A multidisciplinary approach for the characterization of pistacia atlantica desf. subsp atlantica diversity in northwest algeria

Abstract

The current study aims at the characterization of Pistacia atlantica Desf. subsp atlantica diversity from Northwest Algeria Sixteen natural populations of Pistacia atlantica Desf. subsp. atlantica were investigated on a longitudinal gradient. The pluviothermal data of sampling sites were used for the climatic characterization. The study examined morphologically ten mature trees per sites (males and females) and biometrically 3520 mature compound leaves. A significant correlation between climatic parameters and leaf morphology was showed. Several values were reported for the first time on the species, such as the length and the width of the leaf (reaching up to 24.5 cm/21.9 cm), the leaflets number (up to 18 leaflets/ leaf) and the petiole length of the terminal leaflet (reaching up to 3.4 cm). The original findings of this study were used to update the Pistacia atlantica subsp. atlantica identification key. The intraspecific chemical variability was assessed through the characterization of leaf essential oils and cell wall polysaccharides. Essential oils were isolated by hydrodistillation. Compositions were further investigated by Gas Chromatography (GC) followed by Gas chromatography-mass spectrometry GC/MS and were submitted to multivariate statistical analysis. Essential oils were obtained in yields ranging from 0.1 to 0.42%. Terpinen-4-ol, α-pinene, germacrene D and E-caryophyllene were found as major components. The two last compounds were never mentioned as main constituents in the essential oil of this species. Multivariate cluster analysis of compositional data evidenced three types of essential oil considering the predominant constituents: 1) terpinen-4-ol ; 2) α-pinene / camphene ; 3) α-pinene / germacrene D. For the cell wall polysaccharides dosage, cellulosic and hemicellulosic fractions were isolated and dosed from mature leaves. The wall residue yield varied from 36.2 to 69.5%. The cellulose and hemicelluloses yields varied from 21.25 to 51% and 7 to 27.6% respectively. The genetic diversity was assessed through the genome size estimation, the chromosome count and the genetic polymorphism assessment. The genome size estimation was performed using flow cytometry on 97 samples. The Propidium Iodide (IP) fluorescence intensity showed that all the analyzed samples are diploids (2n=2x). The flow cytometry analysis leaded to estimate a very small genome (2C = 1.21±0.02 pg). No statistical differences were recorded between the genome sizes of the analyzed samples. Squash preparation for chromosome study was carried on fixed buds. Meristematic somatic cells of male floral buds showed the best prometaphase which on (2n =30) chromosome complement was observed. Two chromosomes bearing satellites showed evident and large secondary constrictions. For the genetic diversity assessment, 61 genotypes were characterized using 6 SSR microsatellites. A total of 26 alleles were amplified. The effective number of alleles ranged between 1.92 and 4.34. Allele frequencies ranged from 0.008 to 0.68. Eight alleles (30.8%) observed as rare alleles. The observed heterozygosity (Ho) ranged from 0.35 to 0.67, while the expected heterozygosity (He) ranged from 0.48 to 0.77. Five of six loci showed an overall heterozygote deficiency; however, none of the assayed loci displayed a significant heterozygote deficiency. Two loci showed significant departure from Hardy–Weinberg equilibrium (HWE), while no significant deviations (P<0.01) from the Hardy–Weinberg proportions were found for four loci. Four of six markers were classified as very informative markers. Molecular analysis of variance (AMOVA) showed moderate differentiation in allele frequencies between four groups of populations and that 87.65% of variation is originated from variability between genotypes. Cluster analysis based on the euclidean distance using ‘Unweighted Pair Group Method using Arithmetic Averages’ (UPGMA) leaded to observe divergences between genotypes within site and between sites. The current study indicated an evident diversity probably due to a genetic flow between populations.