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Morphological features of Puccinia chunjii. A. Telia on infected stem. B. Telium cross section. C. teliospores showing various shapes and the severely thickened apical wall. Bars: A. unit 5 1 mm; B, C 5 20 mm.
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A rust specimen with macroscopic similarities to the cereal stem rusts was collected on Elymus sp. from Gansu province, China. Phylogenetic analyses of ITS and COI DNA sequences indicated the fungus was closely related but distinct as a strongly supported sister taxon to the Puccinia graminis species complex. Microscopic examination revealed diagno...
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... with an Olympus DP 70 camera and analyzed by Image-Pro Plus 6.0 image processing and analysis software (MediaCybernetics Inc., Bethesda, Mary- land) to obtain measurements. Teliospore length was measured excluding the thickened apical walls and the projections, that is from the hilum to the base of the thickened apical wall or the projections (FIG. 4C, ...
Citations
... More than 5000 Puccinia species have been recorded across the world, among which approximately 520 species and varieties have been reported in China (Cao et al. 2000;Dai and Zhuang 2015;Liu and Hambleton 2012;Liu et al. 2014;Zhuang 2015, 2016a;Liu et al. 2016b;Zhao 45 et al. 2021;Zhuang and Wei 1999;Zhuang et al. 1998, Museum of the Beijing Forestry University (BJFC), Beijing, China, and Fungarium (HMAS), Institute of Microbiology, Chinese Academy of Sciences. For morphological comparisons with related taxa, we borrowed 75 24 herbarium specimens from Museum of Beijing Forestry University. ...
Members of Puccinia (Pucciniaceae, Pucciniales) are known as plant pathogens worldwide, which are characterized by their morphology, host association, and molecular data of various genes. In the present study, 10 specimens of Puccinia were collected from four herbaceous plants (Anaphalis hancockii, Anthriscus sylvestris, Halenia elliptica, and Pilea pumila) in China and identified based on morphology and phylogeny. As a result, 10 samples represent four undescribed species of Puccinia, viz., P. apdensia, P. decidua, P. dermatis, and P. lianchengensis, spp. nov. P. apdensia is characterized by its smooth teliospores with thickened apex. P. decidua represents the first Puccinia species inhabiting the host Anaphalis hancockii and is distinguished from the other Puccinia species by its telia and uredinia surrounded by the epidermis. P. dermatis from Halenia elliptica differs from the other Puccinia species on the host genus Halenia by the telia that have epidermis and teliospores with sparsely irregular granulated protrusions. P. lianchengensis is characterized by its teliospore surface with fishnet ornamentation and urediniospores without prominent caps. All of the new species are described and illustrated in this study.
... Methods for polymerase chain reaction (PCR) ampli cation and sequencing of the ITS2 plus partial 28S region (when possible) for the rusts, and a portion of the ribulose-1,5-bisphosphate carboxylase/oxygenase large subunit (rbcL) gene for selected hosts, were as described previously (Léveillé-Bourret, Eggertson, Hambleton, & Starr, 2021). Additional sequences for the telial specimen, DAOM 240982, were also generated using the same DNA extract as in the original publication (Liu & Hambleton, 2012) to add partial 28S (the GenBank record was updated) and host rbcL data. The sequence for the host psbA-trnH intergenic spacer discussed in Liu and Hambleton (2012) has since been deposited in GenBank. ...
... Additional sequences for the telial specimen, DAOM 240982, were also generated using the same DNA extract as in the original publication (Liu & Hambleton, 2012) to add partial 28S (the GenBank record was updated) and host rbcL data. The sequence for the host psbA-trnH intergenic spacer discussed in Liu and Hambleton (2012) has since been deposited in GenBank. Sequences were edited using Geneious Prime 2021.2 (Biomatters, Auckland, New Zealand). ...
... Collection information, sequence accession numbers and aeciospore sizes are summarized in Table 1 for the specimens examined. For the detailed description of telia and teliospores, see Liu and Hambleton (2012). Uredinia were not seen. ...
