Color profile: Generic CMYK printer profile Composite Default screen 131 Do rare pines need different conservation strategies? Evidence from three Mexican species Francisco Molina-Freaner, Patricia Delgado, Daniel Piñero, Nidia Perez-Nasser, and Elena Alvarez-Buylla Abstract: Mexico is a major center of diversity for the genus Pinus as it has the greatest number of species in the world. Many species are now restricted to Mexico, and some are represented by very small populations and are in dan- ger of extinction. In this study we examined allozyme variation in three rare species of Mexican pines: Pinus pinceana Gord., Pinus lagunae M.F. Passini, and Pinus muricata D. Don, with the objective of providing conservation guide- lines. The three species had relatively high levels of genetic variation with mean expected heterozygosities of 0.373, 0.386, and 0.346 for P. pinceana, P. lagunae, and P. muricata, respectively. We found marked differentiation among populations and significant inbreeding within populations of the three species. These values are larger than the range reported for most conifers and suggest that conservation strategies of these rare pines require focusing on the viability of several populations. Given that our knowledge about the demographic status of the three species is scarce, we pro- pose a mixed strategy of conservation. For P. lagunae, we propose an in situ strategy, whereas for P. pinceana and P. muricata we propose an ex situ strategy of conservation until permanent protection can be provided for several of their populations. Key words: genetic structure, conservation, rare pines, Pinus pinceana, Pinus lagunae, Pinus muricata. Résumé : Avec le plus grand nombre d’espèces au monde, le Mexique constitue un centre majeur de diversité pour le genre Pinus. Plusieurs espèces sont aujourd’hui restreintes au Mexique, certaines d’entre elles n’étant représentées que par de très petites populations en danger d’extinction. Dans cette étude, afin de jeter les bases pour leur conservation, les auteurs examinent la variation allozymique chez trois espèces de pins rares, du Mexique : Pinus pinceana Gord., Pinus lagunae M.F. Passini et Pinus muricata D. Don. Les trois espèces montrent des degrés relativement élevés de va- riation génétique avec des hétérozygocités moyennes attendues de 0,373, 0,386 et 0,346 pour les P. pinceana, P. la- gunae et P. muricata, respectivement. On observe une différenciation marquée entre les populations et une autofécondation significative dans les populations des trois espèces. Ces valeurs sont plus élevées que celles qui ont été rapportées pour la plupart des conifères et suggèrent que les stratégies de conservation de ces pins rares nécessitent qu’on s’attarde à la viabilité de plusieurs populations. Compte tenu que les connaissances de l’état démographique de ces trois espèces sont limitées, on propose une stratégie mixte de conservation. Pour le P. lagunae, les auteurs suggè- rent une stratégie in situ, alors que pour le P. pinceana et le P. muricata ils proposent une stratégie de conservation ex situ, en attendant qu’on puisse assurer la protection de plusieurs de leurs populations. Mots clés : structure génétique, conservation, pins rares, Pinus pinceana, Pinus lagunae, Pinus muricata. [Traduit par la Rédaction] Molina-Freaner et al. 138 Introduction stance, some knowledge about the genetic structure is im- portant for optimal sampling in ex situ conservation and the The extinction of species is a process that depends on de- identification of critical populations for in situ conservation. mographic and genetic factors. Although demographic fac- Although in situ strategies are generally preferred for long- tors are sometimes of more immediate importance in term conservation, recent case histories of forest genetic re- determining population viability (Lande 1988), long-term sources from North America have revealed that relying on conservation strategies depend upon knowledge of the ge- just in situ programs can be risky and that, in some cases, ex netic structure of the species (Schemske et al. 1994). For in- situ programs are the best alternative (Ledig et al. 1998). Surveys of enzyme polymorphisms in plant populations Received June 20, 2000. Published on the NRC Research have revealed that forest tree species usually have higher ge- Press website on February 8, 2001. netic diversity than other groups of plants (Hamrick et al. F. Molina-Freaner.1 Instituto de Ecologia, Universidad 1979). In conifers, most of this diversity is distributed within Nacional Autonoma de México, Apartado Postal 1354, populations with a small proportion of variation occurring Hermosillo, Sonora, Mexico C.P. 83000. among populations (El-Kassaby 1991; Ledig 1998). In pines P. Delgado, D. Piñero, N. Perez-Nasser, and E. Alvarez- and other coniferous species, usually less than 10% of the Buylla. Instituto de Ecologia Universidad Nacional