Mating behavior and time budget of an androdioecious crustacean,Eulimnadia texana (Crustacea: Conchostraca)

The clam shrimp,Eulimnadia texana (Crustacea, Conchostraca), is found in freshwater ephemeral environments throughtout the United States. Individual clam shrimp of this species are either hermaphroditic or male, a relatively rare mating system for animals known as androdioecy. Comparison of sex ratios between four neighboring populations ofE. texana in Southern New Mexico showed wide variation in the ratio of males to hermaphrodites with males making up as much as 42% of some populations and not occurring at all within others. Since little is known about the behavior of this species, an ethogram and time budget were prepared based on observations of laboratory populations. Males attempt to clasp hermaphrodites prior to mating. Precopulatory mate guarding occurs in this species. Outcrossing generally occurs during mate guarding and after the hermaphrodite molts. Hermaphrodites, however, seem to control the mating process. Successful mating by males never occured if the hermaphrodite struggled with him; hermaphrodite will self in the presence of males.     

Biosystematics of two sympatric species ofEucharis (Amaryllidaceae)

Eucharis candida andE. formosa are two often sympatric species of bulbous geophytes restricted to neotropical rain forest understory. The species are most common in eastern Ecuador, and are the only two east Ecuadorean species of the genus found north of the Pastaza valley. Data from phenetic, karyotypic, and preliminary isozyme electrophoretic analyses of both species are represented. The species are distinguishable phenetically and karyologically, but isozyme-based relationships are more complex. Phenetic resolution of the isozyme phenotypes supports recognition of two species in Ecuador. A Peruvian isolate ofE. formosa, though not morphologically distinct, shows both allozyme and chromosomal divergence from Ecuadorean populations. Cladistic relationships based on overall allozyme data do not support species distinction, but a novel electrophoretic phenotype for glutathione reductase is shared only by individuals ofE. candida. An apparent geographic component within the monophyletic groups resolved in the cladogram suggests that some degree of gene flow between these two species has been maintained without the complete loss of morpholgoical species identity. This may have been mediated either by artificial population structures due to a probable long history of cultivation, or via Pleistocene refugia effects. Both species may have originated in eastern Ecuador from a common ancestral population which has since radiated outward, perhaps several times.     

Variation in DNA microsatellites of the ferruginous pygmy-owl (Glaucidium brasilianum)

Eleven polymorphic microsatellite loci were used to assess genetic variation in the ferruginous pygmy owl (Glaucidium brasilianum) from North America. Analysis of genotypic variation suggests restricted gene flow between pygmy-owl populations in Arizona-Sonora and Sinaloa, and Texas and the remaining states in Mexico. The Arizona-Sonora population showed signs of a recent genetic bottleneck, an observation supported by low population estimates for Arizona (13–117 individuals). Heterozygosity in Arizona, however, was equal to levels recorded throughout Mexico and Texas. Congruent patterns revealed by nuclear (microsatellites) and mitochondrial DNA that indicate Arizona-Sonora and Texas populations are distinct from adjacent populations in Mexico, thus emphasizing need for the design and implementation of separate management plans for recovery and conservation efforts. Revealing evidence of distinct groups within the pygmy-owl populations in North America, results from this study may be used to make management decisions for the recovery and conservation of this species.     

Mitochondrial DNA Variation and Phylogeography of the Ferruginous Pygmy-Owl (Glaucidium brasilianum)

Sequences from the mitochondrial cytochrome b gene were used to examine patterns of variation within and among populations of the ferruginous pygmy-owl (Glaucidium brasilianum) from both North America (including populations from Mexico) and South America. As currently conceived, G. brasilianum is paraphyletic, with North American and South American clades representing two distinct groups that should be recognized as the distinct species G. ridgwayi and G. brasilianum, respectively. Within the G. ridgwayi clade, populations from Arizona, Sonora, and Sinaloa are genetically distinct and share no mitochondrial haplotypes with populations occurring in Texas and other regions of Mexico. According to nested clade analysis this separation may be the consequence of past fragmentation that predates the origin of the Sonora desert. In addition, gene flow between the Arizona/Sonora/Sinaloa populations and the Texas/other Mexico populations is practically nonexistent, with estimates being approximately one individual every 10 generations. Collectively, these data suggest that the Arizona/Sonora/Sinaloa clade should be recognized as either a distinct subspecies or phylospecies from the group containing populations in Texas and the remainder of Mexico. These data should be used as guidelines for pygmy-owl recovery and conservation, as they meet the recommendations of task 3 of the pygmy-owl recovery plan that lists genetic data as essential information for pygmy-owl management.     

