Vulnerability of Barley to African Pathotypes of Puccinia graminis f. sp. tritici and Sources of Resistance.

The emergence of widely virulent pathotypes (e.g., TTKSK in the Ug99 race group) of the stem rust pathogen (Puccinia graminis f. sp. tritici) in Africa threatens wheat production on a global scale. Although intensive research efforts have been advanced to address this threat in wheat, few studies have been conducted on barley, even though pathotypes such as TTKSK are known to attack the crop. The main objectives of this study were to assess the vulnerability of barley to pathotype TTKSK and identify possible sources of resistance. From seedling evaluations of more than 1,924 diverse cultivated barley accessions to pathotype TTKSK, more than 95% (1,844) were found susceptible. A similar high frequency (910 of 934 = 97.4%) of susceptibility was found for the wild progenitor (Hordeum vulgare subsp. spontaneum) of cultivated barley. Additionally, 55 barley lines with characterized or putative introgressions from various wild Hordeum spp. were also tested against pathotype TTKSK but none was found resistant. In total, more than 96% of the 2,913 Hordeum accessions tested were susceptible as seedlings, indicating the extreme vulnerability of the crop to the African pathotypes of P. graminis f. sp. tritici. In total, 32 (1.7% of accessions evaluated) and 13 (1.4%) cultivated and wild barley accessions, respectively, exhibited consistently highly resistant to moderately resistant reactions across all experiments. Molecular assays were conducted on these resistant accessions to determine whether they carried rpg4/Rpg5, the only gene complex known to be highly effective against pathotype TTKSK in barley. Twelve of the 32 (37.5%) resistant cultivated accessions and 11 of the 13 (84.6%) resistant wild barley accessions tested positive for a functional Rpg5 gene, highlighting the narrow genetic base of resistance in Hordeum spp. Other resistant accessions lacking the rpg4/Rpg5 complex were discovered in the evaluated germplasm and may possess useful resistance genes. Combining rpg4/Rpg5 with resistance genes from these other sources should provide more durable resistance against the array of different virulence types in the Ug99 race group.

Genetic Loci Conditioning Adult Plant Resistance to the Ug99 Race Group and Seedling Resistance to Races TRTTF and TTTTF of the Stem Rust Pathogen in Wheat Landrace CItr 15026.

Wheat landrace CItr 15026 previously showed adult plant resistance (APR) to the Ug99 stem rust race group in Kenya and seedling resistance to Puccinia graminis f. sp. tritici races QFCSC, TTTTF, and TRTTF. CItr 15026 was crossed to susceptible accessions LMPG-6 and Red Bobs, and 180 double haploid (DH) lines and 140 recombinant inbred lines (RIL), respectively, were developed. The 90K wheat iSelect single-nucleotide polymorphism platform was used to genotype the parents and populations. Parents and 180 DH lines were evaluated in the field in Kenya for three seasons. A major quantitative trait locus (QTL) for APR was consistently detected on chromosome arm 6AS. This QTL was further detected in the RIL population screened in Kenya for one season. Parents, F1, and the two populations were tested as seedlings against races TRTTF and TTTTF. In addition, the DH population was tested against race QFCSC. Goodness-of-fit tests indicated that the TRTTF resistance in CItr 15026 was controlled by two complementary genes whereas the TTTTF and QFCSC resistance was conditioned by one dominant gene. The TRTTF resistance loci mapped to chromosome arms 6AS and 6DS, whereas the TTTTF and QFCSC resistance locus mapped to the same region on 6DS as the TRTTF resistance. The APR identified in CItr 15026 should be useful in developing cultivars with durable stem rust resistance.

Molecular Mapping of Stem Rust Resistance Loci Effective Against the Ug99 Race Group of the Stem Rust Pathogen and Validation of a Single Nucleotide Polymorphism Marker Linked to Stem Rust Resistance Gene Sr28.

