Russian Wheat Aphid (RWA) Resistant Germplasm Lines

Three new RWA-resistant barley germplasm lines have been selected and increased from previously identified RWA-resistant accessions from the National Small Grains Collection. R046, a 6-row line, and R047, a 2-row line are both highly resistant while MR059 is a 6-row line with a moderate level of RWA resistance.

Prebreeding for adapted germplasm lines

The prebreeding program is an attempt to bring resistance genes from unadapted germplasm lines into adapted malting and feed barley backgrounds for all barley growing regions in the U.S. It involves repeated backcrossing with intermitent screening with a time commitment of approximately seven years from the first cross until homozygous resistant BC3F3 lines can be evaluated in the field. This is an ongoing process involving various resistant lines and adapted cultivars currently at all phases of the program. The summer of 1997 saw the first 300 lines from this program in the field in Aberdeen, ID with the cooperation of Darrell Wesenberg and Phil Bregitizer of the USDA-ARS. These lines involved three different resistant sources and three adapted cultivars and were evaluated for agronomics with the potential for release of adapted germplasm lines or cultivar release. 150 lines were selected for head rows in 1998 and 29 lines were selected for preliminary yield testing in 1998. Continuing with this program, 2,824 BC3F2/BC3F3 involving 7 adapted cultivars and 22 resistant germplasm lines were increased in the spring of 1996 and 1997 in Stillwater and the resultant BC3F3/BC3F4 lines were screened for homozygous resistance in the fall of 1997. Approximately 800 homozygous resistant lines will be evaluated agromnomically in Abdereen in the summer of 1998. 17 BC3F1 populations were screened and 12 plants transplanted for increase in the greenhouse in the spring of 1998. These BC3F1 populations involved three cultivars and 16 resistant germplasm lines. Another 20 BC3F1 populations were screened, 12 resistant plants increased from each population, and 300 BC3F2 progeny from each population screened, with 100 resistant BC3F2 identified for each population and transplanted for increase in the spring of 1998 in Stillwater and Aberdeen. Five previously increased BC3F2 populations were also screened and 100 resistant plants transplanted for increase in Stillwater in the spring of 1998. These 25 BC3F2 populations involved seven cultivars and 18 resistant lines. Fifty-one BC3 to 10cultivars, 50 BC2 to 15 cultivars, 5 BC1 to three cultivars and three F1 crosses were made towards future adapted RWA-resistant lines to be evaluated in the field in 1999, 2000, 2001, and 2002.

Genetic Studies

Parents, F1, RF1, 150 BC to each parent, 200 F2, and 300 F2-derived F3 families were evaluated for one resistant line to determine the number of genes and genetic control for RWA resistance in this line. Analysis is on going. Complete genetic analysis has been performed for 9 out of a total of 108 RWA -resistant lines developed by the USDA-ARS in Stillwater. All 9 lines have shown multiple gene control for RWA resistance. Forty F1 and 47 BC1 were made, 47 F2 populations were increased, and 300 F2-derived F3 families were increased for each of six populations for future genetic studies. Genetic diversity studies are planned for resistant lines to determine if these lines carry different genes for resistance. This type of analysis requires three years for development of seed necessary for each test. 12 testcrosses of a resistant x resistant F1 on a susceptible were made, 18 TC1F1 populations were increased to TC1F2, and 2,170 TC1F1-derived TC1F2 plants were increased for future genetic diversity studies.

Field studies

Previous field studies indicated that RWA feeding may have a negative effect on malting quality of RWA-resistant germplasm lines. Four agronomically adapted advanced generation RWA-resistant germplasm lines developed by Phil Bregitzer and Darrell Wesenberg with USDA-ARS in Aberdeen, ID in cooperation with the USDA-ARS lab in Stillwater, 6 unadapted RWA-resistant germplasm lines developed by the Stillwater lab, and 2 adapted malting barley cultivars were grown at two locations in Idaho and infested artifically with RWA to study the effect of RWA feeding on agronomics and malting quality of these lines. Only agronomic data was collected because the aphids did not persist in the field at either location. A very similar experiment is being planted again this spring at one location in Colorado where RWA is found every year and again in one location in Idaho. More adapted lines are in this test and fewer unadapted .

Cooperative screening

One thousand two hundred and twenty-nine advanced lines and 273 doubled haploids were screened for homozygous resistance for 4 barley breeders.

