Cytospora Canker and Dieback of Apricots

Cytosporal canker and dieback (Cytospora cincta Sacc.) is a widespread disease in apricot growing areas of Europe, excluding the Mediterranean. The pathogen attacks xylem and phloem. If the necrotic phloem completely girdles the branches or trunk, the section above the girdled region dies. When cambium necrosis does not girdle the branch or trunk, canker development ensues. Apricot trees are susceptible to Cytospora infection between July and the next bud opening.     

栗疫病菌侵染板栗枝条的显微观察

栗疫病是一种严重危害栗属植物的病害。为了明确栗疫病菌侵染板栗枝条的过程及侵染的关键时间点,本研究利用病理组织切片技术、显微镜和扫描电镜技术对栗疫病菌侵染板栗枝条的过程进行了观察。结果表明:接种栗疫病菌后0～5 h,菌丝先降解枝条表皮,进行横向营养生长的同时沿着伤口纵向侵染,为进入皮层做准备;接种后6 h病菌开始在表皮定殖,并侵入皮层;接种后9 h在皮层可观察到侵染性菌丝沿着细胞间隙向相邻细胞延伸;接种后12 h栗疫病菌侵入韧皮部,在皮层的侵染面积扩大。随着侵染程度加深,皮层、韧皮部等处细胞被菌丝降解,最终在形成层附近聚集。接菌后9 h为栗疫病菌侵染板栗枝条的关键时间点。     

Infection of horse chestnut (Aesculus hippocastanum) by Pseudomonas syringae pv. aesculi and its detection by quantitative real-time PCR

Pseudomonas syringae pv. aesculi is a pathogenic bacterium causing bleeding canker disease of horse chestnut ( Aesculus hippocastanum ). This is a serious disease which has been affecting horse chestnut in several European countries over the last five years; however, very little is known about the biology of the causal agent. One of the obstacles to studying this pathogen is the lengthy procedure associated with confirming its presence on the host. In this study, P. syringae pv. aesculi was isolated from lesions on different parts of horse chestnut and its pathogenicity confirmed on horse chestnut saplings using two inoculation techniques. Real-time PCR primers were developed based on gyrase B gene sequence data for the specific detection of P. syringae pv. aesculi . Primer specificity was tested on isolates of the target pathogen as well as on a broad range of related non-target bacteria and other bacterial spp. which inhabit horse chestnut. The real-time primers reliably amplified P. syringae pv. aesculi down to 1 pg of extracted DNA, with and without the presence of host DNA, and also amplified unextracted DNA in whole cells of the bacterium down to at least 160 colony forming units. Detection and quantification of the target pathogen in phloem and xylem of both naturally infected and inoculated horse chestnut tissues was also demonstrated. This quantitative real-time PCR assay provides the facility to study several important aspects of the biology of P. syringae pv. aesculi on horse chestnut including its potential for dissemination in different substrates.     

The effect of Cryphonectria parasitica on water relations of American chestnut

Chestnut blight has commonly been regarded as a phloem disease due to conspicuous stem cankers that result from infection by the fungal pathogen Cryphonectria (Endothia) parasitica. Stomatal conductance (g_s), transpiration rate (E) and leaf water potential were measured throughout the day on leaves distal to naturally-occurring virulent (sunken bark with abundant stromata) or hypovirulent (swollen bark lacking stromata) cankers and cankers induced by inoculation with virulent or hypovirulent strains of C. parasitica. Relative to control stems, hydraulic conductivity (K_h), g_s and E were reduced significantly (α = 0·05) for both natural virulent cankers, and cankers that were induced by a virulent strain. These effects were less pronounced for both natural and induced hypovirulent cankers. Isolation experiments revealed that the percentage of xylem tissue chips yielding C. parasitica was greater for virulent than for hypovirulent cankers. The data provide evidence that the localized presence of C. parasitica in cankers of American chestnut results in stomatal closure, possibly as a direct result of xylem dysfunction.     

