Delineating the elusive BaMMV resistance gene rym15 in barley by medium-resolution mapping

Barley mild mosaic virus (BaMMV), transmitted by the soil-borne protist Polymyxa graminis, has a serious impact on winter barley production. Previously, the BaMMV resistance gene rym15 was mapped on chromosome 6HS, but the order of flanking markers was non-collinear between different maps. To resolve the position of the flanking markers and to enable map-based cloning of rym15, two medium-resolution mapping populations Igri (susceptible) × Chikurin Ibaraki 1 (resistant) (I × C) and Chikurin Ibaraki 1 × Uschi (susceptible) (C × U), consisting of 342 and 180 F2 plants, respectively, were developed. Efficiency of the mechanical inoculation of susceptible standards varied from 87.5 to 100% and in F2 populations from 90.56 to 93.23%. Phenotyping of F2 plants and corresponding F3 families revealed segregation ratios of 250 s:92r (I × C, χ2 = 0.659) and 140 s:40r (C × U, χ2 = 0.741), suggesting the presence of a single recessive resistance gene. After screening the parents with the 50 K Infinium chip and anchoring corresponding SNPs to the barley reference genome, 8 KASP assays were developed and used to remap the gene. Newly constructed maps revealed a collinear order of markers, thereby allowing the identification of high throughput flanking markers. This study demonstrates how construction of medium-resolution mapping populations in combination with robust phenotyping can efficiently resolve conflicting marker ordering and reduce the size of the target interval. In the reference genome era and genome-wide genotyping era, medium-resolution mapping will help accelerate candidate gene identification for traits where phenotyping is difficult.

Delineating the Elusive BaMMV Resistance Gene rym15 in Barley by Medium-Resolution Mapping

Barley mild mosaic virus (BaMMV), transmitted by the soil-borne protist Polymyxa graminis, has a serious impact on winter barley production. Previously, the BaMMV resistance gene rym15 was mapped on chromosome 6HS, but the order of flanking markers was non-collinear between different maps. To resolve the position of the flanking markers and to enable map-based cloning of rym15, two medium-resolution mapping populations Igri (susceptible) × Chikurin Ibaraki 1 (resistant) (I×C) and Chikurin Ibaraki 1 × Uschi (susceptible) (C×U), consisting of 342 and 180 F2 plants, respectively, were developed. Efficiency of the mechanical inoculation at susceptible standards varied from 87.5–100% and in F2 populations from 90.56–93.23%. Phenotyping of F2 plants and corresponding F3 families revealed segregation ratios of 250s:92r (I×C, χ2 = 0.659) and 140s:40r (C×U, χ2 = 0.741), suggesting the presence of a single recessive resistance gene. Eight KASP assays, developed after screening the parents with the 50K Infinium chip and anchoring corresponding SNPs to the barley reference genome, were used to remap the gene. Newly constructed maps revealed a collinear order of markers, thereby allowing the identification of high throughput flanking markers. This study demonstrates how construction of medium-resolution mapping populations in combination with robust phenotyping can efficiently resolve conflicting marker ordering and reduce the size of the target interval. In an era of reference genomes and high throughput marker platforms, medium-resolution mapping will help accelerate candidate gene identification for traits where phenotyping is difficult.

Comparative proteomic analysis of Nicotiana benthamiana plants under Chinese wheat mosaic virus infection

Background Chinese wheat mosaic virus (CWMV) is a severe threat to winter wheat and is transmitted by Polymyxa graminis. The mechanisms of interactions between CWMV and plants are poorly understood. In this study, a comparative proteomics analysis based on nanoliquid chromatography mass spectrometry (MS)/MS was conducted to characterize proteomic changes in plants responding to CWMV infection. Results In total, 2751 host proteins were identified, 1496 of which were quantified and 146 up-regulated and 244 down-regulated proteins were identified as differentially expressed proteins (DEPs). Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis showed that DEPs were most strongly associated with photosynthesis antenna proteins, MAPK signaling plant and glyoxylate and dicarboxylate metabolism pathways. Subcellular localization analysis predicted that more than half of the DEPs were localized in the chloroplast, an organelle indispensable for abscisic acid (ABA) synthesis. Our results suggest that CWMV infection interrupts normal chloroplast functions and decreases ABA concentrations in Nicotiana benthamiana. Further analysis showed that the ABA pathway was suppressed during CWMV infection and that ABA treatment induced plant hosts defenses against CWMV. Conclusions We identified several candidate proteins expressed during CWMV infection, and the ABA pathway was strongly associated with responses to CWMV infection in N. benthamiana.