Wheat landrace PI 177906 has seedling resistance to stem rust caused by Puccinia graminis f. sp. tritici races TTKSK, TTKST, and BCCBC and field resistance to the Ug99 race group. Parents, 140 recombinant inbred lines, and 138 double haploid (DH) lines were evaluated for seedling resistance to races TTKSK and BCCBC. Parents and the DH population were evaluated for field resistance to Ug99 in Kenya. The 90K wheat single nucleotide polymorphism (SNP) genotyping platform was used to genotype the parents and populations. Goodness-of-fit tests indicated that two dominant genes in PI 177906 conditioned seedling resistance to TTKSK. Two major loci for seedling resistance were consistently mapped to the chromosome arms 2BL and 6DS. The BCCBC resistance was mapped to the same location on 2BL as the TTKSK resistance. Using field data from the three seasons, two major QTL were consistently detected at the same regions on 2BL and 6DS. Based on the mapping result, race specificity, and the infection type observed in PI 177906, the TTKSK resistance on 2BL is likely due to Sr28. One SNP marker (KASP_IWB1208) was found to be predictive for the presence of the TTKSK resistance locus on 2BL and Sr28.

Genetic mapping of resistance to the Ug99 race group of Puccinia graminis f. sp. tritici in a spring wheat landrace CItr 4311.

KEY MESSAGE: A gene for Ug99 resistance from wheat landrace CItr 4311 was detected on the long arm of chromosome 2B. Wheat landrace CItr 4311 has seedling resistance to stem rust caused by Puccinia graminis f. sp. tritici race TTKSK and field resistance to the Ug99 race group. Parents, F1 seedlings, 121 doubled haploid (DH) lines, and 124 recombinant inbred lines (RILs) developed from a cross between CItr 4311 and the susceptible line LMPG-6 were evaluated for seedling resistance to race TTKSK. Goodness-of-fit tests indicated that a single dominant gene in CItr 4311 conditioned the TTKSK resistance. The 90 K wheat iSelect SNP platform was used to genotype parents and the DH population. The seedling resistance locus was mapped to the chromosome arm 2BL. Parents and the DH population were evaluated for field resistance in Kenya. One major QTL for the field resistance was consistently detected in the same region on 2BL as the seedling resistance. Using KASP assays, five linked SNP markers were used to verify the result in the 124 RIL, 35 wheat accessions, 46 DH lines from the LMPG-6/PI 165194 cross and F1 seedlings, and susceptible bulks derived from crosses between six resistant landraces with LMPG-6. Race specificity, mapping results, and haplotype similarity with lines with Sr9h (Gabo 56, Timstein, and PI 670015), support the hypothesis that the Sr gene in CItr 4311 and the landraces is Sr9h. The KASP assays developed in this study will be useful for pyramiding the TTKSK resistance from CItr 4311 with other Sr genes effective against Ug99.

Association mapping of North American spring wheat breeding germplasm reveals loci conferring resistance to Ug99 and other African stem rust races.

BACKGROUND: The recently identified Puccinia graminis f. sp. tritici (Pgt) race TTKSK (Ug99) poses a severe threat to global wheat production because of its broad virulence on several widely deployed resistance genes. Additional virulences have been detected in the Ug99 group of races, and the spread of this race group has been documented across wheat growing regions in Africa, the Middle East (Yemen), and West Asia (Iran). Other broadly virulent Pgt races, such as TRTTF and TKTTF, present further difficulties in maintaining abundant genetic resistance for their effective use in wheat breeding against this destructive fungal disease of wheat. In an effort to identify loci conferring resistance to these races, a genome-wide association study was carried out on a panel of 250 spring wheat breeding lines from the International Maize and Wheat Improvement Center (CIMMYT), six wheat breeding programs in the United States and three wheat breeding programs in Canada. RESULTS: The lines included in this study were grouped into two major clusters, based on the results of principal component analysis using 23,976 SNP markers. Upon screening for adult plant resistance (APR) to Ug99 during 2013 and 2014 in artificial stem rust screening nurseries at Njoro, Kenya and at Debre Zeit, Ethiopia, several wheat lines were found to exhibit APR. The lines were also screened for resistance at the seedling stage against races TTKSK, TRTTF, and TKTTF at USDA-ARS Cereal Disease Laboratory in St. Paul, Minnesota; and only 9 of the 250 lines displayed seedling resistance to all the races. Using a mixed linear model, 27 SNP markers associated with APR against Ug99 were detected, including markers linked with the known APR gene Sr2. Using the same model, 23, 86, and 111 SNP markers associated with seedling resistance against races TTKSK, TRTTF, and TKTTF were identified, respectively. These included markers linked to the genes Sr8a and Sr11 providing seedling resistance to races TRTTF and TKTTF, respectively. We also identified putatively novel Sr resistance genes on chromosomes 3B, 4D, 5A, 5B, 6A, 7A, and 7B. CONCLUSION: Our results demonstrate that the North American wheat breeding lines have several resistance loci that provide APR and seedling resistance to highly virulent Pgt races. Using the resistant lines and the SNP markers identified in this study, marker-assisted resistance breeding can assist in development of varieties with elevated levels of resistance to virulent stem rust races including TTKSK.

