Genome size and GC content evolution of Festuca: ancestral expansion and subsequent reduction

Background and Aims: Plant evolution is well known to be frequently associated withremarkable changes in genome size and composition; however,the knowledge of long-term evolutionary dynamics of these processesstill remains very limited. Here a study is made of the finedynamics of quantitative genome evolution in Festuca (fescue),the largest genus in Poaceae (grasses). Methods: Using flow cytometry (PI, DAPI), measurements were made of DNAcontent (2C-value), monoploid genome size (Cx-value), averagechromosome size (C/n-value) and cytosine + guanine (GC) contentof 101 Festuca taxa and 14 of their close relatives. The resultswere compared with the existing phylogeny based on ITS and trnL-Fsequences. Key Results: The divergence of the fescue lineage from related Poeae waspredated by about a 2-fold monoploid genome and chromosome sizeenlargement, and apparent GC content enrichment. The backwardreduction of these parameters, running parallel in both mainevolutionary lineages of fine-leaved and broad-leaved fescues,appears to diverge among the existing species groups. The mostdramatic reductions are associated with the most recently andrapidly evolving groups which, in combination with recent intraspecificgenome size variability, indicate that the reduction processis probably ongoing and evolutionarily young. This dynamicsmay be a consequence of GC-rich retrotransposon proliferationand removal. Polyploids derived from parents with a large genomesize and high GC content (mostly allopolyploids) had smallerCx- and C/n-values and only slightly deviated from parentalGC content, whereas polyploids derived from parents with smallgenome and low GC content (mostly autopolyploids) generallyhad a markedly increased GC content and slightly higher Cx-and C/n-values. Conclusions: The present study indicates the high potential of general quantitativecharacters of the genome for understanding the long-term processesof genome evolution, testing evolutionary hypotheses and theirusefulness for large-scale genomic projects. Taken together,the results suggest that there is an evolutionary advantagefor small genomes in Festuca.     

Mounding as a Technique for Restoration of Prairie on a Capped Landfill in the Puget Sound Lowlands

Closed landfills create large open spaces that are often proposed as sites for restored or created ecosystems. Grasslands are probably prescribed most often because of the presumption that grass root systems will not breach the landfill cap. Capped landfills have a number of soil degradation problems, including compaction, decreased permeability, lack of organic material, diminished soil fauna, inappropriate texture, and lack of structure. In this study in the Puget Sound lowlands, Washington, U.S.A., mounding (low sandy‐loam mounds, about 20 cm high and 2 m in diameter), addition of fertilizer, and mulching with yard‐waste compost were applied to landfill sites as treatments in a factorial‐design experiment. Prairie plants (1,344 individuals, 7 species) were planted into 4‐m² plots (n = 48), and plant growth and survival and the increase in weed biomass were monitored for 3 years. Mulching had no effect on plant survival or growth. Fertilization had a negative effect on Lupinus lepidus, a nitrogen‐fixing species. Mounding had a positive effect on growth and survival of Eriophyllum lanatum, Festuca idahoensis, and Aster curtus. Potentilla pacifica was indifferent to mounding, and Carex inops responded negatively. Mounds should probably be used as one element of a complex of habitats on restored landfills.     

Genome mapping of polyploid tall fescue (Festuca arundinacea Schreb.) with RFLP markers

Genetic mapping using molecular markers such as restriction fragment length polymorphisms (RFLPs) has become a powerful tool for plant geneticists and breeders. Like many economically important polyploid plant species, detailed genetic studies of hexaploid tall fescue (Festuca arundinacea Schreb.) are complicated, and no genetic map has been established. We report here the first tall fescue genetic map. This map was generated from an F₂ population of HD28-56 by Kentucky-31 and contains 108 RFLP markers. Although the two parental plants were heterozygous, the perennial and tillering growth habit, high degree of RFLP, and disomic inheritance of tall fescue enabled us to identify the segregating homologous alleles. The map covers 1274 cM on 19 linkage groups with an average of 5 loci per linkage group (LG) and 17.9 cM between loci. Mapping the homoeologous loci detected by the same probe allowed us to identify five homoeologous groups within which the gene orders were found to be generally conserved among homoeologous chromosomes. An exception was homoeologous group 5, in which only 2 of the 3 homoeologous chromosomes were identified. Using 12 genome-specific probes, we were able to assign several linkage groups to one of the three genomes (PG₁G₂) in tall fescue. All the loci detected by the 11 probes specific to the G₁ and/or G₂ genomes, with one exception, identified loci located on 4 chromosomes of two homoeologous groups (LG2a, LG2c, LG3a, and LG3c). A P-genome-specific probe was used to map a locus on LG5c. Comparative genome mapping with maize probes indicated that homoeologous group 3 and 2 chromosomes in tall fescue corresponded to maize chromosome 1. Difficulties and advantages of applying RFLP technology in polyploids with high levels of heterozygosity are discussed.Journal Series No. 12, 190     

