To investigate the genotype of Orientia tsutsugamushi (O. tsutsugamushi) infection in Zhejiang and Fujian provinces, southeastern China and to analyze the gene variation.

Strains of O. tsutsugamushi were isolated from the blood of 5 scrub typhus patients of febrile phase and then cell culture were carried out. DNA from the infected cells were extracted and full-length 56-kD type-specific antigen (TSA) gene and full-length heat shock protein gene (groESL) were amplified. The gene products were sequenced. Multiple sequence alignment and phylogenetic analyses were conducted by Mega 7.0 software.

Five febrile patients with O. tsutsugamushi infection were confirmed etiologically and five O. tsutsugamushi strains were isolated. The sequence alignment of 56-kD TSA gene and groESL gene showed that 2 among the 5 strains had 100% identity; while other two strains also had 100% identity of these two genes. These two groups were temporarily named Zhejiang-1 and Zhejiang-2. The 56-kDa TSA gene alignment and phylogenetic analysis showed that Zhejiang-1 was closely related to Taiwan-A genotype (98.45%) and Zhejiang-2 was closely related to Gilliam-C genotype (98.50%), but there were obvious variations. The other 1 strain of Wuj/2014 was significantly similar to Taiwan-A genotype (99.94%). Based on 56-kDa TSA gene, the timetree based on the RelTime method showed that Taiwan-A genotype, Zhejiang-1 and especially Zhejiang-2 genotype were later than other prototypes on the divergence time from their ancestors of each genotypic clade. These genotypes or subtypes emerged later in evolutionary process of O. tsutsugamushi.

There may be unrecognized novel O. tsutsugamushi variants among the patients in this study. The relationship between genetic variation and severity of scrub typhus also should be discussed.