Aggressiveness,cultural characteristics and genetic variation of Ceratocystis fimbriata on Eucalyptus spp.

Ceratocystis wilt caused by Ceratocystis fimbriata is currently one of the most important diseases affecting Eucalyptus in Brazil. This disease is controlled by planting resistant clones; however, possible variability in the pathogen population may compromise the selection of resistant genotypes. Therefore, this study aimed to evaluate the aggressiveness of C. fimbriata isolates obtained from Eucalyptus spp, as well as their cultural characteristics and genetic variation of their ITS rDNA gene region. We found a significant isolate × clone interaction, with the isolate RM35 being the most aggressive and presenting a broader spectrum of aggressiveness, causing greater xylem discoloration on a larger number of clones. This isolate is the most suitable for artificial inoculations focusing on the selection of resistant materials. Clones CLR‐236 and CLR‐212 were identified as the most resistant and clones CLR‐223 and CLR‐240 as the most susceptible and those that are recommended as reliable comparators in artificial inoculations. All isolates were morphologically similar and differed from C. fimbriata from sweet potato by the formation of a wide mouth endoconidiophore that produces doliform endoconidia. According to the culture media and temperature applied, the most favourable conditions for mycelial growth were observed using malt extract agar (MEA) and temperatures ranging from 24 to 26°C. There was no correlation between sporulation and aggressiveness. Great variation in ITS sequences was observed, and a total of five ITS genotypes were identified among the ten isolates tested.     

Spatial–temporal patterns of Ceratocystis wilt in Eucalyptus plantations in Brazil

Ceratocystis wilt, caused by Ceratocystis fimbriata, has become the most important disease in eucalyptus (Eucalyptus spp. and hybrids) plantations in Brazil. To further our understanding of the epidemiology of this disease, we surveyed eucalyptus plantations in the states of Minas Gerais and Bahia that were known to have Ceratocystis wilt or were thought to have been planted with infected rooted cuttings. There was generally higher disease incidence in the Minas Gerais plantations, which were on former Cerrado forest sites and likely had soilborne inoculum prior to planting eucalyptus. In such plantations, disease incidence was not evident before 20 months after planting but slowly increased up to 50% at 74 months. The symptomatic and killed trees were aggregated, perhaps from uneven distribution of inoculum in the soil. Also, the progression of cumulative disease incidence best fit a monomolecular model, which is typical of soilborne diseases (fixed level of initial inoculum with little or no secondary inoculum during the crop rotation). However, plots where some trees had been harvested during the rotation showed very high levels of disease incidence in the sprouts that arose from stumps, suggesting secondary spread of the pathogen on harvesting tools or machinery. Most of the Bahia plantations were on pastureland prior to eucalyptus cultivation, and the pathogen was likely introduced with infected nursery stock. In such plots, symptoms were evident as soon as 7 months after planting, and most of the mortality occurred within 12 months. The diseased trees on former pastureland sites were sometimes aggregated within planting rows, suggesting that bunches of infected nursery stock were planted together within the rows. Care should be taken in planting disease‐free planting material and spreading the pathogen on tools, but on sites with soilborne inoculum, use of resistant clones may be the only management option.     

A serious new wilt disease of Eucalyptus caused by Ceratocystis fimbriata in Central Africa

In a recent survey of Eucalyptus clones in the Republic of Congo, Central Africa, a serious wilt and die-back disease of two different hybrid clones was observed. Affected trees ranged in age from approximately 6 months to 4 years. Isolations from symptomatic plant material consistently yielded a Ceratocystis species. On the basis of morphology and sequence data this fungus was identified as Ceratocystis fimbriata, a well-known wilt and canker pathogen of many economically important plants. The Eucalyptus isolates were compared with other Ceratocystis spp. based on sequence data generated from the ITS and 5.8S region of the rRNA operon. The results confirmed the identity of the Ceratocystis isolates from Eucalyptus as C. fimbriata and showed that they group with other C. fimbriata isolates from Brazil, South Africa and Europe. Inoculations on young Eucalyptus plants were conducted in the greenhouse and all three of the Congolese isolates tested, produced typical lesions in the bark and xylem. This study represents the first report of C. fimbriata as a pathogen of Eucalyptus in Africa. This is a serious new disease that will require considerable study in order to ensure that losses, caused by C. fimbriata, do not continue.     

Development of the fungus Ceratocystis fimbriata in shoots of poplar clones with differing resistance1

Differences in the development and rate of spread of C. fimbriata hyphae between susceptible, moderately resistant and resistant clones were observed. Conidia of various shapes and ascospores appeared in necrotic parts of shoots of susceptible and moderately resistant clones while in the resistant one only cylindrical conidia were present.     

