Inter-island divergence within Drosophila mauritiana, a species of the D. simulans complex: Past history and/or speciation in progress?

Speciation with gene flow may be more common than generally thought, which makes detailed understanding of the extent and pattern of genetic divergence between geographically isolated populations useful. Species of the Drosophila simulans complex provide a good model for speciation and evolutionary studies, and hence understanding their population genetic structure will increase our understanding of the context in which speciation has occurred. Here, we describe genetic diversity and genetic differentiation of two distant populations of D. mauritiana (Mauritius and Rodrigues Islands) at mitochondrial and nuclear loci. We surveyed the two populations for their mitochondrial haplotypes, eight nuclear genes and 18 microsatellite loci. A new mitochondrial type is fixed in the Rodrigues population of D. mauritiana. The two populations are highly differentiated, their divergence appears relatively ancient (100,000 years) compared to the origin of the species, around 0.25MYA, and they exhibit very limited gene flow. However, they have similar levels of divergence from their sibling, D. simulans. Both nuclear genes and microsatellites revealed contrasting demographic histories between the two populations, expansion for the Mauritius population and stable population size for the Rodrigues Island population. The discovery of pronounced geographic structure within D. mauritiana combined to genetic structuring and low gene flow between the two island populations illuminates the evolutionary history of the species and clearly merits further attention in the broad context of speciation.

First evidence for diploidy and genetic recombination in free-living amoebae of the genus naegleria on the basis of electrophoretic variation.

Electrophoretic variation for 15 enzyme-coding genes was studied in various Naegleria (Rhizopoda, Vahlkampfiidae) species. The occurrence of complex banding patterns provided the first evidence of a diploid structure of the genome of these amoebae. The putative loci identified were found not to be linked and the genotypic distribution suggested chromosomal recombination for one species (Naegleria lovaniensis).

Cloning and characterization of KM190, a specific satellite DNA family of Drosophila kitumensis and D. microlabis.

The nucleotide sequences of nine clones, pKA191/1-4 from Drosophila kitumensis and pMR190/1-5 from D. microlabis, were determined. They represent a tandemly arranged and highly repetitive satellite DNA family, KM190, which is specific for the two species.

Evolutionary novelties in islands: Drosophila santomea, a new melanogaster sister species from São Tomé.

The finding of new melanogaster sister species may help us in understanding more about how the emergence of genetic novelties, particularly in insular habitats, can result in speciation. Here we report on the discovery of Drosophila santomea, which is the first melanogaster sibling found off West-equatorial Africa, on São Tomé, one of the Gulf of Guinea islands. Although the eight other melanogaster sister species are remarkably conservative in their morphology except for their terminalia, the new find has a morphological trait distinguishing it from all of these: a pure yellow body coloration of both sexes without the normal black abdominal banding. Evidence from the terminalia, polytene and mitotic chromosomes, period gene and allozymes are provided indicating that it is nonetheless the nearest relative of Drosophila yakuba with which it coexists on the island. The new find is a clear-cut taxon as shown by the production of sterile male hybrids, eventually with developmental defects, in both directions of cross with yakuba and by the existence of an altitudinal divide accompanied by a hybrid zone at mid-elevation on the island. Molecular and karyotypic data further support this conclusion. In contrast to the significant divergence of their nuclear DNAs, an intriguing similarity in their cytochrome b sequences was observed indicating a recent coalescence common to santomea, yakuba and also teissieri cytoplasms. These were shown to harbour the same Wolbachia endosymbiotic bacteria which could possibly be responsible for mitochondrial DNA hitchhiking across the species barrier.

Genetic structure of natural populations of the free-living amoeba, Naegleria lovaniensis. Evidence for sexual reproduction.

The genetic structure of two populations of Naegleria lovaniensis, comprising 71 isolates collected from the same local geographical area was investigated by isoenzyme analysis. Allelic variation at seven polymorphic enzymatic loci allowed identification of 45 distinctive genotype associations. Analysis of single locus variation reveals that most of them are close to Hardy-Weinberg equilibrium, which indicates segregation and free recombination between alleles. The recovery of a relatively high number of distinct genotypic associations (most of them being unique), and the absence of linkage disequilibrium between genotypes at the different loci also support the existence of recombination. Although we have no idea about the process involved, the results clearly indicate that genetic exchanges occur, at least occasionally, in natural populations of N. lovaniensis.

