Barley Yellow Dwarf occurrence and resistance stability in wheat
Transcription
Barley Yellow Dwarf occurrence and resistance stability in wheat
International Journal of Farming and Allied Sciences Available online at www.ijfas.com ©2015 IJFAS Journal-2015-4-3/207-214/ 31 March, 2015 ISSN 2322-4134 ©2015 IJFAS Barley Yellow Dwarf occurrence and resistance stability in wheat varieties of Pakistan Muhammad Ibrahim* and Syed Jawad Ahmad Shah Nuclear Institute for Food and Agriculture (NIFA), Tarnab, Peshawar Corresponding author: Muhammad Ibrahim ABSTRACT: Barley Yellow Dwarf Virus (BYDV) occurrence and resistance stability were studied in a set of 17 varieties along with a susceptible control (Fakhr-e- Sarhad) over four years (2005-08) at Peshawar, Pakistan. Consistent and uniform BYDV symptoms were observed over study years. Enzyme Linked Immunosorbent Assay (ELISA) of 2008 experiment indicated the presence of BYDV-PAV. Phenotypic assessments with regard to incidence (infected tillers/plot) and percent BYDV severity varied among varieties. Maximum incidence (120) and severity (80%) was recorded during 2006 cropping cycle. BYDV incidence and severity was positively correlated (r = 0.935) over four years. Varieties and years remained significantly different while variety-year interaction was non-significant (P<0.05). Cvs.Punjab-96, Suleman96, Bakhtawar-93, Faisalabad-83 and Bahawalpur-95 displayed stability in severity levels and also expressed a phenotypic marker of Leaf Tip Necrosis (LTN) more frequently over four year experimentations which is linked with BYDV tolerance gene (Bdv1). Comparatively stable and tolerant varieties identified through level of BYDV infection could be improved further by incorporating Bdv2 and Bdv3 genes to manage this serious problem in Khyber Pakhtunkhwa province of Pakistan. Keywords: Barley, stability, wheat, Pakistan INTRODUCTION Barley Yellow Dwarf (BYDV) is an economically important viral disease of cereals in the world (Singh., 1993). Occurrence of BYDV in developing countries is well documented (Lister and Ranieri., 1995). In Pakistan, BYDV like symptoms were known since 1964 in the border areas of Pakistan and Afghanistan where it was confirmed for the first time in the country during 1987 (Aslam and Ahmad., 1987). Prevalence of BYDV symptoms with varying level of chlorosis on commercial wheat varieties were recorded (Ibrahim , 2009, unpublished data) and is presented in Fig.1. BYDV has gradually attained an important position among wheat diseases (Khalid., 1999) and reported from Khyber Pakhtonkhaw Province of Pakistan (Bashir., 1997; Siddiqui., 2011) and Northern Punjab (Bashir., 1997). BYD losses in wheat ranged from 20 to 30% (Pike, 1990) while in severe epidemic conditions, reduction in yield could reach up to 86% (Gill., 1980). BYDVs are members of the Luteoviridae family of plant viruses, including BYDV-PAV, BYDVMAV, BYDV-RMV, BYDV-PAS, BYDV-GPV and BYDV-SGV and Cereal yellow dwarf virus (CYDV)-RPV and CYDVRPS. BYDVs and CYDVs are phloem limited and persistently transmitted by at least 25 aphid species (Halbert and Voegtlin., 1995). Previously five BYDV strains i.e. PAV, MAV, RPV, RMV and SGV (Bashir., 1997) which are now known as independent viruses (D’Arcy and Domier., 2005) are reported from the region alongwith their aphid vectors (Anonymous., 1998; Khan., 2006). Intl J Farm & Alli Sci. Vol., 4 (3): 207-214, 2015 Figure 1. Locations in Khyber Pakhtonkhaw where BYDV symptoms occur regularly under