Identification of RAPD markers for 11 Hessian fly resistance genes in wheat

 The pyramiding of genes that confer race- or biotype-specific resistance has become increasingly attractive as a breeding strategy now that DNA-based marker-assisted selection is feasible. Our objective here was to identify DNA markers closely linked to genes in wheat (Triticum aestivum L.) that condition resistance to Hessian fly [Mayetiola destructor (Say)]. We used a set of near-isogenic wheat lines, each carrying a resistance gene at 1 of 11 loci (H3, H5, H6, H9, H10, H11, H12, H13, H14, H16 or H17) and developed by backcrossing to the Hessian fly-susceptible wheat cultivar ‘Newton’. Using genomic DNA of these 11 lines and ‘Newton’, we have identified 18 randomly amplified polymorphic DNA (RAPD) markers linked to the 11 resistance genes. Seven of these markers were identified by denaturing gradient gel electrophoresis and the others by agarose gel electrophoresis. We confirmed linkage to the Hessian fly resistance loci by cosegregation analysis in F₂ populations of 50–120 plants for each different gene. Several of the DNA markers were used to determine the presence/absence of specific Hessian fly resistance genes in resistant wheat lines that have 1 or possibly multiple genes for resistance. The use of RAPD markers presents a valuable strategy for selection of single and combined Hessian fly resistance genes in wheat improvement. Received: 20 March 1996 / Accepted: 6 September 1996     

Identification and mapping of H32, a new wheat gene conferring resistance to Hessian fly

H32 is a newly identified gene that confers resistance to the highly pervasive Biotype L of the Hessian fly [ Mayetiola destructor (Say)]. The gene was identified in a synthetic amphihexaploid wheat, W-7984, that was constructed from the durum ‘Altar 84’ and Aegilops tauschii. This synthetic wheat is one of the parents of the marker-rich ITMI population, which consists of 150 recombinant inbred lines (RILs) derived by single-seed descent from a cross with ‘Opata 85’. Linkage analysis of the H32 locus in the ITMI population placed the gene between flanking microsatellite (SSR) markers, Xgwm3 and Xcfd223, at distances of 3.7 and 1.7 cM, respectively, on the long arm of chromosome 3D. The Xgwm3 primers amplified codominant SSR alleles, a 72 bp fragment linked in coupling to the resistance allele and an 84 bp fragment linked in repulsion. Primers for the SSR Xcfd223 amplified a 153 bp fragment from the resistant Synthetic parent and a 183 bp fragment from the susceptible Opata line. Deletion mapping of the flanking Xgwm3 and Xcfd223 markers located them within the 3DL-3 deletion on the distal 19% of the long arm of chromosome 3D. This location is at least 20 cM proximal to the reported 3DL location of H24, a gene that confers resistance to Biotype D of the Hessian fly. Tight linkage of the markers will provide a means of detecting H32 presence in marker-assisted selection and gene pyramiding as an effective strategy for extending durability of deployed resistance.     

Obviation of wheat resistance to the Hessian fly through systemic induced susceptibility

Unlike most documented plant-insect interactions, Hessian fly-resistance [Mayetiola destructor (Say)] in wheat (Triticum aestivum L.) is initiated by a gene-for-gene recognition event in which plants carrying a specific R gene recognize salivary effectors encoded by a corresponding larval avirulence gene. However, dual infestation resulting from oviposition by virulent insects from 5 d before to 3 d after oviposition by avirulent insects on the same host plant, lead to systemic induced susceptibility, obviation of resistance, and ultimately the survival of both virulent and genetically avirulent progeny to adulthood. Simultaneous oviposition allowed greater survival of avirulent progeny than ovipositions separated by larger intervals. Because of the induction of plant resistance, hatch of avirulent larvae before virulent was more detrimental to rate of development than hatch of virulent before avirulent larvae. Obviation of resistance was not localized to the leaf being attacked by the virulent larvae, but also functioned across spatial distance into younger leaves. This research suggests that virulent Hessian fly larvae directly suppress the defense response of wheat, thus providing a refuge for avirulent genotypes, preserving diversity in field populations and increasing durability of deployed resistance genes.     

