The annual killifish genus Austrolebias includes approximately 38 species distributed throughout the Paraná-Plata basin and Patos-Merín system. Within the Austrolebias adloffi species complex, the Uruguayan populations of Austrolebias charrua were considered as an intergradation between A. adloffi and Austrolebias viarius populations. Austrolebias charrua presents an intermediate phenotype between both taxa and high levels of morphological and chromatic variability. In the present study, we incorporate different methodological approaches (molecular, morphology, and gamete ultrastructure) to elucidate the pattern of differentiation among the parapatric taxa (A. charrua, Austrolebias reicherti, A. viarius) distributed in a Biosphere Reserve Site. Analyses of cytochrome b sequences show high values of DNA polymorphism, in particular for A. charrua. This is in accordance with both morphological and gametic variation. Using a statistical parsimony network based on these sequences and analysis of morphological data, past fragmentation and range expansion involving perhaps secondary contact between A. charrua and A. reicherti could be proposed. Coloration pattern and morphometric analyses showed an unexpected higher similarity between the most distantly-related taxa, A. viarius and A. charrua. This could be the result of retention of ancestral polymorphisms, especially in A. charrua populations from ponds of higher elevation, or to directional selection acting in similar ecological environments. Because these annual killifish species are considered endangered, our work reinforces the high priority need to include them in a conservation programme.

Annual fish are freshwater teleosts found in South America and Africa that are exposed to an extremely variable environment. They develop and reproduce in seasonal ponds that dry during the summer eliminating the entire adult population. Remarkably, desiccation-resistant embryos survive in these dry ponds that hatch during the next rainy season when the ponds are recreated. Among vertebrates, they represent one of the most remarkable extremophiles. They share several features with other fish models; however, they exhibit unique traits related to their peculiar life cycle. Epiboly is temporally and spatially uncoupled from organogenesis, and the embryos can undergo reversible developmental arrests (diapauses). These attributes make them a useful model to study diverse topics in developmental biology using a comparative and evolutionary approach. In this article, different aspects related to annual fish biology, taxonomy and phylogenetic considerations, reproductive strategy, and developmental characteristics with special focus on arrests, are summarized. The current challenge is to document and determine the factors that generate such high diversity and unique adaptations of annual fish. To understand this complexity, interdisciplinary approaches are being employed taking into consideration evolutionary biology, ethology, reproductive strategies, regulation of developmental mechanisms, and senescence.

These phenotypes are alternative diapause I stages and may have ecological relevance because both were found in embryos in natural ponds. Developmental Dynamics 246:848-857, 2017. © 2017 Wiley Periodicals, Inc.

Diapause is a reversible developmental arrest faced by many organisms in harsh environments. Annual killifish present this mechanism in three possible stages of development. Killifish are freshwater teleosts from Africa and America that live in ephemeral ponds, which dry up in the dry season. The juvenile and adult populations die, and the embryos remain buried in the bottom mud until the next rainy season. Thus, species survival is entirely embryo-dependent, and they are perhaps the most remarkable extremophile organisms among vertebrates. The aim of the present study was to gather information about embryonic diapauses with the use of a “shotgun” proteomics approach in diapause III and prehatching Austrolebias charrua embryos. Our results provide insight into the molecular mechanisms of diapause III. Data are available via ProteomeXchange with identifier PXD025196. We detected a diapause-dependent change in a large group of proteins involved in different functions, such as metabolic pathways and stress tolerance, as well as proteins related to DNA repair and epigenetic modifications. Furthermore, we observed a diapause-associated switch in cytoskeletal proteins. This first glance into global protein expression differences between prehatching and diapause III could provide clues regarding the induction/maintenance of this developmental arrest in A. charrua embryos. There appears to be no single mechanism underlying diapause and the present data expand our knowledge of the molecular basis of diapause regulation. This information will be useful for future comparative approaches among different diapauses in annual killifish and/or other organisms that experience developmental arrest.