Knockout of SlDCL2b attenuates the resistance of tomato to potato spindle tuber viroid infection.

RNA interference, or RNA silencing, is an important defence mechanism against viroid infection in plants. Plants encode multiple DICER-LIKE (DCL) proteins that are key components of the RNA silencing pathway. However, the roles of different DCLs in defence responses against viroid infection remain unclear. Here, we determined the function of tomato DCL2b (SlDCL2b) in defence responses against potato spindle tuber viroid (PSTVd) infection using SlDCL2b loss-of-function tomato mutant plants. Compared with wild-type plants, mutant plants were more susceptible to PSTVd infection, developing more severe symptoms earlier and accumulating higher levels of PSTVd RNAs. Moreover, we verified the feedback mechanism for the regulation of SlDCL2b expression by miR6026. Functional blocking of tomato miR6026, by expressing its target mimics, can enhance resistance to PSTVd infection in tomato plants. These findings deepen the current understanding of RNAi-based resistance against viroid infection and provide a potentially new strategy for viroid control.

Genetic Characteristics and Linkage of Virulence Genes of the Puccinia striiformis f. sp. tritici TSA-6 Isolate to Yr5 Host Resistance.

To understand the inheritance of the TSA-6 Puccinia striiformis f. sp. tritici (Pst) isolate that is virulent to Yr5 and was recently detected in China, we analyzed avirulence and virulence of 120 selfed progeny lines from Berberis shensiana. The results showed that the TSA-6 isolate is virulent against the Yr5 resistance gene, and overall progeny lines were categorized into 73 virulence phenotypes (VPs); of these, 72 VPs differed from the isolate TSA-6, and only one VP, including three progeny, was identical to the parental isolate. The analyses indicated that the TSA-6 isolate is homozygous for avirulence at the Yr10, Yr15, and Yr26 resistance loci and virulence at the YrA resistance locus. The TSA-6 isolate is heterozygous for avirulence at the Yr2, Yr3, Yr5, Yr7, and Yr8 resistance loci, which are controlled by a dominant/recessive relationship. The Yr1, Yr6, Yr9, Yr17, Yr27, Yr25, Yr28, Yr29, Yr32, YrTr1, and YrSP resistance loci are governed by two complementary dominant/recessive genes. Avirulence against heterozygous Yr4, Yr43, Yr44, Yr76, and YrExp2 resistance loci is regulated by a dominant and recessive or a dominant and suppressor gene pair. In total, 117 multilocus genotypes were detected at 24 KASP-SNP marker loci among the 120 progenies. Using these marker loci, we constructed a linkage map with a genetic distance interval spanning 624.5 cM. Quantitative trait loci corresponding to phenotypic segregation for virulence at 20 Yr resistance loci in addition to the Yr1 resistance locus were identified. These results facilitate our understanding of Pst virulence evolution and simplify breeding of wheat cultivars with effective resistance to wheat stripe rust.

Countrywide inter-epidemic region migration pattern suggests the role of southwestern population in wheat stripe rust epidemics in China.

Understanding countrywide pathogen population structure and inter-epidemic region spread is crucial for deciphering crop potential losses. Wheat stripe rust caused by Puccinia striiformis f. sp. tritici is a destructive disease that affects worldwide wheat production, widespread in China, representing largest epidemic region globally. This study aimed to understand the population structure and migration route of P. striiformis f. sp. tritici across China based on sampling from 15 provinces representing six epidemic zones, viz., over-summering, over-wintering, eastern, Yun-Gui, Xinjiang and Tibet epidemic regions. High genotypic diversity was recorded in over-summering, Tibet and over-wintering epidemic regions. Epidemic regions partly explain population subdivision with variable divergence (FST  = 0.005-0.344). Xinjiang and Tibet epidemic regions were independent epidemic zones with least sharing of genotypes. Among other epidemic zones, i.e. over-summering, over-wintering, eastern and Yun-Gui epidemic zones, re-sampling MLGs, clustering-based structure, DAPC analyses, relative migration and low divergence (FST from 0.006 to 0.073) revealed frequent geneflow. Yun-Gui epidemic regions, with a potential for both over-summering and over-wintering, could play an important role in causing epidemics in main wheat-cultivating areas of China. High diversity, recombination signatures and inter-epidemic region migration patterns need to be considered in host-resistant cultivar development in China and neighbouring countries, considering risk of long-distance migration capacity of pathogen.

Evaluation of the Potential Risk of the Emerging Yr5-Virulent Races of Puccinia striiformis f. sp. tritici to 165 Chinese Wheat Cultivars.