Development of STS markers linked to Hessian fly resistance gene H6 in wheat

Hessian fly is one of the world's most destructive insect pests of wheat Triticum aestivum L. We have used the combination of near-isogenic lines (NIL) and random amplified polymorphic DNA (RAPD) analysis to screen up to 2,000 primers to identify DNA markers that are linked to gene H6 that confers resistance to biotype B of the insect. This screen produced six primers that show polymorphic fragments associated with resistance by H6. We have screened 440 F₂ individuals from a cross of the susceptible cultivar Newton and a NIL that contains H6 to verify the linkage between these markers and the resistance gene. A high-resolution genetic map was constructed based on recombination frequency. Two of the markers were tightly linked to the gene with no recombination observed, three were within 2.0 cM, and one was 11 cM from the gene. Three of the six markers were successfully converted to sequence tagged site (STS) markers. Both RAPD and STS primers were used to screen for the presence or absence of the resistance gene in wheat varieties. The identification of markers and construction of the genetic high resolution map provide the first steps toward localization of this resistance gene.     

Association of a DNA marker with Hessian fly resistance gene H9 in wheat

The Hessian fly [Mayetiola destructor (Say)] is a major pest of wheat (Triticum aestivum L.) and genetic resistance has been used effectively over the past 30 years to protect wheat against serious damage by the fly. To-date, 25 Hessian fly resistance genes, designated H1 to H25, have been identified in wheat. With near-isogenic wheat lines differing for the presence of an individual Hessian fly resistance gene, in conjunction with random amplified polymorphic DNA (RAPD) analysis and denaturing gradient-gel electrophoresis (DGGE), we have identified a DNA marker associated with the H9 resistance gene. The H9 gene confers resistance against biotype L of the Hessian fly, the most virulent biotype. The RAPD marker cosegregates with resistance in a segregating F₂ population, remains associated with H9 resistance in a number of different T. aestivum and T. durum L. genetic backgrounds, and is readily detected by either DGGE or DNA gel-blot hybridization.Purdue University, Agric. Exp. Stn. Journal paper No. 14440     

First combined resistance to Hessian fly and Sunn pest identified in synthetic hexaploid wheat

Hessian fly, Mayetiola destructor (Say), and Sunn pest, Eurygaster integriceps (Puton), are the two most damaging insect pests of wheat in North Africa, West and Central Asia. Host plant resistance is the most environmental friendly, cost-effective and practical means of controlling insect pests. Twenty synthetic hexaploid wheat lines selected as resistant to Syrian Sunn pest in 2010 were screened for resistance to Moroccan Hessian fly biotype in 2016. The Hessian fly screening was carried out in standard greenhouse flats using a randomized complete block design with three replications, with susceptible and resistant checks in every test flat. The results showed that three synthetic hexaploid wheat lines exhibited resistance to both Moroccan Hessian fly biotype and Syrian Sunn pest. This is the first record of combined resistance to these two pests in wheat. Mapping populations using these sources of resistance are being developed using double haploid techniques for subsequent genetic characterization and identification of linked molecular markers for marker assisted selection.     

A molecular marker associated with the H21 Hessian fly resistance gene in wheat

Near-isogenic lines in conjunction with bulked segregant analysis were used to identify a DNA marker in wheat (Triticum aestivum L.) associated with the H21 gene conferring resistance to biotype L of Hessian fly [Mayetiola destructor (Say)] larvae. Near-isogenic lines were developed by backcross introgression BC3F3:4 (Coker 797 * 4 / Hamlet) and differed by the presence or absence of H21 (on 2RL) derived from Chaupon rye (Secale cereale L.). Bulked DNA samples were prepared from near-isogenic lines and BC3F2 population individuals segregating for reaction to Hessian fly biotype L and screened for random amplified polymorphic DNA markers using 46 10mer primers. Random-amplified polymorphic DNA markers from resistant and susceptible individuals and parental lines were scored and these data were used to identify a 3 kb DNA fragment that was related to the occurrence of H21. This fragment was amplified from DNA isolated from Hamlet, a near-isogenic line carrying 2RL, and bulked DNA from resistant BC3F2 individuals, but not from the recurrent parent Coker 797 or DNA bulks from susceptible BC3F2 plants. Analysis of 111 BC3F2 segregating individuals and BC3F2:3 segregants confirmed the co-segregation of the 3 kb DNA marker with the H21 resistance gene to Hessian fly. Use of this marker could facilitate more rapid screening of plant populations for Hessian fly resistance and monitoring the introgression of H21.     

