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Study of the Barley Yellow Dwarf Virus Species PAV and PAS: Distribution, Virus Quantification and Host Resistance

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Study of the Barley Yellow Dwarf Virus Species PAV and PAS: Distribution, Virus Quantification and Host Resistance Study of the Barley yellow dwarf virus species PAV and PAS: distribution, virus quantification and host resistance Barley Wheat Oat Jana Jarošová, Jana Chrpová, Václav Šíp, Jiban K. Kundu* WDV Crop Research Institute, Prague, the Czech Republic. *E-mail: [email protected], +420-233022402 Background. Barley yellow dwarf (BYD) is one of the most widespread and damaging viral diseases of grasses and cereal crops worldwide. (BYDVs) are a group of related single-stranded RNA viruses assigned to the Luteovirus and Polerovirus genera or unassigned to genera in the family Luteoviridae . Luteoviruses include Barley yellow dwarf virus (BYDV)-PAV, BYDV-PAS, BYDV-MAV, and BYDV-GAV. Poleroviruses include Cereal yellow dwarf virus (CYDV)- RPV. BYDV-SGV, BYDV-RMV and BYDV-GPV are not currently assigned genera. The BYDV species vary strongly in their symptom manifestation, species of efficient aphid vectors, host preferences and in their serological and molecular properties. The BYDV-PAS has been recently separated from BYDV-PAV, the most commonly studied BYDV species. This study was focused on the distribution and characteristics of the PAS. A comparative study of the species PAV and PAS on distribution, virus accumulation and host resistance are described. BYDV quantification. A SYBER green based real time RT-qPCR assay was BYDV distribution. The percentage distribution of individual BYDV species developed for relative BYDV quantification. In the RT-qPCR assay GAPDH, as in comparison with different aspects of samples. 18S rRNA, TUBA and TUBB as normalisation genes of BYDV quantification in barley and oat tissues and a combination of GAPDH, 18S rRNA, TUBB, EIF4A and E1FA for wheat tissues were used. At least three normalization genes are recommended for virus quantification. A: The relative BYDV titre levels in all the oat samples as determined by relative quantification using GAPDH, Ct values distribution of candidate TUBB and 18S rRNA (GAPDH etc), TUBA, EIF4A and EEF1A reference genes in different (EIF4A etc.) and by absolute quantification with cloned species. Values are given as real- standards (absolute). time PCR cycle threshold (Ct) B: The relative BYDV titre levels in the two oat varieties - values. Boxes represent the lower Atego (tolerant) and Saul (susceptible) as determined by A) shows the distribution of BYDV species and upper quartiles with medians; relative quantification using GAPDH, TUBB and 18S rRNA among aphid species (S.a. stands for whiskers illustrate the 10 to 90 (GAPDH), TUBA, EIF4A and EF1A (EIF4A) and by absolute Sitobion avenae; M.d. for Metopolophium percentiles of the samples. All Ct quantification with cloned standards (absolute). The dirhodum; R.m. for Rhopalosiphum values significantly differed absolute numbers were transformed into relative ratios maidis; R.p. for Rhopalosiphum padi, between species (One way so they could be compared. B) shows distribution of individual aphid species among samples, C) shows the effect of plant ANOVA test). being a volunteer plant or not on BYDV species distribution , D) shows the effect of sowing time on BYDV species distribution. Quantitative analysis of BYDV –PAS and BYDV–PAV uptake in winter barley Throughout three years of experimental monitoring of BYDV incidence, PAV resulted in the PAS was the most frequently occurring species