Allozyme diversity in natural populations of Viola palmensis Webb & Berth. (Violaceae) from La Palma (Canary Islands): implications for conservation genetics

Genetic diversity was measured by allozyme electrophoresis in eight natural populations of the threatened Canarian endemic Viola palmensis Webb & Berth. (Violaceae). Nineteen alleles corresponding to 11 gene loci were detected. High levels of genetic diversity were found, ranging from 36.3 to 45.4 % for the percentage of polymorphic loci (P), from 1.45 to 1.60 for the average number of alleles per locus (A) and from 0.128 to 0.200 for the expected heterozygosity (H(e)). Between 85.5 and 96.6 % of genetic variability was apportioned within populations. As a whole, populations were not at Hardy-Weinberg equilibrium, with a deficit of heterozygous individuals attributable to the existence of genetic structuring in the populations analysed. The levels of interpopulation genetic differentiation were low (mean F(ST) = 0.100), while genetic identity pair-wise comparisons were high (mean I = 0.973) suggesting considerable levels of gene flow among populations. No relationship was detected between genetic differentiation and geographical distances between populations. An outcrossing insect-mediated breeding system might contribute to pollen dispersion of this species. For conservation genetics we suggest in situ preservation areas are defined that are free of disturbance and that include populations with the highest genetic diversity.     

Use of RAPD analysis in devising conservation strategies for the rare and endangered Grevillea scapigera (Proteaceae)

Extensive human impact in south western Australia has resulted in a high incidence of rarity throughout the highly endemic flora of the region. Grevillea scapigera (Proteaceae) is a typical example, with 27 plants (represented by four extant populations) remaining in the wild. In order to devise an appropriate strategy for the conservation of this species, its population genetics were studied using RAPD analysis, which enabled the discrimination of individual plants and the detection of a relatively high amount of variability (V = 0.32) within G. scapigera. This variability was found to be evenly distributed within the plants analysed despite the clear distinction between most populations (87% of the variability being attributable to single plant difference and 13% to population difference). Finally, RAPD analysis was used to select a small group of plants that captured maximum genetic variability to be used in the recovery program of the species. Because of the low genetic difference between populations, the mixing of these selected plants during the recovery process should not create genetic imbalances. The methods used in this study provide a useful model for future projects involving the recovery of rare flora.     

Population genetic structure of rock ptarmigan in the 'sky islands' of French Pyrenees: implications for conservation

Expected consequences of global warming include habitat reduction in many cool climate species. Rock ptarmigan is a Holarctic grouse that inhabits arctic and alpine tundra. In Europe, the Pyrenean ptarmigan inhabits the southern edge of the species' range and since the last glacial maximum its habitat has been severely fragmented and is restricted to high-alpine zones or 'sky islands'. A recent study of rock ptarmigan population genetic in Europe found that the Pyrenean ptarmigan had very low genetic diversity compared with that found in the Alps and Scandinavia. Habitat fragmentation and reduced genetic diversity raises concerns about the viability of ptarmigan populations in the Pyrenees. However, information on population structuring and gene flow across the Pyrenees, which is essential for designing a sound management plan, is absent. In this study, we use seven microsatellites and mitochondrial control region sequences to investigate genetic variation and differentiation among five localities across the Pyrenees. Our analyses reveal the presence of genetic differentiation among all five localities and a significant isolation-by-distance effect that is likely the result of short dispersal distances and high natal and breeding philopatry of Pyrenean ptarmigan coupled with severe habitat fragmentation. Furthermore, analysis of molecular variance, principal component analysis and Bayesian analysis of genetic structuring identified the greatest amount of differentiation between the eastern and main parts of the Pyrenean chain separated by the Sègre Valley. Our data also show that the Canigou massif may host an isolated population and requires special conservation attention. We propose a management plan which includes the translocation of birds. If a sky island structure affects genetic divergence in rock ptarmigan, it may also affect the genetic structure of other sky island species having low dispersal abilities.     

