P. Nichols

Genetic improvement of subterranean clover (Trifolium subterraneum L.). 1. Germplasm, traits and future prospects

Abstract. Subterranean clover (Trifolium subterraneum L.) is the most widely sown annual pasture legume species in southern Australia, valued in the livestock and grains industries as a source of high-quality forage and for its ability to fix atmospheric nitrogen. From its initial accidental introduction into Australia in the 19th Century and subsequent commercialisation in the early 1900s, 45 cultivars have been registered in Australia. These consist of 32 cultivars of ssp. subterraneum, eight of ssp. yanninicum, and five of ssp. brachycalycinum and range in flowering time from 77 to 163 days from sowing, enabling the species to be grown in a diversity of rainfall environments, soil types, and farming systems. Eleven of these cultivars are introductions from the Mediterranean region, 15 are naturalised strains collected in Australia, 18 are the products of crossbreeding, and one is derived from mutagenesis. Cultivars developed in Italy have been commercialised for the local market, whereas other cultivars developed in Spain, Portugal, and France have not had commercial seed production. Important traits exploited include: (i) selection for low levels of the oestrogenic isoflavone formononetin, which causes reduced ewe fertility; (ii) increased levels of dormancy imposed by seed-coat impermeability (hard seeds) for cultivars aimed at crop rotations or unreliable rainfall environments; (iii) strong burr-burial ability to maximise seed production; (iv) resistance to important disease pathogens for cultivars aimed at medium- and high-rainfall environments, particularly to Kabatiella caulivora and root rot pathogens; (v) resistance to pests, particularly redlegged earth mites; and (vi) selection for unique leaf markings and other morphological traits (where possible) to aid cultivar identification. Cultivar development has been aided by a large genetic resource of ∼10 000 accessions, assembled from its centre of origin in the Mediterranean Basin, West Asia, and the Atlantic coast of Western Europe, in addition to naturalised strains collected in Australia. The development of a core collection of 97 accessions, representing almost 80% of the genetic diversity of the species, and a genetic map, provides a platform for development of future cultivars with new traits to benefit the livestock and grains industries. New traits being examined include increased phosphorous-use efficiency and reduced methane emissions from grazing ruminant livestock. Economic analyses indicate that future trait development should focus on traits contributing to increased persistence and autumn–winter productivity, while other potential traits include increased nutritive value (particularly of senesced material), increased N2 fixation ability, and tolerance to cheap herbicides. Beneficial compounds for animal and human health may also be present within the species for exploitation.

Variation in seed softening patterns and impact of seed production environment on hardseededness in early-maturing genotypes of subterranean clover

This paper describes variation in the dynamics of seed softening (loss of impermeability) in 20 early-maturing genotypes, including 6 cultivars, of subterranean clover (Trifolium subterraneum L.). It reports the effect of 3 sites of seed production in south-western Australia on the pattern of softening in the first summer–autumn and on total softening over the subsequent 2 years. Seeds were softened at a single field location and in a diurnally fluctuating cabinet (60°C/15°C). There was significant variation among genotypes in the pattern of seed softening over the first 5 months after senescence. Cultivars Nungarin, Dwalganup, and Geraldton softened most rapidly in late February, whereas cultivars Dalkeith, Urana, and Izmir softened most rapidly in late March. The duration of field exposure required in order for 50% of the first season’s softening to occur ranged from 44 to 108 days among the 20 genotypes. Persistence of hard seeds into the second and third years also varied among genotypes. Of the cultivars, Nungarin and Izmir had the highest levels of residual hard seed after 30 months (5.3% and 3.9%, respectively), whereas Dalkeith had the lowest (0.9%). Site of seed production had a small but significant effect on both the pattern of softening in the first summer–autumn and the persistence of hard seeds in subsequent years. Seeds produced in a relatively high-rainfall site (768 mm of growing-season rainfall plus supplementary irrigation) had a slower rate of hard seed breakdown than those from either of 2 sites located in the wheatbelt (217 and 423 mm growing-season rainfall). Seed softening through exposure in the field and in a 60°C/15°C fluctuating-temperature cabinet was compared for all genotypes. The cabinet treatment was fairly successful in ranking genotypes for relative between-season hardseededness, although it underestimated total softening by an average of 16%. However, the cabinet treatment was a poor predictor of the within-season pattern of seed softening.

Genetic improvement of subterranean clover (Trifolium subterraneum L.). 2. Breeding for disease and pest resistance