Metabolic evolution in alpha-amylases from Drosophila virilis and D. repleta, two species with different ecological niches.

alpha-Amylases from Drosophila virilis and D. repleta were partially purified by ion exchange chromatography. The two amylases share common characteristics for pH and cations effects, although with slight differences. D. virilis has optimal activity at pH 6.6 and D. repleta at pH 7.2. Calcium, sodium, and potassium cations activate amylolytic activity in both species but Ba2+ has an activation effect in D. repleta only. In contrast, there are major differences in thermal offbility and kinetics among amylases of the two species. D. virilis amylase is much more stable at high temperature and the optimal temperatures are very different between the two species, respectively, 45 degrees C and 30 degrees C for D. virilis and D. repleta. alpha-Amylase activity using different substrates is greater on starch than on glycogen in both species and still higher on amylose for D. virilis, the nonfungus feeder species. alpha-Amylase of D. repleta, the mycophagous species, has a better affinity to amylopectin and glycogen. Such differences in substrate specificity suggest adaptation to different resources in these species living in different habitats. Metabolic evolution seems to have occurred through a "tradeoff" between kinetic effectiveness and the nature of substrate, with a higher Vmax on amylose for D. virilis and a lower K(m) on glycogen for D. repleta.

Large genetic heterogeneity within amoebas of the species Naegleria gruberi and evolutionary affinities to the other species of the genus.

The allozyme survey was extended to 7 strains of Naegleria gruberi and N. jadini in order to further characterize the genetic structure of these free-living amoebas. As formerly known for several characters the electrophoretic evidence reveals considerable heterogeneity at the genetic level among N. gruberi strains. Moreover, 2 distinct gene pools, that might likely represent natural taxa, are clearly identified. The single strain of N. jadini appears evolutionarily related to 1 group of N. gruberi which is also related to N. a. australiensis.

Biochemical identification and phylogenetic relationships in free-living amoebas of the genus Naegleria.

Using isoelectric focusing, the zymograms of 23 pathogenic and nonpathogenic Naegleria strains were studied for the activity of 16 enzymes. Certain enzymes (lactate dehydrogenase, L-threonine dehydrogenase, superoxide dismutase, acid phosphatase, malic enzyme, and leucine aminopeptidase) proved particularly useful from a practical point of view as they allow easy and reliable identification of pathogenic N. fowleri and N. australiensis as well as nonpathogenic N. lovaniensis strains. Genetic interpretation of these zymograms gave estimates of genetic distances that largely confirmed the taxonomic position of the Naegleria species. In addition, the genetic data suggest that there are two main phylogenetic groups in the genus Naegleria.

Isozyme variability and differentiation between Rhodnius prolixus, R.robustus and R.pictipes, vectors of Chagas disease in Venezuela.

Enzyme polymorphism in triatomine bugs of the genus Rhodnius (Hemiptera: Reduviidae), vectors of Chagas disease, is analysed using both starch and polyacrylamide gel electrophoresis. Out of forty-five enzymes assayed, the electromorphs of seventeen of them: AO, CA, DIA, ES, ES-A, FH, GPD, G6PD, GPI, MDH, ME, 6PGD, PGM, ACON, ACPH, LAP and SOD, involving twenty-two putative structural loci, were scorable. These gene-enzyme systems were therefore selected for routine characterization of R.prolixus Stal adults from different strains. The first thirteen enzymes, involving sixteen structural loci, were also analysed in first instar nymphs of the three species, R.prolixus, R.robustus Larrousse and R.pictipes Stal. Allelic frequencies were calculated for three R.prolixus strains: three to five loci appeared to be polymorphic. The proportion of polymorphic loci (22%) and the average heterozygosity (0.06) indicated low genetic variability, with significant differences between the strains at individual loci. Rhodnius prolixus and R.robustus were found to have identical isozymic patterns. R.pictipes was genetically well differentiated, with twelve diagnostic loci.

Molecular characterization and evolution of the amylase multigene family of Drosophila ananassae.

Drosophila ananassae is known to produce numerous alpha-amylase variants. We have cloned seven different Amy genes in an African strain homozygous for the AMY1,2,3,4 electrophoretic pattern. These genes are organized as two main clusters: the first one contains three intronless copies on the 2L chromosome arm, two of which are tandemly arranged. The other cluster, on the 3L arm, contains two intron-bearing copies. The amylase variants AMY1 and AMY2 have been assigned to the intronless cluster, and AMY3 and AMY4 to the second one. The divergence of coding sequences between clusters is moderate (6.1% in amino acids), but the flanking regions are very different, which could explain their differential regulation. Within each cluster, coding and noncoding regions are conserved. Two very divergent genes were also cloned, both on chromosome 3L, but very distant from each other and from the other genes. One is the Amyrel homologous (41% divergent), the second one, Amyc1 (21.6% divergent) is unknown outside the D. ananassae subgroup. These two genes have unknown functions.

