Allelopathic Effects of Pinus halepensis Needles on Turfgrasses and Biosensor Plants

Fresh, senesced, and decaying pine needles from Pinus halepensis were evaluated for their allelopathic potential on Festuca arundinacea, Cynodon dactylon and the biosensor plants Avena sativa and Lemna minor through in vivo and in vitro studies. The in vivo study was performed in growth chambers, using 6, 12, and 18 g of pine needle tissue mixed with screened perlite as a substrate. The effects of the different pine needle types were evaluated by determining the total root length, total root surface, root dry weight, total shoot length, total shoot surface, and shoot dry weight. The in vitro study was performed in Petri dishes where seeds from each species were subjected to an increasing concentration of pine needle extract. The extracts were obtained from pine needle ground tissue that was diluted with water and either shacked at room temperature or placed in water bath at 40 °C for 24 h. The evaluation of the allelopathic potential was performed with the determination of radicle length. The allelopathic potential of the pine needle tissues was confirmed with bioassays using oat (A. sativa) and duckweed (L. minor). The results strongly suggested the allelopathic potential of the pine needle tissue, being more pronounced in the fresh, moderate in the senesced, and low in the decaying pine needles. The allelopathic substances were species-specific, and the inhibition resistance of the species tested followed the order F. arundinacea > C. dactylon > A. sativa. The inhibition of the L. minor suggested that the water soluble phytotoxic compounds were inhibitors of Photosystem II. Plants produce many compounds that have no apparent metabolic, physiological, or structural role for the producer. These secondary metabolites might exhibit allelopathic effects and influence other organisms in the vicinity of the producer (Rice, 1984). In many cases, allelochemicals function as biochemical defence mechanisms or possess insecticidal, repellent, antifeedant, and antimicrobial activities (Grainge and Ahmed, 1988; Peterson et al., 1989). In turfgrasses, allelopathy has been investigated as an interactive mechanism to explain competition between species (Lickfeldt et al., 2001). Brede (1991) was unable to determine allelopathic inhibitory action when leachate collected from a Poa annua putting green was applied onto bentgrass (Agrostis stolonifera ssp. palustris Huds.). In contrast, Buta et al. (1987) found different degrees of lettuce seedling growth inhibition when seed leachate from various turfgrasses was applied. Tall fescue (Festuca arundinacea Schreb.) has been found to exhibit allelopathic effects on birdsfoot trefoil (Lotus corniculatus L.) (Luu et al., 1989; Peters, 1968) while tree seedlings of loblolly pine (Pinus taeda) were inhibited more by tall fescue and less by bermudagrass (Cynodon dactylon [L.] Pers.) (Smith, 1989). However, allelopathy can also be exerted by the tree species to turfgrasses that are growing in the understory. In the Mediterranean region, pine trees and pine forests are found in abundance from sea level to the higher elevations of the mountains. Turfgrasses that are used as groundcovers in urban parks, golf courses, as well as in private gardens are frequently established proximately to pine trees. It has long been noticed that turfgrass growth is severely restricted under pine trees. Even though research evidence is nonexistent, turfgrass growth reduction under the canopy of the pines has been attributed to various reasons such as 1) light exclusion by the dense canopy of the pines hindering plant photosynthesis, 2) soil pH reduction under the canopy of the pine trees due to the fallen pine needles (Pallant and Riha, 1990), and 3) the accumulation of an organic matter layer due to the fallen pine needles, which hinders the growth of the turf sward. However, the allelopathic capacity of phytotoxic compounds that are produced and accumulated within the pine needle tissue has been postulated as an additional reason for the restricted turfgrass growth under pine trees. Wilt et al. (1993a) found several volatile monoterpene hydrocarbons in the understory litter of P. monophylla Torr. & Frem., which might act as phytotoxins. The monoterpene concentration was higher in fresh, remained relatively high in senesced, but diminished in the decaying needles of the litter material (Wilt et al., 1993b). Therefore, the aim of the present study is 1) to investigate the allelochemical characteristics of P. halepensis L. and to evaluate the allelopathic activity of pine tissues on the growth and development of F. arundinacea, C. dactylon, and A. sativa; 2) to determine the toxicity of the pine needle extracts received at different temperatures to the development of the above-mentioned turfgrass species; and 3) to determine the toxicity potential and characterize the aqueous extracts of the pine needles based on their inhibition effect on plant biosensors. Materials and Methods To evaluate the allelopathic potential of pine needles, three studies were performed: Study I: Pine needle effects on plant growth. The study was performed in growth chambers with the use of small plastic pots (70 mm ID and 110 mm high), filled with sieved perlite (Isocon S.A., Athens, Greece) having a uniform particle distribution (0.5 to 1.0 mm). Pine needles were collected from P. halepensis trees found in the area of Ilioupoli, a suburb of Athens, Greece. For the purposes of allelopathic assessment, 3 different physiological stages of the pine needles were used: 1) fresh pine needles (F) were collected from the pine trees; 2) senesced (S) pine needles, which had a yellowish color, were collected from the top of the thatch layer that was formed under the canopy of the trees, and 3) decaying (D) pine needles having a dark grey color were collected from the bottom of the thatch layer. The pine needles were cut into 5-mm segments and placed at –18 °C for 7 d. The frozen tissues were incorporated in the perlite substrate at 3 different rates namely 6, 12, and 18 g of fresh weight per pot. The pots were then seeded either with tall fescue (F. arundinacea ‘Tomahawk’), bermudagrass (C. dactylon ‘Bluemuda’), or oat (A. sativa) at a rate of 20, 20, and 3 seeds per pot, respectively. Oat was included in this study since it has been used extensively in allelopathy research as the receiver plant to test compounds released by a donor plant. Oat seeds germinate evenly, resulting in a uniform and rapid plant growth that enables qualification of biological response in plants. In addition, oat biotest is considered as a sensitive and easily facilitated method (Rice, 1984). The seeded pots were placed in the growth chamber with a constant temperature of 24 °C for tall fescue and oat and 30 °C for bermudagrass. All pots were irrigated with deionised water, and artificial light was supplied with a photoperiod of 16 to 8 h light and dark, respectively. The seedlings were fertilized with foliar fertilizer (Nutrileaf 20–20–20; Miller, Hanover, Pa.) at a rate of 2.5 g·L at weekly intervals after the emergence of the second leaf. The seedlings were destructively sampled 12, 43, and 31 d HORTSCIENCE 40(1):246–250. 2005. Received for publication 19 Dec. 2003. Accepted for publication 25 June 2004. The authors wish to thank Susan Coward for her help in improving the manuscript. Assistant professor. Lecturer. Professor.