natural conditions High level of BYDV resistance is not available in wheat while tolerance which is known since 1950s (Burnett., 1995) is still one of the most promising mode of plant protection to limit yield losses in developing countries (Ginkel and Henry., 2002). Almost all Pakistani wheat cultivars grown in Northern Punjab and KPK were susceptible to BYDV (Khalid., 1999). Adult plant leaf tip necrosis could be used as a phenotypic marker for the presence of Bdv1 gene (Singh., 1993) responsible for BYDV tolerant in wheat. In characterizing BYDV tolerance, environmental conditions and infection level are important to be considered (Comeau and Jedlinski., 1990) because a cultivar which is considered to be tolerant in one environment may be less tolerant in an other environment, thus complicating selection for tolerance. Singh and Chaudhary (1977) reported that performance stability is one of the most desirable traits of a genotype to be released as variety for general cultivation. Keeping in view the national importance of BYDV, varietal screening efforts for the identification of BYD tolerant wheat varieties through phenotypic marker with low symptoms severity, disease identification through DAS-ELISA and stability across different growing seasons was carried out and findings are reported here. MATERIALS AND METHODS Site selection and experimental detail Nuclear Institute for Food and Agriculture (NIFA), Peshawar was selected due to its unique geographical position (34° 0'N, 71°42'E, elevation: 304 m), soil fertility, irrigated conditions and BYD hotspot where it occur regularly in the region (Aslam and Ahmad., 1987; Bashir., 1997; Siddiqui., 2011). Seventeen commercial wheat varieties along with a susceptible check Fakhr e Sarhad (Table 3) were evaluated over a period of four years in a Randomized Complete Block Design (RCB) with three replications where experimental unit consisted of 3.6 m 2 plot having 4 rows each 3 m length with row to row distance of 30 cm. Disease Assessment and leaf tip necrosis Scoring of BYD incidence (number of infected tillers/plot) and severity (percentage of foliage with symptoms) was carried out according to 0-9 scale as described by Singh., (1993) in early March each year due to the expression of BYD symptoms and peak activity period of its vectors (i.e. Rhopalosiphum & Sitobion spp) in the area (Anonymous, 1997-98; Ibrahim., 2006) with the help of pictorial guide for identification of field wheat diseases and pests (Prescott., 1986). Presence or absence of leaf tip necrosis (LTN) on flag leaf of each variety was recorded over four years (200508)/12 replications as mentioned by (Shah., 2010; Shah., 2011). ELISA Analysis Double Antibody Sandwich-Enzyme Linked Immunosorbent Assay (DAS-ELISA) was used as described by Clark and Adams (1977), with little modifications in the incubation times, antibody concentration, and sample dilution. The volume of sample and incubating solutions used was 100 μl in all steps, as described by Ayala., (2001). Polystyrene microtiter plates were incubated at 37°C for 3 h with a coating of polyclonal antibodies (1 ng/μl) directed against BYDPAV (provided by Agdia Inc.). Plant sap (1:10, in 0.1 M phosphate buffer pH 7.0) was incubated for 3 h at 37°C. Controls used consisted of phosphate buffer (blank) and infected PAV plant extract (leaf material containing BYDVPAV serotype from Agdia Inc.). Alkaline phosphatase-labeled antibodies (1:1000), anti-PAV, were incubated 208 Intl J Farm & Alli