Allozyme diversity and systematics inAnnonaceae—a pilot project

Five species ofAnnona and one species fromArtabotrys, Cananga, Polyalthia, andRollinia were investigated in regard to 11 different allozyme loci. Preliminary studies on small population samples ofAnnona suggest genetic uniformity in three species and variability within and between populations in two other species. The allotetraploid origin ofA. glabra is clearly shown by its hybrid enzyme bands. The genetic distance between fiveAnnona species partly corresponds with their morphological relationships; onlyA. muricata appears more separated than is suggested by morphology. A comparison of the five genera demonstrates close relationship betweenAnnona andRollinia. Two enzyme loci are identical within all taxa investigated and possibly may serve as a genetic marker for the family.     

MAJOR GENETIC DIFFERENCES BETWEEN CROWN-OF-THORNS STARFISH (ACANTHASTER PLANCI) POPULATIONS IN THE INDIAN AND PACIFIC OCEANS

Spatial variation in allelic frequencies at nine allozyme loci were assayed in 20 populations of the crown-of-thorns starfish, Acanthaster planci, collected throughout the Pacific and Indian Oceans. These data were analyzed together with published data, for the same loci, from an additional 19 populations, giving a total sample size of approximately 1800 individuals. There was a marked discontinuity between the Indian and Pacific Ocean populations, but those off Western Australia and from the Southeast Asian region had a strong Pacific affinity. The genetic groups were congruent with the distributions of two color morph groups: gray-green to red-brown forms in the Pacific and a blue to pale red form in the Indian Ocean. These patterns of genetic structure are similar to those described for the starfish Linckia laevigata, which has similar life-history characteristics. Vicariant events may have influenced some populations within the Pacific, but the allozyme data cannot resolve the effects of these events clearly. Patterns of variation within regions were consistent with isolation by distance, but, at larger scales, were obscured by regional vicariance and some outliers, particularly by apparently high levels of gene flow between Japan and the Great Barrier Reef, Australia. Apparent gene flow between population pairs was not closely related to present-day ocean currents. The results demonstrate a strong influence of allopatric separation on genetic divergence at large geographic scales, but also show evidence of slow rates of change in gene frequencies consistent with the large population sizes of this species. Low levels of divergence between groups demonstrate the genetic structure is recent (Pleistocene) and are likely responses to changes in climate and sea level.     

Genetic variation in populations of the morphologically and ecologically variable fern Stegnogramma pozoi subsp. mollissima (Thelypteridaceae) in Japan

In Japanese Stegnogramma pozoi subsp. mollissima (Fisher ex Kunze) K. Iwats. there is the intrasubspecific variation among rbcL sequences. Northern and southern plants are genetically differentiated for maternally inherited cpDNA. In the present study we examined allozyme polymorphisms to test the hypothesis that northern and southern plants may be separate species. Based on allozyme data, the degree of gene flow among populations was estimated to be large. The artificial crossing experiments between cpDNA haplotypes also suggested that isolation has not developed among these cpDNA haplotypes. However, interpopulation genetic differentiation in cpDNA was observed even in the small area at the foot of Mt. Hakone, and the cpDNA haplotypes appear to have different habitat preferences.     

Levels of genetic polymorphism: marker loci versus quantitative traits.

Species are the units used to measure ecological diversity and alleles are the units of genetic diversity. Genetic variation within and among species has been documented most extensively using allozyme electrophoresis. This reveals wide differences in genetic variability within, and genetic distances among, species, demonstrating that species are not equivalent units of diversity. The extent to which the pattern observed for allozymes can be used to infer patterns of genetic variation in quantitative traits depends on the forces generating and maintaining variability. Allozyme variation is probably not strictly neutral but, nevertheless, heterozygosity is expected to be influenced by population size and genetic distance will be affected by time since divergence. The same is true for quantitative traits influenced by many genes and under weak stabilizing selection. However, the limited data available suggest that allozyme variability is a poor predictor of genetic variation in quantitative traits within populations. It is a better predictor of general phenotypic divergence and of postzygotic isolation between populations or species, but is only weakly correlated with prezygotic isolation. Studies of grasshopper and planthopper mating signal variation and assortative mating illustrate how these characters evolve independently of general genetic and morphological variation. The role of such traits in prezygotic isolation, and hence speciation, means that they will contribute significantly to the diversity of levels of genetic variation within and among species.     

