Clotted blood samples of 240 scrub typhus patients were collected from 8 Regional Medical Sciences Centers in Thailand during 1999 to 2002. The serotypes of Orientia tsutsugamushi and their geographic distribution were determined. A nested polymerase chain reaction (PCR) was used to identify the serotypes of O. tsutsugamushi. The number of patients with positive results for O. tsutsugamushi was 25.0%. Two serotypes, Karp and Kato, were detected in these samples. No Gilliam serotype was detected from any of the study locations. The PCR products were sequenced using an automated DNA sequencer. The nucleotide sequence of gene encoding 56-kDa protein from these samples showed a high sequence homology with the reference sequence of O. tsutsugamushi Karp and Kato serotypes. O. tsutsugamushi Karp serotype was predominant throughout Thailand with the percentage of 96.8% of the total serotype-positive patients, whereas 3.2% for Kato serotype was observed only in the south. The highest number among the region of Karp serotype-positive patients of 31.6% was found in the northeast. © 2006 Elsevier Inc. All rights reserved.