A review of the allozyme data set for the Canarian endemic flora: causes of the high genetic diversity levels and implications for conservation

Background and Aims

Allozyme and reproductive data sets for the Canarian flora are updated in order to assess how the present levels and structuring of genetic variation have been influenced by the abiotic island traits and by phylogenetically determined biotic traits of the corresponding taxa; and in order to suggest conservation guidelines.

Methods

Kruskal–Wallis tests are conducted to assess the relationships of 27 variables with genetic diversity (estimated by A, P, H_o and H_e) and structuring (G_ST) of 123 taxa representing 309 populations and 16 families. Multiple linear regression analyses (MLRAs) are carried out to determine the relative influence of the less correlated significant abiotic and biotic factors on the genetic diversity levels.

Key Results and Conclusions

The interactions between biotic features of the colonizing taxa and the abiotic island features drive plant diversification in the Canarian flora. However, the lower weight of closeness to the mainland than of (respectively) high basic chromosome number, partial or total self-incompatibility and polyploidy in the MLRAs indicates substantial phylogenetic constraint; the importance of a high chromosome number is feasibly due to the generation of a larger number of linkage groups, which increase gametic and genotypic diversity. Genetic structure is also more influenced by biotic factors (long-range seed dispersal, basic chromosome number and partial or total self-incompatibility) than by distance to the mainland. Conservation-wise, genetic structure estimates (F_ST/G_ST) only reflect endangerment under intensive population sampling designs, and neutral genetic variation levels do not directly relate to threat status or to small population sizes. Habitat protection is emphasized, but the results suggest the need for urgent implementation of elementary reproductive studies in all cases, and for ex situ conservation measures for the most endangered taxa, even without prior studies. In non-endangered endemics, multidisciplinary research is needed before suggesting case-specific conservation strategies. The molecular information relevant for conservation should be conserved in a standardized format to facilitate further insight.     

Allozymic variation and divergence in three species of Antirrhinum L. (Scrophulariaceae-Antirrhineae)

An allozymic study of three wild species of Antirrhinum L. A. lopesiamum Rothm., A. mollissimum Roihni. and A. microphyllum Rothm. is described. All are members of subsection Kickiella Rothm., and are narrow-range endemics of the Iberian Peninsula. The variability of the different loci, as well as the number and mobility of the alleles, differ among the three species, a demonstration of the usefulness of allozymes for the systematics of the genus. The finding of alleles unique to each species indicates high divergence among species suggesting ancient diversification, and supports the hypothesis of a geographical model of speciation. All three species show high levels of within-species variability, mainly partitioned within populations, while between populations genetic differentiation is low. Correlations between population size, sample size and genetic variability, and the usefulness of allozymic data for conservation purposes, are discussed.     

Genetic variation in populations of the endangered fish Ladigesocypris ghigii and its implications for conservation

1. The genetic variation of the endangered freshwater fish Ladigesocypris ghigii, endemic to the island of Rhodes (Greece), was investigated for nine populations, originating from seven different stream systems and a reservoir, both at the mtDNA and nuclear level, in order to suggest conservation actions. 2. Both restriction fragment length polymorphism analysis of five segments of mitochondrial DNA (ND‐5/6, COI and 12S‐16S rRNA) amplified by polymerase chain reaction, and random amplified polymorphic DNA analysis, revealed extremely low levels of intra‐population polymorphism. It is highly likely that the low intra‐population variability is the result of successive bottleneck events evident in shrinkage and expansion of the populations year after year, which may have led to a complete loss of several genotypes and haplotypes, and an increased degree of inbreeding. 3. Inter‐population genetic structuring was high, with fixation of haplotypes within six of the nine populations and fixation of alleles within populations originating from different waterbodies. It is probable that all haplotypes and/or alleles found were initially represented in all populations. However, because of the long time of isolation coupled with successive bottleneck and subsequent genetic drift, common mtDNA haplotypes and alleles among the populations may have become rare or extinct through stochastic lineage loss. 4. Although nucleotide divergence among haplotypes was very shallow, half of the haplotypes recorded (three of six), resulted from nucleotide changes on the 12S–16S rRNA segments, which are the most conserved part of the mitochondrial genome. This fact may indicate that the observed genetic variation did not necessarily result only from the retention of ancestral polymorphism, but may have arisen through mutation and complete lineage sorting over a relatively small number of generations, once the populations had become isolated from one another. 5. Our data suggest that two of the L. ghigii populations may be on independent evolutionary trajectories. Considering that each population appears so far well adapted within each site, all populations should be managed and conserved separately.     

