| Wheat stripe(yellow)rust,caused by Puccinia striiformis f.sp.tritici(Pst),is one of the most damaging diseases of wheat worldwide.It can cause an annual loss of wheat yield to be about 1 million tons.In China,the wheat crop is the second most important cereal and food crop both in production and area,so it has the largest stripe rust regions,affected by this disease.Recently,in 2017,wheat stripe rust epidemic spread to twelve provinces in China and affected about 1.65 million hectares of wheat area.In Pakistan,about 70%(5.8 m ha)wheat growing area is prone to Pst and huge wheat yield losses are reported from Pakistan almost every year.Since 1948,thirteen stripe rust epidemics have been reported from Pakistan causing huge yield losses.The fungus is able to produce a large number of pathogenic races of high virulence genes through mutation,introduction,genetic drift,asexual or sexual recombination,and host selection.These more virulent races are capable to overcome host resistance genes.Some races are highly aggressive and become widespread and dominant in a particular region or several regions.For example,in China,CRY32 is the most dominant and virulent race of Pst.Similarly,in Pakistan,the most frequent races identified are 574212,574232,474232,474233,574213 and 434232.Among these dominant races,574232 having avirulence/virulence formula: Yr5,Yr10,Yr15,Yr 41,Yr 43,Yr 46,Yr Tr1,Yr Tye/Yr1,Yr6,Yr7,and Yr8,Yr9,Yr17,Yr42,Yr44,Yr45 was found to be the most dominant race.The study of the genetics of the pathogen can help to understand the interaction of different alleles at the same locus and different loci,which are the determinants of genetic diversity either through sexual recombination,somatic recombination,or mutation.The homozygosity or heterozygosity of virulence loci and dominance or recessiveness of the opposite phenotypes in a race or population can be used to predict the likelihood of the emergence of new virulent races or genes.The stripe rust pathogen infecting cereal crops and grasses was believed to have a hemicyclic life cycle consisting of uredinial and telial stages before the recent discovery of barberry(Berberis spp.)as an alternate(aecial)host for the fungus.This discovery has improved the understanding of the biology of the stripe rust pathogen.The Himalayan and near-Himalayan regions of Pakistan,China,and Nepal are considered as the center of diversity for Pst pathogen.The high genetic diversity has been reported in these areas,probably resulting from the sexual reproduction of the stripe rust fungus.To determine if Berberis species growing in Pakistan are susceptible to Pst,we collected seeds of five species and two subspecies from the Himalayan region in 2016 and inoculated the seedlings with germinated teliospores of a Pakistani dominant race(574232)of Pst under controlled conditions.Pycnia and aecia were produced on all inoculated plants of these species and subspecies and demonstrated as Pst by successful infection of wheat plants with aeciospores.This study showed that the tested Pakistani Berberis species and subspecies were susceptible to Pst under controlled conditions.Wheat stripe rust disease seriously threatens wheat production in almost all wheat growing regions in the world.Being an obligate biotrophic fungus,it produces new virulent races which can break the resistance of cultivating wheat Yr resistant lines.The discovery of barberry as its aecial host made it possible to study the inheritance of Pst using the classical genetic approach.We used a total of 115 single-spore isolates,generated through sexual reproduction by artificially inoculated susceptible Himalayan barberry(B.pseudumbellata)with a single isolate [Pak-1-(A)-9] of Pakistani dominant race(574232)of Pst.The progeny isolates were homozygous avirulent to 5 single-gene lines(Yr5,Yr15,Yr24,Yr26,and Yr Tye),homozygous virulent to 9 single-gene lines(Yr3,Yr6,Yr7,Yr8,Yr25,Yr27,Yr28,Yr43,Yr44)and segregated on 10 single-gene lines(Yr1,Yr2,Yr9,Yr17,Yr47,Yr A,Yr Sp,Yr Tr1,Yr Exp2 and Yr10).The progeny isolates generated through sexual reproduction on barberry plants were considered as