Redefining the global populations of Pseudomonas syringae pv. actinidiae based on pathogenic,molecular and phenotypic characteristics

Knowing the population structure of a pathogen is fundamental for developing reliable phytosanitary legislation, detection techniques, and control strategies based on the actual aggressiveness and distribution of the pathogen. Currently, four populations of Pseudomonas syringae pv. actinidiae (Psa) have been described: Psa 1, Psa 2, Psa 3 and Psa 4. However, diagnostic assays specific for Psa populations do not detect Psa 4, the less virulent (LV) strains isolated in New Zealand. Similarly, multilocus sequence typing (MLST) of housekeeping genes, or broad Psa strain genome comparisons, revealed that Psa 4‐LV strains clustered separately from other Psa populations. In order to examine whether the placement of Psa 4 in the pathovar actinidiae was appropriate, various tests were carried out. It was shown that the Psa 4‐LV strains induced leaf and shoot wilting in Prunus cerasus, extensive necrotic lesions in Capsicum annuum fruits, and no significant symptoms in Actinidia deliciosa. Moreover, repetitive‐sequence PCR fingerprinting, type III secretion system effector protein genes detection and colony morphology clearly indicated the distinctiveness of Psa 4‐LV strains from the other three Psa populations. Rep‐PCR molecular typing revealed a high similarity of the Psa 4‐LV strains with members of Pseudomonas avellanae species. The Psa 4‐LV strains, most probably, belong to a new, still unnamed pathovar. It was concluded that the Psa 4‐LV strains isolated in New Zealand do not belong to the pathovar actinidiae, and, consequently, three Psa populations pathogenic to Actinidia spp. should currently include Psa 1, Psa 2 and Psa 3.     

苹果树腐烂病的发病过程和药剂防治研究

本文分两部分。第一部分概述了陕西风县秦岭高地苹果树腐烂病的发生发展过程,指出腐烂病菌主要是先在树皮的落皮层内潜伏侵染,进而侵害健组织。第二部分为防治试验。根据腐烂病的发生发展过程,于6月下旬和11月中旬对主干和大枝下部涂40%福美砷可湿性粉剂50倍悬浮液二次,获得了施药部位新病减少90%,“病疤重犯”减少80%的防治效果。与在辽宁兴城的试验,防效近似。     

Survival,cold tolerance and seasonality of infection of European horse chestnut (Aesculus hippocastanum) by Pseudomonas syringae pv. aesculi

Bleeding canker of European horse chestnut, caused by the pathogenic bacterium Pseudomonas syringae pv. aesculi (Pae), is now an established disease throughout several countries in northwest Europe after first emerging in 2001–2002. Pae infects the woody branches of horse chestnut directly via natural discontinuities in the bark, such as lenticels, leaf scars and leaf traces, and nodes. However, the timing of infection in relation to seasonality of host development, and the dispersal mechanisms of Pae, in particular its ability to survive and spread in soil and water, remains unknown. In this study, infection of freshly cut horse chestnut shoots by Pae was assessed at monthly intervals over a 12 month period. Pae infected the greatest numbers of lenticels and leaf scars of horse chestnut when inoculated onto actively elongating shoots in spring and early summer, whereas lesion extension from artificial wound sites was greatest after inoculation during early dormancy. Soil survival experiments showed that Pae was still detectable and viable after 50 weeks' incubation in sterile soil and 41 weeks' incubation in nonsterile soil in the absence of host debris. Pae also remained viable and pathogenic after 1 year's storage in King's B broth at ?20 and ?80°C, and was not killed by freeze/thaw treatments. Thus Pae is able to survive independently for extended periods in soil and water, and can tolerate lengthy periods of freezing at very low temperatures. Such information may facilitate a greater understanding of the epidemiology and spread of Pae in northern Europe.     