Isozyme analysis of Galaxias species (Teleostei: Galaxiidae) from the Taieri River, South Island, New Zealand: a species complex revealed

We examined genetic differentiation among 23 samples of non-migratory river galaxias from 17 streams in the Taieri River system, South Island, New Zealand. Four major genetic types were found, two of which occur in narrow sympatry in one location. These were compared with topotypical material representing Galaxias anomalus from the Clutha system (Otago) and G. vulgaris from the Waimakariri system (Canterbury) in order to establish identity. Morphological examination of these four major genetic types revealed consistent concomitant differences. The results suggest that there are at least three species of river galaxias in the Taieri system: G. anomalus, G. vulgaris and at least one previously undescribed species. We propose that the genetic structuring and subsequent speciation of this group has been promoted by the absence of the marine juvenile phase that is found in five other members of the genus native to New Zealand. This structuring may be exacerbated by population fragmentation over the last century owing to the negative influence of introduced trout. The phylogenetic diversity within the river system mirrors the diverse flora and invertebrate fauna of the region, and has conservation implications that parallel those resulting from our improved knowledge of the New Zealand herpetofauna through the application of genetic analysis.     

Extremely low effective population sizes, genetic structuring and reduced genetic diversity in a threatened bumblebee species, Bombus sylvarum (Hymenoptera: Apidae)

Habitat fragmentation may severely affect survival of social insect populations as the number of nests per population, not the number of individuals, represents population size, hence they may be particularly prone to loss of genetic diversity. Erosion of genetic diversity may be particularly significant among social Hymenoptera such as bumblebees (Bombus spp.), as this group may be susceptible to diploid male production, a suggested direct cost of inbreeding. Here, for the first time, we assess genetic diversity and population structuring of a threatened bumblebee species (Bombus sylvarum) which exists in highly fragmented habitat (rather than oceanic) islands. Effective population sizes, estimated from identified sisterhoods, were very low (range 21-72) suggesting that isolated populations will be vulnerable to loss of genetic variation through drift. Evidence of significant genetic structuring between populations (theta = 0.084) was found, but evidence of a bottleneck was detected in only one population. Comparison across highly fragmented UK populations and a continental population (where this species is more widespread) revealed significant differences in allelic richness attributable to a high degree of genetic diversity in the continental population. While not directly related to population size, this is perhaps explained by the high degree of isolation between UK populations relative to continental populations. We suggest that populations now existing on isolated habitat islands were probably linked by stepping-stone populations prior to recent habitat loss.     

Significant patterns of population genetic structure and limited gene flow in a threatened macropodid marsupial despite continuous habitat in southeast Queensland, Australia

Many endangered species worldwide are found in remnant populations, often within fragmented landscapes. However, when possible, an understanding of the natural extent of population structure and dispersal behaviour of threatened species would assist in their conservation and management. The brush-tailed rock-wallaby (Petrogale penicillata), a once abundant and widespread rock-wallaby species across southeastern Australia, has become nearly extinct across much of the southern part of its range. However, the northern part of the species’ range still sustains many small colonies closely distributed across suitable habitat, providing a rare opportunity to investigate the natural population dynamics of a listed threatened species. We used 12 microsatellite markers to investigate genetic diversity, population structure and gene flow among brush-tailed rock-wallaby colonies within and among two valley regions with continuous habitat in southeast Queensland. We documented high and significant levels of population genetic structure between rock-wallaby colonies embedded in continuous escarpment habitat and forest. We found a strong and significant pattern of isolation-by-distance among colonies indicating restricted gene flow over a small geographic scale ( <10 km) and conclude that gene flow is more likely limited by intrinsic factors rather than environmental factors. In addition, we provide evidence that genetic diversity was significantly lower in colonies located in a more isolated valley region compared to colonies located in a valley region surrounded by continuous habitat. These findings shed light on the processes that have resulted in the endangered status of rock-wallaby species in Australia and they have strong implications for the conservation and management of both the remaining ‘connected’8 brush-tailed rock-wallaby colonies in the northern parts of the species’8 range and the remnant endangered populations in the south.     

