Phylogenetic investigations in the downy mildew genus <Emphasis Type="Italic">Bremia</Emphasis> reveal several distinct lineages and a species with a presumably exceptional wide host range

Bremia lactucae is one of the most devastating and widespread pathogens in lettuce production worldwide. Despite its economical importance, uncertainty prevails about the species delimitation in the genus Bremia. Commonly, Bremia is considered to be monotypic, containing only Bremia lactucae, while taxonomists have described additional species, and molecular phylogenetic studies have shown significant sequence divergence between accessions from different hosts. Here, we report that several previously described species are genetically highly distinct from Bremia lactucae parasitic to Lactuca sativa. These include Bremia lapsanae, Bremia sonchicola, and Bremia taraxaci. In addition to these host-specific species, a plurivorous species is revealed, which infects hosts from three different tribes in the Asteraceae subfamilies Asteroideae and Carduoideae. The broad host range of clade 1 is exceptional for downy mildews and only paralleled by Pseudoperonospora cubensis, which infects a broad range of Cucurbitaceae. The taxonomic status of Bremia cirsii and of Bremia centaureae remains unresolved, as the accessions from Cirsium and Centaurea, respectively, did not form a monophylum but were partly contained in the plurivorous clade 1. Bremia lactucae was found to be restricted to Lactuca sativa and Lactuca serriola. Thus, it can be assumed that Bremia infections on weeds apart from Lactuca species do not pose a significant risk for lettuce production. However, it is unlikely that breeding resistance genes from Lactuca serriola into Lactuca sativa will result in durable resistance of lettuce to downy mildew disease, because the current study provides additional evidence that Bremia accessions from both hosts form a population continuum.     

Phylogenetic investigations in the genus <Emphasis Type="Italic">Pseudoperonospora</Emphasis> reveal overlooked species and cryptic diversity in the <Emphasis Type="Italic">P. cubensis</Emphasis> species cluster

Pseudoperonospora cubensis is one of the most devastating diseases of cucurbitaceous crops. The pathogen has a worldwide distribution and occurs in all major cucurbit growing areas. It had been noticed for the first time at the end of the 19th century, but it became a globally severe disease as recently as 1984 in Europe and 2004 in North America. Despite its economic importance, species concepts in Pseudoperonospora are debated. Here, we report that the genus Pseudoperonospora contains cryptic species distinct from the currently accepted ones. Pseudoperonospora on Celtis is split into two phylogenetic lineages and Pseudoperonospora humuli is confirmed as a species distinct from the Cucurbitaceae-infecting lineages. A cryptic species occupying a basal position within the Pseudoperonospora cubensis complex is revealed to be present on Humulus japonicus, thus providing evidence that the host jump that gave rise to Pseudoperonospora cubensis likely occurred from hops. Notably, Cucurbitaceae infecting pathogens are present in two cryptic sister species or subspecies. Clade 1 contains primarily specimens from North America and likely represents Pseudoperonospora cubensis s.str.. Pre-epidemic isolates in clade 2 originate from Japan and Korea, suggesting this cryptic species or subspecies is indigenous to East Asia. Recent samples of this lineage from epidemics in Europe and the United States cluster together with clade 2. It thus seems possible that this lineage is associated with the recent severe epidemics of cucurbit downy mildew and is now naturalised in North America and Europe.     

What causes flag smut of wheat?

