Sex-chromosome evolution of Palearctic tree frogs in space and time

Sexual reproduction is nearly universal in eukaryotes and genetic determination of sex prevails among animals. The astonishing diversity of sex-determining systems and sex chromosomes is yet bewildering. Some taxonomic groups possess conserved and dimorphic sex chromosomes, involving a functional copy (e.g. mammals' X, birds' Z) and a degenerated copy (mammals' Y, birds' W), implying that sex- chromosomes are expected to decay. In contrast, others like amphibians, reptiles and fishes yet maintained undifferentiated sex chromosomes. Why such different evolutionary trajectories? In this thesis, we empirically test and characterize the main hypotheses proposed to prevent the genetic decay of sex chromosomes, namely occasional X-Y recombination and frequent sex-chromosome transitions, using the Palearctic radiation of Hyla tree frogs as a model system. We take a phylogeographic and phylogenetic approach to relate sex-chromosome recombination, differentiation, and transitions in a spatial and temporal framework. By reconstructing the recent evolutionary history of the widespread European tree frog H. arborea, we showed that sex chromosomes can recombine in males, preventing their differentiation, a situation that potentially evolves rapidly. At the scale of the entire radiation, X-Y recombination combines with frequent transitions to prevent sex-chromosome degeneration in Hyla: we traced several turnovers of sex-determining system within the last 10My. These rapid changes seem less random than usually assumed: we gathered evidences that one chromosome pair is a sex expert, carrying genes with key role in animal sex determination, and which probably specialized through frequent reuse as a sex chromosome in Hyla and other amphibians. Finally, we took advantage of secondary contact zones between closely-related Hyla lineages to evaluate the consequences of sex chromosome homomorphy on the genetics of speciation. In comparison with other systems, the evolution of sex chromosomes in Hyla emphasized the existence of consistent evolutionary patterns within the chaotic diversity of flexibility of cold-blooded vertebrates' sex-determining systems, and provides insights into the evolution of recombination. Beyond sex-chromosome evolution, this work also significantly contributed to speciation, phylogeography and applied conservation research. -- La reproduction sexuée est quasi-universelle chez les eucaryotes et le sexe est le plus souvent déterminé génétiquement au sein du règne animal. L'incroyable diversité des systèmes de reproduction et des chromosomes sexuels est particulièrement étonnante. Certains groupes taxonomiques possèdent des chromosomes sexuels dimorphiques et très conservés, avec une copie entièrement fonctionnelle (ex : le X des mammifères, le Z des oiseaux) et une copie dégénérée (ex : le Y des mammifères, le W des oiseaux), suggérant que les chromosomes sexuels sont voués à se détériorer. Cependant les chromosomes sexuels d'autres groupes tels que les amphibiens, les reptiles et les poissons sont pour la plupart indifférenciés. Comment expliquer des trajectoires évolutives si différentes? Au cours de cette thèse, nous avons étudié empiriquement les processus évolutifs pouvant maintenir les chromosomes sexuels intacts, à savoir la recombinaison X-Y occasionnel ainsi que les substitutions fréquentes de chromosomes sexuels, en utilisant les rainettes Paléarctiques du genre Hyla comme modèle d'étude. Nous avons adopté une approche phylogéographique et phylogénétique pour appréhender les événements de recombinaison, de différenciation et de transitions de chromosomes sexuels dans un contexte spatio-temporel. En retraçant l'histoire évolutive récente de la rainette verte H. arborea, nous avons mis en évidence que les chromosomes sexuels pouvaient recombiner chez les mâles, empêchant ainsi leur différenciation, et que ce processus avait le potentiel d'évoluer très rapidement. A l'échelle plus globale de la radiation, il apparait que les phénomènes de recombinaison X-Y soient également accompagnés de substitutions de chromosomes sexuels, et participent de concert au maintien de chromosomes sexuels intacts dans les populations: le système de détermination du sexe des rainettes a changé plusieurs fois au cours des 10 derniers millions d'années. Ces transitions fréquentes ne semblent pas aléatoires: nous avons identifié une paire de chromosomes qui présente des caractéristiques présageant d'une spécialisation dans le déterminisme du sexe (notamment car elle possède des gènes importants pour cette fonction), et qui a été réutilisée plusieurs fois comme tel chez les rainettes ainsi que d'autres amphibiens. Enfin, nous avons étudié l'hybridation entre différentes espèces dans leurs zones de contact, afin d'évaluer si l'absence de différenciation entre X et Y jouaient un rôle dans les processus génétiques de spéciation. Outre son intérêt pour la compréhension de l'évolution des chromosomes sexuels, ce travail contribue de manière significative à d'autres domaines de recherche tels que la spéciation, la phylogéographie, ainsi que la biologie de la conservation.

