Identification and characterization of fungi associated with internal wood lesions and decline disease of willow and poplar trees in Iran

During the study of fungal trunk pathogens associated with urban trees decline in Shiraz (Iran), a serious decline of willow and poplar trees was observed. Therefore, an investigation was conducted on these trees in some areas of this city during spring and summer 2012 and 2013, to determine the main fungal trunk pathogens associated with these ornamental plants. Plant materials were collected from trees exhibiting disease symptoms such as yellowing, shoot canker, shoot dieback, defoliation and internal wood necrosis and decayed wood. Fungal isolations were made from discoloured or decayed wood tissue onto 2% malt extract agar (MEA) amended with streptomycin sulphate. Nine species, Fomes fomentarius, Diplodia seriata, Lasiodiplodia theobromae, Dothiorella sarmentorum, Neoscytalidium hyalinum, Diatrype whitmanensis, Phaeoacremonium rubrigenum, P. aleophilum and P. parasiticum, were identified based on morphology and DNA sequence comparisons. Pathogenicity tests were performed on detached shoots of willow and poplar trees under greenhouse conditions. Lasiodiplodia theobromae caused the longest lesions on willow. On poplar shoots, the longest lesions were caused by P. parasiticum. Diplodia seriata produced the smallest lesions on both woody hosts. First reports from willow wood include P. parasiticum, P. rubrigenum, D. whitmanensis, L. theobromae, D. seriata and N. hyalinum, while new reports from poplar wood include P. parasiticum and Do. sarmentorum. Based on our knowledge, this is also the first report of D. whitmanensis in Iran.     

Symptoms and fungi associated with elm trees decline in Iran

Elm (Ulmus, Ulmaceae) is one of the most popular and important trees in urban and landscape areas in Iran. A severe decline of elm trees has recently been observed in some areas of Kerman and Shiraz cities. However, the identity of the causal agents has not yet been investigated. The purpose of this study, carried out in the years 2012–2014, was to isolate and identify fungal trunk pathogens associated with decline symptoms observed in elm trees. Samples were collected from trunks and branches of trees showing various disease symptoms and internal wood lesions. Fungal isolations were made from discolored or decayed wood tissue. In Kerman some elm trees were attacked by the beetle Aeolesthes sarta; therefore, samples were also collected from larvae and adults of beetles associated with trees showing wood damage. Fungal isolates were identified by morphological, cultural and molecular characteristics. Thirteen fungal species, Phaeoacremonium (P.) minimum, P. parasiticum, P. sicilianum, P. alvesii, P. fraxinopennsylvanicum, Spencermartinsia viticola, Dothiorella (Do.) sarmentorum, Neoscytalidium hyalinum, Diatrype (Di.) whitmanensis, Cosmospora viridescens, Phoma (Pho.) herbarum, Phellinus (Phe.) tuberculosus and Inonotus levis were identified. Phaeoacremonium parasiticum was the only species isolated from both larvae and adult beetles. Pathogenicity trials were performed on detached shoots of elm under greenhouse conditions. Dothiorella sarmentorum was the most virulent species based on the length of wood necrosis. This study is the first report of P. sicilianum, C. viridescens and Phe. tuberculosus in Iran. In addition, most of the isolated species are reported for the first time on elm trees in the world.     

Mycobiota in needles of Abies alba with and without symptoms of Herpotrichia needle browning

The ascomycete Nematostoma parasiticum (syn.: Herpotrichia parasitica) is commonly perceived as the causal agent of the so‐called Herpotrichia needle browning in silver fir (Abies alba). However, its fruitbodies are rarely present on symptomatic needles, which are also colonized by many presumably saprotrophic fungi. We compared the internal colonization of healthy and symptomatic needles on two sites in Poland. In addition, the endophytic mycobiota in needles of various age was recorded on two other sites without disease symptoms. Fungi were isolated from 95.6% of the dead needles and from 62.9% of the living needles on symptomatic trees, whereas on healthy trees, only 45.0% of the needles were colonized internally. Colonization frequency increased with needle age. From a total of 2017 isolates, 116 fungal taxa were identified. Frequency of many species was influenced by needle type. Anthostomella formosa, Gloeosporidiella sp., Hypoxylon fragiforme, Xylaria hypoxylon and X. polymorpha were the most common fungi isolated from living needles. In symptomless living needles, fungi occurred significantly more often in the basal than in the apical parts. In dead needles, the most common fungi were Alternaria alternata, Paraconiothyrium sporulosum, Fusarium sp., Mollisia cinerea, Rhizoctonia sp., Rhizosphaera oudemansii, Thysanophora penicillioides, Xylaria hypoxylon and X. polymorpha. Rhizoctonia sp. was the most frequently isolated fungus in dead needles (23.4%) but occurred rarely also in living needles (0.3–1.1%). The supposed pathogen N. parasiticum was detected only sporadically (at most in 0.6% of the needles). Our findings demonstrate the need for understanding the role of Rhizoctonia sp. in Herpotrichia needle browning disease aetiology.     

