A secreted catalase contributes to Puccinia striiformis resistance to host-derived oxidative stress

Plants can produce reactive oxygen species (ROS) to counteract pathogen invasion, and pathogens have also evolved corresponding ROS scavenging strategies to promote infection and pathogenicity. Catalases (CATs) have been found to play pivotal roles in detoxifying H2O2 formed by superoxide anion catalyzed by superoxide dismutases (SODs). However, few studies have addressed H2O2 removing during rust fungi infection of wheat. In this study, we cloned a CAT gene PsCAT1 from Puccinia striiformis f. sp. tritici (Pst), which encodes a monofunctional heme-containing catalase. PsCAT1 exhibited a high degree of tolerance to pH and temperature, and forms high homopolymers.Heterologous complementation assays in Saccharomyces cerevisiae reveal that the signal peptide of PsCAT1 is functional. Overexpression of PsCAT1 enhanced S. cerevisiae resistance to H2O2. Transient expression of PsCAT1 in Nicotiana benthamiana suppressed Bax-induced cell death. Knockdown of PsCAT1 using a host-induced gene silencing (HIGS) system led to the reduced virulence of Pst, which was correlated to H2O2 accumulation in HIGS plants. These results indicate that PsCAT1 acts as an important pathogenicity factor that facilitates Pst infection by scavenging host-derived H2O2.

Two new species and one new record of Hyalopsora (Pucciniastraceae) on ferns in China

Three rust fungi belonging to the genus Hyalopsora occurring on ferns were found in China. Hyalopsora minispora and H. tibetica are described as new species in this paper. Hyalopsora minispora can be distinguished from known species by its relatively small urediniospores (19.5–23.0 × 12.0–19.0 μm) with 4–6 scattered germ pores and amphispores in a common pustule. Hyalopsora tibetica is mainly characterized by urediniospores with 2–6 scattered germ pores and amphispores. Hyalopsora neocheilanthis is described and reported as a new record for China. The phylogenetic relationships of Hyalopsora species and related taxa were examined by comparing the sequences of their internal transcribed spacer 2 (ITS2) barcode and 28S rDNA gene segments. Hyalopsora is supported with strong bootstrap support and it differs from the two other fern-infecting genera viz. Milesina and Uredinopsis.

Advances in understanding the biology and epidemiology of rust diseases of cereals

Rust fungi (order: Pucciniales) constitute the largest group of plant parasitic fungi and include many species of agricultural importance. This includes the three wheat rust fungi (Puccinia graminis f. sp. tritici, Puccinia striiformis f. sp. tritici and Puccinia triticina) that have posed a threat to crop production throughout history. This chapter provides an overview of the wheat rust pathogen lifecycle that has been critical to the design of effective disease management strategies and discusses recent integration of basic biological knowledge and genomic-led tools within an epidemiological framework. Furthermore, we include a case study on the “field pathogenomics” technique, illustrating the value of genomic-based tools in disease surveillance activities. Bringing together advances in understanding basic pathogen biology, developments in modelling for disease forecasting and identification, alongside genomic-led advances in surveillance and resistance gene cloning, holds great promise for curtailing the threat of these notorious pathogens.

Two new species of rust fungi (Pucciniales) from the Brazilian Amazon: Aecidium margaritariae on Margaritaria and Uromyces amapaensis on Jatropha

ABSTRACT Two new species of Pucciniales fungi on plants of the Fabid clade are described from samples deposited in the herbarium of Museu Paraense Emilio Goeldi, collected in the state of Amapá, in the Brazilian Amazon. They are Aecidium margaritariae found on Margaritaria nobilis (Phyllanthaceae), and Uromyces amapaensis on Jatropha gossypiifolia (Euphorbiaceae). The microstructures of the specimens were analyzed using optical and scanning electron microscopy. Aecidium margaritariae is characterized by the presence of globose, subglobose to slightly ellipsoid aeciospores with warty walls and smooth areas usually in the basal portion. Uromyces amapaensis is distinguished by the presence of uredinia with paraphyses which are thickened and rounded at the tip, and pedicellate and smooth teliospores. Descriptions, illustrations, and taxonomic comments are presented for each species.