Association of ‘Candidatus Liberibacter asiaticus’ root infection,but not phloem plugging with root loss on huanglongbing‐affected trees prior to appearance of foliar symptoms

Huanglongbing (HLB) is a systemic disease of citrus caused by phloem‐limited bacteria ‘Candidatus Liberibacter’ spp. with ‘Ca. Liberibacter asiaticus’ (Las) the most widespread. Phloem‐limited bacteria such as liberibacters and phytoplasmas are emerging as major pathogens of woody and herbaceous plants. Little is known about their systemic movement within a plant and the disease process in these tissues. Las movement after initial infection was monitored in leaves and roots of greenhouse trees. Root density, storage starch content, and vascular system anatomy in relation to Las presence in field and greenhouse trees, both with and without symptoms, showed the importance of root infection in disease development. Las preferentially colonized roots before leaves, where it multiplied and quickly invaded leaves when new foliar flush became a sink tissue for phloem flow. This led to the discovery that roots were damaged by root infection prior to development of visible foliar symptoms and was not associated with carbohydrate starvation caused by phloem‐plugging as previously hypothesized. The role of root infection in systemic insect‐vectored bacterial pathogens has been underestimated. These findings demonstrate the significance of early root infection to tree health and suggest a model for phloem‐limited bacterial movement from the initial insect feeding site to the roots where it replicates, damages the host root system, and then spreads to the rest of the canopy during subsequent leaf flushes. This model provides a framework for testing movement of phloem‐limited bacteria to gain greater understanding of how these pathogens cause disease and spread.     

Bacterial Canker and Dieback Disease of Apricots (Pseudomonas syringae van Hall)

Bacterial canker and dieback (Pseudomonas syringae van Hall) of apricot is a widespread disease in Europe, except for the Mediterranean areas. Cankers usually develop at pruning wounds or other points of injury. Phloem and cambium become susceptible from just after leaf drop until budding. If the phloem necrosis does not girdle the branch or trunk, cankers develop by the middle or the end of summer. In early summer, the bacterium actually dies out in the infected tissue, and the tree remains resistant during the summer. During this period, the pathogen lives epiphytically on the surface of the leaves, without causing any symptoms. The extent of bacterial necrosis of the phloem depends on the severity of winter frost. Without winter frost, bacterial induced tissue necrosis does not occur. Necrosis is found only if P. syringae has enough time to proliferate before the onset of frost. The most effective method of control is to carry out spring rather than winter pruning.     

Monilia Disease

Monilia is a well-known parasite of stone fruit trees. It normally attacks flowers, buds and fruits, producing the diseases known as “brown rot” and “mummified fruit”. In certain circumstances, however, Monilia can also attack older branches, producing chronic or acute symptoms. In the first case, the affected branches wilt after a certain time, while in the second, this wilting is extremely fast; these are the classical symptoms of dieback. The parasite penetrates through wounds in the branches and first damages the phloem. If it reaches the cambium and girdles it, the branch dies from the area of attack to the tip. If there is no, or only moderate damage to the cambium, the tree can recover, or the disease may become chronic. The speed of disease development depends, primarily, on environmental conditions.     

Early investigations into the infection courts used by <Emphasis Type="Italic">Neonectria fuckeliana</Emphasis> to enter <Emphasis Type="Italic">Pinus radiata</Emphasis> stems

Nectria flute canker is a disease of Pinus radiata stems caused by the pathogen Neonectria fuckeliana occurring in the southern parts of New Zealand. In Northern Hemisphere countries where N. fuckeliana is endemic, it is commonly found in Picea and Abies spp. Open wounds, dead attached branches and branch stubs have been identified as the primary infection courts. Although in New Zealand the development of Nectria flute canker disease is associated with pruned branch stubs, recent studies suggest that this is not the only possible entry method as the fungus has been found in trees prior to pruning. Three field trials were established to examine the potential infection mechanisms for N. fuckeliana in P. radiata in New Zealand; including stem wounds and branch stubs. The difference between inoculations into the stem and into branch wood was clear. Inoculation of deep stem wounds resulted in the greatest fluting with 76% of trees inoculated developing cankers. Inoculation directly into stubs resulted in only small stem depressions that occurred in 17% of cases and the fungus was largely contained within the branch trace. Tree response to inoculation with either ascospores or conidia of the Acremonium anamorph gave similar results in terms of canker development and fungal spread within the stem. Tree response to inoculation was highly variable however: in one study 6% of trees did not respond to inoculation at all, while 26% produced severe cankers regardless of inoculation method. A more thorough understanding of the infection mechanisms of N. fuckeliana will contribute to the development of better disease management protocols to prevent infection and disease development in future plantation stock.     

