Genetic sequencing of the 56-kDa surface protein antigen gene of Orientia tsutsugamushi causing contemporary human scrub typhus at two sites in Sri Lanka

Abstract

INTRODUCTION AND OBJECTIVES: Genetic and antigenic characterisation of Orientia tsutsugamushi (OT) causing human disease in OT-endemic regions of Sri Lanka is not known. METHODS: Eschar biopsies (in age> 18 years} and buffy coat samples {in age<18 years) collected from patients with OT confirmed by IFA were further studied at the Rickettsia! Diseases Research Laboratory, Texas, USA by real time PCR (Orientia 47 kD} and nested PCR (Orientia 56 kD) amplification. DNA sequences were obtained for 56 kD gene amplicons and phylogenetic comparisons were analysed using currently available data in GenBank [Neucleotide substitution per 100 residues, 1000 Bootstrap Trials]. RESULTS: Twenty eschar biopsies (Balapitiya:19, Ragama:1} and eight buffy coat samples (Balapitiya:6, Ragama:2} examined by real time PCR revealed Orientiaamplicons in 16 samples. DNAsequences were obtained for the 56 kD gene amplicons in 12 eschars and 4 buffy coat samples. The genotypes of the Balapitiya samples revealed that, 7 exhibiting close homology with JP1 [distantly related to UT177 Thai (Karp related}], two had close homology with JGv and JG AF [distantly related to Kawasaki M63383], one had close homology with Gilliam strain, and five had close homology with Kato strain. The Ragama strain was closely related to Kuroki-Boryong L04956. CONCLUSIONS: We observed all three main OT genotypes in Sri Lanka, and the majority fell into Thai Karp related clade. These results demonstrate antigenic diversity of OT in the studied areas of Sri Lanka.