Phytophthora Crown and Root Rot of Apricot Caused by Phytophthora palmivora in Turkey

Forty‐nine Phytophthora isolates were obtained from roots and crown of apricot trees with symptoms of decline grown in commercial orchards in Malatya, Elaz?? and Diyarbak?r provinces, Turkey, in 2011 and 2013. All of the recovered isolates were identified as Phytophthora palmivora on the basis of morphological characteristics. Blast analysis of ITS region sequences of rDNA of 5 isolates revealed 100% identity with a reference isolates of P. palmivora from GenBank. Isolates of P. palmivora were pathogenic on 12‐month‐old wild apricot rootstock ‘Zerdali’ plants that were wound inoculated on the roots and on the crown. This study demonstrated that P. palmivora is the cause of the crown and root rot found on apricot in Turkey. To our knowledge, this is the first report of P. palmivora on this host plant.     

Phytophthora cinnamomi Associated with Root and Crown Rot of Walnut in Turkey

Walnut decline caused by Phytophthora sp. occurred in an orchard in Sakarya province in Turkey. Affected young trees showed poor growth, leaf discolouration, root and crown rot and eventual death. A Phytophthora sp. isolated from necrotic taproots and crown tissues. The causal agent of the disease was identified as Phytophthora cinnamomi by morphological characteristics and comparing sequences of internal transcribed spacer (ITS) region. Upon conducting pathogenicity test, averaging 7.8‐cm‐long canker developed on basal stem within 2 weeks, while no cankers developed in the control plants.     

Outbreak of Phytophthora cinnamomi causing severe decline of avocado trees in southern Turkey

Since the summer 2017, severe decline symptoms have been observed on 10- to 25-year-old avocado trees in almost all commercial orchards planted in the Mediterranean coastal region of Turkey. Young, newly planted trees in infected orchards were also affected by the disease. Affected trees showed wilting, leaf discoloration, defoliation and severe dieback. Some trees were completely desiccated. Although fine roots of symptomatic trees usually were decayed, reddish brown cankers also occurred on taproots and lateral roots of heavily infected trees. The pathogens were isolated from necrotic root and soil samples of symptomatic trees, using selective medium and soil baiting, and were identified based on morphological features and DNA sequences of the internal transcribed spacer (ITS) region. One isolate each of Phytophthora cryptogea and P. palmivora was identified, while all other isolates were P. cinnamomi. In addition, a subcortical fan-shaped mycelium, characteristic of Armillaria spp., was observed in the stem base of a symptomatic tree and identified as Armillaria gallica by DNA sequences of the internal transcribed spacer (ITS) and the translational elongation factor 1-α (EF 1-α) gene regions. Pathogenicity of Phytophthora isolates was tested by stem inoculation on one-year-old avocado seedlings. Two months after inoculation, canker lesions developed on stems of seedlings inoculated by any of the three Phytophthora spp. In contrast, collenchyma callus formed over the wound points on control plants over the same time period. This is the first report of P. cinnamomi, P. cryptogea, P. palmivora and A. gallica causing root rot of avocado trees in Turkey. In addition, P. cryptogea and A. gallica are reported for the first time associated with disease on this host. Due to the severe symptoms and widespread occurrence, P. cinnamomi should be considered a potential threat to avocado cultivation and natural ecosystems of this region of Turkey.     

Characterization and Pathogenicity of Phomopsis theicola Anamorph of Diaporthe foeniculina Causing Stem and Shoot Cankers on Sweet Chestnut in Italy

In March 2014, an outbreak of shoot cankers was observed on grafted Castanea sativa plants in a glasshouse in central Italy. Morphological characteristics led to the identification of isolates of Phomopsis recovered from cankered stems and shoots. Based on the morphological characteristics of colony appearance, shape of conidia and conidiomata as well as sequences of internal transcribed spacer regions (ITS), actin (ACT) and translation elongation factor (TEF‐1α), the fungus was identified as Phomopsis theicola/Diaporthe foeniculina. Pathogenicity test showed that P. theicola isolates were pathogenic to C. sativa when artificially inoculated, reproducing the symptoms originally observed. Koch's postulates were fulfilled by re‐isolating the pathogen. This is the first report of P. theicola/D. foeniculina causing stem and shoot cankers and dieback on C. sativa in Italy or elsewhere.     