Sci. Vol., 4 (3): 207-214, 2015 overnight at 4°C; P-nitrophenyl phosphate substrate (Sigma) was then added. Assay was by visual observation of the yellow nitrophenolate, based on a “plus” and “minus” classification (Fargette., 1982). Statistical Analysis Symptoms severity data of BYD was analyzed using Analysis of Variance (ANOVA) as reported by Steel and Torrie (1960). Means were separated for comparison through LSD test by following the method of Steel and Torrie (1980) in MSTATC software (Michigan state University, 1991). Stability parameters were worked out as suggested by Eberhart and Russel (1966) using SPSS. Percent BYD severity was expressed through Box plots (Tukey., 1990) for % BYD foliage disease severity were also produced through SPSS for an overall descriptive presentation of the data. RESULTS AND DISCUSSION BYDV Occurrence and leaf tip necrosis BYDV occurrence varied over seasons in the tested varieties. Disease pressure as assessed by infected tillers/plot of susceptible check (Fig.3) and %severity of commercial varieties (Fig.4) were highest in 2006 which were followed by 2007, 2008 and 2005, respectively. Several factors involving virus, vectors, host and environment (Ginkel and Henry., 2002; Harrington., 2002; Hoffman and Kolb., 1998) may have contributed to this observed variation. Highest incidence (120) was recorded on susceptible check during 2006, followed by 2007 (55), 2008 (35), and 2005 (07). Mean number of infected tillers/plot were highest (i.e.38) during 2006 which was followed by 23, 06 and 02 in 2007, 2008 and 2005 respectively (Fig. 2). Similarly, symptoms severity varied in different seasons where the data distribution was not normal (Fig. 4). Maximum variability in BYDV severity was observed in 2008 which was followed by 2006, 2007, and 2005 respectively (Fig 4). BYDV severity was normally distributed in 2007 while in each 2005 and 2006 it was positively skewed whereas in 2008 it was negatively skewed. Highest median value of 35 was followed by 30, 18 and 04 in 2006, 2007, 2008 and 2005 seasons, respectively. This fluctuation in BYD severity could be attributed to (i) high mean temperature in the months of January to April (Table 2) during season 2006 (16.81°C) as compare to 2007 (15.81°C), 2008 (14.83°C) and 2005 (14.56°C) in the peak activity period of aphid vectors resulted in BYDV spread through vectors in the region (ii) low mean precipitation (121 mm) coupled with low % relative humidity (%56.88) during peak aphid vector movement and BYDV transmission activity in 2006 enhanced disease development followed by 2007, 2008, and least during 2005 with precipitation ranged from 194.8 to 285.9 mm and % relative humidity up to 64.8%. (iii) high frequencies (50) of temperature range i.e. 16-25°C during 2006 peak disease activity period enhanced BYD symptoms expression compared to 2007, 2008, and 2005 with frequency ranged from 16 to 33. Environmental factors are reported to contribute in the aphid movement and BYDV transmission (Smyrnioudis., 2001; Smyrnioudis., 2000). Cooler temperature (15 to 18°C) is reported to favor symptoms expression and attract vectors to disease plants (D’ Arcy and Domier., 2000). Drought/low rainfall with is reported to increase aphid vector movement and BYDV spread among host plants (Bailey., 1995). High temperature (30°C) is reported to increase transmission abilities of inefficient vectors (D’ Arcy and Domier., 2000). The uneven distribution of viruliferous aphid vectors and spreading virus from infection foci under natural field conditions might have caused variation in BYDV incidence and severity level among wheat varieties both within and between seasons. Such complications that a wheat variety appeared tolerant to BYDV in one season showed less tolerance in an other season or environment was also reported by researchers (Comeau and Jedlinski., 1990). 