Several staff members required treatment for their symptoms, spokeswoman Heather Nauert said.
The attacks appeared to have ceased, Ms Nauert added.
Cuba has denied targeting foreign diplomats and said it was investigating the allegations.
The Associated Press reported that the diplomats' hearing loss could have been linked to sonic devices emitting inaudible sound waves that can cause deafness.
"We can confirm that at least 16 US government employees, members of our embassy community, have experienced some kind of symptoms," Ms Nauert said.
"We take the situation extremely seriously," she added.
US embassy staff and at least one Canadian began to notice symptoms late last year.
The victims, some of whom have been withdrawn from Cuba, have been treated in the US and by US doctors in Cuba.
US Secretary of State Rex Tillerson has described the incident as a "health attack".
What stands out is that the incidents appeared to continue even after the US staff complained of them and as the Cuban authorities were investigating the cause, the BBC's Will Grant in Havana says.
There are currently investigations under way by the Americans, the Canadians and the Cubans, and security has been stepped up around the diplomats' residences in Havana.
Security analysts believe a third country with hostile relations with the US may be involved but no clear picture has yet been established.
Washington expelled two Cuban diplomats from the US in response to the incident.
Washington and Havana only re-established ties in 2015, following 50 years of hostilities between the two countries.