Phylogeny, Morphology, Distribution, and Pathogenicity of Seven Calonectria Species from Leaf-Blighted Eucalyptus in HaiNan Island, China.

Leaf blight caused by Calonectria species constrains Eucalyptus trees in China. Calonectria leaf disease on Eucalyptus in China was first reported in HaiNan Island in 1985. No systematic investigation of Calonectria species associated with diseased Eucalyptus in HaiNan has been performed. To understand the species diversity, distribution, and pathogenicity of these Calonectria, 400 Calonectria isolates were obtained from 278 diseased Eucalyptus planted in 17 sites in five regions. All 400 isolates were identified by DNA sequences of translation elongation factor 1-alpha, ß-tubulin, calmodulin, and histone H3 gene regions and on morphology. Seven species, C. acaciicola (198 isolates), C. pseudoreteaudii (161 isolates), C. reteaudii (29 isolates), C. hawksworthii (6 isolates), C. hongkongensis (4 isolates), C. auriculiformis (1 isolate), and C. chinensis (1 isolate), were identified. This is the first report of C. acaciicola in China. C. acaciicola, C. pseudoreteaudii, and C. reteaudii belong to the C. reteaudii species complex and accounted for 97% of all isolates. The three species overlapped in vesicle shape, macroconidia size, and macroconidia septa number. Region significantly influenced C. acaciicola and C. pseudoreteaudii distribution. Representative isolates of C. acaciicola, C. pseudoreteaudii, C. reteaudii, and C. hawksworthii producing abundant macroconidia were used in conidial suspension inoculation on Eucalyptus seedlings; all were highly pathogenic to the two tested genotypes. The tolerances of two Eucalyptus genotypes were significantly different. This first systematic investigation of Calonectria species associated with Eucalyptus leaf blight in HaiNan will aid selection of disease-resistant genotypes for managing Eucalyptus leaf blight caused by Calonectria species in China.

Calonectria queenslandica: Causal Agent of Eucalyptus Leaf Blight in Southern China.

Calonectria leaf blight caused by Calonectria spp. is among the most serious diseases affecting the health and sustainability of Eucalyptus plantations in southern China. Recent outbreaks of this disease in GuangDong Province prompted a need to identify the species involved. Typical symptoms of Calonectria leaf blight were observed on 2-year-old Eucalyptus urophylla × E. grandis trees in a plantation in the ZhaoQing region. In total, 38 Calonectria isolates were collected from 32 diseased trees. All isolates were identified using DNA sequence analyses of the translation elongation factor 1-α (tef1), ß-tubulin (tub2), calmodulin (cmdA), and histone H3 (his3) gene regions. Phylogenetic analyses revealed that Calonectria queenslandica was the dominant species, accounting for 81.6% of the isolates collected. Other species isolated included C. pseudoreteaudii (10.5%), C. reteaudii (5.3%), and C. aconidialis (2.6%). This is the first report of C. queenslandica in China and all isolates had identical sequences in all four gene regions. PCR amplification using primers targeting the MAT1-1-1 and MAT1-2-1 genes in all C. queenslandica isolates revealed that only the MAT1-2 idiomorph was present. The results suggest that C. queenslandica was introduced into the sampled area with very limited genetic diversity. Pathogenicity tests were conducted on two Eucalyptus genotypes widely planted in the GuangDong Province using isolates representing all species collected. The results showed that these species could all cause disease but the predominance of C. queenslandica on infected trees suggests that it is the major driver of the disease problem studied. Different Eucalyptus genotypes used in the pathogenicity tests differed in susceptibility to infection by the Calonectria spp. tested, providing opportunities to avoid leaf blight by deploying disease-tolerant planting stock.

Comparison of Hyphal Fragments and Spores to Evaluate the Pathogenicity of the Eucalyptus Leaf and Shoot Pathogen Calonectria pseudoreteaudii.