209 Intl J Farm & Alli Sci. Vol., 4 (3): 207-214, 2015 Table 2. Weather data recorded during 2005-08 wheat growing seasons Month s Weather Data 2005 Mean Tem p. (°C) Mean % Relativ e Humidit y Jan 9.1 73.55 Feb 9.7 75.2 March 16.95 66.55 April 22.5 43.9 Mean 14.56 64.8 50.46 Total 194.8 Total Precipitati on (mm) Weather Data 2006 Weather Data 2007 Weather Data 2008 Mean Tem p. (°C) Mean Tem p. (°C) Mean Tem p. (°C) Mean % Relativ e Humidit y 64.4 9.95 68.4 109.9 16.4 58.2 97.1 17.6 59.02 14.5 23.3 41.9 Total 285.9 Mean 16.81 Total Precipitati on (mm) 56.4 15.2 32.4 17 56.88 Mean Total % Precipitati Relativ on e (mm) Humidit y 10.3 57.35 0 12.25 69.75 177 15.9 62.65 82.4 24.8 45.7 20.4 Total 121 Mean 15.81 58.86 Mean Total % Precipitati Relativ on e (mm) Humidit y 7.65 54.7 57.2 10.75 51.8 21.2 19.6 47.25 03 21.35 48.1 113.4 Total 279.8 Mean 14.83 Figure 2. Mean BYD incidence on Commercial Varieties during 2005-08 Figure 3. Mean BYD incidence on commercial wheat varieties during 2005-2008 Figure 4. Box plot depicting BYD % severity on commercial varieties during 2005-08 at NIFA, Peshawar, Pakistan 210 Intl J Farm & Alli Sci. Vol., 4 (3): 207-214, 2015 Varieties were classified on the bases of LTN occurrence into three groups (Table 3). Group 1 varieties possessed LTN symptoms which were less frequent in group 2 but not observed in group 3. In the present study, 12 replications (3 each year) were used to evaluate the presence or absence of LTN symptoms. Varieties in group 1(Faisalabad-83, Faisalabad-85, Suleman-96, Bahawalpur-95, Punjab-96 and Bakhtawar-93) expressed LTN phenotypes in eight or more of the replications and were therefore considered positive for LTN. Relative BYDV severities of these six varieties of group 1 were 42-70% less infected than the susceptible check. Varieties of group 2 (Inqilab-91, Zardana-89 and Rohtas-90) expressed LTN less frequently and inconsistently in four or fewer of the replications and was therefore considered absent while BYDV severity was reduced from 52-64% of the susceptible check. LTN was not displayed by varieties of group 3 (Sarsabz, Shaheen-94, Kaghan-93, Watan-94, Soughat-90, Shahkar-95, Kohsar-95, Local White and Fakhar-e-Sarhad) which were able to reduce BYDV severity from 45-73% of the susceptible check. Bdv1 gene was first identified in Anza wheat variety as BYDV tolerant gene which confers slow yellowing (Singh , 1993). Leaf tip necrosis (LTN), a morphological trait reported of showing complete linkage or pleiotropism with Bdv1, Yr18 and Lr34 genes (Singh., 1992a, b) which was suggested to use as phenotypic marker to identify wheat varieties carrying the gene. Adult leaf tip necrosis, a phenotypic marker was observed at adult plant stage. In the present studies high LTN frequencies were observed in group-1 varieties and were considered positive, less frequent in group-2 while this trait was completely absent in group-3 varieties. Varieties which expressed LTN phenotypes ≥ 8 are considered carrying Bdv1 gene and this gene is believed to occur very frequently in CIMMYT wheats (Singh., 1993). Similar findings regarding LTN