Molecular genetic mapping of three X-linked avirulence genes, vH6, vH9 and vH13, in the Hessian fly.

Three X-linked avirulence genes, vH6, vH9, and vH13 in the Hessian fly, Mayetiola destructor, confer avirulence to Hessian fly resistance genes H6, H9, and H13 in wheat. We used a combination of two- and three-point crosses to determine the order of these genes with respect to each other, the white eye mutation and three X-linked molecular markers, G15-1, 020, and 021, developed from genomic lambda clones, lambda G15-1, lambda 020, and lambda 021. The gene order was determined to be vH9-vH6-G15-1-w-vH13-020-021. In situ hybridization of lambda G15-1, lambda 020, and lambda 021, on the polytene chromosomes of the Hessian fly salivary gland established their orientation on Hessian fly chromosome X1. Based on the size of the Hessian fly genome, and the genetic distances between markers, the relationship of physical to genetic distance was estimated at no more than 300 kb/cM along Hessian fly chromosome X1, suggesting that map-based cloning of these avirulence genes will be feasible.     

Chromosome landing near avirulence gene vH13 in the Hessian fly.

AFLP markers in linkage disequilibrium with vH13, an avirulence gene in the Hessian fly (Mayetiola destructor) that conditions avirulence to resistance gene H13 in wheat (Triticum spp.), were discovered by bulked segregant analysis. Five AFLPs were converted into codominant site-specific markers that genetically mapped within 13 cM of this gene. Flanking markers used as probes positioned vH13 near the telomere of the short arm of Hessian fly chromosome X2. These results suggest that the X-linked avirulence genes vH6, vH9, and vH13 are present on Hessian fly chromosome X2 rather than on chromosome X1 as reported previously. Genetic complementation demonstrated that recessive alleles of vH13 were responsible for the H13-virulence observed in populations derived from four different states in the U.S.A.: Georgia, Maryland, Virginia, and Washington. Results support the hypothesis that a gene-for-gene interaction exists between wheat and Hessian fly.     

Biotype composition of Hessian fly (Diptera: Cecidomyiidae) populations from the southeastern, midwestern, and northwestern United States and virulence to resistance genes in wheat

Twenty-three Hessian fly, Mayetiola destructor (Say), populations collected in the southeastern (Alabama and Mississippi), midwestern (Indiana), and northwestern (Idaho and Washington) United States from 1995 to 1999 were evaluated for biotype composition based on response to Hessian fly resistance genes H3, H5, H6, and H7H8 in wheat, Triticum aestivum L. Biotypes L and O, combined, made up at least 60% of all Alabama populations. Biotype L was predominant in the northern third of Alabama and biotype O in the southern two-thirds of the state. Based on biotype data, wheat cultivars with H7H8 resistance should be highly effective in central and southern Alabama. Fifty-four percent of the Mississippi population consisted of biotype L, and the remaining virulent biotypes (B, D, E, G, J, and O) ranged in frequency from 1 to 17%. The Mississippi population also contained 4% of the avirulent biotype GP. Only biotypes D and L were found in Indiana populations, but biotype L was predominant. Hessian fly populations from Idaho and Washington contained one or more of the virulent biotypes D-H, J, and L-O; however, only biotypes E, F, and G occurred at frequencies > 12%. The avirulent biotype GP made up 25-57% of Idaho and Washington populations, a much higher percentage than found in populations from the eastern United States. Although the highest level of virulence in Idaho and Washington populations was found to resistance genes H3 and H6, the frequency of biotype GP would indicate that the currently deployed gene H3 would provide a moderate to high level of resistance, depending on location. Nine of the populations, plus populations collected from the mid-Atlantic state area in 1989 and 1996, also were tested against the wheat cultivar 'INW9811' that carries H13 resistance to Hessian fly biotype L and two Purdue wheat lines with unidentified genes for resistance. The H13 resistance in INW9811 was highly effective against all populations tested from the eastern and northwestern U.S. wheat production areas, except Maryland and Virginia. Population studies also indicated that wheat line CI 17960-1-1-2-4-2-10 likely carries the H13 resistance gene, based on the similarity of its response and that of INW9811 to eight fly populations. Continued monitoring of biotype frequency in Hessian fly populations is required for optimal deployment and management of resistance genes in all wheat production areas.     