F2 generation and the heterokaryotic urediniospores as diploid for the genetic studies.The degree of fit of observed numbers to theoretical segregation ratios was calculated by χ2 tests to determine both virulence and SSR markers.We found complex interaction between the resistant genes of wheat Yr single-gene lines and virulent genes of the pathogen.The results show that the parental Pakistani isolate was homozygous avirulent for the first group of four avirulence loci,homozygous virulent for the second group of ten virulence loci,and heterozygous for the third group of ten loci.From the 115 progeny isolates,25 VPs were identified.The VP1 was comprised of 42(36.5%)isolates had the same virulence pattern as the parental isolate.While all other 24 VPs were different from that of the parental isolate in virulence/avirulence on 10 wheat Yr single-gene lines.Six VPs(VP2 to VP7),comprising of 39 isolates,had the same number of virulences,compared with the parental isolate.11 VPs(VP8 to VP18),comprised of 24 isolates had shown an increased number of virulences;while the remaining 7 VPs(VP19 to VP25)had shown a decreased number of virulences.The results show that the single isolate of a dominant Pakistani race of Pst produced a large number of races with more virulences.For individual responses of virulences and avirulence,the parental isolate and progeny isolates were all homozygous virulent to Yr1,Yr2,Yr9,Yr17,Yr47 and Yr A;and homozygous avirulent to Yr Sp,Yr Tr1,Yr Exp2,and Yr10,indicating that the parental isolate was homozygous in virulence to the first group of resistance genes and homozygous in avirulence to the latter group of wheat Yr single-gene lines.Segregations were observed for virulences to Yr1,Yr2,Yr9,Yr17,Yr27 and Yr A;and for avirulence to Yr Sp,Yr Tr1,Yr Exp2,and Yr10.A total of 141 SSR markers,specific for Pst,were screened.Of the 141 SSR markers screened,10 produced co-dominant amplicons in the parental isolate and segregated in the progeny population.We found different segregation ratios on virulence/avirulence loci.The segregations of the 115 progeny isolate best-fit avirulence to virulence ratio(1:3)for avirulence to Yr Sp,Yr Tr1,Yr Exp2,and Yr10 indicating that the parental isolate was controlled by a single avirulence dominant gene.The virulences to Yr1 and Yr9 of the progeny isolates best fit to avirulence and virulence ratios(1:15)indicating that two complementary recessive genes are involved in controlling avirulence phenotypes.For virulence to Yr2,Yr17,Yr47,and Yr A,the avirulence to virulence ratio(3:13)fits best,indicating that the avirulence phenotypes were controlled by a recessive gene and the parental isolate has a complementary dominant gene for avirulence phenotypes.The calculated frequency distribution of the number of marker loci becoming homozygous was close to the normal distribution based on progeny isolates and MLGs.This shows that the 115 progeny isolates were randomly generated and the selected SSR markers are suitable to construct the linkage map.Using all the 10 segregating SSR markers and VPs,a map consisting of two linkages was constructed.The linkage 1 consisted of 9 SSR markers and linkage 2 consisted of 10 virulence/avirulence genes and 1 SSR marker.The genetic distance between two neighboring virulence loci ranged from 3.90 c M between avir10 and Avir Sp,and 20.02 c M between Avir17 and Avir47.The linkage 2 of 9 SSR markers spanning 131.58 c M shows genetic distances ranged from 12.65 to 16.26 c M between SSR markers SUNIPst11-10 and SUNIPst10-06.These results show that the barberry plants may play a role in the diversity of the pathogen through sexual reproduction under natural conditions.As in this study,the artificially generated sexual progeny population from a single isolate of Pst had a diversity of virulence and these results can be used to identify resistant genes in developing wheat lines for future breeding programs.However,there is a need to study the genetic inheritance of Pst populations collected from naturally infected barberry species in the Himalayan region. |
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