Population Genetic Structure of Endemic and Endangered Yellow Catfish, Horabagrus brachysoma, Using Allozyme Markers

The allozyme variation and population genetic structure of Horabagrus brachysoma in three natural populations from the southern part of the Western Ghats region, India, were investigated by polyacrylamide gel electrophoresis. Variations at 14 loci from 14 enzyme systems were analyzed. The allozyme analysis revealed a high level of genetic variation in this species, with an average of observed alleles per locus of 2.357 and observed heterozygosity of 0.178. The positive value of the fixation index (FIS=0.507) implied a significant deficiency of heterozygosity at the population level. The highly significant probability (P<0.0001) for the overall loci suggested that the three sample sets were not part of the same gene pool.     

Bottlenecks, drift and differentiation: the fragmented population structureof the saltmarsh beetle Pogonus chalceus

We investigated the distribution of genetic variation within and between 10 fragmented populations of the saltmarsh beetle Pogonus chalceus in the region of Flanders (Belgium) representing all extant populations of the species in that region by using allozyme and microsatellite markers. Beetle population size and habitat area failed to explain a significant part of the genetic variability. Microsatellite allelic diversity was sensitive to population size differences but not to saltmarsh area estimates. Heterozygosities of both marker types and allozyme allelic diversity on the other hand showed no significant correlation to population size and saltmarsh area. There was also no correlation between geographical and genetic distances among populations. Evidence was found for past bottlenecks in some of the smallest populations. Maximum likelihood methods using the coalescent approach revealed that the proportion of common ancestors was also high in those small populations. 35% of our studied individuals, especially in the largest populations showed a relative wing size smaller than 70%. Moreover, only six out of the 10 studied populations showed a few individuals with functional flight musculature. In conclusion, the overall pattern of distribution of genetic variation and the low flight capacity did not support an equilibrium model of population structure in P. chalceus, but mainly suggested a lack of regional equilibrium with both drift and gene flow influences.     

Regional subdivision in wild barley allozyme variation: adaptive or neutral?

We examined the adaptive importance of allozyme variation in wild barley (Hordeum spontaneum). The test involved a nested sampling design with four population groups, each representing a different environment, and a comparison of observed allozyme variation with that expected under the assumption that allozymes are not neutral. Measurements of plant fitness in indigenous and alien environments in reciprocal introductions of seeds and seedlings in the four environments provided a guideline for the expected pattern of allozyme variation. The results showed considerable variation in both the degree of regional and population subdivision and the pattern of the subdivision among loci. The observed pattern of variation was ambiguous. Although two alleles exhibited a pattern of distribution that cannot be explained by genetic drift as a function of geographic distance, we failed to detect either a significant relationship between genetic distance and environmental similarity or any favored epistatic allele combinations across the four environments. Our results suggest that interpretation of allozyme variation in wild barley as adaptive and directly related to local environment still needs justification. Although we could not reject the null hypothesis, a proposed methodology seeking a concordance between observed and "adaptive" (i.e., expected under hypothesis that allozymes are not neutral) allozyme variation may prove to be effective in resolving the neutralist-selectionist debate when applied to other species.     