Brenneria spp. and Rahnella victoriana associated with acute oak decline symptoms on oak and hornbeam in Iran

Decline diseases of forest trees are complex syndromes not attributable to single causal factors. In Iran, symptoms of decline disease have been observed in a number of native forest species including Quercus castaneifolia (chestnut‐leaved oak), Q. brantii (Persian oak) and Carpinus betulus (hornbeam). The symptoms are prevalent in the northern forests and the Zagros mountain forests. There are parallels between the disease in Iran and acute oak decline (AOD) reported in the UK, specifically the presence of weeping cankers, which have been attributed to a polybacterial complex wherein Brenneria goodwinii is considered a key necrogen. Based on the AOD symptomatology, and as a first step towards discovering potential causal agents of the stem weeping symptoms of affected trees in Iran, necrotic tissues were tested primarily for the presence of B. goodwinii. Symptomatic Q. castaneifolia and C. betulus from the Mazandaran Province and symptomatic Q. brantii from Kohgiluyeh and Boyerahmad Province were sampled. Isolation and culture on a selective medium yielded uniform bacterial colonies. Isolates were characterized using phenotypic and genotypic (DNA sequencing) tests. The isolates were phenotypically identical to members of Pectobacteriaceae and Yersiniaceae, specifically Brenneria and Rahnella spp. The nucleotide sequences of the 16S rRNA and housekeeping genes gyrB, infB and atpD (MLSA) amplified via PCR demonstrated that the isolates from the trees in Iran were indeed B. goodwinii, B. roseae subsp. roseae, Rahnella victoriana and an unknown species of Brenneria. Most bacteria isolated from non‐symptomatic trees were Gram‐positive, and Pseudomonas spp. were dominant, but bacterial species isolated from the diseased trees were not detected in healthy trees. Hypersensitivity response tests were positive, but inoculation on saplings was more variable with internal necrosis developing only once in the test period. Therefore, further testing is required. This is the first report of the incidence of B. goodwinii, B. roseae subsp. roseae, R. victoriana and Brenneria sp. associated with acute oak decline‐like symptoms on Q. castaneifolia, Q. brantii and C. betulus across the western forests of Iran and in the world.     

Diplodia corticola and Phytophthora cinnamomi: the main pathogens involved in holm oak decline on Caprera Island (Italy)

Since 2008, severe and widespread tree decline and mortality has been observed at the main growing Quercus ilex L. (holm oak) forest on Caprera Island, Italy. To clarify the symptomatology and aetiology of this phenomenon, field surveys and isolations from symptomatic trees were carried out in summer 2010. Affected trees exhibited crown thinning, branch dieback, sunken cankers, epicormic shoots, exudates on branches and trunk, root losses and sudden death symptoms. Four fungal species belonging to Botryosphaeriaceae family, namely Botryosphaeria dothidea, Diplodia corticola, D. seriata and Neofusicoccum parvum, were isolated from cankers on trunk and branches, whereas three species of Phytophthora, namely P. cinnamomi, P. cryptogea and P. gonapodyides, were isolated from fine roots and rhizosphere soil samples. Isolates were identified using both morphological analysis and DNA‐based techniques. Pathogenicity trials on holm oak seedlings showed that all the isolated species are pathogenic. D. corticola proved to be the most aggressive species. Our results provide the first evidence for a combined involvement of D. corticola and P. cinnamomi in the aetiology of holm oak decline in Italy and suggest that these pathogens are not only important contributing factors in the onset of long‐term tree decline, but also may cause the rapid devastation of extensive oak ecosystems.     