Hypersensitive response of wheat to the Hessian fly

Hessian flyMayetiola destructor (Say) larvae are able to obtain food from their host plant without inflicting mechanical damage to the plant surface, apparently by secreting substances which elicit release of nutrients from plant cells surrounding the feeding site. Cells of fully susceptible plants retain their normal appearances, while in resistant plants extensive areas of cellular collapse occur. These responses indicate that hypersensitivity is the basis of wheat's resistance to the Hessian fly. The fly's feeding mechanism more closely resembles that of a pathogen than of a phytophagous insect; correspondingly, both the genetic relationship and resistance mechanism of the host plant to the parasite are of the sorts commonly associated with bacterial and fungal pathogens.     

Precisely mapping a major gene conferring resistance to Hessian fly in bread wheat using genotyping-by-sequencing

Background

One of the reasons hard red winter wheat cultivar ‘Duster’ (PI 644016) is widely grown in the southern Great Plains is that it confers a consistently high level of resistance to biotype GP of Hessian fly (Hf). However, little is known about the genetic mechanism underlying Hf resistance in Duster. This study aimed to unravel complex structures of the Hf region on chromosome 1AS in wheat by using genotyping-by-sequencing (GBS) markers and single nucleotide polymorphism (SNP) markers.

Results

Doubled haploid (DH) lines generated from a cross between two winter wheat cultivars, ‘Duster’ and ‘Billings’ , were used to identify genes in Duster responsible for effective and consistent resistance to Hf. Segregation in reaction of the 282 DH lines to Hf biotype GP fit a one-gene model. The DH population was genotyped using 2,358 markers developed using the GBS approach. A major QTL, explaining 88% of the total phenotypic variation, was mapped to a chromosome region that spanned 178 cM and contained 205 GBS markers plus 1 SSR marker and 1 gene marker, with 0.86 cM per marker in genetic distance. The analyses of GBS marker sequences and further mapping of SSR and gene markers enabled location of the QTL-containing linkage group on the short arm of chromosome 1A. Comparative mapping of the common markers for the gene for QHf.osu-1A^d in Duster and the Hf-resistance gene for QHf.osu-1A⁷⁴ in cultivar ‘2174’ showed that the two Hf resistance genes are located on the same chromosome arm 1AS, only 11.2 cM apart in genetic distance. The gene at QHf.osu-1A^d in Duster has been delimited within a 2.7 cM region.

Conclusion

Two distinct resistance genes exist on the short arm of chromosome 1A as found in the two hard red winter cultivars, 2174 and Duster. Whereas the Hf resistance gene in 2174 is likely allelic to one or more of the previously mapped resistance genes (H9, H10, H11, H16, or H17) in wheat, the gene in Duster is novel and confers a more consistent phenotype than 2174 in response to biotype GP infestation in controlled-environment assays.

Electronic supplementary material

The online version of this article (doi:10.1186/s12864-015-1297-7) contains supplementary material, which is available to authorized users.     