phenotypes and their molecular confirmation of Yr18 presence/absence were reported on the same commercial varieties by Shah (2010). The phenotypic marker (LTN) for Bdv1 gene was not 100% expressed in the LTN positive varieties across different replications and is reported to be obscured by genetic background (Singh., 1999) and variable influences of environment (Dyck., 1991). Watan-94 and Shahkar-95 of group-3 developed less foliar chlorosis but could not express LTN. There may be the possibility that varieties that could not expressed LTN may even carry Bdv1 as Singh (1999) reported that varieties which don’t express LTN may still carry Lr34. Table 3. Pedigrees, BYDV severity (%), leaf tip necrosis (LTN), stability parameters of severity trait and indexing through DASELISA of tested wheat varieties Varieties Pedigree Mean BYDV Severity Group-1(Possess LTN) Faisalabad-83 FURY/KAL/BB Faisalabad-85 MAYA/MON//KVZ/TRM Suleman-96 F6.74/BUN//SIS/3/VEE#7 Bahawalpur95 Punjab-96 AVRORA/UP301//GALLO/SUPER-X/3/(SIB) PEWEE/4/MAIPO(SIB)/ (SIB)MAYA-74//PEWEE SA 42 *2/4CC/INIA//BB/3/ INIA/HD832 Bakhtawar-93 JUP/BJYG//URES Group-2(LTN less frequent) Inqilab-91 WL 711/CROWS’ Zardana-89 CNO S/8156 TOB 66 CNO6-PVN Rohtas-90 INIA F 66/ A.DISTCHUM//INIA66/3/GEN Group-3(LTN not observed) Sarsabz P1/FRND//MXP/3/P1/M20/79 Shaheen-94 MLT “S” Kaghan-93 TTR/JUN 31.36 Watan-94 Lu26/HD2179 26.6 Soughat-90 Pavon Mutant-3 31.51 35.6 Resistance class LTN Stability Parameters bi S2d ELISA 32.00 (58)* 16.00 (30) 26.00 (47) 24.00 (44) 17.00 (32) 23.00 (41) LR** 8 1.03 -1.09 ++ R 7 0.53 0.54 + LR 9 1.19 4.59 + LR 10 0.80 -1.36 ++ R 10 1.00 5.25 ++ LR 9 1.47 - 0.99 + 20.00 (36) 27.00 (48) 27.00 (48) R 3 1.21 - 0.13 + LR 4 0.96 -1.39 ++ LR 3 1.20 6.59 + 15.00 (28) 20.00 (36) 17.00 (31) 15.00 (27) 17.00 (32) R Absent 1.06 - 0.32 ++ R Absent 1.31 3.69 + R Absent 1.09 - 0.57 + R Absent 1.03 3.40 ++ R Absent 0.43 - 1.31 ++ 211 Intl J Farm & Alli Sci. Vol., 4 (3): 207-214, 2015 Shahkar-95 WL 711//F3.71/TRM Kohsar-95 PSN/BOW Local White Land race FakhrSarhad e- 35.45 35.76 54.85 PFAU “S”/SERI/BOW “S” 100.00 20.00 (35) 20.00 (35) 30.00 (55) 55.00 (100) R Absent 0.79 5.09 ++ R Absent 1.60 1.51 ++ LR Absent 0.64 - 1.22 ++ I Absent 0.63 - 0.98 ++ BYD Symptoms Stability and ELISA analysis Analysis of variance for percent BYDV severity showed highly significant (P<0.01) differences for both genotypes and years while genotypes x years interaction remained non-significant (Table 1). BYDV incidence and severity was positively correlated (r = 0.935) over four years. Performance of individual varieties with regard to comparative stability of BYDV severity is presented in Table 3. Stability parameter, coefficient of regression (bi) value deviated from unity (bi=1) and ranged from 0.43 to 1.60 in the tested varieties. Based on mean BYDV severity over years, tested varieties were classified into resistant (1-20% severity), low resistance (21-50% severity) and intermediate (51-70% severity). Five varieties (Faisalabad-83, Punjab-96, Sarsabz, Kaghan-93 and Watan-94) were found stable for this trait over four years of study as their bi value was 1 or near to 1. Apart from the check variety (Fakhr-eSarhad), all remaining genotypes fell into resistant or low resistance classes but remained highly unstable for severity trait over four years. Wheat commercial varieties were assessed against BYD natural infection across different seasons to know disease symptoms stability (tolerance) on the tested varieties. BYD