Systemic applications of neem in the control ofCameraria ohridella, a pest of horse chestnut (Aesculus hippocastanum)

Systemic injection of neem (azadirachtin) was tested in the control of horse chestnut leafminer,Cameraria ohridella Deschka & Dimic (Lepidoptera: Gracillariidae), a pest of horse chestnut trees (Aesculus hippocastanum L.: Hippocastanaceae). Three concentrations, 0.08, 0.15 and 0.25 g of active ingredient (a.i.) per cm of diameter at breast height (dbh), provided long-lasting insecticidal activity. The 0.15 and 0.25 g a.i. concentrations resulted in 100% pupae reduction in all generations; the 0.08 g a.i. concentration was effective against the horse chestnut leafminer for at least 23 weeks and led to 70–80% pupae reduction. Treated trees did not lose their leaves before winter. Neem applicationvia injection is cheap, with no environmental hazards, and thus this treatment is recommended for the control ofC. ohridella in horse chestnut trees. http://www.phytoparasitica.org posting Dec. 19, 2004.     

Pewenomyces kutranfy gen. nov. et sp. nov. causal agent of an important canker disease on Araucaria araucana in Chile

Araucaria araucana, (commonly referred to as araucaria, pewen, or monkey puzzle tree) is an ancient conifer endemic to the Chilean and Argentinian mountain ranges where it has a sacred relevance to indigenous communities. During 2015, a serious disease was noticed on trees of all ages in most of the natural distribution of this iconic tree. Four areas were surveyed, and the most important symptoms of the disease were cankers on branches and stems resulting in copious resin exudation. Trees were monitored for a period of two years and isolations were made from the cankers. Field observations showed that the disease typically begins on the leaves or at the leaf bases and progresses downwards to initiate cankers that can girdle branches or stems within a two-year period. Black ascomata, resembling those of Caliciopsis species previously described from A. araucana, were consistently found developing in the cankers from which isolations were made. Phylogenetic analyses of the ITS, nucSSU, and nucLSU gene regions showed that the fungus resides in the Coryneliaceae but is distinct from other genera in that family. The morphological characteristics and phylogenetic position of the fungus show that it represents a new genus and species, described here as Pewenomyces kutranfy gen. nov. et sp. nov. Pathogenicity trials on trees under field conditions confirmed that this newly described fungus is able to cause cankers on A. araucana similar to those found under natural conditions.     

Alternative inoculum sources for fire blight: the potential role of fruit mummies and non‐host plants

Fire blight is the most damaging bacterial disease in apple production worldwide. Cankers and symptomless infected shoots are known as sites for the overwintering of Erwinia amylovora, subsequently providing primary inoculum for infection in the spring. In the present work, further potential sources of inoculum were investigated. Real‐time PCR assays covering a 3‐year‐period classified 19·9% of samples taken from fruit mummies as positive. Bacterial abundance in fruit mummies during autumn, winter and spring was up to 10⁹ cells per gram of tissue and correlated well with later infection rates of blossoms. Blossoms of non‐host plants growing close to infected trees were also shown to be colonized by E. amylovora and to enable epiphytic survival and propagation of bacteria. The results indicate a potential role of fruit mummies and buds in overwintering and as a source of primary inoculum for dissemination of the pathogen early in the growing season. Non‐host blossoms may also serve as an inoculum source in the build‐up of the pathogen population. Both aspects may contribute significantly to the epidemiology of E. amylovora. The significance of infected rootstocks as an inoculum source is also discussed. Fruit mummies might be used to determine pathogen pressure in an orchard before the beginning of the blooming period.     

