An ELISA assay using a combination of recombinant proteins from multiple strains of Orientia tsutsugamushi offers an accurate diagnosis for scrub typhus

Chien-Chung Chao,Wei-Mei Ching,Zhiwen Zhang,Ekkarat Wongsawat,Saowaluk Silpasakorn,Wilawan Thipmontree,Wiwit Tantibhedyangkul,Tatyana Belinskaya,Yupin Suputtamongkol

doi:10.1186/s12879-017-2512-8

Abstract

BackgroundScrub typhus (ST) is a disease caused by an obligate intracellular bacterium, Orientia tsutsugamushi, an organism that requires a BSL3 laboratory for propagation. The disease is hallmarked by an eschar at the site of the chigger bite, followed by the development of fever, malaise, myalgia, anorexia, and papulomacular rash. Indirect immunofluorescent assay (IFA) is the gold standard for scrub typhus diagnosis, however, the subjectivity of the assay, the need for a specialized laboratory and instruments has limited the wide use of the test in resource limited areas.MethodsA recombinant-protein based enzyme linked immunosorbent assay (ELISA) using the most abundant and immunodominant protein for the detection of Orientia specific antibodies in serum has been developed. The performance of the assay was evaluated using prospectively collected acute sera from 248 randomly selected patients in Thailand. The ELISA assay was evaluated using two different cutoff values.ResultsThe receiver operating characteristic (ROC) curve generated cutoff values gave slightly better consistency with diagnosis of ST than those cutoff values established by averaging ELISA optical density of known negatives at 99% confidence interval. Both cutoff values provided similar statistical parameters when compared with the diagnosis of ST, indicating the validity of both calculations to derive cutoff values. These results suggest that both IgG and IgM ELISA performed well to accurately diagnose scrub typhus cases in endemic areas using only acute serum samples.ConclusionsWe have successfully developed an ELISA assay for the detection of Orientia-specific antibodies in serum that could provide effective screening of acute sera under clinical setup and it is also a useful assay to estimate seroprevalence in various endemic areas.

Highlights

Scrub typhus (ST) is a disease caused by an obligate intracellular bacterium, Orientia tsutsugamushi, an organism that requires a BSL3 laboratory for propagation
The enzyme linked immunosorbent assay (ELISA) assay has similar performance using the receiver operating characteristic (ROC) curved derived cutoffs in comparison with those using known negative samples. Both IgG and IgM have good Likelihood ratio positive (LR)+ (7.27–16.3) and very low LR- (0.07–0.20), strongly suggest both are very good diagnostic assays [35] even though diagnosis criteria were based on results from both acute and convalescent sera and only acute sera were used for ELISA
Since only the acute specimens from individual patients was used for IgG or IgM ELISA with sensitivity and specificity suitable as diagnostic assays, these results support the clinical utility of either IgG or IgM ELISA to be used for ST diagnosis using only the acute specimens and some known negatives

Summary

Introduction

Scrub typhus (ST) is a disease caused by an obligate intracellular bacterium, Orientia tsutsugamushi, an organism that requires a BSL3 laboratory for propagation. Scrub typhus (ST) is one of the most common rickettsial diseases caused by the infection of Orientia tsutsugamushi, an obligate intracellular Gram-negative bacterium [1, 2]. The disease is characterized by fever, rash, eschar, pneumonitis, meningitis, and in some cases, disseminated intravascular coagulation that may lead to circulatory failure [3]. Reports from India documented 12–17% case fatality rate in recent years [5, 6]. Scrub typhus cases usually were reported primarily from the rural populace, but recently it is being increasingly detected in people residing in expanding cities [11]

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