Background and aims Low temperature stress is a common hazard during plant growth. Endophyte infection has been shown to increase cold tolerance in host plants. Many Lolium perenne cultivars contain low to moderate levels of endophyte. This study was done to explore cultivar improvement by segregation of endophyte containing individuals from the original cultivar to create a high endophyte subpopulation.

Methods Endophyte-infected plants were segregated over the first three years to produce high-endophyte subpopulation, and field and greenhouse experiments were carried out in the forth and fifth to determine the cold tolerance of the L. perenne subpopulation with high endophyte infection rates (N), the parent (F), the control endophyte-free subpopulation (E) and the control local variety (L).

Results (1) After three years of screening, high endophyte infection rates in the tillers and seeds of plants were still observed (96.5%), and agronomic traits (crown width, plant height, panicle number, withering, regreen-up, the growth cycle and the over-wintering rate) was also improved with increased Epichlo? colonization of host plant. (2) The subpopulation with high endophyte infection rates and improved agronomic traits had better cold tolerance than the parent, the control endophyte-free subpopulation and the control local variety. The possible mechanisms by which high endophyte infection enhances cold resistance in the field include increased root system, increased the over-wintering rate, reduced regrowth periods with the sowing date being October 15^th. (3) The high-endophyte subpopulation significantly increased SOD, POD, CAT, and APX activities at 0, 5, and 10 °C by 11.8%–44.6%, compared with the parent population.