The genus Calonectria includes many aggressive plant-pathogenic species with a worldwide distribution. Calonectria leaf blight is one of the most prominent diseases of Eucalyptus trees in Southeast Asian and South American plantations. Inoculation trials to evaluate pathogenicity of Calonectria spp. typically use conidial suspensions but this is not possible for species that do not sporulate sufficiently in culture. Calonectria pseudoreteaudii is one of the species that is most aggressive to Eucalyptus in China but most isolates fail to produce conidia in culture, requiring an alternative procedure for artificial inoculation. This study compared inoculations utilizing conidial and hyphal fragment suspensions. Two Eucalyptus genotypes were used, and these were inoculated with different concentrations of hyphal fragments or conidia of three C. pseudoreteaudii isolates. Three days after inoculation, the treated Eucalyptus plants displayed similar disease symptoms, regardless of whether they had been inoculated with conidia or hyphal fragments. This was consistent for all C. pseudoreteaudii isolates and also the different Eucalyptus genotypes. The results demonstrate that hyphal fragment suspensions can be used to provide a reliable indication of C. pseudoreteaudii isolate pathogenicity when conidia are not available for inoculation studies.

Global Genetic Diversity and Mating Type Distribution of Calonectria pauciramosa: An Important Wide-Host-Range Plant Pathogen.

The fungal pathogen, Calonectria pauciramosa, has caused serious diseases of many important plants worldwide. Understanding the genetic diversity and mating type distribution of this pathogen provides an essential step toward the development of disease control measures. In this study, we designed 15 polymorphic microsatellite markers by using genome sequences of two Ca. pauciramosa isolates having opposite mating type and from different countries. These markers were used to determine the genetic diversity of 145 isolates representing 13 different hosts (12 plant hosts residing in 12 genera, and soil) from 10 countries. In addition, mating type genes were amplified to investigate the reproduction mode of the pathogens in these populations by using mating type primers designed for Calonectria spp. Results revealed that a single dominant genotype, isolated from 11 plant genera residing in eight families, was present in seven countries across five continents. Only mating type MAT1-1 or MAT1-2 was amplified in each of the isolates, confirming that Ca. pauciramosa is heterothallic. Both mating types were detected in isolates from Eucalyptus in South Africa and Uruguay. The MAT1-2 phenotype was widely distributed in isolates from 12 different hosts (11 plant hosts and soil) collected in 10 countries. Overall, the results suggest that there has been substantial global movement of Ca. pauciramosa and that this has shaped its current population structure.

Mating strategy and mating type distribution in six global populations of the Eucalyptus foliar pathogen Teratosphaeria destructans.

Teratosphaeria destructans is an aggressive fungal pathogen causing leaf and shoot blight on young Eucalyptus trees in plantations. The disease occurs across tropical and subtropical regions of South East Asia and has recently been found in South Africa. Asexual structures of the pathogen are produced on infected tissues, but sexual structures have never been observed. The aim of this study was to investigate the reproductive biology of T. destructans by characterising its mating type (MAT1) locus and investigating its potential for sexual recombination. We found that T. destructans has a heterothallic mating system, with either the MAT1-1-1 and MAT1-1-10 genes (MAT1-1 idiomorph) or the MAT1-2-1 and MAT1-2-12 genes (MAT1-2 idiomorph) present in a single individual. With a multiplex PCR assay, it was possible to distinguish the two MAT idiomorphs in several Teratosphaeria species and this approach was applied to six global populations of T. destructans. Although both mating types occurred in the South East Asian populations, a single mating type dominated each population. Isolates from the recent disease outbreak in South Africa comprised only a single mating type. Attempts to induce a sexual cycle in vitro using strains of opposite mating type were not successful. The uneven distribution of mating types in populations of T. destructans and the presence of only an asexual state on infected tissues suggests the absence of or at least a minor role for sexual reproduction where the pathogen occurs on non-native Eucalyptus in plantations.

Calonectria pentaseptata Causes Severe Leaf Disease of Cultivated Eucalyptus on the Leizhou Peninsula of Southern China.