The telial stage of Puccinia chunjiei was described in 2012 from a single specimen collected in China (DAOM 240982). This species is the closest relative of P. graminis but differs in telial morphology on their grass hosts and in DNA sequences. As part of a DNA-barcoding project for rust herbarium specimens, collections of the aecial stage of P. graminis on Berberis were processed. For one specimen, BPI 1103856, the ITS sequence matched that of P. chunjiei and the aecial morphology differed from P. graminis. An expanded description of P. chunjiei is presented with photographs of the aecial stages of both species.
... Supplementary Data S1: A. Sequence alignment of the genic region OG08220 for Tilletia walkeri and T. indica isolates sequenced by Nguyen et al. [43]; B. Sequence alignment of the isolates sequenced by Nguyen et al. [43] presenting part of the region OG01193 unique to Tilletia walkeri and T. indica used for the qPCR assay design. References [68][69][70] are cited in the supplementary materials. Funding: This research was funded by Agriculture and Agri-Food Canada grant number J-002272 "Fungal and Bacterial Biosystematics" with additional support from the Canadian Food Inspection Agency, N-000277 "Advanced pre-screening of export grain and oilseed commodities using machine learning on long read sequencing"; and N-000095 "Identification of molecular markers for development of detection and genotyping tools for regulated plant pathogens". ...
Several fungi classified in the genus Tilletia are well-known to infect grass species including wheat (Triticum). Tilletia indica is a highly unwanted wheat pathogen causing Karnal bunt, subject to quarantine regulations in many countries. Historically, suspected Karnal bunt infections were identified by morphology, a labour-intensive process to rule out other tuberculate-spored species that may be found as contaminants in grain shipments, and the closely-related pathogen T. walkeri on ryegrass (Lolium). Molecular biology advances have brought numerous detection tools to discriminate Tilletia congeners (PCR, qPCR, etc.). While those tests may help to identify T. indica more rapidly, they share weaknesses of targeting insufficiently variable markers or lacking sensitivity in a zero-tolerance context. A recent approach used comparative genomics to identify unique regions within target species, and qPCR assays were designed in silico. This study validated four qPCR tests based on single-copy genomic regions and with highly sensitive limits of detection (~200 fg), two to detect T. indica and T. walkeri separately, and two newly designed, targeting both species as a complex. The assays were challenged with reference DNA of the targets, their close relatives, other crop pathogens, the wheat host, and environmental specimens, ensuring a high level of specificity for accurate discrimination.
... Rust fungi (>8400 species; Pucciniales, Basidiomycota) take their name from their usually bright-orange replicative spores (Aime et al., 2014). They are responsible for several economically devastating crop diseases such as crown rust of oats, stem rust of wheat, and white pine blister rust (Saari & Prescott, 1985;Geils et al., 2010;Liu & Hambleton, 2012, but they are also ecologically important as mediators of plant competition in natural environments (Barnes et al., 2005;Price et al., 1988; Accepted Article Rice & Westoby, 1982), and as food for many fungal and arthropod hyper-parasites (Lutz et al., 2004a(Lutz et al., , 2004bNischwitz et al., 2005;Nelsen, 2010;Henk et al., 2011;Trakunyingcharoen et al., 2014). ...
Plants play important roles as habitat and food for a tremendous diversity of specialist animals and fungi. The disappearance of any plant species can lead to extinction cascades of its associated biota. In consequence, documenting the diversity and specificity of plant‐associated organisms is of high practical relevance in biodiversity conservation. Here we present the first large‐scale molecular investigation into the diversity, host‐specificity, and cophylogenetic congruence of an especially rich plant‐fungal association, the rust fungi (Pucciniaceae) of Cyperaceae and Juncaceae. Using the largest rust fungi DNA barcoding dataset published to date (252 sequences, 82 taxa), we reject the presence of a global ITS2‐28S barcode gap, but find a local gap in Cyperaceae‐Juncaceae rusts, and suggest the existence of many cryptic species in North America, with some broadly‐circumscribed species possibly corresponding to >10 cryptic species. We test previous hypotheses of correlations between the phylogenies of rust fungi and their Cyperaceae‐Juncaceae hosts using a combination of global‐fit and event‐based cophylogenetic methods. Significant cophylogenetic signal is detected between rusts and their hosts, but the small number of cospeciations argues for preferential host jumps as the driving process behind these correlations. In addition, temporal congruence between the origin of major Carex clades and their rusts suggests that host diversification may have promoted parasite diversification. Finally, we discuss the relevance of rust infection patterns to the systematics of Cyperaceae, highlight some taxonomic problems uncovered by the analyses, and call attention to the promise of DNA barcoding for bridging knowledge gaps in poorly studied plant‐associated microorganisms.