Sex-Chromosome Homomorphy in Palearctic Tree Frogs Results from Both Turnovers and X-Y Recombination.

Contrasting with birds and mammals, poikilothermic vertebrates often have homomorphic sex chromosomes, possibly resulting from high rates of sex-chromosome turnovers and/or occasional X-Y recombination. Strong support for the latter mechanism was provided by four species of European tree frogs, which inherited from a common ancestor (∼5 Ma) the same pair of homomorphic sex chromosomes (linkage group 1, LG1), harboring the candidate sex-determining gene Dmrt1. Here, we test sex linkage of LG1 across six additional species of the Eurasian Hyla radiation with divergence times ranging from 6 to 40 Ma. LG1 turns out to be sex linked in six of nine resolved cases. Mapping the patterns of sex linkage to the Hyla phylogeny reveals several transitions in sex-determination systems within the last 10 My, including one switch in heterogamety. Phylogenetic trees of DNA sequences along LG1 are consistent with occasional X-Y recombination in all species where LG1 is sex linked. These patterns argue against one of the main potential causes for turnovers, namely the accumulation of deleterious mutations on nonrecombining chromosomes. Sibship analyses show that LG1 recombination is strongly reduced in males from most species investigated, including some in which it is autosomal. Intrinsically low male recombination might facilitate the evolution of male heterogamety, and the presence of important genes from the sex-determination cascade might predispose LG1 to become a sex chromosome.

Introgressive hybridization of threatened European tree frogs (Hyla arborea) by introduced H. intermedia in Western Switzerland

Hybridization by introduced taxa is a major threat to native species. Characterizing human introductions is thus one of the missions of conservation geneticists. Here we survey a declining population of the regionally endangered European tree frog (Hyla arborea) in the Grangettes natural reserve (Rhone valley, Western Switzerland), where previous evidence indicated human introduction of the Italian taxon H. intermedia. We combined fast-evolving mitochondrial and nuclear markers and an extended sampling to conduct population genetic analyses of the Grangettes and putative source areas. We show that the Grangettes population is a hybrid swarm, with all individuals featuring recent nuclear admixture and mitochondrial DNA of introduced H. intermedia, most likely of proximate south Alpine origin. In contrast, H. arborea and H. intermedia hardly introgress in their natural parapatric ranges, consistent with an advanced reproductive isolation. Thus, potential hybrid incompatibilities may account for the strong decline of this population, despite important conservation efforts. Although their hybrid nature makes them a priori unworthy of any protection, we propose specific measures to recover local H. arborea gene pool and preserve tree frogs in the Grangettes, the last population remaining from this heavily impacted part of the Alps.

Divergence with gene flow and fine-scale phylogeographical structure in the wedge-billed woodcreeper, Glyphorynchus spirurus, a Neotropical rainforest bird.

Determining the relative roles of vicariance and selection in restricting gene flow between populations is of central importance to the evolutionary process of population divergence and speciation. Here we use molecular and morphological data to contrast the effect of isolation (by mountains and geographical distance) with that of ecological factors (altitudinal gradients) in promoting differentiation in the wedge-billed woodcreeper, Glyphorynchus spirurus, a tropical forest bird, in Ecuador. Tarsus length and beak size increased relative to body size with altitude on both sides of the Andes, and were correlated with the amount of moss on tree trunks, suggesting the role of selection in driving adaptive divergence. In contrast, molecular data revealed a considerable degree of admixture along these altitudinal gradients, suggesting that adaptive divergence in morphological traits has occurred in the presence of gene flow. As suggested by mitochondrial DNA sequence data, the Andes act as a barrier to gene flow between ancient subspecific lineages. Genome-wide amplified fragment length polymorphism markers reflected more recent patterns of gene flow and revealed fine-scale patterns of population differentiation that were not detectable with mitochondrial DNA, including the differentiation of isolated coastal populations west of the Andes. Our results support the predominant role of geographical isolation in driving genetic differentiation in G. spirurus, yet suggest the role of selection in driving parallel morphological divergence along ecological gradients.

