Gill evolution in Neotropical electric fishes: Comparative phylogenetic evidence for hypoxia-driven adaptation

Tropical freshwater fishes are often exposed to hypoxia, which limits aerobic metabolism and drives the evolution of diverse physiological, behavioral, and morphological adaptations. In species from chronically hypoxic habitats, gill morphology is frequently modified through traits that increase branchial surface area. However, the adaptive basis of these traits and the evolutionary origins of hypoxia tolerance remain poorly understood, partly because few studies have examined species-rich clades in a phylogenetic context. The Neotropical electric knifefish family Hypopomidae, which inhabits environments with a broad range of dissolved oxygen (DO) conditions, represents an ideal model for examining respiratory adaptations to DO variability. Using phylogenetic comparative methods, we examined correlations between environmental DO and gill morphology in 27 species, focusing on three independently varying traits associated with branchial surface area: mean filament length (MFL), mean hemibranch length (MHL), and total filament number (TFN). Species specialized to hypoxic habitats exhibited significantly greater MFL (but not MHL or TFN) after phylogenetic corrections, supporting an adaptive basis for increased gill filament length. In contrast, species specialized to normoxic habitats had lower MFL, likely reflecting trade-offs involving the energetic costs of gill development or functional constraints from adjacent morphological structures. Many hypopomid species are eurytopic, with demes inhabiting both normoxic and hypoxic habitats. Like species specialized to hypoxic systems, hypoxia-tolerant demes exhibit elevated MFL, but not MHL or TFN, suggesting a shared pathway for hypoxia tolerance across different evolutionary scales. Ancestral state reconstructions indicate that ancestors with lower MFL occupied normoxic environments, whereas those with elevated MFL likely evolved in hypoxic systems. A marked transition to greater MFL, accompanied by a simultaneous shift in ancestral habitat, occurs at the base of the Brachyhypopomus beebei group. The emergence of hypoxia tolerance and the onset of diversification in the B. beebei group occurred approximately 3.9 Ma, coincident with a Pliocene expansion of seasonally hypoxic Amazon whitewater floodplains—likely driven by increased Andean sedimentation during the Amazon’s transition to its modern transcontinental course. Our findings highlight how environmental changes shaped by ancient geological events can drive ecomorphological innovation, potentially accelerating species diversification.

Methods
Following Chapman et al. (2008), three independently varying morphological traits correlated with total lamellar surface area were measured: (1) Mean filament length (MFL)—the average length of gill filaments across the eight left hemibranchs; (2) Mean hemibranch length (MHL)—the average distance between the bases of the first and last gill filaments, calculated across all eight left hemibranchs; (3) Total filament number (TFN)—the total count of primary gill filaments on the eight left hemibranchs multiplied by two. Filament counts were taken at the tapering end (every second filament) and central portion (every fifth filament), with the remainder interpolated (Fig. 1).

All specimens were fixed in 10% formalin and preserved in 70% ethanol. Only adult specimens, defined as those exceeding the minimum size for gonadal expression (Crampton et al. 2016) were measured. Body mass was recorded to ±0.01g using a digital scale after blotting excess ethanol. The branchial arches were dissected, and the left and medial hemibranchs from each arch were mounted on slides and imaged with an SDCM-3 digital camera attached to a Meiji-RZ stereo microscope fitted with an ocular micrometer. Measurements were obtained using Motic 2.0 software (www.motic.com).

References:

Chapman, C.A., Albert, J.S. & Galis, F. (2008) Developmental plasticity, genetic differentiation,and hypoxia-induced trade-offs in an African cichlid fish. The Open Evolution Journal, 2, 75-88. DOI: 10.2174/1874404400802010075.

Crampton, W.G.R., de Santana, C.D., Waddell, J.C. & Lovejoy, N.R. (2016) A taxonomic revision of the Neotropical electric fish genus Brachyhypopomus (Ostariophysi: Gymnotiformes: Hypopomidae), with descriptions of 15 new species. Neotropical Ichthyology, 14, 639-790. DOI: 10.1590/1982-0224-20150146.