Aspergillus terreus and fescue toxicosis

The incidence ofAspergillus terreus recovered fromAcremonium coenophialum-infected and non-infected tall fescue grass and from the rumens of heifers grazing on the grasses was determined. The recovery ofA. terreus fromA. coenophialum-infected grass was similar to that from non-infected grass. The same was true of the recovery from the rumens of heifers on infected and non-infected grass. All heifers grazing onA. coenophialum-infected grass showed symptoms of the summer syndrome manifestation of fescue toxicosis while those grazing on non-infected grass did not;A. terreus is not a factor in fescue toxicosis in cattle. Hatch Project #630, Alabama Agricultural Experiment Station, Auburn, Alabama 36849. AAES Journal No. 18-881477P.     

Mealybug, Phenococcus solani, and barley aphid, Sipha maydis, response to endophyte-infected tall and meadow fescues

Mealybugs and aphids are insects which damage grass species. The effects of fungal endophytes on the feeding of the mealybug, Phenococcus solani Ferris (Homoptera: Pseudococcidae), and barley aphid, Sipha maydis Passerini (Homoptera: Aphididae), on tall fescue, Festuca arundinacea Schreb. and meadow fescue, Festuca pratensis Huds., were studied under greenhouse conditions. Mealybugs preferred endophyte‐free (E–) clones over their endophyte‐infected (E+) counterparts. E+ plants had a significantly lower number of mealybugs than E– plants. A mixture of E+ and E– plants supported intermediate mealybug numbers, between pure plantings of E+ and E– grasses. Barley aphids released on to plant materials were deterred from feeding and could not persist on E+ plants. E– plants did not survive because of aphid damage, while E+ plants generally re‐grew, but were damaged to some degree. The results showed that the use of pure stands of endophyte‐infected grasses or a mixed stand of infected and non‐infected plants may increase the persistence and durability of turf and forage grass species in the presence of foliar damaging insects.     

Agrobacterium-mediated high efficiency transformation of tall fescue (Festuca arundinacea)

Tall fescue (Festuca arundinacea) is the predominant cool-season pasture grass in the USA. Embryogenic calluses were induced from seeds/caryopsis of elite tall fescue cultivars Jesup and Kentucky-31, and were broken up into small pieces and used for Agrobacterium tumefaciens-mediated transformation. Agrobacterium strains LBA4404 and EHA105 harboring pCAMBIA vectors or the super-binary vector pTOK233 were used to infect the embryogenic callus pieces. The number of hygromycin resistant calluses obtained per dish of infected callus pieces was in the range of 2.0-5.8, and the number of transgenic plants recovered per dish of infected callus pieces was in the range of 0.4-1.7. When transformation efficiency was calculated based on the number of transgenic plants recovered and the number of original intact calluses used, the transformation frequency was in the range of 1.9-8.7%. The use of the easily available pCAMBIA vectors could produce equivalent results as the superbinary vector pTOK233. The transgenic nature of the regenerated plants was demonstrated by Southern hybridization analysis using undigested and digested genomic DNA samples. Expression of the transgenes was confirmed by northern hybridization analysis, GUS staining, and detection of GFP signals. Fertile transgenic plants were obtained after vernalization in the greenhouse. Progeny analysis revealed Mendelian inheritance of the transgenes.     

Endophyte–grass complexes and the relationship between feeding preference and performance in a grass herbivore

Overseeding non‐endophytic turfgrass lawns with endophytic turfgrass is considered promising for the development of integrated pest management programs in urban landscapes. In this context, a better understanding of the variability in the preference–performance response of insect herbivores exposed to endophytic grasses could be useful to develop efficient practices. Specifically, while endophytic varieties that are strongly toxic and avoided could provide good control of mobile pests, varieties that are toxic but not avoided could be more suitable for the control of sedentary pests. In this laboratory study, we investigated how the infection of 10 grass varieties by Neotyphodium endophytes affected the feeding performance and preference of newly hatched nymphs of the hairy chinch bug, Blissus leucopterus hirtus Montandon (Heteroptera: Lygaeidae), a common turfgrass pest in north‐eastern USA. We found that endophytes generally induced a strong relationship between feeding performance and preference in this herbivore. However, two endophytic varieties did not conform to this relationship, with one variety being highly toxic but not avoided and the other less toxic but highly avoided. These results provide a solid basis to further explore the impacts of endophytic grasses on the dispersal and survival of insect pests in mixed stands of endophytic and non‐endophytic plants.     