Autonoma genetic variation is distributed among populations (El- de México, Apartado Postal 70-275, Mexico D.F. C.P. 04510. Kassaby 1991; Ledig 1998). Large population sizes, a pre- 1Corresponding author (e-mail: [email protected]). dominantly outcrossing mating system, and the potential for Can. J. Bot. 79: 131–138 (2001) DOI: 10.1139/cjb-79-2-131 © 2001 NRC Canada J:\cjb\cjb79\cjb-02\B00-155.vp Friday, February 02, 2001 2:48:46 PM Color profile: Generic CMYK printer profile Composite Default screen 132 Can. J. Bot. Vol. 79, 2001 long-distance gene flow are thought to be potential contribu- 500 km, and the nearest pine forest on the peninsula is found tors to the observed pattern in the genetic structure of coni- 800 km north, in the Sierra de San Pedro Mártir (Arriaga et al. fer populations (Loveless and Hamrick 1984). However, 1994). despite many studies on the genetic structure of pines, the Pinus muricata is a small tree that grows on coastal terraces and effect of the number, size, and distributional range of popu- slopes within 10 km of the ocean and on sites where fog is com- mon (Millar 1986). The species comprises nine disjunct popula- lations on the distribution of genetic diversity remains tions that extend from San Vicente in northern Baja California to poorly documented (Moran et al. 1988; Ledig 1998). There Trinidad in northern California and to two of the Channel Islands, is some evidence that pine species with very disjunct distri- Santa Cruz and Santa Rosa (Millar and Critchfield 1988). It be- butions exhibit higher interpopulation variation (Moran et al. longs to the subgenus Pinus, subsection Attenuatae (Price et al. 1988; Gibson and Hamrick 1991). The few studies dealing 1998). In Mexico, its distribution is discontinuous, and it is known with rare endemic pines have shown greater levels of differ- only from the foggy coastal foothills near San Vicente in the state entiation among populations (Ledig and Conkle 1983; of Baja California (Minnich 1987). Although this pine is not in Delgado et al. 1999). However, it is not clear if such a pat- global danger of extinction, its status in Mexico is considered as tern is common among all rare pine species. The distribution endangered in the Norma Oficial Mexicana (SEDESOL 1994). of genetic variation within and among populations of rare This species has serotinous cones and winged seeds (Perry 1991). pines may have important implications for their conservation strategies. If most of the variation in rare pines is distributed Sampling within populations, the preservation of just one large popula- Vegetative buds were collected from several populations of the tion might be sufficient to conserve the species. However, if three pine species. Table 1 shows the populations and the sampling a substantial amount of variation is present among popula- localities from each species. Coordinates from Table 1 were taken with a Magellan (Meridian XL) GPS. The number of plants col- tions, the conservation of rare pines may require a different lected in each population is given in Table 2. At least three buds strategy. were collected per plant, placed in individual plastic bags, trans- Pinus, with approximately 110 species (Price et al. 1998), ported to Mexico City on ice, and stored in an ultrafreezer at is the largest genus of conifers and the most widespread ge- –70°C. nus of trees in the Northern Hemisphere. Mexico is a major center of diversity for the genus, because it has more species Electrophoretic procedures of pines than any other country in the world (Perry 1991; Buds were ground using three parts YO (Yeh and O’Malley Styles 1993). Some subsections of the genus seem to have 1980) extraction buffer and one part Veg II buffer (Cheliak and originated in Mexico and many species are now restricted to Pitel 1984). Crude extracts were absorbed onto filter paper wicks this country. At least nine of these endemic species are rep- and immediately inserted into 12% starch gels, using four buffer resented by very small populations and are in danger of ex- systems. Buffer system number 2 of Miles et al. (1977) was used tinction (Perry 1991). This group of pines offers an excellent to resolve GDH, IDH, and SDH; buffer system number 2 of opportunity to determine whether small population sizes and Wendel and Weeden (1989), to resolve ACPH, MDH, and 6-PGD; different degrees of isolation affects the amount and distribu- the tris–citrate–borate system of Mitton et al. (1977), to resolve APX, CPX, GOT, and MNR; and buffer system R of May (1992), tion of genetic variation. In this study, we examined allozyme to resolve EST, LAP, NADH, and RUB. Staining procedures fol- variation in three rare species of Mexican pines from this lowed Hakim-Elahi (1976) for ACPH, APX, and CPX; Conkle et group: Pinus pinceana Gord., Pinus lagunae M.F. Passini, al. (1982) for GDH, MDH, and MNR; Soltis et al. (1983) for IDH and Pinus muricata D.