The Role of Phytophthora cryptogea and Waterlogging in Root Rot Kiwifruit

A 48 h or 96 h period of waterlogging induced a partial rotting of the root system of 18 month old potted kiwifruit cuttings. However, root regeneration compensated for the loss of damaged roots and the plants remained healthy. Inoculation with Phytophthora cryptogea under low soil moisture conditions caused a small amount of root rotting (11 %) and regeneration of the roots occurred. Root rot was significantly increased when inoculations were combined with extended periods of waterlogging. Root rot incidence was highest (89 %) in inoculated plants exposed to periods of 96 h waterlogging. Regeneration did not occur with any of these plants and consequently, the shoot systems were severely damaged.     

Phytophthora cryptogea root rot of tomato in rockwool nutrient culture. I. Analysis of root infection

The spatial and temporal colonisation of tomato plants by Phytophthora cryptogea was studied in rockwool nutrient culture. Root growth and the distribution and progress of infection were measured on dissected root fragments obtained from a detailed destructive sampling of the substrate. Dissected fragments of root or roots and fibres were plated on BNPRA agar, a selective Phytophthora medium. Roots colonised all parts of the rockwool substrate 60 days after planting with the exception of surface marginal and central areas of the slab which had a lower solution content. Most root biomass occurred in and immediately beneath the original growing block. The distribution of P. cryptogea closely followed the pattern of root colonisation. An alternative, novel method for root analysis involved the dissolution of the mineral fibre and its formate bonding resin by digestion in 1 m H₃PO₄ for 45 min. Comparative recoveries of P. cryptogea from plated fragments of dissected root and fibre or comminuted samples of acid-released or dissected root showed that the acid treatment initially reduced the number of Phytophthora colonies in block and slab roots by 67% and 61% respectively. After 28 days, colony recovery from acid-released roots in the rockwool slab increased and was between 4% and 13% lower than from other plating methods. Since 1 m H₃PO₄ was lethal to zoospores and surface sporangia, the colonies recovered were interpreted as originating exclusively from root lesions. Root fresh weight of healthy and inoculated plants declined during the initial period of fruit formation. P. cryptogea infection led to a progressive reduction in root weight in the growing block and main slab and 28 days after inoculation, was approximately 50% of the controls. The acid digestion of rockwool fibre is proposed as a new approach to the problem of root and pathogen analysis in this substrate.     

The effect of chemical and biological treatment on root and crown rot disease caused by Phytophthora cryptogea Pethybr. & Lafferty in Gerbera

Phytophthora root and crown rot (Phytophthora cryptogea) on gerbera is difficult to manage because most gerbera cultivars are susceptible to P. cryptogea. This study was conducted in order to determine the in vivo (pot experiment) efficacy of some fungicides and biofungicides. In pot experiments, fungicides were applied 7 days after inoculation with P. cryptogea, while biofungicide was applied 7 days before inoculation. In this study, soil drenches of five fungicides were tested. “Ametoctradin+dimethomorph (100 ml/day),” “mandipropamid+difenoconazole (60 ml/day),” “propamocarb+fosetyl‐Al (200 ml/day),” “mancozeb+metalaxyl‐M (250 g/day)” and “azoxystrobin+difenoconazole (100 ml/day)” active substances were used. Similarly, one biofungicide Bacillus amyloliquefaciens syn. MBI 600 (50 g/100 L) was applied by soil drenching. Efficacy of treatments was assessed according to the percentage of the root system which was visibly rotten at the end of the experiment. Root and crown rot severity was rated on a scale of 0 = 0% root system necrotic, 1 = 1%‐25% necrotic, 2 = 26%‐50% necrotic, 3 = 51%‐75% necrotic and 4 = 76%‐100% necrotic from 12 to 21 days. In this experiment, “azoxystrobin 200 g/L + difenoconazole 125 g/L” exhibited the highest efficacy against P. cryptogea with a ratio of 43.75%. The other fungicides and biofungicides ametoctradin 300 g/L + dimethomorph 225 g/L, mandipropamid 250 g/L + difenoconazole 250 g/L, propamocarb 530 g/L + fosety‐Al 310 g/L, mancozeb 64%+metalaxyl‐M 4% and Bacillus amyloliquefaciens syn. MBI 600 11% were ineffective. Importance should be given to management strategies of P. cryptogea of and more experiments should be carried out for a better understanding of the use of registered fungicides and biofungicides.     