Eucalyptus (Myrtaceae, Myrtales) trees are widely cultivated for commercial purposes worldwide. Calonectria leaf blight is one of the most prominent diseases associated with Eucalyptus trees grown in plantations in Asia and South America. Recently, symptoms of leaf blight, shoot blight, tree death, and seedling rot caused by Calonectria species have been observed in commercial Eucalyptus plantations and nurseries in Leizhou Peninsula, which is one of the most densely Eucalyptus-planted areas in southern China. Disease samples were collected from 10 Eucalyptus species and a number of Eucalyptus grandis, E. tereticornis, and E. urophylla hybrid genotypes that were planted on plantations at 13 sites and one experimental nursery. A total of 773 isolates of Calonectria were obtained from 683 plantation trees and nursery seedlings. Fifty-five representative isolates from all the surveyed sites and Eucalyptus species/genotypes were selected for molecular identification. These 55 isolates were identified by DNA sequence analyses based on the calmodulin (cmdA), histone H3 (his3), translation elongation factor 1-alpha (tef1), and ß-tubulin (tub2) gene regions, as well as a combination of morphological characteristics. The results indicated that these 55 isolates present one single species, Calonectria pentaseptata. Determined by sequences of cmdA, his3, tef1, and tub2 gene regions, only two genotypes were identified among the 55 representative isolates; 54 of these isolates share the same genotype, suggesting that the genetic diversity of Ca. pentaseptata collected during this study was relatively low. A growth study indicated that Ca. pentaseptata is a high-temperature species. The mating test results suggested that Ca. pentaseptata is heterothallic or lacks the ability to recombine to produce fertile progeny. Inoculation results showed that Ca. pentaseptata causes leaf blight and stem rot, resulting in tree death of the two widely planted Eucalyptus genotypes in southern China, and that the two genotypes differ significantly in their susceptibility to infection by Ca. pentaseptata. A selection program to develop Eucalyptus planting stocks with high levels of resistance to Calonectria leaf blight in China during the long-term should be urgently initiated.[Formula: see text] Copyright © 2020 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Three genetic groups of the Eucalyptus stem canker pathogen Teratosphaeria zuluensis introduced into Africa from an unknown source.

The Eucalyptus stem canker pathogen Teratosphaeria zuluensis was discovered in South Africa in 1988 and it has subsequently been found in several other African countries as well as globally. In this study, the population structure, genetic diversity and evolutionary history of T. z uluensis were analysed using microsatellite markers to gain an enhanced understanding of its movement in Africa. Isolates were collected from several sites in Malawi, Mozambique, Uganda and Zambia. Data obtained were compared with those previously published for a South African population. The data obtained from 334 isolates, amplified across eight microsatellite loci, were used for assignment, differentiation and genetic diversity tests. STRUCTURE analyses, θ st and genetic distances revealed the existence of two clusters, one dominated by isolates from South Africa and the other by isolates from the Zambezi basin including Malawi, Mozambique and Zambia. High levels of admixture were found within and among populations, dominated by the Mulanje population in Malawi. Moderate to low genetic diversity of the populations supports the previously held view that the pathogen was introduced into Africa. The clonal nature of the Ugandan population suggests a very recent introduction, most likely from southern Africa.

New Ceratocystis species from Eucalyptus and Cunninghamia in South China.

During routine surveys for possible fungal pathogens in the rapidly expanding plantations of Eucalyptus and Cunninghamia lanceolata in China, numerous isolates of unknown species in the genus Ceratocystis (Microascales) were obtained from tree wounds. In this study we identified the Ceratocystis isolates from Eucalyptus and Cunninghamia in the GuangDong, GuangXi, FuJian and HaiNan Provinces of South China based on morphology and through comparisons of DNA sequence data for the ITS, partial ß-tubulin and TEF-1α gene regions. Morphological and DNA sequence comparisons revealed two previously unknown species residing in the Indo-Pacific Clade. These are described here as Ceratocystis cercfabiensis sp. nov. and Ceratocystis collisensis sp. nov. Isolates of Ceratocystis cercfabiensis showed intragenomic variation in their ITS sequences and four strains were selected for cloning of the ITS gene region. Twelve ITS haplotypes were obtained from 17 clones selected for sequencing, differing in up to seven base positions and representing two separate phylogenetic groups. This is the first evidence of multiple ITS types in isolates of Ceratocystis residing in the Indo-Pacific Clade. Caution should thus be exercised when using the ITS gene region as a barcoding marker for Ceratocystis species in this clade. This study also represents the first record of a species of Ceratocystis from Cunninghamia.

Novel species of Botryosphaeriaceae associated with shoot blight of pistachio.