severity fluctuated between years and greatly influenced by environment which is important for its tolerance characterization through foliar chlorosis (Comeau and Jedlinski., 1990). Faisalabad-83, Sarsabz, Shaheen-94, Suleman-96, Kaghan-93, Rohtas90, Inqilab-91, Kohsar-93, Watan-94 and Bakhtawar-93 showed regression of coefficient value greater than unity (b>1) and thus produced symptoms under BYD favorable environmental conditions. While Bahawalpur-95, Zardana89, Faisalabad-85, Soughat, Shahkar-95, Local white and susceptible check Fakhr e sarhad with coefficient of regression value less than unity (b<1) therefore, produced symptoms under less favorable conditions (Eberhart and Russel., 1966). Furthermore, Punjab-96 remained stable (b=1) with coefficient of regression value equal to unity (Eberhart and Russel, 1966) and expressed similar disease symptoms at different environmental conditions. BYDVPAV was detected through DAS-ELISA in the infected leaves of wheat exhibiting typical yellowing symptoms of BYD. BYD-PAV infection was dominant due to abundance of their aphid vectors i.e. Rhopalosiphum padi and Sitobion avenae and was frequently detected during peak aphid BYD activity period from the region in earlier studies (Bashir et at., 1997; Ibrahim., 2006; Suddiqui., 2011). Table 1. Pooled analysis of variance of BYD % severity during 2004-2008 at NIFA, Peshawar Pakistan Source Replications Genotypes (G) Years (Y) GxY Error Degrees of Freedom 2 17 3 51 142 Sum of Squares 1.349 223.060 650.684 309.446 623.517 Mean Square 0.674 13.121 216.895 6.068 4.391 F value 0.1536 2.9882 49.3957 1.3818 Prob 0.0002** 0.0001** 0.0512 Coefficient of Variation: 19.20% Tested varieties were indexed for the infection of Barley Yellow Dwarf Virus (BYDV) through DAS-ELISA during 2007-08. The overnight incubation of conjugate with antigen gave lower background values and allowed easier visual discrimination between positive and negative reactions. Assay was done by visual observation of the yellow nitrophenolate, based on “+” and “++” as presented in Table 3. Presence of BYDV-PAV was confirmed in the infected leaves of all tested varieties, of which ten (Faisalabad-83, Sarsabz, Bahawalpur-95, Zardana, Kohsar-93, Watan-94, Shahkar-95, Soughat, Local white and Fakhr e Sarhad) had strong positive reaction. In our environment varieties possessing Bdv1 gene identified through phenotypic marker did not show tolerance against BYD. LTN positive varieties showed high level of disease severity as compared to varieties with less frequent LTN or with no LTN. Thus the hypothesis that common wheat possessing Lr34/Yr18/Bdv1 increases tolerance against BYD was not confirmed. Similar results regarding the Bdv1 tolerance was reported by Ayala. (2002) and Slamova., (2009) that the effect of the Bdv1 gene is not as effective as expected in the previous study by Singh. (1993). As the Bdv1 gene does not provide true resistance, therefore breeders identified Thinopyrum intermedium which is a wild grass and show a very high level of resistance possessing Bdv2 and Bdv3genes. These genes present 212 Intl J Farm & Alli Sci. Vol., 4 (3): 207-214, 2015 in many translocated wheat lines conferring different levels of tolerance to BYD infection should be incorporated in national breeding program of the country to instill the crop with an inbuilt ability to avoid losses due to this serious disease. REFERENCES Anonymous. 