The causal agent of flag smut of wheat is currently subject to strict quarantine regulations in many countries and is believed to have a wide host range on wild and cultivated grasses. This fungus has been classified as both Urocystis agropyri and Urocystis tritici. Urocystis agropyri was first described from Elymus repens in Germany and U. tritici was first described from Triticum vulgare (=T. aestivum). In 1953, G. W. Fischer placed U. tritici and a large number of other Urocystis species in synonymy with U. agropyri. The present study is the first attempt to clarify the taxonomy and phylogeny of flag smut pathogens of grasses using molecular analyses. Three loci, the internal transcribed spacer (ITS) region of rDNA, the RNA polymerase II subunit 2 (RPB2), and translation elongation factor (TEF) protein‐coding regions were used for phylogenetic reconstruction to determine the species boundaries of 24 Urocystis specimens from triticoid hosts. Results indicate that there are several distinct lineages of flag smut pathogens, including the causal agent of flag smut of wheat, which is supported as a separate species, U. tritici. Sequences from specimens on E. repens, which are retained as U. agropyri, grouped in a clade distinct from those on wheat and rye. The closest relatives of U. tritici were found to be U. hispanica from Aegilops and Urocystis sp. from Thinopyrum junceiforme and Elymus trachycaulis. Recognition that U. tritici is genetically distinct from U. agropyri sensu stricto will impact regulatory policy and facilitate the development of diagnostic tests.     

Host matrix has major impact on the morphology of <Emphasis Type="Italic">Pseudoperonospora cubensis</Emphasis>

Oomycetes contain some of the economically most important pathogens of flowering plants. Most have a rather narrow host range, often being restricted to single host species. In downy mildews and other obligate biotrophic plant parasites, like powdery mildews and rusts, delimitating species on grounds of morphological characteristics is often hardly possible and thus often based on only subtle differences. This has led to the widespread application of a broad species concept for these organisms. Consequently, despite the fact that morphological differences were reported for Pseudoperonospora cubensis from different host species, the corresponding new pathogen species were not accepted as being independent, and the host range of Pseudoperonospora cubensis is reported to encompass more than 50 host species in the Cucurbitaceae in temperate to tropical climates. However, recent studies have reported narrow host ranges for other downy mildew genera and advocated a narrow species concept. Here, we report successful colonisation of five different tribes of the Cucurbitaceae by a strain of Pseudoperonospora cubensis and demonstrate that the host matrix has a major impact on the morphology of the pathogen. On the basis of five morphological criteria significant differences could be found for all hosts. These differences were more pronounced in phylogenetically unrelated than in related hosts. Our results provide evidence for a broad host range of Pseudoperonospora cubensis and demonstrate that species delimitation based on morphological characters is not feasible in Pseudoperonospora on Cucurbitaceae. Also in other biotrophic plant pathogens, the situation could be similar, thus necessitating thorough morphological, molecular phylogenetic and cross inoculation experiments for species recognition.     

Modern approaches to species concepts in downy mildews

Inadequate species definitions present a serious problem to the pathologist working in plant hygiene or quarantine which demands urgent attention. Species concepts in the downy mildews have not kept pace with developments in evolutionary and molecular biology, or with advances in ecological genetics, because the downy mildews are obligate biotrophs that are not easily cultured in the laboratory. Existing approaches to species concepts in this group (morphometric, Ga¨umann's 'biological species' and Skalicý's 'eco–physio–phentic' concepts) are examined and found to be inadequate and potentially misleading. The systematic treatment of the downy mildews is beginning to benefit from the application of modern methods of systematic analysis. The contribution and potential of ultrastructure, karyotyping, sterol and fatty acid composition, isoenzyme patterns, molecular biology, numerical methods, immunoassay and hypotheses of coevolution to the development of species concepts are reviewed and their wider application is seen as a priority. The application to the downy mildews of two widespread species concepts, the biological and phylogenetic concepts, is examined in the light of the information gained from modern methods of analysis, but neither is found adequate to describe species of downy mildews as they occur in nature. Modern methods can suggest phylogenetic relationships on the basis of statistical probabilities and may also detect microevolutionary change, but it is concluded that much more information is required about individual breeding systems, gene flow, ecology, phylogeny and distribution before informed decisions about the delimitation of most species can be made. Until patterns of genetic diversity can be established, a modified phylogenetic species concept may offer one interim solution to the problem of species definition in the downy mildews.     