Various species of phytopathogenic Botryosphaeriaceae were identified previously from pistachio trees worldwide. Disease symptoms caused by pathogens in Botryosphaeriaceae on pistachio include panicle and shoot blight, leaf defoliation, fruit discoloration and decay. In this study species of Botryosphaeriaceae were collected from blighted pistachio shoots in Arizona, USA, and Greece. The aims of this study were to identify these Botryosphaeriaceae isolates and to test their pathogenicity to pistachio. The fungi were identified based on comparisons of DNA sequence data of the nuclear rDNA internal transcribed spacer region (ITS), a partial translation elongation factor 1-alpha gene (TEF1), a partial ß-tubulin gene (TUB2) and morphological characteristics. Results indicated that some isolates collected from pistachio represent two previously undescribed species, which we described here as Lasiodiplodia americana sp. nov. from the United States and Neofusicoccum hellenicum sp. nov. from Greece. Field inoculations of L. americana and N. hellenicum on branches of four pistachio cultivars showed that both L. americana and N. hellenicum are pathogenic on pistachio. The four pistachio cultivars differed in their susceptibility to the Botryosphaeriaceae species. Results of this study suggested that the two new species of Botryosphaeriaceae need to be monitored carefully to determine the distribution of these pathogens and the possible spread to other areas.

Phylogeny, Morphology, Distribution, and Pathogenicity of Botryosphaeriaceae and Diaporthaceae from English Walnut in California.

Species of family Botryosphaeriaceae and genus Diaporthe (anamorph: genus Phomopsis, family Diaporthaceae) were reported and caused diseases on various fruit and nut trees in California. In the last several years, diseases on English walnut (Juglans regia) caused by species of Botryosphaeriaceae and Diaporthe were observed frequently in California. Disease symptoms include stem canker; shoot canker and blight; twig, leaf, and fruit blight; and necrotic leaf lesions. Isolates of the pathogen were collected from English walnut in 13 counties in California. The aims of this study were to identify these isolates and to test their pathogenicity to English walnut cultivars. In total, 159 California isolates were identified based on comparisons of DNA sequence data of the internal transcribed spacer, translation elongation factor 1-α, and ß-tubulin gene regions, and combined with the morphological features of the cultures and conidia. Research results revealed that isolates represent 10 species of Botryosphaeriaceae and two species of Diaporthe. These species include Botryosphaeria dothidea, Diplodia mutila, D. seriata, Dothiorella iberica, Lasiodiplodia citricola, Neofusicoccum mediterraneum, N. nonquaesitum, N. parvum, N. vitifusiforme, Neoscytalidium dimidiatum, Diaporthe neotheicola, and D. rhusicola. Pathogenicity on three English walnut cultivars ('Chandler', 'Tulare', and 'Vina') using a mycelium plug inoculation method revealed that all these species are pathogenic to all the tested cultivars, with L. citricola and N. parvum being the most pathogenic species, followed by N. mediterraneum, N. dimidiatum, and B. dothidea. Chandler was more tolerant to infection than Tulare and Vina. Results in this study determined that multiple numbers of the Botryosphaeriaceae fungi and two Diaporthe spp. cause cankers and blights of English walnut and vary in their virulence from highly to slightly virulent, respectively.

Wide Distribution of Teratosphaeria epicoccoides and T. destructans Associated with Diseased Eucalyptus Leaves in Plantations in Southern China.

Species of Mycosphaerellaceae and Teratosphaeriaceae represent over 40% of the fungi identified on eucalypt leaves worldwide. These include some important pathogens that mainly cause leaf blight and spot, and result in increasingly negative impacts on global commercial eucalypt industries. Eucalyptus plantations are commonly cultivated in southern China for solid wood and pulp products. However, the species diversity and geographic distribution of Mycosphaerellaceae and Teratosphaeriaceae, associated with diseased plantation Eucalyptus leaves in China, have not been clarified. In this study, we conducted the first systematic surveys and sample collections of Mycosphaerellaceae- and Teratosphaeriaceae-like fungi from diseased plantation Eucalyptus leaves in southern China. In total, 558 isolates were obtained from 59 sampled sites in five provinces. One isolate was isolated from each tree. According to the disease symptoms, conidia morphological characteristics, and DNA sequence comparisons of ITS, tef1 and tub2 gene regions. The 558 isolates were identified as Teratosphaeria epicoccoides (312 isolates; 55.9%) and T. destructans (246 isolates, 44.1%). Both species were widely distributed in the sampled regions in southern China. The genotypes of T. epicoccoides and T. destructans were determined based on ITS, tef1, and tub2 sequences. The results showed that multiple genotypes of each species of T. epicoccoides and T. destructans exist in China. Additionally, isolates with multiple genotypes were obtained in all five sampled provinces. These results suggest that both T. epicoccoides and T. destructans are not clonal. This study proved that both T. epicoccoides and T. destructans are dominant species and widely distributed on diseased Eucalyptus leaves in southern China. The wide geographic distribution and potential high genetic diversity pose challenges for the disease management of Teratosphaeria leaf blight and leaf spot in China.

