The initial peopling of the Americas is a contested and evolving topic 1 , with the exact timing of the first arrivals still unknown. Historically, Mexico's understudied and controversial archaeological record has remained on the periphery of First Americans research 2 . However, in recent years, investigations have shown reliable evidence of a late-Pleistocene and early-Holocene human presence in the northwest region 3,4 , the Chiapas Highlands 5 , Central Mexico 6 , and the Caribbean coast [7][8][9] . Evidence of human presence at Chiquihuite Cave extends this antiquity and attests to the cultural variability of older-than-Clovis sites [10][11][12][13][14][15][16][17] and the earliest humans on the continent. Site settings and excavation context.

Highlights d We recovered genomic data from ancient bears directly from cave sediments d Ancient Mexican black bears are ancestrally related to Eastern American black bears d The Mexican and Yukon populations of extinct giant shortfaced bears were distinct d Our results bring ancient eDNA into the era of population genomics

Highlights d We recovered genomic data from ancient bears directly from cave sediments d Ancient Mexican black bears are ancestrally related to Eastern American black bears d The Mexican and Yukon populations of extinct giant shortfaced bears were distinct d Our results bring ancient eDNA into the era of population genomics

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