Biotypes of Hessian fly (Dipt., Cecidomyiidae) in Morocco

Hessian fly, Mayetiola destructor (Say), is the most important insect pest of wheat in Morocco, where host plant resistance has been used successfully for control. Our objective was to determine the frequency of Hessian fly virulence on H5, H13 and H22 resistance genes. Five Hessian fly populations from the principal cereal‐growing regions in Morocco were studied. The variability in percentage of susceptible plants across Hessian fly populations was highly significant (P < 0.01), indicating differences in virulence frequencies. Plants with the H13 gene had the lowest percentage of susceptible plants, 1.77 and 1.51%, when infested with Hessian flies from Fes and Marchouch, respectively. A low level of virulence to H22 was detected in Fes, Abda and Marchouch populations, 1.87, 1.54 and 1.99% susceptible plants, respectively. The level of virulence to H5 was low in all the five populations. The Beni Mellal population gave the highest percentage of susceptible plants carrying H13 and H22 genes, 6.43 and 7.28%, respectively. The size of live larvae on susceptible plants of the three cultivars carrying H5, H13 and H22 was similar to that of the susceptible check, indicating that a true virulence (biotype) is developing in Hessian fly populations in Morocco. Thus, continuous monitoring of the development of Hessian fly biotypes is essential for optimal deployment of resistance genes.     

Virulence in Hessian fly (Diptera: Cecidomyiidae) field collections from the southeastern United States to 21 resistance genes in wheat

Genetic resistance in wheat, Triticum aestivum L., is the most efficacious method for control of Hessian fly, Mayetiola destructor (Say) (Diptera: Cecidomyiidae). However, because of the appearance of new genotypes (biotypes) in response to deployment of resistance, field collections of Hessian fly need to be evaluated on a regular basis to provide breeders and producers information on the efficacy of resistance (R) genes with respect to the genotype composition of Hessian fly in regional areas. We report here on the efficacy of 21 R genes in wheat to field collections of Hessian fly from the southeastern United States. Results documented that of the 21 R genes evaluated only five would provide effective protection of wheat from Hessian fly in the southeastern United States. These genes were H12, H18, H24, H25, and H26. Although not all of the 33 identified R genes were evaluated in the current study, these results indicate that identified genetic resistance to protect wheat from Hessian attack in the southeastern United States is a limited resource. Historically, R genes for Hessian fly resistance in wheat have been deployed as single gene releases. Although this strategy has been successful in the past, we recommend that in the future deployment of combinations of highly effective previously undeployed genes, such as H24 and H26, be considered. Our study also highlights the need to identify new and effective sources of resistance in wheat to Hessian fly if genetic resistance is to continue as a viable option for protection of wheat in the southeastern United States.     

Gene-for-gene defense of wheat against the Hessian fly lacks a classical oxidative burst

Genetic similarities between plant interactions with microbial pathogens and wheat interactions with Hessian fly larvae prompted us to investigate defense and counterdefense mechanisms. Plant oxidative burst, a rapid increase in the levels of active oxygen species (AOS) within the initial 24 h of an interaction with pathogens, commonly is associated with defenses that are triggered by gene-for-gene recognition events similar to those involving wheat and Hessian fly larvae. RNAs encoded by Hessian fly superoxide dismutase (SOD) and catalase (CAT) genes, involved in detoxification of AOS, increased in first-instar larvae during both compatible and incompatible interactions. However, mRNA levels of a wheat NADPH oxidase (NOX) gene that generates superoxide (O2-) did not increase. In addition, inhibiting wheat NOX enzyme with diphenyleneiodonium did not result in increased survival of avirulent larvae. However, nitro blue tetrazolium staining indicated that basal levels of O2- are present in both uninfested and infested wheat tissue. mRNA encoded by wheat genes involved in detoxification of the cellular environment, SOD, CAT, and glutathione-S-transferase did not increase in abundance. Histochemical staining with 3,3-diaminobenzidine revealed no increases in wheat hydrogen peroxide (H2O2) during infestation that were correlated with the changes in larval SOD and CAT mRNA. However, treatment with 2',7'-dichlorofluorescin demonstrated the presence of basal levels of H2O2 in the elongation zone of both infested and uninfested plants. The accumulation of a wheat flavanone 3-hydroxylase mRNA did show some parallels with larval gene mRNA profiles. These results suggested that larvae encounter stresses imposed by mechanisms other than an oxidative burst in wheat seedlings.     

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.     

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     

Characterization of Wheat Random Amplified Polymorphic DNA Markers Associated with the H11 Hessian Fly Resistance Gene

In Tunisia, the Hessian fly Mayetiola destructor Say is a major pest of durum wheat （Triticum durum Desf.） and bread wheat （T. aestivum L.）. Genetic resistance is the most efficient and economical method of control of this pest. To date, 31 resistance genes, designated H1-H31, have been identified in wheat. These genes condition resistance to the insect genes responsible for virulence. Using wheat cultivars differing for the presence of an individual Hessian fly resistance gene and random amplified polymorphic DNA （RAPD） analysis, we have identified a polymorphic 386-bp DNA marker （Xgmib1-1A.1） associated with the H11 Hessian fly resistance gene. Blast analysis showed a high identity with a short region in the wild wheat （T. monococcum） genome, adjacent to the leaf rust resistance Lr10 gene. A genetic linkage was reported between this gene （Lr10） and Hessian fly response in wheat. These data were used for screening Hessian fly resistance in Tunisian wheat germplasm. Xgmib1-1A.1-like fragments were detected in four Tunisian durum and bread wheat varieties. Using these varieties in Hessian fly breeding programs in Tunisia would be of benefit in reducing the damage caused by this fly.     