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... These novelties were from 41 different genera, ten of which represent new genera, and therefore entirely new genetic lineages. Most of the new rust fungi species were described in the genera Puccinia (66 new species; Abbasi et al. 2002;Abbasi and Darvishnia 2015;Afshan and Khalid 2008Aliabadi and Abbasi 2012;Bahcecioglu andGjaerum 2003, 2004;Bahcecioglu et al. 2005Bahcecioglu et al. , 2009Berndt 2007Berndt , 2009Berndt , 2010Berndt , 2013aBerndt and Freire 2004;Berndt and Hüseyin and Kirbag 2003;Iqbal et al. 2009;Kabaktepe 2015;Khalid and Afshan 2009;Kirbag et al. 2001Kirbag et al. , 2011Liu and Hambleton 2012;McKenzie 2008;McKenzie and Johnston 2004;Mennicken and Oberwinkler 2004;Okane et al. 2014;Perdomo-Sanchez and Piepenbring 2008;Sotao et al. 2007;Thaung 2011;Wei 2001, 2011), Uromyces (28 new species ;Agarwal 2003;Bahcecioglu 2014;Bahcecıoglu and Gjaerum 2004;Berndt 2002aBerndt , 2004Berndt , 2009Berndt , 2013bBerndt and Baiswar 2009;Berndt and Uhlmann 2006;Berndt et al. 2007;Doungsaard et al. 2014;Hernandez et al. 2005;Mennicken and Oberwinkler 2004;Perdomo-Sanchez and Piepenbring 2014;Rezende and Dianese 2003;Thaung 2009;Walker and van der Merwe 2009;Wood and Scholler 2005;Zhuang and Wei 2003), Uredo (16 new species ;Berndt 2002bBerndt , 2004Berndt , 2009Berndt and Freire 2004;Berndt and Uhlmann 2006;Berndt and Wood 2012;Berndt et al. 2007;Cao et al. 2000;Hernandez et al. 2005;Mennicken and Oberwinkler 2004;Wei 2011, 2012), Prospodium (12 new species; Berndt 2002b; Berndt et al. 2007;de Carvalho and Hennen 2010), and Phakopsora (11 new species; Bagyanarayana et al. 2001;Beenken 2014;Berndt and Wood 2012;Berndt et al. 2008;Ferreiea et al. 2001;Maier et al. 2015;Ono 2000;Ono et al. 2012;Ritschel et al. 2007). Interestingly, species descriptions for rust genera follow the same trend as was seen for classes, i.e. the most species-rich genera (Puccinia, Uredo, and Uromyces) had the highest number of new species discovered. ...
Pucciniomycotina, one of the three subphyla of Basidiomycota, contains a range of microfungi from various habitats and with different lifestyles. In addition to familiar plant pathogenic rusts and anther smuts, the group also contains saprobic and pathogenic yeasts, minute sporocarp-forming fungi, and anamorphic moulds among others. Our knowledge of this group is still improving; over the last 16 years alone, researchers have described 375 new species of Pucciniomycotina, most of which were isolated from less documented areas such as Asia, South America, and Africa. While the majority of these new species belong to the species-rich rust fungi (Pucciniales), exploration in extreme environments such as deep-sea sediments and psychrophilic habitats is uncovering a variety of Pucciniomycotina species, especially yeasts. Molecular phylogenetic studies have greatly improved our understanding of the relationships between these taxa over the last 10 years. As presently circumscribed, the subphylum contains nine classes and 20 orders, the relatedness for most of which was not suspected until recently. Genomic data from members of the subphylum have been scarce but increasing over the last 5 years. We now know, for example, that Pucciniomycotina contains both fungi with the largest known genomes (rust fungi, up to 900 Mb) as well as a fungus with the smallest genome in Basidiomycota (Mixia osmundae, 13 Mb). This chapter discusses these latest developments in Pucciniomycotina research and highlights some challenges still to overcome in order to improve our understanding of this enigmatic group of fungi.