Isozymes in macroalgae (seaweeds): genetic differentiation,genetic variability and applications in systematics

Use of isozymes (including allozymes) in studies of population genetics and systematics of seaweeds has increased sufficiently in the last decade to allow some generalization. Only a single locus has been observed for about half the enzymes analysed in seaweeds, compared with 29% in vascular plants. Compared with higher plants, macroalgal species generally have low amounts of electrophoretically detectable genetic variation; the lowest levels of genetic variation found in natural populations are those reported for seaweeds. Nonetheless, seaweeds show an association between levels of genetic diversity as revealed by isozymes and species-specific attributes, such as mating system and predominance of asexual versus sexual reproduction. In systematic studies, isozymes have revealed cryptic species and identified pairs of sibling taxa. The quaternary structure of enzymes appears to be conserved at the phylum level. With the current availability of improved techniques for enzyme electrophoresis and for data interpretation, we expect future studies utilizing isozyme electrophoresis to provide further insight into population and evolutionary processes in seaweeds.     

Isozyme variation in Calliandra calothyrsus (Leguminosae): its implications for species delimitation and conservation

Patterns of genetic diversity within and among populations of Calliandra calothyrsus, an important multipurpose tree species, were examined using isozyme analysis. C. calothyrsus is a widespread species distributed throughout Central America and southern Mexico, across a variety of environments. Morphologically and ecologically distinct populations can be identified within this range, but they are currently considered to represent a single species. C. calothyrsus has been introduced to many parts of the tropics, where it is cultivated as a source of fuelwood, animal fodder, green manure, and shade by rural communities. Some of these introductions are known to have originated from Guatemala, but very little is known about the genetic diversity of either the native or naturalized populations. Isozyme electrophoresis of 23 loci across 17 populations of C. calothyrsus indicated that the majority of genetic diversity was partitioned between populations (FST = 0.802) and that within-population heterozygosity was low (mean Ho = 0.057). Naturalized populations had lower than expected heterozygosities and were most similar to material from Santa Maria de Jesus, a natural population in southern Guatemala. Four distinct groups of populations were identified on the basis of Nei's genetic distances and Population Aggregation Analysis (PAA), and correlate with the morphological and ecological differences that can be observed within the species. The results are discussed in relation to species delimitation and conservation.     

Genetic diversity in an endangered alpine plant, Eryngium alpinum L. (Apiaceae), inferred from amplified fragment length polymorphism markers

Eryngium alpinum L. is an endangered species found across the European Alps. In order to obtain base-line data for the conservation of this species, we investigated levels of genetic diversity within and among 14 populations from the French Alps. We used the amplified fragment length polymorphism (AFLP) technique with three primer pairs and scored a total of 62 unambiguous, polymorphic markers in 327 individuals. Because AFLP markers are dominant, within-population genetic structure (e.g. FIS) could not be assessed. Analyses based either on the assumption of random-mating or on complete selfing lead to very similar results. Diversity levels within populations were relatively high (mean Nei's expected heterozygosity = 0.198; mean Shannon index = 0.283), and a positive correlation was detected between both genetic diversity measurements and population size (Spearman rank correlation: P = 0. 005 and P = 0.002, respectively). Moreover, FST values and exact tests of differentiation revealed high differentiation among populations (mean pairwise FST = 0.40), which appeared to be independent of geographical distance (nonsignificant Mantel test). Founder events during postglacial colonizations and/or bottlenecks are proposed to explain this high but random genetic differentiation. By contrast, we detected a pattern of isolation by distance within populations and valleys. Predominant local gene flow by pollen or seed is probably responsible for this pattern. Concerning the management of E. alpinum, the high genetic differentiation leads us to recommend the conservation of a maximum number of populations. This study demonstrates that AFLP markers enable a quick and reliable assessment of intraspecific genetic variability in conservation genetics.     