Gene variation and gene flow inOrchis morio (Orchidaceae) from Italy

Data are presented on genetic variation at 27 enzyme loci of the Green-Winged orchid,Orchis morio, in 18 population samples from Italy. The existence in Italy of two subspecies, i.e. subspp.morio andpicta, is not supported by allozyme data. No genetic heterogeneity was found betweenmorio-like andpicta-like samples and specimens. Moreover, morphological transition between the two forms was observed in different Italian populations. The parameters of genetic variability estimated forO. morio populations are consistent with those found among monocotyledon plants, and among those outcrossing, animal-pollinated and with wind-dispersed seeds. Genetic diversity of ItalianO. morio is mostly within populations. Correspondingly, low values of interpopulational genetic distance were found. This appears to be due to high levels of gene flow, which were estimated with different methods. The lack ofO. longicornu from Italian samples, as well as of any hybrid withO. morio (F₁, backcrossed or recombinant individuals) is demonstrated on the basis of genetic data. It is concluded that recurrent reports ofO. longicornu from Italy are due to confusion withO. morio or with otherOrchis species.     

Morphological variation inBemisia endosymbionts

Summary The ultrastructure of the endosymbionts of several populations of whitefly (Homoptera: Aleyrodidae) was examined using transmission electron microscopy. Consistent differences in morphology and relative number of endosymbionts were observed between species and biotypes of whitefly within the Bemisia taxon.Bemisia argentifolii (=B. tabaci B biotype) individuals from Hawaii, Florida, and Arizona contained two morphological types of microorganisms housed within the mycetocyte cells of immature whiteflies. In contrast, individuals from populations ofB. tabaci A biotype from Arizona and Mexico, andB. tabaci Jatropha biotype from Puerto Rico, consistently contained three distinct morphological types of microorganisms within their mycetocytes. Organisms fromB. tabaci A and Jatropha biotypes differed from each other in the relative frequency of each type of microorganism. These observations suggest that different whitefly biotypes may have variable combinations of micro-fauna, with some possibly unique to each group, and furthers the hypothesis that variation in whitefly endosymbionts may be associated with the development of biotypes.     

Molecular evidence of hybridisation between the endemic Phoenix canariensis and the widespread P. dactylifera with Random Amplified Polymorphic DNA (RAPD) markers

We used RAPD markers to test whether morphologically intermediate individuals between the Canarian endemic Phoenix canariensis and the widespread P. dactylifera correspond to hybrids. Consistent with previous allozyme evidence, the scarcity of appropriate RAPD markers to distinguish P. dactylifera and P. canariensis indicated a close genetic relationship among these species. Only two of the 54 ten-mer primers (OPM-8 and OPK-14) tested in 221 individuals from 7 localities in different islands enabled us to unambiguously identify both species. While P. canariensis possesses two exclusive monomorphic bands of 1000 bp and 750 bp (for OPM-8 and OPM-14, respectively), dactylifera is characterised by two bands of 900 bp and 950 bp for the same primers. The additivity of these taxon-specific bands in the individuals that were morphologically intermediate provided, for the first time, firm evidence for their hybrid origin. Because these hybridisation capabilities pose clear threats to the survival of the endemic P. canariensis and some individuals that had been morphologically characterised as pure P. canariensis revealed later a hybrid nature in the RAPD analysis, we suggest that RAPD markers be used to estimate the possible incidence of introgression in the scarce extant natural populations of P. canariensis. This procedure will provide a straightforward means to select target populations to implement the in situ conservation strategies suggested previously on the basis of allozyme research.We thank Michel Ferry for providing samples of Phoenix dactylifera from the Hort del Gat Research Station on Date Palm and Arid Land Farming Systems in Elche (Spain). The authors wish to thank Jaime OShanahan for his wise comments about Phoenix canariensis. This research was funded by the Gobierno de Canarias (94-2614). A PhD Research Fellowship (Direccion General de Universidades, Gobierno de Canarias) to Gonzalez-Perez. is also acknowledged.     