Optimization of Culturing Conditions of a Strain of Phytophthora capsici Pathogenic to Habanero Pepper (Capsicum chinense)

Phytophthora capsici is an oomycete known as the causal agent of wilting disease in Capsicum spp., which causes rotting of roots, crowns, stems, leaves and fruits. To date, little is known about the production of phytotoxic metabolites by P. capsici or their role in the infection process. As part of a project directed towards the isolation and identification of phytotoxins produced by a strain of P. capsici pathogenic to habanero pepper (Capsicum chinense), we have evaluated the effect of factors such as aeration, light and culture medium on the production of mycelium and phytotoxic metabolites by P. capsici. The results showed that culturing P. capsici in potato dextrose broth (PDB) containing habanero pepper leaf infusion, in the dark and under still conditions, results in a high production of mycelium and a high phytotoxicity of the culture filtrate, in the shortest period of time.     

Evaluation of fatty acid methyl ester profile and amplified fragment length polymorphism analyses to distinguish five species of Phytophthora associated with ornamental plants

Fatty acid methyl ester (FAME) profiles and amplified fragment length polymorphisms (AFLPs) were evaluated as tools for identifying species of Phytophthora. Five isolates of each of Phytophthora cactorum, Phytophthora citrophthora, Phytophthora cinnamomi, Phytophthora nicotianae and Phytophthora cryptogea were subjected to both analyses to examine variation among and within species. In FAME analysis, isolates of P. cactorum, P. cinnamomi and P.nicotianae were clustered by species, but isolates of P. citrophthora and P.cryptogea were divided into multiple clusters based on greater variations within these two species. The AFLP analysis differentiated all five species of Phytophthora. The five isolates of each species were grouped in a separate terminal cluster, but diversity within a species cluster varied considerably with variation greater in P. cryptogea and P. citrophthora. Comparing the dendrograms based on FAME and AFLP analyses, the overall patterns of both were similar. The P. cactorum cluster was distinct from clusters of the other four species, which formed one large cluster. The higher values of percentages of polymorphic loci and gene diversity in AFLP analysis substantiated diversity observed among isolates of P. citrophthora and P. cryptogea in FAME and AFLP dendrograms. Both FAME and AFLP appear to be useful tools for identifying species of Phytophthora, but only AFLP analysis has potential to study genetic and phylogenetic relationships within and among species in this genus.     

Occurrence of Phomopsis longicolla β Conidia in Naturally Infected Soybean

Although Phomopsis longicolla is primarily known as a seedborne pathogen, it can be isolated from all parts of the plant. The disease lesions observed on the basal parts of soybean stems were slightly sunken with irregular shapes and sizes, bordered by a thin black margin. Within the lesions themselves, large and diffusely distributed pycnidia with α and β conidia, typical of the genus Phomopsis, were observed. The percentages of the two types of conidia varied considerably, but β conidia were predominant in most of the pycnidia. The presence of these reproductive organs indicated that the symptoms could have been caused by Phomopsis sojae. However, after isolation on a nutritive medium, all cultural and morphological characteristics clearly indicated that the isolated fungus was P. longicolla, whose identification was subsequently confirmed by sequencing three genomic regions. Monosporic isolates, with different ratios of α and β conidia, exhibited a high level of pathogenicity on soybean, after artificial inoculation. Both types of conidia were observed on the stems of the inoculated soybean plants. Beta conidia also formed quickly on medium made of soybean seeds and mature stems after exposure to low temperatures (?10°C). This study suggests that P. longicolla is capable of a massive production of β conidia, not only in old fungal cultures as it had until now been believed, but also in infected soybean plants in the field.     

