Usefulness of rickettsial PCR assays for the molecular diagnosis of human rickettsioses.

BACKGROUND

The effectiveness of PCR methods to amplify rickettsiae from clinical samples has still not been evaluated. Our aim was to determine the sensitivity and usefulness for Rickettsia species identification by PCR methods, targeting 16S rDNA, htrA, gltA, ompA, and ompB genes for molecular diagnosis of rickettsioses.

METHODS

A total of 72 clinical samples (EDTA-blood, skin biopsies and ticks) taken from 52 patients in the early phase of the illness with PCR-confirmed rickettsioses were included. Single [16S rDNA, gltA (5' end), and htrA genes] and sequential (nested or semi-nested) PCR assays [ompB, gltA (central region) and ompA genes] were performed.

RESULTS

For single-stage PCR assays, the greatest sensitivity (33.3%) was obtained using the gltA (5' end), while for sequential assays, the most sensitive results were obtained using the ompB assay (83.3%). The highest sensitivity (100%) was achieved using the three sequential PCRs. The ompA PCR method was the most reliable for identifying Rickettsia species, according to clinical features.

CONCLUSIONS

PCR-based amplification methods are useful rickettsial diagnostic tools in the early phase of the illness. The three sequential PCR assays here investigated (ompB, gltA and ompA) appear to be useful tools for molecular diagnosis of rickettsioses. ompB PCR assay is effective for primary screening, since it detects a high percentage of positive samples. ompA assay is the most useful method to identify a Rickettsia species in human pathology. Nevertheless, epidemiology, clinical symptoms and the vector involved in the infection have to be taken into account for the diagnosis of rickettsioses.