... The phylogenetic trees recovered lineages corresponding to P. Series Striiformis (Liu & Hambleton 2010), P. Series Coronata , P. chunjii (Liu & Hambleton 2012), P. graminis s.l., P. hordei, P. poaenemoralis, P. poarum, P. recondita and P. triticina. Clade numbers in the P. graminis s.l. ...
The rust species Puccinia graminis and P. striiformis sensu stricto are important cereal pathogens, most well-known for the potential of causing disease epidemics on wheat and generating significant yield losses. Early and accurate detection of the pathogens would facilitate effective control of the diseases. In the present study we developed real-time PCR assays to detect the specific lineages that include the wheat pathogens for each species complex, as identified using multi-gene DNA sequence analyses. Four DNA loci, for a comprehensive set of target and closely related fungi collected from diverse hosts and geographic regions, were explored to search for suitable lineage-specific probes: β-tubulin (BT), cytochrome c oxidase subunit 1 (COI), rDNA internal transcribed spacer (ITS), and RNA polymerase II second largest subunit (RPB2). Four TaqMan® real-time PCR assays were designed based on either the BT or RPB2 genes: one targeting Puccinia Series Striiformis (PSBT), one targeting P. striiformis sensu stricto (PSstrRPB2), and two targeting the P. graminis lineages on wheat (Pg2+BT and Pg2RPB2). Sensitivities of the assayswere determined to be 0.65 pg/uL (Pg2) and 5 pg/uL (PS). Specificity of each assay was confirmed using a broad diversity of rusts and selected other wheat-associated fungi. The ITS and COI loci were found to be unsuitable for diagnostic assay development but contributed phylogenetic signal to the multi-gene analyses.
... The COXI gene is used in DNA barcoding in animals (http://www.dnabarcoding101.org/) and has been reported to be valuable for the identification of some fungi, Penicillium species (Seifert et al. 2007) and cereal rusts (Liu and Hambleton 2012). The availability of sequences for a short segment of the mitochondrial COXI gene for a large number of rust species facilitated the use of the short COXI gene segment for an analysis of evolutionary relationships of myrtle rust with other rust species. ...
Using de novo assembly of 46 million paired end sequence reads of length 250 bp for a myrtle rust isolate, we have estimated its genome size to be between 103 and 145 Mb and the number of proteins as >19,000. Annotation of the contigs found a very large percentage of proteins are associated with molecular functions of DNA binding or binding in biological processes for DNA integration and RNA-dependent DNA replication. A large proportion of these activities are attributed to the transposable elements (TEs). These elements are estimated to comprise 27% of the genome with 22% retrotransposons and 5% DNA transposons. The exon and intron boundaries of 46 genes occurring on contigs >20,000 bp have been determined. The number of introns range from 2 to 20 with a mean of 7. Phylogenetic analyses using partial COXI, 18S rRNA and 28S rRNA genes have placed myrtle rust in the Pucciniaceae lineage on a separate taxonomic branch from the families of Pucciniaceae, Phragmidiaceae, Sphaerophragmiaceae, Phragmidiaceae, Uropyxidaceae, Chaconiaceae and Phakopsoraceae. Further work is thus required to determine the family placement of myrtle rust in the Pucciniaceae of Pucciniales.
... The DNA barcoding system uses a short, standardized gene region of 648 bp as the core barcode region for animals (http://www.dnabarcoding101.org/). The COXI gene has been reported to be valuable for the identification of Penicillium species(Serfert et al. 2007) and cereal rusts(Liu and Hambleton 2012). The availability of sequences for a short segment of the mitochondrial COXI gene for a large number of rust species facilitated the use of the short COXI gene segment for the analysis of evolutionary relationships of myrtle rust with other rust species. ...
Using De novo assembly of 46 million paired end sequence reads of length 250 bp for a myrtle rust isolate, we have estimated the genome size to be between 103 ─145 Mb and the number of proteins as >19,000. Mapping and annotation of the contig sequences found a very large percentage of proteins are associated with molecular functions of DNA binding or binding in biological processes for DNA integration and RNA-dependent DNA replication. A large proportion of these activities are attributed to the transposable elements (TEs). These elements are estimated to comprise 27% of the genome with 22 % retrotransposons and 5% DNA transposons. They are thus the major players in the generation of genetic variability in the pathogen’s adaptation to the environment and new hosts.