Phylogenetic relationship of Phytophthora cryptogea Pethybr. & Laff and P. drechsleri Tucker

The phylogeny and taxonomy of Phytophthora cryptogea and Phytophthora drechsleri has long been a matter of controversy. To re-evaluate this, a worldwide collection of 117 isolates assigned to either P. cryptogea, P. drechsleri or their sister taxon, Phytophthora erythroseptica were assessed for morphological, physiological (pathological, cultural, temperature relations, mating) and molecular traits. Multiple gene phylogenetic analysis was performed on DNA sequences of nuclear (internal transcribed spacers (ITS), ß-tubulin, translation elongation factor 1α, elicitin) and mitochondrial (cytochrome c oxidase subunit I) genes. Congruence was observed between the different phylogenetic data sets and established that P. drechsleri and P. cryptogea are distinct species. Isolates of P. drechsleri form a monophyletic grouping with low levels of intraspecific diversity whereas P. cryptogea is more variable. Three distinct phylogenetic groups were noted within P. cryptogea with an intermediate group providing strong evidence for introgression of previously isolated lineages. This evidence suggests that P. cryptogea is an operational taxonomic unit and should remain a single species. Of all the morphological and physiological traits only growth rate at higher temperatures reliably discriminated isolates of P. drechsleri and P. cryptogea. As a homothallic taxon, P. erythroseptica, considered the cause of potato pink rot, is clearly different in mating behaviour from the other two species. Pathogenicity, however, was not a reliable characteristic as all isolates of the three species formed pink rot in potato tubers. The phylogenetic evidence suggests P. erythroseptica has evolved from P. cryptogea more recently than the split from the most recent common ancestor of all three species. However, more data and more isolates of authentic P. erythroseptica are needed to fully evaluate the taxonomic position of this species.     

Inoculation of Paenibacillus illinoisensis alleviates root mortality, activates of lignification-related enzymes, and induction of the isozymes in pepper plants infected by Phytophthora capsici

To investigate the variations of the enzymes responsible for lignification, after inoculation with Phytophthora capsici and/or Paenibacillus illinoisensis KJA-424, in relation to biocontrol of Phytophthora blight in pepper, roots of two-month-old plants were inoculated with P. capsici inoculation (P), and co-inoculation of P. capsici and P. illinoisensis cell cultures (P + A). Root mortality of pepper plants induced by inoculation of P. capsici was completely recovered by co-inoculation with antagonistic KJA-424. At day 7, peroxidase (POD) activity increased by 36.7% in P-treated roots but by 7.1% only in P + A-treated, compared with control. Polyphenol oxidase (PPO) activity increased for 3 days and then drastically decreased in P-treated roots but maintained a constant level in control and P + A-treated. At day 7, PPO activity in P-treated leaves decreased but recovered to the level of control in the P + A-treated. Three major POD isozymes (45, 53, and 114 kDa) were shown in P-treated roots, while two major (53 and 114 kDa) in control and P + A-treated, suggesting that the 45 kDa of POD was actively induced in P-treated roots but not induced in P + A-treated roots. A PPO isozyme of 80 kDa was induced in P-treated roots but not induced by co-treated with KJA-424. In leaves, the POD isozyme of 45 kDa appears to be systemically induced in P-treated only. The PPO isozyme of 80 kDa in leaves was not induced by pathogen challenge but recovered by co-inoculated with P. illinoisensis. All these results suggest that the inoculation of an antagonist, P. illinoisensis alleviates root mortality, activates of lignification-related enzymes and induction of the isozymes in pepper plants infected by P. capsici.     

First Report of Nigrospora oryzae (Berk. & Broome) Petch Causing Stem Blight on Brassica juncea in India

A stem blight disease was observed on the lower portions of Brassica juncea stems during the cropping season (2010–2011). In advanced stages, the lesions were up to 120 cm in length on the stems and also spread to petioles and midribs of leaves. The purified fungus was identified as Nigrospora oryzae (Berk. & Br.) Petch (teleomorph Khuskia oryzae), which produced similar symptoms when healthy B. juncea plants were inoculated, thus proving Koch's postulates. This is the first report of the occurrence of N. oryzae on B. juncea.     