Identification of endophytic fungi of grapevine with emphasis on the causal agents of grapevine trunk disease in the vineyards of Zanjan Province in Iran

Trunk disease is a major problem on grapevine in Zanjan province, causing serious decline, despite which its aetiology remains unknown. The purpose of this study was to investigate factors involved in grapevine decline in vineyards of Zanjan province. Samples were collected from twigs and branches of grapevines in the region between October and November 2018. Fungal isolates were identified based on morphological characteristics and ITS-rDNA sequence data for selected isolates. The frequency and diversity of the fungal community recovered from grapevines in Darreh Sejin in Zanjan province were higher than in other regions. A total of 112 fungal isolates comprising 22 species were recovered. Phaeoacremonium minimum, Microsphaeropsis olivacea and Kalmusia variispora were identified as the dominant species in the region examined and could be considered the main trunk pathogens of grapevines in Zanjan region. In inoculation tests, M. olivacea was proved to be pathogenic on grapevine for the first time in this study.     

Assessment of the pathogenicity of marine Cladosporium spp. towards mangroves

Mangroves play an important role in coastal ecosystems. In this work, Cladosporium spp were isolated from a natural mangrove environment in Zhanjiang Bay, Guangdong Province, China. Ninety‐one isolates were obtained and identified as Cladosporium cladosporioides, C. colocasiae, C. oxysporum, C. perangustum, C. sphaerospermum, C. tenuissimum and C. uredinicola. Forty‐two isolates were selected and tested for pathogenicity towards mangroves. The results showed that pathogenic isolates were found on Avicennia marina, Kandelia candel and Rhizophora stylosa with prevalence of 93.40%, 83.52% and 7.41%, respectively. This indicated that A. marina and K. candel were more vulnerable to Cladosporium ssp. than R. stylosa. Among the seven species, C. colocasiae had the strongest pathogenic effect towards A. marina and K. candel with a prevalence of 100.00%. The rest of the seven species also showed pathogenic effects on the same plants. These results provided valuable insights into the effect of various pathogens on mangroves.     

First report of Cryptosphaeria pullmanensis as causal agent of Cryptosphaeria canker of Populus nigra in Iran

A diatrypaceous fungus was isolated consistently from cankers on Populus nigra trees showing dieback symptoms in Kohgiluyeh Boyer‐Ahmad, Zanjan and Esfahan provinces in Iran. Morphological characteristics and phylogenetic analyses of the ITS region of the rDNA identified the taxon as Cryptosphaeria pullmanensis. Pathogenicity tests conducted in potted 3‐month‐old cuttings of P. nigra confirmed Koch's postulates and revealed that C. pullmanensis caused canker on this host. This is the first report of C. pullmanensis causing a canker disease on P. nigra in Iran.     

Phytophthora pseudosyringae associated with the mortality of Nothofagus obliqua in a pure stand in central‐southern Chile

Mortality of Nothofagus trees in the southern‐central Chile region has been observed for over 30 years. A field survey conducted in 2013 detected partial defoliation and bleeding cankers on Nothofagus obliqua in a pure stand in the Nahuelbuta coastal ranges of the Biobío region. A Phytophthora sp. was isolated from stem cankers and soil samples around symptomatic N. obliqua trees: All isolates were identified as Phytophthora pseudosyringae. These isolates were pathogenic on 1‐year‐old N. obliqua and Nothofagus alpina, and on detached twigs of adult N. obliqua and Nothofagus dombeyi trees. This paper is the first to report association and pathogenicity of P. pseudosyringae with N. obliqua, N. alpina and N. dombeyi native to the Biobío region of Chile. The potential of P. pseudosyringae to cause damage in natural Nothofagus stands in Chile must be determined.     

Pathogenicity testing of four Bursaphelenchus species on conifer seedlings under greenhouse conditions

The pinewood nematode, Bursaphelenchus xylophilus (Steiner and Buhrer, J. Agric Res. 48, 1934, 949), Nickle (J. Nematol. 2, 1970, 375), is the causative agent of the pine wilt disease and causes serious damage to pine forests around the world. During a survey for the pinewood nematode, four other Bursaphelenchus species (Bursaphelenchus mucronatus, B. sexdentati, B. anamurius and B. vallesianus) were isolated from wilted pine trees in Turkey. To understand the effects of these Bursaphelenchus species on wilting of pine trees, a study was conducted under greenhouse conditions. Two‐year‐old seedlings of three pine species (Pinus nigra, P. brutia and P. pinea) and one cedar species (Cedrus libani) were used. Fifteen seedlings of each species were inoculated with nematodes and 10 seedlings of each species served as controls. The inoculum densities used for each seedling contained approximately 1000 (±100) nematodes of all life stages in 0.25 ml of distilled water. The first wilting symptoms were observed in the fifth week in all pine species but not in the cedar seedlings. All seeding mortality occurred between the 5th and 13th weeks of the study; no mortality was observed outside of this period. The most pathogenic nematode species was B. mucronatus, closely followed by the other species. The most susceptible seedling species was P. nigra, and C. libani was the most resistant species.     

