Molecular diagnosis and duration of eschar swab sample positivity post-doxycycline therapy in scrub typhus patients

Background: An eschar is not always present in all scrub typhus patients. Furthermore, such patients may present to tertiary care hospitals after administration of doxycycline. The present study aimed to determine the usefulness of using the swab from eschar sites in the diagnosis of scrub typhus in patients who present post-doxycycline therapy. Methods: Blood, eschar scraping and swab samples were collected daily until patient discharge/death. Realtime SYBR green PCR targeting the groEl gene, TaqMan probe PCR targeting the 47 kDa gene and nested PCR targeting the 56 kDa gene of Orientia tsutsugamushi were carried out. Partial sequences of the 56 kDa gene of O. tsutsugamushi were sequenced by the Sanger method and phylogenetic analysis was performed using Mega X. Results: In total, 42 samples (19 eschar scraping and 23 eschar swab samples) were collected from 22 patients. A high positivity of eschar scraping samples (89.5%, 17/19) in comparison to blood (63.2%, 12/19) was observed. The nested PCR for eschar swab samples was positive in 10 (43.5%), 47 kDa gene in nine (39.1%) and groEl in three (13%) samples. The swabs remained positive for 1-4 d after doxycycline was started. The majority of the sequences clustered with Karp-like strains. Conclusion: The eschar swab is a good alternative sample for the diagnosis and genotyping of scrub typhus. It also has the added advantage of persistent positivity despite doxycycline administration.