The Barcelona defender missed the world and European champions' training session on Saturday and may not feature in Monday's Group B clash in Curitiba.
A statement on the official website for the Spanish Football Federation (RFEF) said: "Gerard Pique had to withdraw due to pain in the adductor muscle region."
Pique was dropped for Spain's 2-0 defeat against Chile on Wednesday on the back of their 5-1 thrashing at the hands of the Netherlands in their first game of the tournament.
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