Genetic relationships between European and Siberian larch,Larix spp. (Pinaceae), studied by allozymes. Is the Polish larch a hybrid between these two species?

Thirteen populations ofLarix decidua subsp.decidua and subsp.polonica, and three populations ofL. sibirica were investigated by starch-gel electrophoresis. In the populations assayed 61 alleles at 17 loci were revealed. The allozyme data support the earlier observations about close relationships between these two larch species. Nei's genetic distances betweenL. decidua andL. sibirica were relatively small (D = 0.057), however, almost five times larger, on average, than those between populations of the same species. Results obtained in this study disagree withBobrov's hypothesis about the hybrid origin of the Polish larch and suggest a direct origin from the European larch.     

One Species or Several? Discordant Patterns of Geographic Variation between Allozymes and mtDNA Sequences among Spiders in the Genus Metepeira (Araneae: Araneidae)

Paradoxically, an allozyme study of Metepeira “spinipes” (sensu lato) demonstrated extensive gene flow among four populations whose members are nevertheless morphologically and behaviorally distinct. Initially, the authors tentatively concluded that the populations exhibited panmixis and suggested that local environmental effects accounted for the apparent morphological and behavioral differences. However, they later concluded that such differences were too great to be accounted for by the environment alone and that the four populations actually represented three different species. To confirm that the allozyme results were, in fact, artifactual, we reexamined the relationships among these populations by sequencing a portion of the 12S mtDNA ribosomal subunit. In contrast to the allozyme result, our results demonstrate good agreement between patterns of genetic and morphological/behavioral variation. We suggest (1) that the allozyme allele frequencies are homogenized by balancing selection, not gene flow as was previously concluded, and therefore (2) that this study provides another instance in which inferences about population structures from allozyme data are misleading.     

On shortcomings of using mtDNA sequence divergence for the systematics of hares (genus Lepus): An example from cape hares

Despite the well-known fact that evolutionary patterns of single genes or sequences are not necessarily paralleled by organismic evolution, using mitochondrial sequence divergence for inferring phylogenetic relationships among hare species is not uncommon. Earlier studies reported interspecific introgression in the genus Lepus that may slur systematic conclusions drawn exclusively from mtDNA data. We examined pairwise divergence in partial mtHV-1 sequences and nuclear gene pools based on microsatellite and allozyme loci separately in a South and North African population of cape hares, Lepus capensis, and did not find significant correspondence on the individual level within either population. Also, the former population had a significantly higher level of mtHV-1 sequence divergence compared to the latter, but levels of individual nuclear gene pool divergence did not differ significantly between the two populations. Hence, the absence of correspondence between mtHV-1 sequence divergence and nuclear gene pool divergence among individuals within a population might be independent of the population-specific level of differentiation among individuals. Our results complement earlier findings in hares, and strongly recommend to include nuclear gene pool evidence for systematic inferences within this genus, even under the absence of mitochondrial introgression.     

