Lucia Conte

Genetic and phytochemical difference between some Indian and Italian plants of Withania somnifera (L.) Dunal

The geographical distribution of Withania somnifera (L.) Dunal is quite wide. However, in Italy, this species is very rare and grows spontaneously only in Sicily and in Sardinia. The PCR–RAPD technique has been utilized in this work to determine the genetic relationship among Sicilian, Sardinian and Indian samples and the HPLC analysis of whitaferin A was used as a marker to evaluate the phytochemical differences. The genetic difference between Indian and Sicilian plants of W. somnifera turned out to be smaller than that between Indian and Sardinian plants of this species. The phytochemical analysis as well showed that the Sardinian specimen strongly differed from the Indian and Sicilian ones in its contents of withaferin A. Our results seem to confirm the hypothesis that the Italian populations of this species may not be indigenous but naturalised. Due to the high withaferin A content of the Sardinian samples, these plants could be used as a source for pharmaceutical purposes.

Genetic diversity in Cytisus aeolicus Guss. (Leguminosae), a rare endemite of the Italian flora

ABSTRACT Cytisus aeolicus Guss, is an endemic plant restricted to the isles of Vulcano, Stromboli and Alicudi in the Aeolian archipelago. All known populations were assayed for genetic variability using enzyme polymorphism. Allozyme variation at 16 loci coding for 10 enzyme systems was examined. The great majority of loci turned out to be monomorphic or fixed heterozygous. The observed genetic depauperation is indicative of historical factors, such as the bottleneck effect associated with migration, and the founder effect in population re-establishment. The low genetic diversity is largely partitioned within rather than among populations, indicating that extant populations have not been established long enough for divergence to have occurred. The genetic uniformity, combined with the scattered distribution of small populations, and the negative shift of population size point to a critically endangered species. Strategies for conservation are considered.

Detection of ephemeral genetic sub-structure in the narrow endemic Abies nebrodensis (Lojac.) Mattei (Pinaceae) using RAPD markers

A. nebrodensis (Nebrodi fir, Sicilian fir) is restricted to a small area of the Madonie Natural Park in Sicily. According to recent estimates, its only population consists of 30 adult individuals and a fluctuating number of juveniles derived from natural regeneration; besides, some hundreds of cultivated plants are preserved as ex situ collection. We used RAPD data from six 10-mer primers to examine the consequences of extensive historical clearance and human pressures on the extant population. Data from multiple life stages and different habitat conditions were considered, affording an opportunity to ascertain for the first time the structure of genetic variation in the extant uneven-aged population on both spatial and temporal scale. Consistent with previous works, we found that the history of disturbance and past land use did not affect drastically the biological diversity of the present-day population. Considerable levels of genetic variation were detected both in adult and juvenile sets. The non-significant correlation between genetic and physical distances (Mantel test; r = − 0.075) revealed a random distribution of genotypes in the natural stand. When the juvenile set was divided in 17 subsets, based on maternal provenance and growing conditions, Shannons index (54%) and AMOVA analysis (84%) indicated that most of genetic variation resides within subsets. Neighbour-joining cluster analysis supported the hypothesis of high cross-fertilization rates expected for a woody perennial species; nevertheless some grouping of related individuals suggested partial inbreeding and a weak pattern of genetic structure. Changes in structure can occur as the progeny ages and offspring thinning is responsible for moulding patterns of genetic diversity and population structure in time.

Isolation and biodiversity in Cytisus villosus Pourret (Fabaceae, Genisteae): enzyme polymorphism in disjunct populations

ABSTRACT The genetic diversity of isolated populations of Cytisus villosus has been studied by means of enzyme polymorphism analysis. Two types of isolated populations were studied: “terrestrial islands” in Sicily, and “true islands” in the Aeolian archipelago. In the populations of “true islands” the number of alleles and the heterozygosity are lower than in “terrestrial islands”. Isolation amongst Sicilian populations seems to be more recent than isolation of the Aeolian populations, and may be attributed to climatic changes which occurred during the Holocene and/or to human activities. The disjunction of the Aeolian populations seems much more recent than the origin of the isles themselves; the colonization of the archipelago is attributed to a single, recent dispersal event not followed by local evolution. In view of the biological structure of the Aeolian populations, C. villosus must be regarded as a locally endangered species.

Molecular evidence for hybrid origin of Quercus crenata Lam. (Fagaceae) from Q. cerris L. and Q. suber L.

Abstract Random Amplified Polymorphic DNA (RAPD) and Inter-Simple Sequence Repeats (ISSR) markers were employed to examine samples from Quercus cerris, Q. suber and Q. crenata in order to test the hypothesis of the hybrid origin of Q. crenata from Q. cerris and Q. suber in a part of its distribution area where the two putative parents do not overlap at present. Leaves from 21 Q. crenata and 37 Q. cerris individual trees were collected at natural sites in northern Italy, where Q. suber is currently lacking; 21 Q. suber and six Q. crenata plants from central Italy were also analysed. Results from Unweighted Pair Group Method with Arithmetic mean (UPGMA) analysis and principal component analysis (PCA) implied that exchange of neutral markers has been considerable between the three species, while differences in morphological characters have remained comparatively stable. The Mantel test indicated low correlation between RAPD- and ISSR-based similarity matrices, showing that the two screening techniques reveal unrelated estimates of genetic relationships. Hybrid indices computed for both markers displayed an intermediate position of Q. crenata individuals between the two putative parents shifted toward Q. cerris. Results from the present study corroborate the hypothesis of a hybrid origin for Q. crenata occurring in northern Italy, and suggest asymmetrical backcrossing with Q. cerris acting as the recurrent parent.