Ceratocystis smalleyi colonization of bitternut hickory and host responses in the xylem

Colonization of Carya cordiformis sapwood by Ceratocystis smalleyi and subsequent host defence responses following artificial inoculation were investigated using anatomical and histological techniques. Hyphae of C. smalleyi were observed in all sapwood xylem features confirming the ability of the pathogen to invade and colonize the xylem tissues of the host species. The fungus was isolated from within and at the margins of discoloured sapwood areas at 2 and 12 months after inoculation. General host defence responses that included vessel occlusion with gels or tyloses, lipid accumulation, and production of phenolic compounds were observed in xylem tissues of inoculated C. cordiformis stems. Pectic substances, lipids, and to a rare extent, phenolic compounds were detected in vascular gels. The lipid‐rich barriers observed likely prevent lateral expansion of the fungus in the sapwood. Furthermore, lack of fungus sporulation within vessels may restrict axial spread of the fungus. C. smalleyi appears to be a limited vascular wilt pathogen of bitternut hickory based on these observations and previously reported sap flow reduction correlated with multiple infections in artificially inoculated trees.     

Impact of Ceratocystis wilt on eucalyptus tree growth and cellulose pulp yield

In this study, we determined the effects of Ceratocystis fimbriata wilt on volumetric growth and cellulose pulp yield of eucalyptus. Seven‐year‐old healthy and infected trees were separated into different classes of severity, and the individual volume of each whole tree with bark was determined. Volumetric losses were found regardless of the class of disease severity, resulting in reduction in volumetric growth of the trees from 65% to 87%, depending on the infection level. Two approaches were used for evaluating the effects of the disease on cellulose pulp yield: (i) evaluation of wood basic density and chemical composition, contents of ethanol/toluene extractives and Klason lignin, and the alkaline load and screen yield across Kraft pulping to kappa number 18 ± 0.5 for both healthy and discoloured materials; (ii) determination of the alkaline load and yield across Kraft pulping to kappa number 18 ± 0.5 of woods with different levels of discoloration (0, 25, 50, 75 and 100% of discoloured wood). Discoloured wood presented lower basic density, higher ethanol/toluene extractives and Klason lignin, higher alkaline load and lower pulp screened yield at kappa number 18 ± 0.5 compared with the healthy ones. Increasing the fraction of infected wood in chip furnishes resulted in proportionally increased alkaline load demand and decreased screen yields. Adding 50% of infected wood in the chip furnish resulted in 23.8% increase in the alkali load and 13.7% reduction in the pulp screen yield at kappa number 18 ± 0.5. The results obtained have proved that the disease reduces volumetric growth, increases wood consumption and reduces yield in eucalyptus pulp production.     

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.     

Possibilités d'infection du platane par Ceratocystis fimbriata f. platani après contamination de l'eau où se développent des racines blessées

Infection of plane tree by Ceratocystis fimbriata f. platani after contamination of water as growing substrate of wounded roots. Plane tree seedlings (Platanus acerifolia) were cultivated during 1. year so as some roots emerged from the culture pot And grow in the water of a container placed below. In April, these roots were wounded and nearby, conidia suspension of C. fimbriata?. platam was injected into the water. 1 month later, foliage of some seedlings began to wilt and after 2 months, 7 out of 12 plane trees were drying. The pathogen was recovered from all parts of the roots above the water level inside the pots and as high as the collar for 4 plants.     

青枯菌对桉树及非寄主树木根部吸附和侵入的比较

通过接种青枯菌与非病原的大肠杆菌,对2个桉树无性系和1种非寄主树木的根表吸附菌量及根内含菌量进行测定.结果表明:青枯菌对感病寄主根表的吸附量多于对抗病寄主和非寄主根表的吸附量,对感病寄主根部的侵入量多于对抗病寄主但少于对非寄主根部的侵入量;病原菌较非病原菌对感病寄主根表的吸附量大,但二者对感病寄主根部的侵入量则依据根部伤口的有无而互有高低.青枯菌接种24 h内,感病寄主根表吸附菌量和根内含菌量呈上升趋势,在抗病寄主及非寄主根表的吸附菌量和根内含菌量却呈下降趋势.从统计学上看,青枯菌对寄主根部的吸附和侵入都显示出了一定的选择性,但由于病原细菌能够大量地吸附和侵入抗病寄主和非寄主根部,非病原细菌也能够大量地吸附和侵入桉树根部,因此这种选择性的生物学意义不大,青枯菌对桉树根表没有表现出明显的识别行为.     

First hybrid plane trees to show resistance against canker stain (Ceratocystis fimbriata f. sp. platani)

Canker stain, caused by the ascomycete Ceratocystis fimbriata f. sp. platani, is a major threat to plane trees in Europe. The American plane tree (Platanus occidentalis) carries some genetic resistance, but this species is not adapted to the climatic conditions in Europe. Therefore, hybrids between susceptible oriental plane trees (Platanus orientalis) and resistant P. occidendalis were screened for disease resistance in France. Among 960 hybrids, we found 18 individuals which survived two successive inoculations. These putatively resistant trees were cut back and, after 18 months, on each tree 12 of the newly produced shoots as well as two roots, were inoculated again. From these trees, only one showed complete and another one partial resistance. This selection process resulting in the resistant clone ‘Vallis clausa’ is described in detail.     