Grafting on CM‐334 controls serrano chili wilting caused by Phytophthora capsici and changes phenology but does not affect fruit yield

Grafting is an alternative method of plant propagation used to prevent soil‐borne diseases. This technique can improve the development of plants and therefore improve fruit yield and quality; however, several studies report that when a plant is grafted, there may be compatibility problems and changes in the phenological pattern of the crop and fruit yield and quality with respect to non‐grafted plants. There are no reports in the literature on the behaviour of serrano chilli grafted on varieties resistant to Phytophthora capsici. In this study, we evaluated the phenological behaviour and response to inoculation with P. capsici in commercial serrano chilli (Camino Real, Harris Moran) grafted or not on CM‐334 as a strategy to explore the possibility of incorporating the use of grafts in the production systems of serrano chilli. The plants were grafted at 55–60 days and maintained for 13 days in a curing chamber for the acclimatization process. At 36 and 43 days after transplantation, they were inoculated with the pathogen (300,000 zoospores/plant). None of plants grafted and inoculated with the pathogen showed wilt symptoms. All plants not grafted and inoculated with P. capsici died. There was a significant reduction in the production of leaves and flowers in the grafted plants, in relation to the non‐inoculated and non‐grafted plants, as well as a temporary delay in the beginning of fruit production with respect to the non‐grafted plants, but this delay did not affect the overall yield of the crop.     

Migration of the Fungal Protein Cryptogein within Tobacco Plants

Cryptogein (CRY), a protein secreted by Phytophthora cryptogea, causes necrosis on tobacco (Nicotiana tabacum) plants at the site of application (the stem or the roots) and also on distant leaves. Autoradiography of plantlets after root absorption of radioiodinated CRY demonstrated a rapid migration of the label to the leaf lamina via the veins. Using an anti-CRY antiserum, a CRY-related antigen was detected in the stem and leaves of CRY-treated plants at a distance from the site of application. This antigen had the same molecular weight as CRY and was detected in the leaves as early as 1 hour after stem treatment, i.e. long before necrosis was detectable. The antigen was also detected in plants inoculated with P. cryptogea. The distant location of the necrosis induced by the fungus or by CRY can be ascribed to the migration of this protein, which is toxic to tobacco cells. It is proposed that CRY, which also elicits defense reactions in tobacco, might contribute to the hypersensitive response of tobacco to P. cryptogea.     

Effect of tissue wounding and time of soil inoculation on pepper root rot development

Five-week-old pepper plants with wounds created on stems and roots were transplanted to soils having inoculum of Phytophthora capsici incorporated for different lengths of time. Disease severity (39.99%) on root trimmed seedlings was not significantly different (P ≤ 0.05) from the severity (36.24%) obtained on stem lacerated seedlings. The wound treatments did not result in significantly different rates of lesion extension per day; stem lacerated seedling had the fastest, 1.99 mm/day lesion extension rate, followed by 1.90 and 1.89 mm/day extension rates obtained on root trimmed and unwounded treatments, respectively. However, time of soil inoculation had significant effect on severity; root trimmed and stem lacerated treatments had 46.3% and 39.8% severities, respectively. Tissue wounding × time of soil inoculation interaction did not have significant effect on disease severity; stem lacerated seedlings transplanted to 1-day and 3-day inoculated soils gave highest severity (49.9%), followed by seedlings inoculated at the time of transplantation. Root trimmed seedlings inoculated at the time of transplantation had highest severity (61.1%), while the lowest severity was obtained on seedlings transplanted to 5-day inoculated soil.     

Biocontrol of Late Blight Disease (Phytophthora capsici) of Pepper and the Plant Growth Promotion by Paenibacillus ehimensis KWN38

For field application of a bacterial strain used to control Phythophthora capsici, we will need a biologically and economically efficient carrier medium. The known antagonist Paenibacillus ehimensisKWN38 was grown in a grass medium where it showed high antifungal and lytic enzyme activities. To demonstrate the potential of P. ehimensisKWN38 for biocontrol of late blight disease in pepper, pot trials were conducted by treating the 1‐month‐old plants with water (W), a selected grass medium (G3), G plus P. ehimensisKWN38 inoculation (G3P) or synthetic fungicide (F). The shoot dry weight in G3P was higher than that in W and F treatments at 15 days after zoospore infection (DZI). The root dry weight in G3P was also higher than that in W. The root mortality of G3 and W increased over 58 and 80% at 15 DZI, and some plants in those treatments wilted due to the failure of root physiology. The plants in G3P and F survived well because of their better root health conditions. Soil cellulase activity of G3P was consistently higher than that of W and F at earlier observation times (0, 2 and 6 DZI). The root β‐1,3‐glucanase activity of G3P promptly increased to maximum shortly after zoospore infection and reached the maximum value of 51.12 unit g^?1 of fresh weight at 2 DZI. All these results indicate that inoculation of P. ehimensisKWN38 to the root zone of potted pepper plants increases plant growth, root and soil enzyme activities and alleviates the root death caused by infection with P. capsici zoospores.     