Genetic diversity of the rare and endangered meadow violet (Viola pumila Chaix) at the southern margin of its range

Viola pumila Chaix is a rare violet with a Eurasian-continental distribution. It has undergone a severe population decline during the last decades, and is endangered in Europe. Its southernmost extrazonal occurrence is in northern Italy, with only four populations. Inter-simple sequence repeat markers were used to detect genetic diversity level and distribution in the Italian stand and to compare it to a small sample from Germany. Estimates of genetic diversity revealed retention of genetic variation and weak genetic structure in the Italian populations of V. pumila. Comparable levels of genetic variation were detected in the German population. Principal component and cluster analyses on the whole data-set grouped individuals regardless of their geographical distribution, pointing out that genetic diversity in the V. pumila populations surveyed is homogeneously distributed. Analysis of molecular variance showed that most of the genetic variation is within populations rather than among them. Our results indicate that habitat fragmentation has not induced genetic depauperation and differentiation in the Italian stand of V. pumila; moreover, they provide evidence of past extensive outcrossing and a common evolutionary history of the examined populations. The high diversity detected in this study suggests effective measures for conservation strategies of this rare species.

Assessment of RAPD variation in Abies nebrodensis (Lojac.) Mattei (Pinaceae) using haploid tissue analysis

RAPD technique using haploid DNA from megagametophyte tissue was applied to detect the pattern of genetic variation within Abies nebrodensis, a species endemic to Sicily. Results were compared with a reference system composed of representatives of five Mediterranean firs: A. alba, A. cephalonica, A. nordmanniana, A. numidica, and A. pinsapo. Diploid genotypes for RAPD loci were inferred from segregation patterns in haploid megagametophytes from individual trees. Therefore, it was possible to distinguish between homo- and heterozygotes carrying a RAPD fragment and to calculate unbiased estimates of genetic parameters. In the interspecific comparison, A. nebrodensis appears intermixed with the other Abies species, supporting the hypothesis of remarkable hybridization processes within the genus. A high level of polymorphism and deviation from panmictic equilibrium as a result of heterozygote excess depict a pattern of population structure in A. nebrodensis that agrees with the expectations for an outcrossing perennial species, in spite of its critical conditions. Implications with respect to the conservation management of the species are also discussed.

Phylogenetic patterns and endemism genesis in Cytisus Desf. (Leguminosae-Cytiseae) and related genera

A cladistic analysis based on morphological characters was performed on Cytisus and related genera. Terminal taxa were three Mediterranean endemics (C. aeolicus, C. emeriflorus, and C. tribracteolatus), all sections of genus Cytisus, and all other genera of the tribe. The main dichotomy within the tribe, between Cytisus group and Genista group, is corroborated. Both genera, Genista and Cytisus, seem to be paraphyletic. Cytisus s.s. is composed of at least three clades. The cladograms indicate a link of C. emeriflorus with sect. Tubocytisus, and of C. aeolicus with C. villosus. C. tribracteolatus belongs to the same clade as the two species above, but is more loosely related to them. The phyletic relations suggested are compatible with phytogeography. On the basis of this analysis, an evolutionary sketch is outlined and some taxonomic realignments are proposed.

Infraspecific diversity of Cytisus emeriflorus Reichenb. (Leguminosae), an endemic plant with disjunct distribution: Evidence from isozyme data

ABSTRACT C. emeriflorus is an endemic species occurring in two disjunct areas some 200 km apart: the Pre-Alps of Lombardia and the Pre-Alps of Friuli. Four populations from the western area and five populations from the eastern area were sampled in order to ascertain the amount and the distribution of variation throughout the range of the species. Phenetic measures of isozyme polymorphism were used. Diversity values showed that the species is polymorphic, despite its restricted distribution and the small population size; phenetic distances revealed that partitioning of diversity was largely within rather than among populations. Moreover, little differentiation was found between the two disjunct areas, although the western populations exhibited a slightly higher diversity compared with the eastern ones. The high level of polymorphism and the overall similarity between populations from the two areas may be ascribed to past contacts, and indicate that, despite the present widespread disjunction, the two areas have not evolved independently long enough for divergence to have occurred. There is no evidence that C. emeriflorus is at present genetically depauperated, although its range of distribution must have been much broader in the past; its present reduced distribution area and the small population size may be mainly due to ecological factors.

Storage globulins in Gnetopsida. 1. Recognition of legumin-like proteins

Abstract Salt soluble storage proteins were extracted from seeds of Ephedra distachya, Ephedra foeminea, Gnetum gnemon, Gnetum montanum and Welwitschia mirabilis and separated by chromatographic procedures. The molecular weight of the main storage globulin ranges from 300 to 350 kD. Denaturation by SDS resolved the holoprotein in monomers of Mr 40 to 60 kD. Oligomers up to 120 kD were observed in Ephedra. Reduction of disulphide bridges by DTE resolved the monomers in paris of polypeptides of Mr 10 to 35 kD. The characters above indicate that the main storage globulin of Gnetopsida is a legumin-like protein.