Reproductive success and genetic divergence among varieties of the rare and endangered <Emphasis Type="Italic">Astragalus cremnophylax</Emphasis> (Fabaceae) from Arizona,USA

Populations of Astragalus cremnophylax from Arizona and related A. humillimus from New Mexico were assessed for morphological, reproductive, genetic, and ecological differences. Results suggest that some of the taxa (i.e., A. cremnophylax var. cremnophylax and A. humillimus) are poor reproducers and environmental fluctuation is not likely a factor contributing to low fecundity in these species. Astragalus cremnophylax var. cremnophylax appears to have low fecundity due to inbreeding depression. Allozyme data from 18 loci show little within-population and taxon genetic variability. Most genetic variability is distributed among populations and taxa. The population of A. cremnophylax var. cremnophylax from the Grand Canyon (South Rim) was genetically depauperate, indicative of the bottleneck caused by historic trampling at this viewpoint. Cape Final, a population of A. cremnophylax var. cremnophylax on the Grand Canyon (North Rim) is genetically different from typical materials of this taxon (South Rim) and is more closely related to other taxa within this species complex. However, the other varieties of A. cremnophylax are more closely related to A. humillimus than the population at Cape Final, suggesting potential species status for Cape Final materials based on allozyme divergence.     

Alcohol dehydrogenase polymorphism in barrel cactus populations of Drosophila mojavensis

Starch gel electrophoresis revealed that the alcohol dehydrogenase (ADH-2) locus was polymorphic in two populations (from Agua Caliente, California and the Grand Canyon, Arizona) of cactophilic Drosophila mojavensis that utilize barrel cactus (Ferocactus acanthodes) as a host plant. Electromorphs representing products of a slow (S) and a fast (F) allele were found in adult flies. The frequency of the slow allele was 0.448 in flies from Agua Caliente and 0.659 in flies from the Grand Canyon. These frequencies were intermediate to those of the low (Baja California peninsula, Mexico) and high (Sonora, Mexico and southern Arizona) frequency Adh-2^S populations of D. mojavensis that utilize different species of host cacti.     

Distribution of parental DNA markers inEncelia virginensis (Asteraceae: Heliantheae), a diploid species of putative hybrid origin

Morphological, geographical and ecological evidence suggests thatEncelia virginensis is a true-breeding diploid species derived from hybrids ofE. actoni andE. frutescens. To test this hypothesis, we examined the chloroplast and nuclear DNA of severalEncelia species. PCR amplification targeted three separate regions of chloroplast DNA:trnK-2621/trnK-11,rbcL/ORF106, andpsbA3/TrnI-51, which amplify 2600bp, 3300bp and 3200bp fragments respectively. Restriction fragment analysis of chloroplast DNA revealed no variation that could be used to discriminate between the parent species. A RAPD analysis using 109 dekamer primers was used to analyze the nuclear genome.Encelia actoni andE. frutescens were distinguished by several high-frequency RAPD markers. In populations ofE. virginensis, these markers were detected in varying proportions, and no unique markers were found. Evidence from the nuclear genome supports the hypothesis thatE. virginensis is of hybrid origin. ThatE. virginensis may have arisen by normal divergent speciation followed by later introgression remains a possibility, however, and is not formally ruled out here. Diploid hybrid speciation inEncelia differs from other documented cases in that there are no discernible chromosome differences between the species, and all interspecific hybrids are fully fertile. In addition, apparent ecological selection against backcross progeny provides an external barrier to reproduction between F₁ progeny and the parental species. These characteristics suggest that hybrid speciation inEncelia may represent an alternative model for homoploid hybrid speciation involving external reproductive barriers. In particular, this may be the case for other proposed diploid hybrid taxa that also exhibit little chromosomal differentiation and have fertile F₁s.     

Genetic introgression between tetraploidDactylis glomerata subspp.reichenbachii andglomerata in the French Alps. Insight from morphological and allozyme variation

The morphology, phenology and allozyme polymorphism in seven tetraploidDactylis glomerata populations growing in the French Alps were studied. Based on habitat and morphological characteristics, two of the populations could be classified asD. glomerata subsp.reichenbachii, which is known to comprise diploid and tetraploid plants growing exclusively on dolomite, on south-facing meadows. This subspecies has been previously recorded in the Italian and Swiss Alps but not in the French Alps. The remaining five populations were morphologically intermediate but more similar to the cosmopolitanD. glomerata subsp.glomerata. On the basis of allozyme variation, all populations were more related to subsp.reichenbachii than to the cosmopolitan subsp.glomerata, suggesting that gene flow has occurred between the two tetraploid subspecies. The possibility that selection has acted differently on morphological and allozyme characters is discussed.     

