The Manchester United captain has returned to Carrington for further assessment on the problem after sitting out Monday's training session.
However, he may not necessarily be out of United's Premier League fixture with Arsenal on Saturday.
The England international was due to be rested in Tuesday's friendly with Spain at Wembley by interim manager Gareth Southgate, so an early release from the Three Lions' camp is a logical precaution.
Jose Mourinho will already be missing striker Zlatan Ibrahimovic against Arsene Wenger's Gunners after the Swede picked up a suspension for a fifth yellow card against Swansea last time out.
Marcus Rashford will be hoping to get the start up front for United, but was forced to play on the wing in the defeat to Fenerbahce while Rooney was in the central role until Ibrahimovic was introduced.
Arsenal will be without Hector Bellerin for the clash, but Alexis Sanchez has been declared fit to play for Chile on Tuesday after an injury scare last week when he suffered a "low grade" muscle tear.
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