‘taxon Agathis’ in soil,pathogen of Kauri in New Zealand

Kauri Agathis australis, an iconic tree of New Zealand, is under threat from an introduced disease‐causing pathogen provisionally named Phytophthora ‘taxon Agathis’ (referred to as PTA). This soilborne, Pythiaceous species belongs to the Chromista and causes a collar rot resulting in yellowing of the foliage and thinning of the canopy, which eventually causes death of the infected tree. The management and containment of this pathogen requires rapid and reliable detection in the soil. The current method for soil detection utilizes a soil bioassay involving lupin baits and soil flooding in a process that takes between ten and twenty days. We describe a real‐time PCR assay based on TaqMan chemistry for the specific detection of PTA, which targets the internal transcribed spacer (ITS) region of the nuclear ribosomal DNA. This TaqMan real‐time PCR assay could be used with DNA extracted directly from bulk soil samples to enable rapid quantification of PTA within soil. The detection limit was 2 fg of PTA DNA from pure culture, or 20 fg in the presence of DNA extracted from soil. The assay was validated using soil samples taken from a PTA‐infested site and soil spiked with a known concentration of oospores. We conclude that the TaqMan real‐time PCR assay offers a more time‐efficient method for detection of PTA in soil than existing methods.