Mapping resistance to the Ug99 race group of the stem rust pathogen in a spring wheat landrace.

KEY MESSAGE: A new gene for Ug99 resistance from wheat landrace PI 374670 was detected on the long arm of chromosome 7A. Wheat landrace PI 374670 has seedling and field resistance to stem rust caused by Puccinia graminis f. sp tritici Eriks. & E. Henn (Pgt) race TTKSK. To elucidate the inheritance of resistance, 216 BC1F2 families, 192 double haploid (DH) lines, and 185 recombinant inbred lines (RILs) were developed by crossing PI 374670 and the susceptible line LMPG-6. The parents and progeny were evaluated for seedling resistance to Pgt races TTKSK, MCCFC, and TPMKC. The DH lines were tested in field stem rust nurseries in Kenya and Ethiopia. The DH lines were genotyped with the 90K wheat iSelect SNP genotyping platform. Goodness-of-fit tests indicated that a single dominant gene in PI 374670 conditioned seedling resistance to the three Pgt races. The seedling resistance locus mapped to the long arm of chromosome 7A and this result was verified in the RIL population screened with the flanking SNP markers using KASP assays. In the same region, a major QTL for field resistance was detected in a 7.7 cM interval and explained 34-54 and 29-36% of the variation in Kenya and Ethiopia, respectively. Results from tests with specific Pgt races and the csIH81 marker showed that the resistance was not due to Sr22. Thus, a new stem rust resistance gene or allele, either closely linked or allelic to Sr15, is responsible for the seedling and field resistance of PI 374670 to Ug99.

Virulence Characterization of International Collections of the Wheat Stripe Rust Pathogen, Puccinia striiformis f. sp. tritici.

Wheat stripe rust (yellow rust [Yr]), caused by Puccinia striiformis f. sp. tritici, is an economically important disease of wheat worldwide. Virulence information on P. striiformis f. sp. tritici populations is important to implement effective disease control with resistant cultivars. In total, 235 P. striiformis f. sp. tritici isolates from Algeria, Australia, Canada, Chile, China, Hungary, Kenya, Nepal, Pakistan, Russia, Spain, Turkey, and Uzbekistan were tested on 20 single Yr-gene lines and the 20 wheat genotypes that are used to differentiate P. striiformis f. sp. tritici races in the United States. The 235 isolates were identified as 129 virulence patterns on the single-gene lines and 169 virulence patterns on the U.S. differentials. Virulences to YrA, Yr2, Yr6, Yr7, Yr8, Yr9, Yr17, Yr25, YrUkn, Yr28, Yr31, YrExp2, Lemhi (Yr21), Paha (YrPa1, YrPa2, YrPa3), Druchamp (Yr3a, YrD, YrDru), Produra (YrPr1, YrPr2), Stephens (Yr3a, YrS, YrSte), Lee (Yr7, Yr22, Yr23), Fielder (Yr6, Yr20), Tyee (YrTye), Tres (YrTr1, YrTr2), Express (YrExp1, YrExp2), Clement (Yr9, YrCle), and Compair (Yr8, Yr19) were detected in all countries. At least 80% of the isolates were virulent on YrA, Yr2, Yr6, Yr7, Yr8, Yr17, YrUkn, Yr31, YrExp2, Yr21, Stephens (Yr3a, YrS, YrSte), Lee (Yr7, Yr22, Yr23), and Fielder (Yr6, Yr20). Virulences to Yr1, Yr9, Yr25, Yr27, Yr28, Heines VII (Yr2, YrHVII), Paha (YrPa1, YrPa2, YrPa3), Druchamp (Yr3a, YrD, YrDru), Produra (YrPr1, YrPr2), Yamhill (Yr2, Yr4a, YrYam), Tyee (YrTye), Tres (YrTr1, YrTr2), Hyak (Yr17, YrTye), Express (YrExp1, YrExp2), Clement (Yr9, YrCle), and Compair (Yr8, Yr19) were moderately frequent (>20 to <80%). Virulence to Yr10, Yr24, Yr32, YrSP, and Moro (Yr10, YrMor) was low (≤20%). Virulence to Moro was absent in Algeria, Australia, Canada, Kenya, Russia, Spain, Turkey, and China, but 5% of the Chinese isolates were virulent to Yr10. None of the isolates from Algeria, Canada, China, Kenya, Russia, and Spain was virulent to Yr24; none of the isolates from Algeria, Australia, Canada, Nepal, Russia, and Spain was virulent to Yr32; none of the isolates from Australia, Canada, Chile, Hungary, Kenya, Kenya, Nepal, Pakistan, Russia, and Spain was virulent to YrSP; and none of the isolates from any country was virulent to Yr5 and Yr15. Although the frequencies of virulence factors were different, most of the P. striiformis f. sp. tritici isolates from these countries shared common virulence factors. The virulences and their frequencies and distributions should be useful in breeding stripe-rust-resistant wheat cultivars and understanding the pathogen migration and evolution.