Saturation and comparative mapping of the genomic region harboring Hessian fly resistance gene H26 in wheat

Resistance gene H26, derived from Aegilops tauschii Coss., is one of the most effective R genes against the Hessian fly [Mayetiola destructor (Say)], an important pest of wheat (Triticum aestivum L.). Using a limited number of PCR-based molecular markers a previous study mapped H26 to the wheat chromosomal deletion bin 3DL3-0.81-1.00. The objectives of this study were to saturate the chromosomal region harboring H26 with newly developed PCR-based markers and to investigate the collinearity of this wheat chromosomal region with rice (Oryza sativa L.) and Brachypodium distachyon genome. A population of 96 F₂ individuals segregating at the H26 gene locus was used for saturation mapping. All wheat ESTs assigned to the deletion bin 3DL3-0.81-1.00 were used to design STS (sequence tagged site) primers. The wheat ESTs mapped near H26 were further used to BLAST rice and B. distachyon genomic sequences for comparative mapping. To date, 26 newly developed STS markers have been mapped to the chromosomal region spanning the H26 locus. Two of them were mapped 1.0 cM away from the H26 locus. Comparative analysis identified genomic regions on rice chromosome 1 and Brachypodium Super contig 13 which are collinear with the genomic region spanning the H26 locus within the distal region of 3DL. The newly developed STS markers closely linked to H26 will be useful for mapped-based cloning of H26 and marker-assisted selection of this gene in wheat breeding. The results will also enhance understanding of this chromosomal region which contains several other Hessian fly resistance genes. Mention of trade names or commercial products in this article is solely for the purpose of providing specific information and does not imply recommendation or endorsement by the U.S. Department of Agriculture.     

Genetic characterization and molecular mapping of Hessian fly resistance genes derived from Aegilops tauschii in synthetic wheat

Two synthetic hexaploid wheat lines (×Aegilotriticum spp., 2n = 6x = 42, genomes AABBDD), SW8 and SW34, developed from the crosses of the durum wheat cultivar Langdon (Triticum turgidum L. var. durum, 2n = 4x = 28, genomes AABB) with two Aegilops tauschii Cosson accessions (2n = 2x = 14, genome DD), were determined to carry Hessian fly [Mayetiola destructor (Say)] resistance genes derived from the Ae. tauschii parents. SW8 was resistant to the Hessian fly biotype Great Plains (GP) and strain vH13 (virulent to H13). SW34 was resistant to biotype GP, but susceptible to strain vH13. Allelism tests indicated that resistance genes in SW8 and SW34 may be allelic to H26 and H13 or correspond to paralogs at both loci, respectively. H26 and H13 were localized to chromosome 4D and 6D, respectively, in previous studies. Molecular mapping in the present study, however, assigned the H26 locus to chromosome 3D rather than 4D. On the other hand, mapping of the resistance gene in SW34 verified the previous assignment of the H13 locus to chromosome 6D. Linkage analysis and physical mapping positioned the H26 locus to the chromosomal deletion bin 3DL3-0.81–1.00. A linkage map for each of these two resistance genes was constructed using simple sequence repeat (SSR) and target region amplification polymorphism (TRAP) markers.     

Transfer of Hessian fly resistance from rye to wheat via radiation-induced terminal and intercalary chromosomal translocations

Summary A new Hessian fly (Mayetiola destructor) resistance gene derived from Balbo rye and its transfer to hexaploid wheat via radiation-induced terminal and intercalary chromosomal translocations are described. Crosses between resistant Balbo rye and susceptible Suwon 92 wheat and between the F₁ amphidiploids and susceptible TAM 106 and Amigo wheats produced resistant BC₂F₃ lines that were identified by C-banding analysis as being 6RL telocentric addition lines. Comparative chromosomal analyses and resistance tests revealed that the resistance gene is located on the 6RL telocentric chromosome. X-irradiated pollen of 6RL addition plants was used to fertilize plants of susceptible wheats TAM 106, TAM 101, and Vona. After several generations of selection for resistance, new sublines were obtained that were homogeneous for resistance. Thirteen of these lines were analyzed by C-banding, and three different wheat-6RL chromosomal translocations (T) were identified. Wheat chromosomes involved in the translocations were 6B, 4B, and 4A. Almost the complete 6RL arm is present in T6BS · 6BL-6RL. Only the distal half of 6RL is present in T4BS · 4BL-6RL, which locates the resistance gene in the distal half of 6RL. Only a very small segment (ca 1.0 m) of the distal region of 6RL is present in an intercalary translocation (Ti) Ti4AS · 4AL-6RL-4AL. The 6RL segment is inserted in the intercalary region between the centromere of chromosome 4A and the large proximal C-band of 4AL. The break-points of the translocations are outside the region of the centromere, indicating that they were induced by the X-ray treatment. All three translocations are cytologically stable and can be used directly in wheat breeding programs.Cooperative investigations of the Kansas Agricultural Experiment Station, Departments of Entomology and Plant Pathology, the Wheat Genetics Resource Center, Kansas State University, and the US Department of Agriculture, Agricultural Research Service. Contribution No. 91-117-JDeceased     