Fungal Planet description sheets: 154-213.

Novel species of microfungi described in the present study include the following from South Africa: Camarosporium aloes, Phaeococcomyces aloes and Phoma aloes from Aloe, C. psoraleae, Diaporthe psoraleae and D. psoraleae-pinnatae from Psoralea, Colletotrichum euphorbiae from Euphorbia, Coniothyrium prosopidis and Peyronellaea prosopidis from Prosopis, Diaporthe cassines from Cassine, D. diospyricola from Diospyros, Diaporthe maytenicola from Maytenus, Harknessia proteae from Protea, Neofusicoccum ursorum and N. cryptoaustrale from Eucalyptus, Ochrocladosporium adansoniae from Adansonia, Pilidium pseudoconcavum from Greyia radlkoferi, Stagonospora pseudopaludosa from Phragmites and Toxicocladosporium ficiniae from Ficinia. Several species were also described from Thailand, namely: Chaetopsina pini and C. pinicola from Pinus spp., Myrmecridium thailandicum from reed litter, Passalora pseudotithoniae from Tithonia, Pallidocercospora ventilago from Ventilago, Pyricularia bothriochloae from Bothriochloa and Sphaerulina rhododendricola from Rhododendron. Novelties from Spain include Cladophialophora multiseptata, Knufia tsunedae and Pleuroascus rectipilus from soil and Cyphellophora catalaunica from river sediments. Species from the USA include Bipolaris drechsleri from Microstegium, Calonectria blephiliae from Blephilia, Kellermania macrospora (epitype) and K. pseudoyuccigena from Yucca. Three new species are described from Mexico, namely Neophaeosphaeria agaves and K. agaves from Agave and Phytophthora ipomoeae from Ipomoea. Other African species include Calonectria mossambicensis from Eucalyptus (Mozambique), Harzia cameroonensis from an unknown creeper (Cameroon), Mastigosporella anisophylleae from Anisophyllea (Zambia) and Teratosphaeria terminaliae from Terminalia (Zimbabwe). Species from Europe include Auxarthron longisporum from forest soil (Portugal), Discosia pseudoartocreas from Tilia (Austria), Paraconiothyrium polonense and P. lycopodinum from Lycopodium (Poland) and Stachybotrys oleronensis from Iris (France). Two species of Chrysosporium are described from Antarctica, namely C. magnasporum and C. oceanitesii. Finally, Licea xanthospora is described from Australia, Hypochnicium huinayensis from Chile and Custingophora blanchettei from Uruguay. Novel genera of Ascomycetes include Neomycosphaerella from Pseudopentameris macrantha (South Africa), and Paramycosphaerella from Brachystegia sp. (Zimbabwe). Novel hyphomycete genera include Pseudocatenomycopsis from Rothmannia (Zambia), Neopseudocercospora from Terminalia (Zambia) and Neodeightoniella from Phragmites (South Africa), while Dimorphiopsis from Brachystegia (Zambia) represents a novel coelomycetous genus. Furthermore, Alanphillipsia is introduced as a new genus in the Botryosphaeriaceae with four species, A. aloes, A. aloeigena and A. aloetica from Aloe spp. and A. euphorbiae from Euphorbia sp. (South Africa). A new combination is also proposed for Brachysporium torulosum (Deightoniella black tip of banana) as Corynespora torulosa. Morphological and culture characteristics along with ITS DNA barcodes are provided for all taxa.

Diversity and Distribution of Calonectria Species in Soils from Eucalyptus urophylla × E. grandis, Pinus massoniana, and Cunninghamia lanceolata Plantations in Four Provinces in Southern China.