Susceptibility of eliteAcacia mearnsii families to Ceratocystis wilt in south Africa

Ceratocystis albofundus is a recently described pathogen infectingAcacia mearnsii in South Africa, and it causes a disease known as Ceratocystis wilt. Symptoms of the disease include die-back, gummosis and wilting of infected trees. In order to select trees tolerant to this fungus, susceptibility tests were conducted on trees representing fourteen families ofA. mearnsii. A virulent isolate ofC. albofundus was selected and inoculated into the stems of twelve-month-old plants in a plantation. Lesion lengths, in the bark, and disease development were assessed after 6 weeks. All fourteen families ofA. mearnsii were susceptible to infection byC. albofundus. Considerable variation was, however, noticed between individual trees within the same family and the incorporation of disease tolerant trees into breeding programmes is proposed.     

Response of in vitro obtained oil palm and interspecific OxG hybrids to inoculation with Phytophthora palmivora

Bud rot (BR) disease caused by Phytophthora palmivora is the most devastating disease in oil palm cultivation in America. Oil palms that survived BR epidemics were found in areas highly devastated by the disease; these palms were introduced into Cenipalma's in vitro micropropagation (cloning programme). A severity scale was developed for in vitro palms from five ortets inoculated with two different P. palmivora isolates. Then, eight ortets of Elaeis guineensis and two ortets of the OxG interspecific hybrid were evaluated in two inoculation trials under chamber growth conditions. The clone performance response was consistent with that reported in the field for the corresponding ortets, and two contrasting ortets, one susceptible (ortet 57) and one resistant (ortet 34) were identified. We monitored and compared defence responses to P. palmivora in the contrasting ortets. An increase in reactive oxygen species (ROS) production after inoculation with the pathogen was observed, with higher accumulation of H₂O₂ in the resistant plants. Catalase (CAT) activity in resistant plants increased after inoculation with the pathogen from 24 hr post‐infection (hpi) and remained high during the observation time. In the susceptible ortets, there was a significant increase on catalase activity only at 48 hpi. Peroxidase (POS) activity increased in clones from both susceptible and resistant ortets, but the increase was much greater in the susceptible ones. Phenylalanine ammonia lyase activity increased in response to inoculation, and these increases were greater in clones of the susceptible ortet than in clones of the resistant ortet.     

Une méthode simple de recherche de Ceratocystis fimbriata f. platani sur arbre en place

A simple method of diagnosing Ceratocystis fimbriata f. platani in standing trees. Bluish or purple streaks are found at the margins of lesions caused by Ceratocystis fimbriata f. platani. These contain the fungus systematically. Cores may be taken with an augur from any such streaks, the core then being cut into discs which are kept in a humid chamber at 20–25°C. C. fimbriata, if present, produces diagnostic endo-conidia after 2–4 days of incubation in 50–90 % of the samples.     

Etude experimentale du transport de Pinoculum de Ceratocystis fimbriata f. platani par ľeau ?une riviere

Dissemination of Ceratocystis fimbriata f. platani inoculum by river water . C. fimbriata was detected up to 108 m downstream from pieces of diseased plane tree placed in a fixed site in a river in Southern France. Assessment of C. fimbriata was realized by a trapping method with healthy wood.     

Intra- and inter-provenance variability in phloem phenols of Picea abies and relationship to a bark beetle-associated fungus

One hundred Norway spruce (Picea abies (L.) Karst.) clones (three ramets per clone) were analyzed for phloem phenol composition and concentration before and 10 days after wound inoculation with sterile malt agar. Fifty clones (Experiment 1) belonged to the same provenance, whereas the remaining clones (Experiment 2) belonged to five provenances from three geographic areas. In Experiment 2, two additional ramets from the same clones were mass inoculated with Ceratocystis polonica (Siem.) C. Moreau to quantify the resistance of each clone. Tree response to wound inoculations was characterized by increased catechin concentration in both experiments, accompanied by increases in astringin and decreases in piceid in Experiment 1. In both experiments, we observed a diverse group of phenolic compounds whose concentrations increased (catechin, astringin) or did not vary (taxifolin glucoside) in response to wound inoculations, whereas concentrations of a homogeneous group of stilbene compounds decreased (piceid) or did not vary (isorhapontin, unidentified stilbene). In Experiment 2, provenances from the alpine and Hercynian-Carpatic areas differed from provenances from the Baltic area with respect to the relative importance of these two groups of compounds, further indicating that the two groupings of phenolic compounds structure the Norway spruce populations. Eighty days after mass inoculation, the percentage of healthy sapwood, which was taken as a measure of tree resistance, indicated that clones from the Baltic area were less resistant to mass inoculations than clones from the alpine and Hercynian-Carpatic areas. We conclude that the degree of resistance of Norway spruce trees to mass inoculations with a bark beetle-associated fungus can be predicted based on the diversity of constitutive phloem phenols and the ability to induce phenol synthesis in response to wounding.     

