Ancient DNA from the extinct New Zealand grayling (Prototroctes oxyrhynchus) reveals evidence for Miocene marine dispersal

O_LIThe evolutionary history of Southern Hemisphere graylings (Retropinnidae) in Aotearoa New Zealand--including the number of colonisation events, the directionality and timing of dispersal, and their relationship to the Australian grayling--is poorly understood. The New Zealand grayling (Prototroctes oxyrhynchus) is the only known freshwater fish species to have gone extinct since human arrival in New Zealand. Despite its historical abundance, only 23 formalin-fixed specimens (both wet and dried) exist in museum collections globally, which were previously non-amenable to palaeogenetic analysis. C_LIO_LIHere, we used high-throughput DNA sequencing techniques, specifically designed for formalin-fixed specimens, to generate mitochondrial genomes of P. oxyrhynchus, and analysed these within a temporal phylogenetic framework of retropinnid and osmerid taxa. C_LIO_LIWe recovered strong evidence for a sister relationship between the New Zealand and Australian grayling (P. mareana), with the two having a common ancestor around 13.8 Mya (95% HPD: 6.1-23.2 Mya), after the height of Oligocene marine inundation in New Zealand. C_LIO_LIOur temporal phylogenetic analysis suggests a single marine dispersal event between New Zealand and Australia, though the direction of dispersal is equivocal, followed by divergence into genetically and morphologically distinguishable species through isolation by distance. C_LIO_LIThis study provides further insights into the possible drivers of the extinction of the New Zealand grayling, and highlights how advancements in palaeogenetic techniques can be used to test evolutionary hypotheses in extinct (and living) fish, which have been comparatively neglected in the field of ancient DNA. C_LI