Abstract
As the only endemic neotropical parrot to have recently lived in the northern hemisphere, the Carolina parakeet (Conuropsis carolinensis) was an iconic North American bird. The last surviving specimen died in the Cincinnati Zoo in 1918 [1]. The cause of its extinction remains contentious: besides excessive mortality associated to habitat destruction and active hunting, their survival could have been negatively affected by its range having become increasingly patchy [2] or by the exposure to poultry pathogens [3, 4]. In addition, the Carolina parakeet showed a predilection for cockleburs, an herbaceous plant that contains a powerful toxin, carboxyatractyloside, or CAT [5], which did not seem to affect them but made the birds notoriously toxic to most predators [3]. To explore the demographic history of this bird, we generated the complete genomic sequence of a preserved specimen held in a private collection in Espinelves (Girona, Spain), as well as of a close extant relative, Aratinga solstitialis. We identified two non-synonymous genetic changes in two highly conserved proteins known to interact with CAT that could underlie a specific dietary adaptation to this toxin. Our genomic analyses did not reveal evidence of a dramatic past demographic decline in the Carolina parakeet; also, its genome did not exhibit the long runs of homozygosity that are signals of recent inbreeding and are typically found in endangered species. As such, our results suggest its extinction was an abrupt process and thus likely solely attributable to human causes.
Highlights
As the only endemic neotropical parrot to have recently lived in the northern hemisphere, the Carolina parakeet (Conuropsis carolinensis) was an iconic North American bird
The Carolina parakeet showed a predilection for cockleburs, an herbaceous plant that contains a powerful toxin, carboxyatractyloside, or CAT [5], which did not seem to affect them but made the birds notoriously toxic to most predators [3]
Our genomic analyses did not reveal evidence of a dramatic past demographic decline in the Carolina parakeet; its genome did not exhibit the long runs of homozygosity that are signals of recent inbreeding and are typically found in endangered species
Summary
METHOD DETAILSConuropsis DNA extraction and sequencing Two different samples of about 100 mg were obtained, one from the femur (leg bones were preserved inside the naturalized specimen) and one from toepads, with the help of a Dremel machine.The two samples were digested using 2mL of extraction buffer containing 10mM Tris-HCL (pH 8), 10mM NaCl, 5mM CaCl2, 2.5mM EDTA, 1% SDS, 1% Proteinase K and 40mM DTT. After incubation, digested samples were centrifuged for 3 min at 3000 3 g and the supernatant was collected and mixed with 1X volume of Phenol. The sample solution was incubated on a rotor for 5min at RT. After, it was centrifuged for 3 min at 5000 3 g and the upper aqueous layer was collected in new low-bind Eppendorf tube. The upper aqueous layer was collected in new tube and mixed with 10X volume of binding buffer prepared as previously described [67].The sample solution mixed with the binding buffer was poured into a binding apparatus constructed by fitting an ZymoV extension reservoir in a MinElute column and set inside a 50mL Falcon tube Samples were centrifuged at 6000 3 g and extracted DNA was quantified using a Qubit instrument
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