Helicobacter pylori-like organisms (HPLOs) were isolated from the gastric mucosa of two Sumatran tigers and identified by polymerase chain reaction analysis as Helicobacter acinonyx. At histological examination, both tigers revealed a chronic gastritis associated with HPLOs as demonstrated by immunolabelling and electron microscopy. This is the first isolation of H. acinonyx from tigers, in which, as previously reported in cheetahs, it may be a cause of gastritis.