New and unexpected host associations for Diplodia sapinea in the Western Balkans

Diplodia sapinea is an important pathogen of pine trees in plantations and urban areas in many parts of the world. This pathogen has recently also been isolated from diseased Cedrus atlantica, C. deodara and Picea omorika planted as ornamentals across the Western Balkans. The aim of this study was to consider the host range of D. sapinea in Serbia and Montenegro. Diplodia sapinea was identified from a broader collection of Botryosphaeriaceae from the Western Balkans region, based on the DNA sequence data for the internal transcribed spacer (ITS) rDNA and the translation elongation factor 1α (TEF 1‐ α). The D. sapinea isolates were obtained from sixteen tree species in the genera Abies, Cedrus, Chamaecyparis, Juniperus, Picea, Pinus, Pseudotsuga and Fagus. Four species represented new hosts in the Balkans, and this is the first report of D. sapinea from F. sylvatica anywhere in the world. Pathogenicity tests were conducted on the tree hosts from which D. sapinea was isolated, as well as on P. abies, Thuja occidentalis, Prunus laurocerasus, Eucalyptus grandis and P. patula. Inoculations were made on seedlings in the field, in the greenhouse or on freshly detached branches. Inoculations on P. pungens, P. omorika, P. abies, P. menziesii, A. concolor, P. nigra and P. sylvestris resulted in death of the seedlings 5–16 weeks after inoculation. Diplodia sapinea produced lesions on J. horizontalis and P. patula seedlings and F. sylvatica cut branches. Reciprocal inoculations showed that D. sapinea is not a pine‐specific pathogen, causing disease on tree species, including those from which it had not been isolated. Not surprisingly, the pathogen was most aggressive on some species of Pinaceae.     

Root‐colonizing ophiostomatoid fungi associated with dying and dead young Scots pine in Poland

Ophiostomatoid fungi are carried by various bark beetles. However, very little is known about the role of these fungi in conifer roots. We studied ophiostomatoid fungi in roots of dying and dead Pinus sylvestris trees and tested the potential phytotoxicity of some isolates using a sensitive bioassay with Lepidium sativum in Poland. Fungi were identified based on their morphology and DNA sequencing. Three ophiostomatoid fungi, Leptographium procerum, Sporothrix inflata and Ophiostoma pallidulum, were isolated from the roots. The most abundant soil‐borne fungus, S. inflata, and relatively rare O. pallidulum were isolated for the first time from roots of dying and dead pine trees. The frequency of S. inflata and O. pallidulum correlated with tree decline. The fungi were isolated more frequently from roots of dead than dying trees. Sporothrix inflata and O. pallidulum slightly reduced the stem and root growth of L. sativum. Leptographium procerum reduced more significantly root than stem growth. This species reduced root elongation 32–54% after 10–17 days of incubation.     

Virulence of Bursaphelenchus mucronatus to pine seedlings and trees under field cownditions

Bursaphelenchus mucronatus is a parasitic nematode of pine that is widely distributed in the natural pine forests of Asia and Europe. It has a very similar morphology and biology to that of Bursaphelenchus xylophilus, the causal agent of pine wilt disease, but has generally been considered to be non‐pathogenic to pine. However, in some provinces of China, B. mucronatus has been isolated from dead pine trees rather than B. xylophilus. Previous studies have shown that B. mucronatus can induce the death of pine seedlings under glasshouse conditions. To investigate the virulence of B. mucronatus, 2‐year‐old seedlings of Pinus massoniana and Pinus elliottii were inoculated with one of six isolates of B. mucronatus under field conditions in April 2014 and their condition was monitored over a year. The virulence of the six B. mucronatus isolates differed on the three host species: P. elliottii seedling mortality ranged from zero to six of the 18 inoculated seedlings, whereas P. massoniana seedling mortality ranged from four to 12 of the 18 inoculated seedlings. Three B. mucronatus isolates that appeared to cause different levels of mortality among the seedlings were used to inoculate 12‐year‐old Pinus thunbergii trees in August 2014. The trees were monitored for a year, during which time between 4 and 12 of the 18 inoculated trees in each treatment wilted and died. The average monthly temperature during the test period appeared to be similar to that of the historical average in the test areas; however, both study sites experienced above‐average rainfall. This study demonstrated that B. mucronatus has potential virulence on pine trees and provided experimental evidence that high temperatures or drought stress is not essential for the virulence of B. mucronatus.     