AMAZON RAINFOREST COSMETICS: CHEMICAL APPROACH FOR QUALITY CONTROL

The market for natural cosmetics featuring ingredients derived from Amazon natural resources is growing worldwide. However, there is neither enough scientific basis nor quality control of these ingredients. This paper is an account of the chemical constituents and their biological activities of fourteen Amazonian species used in cosmetic industry, including açaí (Euterpe oleracea), andiroba (Carapa guianensis), bacuri (Platonia insignis), Brazil nut (Bertholletia excelsa), buriti (Mauritia vinifera or M. flexuosa), cumaru (Dipteryx odorata), cupuaçu (Theobroma grandiflorum), guarana (Paullinia cupana), mulateiro (Calycophyllum spruceanum), murumuru (Astrocaryum murumuru), patawa (Oenocarpus bataua or Jessenia bataua), pracaxi (Pentaclethra macroloba), rosewood (Aniba rosaeodora), and ucuuba (Virola sebifera). Based on the reviewed articles, we selected chemical markers for the quality control purpose and evaluated analytical methods. Even though chromatographic and spectroscopic methods are major analytical techniques in the studies of these species, molecular approaches will also be important as used in food and medicine traceability. Only a little phytochemical study is available about most of the Amazonian species and some species such as açaí and andiroba have many reports on chemical constituents, but studies on biological activities of isolated compounds and sampling with geographical variation are limited.

Genetic analyses of Rhizoctonia solani isolates from Phaseolus vulgaris grown in the Atlantic Rainforest Region of São Paulo, Brazil

Rhizoctonia solani isolates obtained from common beans (Phaseolus vulgaris) grown in the mountainous Atlantic Rainforest (Mata Atlântica) region of São Paulo, Brazil, were analyzed to determine their genetic diversity using internal transcribed spacer (ITS), microsatellite and telomere sequence-based PCR primers. Restriction digestion of the ITS1/5.8S/ITS2 ribosomal regions yielded unique banding patterns specific for AG4 and its subgroups. The ITS restriction digestion (ITS/RFLP), telomere and microsatellite primers identified five to 11 genotypes within the isolates of R. solani. While all isolates were pathogenic on beans, there was no correlation found between genotypic differences and pathogenicity. The different PCR primers revealed a number of isolates that were genetically similar. Some of these genetic groups were supported by more than one of the primers utilized in this study, thus confirming their relationship.

Fungal succession on leaves of Alchornea triplinervia (Spreng.) Muell. Arg. submerged in a stream of an Atlantic Rainforest in the state of São Paulo, Brazil

Leaves of Alchornea triplinervia (Spreng.) Muell. Arg. were submerged in a stream in an Atlantic Rainforest in São Paulo state, Brazil, from July/1988 to June/1989 and from July/1989 to May/1990. Fungi were isolated by the leaf disks washing technique followed by plating on culture media and also by using baiting techniques (using substrates with chitin, keratin and cellulose), what resulted on 565 fungal registers corresponding to 81 taxa. The most common species found during this study of the fungal succession were Trichoderma viride Pers. ex S.F. Gray and Fusarium oxysporum Schlecht emend. Snyd. & Hans. (23 registers), Penicillium hirsutum Dierckx (21 registers), Fusarium solani (Mart.) Appel & Wollenw. emend. Snyd. & Hans. (17), followed by 14 registers of: Cylindrocladium scoparium Morgan, Triscelophorus monosporus Ingold and Polychytrium aggregatum Ajello. Although the monthly obtained mycota had been composed by species of different taxonomic groups, the fungal succession was defined by the initial presence of typical terrestrial leaf inhabiting fungi (mostly Deuteromycotina), followed by species of Mastigomycotina and Zygomycotina. Combining culture methods and baiting techniques, it was possible to verify the presence of terrestrial fungi on the decomposition of submerged leaves and the importance of zoosporic fungi in the fungal succession. This is the first paper about the fungal succession on the decomposition of leaves submerged in a lotic ecosystem in Brazil.