A novel wheat gene encoding a putative chitin-binding lectin is associated with resistance against Hessian fly

The gene-for-gene interaction triggering resistance of wheat against first-instar Hessian fly larvae utilizes specialized defence response genes not previously identified in other interactions with pests or pathogens. We characterized the expression of Hfr-3 , a novel gene encoding a lectin-like protein with 68–70% identity to the wheat germ agglutinins. Within each of the four predicted chitin-binding hevein domains, the HFR-3 translated protein sequence contained five conserved saccharide-binding amino acids. Quantification of Hfr-3 mRNA levels confirmed a rapid response and gradual increase, up to 3000-fold above the uninfested control in the incompatible interaction 3 days after egg hatch. Hfr-3 mRNA abundance was influenced by the number of larvae per plant, suggesting that resistance is localized rather than systemic. In addition, Hfr-3 was responsive to another sucking insect, the bird cherry-oat aphid, but not to fall armyworm attack, wounding or exogenous application of methyl jasmonate, salicylic acid or abscisic acid. Western blot analysis demonstrated that HFR-3 protein increased in parallel to mRNA levels in crown tissues during incompatible interactions. HFR-3 protein was detected in both virulent and avirulent larvae, indicating ingestion. Anti-nutritional proteins, such as lectins, may be responsible for the apparent starvation of avirulent first-instar Hessian fly larvae during the initial few days of incompatible interactions with resistant wheat plants.     

Hessian fly resistance gene H13 is mapped to a distal cluster of resistance genes in chromosome 6DS of wheat

H13 is inherited as a major dominant resistance gene in wheat. It was previously mapped to chromosome 6DL and expresses a high level of antibiosis against Hessian fly (Hf) [Mayetiola destructor (Say)] larvae. The objective of this study was to identify tightly linked molecular markers for marker-assisted selection in wheat breeding and as a starting point toward the map-based cloning of H13. Fifty-two chromosome 6D-specific microsatellite (simple sequence repeat) markers were tested for linkage to H13 using near-isogenic lines Molly (PI 562619) and Newton-207, and a segregating population consisting of 192 F_2:3 families derived from the cross PI 372129 (Dn4) × Molly (H13). Marker Xcfd132 co-segregated with H13, and several other markers were tightly linked to H13 in the distal region of wheat chromosome 6DS. Deletion analysis assigned H13 to a small region closely proximal to the breakpoint of del6DS-6 (FL 0.99). Further evaluation and comparison of the H13-linked markers revealed that the same chromosome region may also contain H23 in KS89WGRC03, an unnamed H gene (H_WGRC4) in KS89WGRC04, the wheat curl mite resistance gene Cmc4, and a defense response gene Ppo for polyphenol oxidase. Thus, these genes comprise a cluster of arthropod resistance genes. Marker analysis also revealed that a very small intercalary chromosomal segment carrying H13 was transferred from the H13 donor parent to the wheat line Molly.Mention of commercial or proprietary product does not constitute an endorsement by the USDA.     