The speciation and distribution characteristics of Cu in Phragmites australis (Cav.) Trin ex. Steudel

• Heavy metal allocation and the mechanism(s) of metal sequestration in different clonal organs, micro‐domains and subcellular structures has not been systematically studied for rhizomatous perennial plants. It is thus pertinent to investigate knowledge of the speciation and distribution characteristics of Cu in Phragmites australis to elucidating the mobility of metals in wetland plants after their uptake via root systems so as to facilitate development of strategies to enhance Cu tolerance.
• This study investigated the distributions of Cu in P. australis root, stem and leaf using ICP‐MS, synchrotron‐based X‐ray micro‐fluorescence and X‐ray absorption spectroscopy, then evaluated the effects of Cu on cellular structure and ultrastructure via transmission electron microscopy.
• The results indicate a clear preferential localisation of Cu in the roots as compared with the shoots (stems and leaves). The intensity of Cu in the vascular bundles was higher than that in the surrounding epidermis and the endodermis and parenchyma outside the medullary cavity. The dominant chemical form of Cu in P. australis was similar to Cu citrate.
• The results suggest that although Cu can be easily transported into the vascular tissues in roots and stems via Cu citrate, most of the metal absorbed by plants is retained in the roots because if its high binding to the cell wall, thus preventing metal translocation to aerial parts of the plants. Therefore, P. australis showed a high capacity to accumulate Cu in roots, being therefore a suitable species for phytostabilisation interventions.

     

Evaluation of Resistance of Stone Fruit Rootstocks to Phytophthora Crown Rot*

The pathogenicity of Phytophthora citrophthora, Phy‐tophthora cactorum and Phytophthora megasperma strains was confirmed on 2‐year‐old stone fruit rootstocks artificially inoculated in the field. Phytophthora mega‐sperma was not virulent or less virulent than P. cactorum or P. citrophthora. The rootstocks Damas and AN 1/7 were resistant whereas the rootstocks Jl, Titan x Nemaguard 5/1, GF 677, Myrandier 613, Italian x Nemaguard 1 and Myrandier 617 were susceptible to P. citrophthora. The rootstocks Tsukuba 9, Tit‐an x Nemaguard 4/7, AN 1/2, AN 1/3, St. Mien 655/2 and Bl showed medium susceptibility to this species. The isolates of P. cactorum induced more extensive necrosis in Myrandier 617 and Italian x Nemaguard than in Myr‐andier 613, GF 677, Jl (ADAFUEL), Tsukuba 9 and St. Julien 655/2. Among seven rootstocks, Tsukuba 5 and St. Julien 655/2 were more resistant than Myrandier 613, Myrandier 617, Jl (ADAFUEL), Tsukuba 9 and St. Julien 655/2.     

Root discoloration and shoot symptoms in silver birch after Phytophthora infection in vitro

• There are no records of established plant pathogenic Phytophthora species in Finnish forests, but they are likely in the future. Therefore, the effects of Phytophthora inoculations on young, ca. 2‐month‐old silver birch (Betula pendula) seedling roots and shoots were investigated.
• Visual inspection of dark discoloration, direct PCR and re‐isolation, and detailed root morphology analyses were used to evaluate the effects of Phytophthora inoculation on roots. Symptoms in leaves and stems were also recorded.
• Phytophthora was successfully re‐isolated from 67% of the surface‐sterilized roots of inoculated seedlings, but not from the non‐inoculated control seedlings. Dark discolorations were found more often in the root segments of inoculated seedlings than in control seedlings. In the Phytophthora‐treated seedlings, discoloured root segments were usually linked and found primarily in the main root or lateral roots attached to it, whereas in the control seedlings a few single discoloured root segments were scattered throughout the root systems. The number of root segments was lower in the inoculated than in the control seedlings, indicating root loss after Phytophthora inoculation. In the shoots of inoculated birches, leaf and shoot wilting was observed.
• The appearance of wilting in shoots without visible dark discoloration in the base of stems indicated that symptoms originated from roots inoculated with Phytophthora.