Interaction between the mineral content and the occurrence number of aquatic fungi in leaves submerged in a stream in the Atlantic rainforest, São Paulo, Brazil

Leaves of Ficus microcarpa L. f., Quercus robur L., and Alchornea triplinervia (Spreng.) Muell. Arg., submerged in a stream of the Atlantic rainforest in the "Reserva Biológica do Alto da Serra de Paranapiacaba", State of São Paulo, Brazil, were collected monthly, from April to November 1990, in order to determine the number of fungal occurrences (zoosporic fungi and aquatic Hyphomycetes), and the content of total N (%), total P (%), K+ (%), Ca+2 (%), Mg+2 (%), S+3 (%), Fe+3 (ppm), Cu+3 (ppm), Mn+2 (ppm), Zn+2 (ppm), Bo (ppm), Na+2 (ppm) and Al+3 (ppm). According to the tests of Kruskal-Wallis, Mann-Whitney and Wilcoxon, the means of the mineral content of the three types of leaves were significantly different, except for Mg+2 (%), Mn+2 (ppm), Zn+2 (ppm) and Na+2 (ppm). On comparing the mineral content with the number of fungal occurrence, an independence test showed a positive correlation between the presence of zoosporic fungi on the leaves of A. triplinervia and the total nitrogen, phosphorus and S+3 content, whereas the aquatic Hyphomycetes depended on the amount of Ca+2 available. Regarding leaves of F. microcarpa, the occurrence of zoosporic fungi was linked to the S+3 level, and the presence of aquatic Hyphomycetes, to the content of K+, Ca+2, S+3 and Bo. On Q. robur leaves, zoosporic fungi showed a positive correlation with the Ca+2 content, but a negative one with Fe+3 and Al+3 levels, while the occurrence of aquatic Hyphomycetes was influenced by the content of Ca+2, Mg+2, Fe+3, Al+3, Mn+2, Zn+2 and Na+2. The correlation between the occurrence number of aquatic Hyphomycetes and a high mineral content indicates that their nutritional requirements may be more complex than those of zoosporic fungi. Further studies are still required to understand the implications of this tendency on the diversity of aquatic native mycota.

Spatial distribution and preferential substrate of Neoregelia johannis (Carriére) L.B. Smith (Bromeliaceae) in a disturbed area of Atlantic Rainforest at Ilha Grande, RJ, Brazil

In bromeliad populations, amount of light and available substrates influence individuals spatial organization. In Atlantic Rainforest of Ilha Grande, the heliophylous bromeliad Neoregelia johannis is a large and abundant species. In this forest, it would be expected that N. johannis would occupy stable substrates, as large trunks, large branches, rock boulders or ground, with high sunlight, enough for the bromeliad survivor. In the present work, we analyzed the distribution and most used substrates of N. johannis in secondary forest. We analyzed the frequency of reproductive modes (sexual and vegetative) used by the bromeliad shoots, registering if the shoots were originated from seeds or by vegetative reproduction. The results indicated an aggregated distribution pattern (Ip = 0.052). The preferred substrate was boulders (91%), whereas tree trunks (6%) and the ground (3%) were rarely used. Small and fragile substrates are unstable to support large adults of this species, which may explain the predominant pattern of establishment over boulders within the secondary forest, as the presence of this substrate also results in more opened canopy cover. Approximately 50% of young individuals entered the population by vegetative reproduction. We conclude that the preferential habit and the aggregated distribution of N. johannis are due to the conjunction of preferred substrate with higher amount of light resulting from breaks in tree canopy over areas with rock blocks, and high frequency of recruitment by vegetative reproduction.