1998. Annual Report of NIFA, Peshawar. p.139. Aslam M and Ahmad I. 1987. Barley Yellow Dwarf in Pakistan. World Perspectives on Barley Yellow Dwarf, Proceedings of the International Workshop July 6-11, Udine, Italy. Ayala L, Henry M, González-de-León D, Ginkel M V, Mujeeb-Kazi A and Khairallah M. 2001. A diagnostic molecular marker allowing the study of Th. intermedium-derived resistance to BYDV in bread wheat segregating populations. Theor Appl Genet 102: 942–949. Ayala L, Henry M, Gingel MV, Singh R, Keller B and Khairallah M. 2002. Identification of QTLs for BYDV tolerance in bread wheat. In: Euphytica, 128: 249–259. Bailey SM, Irwin ME, Kampmeier GE, Eastman CE and Hewings AD. 1995. Physical and biological perturbations: their effect on the movement of apterous Rhopalosiphum padi (Homoptera: Aphididae) and localized spread of barley yellow dwarf virus. Environ. Entomol. 24, 24-33. Bashir M, Bertschinger L, Kissana NS,. Mujahid MY and Hashmi NI. 1997. Detection of five Barley yellow dwarf Luteovirus serotypes in Pakistan. Rachis: 16: 47-49. Burnett PA, Comeau A and Qualset CO. 1995. Host plant tolerance or resistance for control of barley yellow dwarf. Pages 321-343 in: Barley Yellow Dwarf: 40 Years of Progress. C. J. D’Arcy and P. A. Burnett, ed. American Phytopathological Society, St. Paul, MN. Clark MF and Adams AN. 1977. Charactaristics of the microplate method of enzyme- linked immunosorbent assay for the detection of plant viruses. J. Gen. Virol. 34:475-483. Comeau A and Jedlinski H. 1990. Successful breeding for barley yellow dwarf resistance or tolerance: a systematic approach related to other agronomic characteristics. Pages 441-451 in: World Perspectives on Barley Yellow Dwarf. P. A. Burnett, ed. CIMMYT, Mexico, D.F., Mexico. D’Arcy CJ and Domier LL. 2005. Family Luteoviridae. In: Fauquet C.M., Mayo M.A., Maniloff J., Desselberger U., Ball L.A. (eds). Virus Taxonomy. 8th Report of the International Committee on Taxonomy of Viruses, pp. 891-900. Elsevier/Academic Press, London, UK. D’Arcy CJ and Domier LL. 2000 Barley yellow dwarf. The Plant Health Instructor. DOI:10.1094/PHI-I-2000-1103-01Updated2005. Dyck PL. 1991. Genetics of adult plant leaf rust resistance in ‘Chinees Spring’ and Sturdy wheats. Crop Sci. 31: 309-311. Eberhart SA and Russel WA. 1966. Stability parameters for comparing varieties. Crop Sci. 6: 36-40. Fargette DR, Lister M and Hood EL. 1982. Grasses as a Reservoir of Barley Yellow Dwarf Virus in Indiana. Plant Dis. 66: 1041-1045. Gill CC. 1980. Assessment of losses on spring wheat naturally infected with barley yellow dwarf virus. Plant Dis. 64: 197-203. Ginkal MV and Henry M. 2002. Breeding for BYDV Tolerance/Resistance in CIMMYT Bread Wheats Targeted to Developing Countries. Barley Yellow Dwarf Disease: Resent Advances and Future Strategies. Proceeding of an International Symposium September1-5. CIMMYT, Mexico.93-96. Hoffman TK and Kolb FL. 1998. Effects of barley yellow dwarf virus on yield and yield components of drilled winter wheat. Plant Dis. 82:620-624. Halbert S and Voegtlin D. 1995. Biology and taxonomy of vectors of barley yellow dwarf viruses. Pages 217-258 in: Barley Yellow Dwarf: 40 Years of Progress. C. J. D’Arcy and P. A. Burnett, eds. American Phytopathological Society, St. Paul, MN. Harrington R. 2002. BYDV: The Heat Is On. Barley Yellow Dwarf Disease: Recent Advances and Future Strategies. Page 34-39. Proceeding of the International Symposium, September 1-5, (Henry and McNab ed.) CIMMYT Mexico. Ibrahim M, Shah SJA, Muhammad T, Hussain S and Waseemullah AS. 2006. Field Assessment of candidate and commercial varieties naturally infected with barley yellow dwarf virus (BYDV) in Peshawar, Pakistan. Proceeding of International wheat seminar. 