Eutypella canker of maple: first report from Germany and situation in Austria

Eutypella canker of maple, caused by Eutypella parasitica (which is native to North America), is reported for the first time from Germany. From 2013 to 2015, this perennial canker disease was recorded on 105 maple trees in Munich. Six maple species were affected: Acer pseudoplatanus, A. campestre, A. platanoides, A. cappadocicum, A. heldreichii ssp. trautvetteri and A. hyrcanum. Occurrence on the latter three species represents new host records for E. parasitica. In Austria, Eutypella canker was newly discovered on two trees at a second locality in 2011, and it is now known to occur on seven A. pseudoplatanus trees at two localities, which are separated nearly 150 km. A. pseudoplatanus was the most frequent host of E. parasitica in Munich and Austria, which is in agreement with previous studies in Europe. The identity of the causative pathogen as E. parasitica was verified by ITS rDNA sequencing of fungal cultures obtained from cankers in Munich and at both Austrian localities. The presence of large and old cankers in both countries suggests that introduction of E. parasitica dates back a long time, probably several decades. The new records of Eutypella canker in Germany and Austria show that the disease is more widely distributed in central Europe than previously recognized.     

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

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

Fungal root endophytes of Quercus robur subjected to flooding

In August 2011, fungal endophytes were isolated from fine (0.1–0.5 cm diameter) and structural (0.6–2.0 cm) roots from eighteen Q. robur trees at six sites in a Q. robur‐dominated forest displaying symptoms of oak decline. Five sites were subjected to periodic flooding for 3 months in 1997 and 1 month in 2010. Fungal colonization was detected in 94% of 1296 root fragments examined. Fungi representing 126 taxa from Zygomycota (four species), Ascomycota (117 species) and Basidiomycota (five species) were isolated using culture‐based methods and identified by morphotyping and sequencing of the ITS 1 and 2 rDNA. The most frequent eudominants (with colonization frequencies ≥10%) were Aspergillus spp., Cosmospora sp., Ilyonectria radicicola, Pezicula radicicola, Pyrenochaeta cava and six species of Trichoderma. Species composition did not vary greatly between sites, but only 3% of species occurred in all sites. Number of species was higher in roots subjected to floods. Eudominants and dominants present only on flooded sites included Aspergillus spp., Chaetomium globosum, Cosmospora sp., I. radicicola, Sporothrix inflata, Trichoderma harzianum, T. pubescens and T. virens. Structural roots were usually colonized by a higher number of species than fine roots. The majority of species detected belong to Class 2 NC‐endophytes. Pezicula radicicola and Phialocephala fortinii belong to Class 4 NC‐endophytes.     

Occurrence of brown root rot in Ocean Expo Park,Okinawa, Japan: Infection route and preventive control measures

Ocean Expo Park, established in 1976 on subtropical Okinawa Island, Japan, has approximately 8000 planted or naturally grown trees in an area of 77.2 ha. Windfall of these trees occurs mainly due to typhoons; wood chips made from the trunks and roots of windfall trees are commonly used as mulching material throughout the park areas, despite possible infection by pathogenic fungi such as Phellinus noxius, the pathogen causing brown root rot. P. noxius is distributed in tropical and subtropical areas worldwide and has a broad host range. The fungus can infect host plants with mycelia via root-to-root contact or with basidiospores. Here, we report the current status of tree damage caused by brown root rot in the park and infer the infection routes of P. noxius based on microsatellite markers. Among a total of 294 trees surveyed in the park, P. noxius was isolated from 13 trees of five species. P. noxius was observed more frequently on Ficus microcarpa than on other tree species. Ficus superba var. japonica was identified as a new host species, and Bombax ceiba was first recorded as a host plant of P. noxius in Japan. Among the 13 P. noxius isolates from the diseased trees, 2 isolates from adjacent diseased trees were considered to be of the same genet, while the other 11 isolates were different genets, indicating that infection by basidiospores occurred frequently among trees in the entire park, although spread via root systems also occurred between at least two adjacent trees. An identical genet was not detected at distant locations, which suggests that the disease does not spread via chips. However, mulching with chips can lead to wood decay due to decaying fungi other than P. noxius. Therefore, in accordance with the precautionary principle, the park currently prohibits the chipping and mulching of windfall or damaged trees and is considering disposal methods such as burning.     

