During World War II, she operated in the Central Pacific and supported Japanese forces in the Battle of Wake Island until she sank after a collision in December 1941.

For surface running, the submarines were powered by two 1,200-brake-horsepower (895 kW) Vickers diesel engines, each driving one propeller shaft.

On 6 December 1941, Ro-66 got underway from Kwajalein with the commander of Submarine Squadron 27 embarked to conduct a reconnaissance of Wake Island,[3] which the Japanese planned to invade on 8 December 1941, the first day of the war on Wake Island′s side of the International Date Line.

The Battle of Wake Island began on 8 December 1941, with Ro-65, Ro-66, and Ro-67 patrolling in the area supporting Japanese forces attempting to seize the atoll.

[3] United States Marine Corps forces defending Wake drove back the initial Japanese assault that day.

[3] Accordingly, Ro-65 and Ro-67 headed back to Kwajalein, but a radio failure prevented Ro-66 from receiving the orders despite three attempts by Submarine Squadron 7 to contact her.

[4][5][6] Ro-66 was on the surface 25 nautical miles (46 km; 29 mi) southwest of Wake Island — bearing 252 degrees from the atoll — to recharge her batteries in a heavy squall in the predawn darkness of 17 December 1941 when her lookouts suddenly sighted Ro-62, also on the surface and recharging batteries.

[2][3][6] Both submarines attempted to back off, but it was too late to avoid a collision, and Ro-62 rammed Ro-66 at 20:20 Japan Standard Time.