Field Resistance to the Ug99 Race Group of the Stem Rust Pathogen in Spring Wheat Landraces.

Wheat landraces provide a source of genetic variability for breeding. The emergence and spread of highly virulent races of the stem rust pathogen (Ug99 race group of Puccinia graminis f. sp. tritici) threaten wheat production globally. Spring wheat landraces were screened for resistance in eight field seasons at the Kenya Agricultural Research Institute, Njoro, where the Ug99 race group has become endemic. Accessions showing resistance in one season were retested and screened with molecular markers associated with resistance genes Sr2, Sr24, Sr36, and Lr34/Yr18; two height-reducing genes; and a photoperiod insensitivity allele. Of 2,509 accessions tested, 278 were categorized as resistant based on results from at least two seasons. Of these resistant accessions, 32 were positive for one or more markers for Sr2, Sr36, Rht-B1b, or Rht-D1b, indicating that they do not fit the definition of "landrace" because these genes were likely introduced via modern breeding practices. Thus, 246 resistant "landrace" accessions were identified. Of countries with more than five tested accessions, Afghanistan, Iran, Portugal, Ethiopia, Uzbekistan, Greece, Tajikistan, Bosnia and Herzegovina, and Serbia had at least 10% of tested accessions that were resistant to the Ug99 race group. Future research will characterize the resistance to determine its novelty and incorporate novel genes into improved lines.

Sources of Resistance to Stem Rust Race Ug99 in Spring Wheat Germplasm.

Wheat stem rust (Puccinia graminis f. sp. tritici) race TTKSK (Ug99), with virulence to the majority of the world's wheat (Triticum aestivum) cultivars, has spread from Uganda throughout eastern Africa, Yemen, and Iran. The identification and spread of variants of race TTKSK with virulence to additional stem rust resistance genes has reminded breeders and pathologists of the danger of deploying major resistance genes alone. In order to protect wheat from this rapidly spreading and adapting pathogen, multiple resistance genes are needed, preferably from improved germplasm. Preliminary screening of over 700 spring wheat breeding lines and cultivars developed at least 20 years ago identified 88 accessions with field resistance to Ug99. We included these resistant accessions in the stem rust screening nursery in Njoro, Kenya for two additional seasons. The accessions were also screened with a bulk of North American isolates of P. graminis f. sp. tritici in the field in St. Paul, MN. In order to further characterize the resistance in these accessions, we obtained seedling phenotypes for 10 races of P. graminis f. sp. tritici, including two races from the race TTKSK complex. This phenotyping led to the identification of accessions with either adult-plant or all-stage resistance to race TTKSK, and often North American races of P. graminis f. sp. tritici as well. These Ug99 resistant accessions can be obtained by breeders and introgressed into current breeding germplasm.

Foliar Fungicides to Control Wheat Stem Rust, Race TTKS (Ug99), in Kenya.

