Juventus striker Morata was stretchered off after half an hour clutching his lower leg.
Neither will travel now for the final Group C clash with Ukraine.
Early reports suggest the pair could be sidelined for two or three weeks and for Premiership leaders City, Silva's absence is yet another blow coming on the back of Sergio Aguero’s injury playing for Argentina.
"Silva is out of the squad due to a bad sprain of the internal lateral ligament of the right ankle," read a statement on the Spanish national team website.
"Morata also will not travel to Ukraine having suffered bruising to his right leg."
European champions in 2008 and 2012, Spain qualified comfortably for a place in next year’s competition in France and lead the Group C table with 24 points from nine games.
Spain are looking to bounce back from their disappointing performance at the 2014 World Cup in Brazil where they failed to retain the trophy they won four years earlier in South Africa and went out in the group stage.
(Reporting by Tim Hanlon in Barcelona, editing by Amlan Chakraborty)
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