Species variation in the fixation and transfer of nitrogen from legumes to associated grasses

Summary Three legume species (alfalfa, red clover, and birdsfoot trefoil) in combination with five grass species (timothy, bromegrass, red fescue, tall fescue, and orchardgrass) were used to study N transfer in mixtures, using the 15_N dilution technique. The advantage of grass-legume mixtures was apparent. Total herbage and protein yields of grasses in mixtures were higher than those alone, especially at the later cuts. This benefit of mixed cropping is mainly due to N transfer from legumes to associated grasses. N₂-fixation and N transfer by alfalfa rated highest, red clover intermediate, and birdsfoot trefoil lowest. The importance of each pathway of N transfer from legumes appeared to differ between species. Alfalfa and red clover excreted more N than trefoil, while the latter contributed more N from decomposition of dead nodule and root tissue. The greatest advantage from a grass-legume mixture, with respect to the utilization of N released from the legume, varied with early maturing tall fescue (Kentucky 31), orchardgrass (Juno), and bromegrass (Tempo), to intermediate timothy (Climax), and least with late maturing red fescue (Carlawn). Contribution no. 817 of the Ottawa Research Station.     

Dynamics of nitrogen remobilization in defoliated Phleum pratense and Festuca pratensis under short and long photoperiods

The impact of photoperiod on the rate and magnitude of N remobilization relative to uptake of inorganic N during the recovery of shoot growth after a severe defoliation was compared over 18 days in two temperate grass species, timothy (Phleum pratense L. cv. Bodin) and meadow fescue (Festuca pratensis Huds. cv. Salten). Plants were grown in flowing solution culture with N supplied as 20 mM NH₄NO₃ and pre-treated by extending the 11 h photosynthetically significant light period either by 1 h (short-day or SD plants) or 7 h (long-day or LD plants) of very low light intensity, during the 10 days prior to defoliation. Following a single severe defoliation, ¹⁵N-labelled NH₄⁺ or NH₄⁺+ NO₃^? was supplied over a 20-day recovery period under the same SD and LD conditions. Changes in the relative contributions of remobilized N and newly acquired mineral N to shoot regrowth were assessed by sequential harvests. Both absolute and relative rates of N remobilization from root and stubble fractions were higher in LD than SD plants of both species, with the enhancement more acute but of shorter duration in timothy than fescue. Remobilized N was the predominant source of N for shoot regrowth in all treatments between days 0 and 8 after cutting; on average more so for fescue than timothy, because the presence of NO₃^? reduced the proportional contribution of remobilized N to the regrowth of timothy but not of fescue. Net uptake of mineral N began to recover between days 4 and 6 after cutting, with NO₃^? uptake restarting 1 or 2 days earlier than NH₄⁺ uptake, even when NH₄⁺ was the only form of N supply. LD timothy plants supplied solely with NH₄⁺ were slowest to resume uptake of mineral N. Supplying NO₃^? in addition to NH₄⁺ after defoliation promoted shoot regrowth rate but not remobilization of N. Rates of regrowth (shoot dry weight production per plant) were not correlated with rates of N remobilization from stubble either over the short-term (days 0–8) or longer term (days 0–18), interpreted as evidence against a causal dependence of regrowth rate on N remobilization under these conditions. The results are discussed in relation to hypotheses for source/sink-driven rates of N remobilization and their interactions with mineral N uptake following defoliation.     

Characterization of microsatellite markers for rough fescue species (Festuca spp.)

One major challenge in genetic diversity analysis of minor grass species is the lack of informative molecular markers. A set of 210 simple sequence repeat (SSR) markers developed from wheat and barley were evaluated for their transferability to three rough fescue species [Festuca altaica Trinius, F. campestris (Rydb.) and F. hallii (Vassey) Piper]. Twelve SSR primer pairs displayed scorable polymorphism among and within the species. The number of alleles per primer pair ranged from three to 17 with an average of 8.3 for all the species and greatly varied for each species. About 82% of SSR variation resided within the species. Festuca hallii was genetically most distinct among the three species.