Genetic diversity and the reproductive system in related species of antirrhinum

BACKGROUND AND AIMS: Seven related species of Antirrhinum (A. siculum, A. majus, A. latifolium, A. linkianum, A. litigiosum, A. cirrhigherum and A. tortuosum) were studied in order to compare levels of genetic variation and its partitioning in them, and to check relationships between genetic patterns and the reproductive system. METHODS: Eight hundred and fifty-one plants were screened for variability at 13 allozyme loci by means of horizontal starch gel electrophoresis. Parameters of genetic diversity and its partitioning, the inbreeding coefficient as well as an indirect estimate of gene flow based on the equation: Nm = (1 - G(ST))/4G(ST), were calculated. KEY RESULTS: Genetic variability in A. siculum was found to be the lowest known in the genus. Mean values of F(IT) and F(IS) were mostly positive and not significantly different from zero. Population differentiation (F(ST)) ranged between 6.1 in A. tortuosum and 17.6 in A. linkianum. The inbreeding coefficient within populations ranged between F(IS) = -0.5 in A. tortuosum and F(IS) = 1 in A. siculum. Estimates of gene flow ranged between Nm = 15 in A. majus (considered as very high) to Nm = 0.42 in A. siculum (considered as low). CONCLUSIONS: Correlation was found between levels of diversity and differentiation on one hand, and the reproductive system of the studied taxa on the other. Striking differences among species in the inbreeding coefficient (F(IS)) show different reproductive systems, which mostly support previous reports. Strategies for the conservation of A. siculum are recommended, such as preservation of natural populations as well as ex situ preservation of seeds from different populations.     

New chloroplast primers for intraspecific variation in Dicranum scoparium Hedw. (Dicranaceae) and amplification success in other bryophyte species

Primers for four loci that amplify cpDNA regions have been designed for population genetic analyses in Dicranum scoparium Hedw. and compared with trnL(UAA)5'exon-trnF. All loci showed intraspecific variation with a number of haplotypes ranging between two and six. trnH-psbA(Dic) showed an intercontinental disjunction, but no variability within the four Swiss populations surveyed, whereas the three remaining loci displayed intrapopulation variability in at least one population (rps19-rpl2, rpoB, trnT-rps4). These primers were additionally tested on 22 bryophytes and three fern species. The primers amplified mostly in mosses and liverworts, but less well in ferns, pointing to their evolutionary distance from the bryophytes.     

A population genetic study of the endangered plant species Limonium dufourii (Plumbaginaceae) based on amplified fragment length polymorphism (AFLP)

Limonium dufourii ( Plumbaginaceae ) is a triploid species with obligate apomictic reproduction and is endemic to the East Mediterranean coast of Spain, where it is present in only six populations, most of which have a very low number of individuals. Genetic variation and population structure in this species was studied using amplified fragment length polymorphisms (AFLPs) as markers, using the same individuals as in a previous study with random amplified polymorphic DNA (RAPD). Three different primers provided 252 bands of which 51 were polymorphic among the 152 individuals analysed. Those polymorphic bands were able to define 65 different phenotypes, of which all but two were present in only one population. The comparative analyses of data from AFLPs with those from RAPDs show a high degree of concordance. Additionally, and given the nature of these markers, we propose the estimation of nucleotide divergences from AFLP patterns. Relationships among the different AFLP patterns and the estimates of population genetic parameters obtained with this evolutionary distance are in good agreement with previous results. These analyses show that substantial genetic variability and differentiation exist within and among populations of L. dufourii . Their higher reproducibility and the possibility of obtaining estimates of nucleotide divergence make AFLPs a much better DNA fingerprinting technique.     