Modulation of defense-response gene expression in wheat during Hessian fly larval feeding

Abstract

Expression profiles of ten genes commonly up-regulated during plant defense against microbial pathogens were compared temporally during compatible and incompatible interactions with first-instar Hessian fly larvae, in two wheat lines carrying different resistance genes. Quantitative real-time PCR revealed that while a lipoxygenase gene (WCI-2) was strongly up-regulated during the incompatible interactions, genes encoding β-1,3 endoglucanase (GNS) and an integral membrane protein (WIR1) were moderately responsive. Genes for thionin-like protein (WCI-3), PR-17-like protein (WCI-5), MAP kinase (WCK-1), phenylalanine ammonia-lyase (PAL), pathogenesis-related protein-1 (PR-1), receptor-like kinase (LRK10) and heat shock protein 70 (HSP70) were minimally responsive. The application of signaling molecules, salicylic acid (SA), methyl jasmonate (MJ) and abscisic acid (ABA), to insect-free plants demonstrated association of these genes with specific defense-response pathways. SA-induced up-regulation of a gene related to lipoxygenases that are involved in jasmonic acid (JA)-biosynthesis is suggestive of positive cross-talk between SA- and JA-mediated signaling pathways. Data suggest that alternative mechanisms may be involved since few of these classical defense-response genes are significantly up-regulated during incompatible interactions between wheat and Hessian fly.     

Quantitative trait loci for aluminum resistance in wheat

Quantitative trait loci (QTL) for wheat resistance to aluminum (Al) toxicity were analyzed using simple sequence repeats (SSRs) in a population of 192 F₆ recombinant inbred lines (RILs) derived from a cross between an Al-resistant cultivar, Atlas 66 and an Al-sensitive cultivar, Chisholm. Wheat reaction to Al was measured by relative root growth and root response to hematoxylin stain in nutrient-solution culture. After screening 1,028 SSR markers for polymorphisms between the parents and bulks, we identified two QTLs for Al resistance in Atlas 66. One major QTL was mapped on chromosome 4D that co-segregated with the Al-activated malate transporter gene (ALMT1). Another minor QTL was located on chromosome 3BL. Together, these two QTLs accounted for about 57% of the phenotypic variation in hematoxylin staining score and 50% of the variation in net root growth (NRG). Expression of the minor QTL on 3BL was suppressed by the major QTL on 4DL. The two QTLs for Al resistance in Atlas 66 were also verified in an additional RIL population derived from Atlas 66/Century. Several SSR markers closely linked to the QTLs were identified and have potential to be used for marker-assisted selection (MAS) to improve Al-resistance of wheat cultivars in breeding programs.     

Comparative analysis of genetic background in eight near-isogenic wheat lines with different H genes conferring resistance to Hessian fly

Near-isogenic lines (NILs) are useful for plant genetic and genomic studies. However, the strength of conclusions from such studies depends on the similarity of the NILs' genetic backgrounds. In this study, we investigated the genetic similarity for a set of NILs developed in the 1990s to study gene-for-gene interactions between wheat (Triticum aestivum L.) and the Hessian fly (Mayetiola destructor (Say)), an important pest of wheat. Each of the eight NILs carries a single H resistance gene and was created by successive backcrossing for two to six generations to susceptible T. aestivum 'Newton'. We generated 256 target region amplification polymorphism (TRAP) markers and used them to calculate genetic similarity, expressed by the Nei and Li (NL) coefficient. Six of the NILs (H3, H5, H6, H9, H11, and H13) had the highly uniform genetic background of Newton, with NL coefficients from 0.97 to 0.99. However, genotypes with H10 or H12 were less similar to Newton, with NL coefficients of 0.86 and 0.93, respectively. Cluster analysis based on NL coefficients and pedigree analysis showed that the genetic similarity between each of the NILs and Newton was affected by both the number of backcrosses and the genetic similarity between Newton and the H gene donors. We thus generated an equation to predict the number of required backcrosses, given varying similarity of donor and recurrent parent. We also investigated whether the genetic residues of the donor parents that remained in the NILs were related to linkage drag. By using a complete set of 'Chinese Spring' nullisomic-tetrasomic lines, one third of the TRAP markers that showed polymorphism between the NILs and Newton were assigned to a specific chromosome. All of the assigned markers were located on chromosomes other than the chromosome carrying the H gene, suggesting that the genetic residues detected in this study were not due to linkage drag. Results will aid in the development and use of near-isogenic lines for studies of the functional genomics of wheat.