Nematode pests of some major cereals in Nigeria: Need for integration of morphological/morphometrical,biochemical, and molecular diagnostic approaches for accurate identification

Plant-parasitic nematodes pose a major threat to crop production in Nigeria. Comprehensive data on yield losses for most cereal crops are not readily available. Research on maize (Zea mays), rice (Oryza sativa), and sorghum (Sorghum bicolor) has shown an 18%–67% reduction in yield linked to the destructiveness of parasitic nematodes, particularly root-knot and root-lesion nematodes. Continuous monoculture and a conservative approach to agriculture increase the incidence and population density of nematodes, leading to higher levels of damage and losses in most cultivated cereal crops. These losses affect the food security of the growing population of Nigeria and many African countries that are heavily dependent on cereals. The accurate identification of nematodes is fundamental for effective management strategies. Nematode taxonomy based on morphology has been difficult due to intraspecific character variations coupled with the limited expertise of indigenous nematologists. Biochemical and molecular marker-based tools and approaches have allowed efficient diagnosis of most nematode species affecting cereal production globally. These approaches have proved useful due to their practicality, rapidness, accuracy, and cost-effectiveness. This review describes the important economic effects of these parasitic nematodes on cereal production and highlights the need for integrative approaches to taxonomy for accurate species identification that will improve nematode diagnosis, thereby contributing to the increase of food production in Nigeria.     

Detection of botryosphaeriaceous species in environmental samples using a multi‐species primer pair

Botryosphaeriaceous species are significant grapevine trunk pathogens worldwide, which can be difficult to identify to species level using conventional morphological methods. This study developed and optimized a quick, reliable molecular identification method that could facilitate investigations into the epidemiology of these diseases in vineyards. The multi‐species primers, BOT100F and BOT472R, amplified a 371–372 bp portion of the rRNA gene region from the six botryosphaeriaceous species commonly found in New Zealand vineyards. In silico analysis indicated that they would amplify DNA from six of the 12 lineages of the Botryosphaeriaceae, including all of the main species pathogenic to grapevines. A detection sensitivity of 1 and 0·1 pg DNA in standard and nested PCR, respectively, was achieved and this was calculated as equivalent to 2·5 conidia. Validation of the primers for environmental samples showed that their specificity was not compromized by the presence of competing DNA templates extracted from wood and soil. Single stranded conformational polymorphism (SSCP) analysis of the amplicons could resolve Neofusicoccum australe, N. luteum, Diplodia mutila and D. seriata, but did not differentiate between N. parvum and N. ribis. The optimized PCR‐SSCP was used to identify botryosphaeriaceous species present in rainwater traps collected over 1 year in a vineyard known to contain infected vines. It could detect multiple species in individual samples and demonstrated differences in the dispersal patterns of conidia from different species. Given the specificity and sensitivity of this method it could prove useful in epidemiology studies involving the numerous botryosphaeriaceous species that infect a wide range of host species.     

Host range expansion in a powdery mildew fungus (<Emphasis Type="Italic">Golovinomyces</Emphasis> sp.) infecting <Emphasis Type="Italic">Arabidopsis thaliana</Emphasis>: <Emphasis Type="Italic">Torenia fournieri</Emphasis> as a new host

Since 2003, Torenia fournieri plants grown for experimental purposes were repeatedly infected by powdery mildew in a laboratory in Hungary. Based on morphological characteristics, the pathogen belonged to the mitosporic genus Oidium subgen. Reticuloidium, the anamorph stage of Golovinomyces. The rDNA ITS sequence was identical to that of two other powdery mildew fungi, infecting Arabidopsis and Veronica, respectively, in different parts of the world. According to a previous phylogenetic analysis of ITS and 28S rDNA sequences, those two powdery mildews belong to a recently evolved group of Golovinomyces characterized by multiple host range expansions during their evolution. Both the ITS sequence and the morphological data indicate that the powdery mildew anamorph infecting Torenia also belongs to this group. It is likely that the powdery mildew infections of the experimental T. fournieri plants, native to south-east Asia, were the result of a very recent host range expansion of a polyphagous Golovinomyces because (i) T. fournieri is absent from our region, except as an experimental plant grown in the laboratory, (ii) the powdery mildew fungus infecting this exotic plant belongs to a group of Golovinomyces where host range expansion is a frequent evolutionary scenario, (iii) cross-inoculation tests showed that this pathogen is also able to infect other plant species, notably A. thaliana and tobacco, and (iv) no Golovinomyces species are known to infect T. fournieri anywhere in the world. Although host range expansion has often been proposed as a common evolutionary process in the Erysiphales, and also in other biotrophic plant pathogens, this has not been clearly demonstrated in any case studies so far. To our knowledge, this is the first convincing case of a host range expansion event in the Erysiphales.     

