Characterization and Mapping of retr04, retr05 and retr06 Broad-Spectrum Resistances to Turnip Mosaic Virus in Brassica juncea, and the Development of Robust Methods for Utilizing Recalcitrant Genotyping Data

Turnip mosaic virus (TuMV) induces disease in susceptible hosts, notably impacting cultivation of important crop species of the Brassica genus. Few effective plant viral disease management strategies exist with the majority of current approaches aiming to mitigate the virus indirectly through control of aphid vector species. Multiple sources of genetic resistance to TuMV have been identified previously, although the majority are strain-specific and have not been exploited commercially. Here, two Brassica juncea lines (TWBJ14 and TWBJ20) with resistance against important TuMV isolates (UK 1, vVIR24, CDN 1, and GBR 6) representing the most prevalent pathotypes of TuMV (1, 3, 4, and 4, respectively) and known to overcome other sources of resistance, have been identified and characterized. Genetic inheritance of both resistances was determined to be based on a recessive two-gene model. Using both single nucleotide polymorphism (SNP) array and genotyping by sequencing (GBS) methods, quantitative trait loci (QTL) analyses were performed using first backcross (BC1) genetic mapping populations segregating for TuMV resistance. Pairs of statistically significant TuMV resistance-associated QTLs with additive interactive effects were identified on chromosomes A03 and A06 for both TWBJ14 and TWBJ20 material. Complementation testing between these B. juncea lines indicated that one resistance-linked locus was shared. Following established resistance gene nomenclature for recessive TuMV resistance genes, these new resistance-associated loci have been termed retr04 (chromosome A06, TWBJ14, and TWBJ20), retr05 (A03, TWBJ14), and retr06 (A03, TWBJ20). Genotyping by sequencing data investigated in parallel to robust SNP array data was highly suboptimal, with informative data not established for key BC1 parental samples. This necessitated careful consideration and the development of new methods for processing compromised data. Using reductive screening of potential markers according to allelic variation and the recombination observed across BC1 samples genotyped, compromised GBS data was rendered functional with near-equivalent QTL outputs to the SNP array data. The reductive screening strategy employed here offers an alternative to methods relying upon imputation or artificial correction of genotypic data and may prove effective for similar biparental QTL mapping studies.

Biological and Molecular Properties of a Turnip mosaic virus (TuMV) Strain that Breaks TuMV Resistances in Brassica napus

A new resistance-breaking strain of Turnip mosaic virus (TuMV) overcomes TuMV resistance genes that currently suppress spread of this virus in Brassica napus crops in the Liverpool Plains region of eastern Australia. Isolates 12.1 and 12.5 of this strain and three other isolates in TuMV pathotypes 1 (NSW-2), 7 (NSW-1), and 8 (WA-Ap1) were inoculated to plants of 19 B. napus cultivars and one breeding line. All plants of these cultivars and the breeding line proved susceptible to 12.1 and 12.5 but developed only resistance phenotypes with WA-Ap1 or mostly resistance phenotypes with NSW-1 and NSW-2. Five different TuMV resistance phenotypes occurred either alone or segregating in different combinations. When these five isolates were inoculated to plants of nine other crop or wild Brassicaceae spp. and four indicator hosts in other families, 12.1 and 12.5 resembled the other three in inducing TuMV resistance phenotypes in some Brassicaceae spp. but not others, and by inducing extreme resistance phenotypes in all inoculated plants of B. oleracea var. botrytis and Raphanus sativus. Therefore, the overall resistance-breaking properties of 12.1 and 12.5 were restricted to B. napus. When isolates 12.1, 12.5, and WA-Ap1 and additional Australian isolate WA-EP1 were sequenced and complete genomes of each compared, 12.1 and 12.5 grouped separately from the other 2 and from all 23 Australian isolates with complete genomes sequenced previously. In addition, there was evidence for at least six separate TuMV introductions to Australia. Spread of this B. napus resistance-breaking strain poses a significant threat to the B. napus oilseed industry. Breeding B. napus cultivars with resistance to this strain constitutes a critical priority for B. napus breeding programs in Australia and elsewhere.

Inheritance and development of EST-SSR marker associated with turnip mosaic virus resistance in Chinese cabbage

Li, Q., Tong, H., Zhang, Z., Zhao, Z. and Song, X. 2011. Inheritance and development of EST-SSR marker associated with turnip mosaic virus resistance in Chinese cabbage. Can. J. Plant Sci. 91: 707–715. Turnip mosaic virus (TuMV) is one of the major pathogens infecting Brassica crops (including Chinese cabbage), and often causes serious reductions in yield and quality. Breeding for resistant cultivars is complicated by the existence of numerous TuMV strains and isolates. The objective of this research was to determine the mode of inheritance and to develop molecular markers associated with TuMV resistance in Chinese cabbage. F1 and F2 populations were developed from the cross between 71-36-2 (susceptible) and 73 (resistant) lines and mechanically inoculated with TuMV-C4. Inheritance analysis by visual scoring and enzyme-linked immunosorbent assay (ELISA) indicated that resistance to TuMV-C4 in this cross was controlled by one recessive gene. A total of 132 EST-SSR primers were designed from EST sequences available in public databases. Seven primers detected polymorphism between parental genotypes. Marker HCC259 was associated with the TuMV resistance Ph-retr02. The distance between the marker and the TuMV resistance gene retr02 was 3.8 cM. This is the first co-dominant marker linked to the TuMV resistance gene with a distance less than 5.0 cM in Chinese cabbage. This marker was suitable for TuMV-C4 resistance screening in progenies from the cross between lines 73 (resistant) and susceptible line, 71-36-2. The usefulness of this marker was validated in 21 additional resistant and susceptible lines. This marker has the potential to simplify and accelerate breeding Chinese cabbage cultivars resistant to TuMV-C4.

Evaluation of resistance to Turnip mosaic virus in Australian Brassica napus genotypes

Forty-three Australian cultivars or breeding lines of Brassica napus (canola, oilseed rape) and 2 cultivars of Brassica juncea (mustard) were inoculated with infective sap containing isolate WA-Ap of Turnip mosaic virus (TuMV), which belongs to TuMV pathotype 8. The types of reactions obtained were: necrotic spots in inoculated leaves without systemic infection (RN), chlorotic blotches in inoculated leaves without systemic infection (R), and chlorotic blotches in inoculated leaves accompanied by systemic infection that consisted of either necrotic spots (+N) or chlorotic blotches (+). The RN and +N reactions are consistent with those expected in the presence of 4 strain-specific TuMV resistance genes TuRB01 (+N response), TuRB03 (+N response) and TuRB04 with TuRB05 (RN), with + indicating a susceptible response. However, which resistance gene corresponds to the R response is unclear. The RN (TuRB04 with TuRB05) type of response was the commonest. Only one genotype lacked any TuMV resistance, and segregation for more than one different type of resistance response occurred within 22 genotypes and some segregated for resistance and susceptibility. Some genotypes segregated for all 3 types of resistance response found. The reaction of 2 plants of cv. Rivette was atypical as they developed both necrotic spots in inoculated leaves and systemic chlorotic spots. Since breeding for TuMV resistance is not undertaken in Australia, these results indicate frequent but inadvertent crossing with parental lines carrying TuMV resistance. Widespread occurrence of TuMV resistance genes and the possibility that many Australian TuMV isolates may not be well adapted to B. napus may explain the low incidence of this virus found in Australian B. napus crops.