Ultrastructural changes in the midguts of Hessian fly larvae feeding on resistant wheat

The focus of the present study was to compare ultrastructure in the midguts of larvae of the Hessian fly, Mayetiola destructor (Say), under different feeding regimens. Larvae were either fed on Hessian fly-resistant or -susceptible wheat, and each group was compared to starved larvae. Within 3 h of larval Hessian fly feeding on resistant wheat, midgut microvilli were disrupted, and after 6 h, microvilli were absent. The disruption in microvilli in larvae feeding on resistant wheat were similar to those reported for midgut microvilli of European corn borer, Ostrinia nubilasis (Hubner), larvae fed a diet containing wheat germ agglutinin. Results from the present ultrastructural study, coupled with previous studies documenting expression of genes encoding lectin and lectin-like proteins is rapidly up-regulated in resistant wheat to larval Hessian fly, are indications that the midgut is a target of plant resistance compounds. In addition, the midgut of the larval Hessian fly is apparently unique among other dipterans in that no peritrophic membrane was observed. Ultrastructural changes in the midgut are discussed from the prospective of their potential affects on the gut physiology of Hessian fly larvae and the mechanism of antibiosis in the resistance of wheat to Hessian fly attack.     

Virulence of Hessian fly (Diptera: Cecidomyiidae) in the Fertile Crescent

The Hessian fly, Mayetiola destructor (Say), is an important insect pest of wheat (Triticum spp.) in North Africa, North America, southern Europe and northern Kazakhstan. Both wheat and this pest are believed to have originated from West Asia in the Fertile Crescent. The virulence of a Hessian fly population from Syria against a set of cultivars carrying different resistance genes, in addition to other effective sources with unknown genes, was determined in the field and laboratory at the International Center for Agricultural Research in the Dry Areas (ICARDA) during the 2005/2006 cropping season. Only two resistance genes (H25 and H26) were effective against the Syrian Hessian fly population, making it the most virulent worldwide. This high virulence supports the hypothesis that Hessian fly coevolved with wheat in the Fertile Crescent of West Asia. The ICARDA screening programme is using this Hessian fly population to identify new resistance genes to this pest.     

Does R gene resistance allow wheat to prevent plant growth effects associated with Hessian fly (Diptera: Cecidomyiidae) attack?

Resistance genes (R genes) are an important part of the plant's immune system. Among insects, the Hessian fly, Mayetiola destructor (Say) (Diptera: Cecidomyiidae), larva is the target of the greatest number of characterized R genes (H1-H32). The biochemical/molecular mechanism of R gene resistance to Hessian fly is not well understood. In the absence of an effective R gene, larvae caused extensive growth deficits (> 30 cm) in wheat seedlings. In the presence of one of three effective R genes, H6, H9, or H13, larvae caused small growth deficits (approximately 3-4 cm) in two leaves (third and fourth) that were actively growing during the first days of larval attack. After larvae died on R gene plants, the fifth leaf and tiller leaves exhibited small increases in growth (2-4 cm). Growth responses of susceptible and resistant plants diverged at a time when Hessian fly larvae were establishing a nutritive gall tissue at feeding sites. The results of this study support the hypothesis that R gene resistance cannot prevent initial larval attack, but, by stopping the formation of the larval gall, it prevents the most serious consequences of larval attack.     

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.     

A physically anchored genetic map and linkage to avirulence reveals recombination suppression over the proximal region of Hessian fly chromosome A2

Resistance in wheat (Triticum aestivum) to the Hessian fly (Mayetiola destructor), a major insect pest of wheat, is based on a gene-for-gene interaction. Close linkage (3 +/- 2 cM) was discovered between Hessian fly avirulence genes vH3 and vH5. Bulked segregant analysis revealed two DNA markers (28-178 and 23-201) within 10 cM of these loci and only 3 +/- 2 cM apart. However, 28-178 was located in the middle of the short arm of Hessian fly chromosome A2 whereas 23-201 was located in the middle of the long arm of chromosome A2, suggesting the presence of severe recombination suppression over its proximal region. To further test that possibility, an AFLP-based genetic map of the Hessian fly genome was constructed. Fluorescence in situ hybridization of 20 markers on the genetic map to the polytene chromosomes of the Hessian fly indicated good correspondence between the linkage groups and the four Hessian fly chromosomes. The physically anchored genetic map is the first of any gall midge species. The proximal region of mitotic chromosome A2 makes up 30% of its length but corresponded to <3% of the chromosome A2 genetic map.     