292-295. Khalid S. 1999. Research on Plant Viral Diseases in Pakistan. Bibliography and Abstracts. pp. 208. Khan SA, Ullah F, Hussain N, Saljoqi AUR, Hayat Y and Sattar S. 2006. Distribution pattern of the cereal aphids in the wheat growing areas of the North West Frontier Province (NWFP) of Pakistan. Sarhad J. Agric. 22: 655-659. Lowels AJ, Tatchell GM, Harrington R and Clark SJ. 1996. The effect of temperature and inoculation access period on the transmission of Barley yellow dwarf virus by Rhopalosiphum padi (L.) and Sitobion avenae (F.). Ann. Appl. Biol. 128: 45-53. Lister RM and Ranieri R. 1995. Distribution ands economic importance of barley yellow dwarf. In: Barley Yellow Dwarf, 40 Years of Progress. D’Arcy, C.J. and Burnett, P.A. (eds.). APS, St. Paul, Minnesota, USA. pp. 29-53. Prescott JM, Burnett PA and Saari EE. 1986. Wheat diseases and Pests. A guide to field identification CIMMYT Mexico. p.135. Shah SJA, Imtiaz M and Hussain S. 2010. Phenotypic and molecular characterization of wheat for slow rusting resistance against Puccinia striiformis Westend. f.sp. tritici J Phytopathol. 158:393–402. Shah SJA, Hussain S, Ahmad M, Farhatullah, Ali I and Ibrahim M. 2011.Using leaf tip necrosis as a phenotypic marker to predict the presence of durable rust resistance gene pair Lr34/Yr18 in wheat. J Gen Plant Pathol. 77:174–177. Siddiqui NN, Ilyas M, Mansoor S, Azhar A and Saeed M. 2011. Cloning and phylogenetic analysis of coat protein of barley yellow dwarf virus isolates from different regions of Pakistan. J Phytopathol. 160:13–18. Singh RP. 1992 a. Genetic association of leaf rust resistance gene Lr34 with adult plant to stripe rust in bread wheat. Phytopathol. 82: 835-838. Singh RP. 1992b. Association between Lr 34 for leaf rust resistance and leaf tip necrosis in wheat. Crop Sci. 32: 874-878. Singh RP, Burnett PA, Albarran M and Rajaram S. 1993. Bdv1: a gene for tolerance to barley yellow dwarf virus in bread wheats. Crop Sci. 33:231–234. Singh RP, Chen WQ and He ZH.1999. Leaf rust resistance of spring, facultative, and winter wheat cultivars from China. Plant Dis. 83:644–651. Singh RK and Chaudhary BD. 1977. Biometrical Methods in Quantitative Genetic Analysis. Kalyani Publishers, New Delhi, India. Smyrnioudis NI, Harrington R, Hall M, Katis N and Clark SJ. 2001. The effect of temperature on variation in transmission of a BYDVPAV- like isolate by clones of Rhopalosiphum padi and Sitobion avenae. Euro. J. Plant Pathology.107: 167-173. Smyrnioudis IN, Harrington R, Katis N and Clark SJ. 2000. The effect of drought stress and temperature on spread of barley yellow dwarf virus (BYDV). Agric. and For. Entomol. 3:161-166. Slamova L, Chorpova J, Vejl P and Veskrna O. 2009. Evaluation of genetic sources of tolerance of common wheat against BYDV and CYDV. Agriculture (Poľnohospodástvo): 55, 33–41. 213 Intl J Farm & Alli Sci. Vol., 4 (3): 207-214, 2015 Steel RGD and Torrie JH. 1960. Principals and Procedure of Statistics, McGraw-Hill, New Yark, 111-112. Steel RGD and Torrie JH. 1980. Analysis of covariance, In: Principles and Procedures of Statistics: a Biometrical Approach, pp. 401-437. McGrawHill, New York. Tukey JW. 1990. Data based grapics: Visual display in the decades to come. Stat. Sci. 5: 327-339. Wiese MV. 1987. Compendium of wheat diseases. 2nd Edition. APS Press, USA. 112pp. 214