Genetic differentiation between alternate-year cohorts of Xestia tecta (Lepidoptera,Noctuidae) in Finnish Lapland

Several species in the noctuid genus Xestia exhibit periodic dynamics, with two coexisting cohorts flying in alternate years. The population dynamics and two-year life cycle of Xestia moths suggest that the two cohorts are more or less isolated in time. Typically one cohort is abundant and the other one is rare. Knowledge of the extent of isolation between the two cohorts is important to fully understand the population dynamics and the evolution of alternate-year flight in these species. We applied allozyme electrophoresis and mitochondrial genome sequencing to infer the extent of genetic differentiation among different cohorts of Xestia tecta (Hübner) within the same geographical area as well as between cohorts with opposite-phase flight pattern in different geographical regions. We found no evidence for substantial genetic differentiation and isolation between the even- and odd-year cohorts in eastern Lapland, nor between the cohorts in eastern and western Lapland. The most informative markers were the most polymorphic allozyme loci (Pgm and Mpi) and the AT-rich region in the mtDNA. However, owing to the generally low levels of genetic variation it was not possible to establish conclusively the degree of genetic isolation between the different cohorts. We discuss the implications of our results in relation to two different hypotheses which could account for this pattern: ongoing gene flow between different cohorts and recent common ancestry.     

Extremely low levels of genetic diversity in the terrestrial orchid Epipactis thunbergii (Orchidaceae) in South Korea: implications for conservation

Levels of allozyme variation, population genetic structure, and fine-scale genetic structure (FSGS) of the rare, both sexually and clonally reproducing terrestrial orchid Epipactis thunbergii were examined for eight ( N  = 734) populations in a 20 × 20-km area in South Korea. Twenty-three putative allozyme loci resolved from 15 enzyme systems were used. Extremely low levels of allozyme variation were found within populations: the mean frequency of polymorphic loci was 3.8% [isocitrate dehydrogenase ( Idh-2 ) with two alleles was polymorphic across populations], the mean number of alleles per locus was 1.04, and the mean expected heterozygosity was 0.013. The overall fixation index was not significantly different from zero ( F _IS = 0.069), although the species is self-compatible. However, a significantly high degree of population differentiation was found between populations at Idh-2 ( F _ST = 0.388) in the studied area. Furthermore, spatial autocorrelation analyses revealed a significant FSGS (up to 3 m) within populations. These observations suggest that the main explanatory factors for the extremely low levels of genetic diversity and the shaping of the population genetic structure of E. thunbergii are genetic drift as a result of a small effective population size, a restricted gene flow, and the isolation of populations. Considering the current genetic structure of E. thunbergii , three guidelines are suggested for the development of conservation strategies for the species in South Korea: (1) protection of habitats of standing populations; (2) prohibition by law of any collection of E. thunbergii ; and (3) protection of nearby pollinator populations, given the fact that fruit set in natural habitats is very low.  © 2007 The Linnean Society of London, Botanical Journal of the Linnean Society , 2007, 155 , 161–169.     

Enzyme Variability in the DROSOPHILA WILLISTONI Group. III. Amounts of Variability in the Superspecies, D. PAULISTORUM

The semispecies composing the superspecies, Drosophila paulistorum, have been analyzed for genetic variation at 17 enzyme loci. On the average a population of D. paulistorum is polymorphic for 55-67% of its loci and an average individual is heterozygous at 21% of its loci. The pattern of genetic variability found supports the hypothesis that allozyme variation is maintained in natural populations by some form of balancing selection. Evidence is presented which supports the hypothesis that glucose-metabolizing enzymes are less genetically variable than non-glucose-metabolizing enzymes. The known genetic relationships between the semispecies of D. paulistorum are discussed in the light of the frequencies of alleles at allozyme loci.     