In 2017, a new race (TSA-6) of the wheat stripe rust pathogen, Puccinia striiformis f. sp. tritici, virulent to resistance gene Yr5, was detected in China. However, whether Chinese wheat cultivars are resistant to the new race was unknown. In this study, two isolates (TSA-6 and TSA-9) with virulence to Yr5 were tested on other wheat Yr gene lines for their avirulence and virulence patterns and used, together with prevalent races CYR32 and CYR34 without the Yr5 virulence, to evaluate 165 major Chinese wheat cultivars for their reactions. Isolates TSA-6 and TSA-9 had similar but different virulence spectra and therefore should be considered two different races. Their avirulence and virulence patterns were remarkably different from that of CYR34 but quite similar to that of CYR32. Of the 165 wheat cultivars, 21 had all-stage resistance to TSA-6, 34 to TSA-9, and 20 to both races. Adult plant resistance (APR) was detected in 35 cultivars to TSA-6 and 27 to TSA-9, but only three cultivars showed APR to both new races. Slow rusting resistance was observed in 24 cultivars to TSA-6 and 33 to TSA-9. Analysis of variance of disease index indicated a significant difference between cultivars but not between the four races. Based on the molecular marker data, a low percentage of wheat cultivars carried Yr5, Yr7, Yr10, Yr15, Yr26, or YrSP. Because TSA-6 and TSA-9 can be a serious threat to wheat production in China, continual monitoring of TSA-6, TSA-9, and other races is needed.

Evidence of Occurrence of Barley Crown Rust Caused by Puccinia coronata var. hordei and Sexual Reproduction of the Pathogen Under Field Conditions in China.

Crown rust of barley, caused by Puccinia coronata var. hordei, was first reported by Jin and Steffenson in 1992, and the fungus has been reported only in the United States and Hungary. In China, stripe, stem, and leaf rusts have been reported on barley, but not for crown rust. Recently, a sample (HZJ0004) of rust collected from barley in Qilian county in Qinghai, China, appeared different from the three rusts based on color, size, arrangements of uredinia and/or telia. Teliospores had crown-shaped appendages on the top. Based on the disease symptoms and morphology of urediniospores and teliospores, the fungus was identified as P. coronata var. hordei. Using the internal transcribed spacer sequences, the isolates HZJ0004 from barley and POR3 from buckthorn (Rhamnus sp.) were clustered in one clade with P. coronata var. hordei isolates from barley and Elymus repens but in a different clade from the isolate POC8 from wild oat and the varieties of P. coronata from oats and grasses. At the seedling stage, most of the tested cultivars of barley and rye were susceptible to P. coronata var. hordei isolates HZJ0004 and POR3, but the cultivars of oats, triticale, wheat, and most grasses of genera Aegilops, Brachypodium, Bromus, Calamagrostis, Deschampsia, Elymus, Festuca, and Phleum were resistant, indicating their host specialization on barley. To our knowledge, this is the first report of crown rust on barley in China.

Identification of Berberis spp. as Alternate Hosts for Puccinia achnatheri-sibirici Under Controlled Conditions and Morphologic Observations of Sexual Stage Development of the Rust Fungus.

Gramineous grasses are a large group of species, many of which act as accessory (secondary) host for a large number of rust fungi, including devastating rust pathogens of cereals. Among the rust fungi, some are known to be heteroecious and have a complete macrocyclic life cycle with five types of spores on distinct plant species, but for many others the complete life cycle is unknown. Puccinia achnatheri-sibirici, a rust fungus infecting grasses in the genus Achnatherum, has been known for only its uredinial and telial stages; however, the other (pycnial and aecial) stages have not been identified. In this study, we demonstrate that P. achnatheri-sibirici is a heteroecious, macrocyclic fungus with the sexual stage on barberry (Berberis spp.) through inoculation. Pycnia and aecia were successively produced on the inoculated barberry plants. Inoculation of Achnatherum extremiorientale leaves with aeciospores produced yellow-orange uredinia with high infection types, whereas inoculation of wheat variety Mingxian 169, highly susceptible to Puccinia striiformis f. sp. tritici which causes stripe rust on wheat, produced chlorotic flecks but no uredinia. The ITS sequence analysis of P. achnatheri-sibirici did not match with any sequence in the NCBI database and had the highest homology with 94% compared to Puccinia brachypodii and Puccinia aizazii. Observations of the uredinial, telial, and basidial stages on A. extremiorientale and pycnial and aecial stages on Berberis shensiana using light and scanning electron microscopes revealed its characteristics. Morphological characteristics of urediniospores and teliospores are most similar with those of P. achnatheri-sibirici described in the literature. This study proved (1) the life cycle of P. achnatheri-sibirici as heteroecious and macrocyclic and the alternate host as barberry; (2) the description of life stages in the sexual cycle, especially the morphologies of aecial, pycnial, and basidial stages; and (3) the expansion of knowledge on the rust flora on barberry.