Distribution and evolution of introns in Drosophila amylase genes.

While the two amylase genes of Drosophila melanogaster are intronless, the three genes of D. pseudoobscura harbor a short intron. This raises the question of the common structure of the Amy gene in Drosophila species. We have investigated the presence or absence of an intron in the amylase genes of 150 species of Drosophilids. Using polymerase chain reaction (PCR), we have amplified a region that surrounds the intron site reported in D. pseudoobscura and a few other species. The results revealed that most species contain an intron, with a variable size ranging from 50 to 750 bp, although the very majoritary size was around 60-80 bp. Several species belonging to different lineages were found to lack an intron. This loss of intervening sequence was likely due to evolutionarily independent and rather frequent events. Some other species had both types of genes: In the obscura group, and to a lesser extent in the ananassae subgroup, intronless copies had much diverged from intron-containing genes. Base composition of short introns was found to be variable and correlated with that of the surrounding exons, whereas long introns were all A-T rich. We have extended our study to non-Drosophilid insects. In species from other orders of Holometaboles, Lepidoptera and Hymenoptera, an intron was found at an identical position in the Amy gene, suggesting that the intron was ancestral.

Variation in sex-, stage- and tissue-specific expression of the amylase genes in Drosophila ananassae.

Expression of the amylase multigene family of Drosophila ananassae was investigated in third-instar larvae and adults. A developmental differentiation was found between the Amy1-2 and Amy3-4 gene clusters, the former being preferentially expressed in larvae, the latter in adults. During adult life, we observed a decrease in Amy1-2 expression in males of certain strans. We have raised some arguments for the existence of trans-active regulators, acting as repressors of Amy1-2 in adults. The putative repressors might exhibit a geographical polymorphism, with a fixed active form in Pacific regions and a polymorphic pattern in Africa, thus increasing the diversity observed in adult amylase phenotypes. A clear differentiation between the two gene clusters was also found in tissue-specific activity along the third-instar larval midgut. In the anterior midgut, only Amy1-2 is active, while both gene groups are expressed in the posterior midgut, with an additional subzonation within it.

[Genetic interpretation of flagellate zymograms of the genera Trypanosoma and Leishmania]. / Interprétation génétique des zymogrammes de Flagellés des genres Trypanosoma et Leishmania.

A survey of enzyme variability between several strains of Trypanosoma cruzi allows to think that these organisms are diploids. According to the data sexuality, may be only occasional, has been hypothesized, at least in the recent past. The genetic distances between three strains are calculated for twelve loci.

Low allozyme and mtDNA variability in the island endemic species Drosophila sechellia (D. melanogaster complex).

Genetic variability of D. sechellia is investigated at both mitochondrial and nuclear levels. The results reveal the existence of a single main type of mtDNA with very few variants and a very low enzyme polymorphism. This situation is consistent with the small population size of this specialized species.

Amylase gene duplication: an ancestral trait in the Drosophila melanogaster species subgroup.

Electrophoretic polymorphism of amylases was studied in 45 geographic populations of the two cosmopolitan sibling species, D. melanogaster and D. simulans, and in one to three populations or strains of six other species in the D. melanogaster subgroup. Two species, D. erecta and D. orena, for which only a few strains were available were monomorphic. In the other species 2 or 3 amylase variants were identified while in D. melanogaster, 12 electrophoretic variants were characterized. Altogether 17 different amylase isozymes have been observed. The contrast in the level of polymorphism between D. melanogaster and the other species cannot be explained simply by the occurrence of a duplication in the former species. Genetic analysis demonstrated the existence of a duplication in at least 4 other species, namely D. simulans, D. mauritiana, D. yakuba, and D. teissieri. It is therefore suggested that the duplication occurred in a common ancestor and the phylogenetic implications of these observations are discussed.

Morphological and molecular differentiation of invasive freshwater species of the genus Corbicula (Bivalvia, corbiculidea) suggest the presence of three taxa in French rivers.