Genetic variation in populations of the morphologically and ecologically variable fern <Emphasis Type="Italic">Stegnogramma pozoi</Emphasis> subsp. <Emphasis Type="Italic">mollissima</Emphasis> (Thelypteridaceae) in Japan

In Japanese Stegnogramma pozoi subsp. mollissima (Fisher ex Kunze) K. Iwats. there is the intrasubspecific variation among rbcL sequences. Northern and southern plants are genetically differentiated for maternally inherited cpDNA. In the present study we examined allozyme polymorphisms to test the hypothesis that northern and southern plants may be separate species. Based on allozyme data, the degree of gene flow among populations was estimated to be large. The artificial crossing experiments between cpDNA haplotypes also suggested that isolation has not developed among these cpDNA haplotypes. However, interpopulation genetic differentiation in cpDNA was observed even in the small area at the foot of Mt. Hakone, and the cpDNA haplotypes appear to have different habitat preferences. Received: May 1, 2001 / Accepted: September 30, 2001     

Reproductive biology and genetic diversity of a cryptoviviparous mangrove aegiceras corniculatum (Myrsinaceae) using allozyme and intersimple sequence repeat (ISSR) analysis

Mangroves consist of a group of taxonomically diverse species representing about 20 families of angiosperms. However, little is known about their reproductive biology, genetic structure, and the ecological and genetic factors affecting this structure. Comparative studies of various mangrove species are needed to fill such gaps in our knowledge. The pollination biology, outcrossing rate, and genetic diversity of Aegiceras corniculatum were investigated in this study. Pollination experiments suggested that the species is predominantly pollinator-dependent in fruit setting. A quantitative analysis of the mating system was performed using progeny arrays assayed for intersimple sequence repeat (ISSR) markers. The multilocus outcrossing rate (tm) was estimated to be 0.653 in a wild population. Both allozyme and ISSR were used to investigate genetic variation within and among populations. The combined effects of founder events and enhanced local gene flow through seedling dispersal by ocean currents apparently played an important role in shaping the population genetic structure in this mangrove species. Both allozyme variation (P = 4.76%, A = 1.05, HE = 0.024) and ISSR diversity (P = 16.18%, A = 1.061, HE = 0.039) were very low at the species level, in comparison with other woody plants with mixed-mating or outcrossing systems. Gene differentiation among populations was also low: allozyme GST = 0.106 and ISSR GST = 0.178. The unusually high genetic identities (0.997 for allozyme and 0.992 for ISSR loci), however, suggest that these populations are probably all descended from a common ancestral population with low polymorphism.     

Birth of a hotspot of intraspecific genetic diversity: notes from the underground

Hotspots of intraspecific diversity have been observed in most species, often within areas of putative Pleistocene refugia. They have thus mostly been viewed as the outcome of prolonged stability of large populations within the refugia. However, recent evidence has suggested that several other microevolutionary processes could also be involved in their formation. Here, we investigate the contribution of these processes to current range-wide patterns of genetic diversity in the Italian endemic mole Talpa romana, using both nuclear (30 allozyme loci) and mitochondrial markers (cytochrome b sequences). Southern populations of this species showed an allozyme variation that is amongst the highest observed in small mammals (most populations had an expected heterozygosity of 0.10 or above), which was particularly unexpected for a subterranean species. Population genetic, phylogeographic and historical demographic analyses indicated that T. romana populations repeatedly underwent allopatric differentiations followed by secondary admixture within the refugial range in southern Italy. A prolonged demographic stability was reliably inferred from the mitochondrial DNA data only for a population group located north and east of the Calabrian peninsula, showing comparatively lower levels of allozyme variability, and lacking evidence of secondary admixture with other groups. Thus, our results point to the admixture between differentiated lineages as the main cause of the higher levels of diversity of refugial populations. When compared with the Pleistocene evolutionary history recently inferred for species from both the same and other geographic regions, these results suggest the need for a reappraisal of the role of gene exchange in the formation of intraspecific hotspots of genetic diversity.     

Spatial and temporal population genetic structure of the butterfly aglais urticae L. (Lepidoptera, nymphalidae)

The genetic diversity and the temporal and spatial genetic population structure of the butterfly Aglais urticae, a highly mobile species, were studied by allozyme electrophoresis. High levels of allozyme diversity were found. Most of the total genetic diversity occurred at the within-population scale rather than at the between-population scale. This variation could not be accounted for by Wright's model of 'isolation by distance'. No significant temporal variation was observed for those populations that were sampled in different years. A process combining high movement rate between neighbouring patches, long-distance migration and rare extinction/recolonization is suggested to explain the observed genetic structure. This hypothesis is favoured over an island model of population structure because migration in A. urticae is uniform neither with distance nor with time.     

