Holmes has a left quad strain and won't be able to play for the USA in the Concacaf competition.
Midfielder Duane Holmes has been dropped from the U.S. Concacaf Gold Cup roster because of a reoccurrence of a left quadriceps strain, and he has been replaced by midfielder Djordje Mihailovic.
The U.S. Soccer Federation announced the change Saturday, four days after the Americans replaced injured midfielder and defender Tyler Adams with Reggie Cannon.
Holmes made his U.S. debut on June 5 in an exhibition against Jamaica.
Coming off a 1-0 loss to Jamaica and a 3-0 defeat to Venezuela, the U.S. opens its Gold Cup title defense against Guyana on Tuesday at St. Paul, Minnesota, in the Americans' first competitive match since the October 2017 loss at Trinidad and Tobago that ended a streak of seven World Cup appearances. They face Trinidad on June 22 at Cleveland and complete group play against Panama on June 26 at Kansas City, Kansas.
The revised roster:
Goalkeepers: Sean Johnson (NYCFC), Tyler Miller (LAFC), Zack Steffen (Columbus Crew)
Defenders: Reggie Cannon (FC Dallas), Omar Gonzalez (Toronto FC), Nick Lima (San Jose Earthquakes), Aaron Long (New York Red Bulls), Daniel Lovitz (Montreal Impact), Matt Miazga (Chelsea), Tim Ream (Fulham), Walker Zimmerman (LAFC)
Midfielders: Michael Bradley (Toronto), Weston McKennie (Schalke, Germany), Djordje Mihailovic (Chicago Fire), Christian Pulisic (Chelsea), Cristian Roldan (Seattle Sounders), Wil Trapp (Columbus Crew)
Forwards: Jozy Altidore (Toronto FC), Paul Arriola (D.C. United), Tyler Boyd (Vitoria Guimares), Jonathan Lewis (Colorado Rapids), Jordan Morris (Seattle Sounders), Gyasi Zardes (Columbus Crew)