Variation in stem infections caused by aggressive and non-aggressive isolates of Leptosphaeria maculans on Brassica napus var. oleifera

Development of stem cankers in seedlings of Brassica napus var. oleifera inoculated with Leptosphaeria maculans was studied in growth cabinets. As the pathogen progressed from the petiole into the stem, hyphae invaded parenchyma cells for the first time, initiating a necrotrophic phase which resulted in the formation of a lesion. Ultimate containment of the pathogen was associated with a sequence of host reactions including lignification, cambium formation and callose deposition at the perimeter of the lesion. Two isolates of L. maculans differed in the pattern of stem infection. One was confined to the cortex and the other passed through the leaf gap into the pith. Variation in response between cultivars Rapora, Primor and Jet Neuf modified these two colonization patterns but did not eliminate the main differences. There were qualitative and quantitative differences in host cell reactions. The significance of these findings is discussed in relation to differences in resistance of the cultivars in the field.     

Frost promotes the pathogenicity of Pseudomonas syringae pv. actinidiae in Actinidia chinensis and A. deliciosa plants

Frost occurs in all major areas of cultivation, presenting a threat for the production of kiwifruit crops worldwide. A series of experiments were performed on 1‐year‐old, potted plants or excised twigs of Actinidia chinensis and A. deliciosa to verify whether strict relationships exist between bacterial canker outbreaks from Pseudomonas syringae pv. actinidiae (Psa) attacks and the occurrence of autumn and winter frost events. The association between the occurrence of autumn frost and the sudden outbreak of bacterial canker in A. chinensis in central Italy has been confirmed. Both autumn and winter frosts promote Psa multiplication in the inoculated twigs of both species. The day after the frost, reddish exudates oozing from the inoculation sites were consistently observed in both species, and Psa was re‐isolated in some cases. During the thawing of both A. deliciosa and A. chinensis twigs, the 2‐cm upward and downward migration of Psa from the inoculation site was observed within 3 min, and the leaves were consistently colonized with the pathogen. A consistent brown discoloration, accompanied with a sour‐sap odour, was observed throughout the length of the excised twigs of both Actinidia species after Psa inoculation and winter frost. Psa inoculation induced a remarkably higher necrosis in excised twigs that were not frozen compared with P. s. pv. syringae inoculation. Antifreeze protection using irrigation sprinklers did not influence the short‐term period of Psa and P. s. pv. syringae multiplication in both A. deliciosa and A. chinensis twigs. Thus, the damage from frost, freeze thawing and the accumulation of Psa in Actinidia twigs promotes the migration of the pathogen within and between the orchards. Taken together, the results obtained in this study confirmed that A. deliciosa is more frost tolerant than A. chinensis, autumn frosts are more dangerous to these crops than winter frosts, and in the absence of Psa, young kiwifruit plants remain sensitive to frost.     

Understanding the effects of multiple sources of seasonality on the risk of pathogen spread to vineyards: vector pressure,natural infectivity,and host recovery

Seasonality plays an important role in the dynamics of infectious disease. For vector‐borne pathogens, the effects of seasonality may be manifested in the variability in vector abundance, vector infectiousness, and host‐infection dynamics over the year. The relative importance of multiple sources of seasonality on the spread of a plant pathogen, Xylella fastidiosa, into vineyards was explored. Observed seasonal population densities of the primary leafhopper vector, Graphocephala atropunctata, from 8 years of surveys in northern California were incorporated into a model of primary spread to estimate the risk of pathogen infection under different scenarios regarding seasonality in vector natural infectivity (i.e. constant or increasing over the season) and grapevine recovery from infection (i.e. none or seasonal recovery). The extent to which local climatic conditions affect risk estimates via differences in vector abundance was investigated. Seasonal natural infectivity, seasonal recovery, and especially the combination, reduced (up to 8‐fold on average) within‐season and cumulative yearly estimates of pathogen spread. Estimated risk of infection also differed greatly among years due to large differences in vector abundance, with wet and moderate winter and spring conditions favouring higher G. atropunctata abundance. Seasonal variation of the pathogen–vector interaction may play an important role in the dynamics of disease in vineyards, reducing the potential prevalence from what it could be in their absence. Moreover, climate, by affecting sharpshooter leafhopper abundance or activity, may influence Pierce’s disease dynamics.     