The emergence of a new virulent race of stem rust (Puccinia graminis f. sp. tritici) of wheat (Triticum aestivum), TTKS (Ug99), calls for urgent measures to contain the disease, which is a serious threat to wheat production in Kenya. Nine commercial fungicides were evaluated for control of stem rust in the field in three locations in 2005 and 2006. Ten treatments, including untreated control, were applied on field plots in a randomized complete block design with four replications. Assessment of stem rust severity was done at growth stage (GS) 55 (heading), GS 65 (flowering), and GS 75 (late milk) and expressed in terms of mean rust severity and area under the disease progress curve (AUDPC). In general, fungicide treatment significantly (P ≤ 0.05) reduced rust severity, AUDPC, and losses in grain yield and 1,000-kernel weight for cv. Duma compared with the untreated control. Stem rust was severe in both years across all sites, resulting in significantly less grain yield in the control treatment. The 1,000-kernel weight was significantly less at two locations in 2006. Fungicides azoxystrobin at 200 g/liter + cyproconazole at 80 g/liter (AmistarXtra 280 SC), tebuconazole (Folicur 250 EC), tebuconazole + tridimenol (Silvacur 375 EC), and tebuconazole (Orius 25 EW) were more effective in reducing stem rust across the sites. Fungicides trifloxystrobin + propiconazole (Stratego 250 EC), hexaconazole (Cotaf 5 EC), epoxiconazole + carbendazim (Swing 250 EC), cyproconazole at 80 g/liter + propiconazole at 250 g/liter (Artea 330EC), and epoxiconazole at 125 g/liter + carbendazim at 125 g/liter (Soprano C 250 EC) were inconsistent in the reduction of the disease. The results suggest that fungicides can be used effectively in reducing stem rust severity and increasing yield of susceptible wheat cultivars in stem-rust-prone areas.

Detection of Virulence to Resistance Gene Sr36 Within the TTKS Race Lineage of Puccinia graminis f. sp. tritici.

The stem rust resistance gene Sr36 confers a near-immune resistance reaction to many races of Puccinia graminis f. sp. tritici and is highly effective against race TTKSK (syn. Ug99), which possesses unusually broad virulence combinations. Because this gene is widely used in United States soft winter wheat germplasm and cultivars, it has been considered to be an important source of resistance to TTKSK. In 2007, moderately susceptible infection responses were observed on wheat lines and cultivars carrying Sr36 in a field screening nursery for stem rust at Njoro, Kenya. We derived 18 single-pustule isolates from stem rust samples collected from the 2007 Njoro nursery. The isolates were evaluated for virulence on 20 North American stem rust differential lines and on wheat lines and cultivars carrying Sr36, Sr31+Sr36, and Sr24+Sr31. Of the 18 isolates, 10 produced infection types 3+ to 4 on line W2691SrTt-1 (monogenic for Sr36) and other lines that carry Sr36 and belonged to a new virulence phenotype that was not detected in previous years. These isolates were identified as race TTTSK. The remaining eight isolates were identified as races TTKSK (five isolates) and TTKST (three isolates), with avirulence and virulence, respectively, to Sr24. Thirteen simple sequence repeat (SSR) markers were used to examine the genetic relationships among the three races in the TTKS lineage. All isolates in the lineage shared an identical SSR genotype and were clearly different from North American races. In all, 16 wheat cultivars and 60 elite breeding lines, postulated to possess Sr36, were susceptible to race TTTSK. The occurrence of race TTTSK with combined virulence on Sr31 and Sr36 has further broadened the virulence spectrum of the TTKS lineage and rendered an important source of resistance ineffective.

Characterization of Seedling Infection Types and Adult Plant Infection Responses of Monogenic Sr Gene Lines to Race TTKS of Puccinia graminis f. sp. tritici.

Stem rust, caused by Puccinia graminis f. sp. tritici, historically was one of the most destructive diseases of wheat and barley. The disease has been under effective control worldwide through the widespread use of host resistance. A number of stem rust resistance genes in wheat have been characterized for their reactions to specific races of P. graminis f. sp. tritici. Adult plant responses to race TTKS (also known as Ug99) of monogenic lines for Sr genes, a direct measurement of the effectiveness for a given gene, have not been investigated to any extent. This report summarizes adult plant infection responses and seedling infection types for monogenic lines of designated Sr genes challenged with race TTKS. High infection types at the seedling stage and susceptible infection responses in adult plants were observed on monogenic lines carrying Sr5, 6, 7a, 7b, 8a, 8b, 9a, 9b, 9d, 9g, 10, 11, 12, 15, 16, 17, 18, 19, 20, 23, 30, 31, 34, 38, and Wld-1. Monogenic lines of resistance genes Sr13, 22, 24, 25, 26, 27, 28, 32, 33, 35, 36, 37, 39, 40, 44, Tmp, and Tt-3 were effective against TTKS both at the seedling and adult plant stages. The low infection types to race TTKS observed for these resistance genes corresponded to the expected low infections of these genes to other incompatible races of P. graminis f. sp. tritici. The level of resistance conferred by these genes at the adult plant stage varied between highly resistant to moderately susceptible. The results from this study were inconclusive for determining the effectiveness of resistance genes Sr9e, 14, 21, and 29 against race TTKS. The understanding of the effectiveness of individual Sr genes against race TTKS will facilitate the utilization of these genes in breeding for stem rust resistance in wheat.