Lack of evidence on hybrid swarm in the sympatric population of Pinus mugo and P. sylvestris

Natural hybridisation between the taxa from Pinus mugo complex and P. sylvestris was postulated in several sympatric populations of the species in Europe. However, due to the absence of precise methods for identification of hybrid seeds and hybrid trees, the frequency of hybridisation and its influence on the genetic structure of relict P. mugo populations has not been clarified so far. In the present study, the species-specific chloroplast DNA (cpDNA) marker and isozymes were applied to test the hybridisation hypothesis in a postulated hybrid swarm population of the species from “Bór na Czerwonem” reserve at the northern foothills of Tatra Mts., Poland. The cpDNA marker was used to identify hybrids within two groups of polycormic (multi-stemmed) and monocormic (single-stemmed) pines from this population. Allelic frequencies at isozyme loci from both groups were compared to frequencies found in allopatric populations of the species. Additionally, cpDNA haplotypes of seedlings derived from open pollinated seeds were studied to detect the possibility of successful cross-pollination. The mixture of seedlings with P. sylvestris and P. mugo cpDNA haplotypes was derived from one parental tree that indicates hybridisation. However, all the mature polycormic pines had cpDNA haplotypes species specific to P. mugo and the isozyme frequencies were similar to frequencies found in three allopatric populations of P. mugo from Tatra Mts. (mean genetic distance, Dn=.027). The differences were much larger in comparisons with monocormic pines from this area (Dn=.085) and two P. sylvestris samples from distant allopatric populations (Dn=.077). Nearly all monocormic pines had cpDNA species specific to P. sylvestris and isozyme frequencies similar to other populations of this species (mean Dn=.004). Only one P. sylvestris-like monocormic tree had cpDNA of P. mugo and can be considered as a hybrid. The results do not provide evidence supporting the hybrid swarm hypothesis. Rather, the results suggest that mature hybrids have a low frequency within this population and (rare) hybridisation is not reciprocal but unidirectional with P. mugo as pollen donor.     

Evolutionary implications of allozyme and RAPD variation in diploid populations of dioecious buffalograss Buchloë dactyloides

Buffalograss, Buchloë dactyloides, is widely distributed throughout the Great Plains of North America, where it is an important species for rangeland forage and soil conservation. The species consists of two widespread polyploid races, with narrowly endemic diploid populations known from two regions: central Mexico and Gulf Coast Texas. We describe and compare the patterns of allozyme and RAPD variation in the two diploid races, using a set of 48 individuals from Texas and Mexico (four population samples of 12 individuals each). Twelve of 22 allozyme loci were polymorphic, exhibiting 35 alleles, while seven 10-mer RAPD primers revealed 98 polymorphic bands. Strong regional differences were detected in the extent of allozyme polymorphism: Mexican populations exhibited more internal gene diversity (H_e= 0.20, 0.19) than did the Texan populations (H_e= 0.08, 0.06), although the number of RAPD bands in Texas (n= 62) was only marginally smaller than in Mexico (n= 68). F-statistics for the allozyme data, averaged over loci, revealed strong regional differentiation (mean F_RT=+ 0.30), as well as some differentiation among populations within regions (mean F_PR=+ 0.09). In order to describe and compare the partitioning of genetic variation for multiple allozyme and RAPD loci, we performed an Analysis of Molecular Variance (AMOVA). AMOVA for both allozyme and RAPD data revealed similar qualitative patterns: large regional differences and smaller (but significant) population differences within regions. RAPDs revealed greater variation among regions (58.4% of total variance) than allozymes (45.2%), but less variation among individuals within populations (31.9% for RAPDs vs. 45.2% for allozymes); the proportion of genetic variance among populations within regions was similar (9.7% for RAPDs vs. 9.6% for allozymes). Despite this large-scale concordance of allozyme and RAPD variation patterns, multiple correlation Mantel techniques revealed that the correlations were low on an individual by individual basis. Our findings of strong regional differences among the diploid races will facilitate further study of polyploid evolution in buffalograss.     