Confirmation of <Emphasis Type="Italic">Peronospora agrimoniae</Emphasis> as a distinct species

Leaves with typical symptoms of downy mildew were found on common agrimony in the Czech Republic in 2014 and 2015 and at several locations in Germany from 2010 to 2014. The causal agent of downy mildew of agrimony was often reported as Peronospora agrimoniae, but sometimes also as P. sparsa. Morphological characteristics of the pathogens found in both countries are in the range of previous works for P. agrimoniae, but also other downy mildews parasitic on Rosaceae, rendering their discrimination based on published observations difficult. For molecular identification sequencing of several loci (nrITS rDNA, cox1 and cox2) was performed. Phylogenetic analyses based on nrITS rDNA clearly separated P. agrimoniae from other Peronospora species infecting Rosaceae. Thus, considering P. agrimoniae as separate species seems justified. Two German specimens were identical to two Czech samples in both nrITS rDNA and cox1 mtDNA sequences, but differed in a single nucleotide substitution in cox2 region. To our knowledge, this is the first verified record of P. agrimoniae on common agrimony in the Czech Republic.     

Unraveling evolutionary relationships among the divergent lineages of colletotrichum causing anthracnose disease in turfgrass and corn

ABSTRACT Colletotrichum species cause anthracnose diseases on a number of grass hosts and are common inhabitants of many others. They are divided into four species: C. sublineolum is pathogenic to Sorghum spp.; C. caudatum is found on C4 grasses such as indiangrass and big bluestem; C. falcatum causes red rot of sugarcane; and C. graminicola sensu lato is a broadly defined species including isolates that attack maize, wheat, oats, and many forage, turf, and amenity grasses of the subfamily Pooideae. In this paper, a combination of hierarchal- and nonhierarchal-based analyses were employed to examine evolutionary relationships among the grass-infecting Colletotrichum species, with special emphasis on isolates from turf and other grasses in the subfamily Pooideae. Reconstructions performed with data sets from over 100 Colletotrichum isolates at three variable loci using phylogenetic and network-based methodologies unambiguously supported the taxonomic separation of maize-infecting isolates of C. graminicola from the pooid-infecting strains of Colletotrichum. To reflect the evolutionary relationships that exist between these distinct lineages, we propose the resurrection of the species name C. cereale to describe the pooid-infecting isolates. There was also support for further subdivision of C. cereale, but the current data are insufficient to confidently subdivide the species, as there was some evidence of recombination between lineages of this species.     

<Emphasis Type="Italic">RB</Emphasis> and <Emphasis Type="Italic">Ph</Emphasis> resistance genes in potato and tomato minimize risk for oospore production in the presence of mating pairs of <Emphasis Type="Italic">Phytophthora infestans</Emphasis>

Composed mostly of fungivorous species, the genus Aphelenchoides also comprises 14 plant-parasitic species. The most common and devastating, A. besseyi, A. fragariae, A. ritzemabosi and A. subtenuis have been reported on more than 900 plant species. The combination of low inter-specific and high intra-specific morphological variability makes morphology-based identification extremely difficult within this genus, and has led to molecular tools being employed to ensure accurate diagnoses. rDNA markers are widely used for the identification of nematodes while the Cytochrome Oxidase I gene (COI) remains relatively unexplored despite its role as the standard barcode for almost all animal groups. To explore its suitability as a diagnostic tool, we studied a fragment of the mtCOI region of the four main plant-parasitic Aphelenchoides within a phylogenetic framework. We generated 69 mtCOI and 123 rDNA sequences of diverse Aphelenchoides taxa; 67 belong to the main plant-parasitic species including the first mtCOI sequence of A. fragariae and the first mtCOI and 28S sequences of A. subtenuis. mtCOI had a similar success rate for PCR amplification. Phylogenetic trees based on the three studied markers are largely in agreement with one another, validating their use for Aphelenchoides diagnosis; additionally, we were able to locate several misidentified sequences of plant-parasitic Aphelenchoides in existing databases. The concatenated analysis from the three markers resulted in a more robust insight into the phylogeny and evolution of Aphelenchoides, revealing that plant-parasitism has evolved independently at least three times within this genus, presumably from fungal-feeding ancestors.     