infecting cereals, more severe and grasses in the Czech Republic. symptoms and Four species of aphid vectors have been observed in cereal fields: caused a greater Sitobion avenae, Rhopalosiphum padi, Metopolophium dirhodum, R. reduction in maidis. R. maidis and M. dirhodum were recorded as BYDV-PAS vectors, plant height and even though M. dirhodum does not usually transmit BYDV-PAV. yield than PAS. We found a significant difference in virus RNA accumulation between the Comparison of BYDV-PAS and BYDV-PAV in resistance traits PAS and PAV species. In susceptible plants, the virus titre of PAS was greater than that of PAV. Infection with the BYDV-PAV isolate resulted Crop Variety/line VSS PHR (%) GWSR (%) in higher expression of symptoms and also in PAS PAV PAS PAV PAS PAV Range winter WBON116 (Yd2 gene) 1.50 a 2.75 ab 3.9 a 16.0 a 12.6 a 17.2 a 1.7 a greater reduction in plant height and grain barley WBON123 (Yd2 gene) 1.38 a 2.13 a 5.3 a 16.4 a 23.4 ab 29.8 ab 2.3 weight per spike in all tested traits. Doria (Yd2 gene) 3.38 b 3.88 b 0.0 a 21.1 ab 17.4 a 23.8 a 3.0 In general, high levels of resistance were Wysor (Yd2 gene) 1.63 a 2.13 a 9.0 ab 19.6 a 25.4 ab 17.8 a 3.2 Sigra 5.13 c 5.88 c 18.8 bc 35.3 bc 23.9 ab 31.3 ab 5.3 recorded in winter barley breeding lines Finesse 4.88 c 6.25 cd 25.6 c 43.1 c 28.3 abc 44.2 b 6.8 (Wbon116, Wbon123) and varieties (Wysor, Traminer 4.75 c 7.13 de 18.8 bc 40.4 c 36.3 bcd 65.1 c 6.8 Doria) containing the Yd2 gene resistant to Perry 5.88 cd 6.13 cd 21.5 c 43.2 c 48.1 de 62.7 c 7.7 Luran 6.75 de 7.88 ef 17.8 bc 49.4 c 54.8 e 73.2 cd 8.5 both PAV and PAS. Graciosa 7.38 e 8.63 f 26.8 c 68.6 d 44.4 cde 84.0 d 9.7 The winter wheat breeding line PSR3628, a winter PSR 3628 0.75 a 1.17 a 4.3 a 1.8 a 3.0 a 8.0 a 1.0 wheat Svitava 4.13 b 5.63 bcd 6.5 a 11.9 abc 35.4 bcd 48.2 b 3.8 hybrid of wheat and couch grass, was highly Dromos 3.88 b 4.90 b 7.7 a 22.3 bcd 40.0 cd 43.5 b 4.0 resistant to both tested BYDV species. Sparta 4.00 b 4.70 b 12.5 ab 11.0 ab 28.6 bc 51.7 b 4.0 The Svitava, Dromos, Sparta and Banquet Banquet 4.13 b 6.25 d 10.6 a 22.0 bcd 24.0 b 46.0 b 4.3 Roane 4.38 b 6.17 cd 7.4 a 27.8 d 37.1 bcd 57.0 b 6.3 varieties were regarded as moderately Sisson 5.00 bc 6.58 d 11.4 ab 19.9 bcd 38.4 bcd 49.2 b 6.5 resistant in our test. Meritto 4.38 b 5.20 bc 19.3 b 28.6 d 45.3 d 56.9 b 7.5 We noted that the use of two species, Vlada 6.38 cd 6.30 d 11.3 ab 26.1 cd 41.1 d 55.8 b 7.5 SG-S 2703 7.25 d 7.92 e 31.8 c 47.8 e 64.1 e 76.9 c 10.0 namely the varieties carrying the Yd2 gene in Average 4.44 a 5.40 b 13.5 a 27.6 b 33.4 a 46.7 b barley and the line PSR 3628 in wheat, Average VSS (visual symptom score recorded on a scale of 0 to 9, where 0 represents no symptoms), PHR (plant allowed the detection of the sources of height reduction) and GWSR (reduction of grain weight per spike) in barley and wheat varieties (lines) after resistance in all the tested traits. inoculation with BYDV-PAS and BYDV-PAV. Means in the columns followed by the same letter are not significantly different from each other at P=0.05 of the LSD test. Jarošová J, Kundu JK, 2010. Validation of reference genes as internal control for studying viral infections in cereals by quantitative real-time RT-PCR. BMC Plant Biology 10, 146. WSMV Reference: Jarošová et al. 2011. A comparative study of the Barley yellow dwarf virus species PAV and PAS: distribution, virus accumulation and host resistance. Plant Pathology (accepted) C R O P R E S EA R C H I NS T I T U T E, P R A G U E, THE C Z E C H R E P U B L I C The work was funded by projects no. QH81269 Mze ČR and ME10022 MŚMT ČR.
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