Other aphids

The bird cherry oat aphid (BCO) is a constant pest of barley throughout the U.S. Previously the main agronomic concern associated with this aphid was transmission of a serious barley disease Barley Yellow Dwarf Mosaic Virus (BYDMV), but more recently reports have indicated yield loss due to the BCO. BCO damage to seedlings is not easily detected and it has proven impossible, so far, to screen for resistance in the seedling stage. A new screening technique has been developed in Stillwater that can be quickly and easily performed on seedlings. Seeds are planted in transparent seed growth pouches in the greenhouse and germinating seedlings are infested with BCO. In addition, an identical control group is kept aphid free. A visual comparrison of root length and secondary root hairs as well as plant height is made between the infested and noninfested controls and resistant plants selected. Resistance to BCO at the seedling stage has not yet been correlated to grain yield. Approximately 163 accessions have been reported in the literature to have resistance to BCO, in terms of the effect of the plant on aphid biology, or to have resistance to BYDMV. One hundred and ten potential BCO lines selected from these accessions on the basis of agronomics were screened with the new technique. Plants were rescued and transplanted to the greenhouse where yield results will hopefully confirm the success of this new screening technique. At the same time, BCO resistant lines will be identified for future use in the prebreeding program.

Work is currently being conducted to examine the changes in plant hormones, signaling molecules, and primary nitrogen metabolites (i.e. abscisic acid, gibberellins, indole-3-acetic acid, ethylene, jasmonic acid, salicylic acid and GABA) in resistant and susceptible isolines of barley in response to virulent and avirulent biotypes of greenbug. We have also completed a gene expression study that examined the differential expression of wound and pathogen related defense genes in response to virulent and avirulent biotypes of greenbug in resistant and susceptible isolines of barley. In this study we examined the expression of 20 known wound and plant defense-related genes using cDNA clones obtained from individual researchers at Texas Tech University, Kansas State University, and the Swiss Federal Research Station. We demonstrated that greenbugs are capable of infesting resistant and susceptible isolines of barley resulting in phenotypic abnormalities without inducing a broad class of known defense-related or wound genes. We are currently using differential display PCR to isolate and clone unique genes which appear to be associated with plant resistance to greenbugs.

We are using a commercial Differential Display Kit to isolate resistance genes from a pair of near-isolines of barley. However, in using this approach we have developed one key modification to the kit. That is, in one study we have replaced the kits arbitrary PCR primers with a 23 base-pair PCR primer that is homologous to the conserved region of the catalytic domain of a class of protein kinases referred to as serine/threonine receptor kinases (also referred by some as LRR (leucine-rich repeat receptor kinases). This class of genes has been associated with plant resistance to virus in tobacco, bacteria in rice, tomato, and Arabidopsis, fungi in flax, wheat and tomato, as well as nematodes in sugar beet. Using these primers we have identified several candidate cDNA fragments which are present only in the greenbug resistant barley isoline. We are currently using these putative receptor kinase fragments to analyze differential gene expression in the resistant and susceptible isolines. We are determining the segregation pattern of these genes in relation with greenbug resistance in F2 progeny of a resistant versus susceptible cross. These genes, which are expressed at relatively low levels in the plant, are thought to play a role in the perception of plant pathogens. When these receptor kinase perceive plant pathogens they then help to turn on a cascade of signal transduction events which leads to the expression of key wound and pathogen-related genes. It is our hope that these clones which are present only in the resistant isoline may be useful in the genetic engineering of plant resistance to aphids in barley. This same approach is also being used to isolate putative genes which are associated with plant resistance to the Russian wheat aphid.
 

Germplasm Enhancement
Dolores W. Mornhinweg, Geneticist
David R. Porter, Research Geneticist
Dirk B. Hays, Post-doc biochemistry and physiology

Host Plant Resistance
James A. Webster, Research Entomologist
Keith A. Mirkes, Biological Science Technician

Cooperators
Darrell Wesenberg, Research Agronomist, USDA-ARS, Aberdeen, ID
Phil Bregitzer, Research Geneticist, USDA-ARS, Aberdeen, ID
Berne Jones, Research Biochemist, USDA-ARS, Madison, WI
 

Mornhinweg, D.W., D.R. Porter and J.A. Webster. 1997. Field evaluation of RWA resistant barleys. Pg. 87 In Agronomy Abstracts, ASA, Madison, WI.