The species of Calonectria include many notorious plant pathogens and are widely distributed around the world. Leaf blight caused by Calonectria species is considered one of the most prominent diseases in Eucalyptus plantations in China. Some Calonectria species isolated from soils in Eucalyptus plantations are highly pathogenic to inoculated Eucalyptus genotypes. In southern China, the plantation trees Cunninghamia lanceolata, Eucalyptus spp., and Pinus massoniana are always adjacently planted, especially in FuJian, GuangDong, GuangXi, and YunNan Provinces. The aim of this study was to understand the diversity and distribution of Calonectria in soils from plantations of different tree species in different geographic regions. Soil samples were collected from 12 sampling sites in Eucalyptus urophylla × E. grandis, P. massoniana, and C. lanceolata plantations in FuJian, GuangDong, GuangXi, and YunNan Provinces. Approximately 250 soil samples were collected from each sampling site, and a total of 2991 soil samples were obtained. A total of 1270 Calonectria isolates were obtained from 1270 soil samples. The 1270 isolates were identified based on DNA sequence comparisons of the partial gene regions of act, cmdA, his3, rpb2, tef1, and tub2. These isolates were identified as 11 Calonectria species: Calonectria aconidialis (69.50%), C. kyotensis (13.10%), C. hongkongensis (10.80%), C. ilicicola (2.50%), C. asiatica (2.36%), C. curvispora (0.31%), C. chinensis (0.24%), C. pacifica (0.24%), C. yunnanensis (0.16%), and C. canadiana (0.08%) in the C. kyotensis species complex and C. eucalypti (0.71%) in the C. colhounii species complex. The three dominant species, C. aconidialis, C. kyotensis, and C. hongkongensis, were widely distributed. The richness of Calonectria (percentage of soil samples that yielded Calonectria) in soils in the eastern regions (relatively humid regions) was higher than that in the western regions. The Calonectria richness of E. urophylla × E. grandis, P. massoniana, and C. lanceolata plantations decreased gradually. For each of the three dominant species, its richness in the eastern regions was generally higher than that in the western regions; the species richness was highest in E. urophylla × E. grandis plantations for C. aconidialis, while for each of C. kyotensis and C. hongkongensis, its species richness was highest in P. massoniana plantations. The genetic variation in C. aconidialis, C. kyotensis, and C. hongkongensis was more greatly affected by geographic region than by plantation tree species. This study expanded our understanding of the richness, species diversity, and distribution characteristics of Calonectria in soils from the plantations of different tree species in different geographic regions in southern China. Results in this study enhanced our understanding of the influencing characteristics of geographic region and tree species on the species and genetic diversity of soilborne fungi.

Wide Distribution and Intraspecies Diversity in the Pathogenicity of Calonectria in Soil from Eucalyptus Plantations in Southern Guangxi of China.

Eucalyptus spp. are extensively cultivated in southern China because of their adaptability and versatile timber production. Calonectria leaf blight caused by Calonectria species is considered a major threat to Eucalyptus trees planted in China. The GuangXi Zhuang Autonomous Region is the provincial region with the largest distribution of Eucalyptus plantations in China. The present study aimed to expound the species diversity and pathogenicity of Calonectria isolates obtained from the soil of Eucalyptus plantations in GuangXi. A total of 188 Calonectria isolates were recovered from the soil located close to Eucalyptus trees, and the isolates were identified based on the DNA sequence comparisons of the four partial regions of the translation elongation factor 1-alpha (tef1), ß-tubulin (tub2), calmodulin (cmdA), and histone H3 (his3) genes. The isolates were identified as Calonectria aconidialis (74.5%), C. hongkongensis (21.3%), C. pseudoreteaudii (2.1%), C. kyotensis (1.6%), and C. chinensis (0.5%). The inoculation results indicated that 40 isolates representing five Calonectria species were pathogenic to the three Eucalyptus genotypes. Two inoculated experiments consistently showed that the longest lesions were produced by the isolates of C. aconidialis. Some isolates of C. aconidialis, C. hongkongensis, and C. kyotensis produced significantly longer lesions than the positive controls, but not the isolates of C. pseudoreteaudii or C. chinensis. These results indicated that Calonectria isolated from the soil may pose a threat to Eucalyptus plantations. Some Calonectria isolates of the same species differed significantly in their virulence in the tested Eucalyptus genotypes. The resistance of different Eucalyptus genotypes to Calonectria isolates within the same species was inconsistent. The inoculation results in this study suggested that many Calonectria isolates in each species had different levels of pathogenicity, and many Eucalyptus genotypes need to be tested to select disease-resistant Eucalyptus genetic materials in the future. The results of the present study enhance our knowledge of species diversity and the potential damage caused by Calonectria in the soil of Eucalyptus plantations. Our results also provide new insights into the breeding of disease-resistant Eucalyptus genotypes for controlling Calonectria leaf blight in China in the future.