Distribution,population dynamics and damage of Hessian fly,Mayetiola destructor (Diptera: Cecidomyiidae) in North Tunisia

A three years survey and monitoring studies (2013–2014–2015) were carried out through 4 regions of north Tunisia in order to follow the evolution of the distribution, the frequency of occurrence and damage caused by the Hessian fly Mayetiola destructor (Say) to bread wheat (Triticum aestivum L.) and durum wheat (Triticum durum Desf). Moreover, the effectiveness of resistance genes H3, H5, H6, H7, H8, H9, H10, H11, H12, H13, H14, H15, H22, H23, H25 and H26 to protect wheat from Hessian fly attack was assessed in natural field and under controlled laboratory conditions at INRAT-Kef Station. Results showed that Hessian fly was detected in 60.33% and 51.5% of all sampled durum and bread wheat fields, respectively. This pest was more frequent with a higher percentage of infestation in semi-arid regions. Indeed, during 2013, infestation rate attained 12.39% in Kef region against 0.9% registered in Bizerte region. In order to update information about the annual number of generations, we surveyed the population dynamic of Hessian fly in Kef region. Three generations of the fly were counted annually on wheat, with two complete and one incomplete generation. This insect affects host plant growth at different developmental stages. Plant height was the most affected parameter followed by shoot dry weight and tiller number. Field investigations on host resistance revealed that among the 16 tested resistance genes, and only three were strictly effective (H22, H25 and H26). The resistance genes H5, H9, H13 and H9H13 have also conferred high levels of protection against Hessian fly. This work indicated that H22, H25 and H26 genes could be incorporated into Tunisian wheat varieties and released to farmers to manage the threat due to Hessian fly attacks.     

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.     

Characterization of new loci for Hessian fly resistance in common wheat

The discovery of several new loci for resistance to Hessian fly was reported here. QHf.uga-6AL, the late HR61 was recognized from wheat cultivar 26R61 on the distal end of 6AL with resistance to both biotypes E and vH13. It is the first gene or QTL found on this particular chromosome. QHf.uga-3DL and QHf.uga-1AL, physically assigned to the deletion bins 3DL2-0.27–0.81 and 1AL1-0.17–0.61, respectively, were detected for resistance to biotype vH13. Both QTL should represent new loci for Hessian fly resistance and the latter was detectable only in the late seedling stage when tolerance was evident. In addition, QHf.uga-6DS-C and QHf.uga-1AS had minor effect and were identified from the susceptible parent AGS 2000 for resistance to biotype E and vH13, respectively. QHf.uga-6DS-C is different from the known gene H13 on 6DS and QHf.uga-1AS is different from H9 gene cluster on 1AS. These loci also might be new components of Hessian fly resistance, although their LOD values were not highly significant. The QTL detections were all conducted on a RIL mapping population of 26R61/AGS 2000 with good genome coverage of molecular markers. The strategy used in the current study will serve as a good starting point for the discovery and mapping of resistance genes including tolerance to the pest and the closely linked markers will certainly be useful in selecting or pyramiding of these loci in breeding programs.     

No fitness cost for wheat's H gene-mediated resistance to Hessian fly (Diptera: Cecidomyiidae)

Resistance (R) genes have a proven record for protecting plants against biotic stress. A problem is parasite adaptation via Avirulence (Avr) mutations, which allows the parasite to colonize the R gene plant. Scientists hope to make R genes more durable by stacking them in a single cultivar. However, stacking assumes that R gene-mediated resistance has no fitness cost for the plant. We tested this assumption for wheat's resistance to Hessian fly, Mayetiola destructor (Say) (Diptera: Cecidomyiidae). Our study included ten plant fitness measures and four wheat genotypes, one susceptible, and three expressing either the H6, H9, or H13 resistance gene. Because R gene-mediated resistance has two components, we measured two types of costs: the cost of the constitutively-expressed H gene, which functions in plant surveillance, and the cost of the downstream induced responses, which were triggered by Hessian fly larvae rather than a chemical elicitor. For the constitutively expressed Hgene, some measures indicated costs, but a greater number of measures indicated benefits of simply expressing the H gene. For the induced resistance, instead of costs, resistant plants showed benefits of being attacked. Resistant plants were more likely to survive attack than susceptible plants, and surviving resistant plants produced higher yield and quality. We discuss why resistance to the Hessian fly has little or no cost and propose that tolerance is important, with compensatory growth occurring after H gene-mediated resistance kills the larva. We end with a caution: Given that plants were given good growing conditions, fitness costs may be found under conditions of greater biotic or abiotic stress.