AUTHOR=Jiang Ju , Martínez-Valdebenito Constanza , Weitzel Thomas , Farris Christina M. , Acosta-Jamett Gerardo , Abarca Katia , Richards Allen L. 

TITLE=Development of a New Genus-Specific Quantitative Real-Time PCR Assay for the Diagnosis of Scrub Typhus in South America

JOURNAL=Frontiers in Medicine

VOLUME=9

YEAR=2022

URL=https://www.frontiersin.org/journals/medicine/articles/10.3389/fmed.2022.831045

DOI=10.3389/fmed.2022.831045

ISSN=2296-858X

ABSTRACT=<p>Scrub typhus is a potentially severe rickettsiosis, caused by <italic>Orientia tsutsugamushi</italic> in the Asia-Pacific region. Recently, however, two distinct pathogens, “<italic>Candidatus</italic> Orientia chuto” and “<italic>Candidatus</italic> Orientia chiloensis”, have been discovered in the Middle East and South America, respectively. Since the novel pathogens differ significantly from <italic>O. tsutsugamushi</italic>, many established diagnostic methods are unreliable. This work describes the development and validation of a new quantitative real-time PCR (qPCR) assay (Orien16S) for the detection of all known <italic>Orientia</italic> species. Based on a 94 bp sequence of the 16S rRNA gene (<italic>rrs</italic>), Orien16S recognized DNA samples from <italic>O. tsutsugamushi</italic> (<italic>n</italic> = 41), <italic>Ca.</italic> O. chiloensis (<italic>n</italic> = 5), and <italic>Ca.</italic> O. chuto (<italic>n</italic> = 1), but was negative for DNA preparations from closely related rickettsiae and other members of the order Rickettsiales (<italic>n</italic> = 22) as well as unrelated bacterial species (<italic>n</italic> = 11). After its implementation in Chile, the assay was verified, correctly identifying all tested eschar and buffy coat samples (<italic>n</italic> = 28) of clinical suspected cases. Furthermore, Orien16S detected <italic>Orientia</italic> DNA in trombiculid mites collected in endemic regions in southern Chile. The presented novel qPCR assay provides a useful tool for detecting <italic>Orientia</italic> and diagnosing scrub typhus from all geographical regions.</p>