Chloroplast DNA markers for Echinochloa taxa

The genus Echinochloa (Poaceae) includes numerous problematic weeds over a wide range of ecoregions in the world. To date, molecular markers for species identification and assessing phylogenetic relationship are still limited in the genus Echinochloa. In this study, we developed seven chloroplast molecular markers based on divergent chloroplast regions of E. crus‐galli and E. oryzicola. Furthermore, Marker #1 (psbA) was examined in more than 200 Echinochloa accessions and a phylogenetic tree grouped these Echinochloa accessions into four clades. Additionally, two different E. crus‐galli varieties (E. crus‐galli var. crus‐galli, E. crus‐galli var. praticola) and E. colona were successfully distinguished by this marker. The developed molecular markers contribute to better identification of Echinochloa taxa.     

Assessment of phylogenetic signal in the germination ability of Phelipanche ramosa on Brassicaceae hosts

Phelipanche ramosa is a generalist parasitic weed known to cause yield losses in various crops, especially in winter oilseed rape, its new preferred host in France. This parasitic plant is also able to complete its life cycle on many Brassicaceae weeds, which are thus important alternative hosts. We studied a set of 14 common Brassicaceae weeds and characterised their ability to induce the germination of three genetically distinct pathovars of P. ramosa, based on in vitro experiments. We then investigated whether phylogenetic relatedness among Brassicaceae weeds could inform on their ability to induce germination of P. ramosa by testing for a phylogenetic signal in the germination rate of the parasite. In the presence of some phylogenetic signal, phylogenetic distances among species might be used as a surrogate to predict the ability to induce germination of potential additional Brassicaceae hosts. The three pathovars studied showed different germination patterns. Moreover, we found substantial variation in the germination rate of P. ramosa among the different Brassicaceae species, with a significant effect of the clustering of species into two ancient phylogenetic lineages. However, no significant phylogenetic signal was detected overall, that is we could not exclude that germination rates were randomly distributed over the phylogeny. We suggest that further analyses should be conducted across wider sets of potential hosts to better characterise the existence of a phylogenetic signal of the ability of weeds to induce the germination of P. ramosa.     

中国新归化菊科植物——弯喙苣

弯喙苣属(Urospermum)有2种,原产于南欧、北非和西亚。其中,该属的模式种弯喙苣(Urospermum picroides(L.)Scop.ex F.W.Schmidt)目前已在北美洲、南美洲、澳大利亚、南非和巴基斯坦等许多地区归化,我国曾有引种栽培,近来成为宁波地区一种新的归化杂草。本文就弯喙苣的分类地位、形态特征、地理分布、生物学特征、传播途径、入侵危害、防治措施进行了报道,以期为外来有害杂草的鉴定提供借鉴。     

Molecular variability and phylogenetic relationships among different species and populations of <Emphasis Type="Italic">Pratylenchus</Emphasis> (Nematoda: Pratylenchidae) as inferred from the analysis of the ITS rDNA

Sequence comparisons and molecular phylogenetic analyses were used to describe the nucleotide variability of the ITS containing regions of eighteen Pratylenchus species and several populations. Comparative analysis of nucleotide sequences of the rDNA internal transcribed spacers (ITS1 and ITS2) among Pratylenchus species used in the present study demonstrates that ITS sequences can widely vary in primary sequence and length. Alignment of eighty-seven Pratylenchus sequences and one outgroup taxon reveals the presence of ambiguous regions that have the greatest effect on phylogeny reconstruction. Phylogenetic analyses using Bayesian Inference, Neighbour Joining-LogDet, Maximum Likelihood and Maximum Parsimony, distinguished twelve highly or moderately supported major clades within Pratylenchus. Our results support the taxonomic usefulness of the ITS region to identify root-lesion nematode species of the genus Pratylenchus but the high nucleotide variability, sometimes, can preclude its use to resolve relationships among all members of the genus. In addition, the phylogenetic groupings are not congruent with those defined by characters derived by lip patterns and numbers of lip annuli.     