Allozyme Diversity and Population Genetic Structure of Pinus densata Master in Northwestern Yunnan, China

We investigated the levels and patterns of genetic diversity of Pinus densata Master in Yunnan. Horizontal starch-gel electrophoresis was performed on macrogametophytes collected from nine populations in northwestern Yunnan, China. Compared with other gymnosperm species, P. densata has higher mean values for all measures of genetic diversity. Allozyme polymorphism (0.99 criterion) was 97.0% and 71.4% at the species and population levels, respectively. The average number of alleles per locus was 3.1 and 2.0 at the species and population levels. Mean expected heterozygosity was substantially higher in P. densata than average values investigated for other gymnosperms both at the population (H _ep = 0.174±0.031) and at the species (H _es = 0.190) levels. Of the total genetic variation, less than 12% was partitioned among populations (G _ST = 0.112). Our allozyme survey supports the suggestion that the observed higher diversity in P. densata may be attributed partly to its hybrid origin between two genetically distinct species, P. yunnanensis and P. tabulaeformis. In addition, we suggest that introgression would give rise to the increase in genetic diversity occurring in P. densata.     

Selfing versus outcrossing in the androdioecious clam shrimp,Eulimnadia texana (Crustacea,Conchostraca)

The clam shrimpEulimnadia texana is an androdioecious crustacean in which hermaphrodites may self fertilize or outcross with males but cannot outcross with other hermaphrodites. Outcrossing is maintained within most populations of this species despite the high genetic cost of sex, suggesting that compensating factors provide an advantage to outcrossing. We hypothesized that one such benefit would be the production of larger clutch sizes resulting from outcrossed matings. To test this prediction, we recorded the body sizes and clutch sizes of hermaphrodites which mated via selfing or bia outcrossing. Clutch sizes showed significant, almost exponential, increases as body size increased in both selfing and outcrossing hernmaphrodites. The rate of this increase was the same for both groups, and there was no significant difference in clutch size when body size was controlled for between the two fertilization types.     

Identification of hybrids betweenQuercus petraea andQ. robur (Fagaceae): Results obtained with RAPD markers confirm allozyme studies based on theGot-2 locus

RAPDs were employed as genetic markers to detect interspecific hybridization between the closely related oak speciesQuercus robur andQ. petraea. Fourteen primers were used in order to check the genetic status (pure or hybrid) of individuals classified morphologically. Among the 147 PCR fragments obtained 11 appear to be species-specific. In the phenotypically intermediate individuals different combinations of these species-specific bands were obtained. The patterns in these putative hybrids were not additive, which may be either the result of repeated backcrossing and introgression between the two species or of heterozygosity within the parental species. The results of the RAPD study are consistent with morphological analyses and allozyme data obtained for theGot-2 locus. Thus the RAPD markers used in this study may provide a powerful genetic tool for the identification of hybrids and the discrimination between the two pure species.     

Analysis of F ST outliers at allozyme loci in Pacific salmon: implications for natural selection

Natural selection has been invoked to explain the observed geographic distribution of allozyme allele frequencies for a number of teleost species. The effects of selection on allozyme loci in three species of Pacific salmon were tested. A simulation-based approach to estimate the null distribution of population differentiation (F _ST) and test for F _ST outliers was used. This approach showed that a majority of allozyme loci conform to neutral expectations predicted by the simulation model, with relatively few F _ST outliers found. No consistent F _ST outlier loci were found across species. Analysis of population sub-groups based on geography and genetic identity reduced the number of outlier loci for some species, indicating that large geographic groups may include genetically divergent populations and/or that there is geographic heterogeneity in selection pressure upon allozyme loci. Two outlier allozyme loci found in this analysis, lactate dehydrogenase-B and malic enzyme, have been shown to be influenced by selection in other teleost species. This approach is also useful in identifying allozyme loci (or other genetic markers) that meet assumptions for population genetic study.