The Spread of Stem Rust Caused by Puccinia graminis f. sp. tritici, with Virulence on Sr31 in Wheat in Eastern Africa.

Stem rust resistance in wheat cultivars with Sr31 has been effective and durable worldwide for more than 30 years. Isolates of Puccinia graminis f. sp. tritici with virulence to Sr31 were detected in Uganda in 1999 (1). During 2003 and 2004, a majority of current Kenyan cultivars and a large portion of CIMMYT wheat germplasm with gene Sr31 planted in Kenya were susceptible to stem rust. Six isolates collected during 2004 at different locations in Kenya were tested for virulence on the 16 North American stem rust race differentials with the following Sr genes: Sr5, 6, 7b, 8a, 9a, 9b, 9d, 9e, 9g, 10, 11, 17, 21, 30, 36, and Tmp. An extended set of designated Sr genes (Sr13, 19, 22, 24, 25, 26, 29, 31, 32, 33, 35, 37, 39, 40, 44, and Wld-1) was also tested at the seedling stage. An isolate from Uganda collected in 1999 with virulence on Sr31 was used for comparison. Urediniospores suspended in a lightweight mineral oil were inoculated onto 7-day-old seedlings. Inoculated plants were placed in a dew chamber for 14 h at 18°C in the dark and then for an additional period of 3 to 4 h placed under fluorescent light. Plants were incubated in a greenhouse at 18 ± 2°C with a photoperiod of 16 h. Infection types (IT), described by Stakman et al. (3), were assessed after 14 days postinoculation. All isolates from Kenya exhibited a low infection type (IT 0) on line W2691SrTt-1 (donor of Sr36), a low infection type (IT 2) on cv. Triumph 64 (donor of SrTmp), and high infection types (IT 3 or 4) on all other lines in the differential set (2); thus these isolates were keyed to race TTKS. The virulence pattern of the isolate collected in 1999 from Uganda was identical to that from Kenya on the differential set and on the extended set of designated Sr genes. In this study, these isolates produced a high infection type (IT 3) on Einkorn and CnSSr21Tm (a derivative of Triticum monococcum in Chinese Spring background), two sources of Sr21 used in our study, whereas the isolate with Sr31-virulence from Uganda in 1999 was reported to be avirulent on Sr21 (1). These isolates produced high infection types on single gene lines with Sr31 and winter wheat cvs. Custer, Foster, GA-Dozier, Patton, and Pioneer 26R61, which were known to carry the 1BL.1RS translocation with Sr31. These isolates were also virulent on SrWld-1, a gene used in spring wheat for its resistance to North American stem rust isolates. In addition to Sr36 and SrTmp, other stem rust resistance genes that were effective against TTKS at the seedling stage include Sr13, 22, 24, 25, 26, 27, 29, 32, 33, 35, 37, 39, 40, and 44. Cultivars, breeding germplasm, and single gene lines are currently being evaluated for adult plant reaction in Kenya. Results from this study indicated that stem rust isolates with virulence on Sr31 are now wide spread in the Eastern Africa highlands and pose a threat to wheat production in the region, as well as in other wheat production areas where Sr31 resistance is important. A rapid deployment of effective resistance genes to this race in breeding programs throughout Eastern Africa and Asia is needed to reduce this threat. References: (1) Z. A. Pretorius et al. Plant Dis. 84:203, 2000. (2) A. P. Roelfs and J. W. Martens. Phytopathology 78:526, 1988. (3) E. C. Stakman et al. U.S. Department of Agriculture. ARS E-617, 1962.