Genetic diversity of Orientia tsutsugamushi strains from patients in north India

Background: Scrub typhus has emerged as a major cause of acute febrile illness in India in recent years. The causative agent, Orientia tsutsugamushi has more than 20 prototype strains due to a variable 56-kDa outer membrane protein. It is crucial to know the prevailing types in India for the success of diagnostic immunoassays and prospective vaccine candidates. In north India, the principal types circulating are largely unknown. Our tertiary care hospital caters to a large area of north India (around 7 states and one union territory). Therefore, the current study was planned to identify the genotypes of O. tsutsugamushi strains circulating in this wide area of north India. Materials and methods: Adults and children presenting with suspected scrub typhus between July 2013 and December 2013 were included in this study. DNA was extracted from whole blood and a nested PCR was used to amplify a 483-bp region of the 56-kDa antigen gene of O. tsutsugamushi. The PCR products were purified and DNA sequencing was performed and aligned using the CLUSTAL_X2 program. A phylogenetic tree was constructed using neighbour- joining algorithms and analysed using the sequences obtained in this study and those obtained from the GenBank database. Results: A total of 34 samples were positive for PCR. The amplicons were sequenced and analyzed. Karp-like strains predominated in all states studied (64.7%) followed by Gilliam-like (26.47 %) and 8.82% similar to Hualien 1 or S072. We did not find any Kato or Kawasaki-like strain. Karp like strains showed >99% similarity to TH2033, TH2191, TH2208, Xinjiang & Neimeng strains and Gilliam-like strains showed >99% similarity to Clone ISS -11. Conclusion: Orientia tsutsugamushi shows a great diversity in its strains over a large geographical area of north India. This has implications in the production of both diagnostic assays and vaccine for scrub typhus. Keywords: Scrub typhus, Orientia tsutsugamushi, Eschar, North India