First report of the canker pathogen Endothia gyrosa on Eucalyptus in South Africa

During country-wide surveys of Eucalyptus plantations for Cryphonectria canker, cankers distinctly different from those usually associated with Cryphonectria cubensis were observed. These cankers were less severe and were exemplified by cracked and slightly swollen areas on the bark. Endothia gyrosa , a well-known pathogen of woody plants including Eucalyptus spp., was consistently associated with these cankers. The pathogen is easily distinguished from C. cubensis by the presence of orange-brown stromata and non-septate ascospores. Inoculations on Eucalyptus grandis resulted in lesions similar to those observed on naturally infected trees. The disease associated with E. gyrosa is widespread in South Africa, and research is required to establish control strategies.     

Sporulation potential,symptom expression and detection of Phytophthora ramorum on larch needles and other foliar hosts

Phytophthora ramorum has caused extensive dieback and mortality of commercially grown Japanese larch (Larix kaempferi) in many parts of the UK, as infected foliage generates spores that then cause bark lesions and girdling cankers on trees. Following inoculation, individual needles of Japanese, European (L. decidua) and hybrid (L. × eurolepis) larch infected with P. ramorum can produce thousands of sporangia. Mean numbers of sporangia ranged from 806 to 1778 per cm² (hybrid larch and Japanese larch, respectively), surpassing mean sporulation levels on foliar hosts previously associated with P. ramorum outbreaks in Britain, namely Rhododendron ponticum, Castanea sativa and Vaccinium myrtillus. Sporulation on larch even exceeded that of California bay laurel (Umbellularia californica), which drives the sudden oak death epidemic in California. Inoculation of foliage selected at different times of year revealed that foliage age significantly affected sporulation levels, but this varied with host species. However, symptom development and sporulation were often not correlated. Symptoms on larch were frequently insignificant or even absent at certain times of year, with sometimes the only evidence of infection being the emergence of sporangia from needles, without any sign of discolouration or necrosis. Plating infected but symptomless needles onto Phytophthora selective medium also often failed to yield the pathogen. Symptomless infection of larch needles apparently occurs, but is only detectable with microscopy. More generally, it is suggested that diagnosis of Phytophthora infection in conifers is often underestimated due to isolation difficulties and delayed symptom expression.     

Highly specific assays to detect isolates of Pseudomonas syringae pv. actinidiae biovar 3 and Pseudomonas syringae pv. actinidifoliorum directly from plant material

Pseudomonas syringae pv. actinidiae (Psa) is responsible for bacterial canker of kiwifruit. Biovar 3 of Psa (Psa3) has been causing widespread damage to yellow‐ and green‐fleshed kiwifruit (Actinidia spp.) cultivars in all the major kiwifruit‐producing countries in the world. In some areas, including New Zealand, P. syringae pv. actinidifoliorum (Pfm), another bacterial pathogen of kiwifruit, was initially classified as a low virulence biovar of Psa. Ability to rapidly distinguish between these pathovars is vital to the management of bacterial canker. Whole genome sequencing (WGS) data were used to develop PCR assays to specifically detect Psa3 and Pfm from field‐collected material without the need to culture bacteria. Genomic data from 36 strains of Psa, Pfm or related isolates enabled identification of areas of genomic variation suitable for primer design. The developed assays were tested on 147 non‐target bacterial species including strains likely to be found in kiwifruit orchards. A number of assays did not proceed because although they were able to discriminate between the different Psa biovars and Pfm, they also produced amplicons from other unrelated bacteria. This could have resulted in false positives from environmental samples, and demonstrates the care that is required when applying assays devised for pure cultures to field‐collected samples. The strategy described here for developing assays for distinguishing strains of closely related pathogens could be applied to other diseases with characteristics similar to Psa.