Identification and pathogenicity of Aurifilum species (Cryphonectriaceae, Diaporthales) on Terminalia species in Southern China.

The family of Cryphonectriaceae (Diaporthales) contains many important tree pathogens and the hosts are wide-ranging. Tree species of Terminalia were widely planted as ornamental trees alongside city roads and villages in southern China. Recently, stem canker and cracked bark were observed on 2-6 year old Terminalianeotaliala and T.mantaly in several nurseries in Zhanjiang City, Guangdong Province, China. Typical conidiomata of Cryphonectriaceae fungi were observed on the surface of the diseased tissue. In this study, we used DNA sequence data (ITS, BT2/BT1, TEF-1α, rpb2) and morphological characteristics to identify the strains from Terminalia trees. Our results showed that isolates obtained in this study represent two species of Aurifilum, one previously described species, A.terminali, and an unknown species, which we described as A.cerciana sp. nov. Pathogenicity tests demonstrated that both A.terminali and A.cerciana were able to infect T.neotaliala and two tested Eucalyptus clones, suggesting the potential for Aurifilum fungi to become new pathogens of Eucalyptus.

Evidence of High Genetic Diversity and Differences in the Population Diversity of the Eucalyptus Leaf Blight Pathogen Calonectria pseudoreteaudii from Diseased Leaves and Soil in a Plantation in Guangxi, China.

Calonectria pseudoreteaudii is an important causal agent of Eucalyptus leaf blight in southern China. This pathogen causes Eucalyptus tree disease across numerous regions in southern China. In addition to diseased leaves, C. pseudoreteaudii has occasionally been isolated from soil in Eucalyptus plantations. The aim of this study was to clarify whether C. pseudoreteaudii causing Eucalyptus leaf blight in China is mainly clonally reproduced and to determine the potential spreading mechanism of C. pseudoreteaudii between diseased leaves and soil. To this end, 10 polymorphic microsatellite markers were analyzed to detect the genetic diversity of 97 C. pseudoreteaudii isolates from diseased leaves and soil in a Eucalyptus plantation in Guangxi Zhuang Autonomous Region, southern China. The analysis showed that the genetic diversity of the isolates from both the diseased leaves and soil was high. However, the gene and genotype diversity of the C. pseudoreteaudii isolates from diseased leaves were higher than those of the isolates from the soil. Moreover, all genotypes detected in the isolates from the soil were also found in the isolates from the diseased leaves. Structural analyses did not show clear population structures related to the population substrates of the diseased leaves or soil, and molecular variance analyses indicated that no significant genetic differentiation existed between the diseased leaf and soil populations. These results suggest that C. pseudoreteaudii in soil spreads from diseased leaves, and that an asexual cycle is the primary reproductive mode in both diseased leaf and soil populations. This is the first study on the genetic diversity and population structure of C. pseudoreteaudii. The high genetic diversity and spread pathways of this pathogen may pose challenges in controlling the disease. C. pseudoreteaudii from both diseased leaves and soils in Eucalyptus plantations needs to be carefully monitored for disease control and management.

Characterization of Pseudofusicoccum Species from Diseased Plantation-Grown Acacia mangium, Eucalyptus spp., and Pinus massoniana in Southern China.