Systematics of key phytopathogenic Fusarium species: current status and future challenges

This review is intended to provide plant pathologists and other scientists with a current overview of the most important Fusarium phytopathogens and mycotoxin producers. Knowledge of Fusarium species diversity and their evolutionary relationships has increased dramatically due to the application of multilocus molecular phylogenetics and genealogical concordance phylogenetic species recognition over the past 15 years. Currently Fusarium is estimated to comprise at least 300 genealogically exclusive phylogenetic species; however, fewer than half have been formally described. The most important plant pathogens reside in the following four groups: the F. fujikuroi species complex noted for Bakanae of rice, ear rot of maize, pitch canker of pine and several species that contaminate corn and other cereals with fumonisin mycotoxins; the F. graminearum species complex including the primary agents causing Fusarium head blight of wheat and barley that contaminate grain with trichothecene mycotoxins; the F. oxysporum species complex including vascular wilt agents of over 100 agronomically important crops; and the F. solani species complex, which includes many economically destructive foot and root rot pathogens of diverse hosts. Several other Fusarium phytopathogens reported from Japan and nested within other species complexes are reviewed briefly. With the abandonment of dual nomenclature, a broad consensus within the global community of Fusarium researchers has strongly supported the unitary use of the name Fusarium instead of several teleomorph names linked to it. Plant pathologists and other scientists needing accurate identifications of Fusarium isolates are encouraged to use Fusarium-ID and Fusarium MLST, Internet accessible websites dedicated to the molecular identification of Fusarium species.     

肾状线虫属重要种类的鉴定和特征描述

记述了目前世界上认可的肾状线虫属(Rotylenchulus sp.)已知的10个种的重要鉴定特征、世界分布和寄主范围。这10个种包括越南肾状线虫(R.anamictus)、短管肾状线虫(R.brevitubulus)、北方贝肾状线虫(R.borea-lis)、短尾肾状线虫(R.clavicaudatus)、离异肾状线虫(R.leptus)、大囊肾状线虫(R.macrodoratus)、长肾状线虫(R.macrosoma)、微小肾状线虫(R.parvus)、肾形肾状线虫(R.reniformis)和甘蔗肾状线虫(R.sacchari)。并对其最具经济意义的两种线虫的特征进行了描述。     

Host‐range studies,genetic diversity and evolutionary relationships of ACLSV isolates from ornamental,wild and cultivated Rosaceous species

A large‐scale survey was carried out to study the host range and genetic diversity of Apple chlorotic leaf spot virus (ACLSV) in various Rosaceae species, with a special emphasis on ornamentals and wild shrubs. Samples were tested by DAS‐ELISA using two different antisera, and RT‐PCR amplification of part of the CP gene. There was generally a poor correlation between the results obtained with the two sets of serological reagents and between serological and molecular detection assays. Using a nested RT‐PCR assay developed here, ACLSV was found to be widespread among cultivated, ornamental and wild species of the Rosaceae. The virus was detected for the first time in plum, wild cherry, Crataegus monogyna, Prunus spinosa and Prunus cerasifera in Greece. Sequences of a part of the CP encoding gene and the 3′ untranslated region from ACLSV isolates originating from various wild species and ornamentals were compared to those of isolates from cultivated hosts, showing similar divergence levels. Further phylogenetic analysis using the sequenced region indicated that the isolates from wild or ornamental hosts were not more closely related to each other than to isolates from cultivated hosts. The possible role of different factors in the spread of ACLSV on cultivated, ornamental and wild species is discussed.