It is thus postulated that myrtle rust is a complex of genotypes from races
to species and possess the genetic ability to jump across hosts of different
species and genera of Myrtaceae. The exon and intron boundaries of forty
six genes occurring on contigs > 20,000 bp have been determined. The b-
tubulin gene has been annotated on a contig of length 3,439 bp. All these
myrtle rust gene sequences have been submitted to GenBank. The
number of introns range from 2 to 20 with a mean of 7.3. Phylogenetic
analysis using the clathrin associated protein, the b-tubulin gene and the
partial COXI gene from this work has placed myrtle rust in the major clade
of Pucciniaceae in the Pucciniales lineage but on a separate taxonomic
branch from the one for the cereal rust fungi and Phakopsora species.
... Leaf rust collections. Thirty-one P. recondita specimens on genus Elymus (the host of some specimens were labeled as Agropyron, Leymus, and Hystrix; (25)(26)(27). Voucher numbers, hosts, origins, and GenBank accession numbers are listed in Table 1. Sequences for some specimens were determined in previous studies by M. Liu and S. Hambleton, as indicated in Table 1. ...
... samples, amplification of the fragment of 5.8S-ITS2 was attempted for 86 samples, and 82 were successfully amplified, including 9 type specimens. Twenty-eight sequences of reference species from previous studies were also used (25)(26)(27). As a single-copy gene, EF1-α had a lower success rate for amplification, and 58 sequences out of 86 samples were obtained, including 4 type specimens. ...
... g Sequences published by Liu and Hambleton(25). h Sequences published by Liu and Hambleton(27). ...
The classification of brown leaf rust fungi (Puccinia recondite complex and allied species) on wheat (Triticum aestivum), rye (Secale cereale), and other grasses in the family Poaceae has experienced a long history of controversy and uncertainty due to the reduced morphological characteristics available for taxonomy and difficulty of conducting interfertility experiments. However, because these are pathogens on important crops, it is important to clarify the species delimitations reflecting the natural lineages. In this study, phylogenetic analyses were conducted with DNA sequence data from the ribosomal DNA internal transcribed spacer region and elongation factor 1-alpha to elucidate this species complex. Three phylogenetic lineages were recovered within the complex of rye leaf rust fungi, P. recondita sensu stricto, which is congruent with existing classifications based on DNA content, sexual compatibility, and morphological studies. The brown leaf rust fungus on wheat (P. triticina) grouped with the related species P. persistens on Elymus repens and E. intermedia as a strongly supported clade. Collections on other Elymus spp. were separated into six clades. Based on the phylogenetic affinities of nine type specimens and aecial host associations, potential taxonomic names were evaluated for selected lineages.
The dispersion of fungal inocula such as the airborne spores of rust fungi (Pucciniales) can be monitored by metabarcoding the internal transcribed spacer 2 (ITS2) of the rRNA gene in environmental DNAs. This is largely dependent upon a high-quality reference database (refDB) and primers with proper taxonomic coverage and specificity. For this study, a curated ITS2 reference database (named CR-ITS2-refDB) comprising representatives of the major cereal rust fungi and phylogenetically related species was compiled. Inter- and intra-specific variation analyses suggested that the ITS2 region had reasonable discriminating power for the majority of the Puccinia species or species complexes in the database. In silico evaluation of nine forward and seven reverse ITS2 primers, including three newly designed, revealed marked variation in DNA amplification efficiency for the rusts. The theoretical assessment of rust-enhanced (Rust2inv/ITS4var_H) and universal fungal (ITS9F/ITS4) ITS2 primer pairs was validated by profiling the airborne rust fungal communities from environmental samples using a metabarcoding approach. Species or subspecific level identification of the rusts was improved by using CR-ITS2-refDB, and the Automated Oligonucleotide Design Pipeline (AODP), which identified all mutations distinguishing highly conserved DNA markers amongst close relatives. A generic bioinformatics pipeline was developed, including all steps employed in this study from in silico evaluation of primers to accurate identification of short metabarcodes at the level of interest for defining phytopathogens. The results highlighted the importance of primer selection, refDBs that are resolved to reflect phylogenetic relationships, and the use of AODP for improving the reliability of metabarcoding in phytopathogen biosurveillance.