Identification and characterization of bacterial strains associated with diseased oak trees in Northern Iran

Oak decline syndrome is characterized by periodic occurrences of decline and death of oaks over widespread areas. An outbreak of a new emerging disease on oak trees was reported in the Hyrcanian forest of Iran (Mazandaran and Golestan provinces) that showed stem bleeding and canker symptoms. Bacterial isolates were characterized through biochemical and physiological tests, protein electrophoresis, DNA fingerprinting (rep‐PCR, ERIC and BOX primers) and sequencing of 16S rRNA and MLSA (multilocus sequencing analysis) for housekeeping genes (gyrB, infB and atpD). A complex community of the genus Brenneria spp. (Brenneria goodwinii, Brenneria roseae subsp. roseae, Brenneria sp. and Brenneria nigrifluens) and a few isolates in the genus Gibbsiella were identified as major groups involved. Isolate differentiation was more accurate using concatenated partial gene sequences within the main groups. All bacterial isolates showed hypersensitivity reactions (HR) on Pelargonium leaves (Pelargonium × hortorum). Pathogenicity studies of different Brenneria and Gibbsiella strains revealed that they have potential to cause the disease in oak seedlings and devastating oak canker and stem bleeding symptoms in northern Iran. Due to the presence of several potentially pathogenic agent(s) associated with the oak decline, identification of the principal agent(s) is of major interest. To our knowledge, this is the first report of potentially pathogenic bacteria associated with oak bleeding and canker in Iran.     

Variation in susceptibility of six pine species and hybrids to pitch canker caused by Fusarium circinatum

Six pine species or hybrids were tested for susceptibility to pitch canker caused by Fusarium  circinatum. Pinus  densiflora, Pinus  thunbergii, Pinus  x rigitaeda (Pinus  rigida × Pinus  taeda), P. rigida × P. x rigitaeda, Pinus  echinata and Pinus  virginiana were inoculated with three spore loads (50, 500 and 5000 per tree) of F. circinatum. External symptoms, lesion length, and the frequency of reisolation of the fungus were investigated. External symptoms were greatest in P. echinata, followed by P. virginiana, however, P. densiflora was not susceptible to F. circinatum. Based on mean lesion lengths, the six pine species or hybrids differed significantly (p < 0.01) in susceptibility to pitch canker. Pinus  echinata sustained the longest lesions, whereas P. densiflora sustained the shortest lesions. The effect of inoculum density was not significant among three spore treatments within species (p = 0.17), although lesion length was slightly greater at higher spore loads over all pine species. The fungus was reisolated from inoculated stems of all pine species tested, even on trees showing little or no damage from the disease. Additional studies are needed to further explore the basis for resistance to pitch canker.     

Field survey,isolation, identification and pathogenicity of Phytophthora species associated with a Mediterranean‐type tree species

Corymbia calophylla (marri), a keystone tree species in the global biodiversity hot spot of southwestern Australia, is suffering decline and mortality associated with a canker disease caused by the endemic fungus Quambalaria coyrecup. Phytophthora species are frequently isolated from the rhizosphere of C. calophylla, and a hypothesis is that Phytophthora root infection is predisposing C. calophylla to this endemic canker pathogen. Field surveys were conducted in both anthropogenically disturbed and undisturbed C. calophylla stands, from where a total of 100 rhizosphere soil samples, from both healthy and cankered trees, were collected. Phytophthora species were isolated from 26% of the samples collected, with Phytophthora incidence significantly higher on disturbed stands than in natural forests (73% and 27%, respectively). Five Phytophthora species were recovered, including P. cinnamomi, P. elongata, P. multivora, P. pseudocryptogea and P. versiformis. Under‐bark inoculations with the Phytophthora isolates caused significant lesion lengths in excised C. calophylla stems. Corymbia calophylla response to pot infestation trials in the glasshouse varied between Phytophthora species and isolates, with isolates of P. cinnamomi and P. multivora causing a significant reduction in seedling root volume and often leading to seedling death. This study demonstrates that root disease caused by Phytophthora species, especially P. cinnamomi and P. multivora, has the ability to adversely affect C. calophylla health. This study leads the way to do a dual inoculation trial with the canker pathogen Q. coyrecup, and different Phytophthora species to investigate if Phytophthora root infection predisposes C. calophylla to this canker disease.