Eight Pinus nigra Arn. populations from Southern Spain and Northern Morocco were examined using inter-simple sequence repeat markers to characterize the genetic variability amongst populations. Pair-wise population genetic distance ranged from 0.031 to 0.283, with a mean of 0.150 between populations. The highest inter-population average distance was between PaCU from Cuenca and YeCA from Cazorla, while the lowest distance was between TaMO from Morocco and MA Sierra Mágina populations. Analysis of molecular variance (AMOVA) and Nei’s genetic diversity analyses revealed higher genetic variation within the same population than among different populations. Genetic differentiation (Gst) was 0.233. Cuenca showed the highest Nei’s genetic diversity followed by the Moroccan region, Sierra Mágina, and Cazorla region. However, clustering of populations was not in accordance with their geographical locations. Principal component analysis showed the presence of two major groups—Group 1 contained all populations from Cuenca while Group 2 contained populations from Cazorla, Sierra Mágina and Morocco—while Bayesian analysis revealed the presence of three clusters. The low genetic diversity observed in PaCU and YeCA is probably a consequence of inappropriate management since no estimation of genetic variability was performed before the silvicultural treatments. Data indicates that the inter-simple sequence repeat (ISSR) method is sufficiently informative and powerful to assess genetic variability among populations of P. nigra.

Soil storage method may alter enzymatic activity being storage conditions of the soil samples prior to analysis decisive for the results. Studies made on freshly collected soils are generally preferred. However it is always not possible due to practical reasons since for example sampling is often restricted to short period of the year or because a great quantity of microbiological analyses must be made on time and by few people. On this context, soil storage methods are needed, being cold at 4°C the most widely used although sometimes alternative storage methods are also utilized. The aim of this study is to evaluate the effect of two alternative storage methods of soil samples (freezing at –20°C and air drying conservation methods) in comparison to cold at 4°C on the enzymatic activities (dehydrogenase, phosphatase, β-glucosidase and urease soil enzymes). Samples of two forest ecosystems (pine and holm oak forest stand) were taken in two different season of the year (winter and spring 2009). Results showed that enzymatic activities differed when freezing or air drying conservation methods were used in comparison with cold soil samples. Generally, alternative soil storage methods presented lower enzymatic activity than cold at 4°C. However, these changes depend on season and sampling location.