Asiatic Clams are common in brackish and fresh water in Asia, and they were introduced into North America in 1924 and have now spread throughout the continent. During the last two decades they have been reported in Europe, but the number of species here is uncertain. Populations of Corbicula from France and the Netherlands were analysed morphologically and genetically to quantify the degree of species and/or population differentiation. The morphological and genetic data, based on allozymes and mitochondrial sequences, were in full agreement. They indicate that there are two distinct species, identified as C. fluminalis and C. fluminea, in the two countries. Analyses of the mitochondrial COI gene revealed an unexpected divergent population of Corbicula in the Rhône. All these individuals were morphologically identified as C. fluminea, but had a COI sequence different from the two previous species. This population may, therefore, be a more ancient population, or a distinct species introduced via a different colonization route.

Geographical polymorphism of amylase in Drosophila ananassae and its relatives.

Strains of Drosophila ananassae from various places in the Tropics were investigated for their electrophoretical amylase pattern. Eleven isoamylases were found in adult flies. African populations were much more polymorphic than those from the Far East, and showed multibanded phenotypes, suggesting a multiplication of the Amy structural gene, with at least four copies per haploid genome in certain populations. Nine other species of the D. ananassae subgroup had weak amylase activity and only one or two variants were found in each species. D. monieri and D. varians are closely related to D. ananassae and showed a single band, similar to the isoamylase 3 of D. ananassae, which suggests that this might be an ancestral allele.

The phylogeny of nine species of the Drosophila obscura group inferred by the banding homologies of chromosomal regions. I. Element B.

The phylogenetic relationships among nine Drosophila species belonging to the obscura group were investigated by establishing the segments displaying banding homologies in their element B (equivalent to the U element of D. subobscura). The phylogenetic ordering of the species was accomplished using overlapping inversions. Two African species, D. kitumensis and D. microlabis, were investigated. These species are homosequential for their element B gene arrangement but differ from that of D. obscura by several rearrangements. Drosophila obscura seems to be most closely related to D. subsilvestris, from which the respective element B gene arrangements differ at least by six inversions. Three species, D. obscura, D. ambigua, and D. tristis, are closely related and form a cluster. Drosophila obscura displays an element B polymorphism for a pericentric inversion for which D. ambigua is fixed for one gene arrangement and D. tristis for the other. Both D. ambigua and D. tristis share a short distal inversion in the small arm of the chromosome, and differ in this respect from D. obscura. Drosophila madeirensis, D. guanche, and D. subobscura all share the same element B gene arrangement, which is acrocentric, but metacentric in all the other species mentioned. It was found that the gene arrangements of the species from the obscura cluster seem to occupy an intermediate position between those of the species of the D. subobscura cluster and those of the African one. The data reported generally are in good agreement with information provided in the literature.

Divergence between Drosophila santomea and allopatric or sympatric populations of D. yakuba using paralogous amylase genes and migration scenarios along the Cameroon volcanic line.

We have used two paralogous genes (Amyrel and Amy) of the amylase multigene family to reconstruct the phylogeny of the nine Drosophila melanogaster subgroup sister species, including D. santomea, the newly discovered endemic from São Tomé island. The evolutionary divergence of these genes is of special interest as it is suspected to result from physiological evolution via gene duplication. This paper describes the relationship between the geographical origin of the various strains and the patterns of mating and phylogeny, focusing on the evolution of D. santomea and its relationship to other species and their niches. The Amyrel and Amy data indicate that, contrary to expectations, the sympatric insular D. yakuba population is less closely related to D. santomea than allopatric mainland ones, suggesting that the extant insular D. yakuba population on São Tomé results from a recent secondary colonization. Data for sympatric and allopatric D. yakuba suggest that D. santomea arose from a mainland D. yakuba parental stock when montane habitats of the Cameroon volcanic line extended to lower altitudes during colder and less humid periods. Despite their different modes of evolution and different functions, the Amyrel and Amy genes provide remarkably consistent topologies and hence reflect the same history, that of the species.

Multiple amylase genes in Drosophila ananassae and related species.

The number and organization of amylase genes in Drosophila ananassae were investigated through classical genetic methods and in situ and filter hybridizations. At least four genes may be active in D. ananassae, organized as two independent pairs of closely linked copies on the 2L and 3L chromosomal arms. Several other species of the D. ananassae subgroup were studied and show the same chromosomal locations, suggesting an ancient duplication event. However, the number of Amy copies seems to be higher in the D. ananassae multigene family, and there is a striking intraspecific molecular differentiation.