Riverscape genetics: linking river environments with adaptive potential in Neotropical freshwater fish

3. Background. Orinoco basin ichthyofauna and environmental characteristics: The Orinoco River basin is an epicenter of freshwater fish diversity, being the second most diverse in the Neotropics18,19 (~1000 spp.). This basin, more precisely the Meta Sub-basin20 (> 658 spp.), is ideal to study the local adaptation of freshwater fish because there are prominent shifts in precipitation and perennial condition21 across the length of each tributary providing an environmental gradient. Specifically, rivers draining the Llanos ecosystem on the Meta River basin (Fig. 1A) receive year-round high levels of rainfall in the piedmont area, while in the lowlands are characterized by a dry and a wet season21,22 (Fig. 1B and C). During the dry season, rivers might become ephemeral, triggering the migration of fishes downstream to enduring rivers22,23. Alternatively, a subset of the ichthyofauna persists in fragmented pools under environmentally stressed conditions of drought23,24, such as hypoxia. These conditions have an impact on the differential survival of organisms - promoting local adaptation. Morphological evidence for adaptation to drought: Several morphological, behavioral and live-history strategies have been associated with adaptation to drought and hypoxic environments in stream organisms25-27. In fishes there are physiological responses to oxygen storage and transportation that promote observable changes in the morphology of gill filaments28. Another morphological adaptation, restricted to Characiformes fishes and relatively easy to assess in nature, is the ability to develop dermal extensions on the lower jaw (Fig. 2) in some South American species29. This feature is thought to improve surface respiration under hypoxic conditions (dissolved oxygen: DO < 1.4 ppm) by increasing oxygen flow to gills directly from the oxygen-rich water surface layer. This condition is observed at different frequencies in populations, depending, not just on DO, but also on the riverine location, indicating that there is agenetic basis underlying this plastic response29,30. Although several characids present morphological responses to hypoxia (> 16 species), to understand how the frequency of these adaptations interacts with the environmental gradient at each tributary, the species focus need to be broadly distributed and abundant in all environments. Because of its broad distribution in tributary rivers from the Andes piedmont to the Llanos, we aim to study the species Gephyrocharax valencia (Eigenmann, 1920) (Characidae; Fig. 1 and 2). Other potential candidates are Odontostilbe pulchra (Gill, 1858) and Aphyocharax alburnus (Günther, 1869) ¿ but their occurrence in the Meta River main channel might indicate strong gene flow among tributaries23.