Patterns of allozyme diversity in several selected rare species in Korea and implications for conservation

Because some endemic plants appear to be adapted to a narrow setof environmental conditions with limited genetic diversity, an analysis ofpopulation genetic structure is necessary to fully evaluate the impact of rarityon genetic variation. Listed as endangered species in Korea, only fewpopulations of Abeliophyllum distichum, Leonticemicrorhyncha, Bupleurum euphorbioides, andBerchemia berchmiaefolia were found. A reduced level ofgenetic variation in B. berchemiaefolia is consistent withthe occurrence of a genetic bottleneck and inbreeding. Leonticemicrorhyncha differed dramatically from other taxa in its observed level of geneticvariation, probably due to its predominant selfing. The level ofallozyme variation maintained by A. distichum was high forendemic species. Compared to species with similar traits, A.distichum maintained a relatively higher genetic diversity, probably dueto floral heteromorphism and preferred outcrossing. Bupleurumeuphorbioides maintained a higher genetic diversity due to outcrossing,but at the individual locus, deficiency of heterozygosity prevailed. Probablyinbreeding between local neighborhoods was frequent because A.distichum and B. euphorbioides were pollinatedby small fly species which might be less effective as a pollen dispersal, andtheir visits were extremely scarce and controlled by the weather conditions.Since much of the species-to-species variation in genetic diversity is due tothe specific ecological and evolutionary history of a species, any managementplan developed should be based on historical changes in the population size anddistribution to better predict the amounts and patterns of genetic diversity.     

A prime inference on genetic diversity (RAPDs) in the marine fish Atherinella brasiliensis (Teleostei,Atherinopsidae) from Southern Brazil

Da Silva Cortinhas, M. C., Glienke, C., Prioli, A. J., Noleto, R. B., Matoso, D. A. and Cestari, M. M. 2010. A prime inference on genetic diversity (RAPDs) in the marine fish Atherinella brasiliensis (Teleostei, Atherinopsidae) from Southern Brazil. —Acta Zoologica (Stockholm) 91 : 242–248 As a result of the importance of Atherinella brasiliensis in estuarine environments, random amplified polymorphic DNA (RAPD) markers were used to verify the genetic diversity in A. brasiliensis from two different places in Paranaguá Bay (Paraná State) and one from the Conceição Lagoon (Santa Catarina State). Cytogenetic data have shown a high karyotypic diversity in some populations, although in others this peculiarity demonstrates rearrangements such as heterochromatinization. In the present study, a low level of genetic structuring between the samples from Conceição Lagoon compared with the others was observed through principal coordinate analysis (PCO), analysis of molecular variance and Mantel test according to 79 RAPD markers. As this specie does not perform horizontal migration and the individuals of Conceição Lagoon are isolated, three hypotheses are proposed to explain the results: (i) similar environments may show homogeneous populations not depending on the geographical distance, (ii) because vicariant events that formed the bays occurred in a recent period, the fragmentation effects over the structuring of the genetic diversity may still be low and not totally detectable by the RAPD technique and (iii) the isolation time or the number of generations may not be enough to promote a possible differentiation and genetic structuring between the specimens of these three places. The specimens of these places present a low level of differentiation and genetic structuring so we can consider them as a unique homogeneous population.     

Genetic structure of populations of the threatened eastern massasauga rattlesnake, Sistrurus c. catenatus: evidence from microsatellite DNA markers

Throughout its distribution in North America, the threatened eastern massasauga rattlesnake ( Sistrurus c. catenatus ) persists in a series of habitat-isolated disjunct populations of varying size. Here, we use six microsatellite DNA loci to generate information on the degree of genetic differentiation between, and the levels of inbreeding within populations to understand how evolutionary processes operate in these populations and aid the development of conservation plans for this species. Samples were collected from 199 individuals from five populations in Ontario, New York and Ohio. Our results show that all sampled populations: (i) differ significantly in allele frequencies even though some populations are < 50 km apart, and may contain genetically distinct subpopulations < 2 km apart; (ii) have an average of 23% of alleles that are population specific; and (iii) have significant F _IS values (mean overall F _IS= 0.194) probably due to a combination of Wahlund effects resulting from fine-scale genetic differentiation within populations and the presence of null alleles. Our results imply that massasauga populations may be genetically structured on an extremely fine scale even within continuous populations, possibly due to limited dispersal. Additional information is needed to determine if dispersal and mating behaviour within populations can account for this structure and whether the observed differentiation is due to random processes such as drift or to local adaptation. From a conservation perspective, our results imply that these massasauga populations should be managed as demographically independent units and that each has high conservation value in terms of containing unique genetic variation.