Growth characteristics of endophyte-infected and endophyte-free Lolium perenne L. seedlings under osmotic stress conditions

In their evolution, grasses have developed symbiotic associations with fungi including mycorrhizal fungi that grow in or on the roots, and endophytes that live their entire life cycle within the aerial portion of the host grass, forming nonpathogenic, systemic and usually intercellular associations. Currently, seven genera have been identified, namely, Atkinsonella, Balansia, Balansiopsis, Echinodothis, Epichlo e ¨, Myriogenospora and Parepichlo e ¨, among which the anamorphic (imperfect) stage of Epichlo e ¨ spp. is most closely related to cultivated grasses. In 1982 Morgan Jones Gams erected the section Albo lanosa to accommodate anamorphic stage of Epichlo e ¨ spp. in the genus Acremonium . In 1996 Glenn et al . reclassified the genus Acremonium section Albo lanosa in a new genus Neotyphodium to separate clavicipitaceous Acremonium from Acremonium with other affinities (usually saprotrophs). The most widely known Neotyphodium endophytes are N. lolii (Latch, Christensen and Samuels) Glenn, Bacon and Hanlin, and N. coenophialum (Morgan Jones Gams) Glenn, Bacon and Hanlin, which colonize perennial ryegrass (Lolium perenne L.) and tall fescue ( Festuca arundinaceae Schreb.), respectively. Endophytes and their host grasses are mutually symbiotic. On one hand, the grasses provide photosynthates for the fungi; on the other hand, the endophytes may enhance the host's growth and protect the host from biotic stresses (such as mammalian, insect, and nematode herbivores) and abiotic stresses (such as drought and high temperature). Since Read Camp first reported the close relationship of endophyte infection to drought resistance of tall fescue in 1986, a lot of research work has been done on the effect of endophyte infection on drought resistance of tall fescue, while rather less work has been done on the effect of endophyte infection on drought resistance of perennial ryegrass. Thus in this paper Lolium perenne cv SR4000 infected by Neotyphodium lolii was chosen as experimental material. Seed germination and seedling growth in normal and osmotic stress conditions were compared between endophyte infected (EI) and endophyte free (EF) populations. Seeds of SR4000 were stored in two different ways: some were stored at 4℃ in refrigerator while others were stored at 20℃ in illuminating incubator for 18 months to eradicate endophytes in the seeds. After being soaked in distilled water for 4 hours, the seeds treated in the above ways were placed on filter papers moistened with distilled water, 5% PEG, 10% PEG and 15% PEG solutions, respectively. Seed germination was daily recorded from the 3 rd day until the number of germinated seeds no longer increased. In seedling growth experiment, EI and EF plants were transplanted into culture pots filled with 500mL Hoagland solution. One week later, PEG6000 was added as osmoticum to impose osmotic stress (control, mild and severe stress). The PEG contents for the above stresses were 0, 10% and 20%, respectively. Leaf elongation of marked shoots was measured every three days, and the number of leaves and number of tillers were recorded every six days. By the end of the experiment, green leaves, wilted leaves, pseudostems and roots were harvested separately. All data were statistically analyzed with SAS package. Endophyte infection accelerated seed germination under normal and osmotic stress conditions. Only under severe osmotic stress, however, seed germination rate and germination index of EI seeds were significantly higher than those of EF seeds. For example, under 10% PEG stress, EI seeds began to germinate from the 3 rd day, and the germination rate was 79 6% on the 6 th day, and 90 4% on the 8 th day; while EF seeds began to germinate from the 4 th day, and the germination rate was 73 4% on the 6 th day, and 89 1% on the 8 th day, which was close to the germination rate of EI seeds. Under 15% PEG stress, however, germination rate of EI seeds was 10%～16% highe