Allozyme Diversity in Populations of Cymbidium goeringii (Orchidaceae)

Abstract: Using 14 allozyme loci, we investigated levels of genetic diversity within populations, and degree of genetic divergence among 24 populations of Cymbidium goeringii (Orchidaceae) in Korea and Japan. Cymbidium goeringii maintains high levels of genetic diversity both at population (mean expected heterozygosity, H _e = 0.238) and species levels (0.260). Means of H _e found in 24 populations were not significantly different from each other. About 90 % of the total variation in the species is common to all populations (mean G _ST = 0.108). No unique allele was found in any population. The indirect estimate of gene flow based on the mean G _ST was high ( Nm = 2.06). Nei's genetic identities for pairs of populations had high values (mean = 0.974 [SD = 0.013]). The Mantel-Z test showed a significant correlation between genetic distance and geographic distance. However, the mean G _ST value between 17 populations in Korea and seven Japanese populations was relatively low (0.029), even though the land connection between the southern Korean peninsula and southern Japanese archipelagos has not existed since the middle Pleistocene. Large numbers of small seeds of C. goeringii might travel long distances by wind from populations to populations both in Korea and Japan, increasing genetic diversity within populations and maintaining low genetic differentiation among populations.     

EVIDENCE OF A BIOGEOGRAPHIC BREAK BETWEEN POPULATIONS OF A HIGH DISPERSAL STARFISH: CONGRUENT REGIONS WITHIN THE INDO-WEST PACIFIC DEFINED BY COLOR MORPHS,mtDNA, AND ALLOZYME DATA

Both mtDNA variation and allozyme data demonstrate that geographic groupings of different color morphs of the starfish Linckia laevigata are congruent with a genetic discontinuity between the Indian and Pacific Oceans. Populations of L. laevigata sampled from Thailand and South Africa, where an orange color morph predominates, were surveyed using seven polymorphic enzyme loci and restriction fragment analysis of a portion of the mtDNA including the control region. Both allozyme and DNA data demonstrated that these populations were significantly genetically differentiated from each other and to a greater degree from 23 populations throughout the West Pacific Ocean, where a blue color morph is predominant. The genetic structure observed in L. laevigata is consistent with traditional ideas of a biogeographic boundary between the Indian and Pacific Oceans except that populations several hundreds kilometers off the coast of north Western Australia (Indian Ocean) were genetically similar to and had the same color morphs as Pacific populations. It is suggested that gene flow may have continued (possibly at a reduced rate) between these offshore reefs in Western Australia and the West Pacific during Pleistocene falls in sea level, but at the same time gene flow was restricted between these Western Australian populations and those in both Thailand and South Africa, possibly by upwellings. The molecular data in this study suggest that vicariant events have played an important role in shaping the broadscale genetic structure of L. laevigata. Additionally, greater genetic structure was observed among Indian Ocean populations than among Pacific Ocean populations, probably because there are fewer reefs and island archipelagos in the Indian Ocean than in the Pacific, and because present-day surface ocean currents do not facilitate long-distance dispersal.     

Patterns of variation of a common fern (Athyrium filix-femina; Woodsiaceae): population structure along and between altitudinal gradients

Genetic variability of Athyrium filix-femina populations was evaluated with regard to phenotypic, allozyme, and RAPD variation in 20 Swiss populations along five altitudinal gradients at four different elevations in the northern Swiss Alps. Additionally, allozyme and phenotypic variations in one Italian and two Spanish populations were compared with the variation in the Swiss populations. We hypothesized that there will be statistically significant genetic differences among populations of different altitudes and sites. The results showed no substantial correlation between genetic variation and phenotypic variation among Swiss populations. These results imply that outbreeding and effective gene exchange (long-distance spore dispersal) are the keys to population structure in this fern species, and as a consequence, phenotypic plasticity is assumed to be favored. This contrasts with results found in similar studies of herbaceous flowering plants where genetic adaptation to gradients like altitude is common. However, when data from the more distant Italian and Spanish populations of A. filix-femina were included, significant variation was detected.     