Abstract. Subterranean clover (Trifolium subterraneum L.) is the most widely sown pasture legume in southern Australia and resistance to important diseases and pests has been a major plant-breeding objective. Kabatiella caulivora, the cause of clover scorch, is the most important foliar fungal pathogen, and several cultivars have been developed with resistance to both known races. Screening of advanced breeding lines has been conducted to prevent release of cultivars with high susceptibility to other important fungal foliar disease pathogens, including rust (Uromyces trifolii-repentis), powdery mildew (Oidium sp.), cercospora (Cercospora zebrina) and common leaf spot (Pseudopeziza trifolii). Several oomycete and fungal species cause root rots of subterranean clover, including Phytophthora clandestina, Pythium irregulare, Aphanomyces trifolii, Fusarium avenaceum and Rhizoctonia solani. Most breeding efforts have been devoted to resistance to P. clandestina, but the existence of different races has confounded selection. The most economically important virus diseases in subterranean clover pastures are Subterranean clover mottle virus and Bean yellow mosaic virus, while Subterranean clover stunt virus, Subterranean clover red leaf virus (local synonym for Soybean dwarf virus), Cucumber mosaic virus, Alfalfa mosaic virus, Clover yellow vein virus, Beet western yellows virus and Bean leaf roll virus also cause losses. Genotypic differences for resistance have been found to several of these fungal, oomycete and viral pathogens, highlighting the potential to develop cultivars with improved resistance. The most important pests of subterranean clover are redlegged earth mite (RLEM) (Halotydeus destructor), blue oat mite (Penthaleus major), blue-green aphid (Acyrthosiphon kondoi) and lucerne flea (Sminthurus viridis). New cultivars have been bred with increased RLEM cotyledon resistance, but limited selection has been conducted for resistance to other pests. Screening for disease and pest resistance has largely ceased, but recent molecular biology advances in subterranean clover provide a new platform for development of future cultivars with multiple resistances to important diseases and pests. However, this can only be realised if skills in pasture plant pathology, entomology, pre-breeding and plant breeding are maintained and adequately resourced. In particular, supporting phenotypic disease and pest resistance studies and understanding their significance is critical to enable molecular technology investments achieve practical outcomes and deliver subterranean clover cultivars with sufficient pathogen and pest resistance to ensure productive pastures across southern Australia.

Coolamon subterranean clover (Trifolium subterraneum L. var. subterraneum)

Izmir is a hardseeded, early flowering, subterranean clover of var. subterraneum (Katz. et Morley) Zohary and Heller collected from Turkey and developed by the collaborating organisations of the National Annual Pasture Legume Improvement Program. It is a more hardseeded replacement for Nungarin and best suited to well-drained, moderately acidic soils in areas with a growing season of less than 4.5 months. Izmir seed production and regeneration densities in 3-year pasture phases were similar to Nungarin in 21 trials across southern Australia, but markedly greater in years following a crop or no seed set. Over all measurements, Izmir produced 10% more winter herbage and 7% more spring herbage than Nungarin. Its greater hardseededness and good seed production, makes it better suited to cropping rotations than Nungarin. Softening of Izmir hard seeds occurs later in the summer–autumn period than Nungarin, giving it slightly greater protection from seed losses following false breaks to the season. Izmir is recommended for sowing in Western Australia, New South Wales, Victoria, South Australia and Queensland. Izmir has been granted Plant Breeders Rights in Australia.

New sources of resistance in Trifolium subterraneum to rust (Uromyces trifolii-repentis)

Fifty-seven genotypes, including 10 cultivars, of Trifolium subterraneum var. subterraneum and var. yanninicum were screened in the field for resistance to rust (Uromyces trifolii-repentis) using artificial inoculation. There was outstanding resistance among the var. yanninicum, types with all but 1 genotype showing no rust symptoms. Several var. subterraneum genotypes also showed only a low rust incidence (≤3.5 on a 0–10 scale) with little or no leaf collapse from rust infection, including 83S19–07, CPI 103906F, EP132Sub-E, 84S20–02, and 84S20–01. Several other lines had a significant incidence of rust, while little leaf collapse from the disease was evident. Several highly susceptible lines were identified, including cultivars Green Range, Seaton Park and York, all with 100% of leaves affected by rust and extensive leaf collapse. There was excellent positive correlation between rust incidence and leaf collapse across the genotypes tested (R2 = 0.91). The excellent rust resistance observed in the majority of var. yanninicum lines and the good resistance in some var. subterraneum lines, indicates that these are useful sources of resistance that can be exploited, either directly as new cultivars to minimise leaf collapse from this disease or as parents in breeding programmes to develop more rust-resistant cultivars.

Coolamon subterranean clover (Trifolium subterraneumL. var.subterraneum)

Coolamon is a mid-season to late-season flowering F4-derived crossbred subterranean clover of var. subterraneum, developed by the collaborating organisations of the National Annual Pasture Legume Improvement Program. It is a replacement for Junee and has been selected for release on the basis of its greater herbage production and persistence, and its resistance to both known races of clover scorch. Coolamon is recommended for sowing in Western Australia, New South Wales, Victoria, South Australia and Queensland. It is best suited to well-drained, moderately acidic soils in areas with a growing season of 6.5–8 months that extends into November. Coolamon is best suited to phase farming and permanent pasture systems. It can also be used in cropping rotations, but at least 2 years of pasture are required between crops. Coolamon has been granted Plant Breeders Rights in Australia.

Napier subterranean clover (Trifolium subterraneum L. var. yanninicum)

Napier is a late flowering F6-derived crossbred subterranean clover of var. yanninicum [(Katz. et Morley) Zohary and Heller] developed by the collaborating organisations of the National Annual Pasture Legume Improvement Program. It is a replacement for both Larisa and Meteora and has been selected for release on the basis of its greater herbage and seed production and disease resistance to both known races of clover scorch and 2 of the common races of Phytophthora root rot. Napier is recommended for sowing in Victoria, Western Australia, New South Wales, and South Australia. It is best suited to moderately acidic soils prone to water-logging and to loamy and clay soils with good water-holding capacity in areas with a minimum growing season length of 7.5 months, which extends into late November. Napier is well adapted to the permanent pasture systems found in the areas in which it will be grown. Its upright, vigorous growth makes it well suited to grazing by cattle or sheep and to fodder conservation. Napier has been granted Plant Breeders Rights in Australia.