Mycobacterium infections in nonhuman primates can devastate the colonies and place human handlers at risk. Despite conservative measures to prevent exposure, infections occur even in closed colonies. Here we describe a recent case of M. tuberculosis within a closed colony of rhesus macaques at our Thailand facility and the procedures instituted to prevent subsequent infections. Investigation of the outbreak did not confirm the source of the infection, but even with intensive occupational safety measures in place, human contact remains the most likely possibility.