Comparative Proteomic Analysis of Nicotiana Benthamiana Seedings Under Chinese Wheat Mosaic Virus Infection

Background: Chinese wheat mosaic virus (CWMV) is a severe threat to winter wheat, and it is transmitted by Polymyxa graminis. However, the mechanisms of interactions between CWMV and plants are poorly understood. In this study, a comparative proteomics analysis based on nanoliquid chromatography (LC)- mass spectrometry (MS)/MS technique was conducted to characterize the proteomic changes of plants in response to CWMV infection. Results: A total of 2,751 host proteins were identified, 1,496 of which were quantified. 146 up-regulated and 248 down-regulated proteins were identified as differentially expressed proteins (DEPs). KEGG enrichment analysis showed that the DEPs were most strongly associated with Photosynthesis - antenna proteins, MAPK signaling – plant and Glyoxylate and dicarboxylate metabolism pathways. Subcellular localization analysis showed that more than half of DEPs were predicted to be localized in the chloroplast, an organelle indispensable for abscisic acid (ABA) synthesis. Our results suggested that CWMV infection interrupts normal chloroplast functions and decreased the contents of ABA in Nicotiana benthamiana. Further analysis showed that the ABA pathway was suppressed under CWMV infection and ABA treatment did induce plant hosts defense against CWMV. Conclusions: In summary, our results identified several candidate proteins under CWMV infection, and the ABA pathway was deeply involved in the responses to CWMV infection in N. benthamiana.

Ribotypes of Polymyxa graminis in Wheat Samples Infected with Soilborne Wheat Viruses in China

Polymyxa graminis is an obligate parasite and important vector of more than 14 soilborne plant viruses that pose a significant threat to cereal crops in Europe, North America, and Asia. Different ribotypes or formae speciales of P. graminis have been recognized and these may be associated with different cereal hosts or with transmission of different viruses. Two soilborne viruses infecting winter wheat in China have been reported and well studied (Wheat yellow mosaic virus [WYMV, genus Bymovirus] and Chinese wheat mosaic virus [CWMV, genus Furovirus]) but there has been no reported characterization of P. graminis isolates associated with them. In this study, the ribosomal DNA internal transcribed spacer (ITS) regions of P. graminis were examined from 63 wheat samples with apparent virus symptoms obtained from 16 sites within six Chinese provinces. Their associations with soilborne viruses were investigated. Ribotype I (P. graminis f. sp. temperata) and ribotype II (P. graminis f. sp. tepida) were confirmed in winter wheat regions of China for the first time. All 63 wheat root samples were infected with ribotype I of P. graminis and 11 were also infected with ribotype II. There was no obvious association between the ribotypes and infection by either WYMV or CWMV (or double infection). Phylogenetic analysis of the P. graminis ITS1-5.8S-ITS2 sequences revealed that ribotype I in China belongs to previously reported subgroup Ib, whereas ribotype II belongs to IIa. There was considerable sequence variation (pairwise distances from 0.0219 to 0.0319) between Chinese ribotype I isolates of different regions and previously reported ribotype I isolate Ken5 (accession number HE860055.1).

Diagnostic Guide: Wheat Soil-Borne Mosaic

Wheat soil-borne mosaic is caused by the rod-shaped Soil-borne wheat mosaic virus (SBWMV), which is in the genus Furovirus. SBWMV is vectored by Polymyxa graminis. Economic hosts of SBWMV include wheat, barley, and rye. Winter wheat is the most economically important host; spring wheat infections are possible but rare owing to unfavorable soil temperatures slowing virus replication. Yield loss owing to SBWMV infection is variable among season and geographic location.

Distribution of Two Formae Speciales of Polymyxa Graminis in the Czech Republic

It has been shown that two formae speciales of P. graminis, namely f. sp. temperata (ribotype Pg-I) and f. sp. tepida (ribotype Pg-II), are widely distributed throughout temperate areas of Europe. In this study, the presence of both forms of the temperate Polymyxa spp. was identified in soil samples from different locations of the Czech Republic during a survey performed in 2012 and 2013. Based on polymerase chain reaction results, of the total 58 tested samples, 67.2% contained at least one monitored forma specialis. Specifically, P. graminis f. sp. temperata was detected in 48.3% of soil samples, while P. graminis f. sp. tepida was detected in 44.8% of samples. Mixed populations were found in 25.9% of the tested areas. This plasmodiophorid was confirmed not only in crop fields but also in meadows and forests in all explored regions. Our results extend the knowledge on the distribution of both ribotypes of P. graminis and provide the first evidence of f. sp. tepida within the Czech Republic.