Virulence of pathogenic bacteria is a tightly controlled process to facilitate invasion and survival in host tissues. Although pathways controlling virulence have been defined in detail, signals modulating these processes are poorly understood. The opportunistic pathogen Pseudomonas aeruginosa causes acute and chronic infections in humans. Disease progression is typically associated with a loss of acute virulence and the emergence of biofilms and chronic behaviour. The acute-to-chronic switch is governed by the global Gac/Rsm pathway. Using a newly developed acute-chronic dual reporter system we show that calcium stimulates the Gac/Rsm pathway via the Gac-associated hybrid histidine kinase LadS. We show that calcium binds to the periplasmic DISMED2 sensor domain of LadS to activate its kinase activity. Activation of the Gac/Rsm pathway by calcium leads to a switch to the chronic program and confers drug tolerance by reducing P. aeruginosa growth rate. Clinical isolates from cystic fibrosis airways retain their calcium response during chronic infections. Our data imply that calcium sensing evolved as an adaptation to the opportunistic lifestyle of P. aeruginosa and that calcium serves as a host signal to balance acute-to-chronic behaviour during infections. Establishing calcium signalling in host-pathogen interaction adds to growing evidence indicating key roles for calcium in bacterial signalling.