Enzyme polymorphism, morphometric variation and population structure in a restricted endemic, Silene diclinis (Caryophyllaceae)

Silene diclinis is a dioecious perennial herb, restricted to a small area of the SE Spanish province of Valencia. Polyacrylamide gel electrophoresis and morphometric analysis were carried out on cultivated material derived from seed collected from the main population of S. diclinis at Játiva. Only two out of 26 inferred enzyme loci showed substantial variation. There was some indication of population subdivision in both. The esterase locus also showed homozygote excess within the population as a whole and within one subpopulation. Morphological characters showed significant differentiation between subpopulations.
Low allozyme variability characterizes populations of most endemic species that have been investigated. Isolation and historical events may have been as important as population size or edaphic specialization in causing this low variability. But S. diclinis resembles other outbreeding plants in showing population structure and homozygote excess, suggesting restricted within-population gene flow. Mechanisms that may restrict gene flow in S. diclinis include environmentally-induced differences in flowering time, poor seed dispersal and possibly short pollinator foraging distances. Morphological variability and the incidence of developmental abnormalities are speculatively linked with homozygosity. Storage of genetic diversity in space can occur in restricted as well as widespread species and sampling for gene banking should then be spatially representative. Silene diclinis is threatened by continued habitat loss and consequent genetic erosion; its long-term future may depend on the gene bank and the botanic garden.     

Genetic evidence for population expansion in Hydrotaea irritans (Fallèn) (Diptera: Muscidae)

Allozyme variation in the sheep headfly Hydrotaea irritans was studied on two spatial scales. Geographic variation among seven Danish and one Dutch population revealed significant but rather low genetic differentiation with F _ST = 0.01 over all loci. The Dutch population was on average not more different from the Danish populations than the Danish populations from each other. Allele frequencies were very skewed with the most common allele always exceeding 0.85 and usually 0.9 in frequency, but with many rare alleles at some loci. Tests for neutrality of the variation at the nine polymorphic loci revealed highly significant deviations from expected homozygosity in this species, which was not found in a comparative analysis of allozyme variation at similar loci of seven other Hydrotaea species. To explain the peculiar observed pattern of allozyme variation in H. irritans , it is suggested that this species has successfully expanded its range and spread through northern and central Europe in the recent past. Alternatively, H. irritans may have recently invaded a new niche, resulting in increased abundance of the species and subsequent dispersal to former areas of the species distribution.     

Evidence of sexuality in European Rubus (Rosaceae) species based on AFLP and allozyme analysis

Reproduction of polyploid Rubus species is described as facultatively apomictic. Pollination is needed for seed set, but most seedlings are produced asexually by pseudogamy. Although sexual processes may occur, clonal diversity can be extremely low. We performed a pollination experiment to investigate the breeding system and used allozyme and AFLP markers to analyze genetic variation among and within seed families in R. armeniacus and R. bifrons. Pollination either with self or outcross pollen was necessary to trigger seed set. Outbreeding marginally increased the number and quality of seeds compared with selfing. The enzyme PGI revealed some genetic variation within seed families. Seven other enzyme systems were monomorphic. The more detailed AFLP analyses with five primer pairs detected the same rate of genetic variation (14-17% of seedlings were genetically distinct) and confirmed the allozyme results for the same individuals. No genetic variation was found between the seed families from within a species collected in widely separated populations, but clear species-specific differences were observed. The results support the view that polyploid Rubus species are pseudogamous apomicts with low genetic diversity among and within seed families. However, sexual reproduction occasionally occurs and contributes to the maintenance of genetic variation within natural populations.