Fungi from Pseudofusicoccum (Phyllostictaceae, Botryosphaeriales) have been reported as pathogens, endophytes, or saprophytes from various woody plants in different countries. Recently, Botryosphaeriales isolates were obtained from the dead twigs of Acacia mangium, Eucalyptus spp., Pinus massoniana, and Cunninghamia lanceolata in Guangdong, Guangxi, Hainan, and Fujian Provinces in southern China. This study aimed to understand the diversity, distribution, and virulence of these Pseudofusicoccum species on these trees. A total of 126 Pseudofusicoccum isolates were obtained, and the incidences of Pseudofusicoccum (percentage of trees that yielded Pseudofusicoccum) on A. mangium, P. massoniana, Eucalyptus spp., and C. lanceolata were 21%, 2.6%, 0.5%, and 0%, respectively. Based on the internal transcribed spacer (ITS), translation elongation factor 1-alpha (tef1), and ß-tubulin (tub2) loci, 75% of the total isolates were identified as P. kimberleyense, and the remaining isolates were identified as P. violaceum. For P. kimberleyense, the majority of isolates (83%) were from A. mangium, and the rest were from P. massoniana (14%) and Eucalyptus spp. (3%). Similarly, the proportion of isolates of P. violaceum from A. mangium, P. massoniana, and Eucalyptus spp. were 84%, 13%, and 3%, respectively. Inoculation trials showed that the two species produced expected lesions on the tested seedlings of A. mangium, E. urophylla × E. grandis, and P. elliottii. This study provides fundamental information on Pseudofusicoccum associated with diseases in main plantations in southern China.

Calonectria in the age of genes and genomes: Towards understanding an important but relatively unknown group of pathogens.

The genus Calonectria includes many aggressive plant pathogens causing diseases on various agricultural crops as well as forestry and ornamental tree species. Some species have been accidentally introduced into new environments via international trade of putatively asymptomatic plant germplasm or contaminated soil, resulting in significant economic losses. This review provides an overview of the taxonomy, population biology, and pathology of Calonectria species, specifically emerging from contemporary studies that have relied on DNA-based technologies. The growing importance of genomics in future research is highlighted. A life cycle is proposed for Calonectria species, aimed at improving our ability to manage diseases caused by these pathogens.

Diversity and Distribution of Calonectria Species from Plantation and Forest Soils in Fujian Province, China.

To meet the growing demand for wood and pulp products, Eucalyptus plantations have expanded rapidly during the past two decades, becoming an integral part of the southern China landscape. Leaf blight caused by various Calonectria spp., is a serious threat to these plantations. In order to explore the diversity and distribution of Calonectria spp. in Fujian Province soils, samples were collected in Eucalyptus plantations and adjacent plantings of Cunninghamia lanceolata, Phyllostachys heterocycle and Pinus massoniana as well as in natural forests. Three hundred and fifty-three Calonectria isolates were recovered from soil samples and they were identified based on a comparison of multilocus DNA sequence data for the act (actin), cmdA (calmodulin), his3 (histone H3), rpb2 (the second largest subunit of RNA polymerase), tef1 (translation elongation factor 1-alpha) and tub2 (ß-tubulin) gene regions, as well as morphological characteristics. Six known taxa including Calonectria aconidialis, Ca. hongkongensis, Ca. ilicicola, Ca. kyotensis, Ca. pacifica, Ca. pseudoreteaudii and one novel species described here as Ca. minensis sp. nov. were identified. Of these, Ca. aconidialis and Ca. kyotensis were the most prevalent species, and found in eight and seven sites, and four and five forest types, respectively. Calonectria spp. were most abundant in soils from Eucalyptus stands, followed by P. heterocycle and natural forests. Relatively few species were found in the soils associated with Cunninghamia lanceolata and Pinus massoniana. The abundance of known Calonectria spp. suggests that these fungi have been relatively well sampled in Fujian. The results are also consistent with the fact that most Calonectria diseases are found on Angiosperm as opposed to Gymnosperm plants.

Soil-borne Calonectria (Hypocreales, Nectriaceae) associated with Eucalyptus plantations in Colombia.

Eucalyptus spp. are widely planted in Colombia as an important component of a growing paper and pulp industry. Leaf and shoot blight caused by Calonectria spp. was one of the first disease problems to emerge in these plantations. A survey of Eucalyptus plantations in four forestry regions of Colombia during 2016 resulted in a large number of Calonectria isolates from soil samples collected in the understories of trees having symptoms of Calonectria leaf and shoot blight. The aim of this study was to identify and resolve the phylogenetic relationships for these isolates using DNA sequence comparisons of six gene regions as well as morphological characters. From a collection of 107 isolates, seven Calonectria species residing in three species complexes were identified. Two of these represented undescribed species, namely C.exiguispora sp. nov. and C.guahibo sp. nov. Calonectriaparvispora and C.spathulata were the most commonly isolated species, each of which accounted for approximately 30% of the isolates. The results suggest that Colombia has a wide diversity of Calonectria spp. and that these could challenge Eucalyptus plantation forestry in the future.