A kiwellin protein-like fold containing rust effector protein localizes to chloroplast and suppress cell death in plants

The effector proteins expressed by plant pathogens are one of the essential components of the host-pathogen interaction. Despite being important, most of the effector proteins remain unexplored due to the lack of conserved features and huge diversity in their primary sequence. In the present study, extensive secretome analysis was performed in sixteen major plant fungal pathogens to find the conserved features in the candidate secretory effector proteins (CSEPs) using homology and ab initio modeling approaches. Interestingly, a variable number of plant kiwellin proteins fold like secretory proteins were found in all the major rust fungal pathogens. Many of them are predicted as potential effector proteins. For instance, 26 out of 35 Kiwellin like proteins identified in Puccinia striiformis race 104E 137A were predicted as potential effector proteins. In addition, a kiwellin predicted effector gene, Pst_13960, from the Indian Puccinia striiformis race Yr9 was characterized using overexpression, localization, and deletion studies in Nicotiana benthamiana. The Pst_13960 suppressed the BAX-induced cell death and localized in the chloroplast. Furthermore, the expression of the kiwellin matching region (Pst_13960_kiwi) alone suppressed the BAX-induced cell death in N. benthamiana despite the change of location to the cytoplasm and nucleus, suggesting the novel function of the kiwellin fold in rust fungi. Further analysis of these proteins predicted these candidates to contain N-terminal Intrinsically disordered regions (IDRs) putatively associated with chloroplast translocation as deletion of region abolished the chloroplast localization of Pstr_13960. Overall, the current study reports the presence of kiwellin like proteins in rust fungi that act as a novel effector in plants.

Comparative secretomics identifies conserved WAxR motif-containing effectors in rust fungi that suppress cell death in plants

Identification of novel effectors with conserved features has always remained a challenge in plant-pathogen interaction studies. The introduction of the genomics era in plant-pathogen studies has led to the identification of significant candidate effectors with novel motifs such as RxLR and dEER motifs. However, in the case of fungal pathogens, limited conserved motifs associated with effectors have been discovered yet. In the present study, we have performed comparative secretome analysis for major plant pathogens of diverse nutrition mechanisms with the aim of dissecting the features underlying their corresponding secretome and conserved motifs. We showed that rust fungi possess the lowest Cell wall degrading enzymes (CWDEs) consortium lower than other biotrophic pathogens. We also showed rust fungi possess the highest secretory superoxide dismutase (SOD) than other studied plant pathogens. Further, we prioritized the candidate secretory effectors proteins (CSEPs) of all the studied pathogens by combining various effector mining parameters to highlight the candidates with potential effector features. A novel WAxR motif in conjugation with the Y/F/WxC (FGC) motif was identified in the effectors of various P. striiformis races present globally. The WAxR/WAxR like motifs ( WxxR, WAxx, xAxR) containing effectors were also found in the secretome of other rust fungi. Further, the functional validation of two candidate effectors with WAxR motif from P. striiformis Yr9 showed that these effectors localize to the nucleus as well as cytoplasm, and are able to suppress BAX induced cell death in Nicotiana benthamiana. The mutation analysis of individual residues of the WAxR motif (W, A, R ) however did not affect the cell death suppression nor subcellular localization of these effectors. Overall, the current study reports the presence of novel motifs in large numbers of effectors of rust fungi with cell death suppression features.

Towards improved detection and identification of rust fungal pathogens in environmental samples using a metabarcoding approach

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.

Host Adaptation and Virulence in Heteroecious Rust Fungi

Rust fungi (Pucciniales, Basidiomycota) are obligate biotrophic pathogens that cause rust diseases in plants, inflicting severe damage to agricultural crops. Pucciniales possess the most complex life cycles known in fungi. These include an alternation of generations, the development of up to five different sporulating stages, and, for many species, the requirement of infecting two unrelated host plants during different parts of their life cycle, termed heteroecism. These fungi have been extensively studied in the past century through microscopy and inoculation studies, providing precise descriptions of their infection processes, although the molecular mechanisms underlying their unique biology are poorly understood. In this review, we cover recent genomic and life cycle transcriptomic studies in several heteroecious rust species, which provide insights into the genetic tool kits associated with host adaptation and virulence, opening new avenues for unraveling their unique evolution. Expected final online publication date for the Annual Review of Phytopathology, Volume 59 is August 2021. Please see http://www.annualreviews.org/page/journal/pubdates for revised estimates.