Détection par piégeage du Ceratocystis fimbriata f. platani

Application of the technique of trapping Ceratocystis fimbriata f. platani for the study of the persistance in affected trees . The study of the persistance of Ceratocystis fimbriata f. platani in affected trees is achieved by trapping of the fungus which is carried out on small branches of London plane, stripped of their bark. If C. fimbriata is present in samples of wood to be tested, it becomes visible after a few days when numerous perithecia form on the trap. Trapping the fungus by this method has been systematically achieved with young, artificially inoculated trees: in many cases, C. fimbriata persists in the wood after a period of about 19 to 22 months.     

Sphaeropsis canker,a new disease of cypress in Greece

The fungus Sphaeropsis sp. is reported for first time in Greece to cause cankers on Cupressus sempervirens. The cultural characteristics on PDA, spore shape and size and canker morphology were identical to those of the fungus described as Diplodia pinea f. sp. cupressi in Israel. The cankers of Sphaeropsis sp. are characterized by resin exudation, with fissuring of the bark over a dry sector of the wood. The pathogenicity of the Sphaeropsis sp. was proved by artificial inoculations on selected cypress clones resistant and susceptible to Seiridium cardinale. The mean canker length of the susceptible clone was significantly more than that of the resistant clones. It appears that the defence reaction in cypress against both fungi is similar. In artificial inoculations on Pinus halepensis and Pinus pinea, Sphaeropsis sp. was nonpathogenic. During a 3‐year period, in an experimental plot in western Peloponnese, the fungus spread from one, initially, to 13 cypress clones. Drought stress during the summer appears to be the main factor predisposing the cypress plants to become susceptible to the fungus.     

Electrophoretic protein profiles of strains of Ceratocystis fimbriata f. sp. platani

Soluble proteins extracted from the mycelium of strains of Ceratocystis fimbriata (Ell. et Halst) Davidson f. sp. platani Walter were separated on native and SDS-polyacrylamide gradient gel electrophoresis. The protein patterns were compared and the electrophoretic profiles of all the strains examined showed striking intraspecific similarities. The use or protein banding patterns as a diagnostic criterion to distinguish strains of C. fimbriata is discussed.     

Physiological and biochemical differences among Ulmus minor genotypes showing a gradient of resistance to Dutch elm disease

Dutch elm disease (DED) spread across Europe and North America in the 20th century killing most natural elm populations. Today, breeding programmes aim at identifying, propagating and studying elm clones resistant to DED. Here, we have compared the physiology and biochemistry of six genotypes of Ulmus minor of variable DED resistance. Leaf gas exchange, water potential, stem hydraulic conductivity and biochemical status were studied in 5‐year‐old trees of AB‐AM2.4, M‐DV2.3, M‐DV2 × M‐CC1.5 and M‐DV1 and 6‐year‐old trees of VA‐AP38 and BU‐FL7 before and after inoculation with Ophiostoma novo‐ulmi. Leaf water potential and net photosynthesis rates declined, while the percentage loss of hydraulic conductivity (PLC) increased after the inoculation in susceptible trees. By the 21st day, leaf predawn and midday water potential, stomatal conductance to water vapour and net photosynthesis rates were lower, and PLC was higher in trees of susceptible (S) genotypes inoculated with the pathogen than in control trees inoculated with water, whereas no significant treatment effect was observed on these variables in the resistant (R) genotypes. Fourier transform infrared spectroscopy analyses revealed a different biochemical profile for branches of R and S clones. R clones showed higher absorption peaks that could be assigned to phenolic compounds, saturated hydrocarbons, cellulose and hemicellulose than S clones. The differences were more marked at the end of the experiment than at the beginning, suggesting that R and S clones responded differently to the inevitable wounding from inoculation and repeated sampling over the experimental course. We hypothesize that a weak activation of the defence system in response to experimental wounding can contribute to the susceptibility of some genotypes to O. novo‐ulmi. In turn, the decline in shoot hydraulic conductivity and leaf carbon uptake caused by the infection further exacerbates tree susceptibility to the fungus.     

Vascular modifications in Platanus acerifolia seedlings inoculated with Ceratocystis fimbriata f. sp. platani

Vascular modification of plane tree seedlings inoculated with Ceratocystis fimbriata f. sp. platani, the canker stain disease agent, were characterized using histochemical and scanning electron microscopy methods.