Community structure of vascular plants in treefall gaps and fire-disturbed habitats in the Atlantic rainforest, southern Bahia, Brazil

The effects of disturbances on plant community structure in tropical forests have been widely investigated. However, a majority of these studies examined only woody species, principally trees, whereas the effects of disturbances on the whole assemblage of vascular plants remain largely unexplored. At the present study, all vascular plants < 5m tall were surveyed in four habitats: natural treefall gaps, burned forest, and their adjacent understorey. The burned area differed from the other habitats in terms of species composition. However, species richness and plant density did not differ between burned area and the adjacent understorey, which is in accordance to the succession model that predict a rapid recovery of species richness, but with a different species composition in areas under moderate disturbance. The treefall gaps and the two areas of understorey did not differ among themselves in terms of the number of individuals, number of species, nor in species composition. The absence of differences between the vegetation in treefall gaps and in understorey areas seems to be in agreement with the current idea that the species present in treefall gaps are directly related to the vegetation composition before gap formation. Only minimal differences were observed between the analyses that considered only tree species and those that considered all growth habits. This suggests that the same processes acting on tree species (the best studied group of plants in tropical forests) are also acting on the whole assemblage of vascular plants in these communities.

A new species of Lemmermanniella (Cyanobacteria) from the Atlantic Rainforest, Brazil

The Brazilian Atlantic Rainforest is a highly heterogeneous ecosystem comprising large numbers of tropical and subtropical habitats favorable to the development of cyanobacteria. Studies on cyanobacteria in this ecosystem are still rare, however, especially those involving unicellular and colonial types. The high biodiversity and endemism of this biome has been extremely impacted and fragmented, and less than 10% of its original vegetation cover remains today. We describe here a new species of a colonial cyanobacteria, Lemmermanniella terrestris, found on dry soils in a subtropical region of the Atlantic Rainforest in the municipality of Cananéia in southern São Paulo State, Brazil. This new taxon demonstrated all of the diacritical features of the genus Lemmermanniella but, unlike the other species of the genus, it was growing on the soil surface and not in an aquatic environment. A set of morphological features, including colonies composed of subcolonies, and cell dimensions, shapes and contents distinguish it from other species of the genus. Considering that species of Lemmermanniella are found in very distinct habitats (such as thermal and brackish waters) and that they maintain the same life cycle described for the genus in all of those environments, the morphological structures of the colonies can be used as reliable markers for identifying the genus, and its species differ primarily in relation to the habitats they occupy.

Use of conventional PCR and smear microscopy to diagnose pulmonary tuberculosis in the Amazonian rainforest area

The diagnostic usefulness of Ziehl-Neelsen (ZN)-stained sputum smears combined with conventional polymerase chain reaction (ZN/PCR) to amplify IS6110 region DNA extracted from ZN slides was evaluated. The objective was to verify if this association could improve tuberculosis (TB) diagnosis in patients at remote sites. The study was carried out in 89 patients with culture-confirmed pulmonary TB as defined by the Brazilian Manual for TB Treatment. The participants were recruited in a reference unit for TB treatment in Rondônia, a state in the Amazonian area in northern Brazil. ZN, PCR, and culture performed in the sputum samples from these patients were analyzed in different combinations (i.e., ZN plus PCR and ZN plus culture). The prevalence rates of pulmonary TB in these patients were 32.6 and 28.1% considering culture and ZN/PCR, respectively. The sensitivity and specificity of ZN/PCR were 86 and 93%, respectively. ZN/PCR was able to detect more TB cases than ZN alone. This method could offer a new approach for accurate tuberculosis diagnosis, especially in remote regions of the world where culture is not available.

Frogs and toads.

Aprobada por expertos en educación la obra trata temas de la vida real. Texto con frases sencillas y repetición de palabras para motivar al niño a desarrollar su interés por la lectura y la curiosidad por el mundo en que vive.Tiene glosario alfabético.

Frogs.

Escrito para responder las preguntas que los niños pequeños hacen sobre las ranas. Proporciona comparaciones visuales, pasatiempos, chistes, acertijos y rompecabezas. El texto tiene dos tamaños de letra y dos niveles de dificultad para los lectores más expertos.