Accession Number:

ADA433727

Title:

Real-Time Polymerase Chain Reaction Assays for Rickettsial Diseases

Descriptive Note:

Conference paper

Corporate Author:

NAVAL MEDICAL RESEARCH CENTER SILVER SPRING MD

Personal Author(s):

Report Date:

2004-06-01

Pagination or Media Count:

13.0

Abstract:

Arthropod-borne rickettsial diseases are found worldwide and have been the cause of significant amounts of suffering, disability and fatalities among both military and civilian populations throughout history. Because of the similarity to many infectious diseases in signs and symptoms, rickettsial diseases are difficult to diagnose clinically. Moreover, due to the time it takes for antibodies to develop and the low concentration of rickettsial agents in the blood stream the diseases are also difficult to diagnose by laboratory methods. For that reason we have developed real-time PCR assays to detect rickettsial disease agents both at the genus and the species level. Real-time PCR assays were developed to identify 1 pathogenic Rickettsia 2 Rickettsia prowazekii and R. rickettsii, the etiological agents for epidemic typhus and Rocky Mountain spotted fever RMSF and potential BW agents R. typhi and R. felis, the flea-borne typhus disease agents, and Orientia formerly Rickettsia tsutsugamushi, the scrub typhus agent. The assays utilize molecular beacon probes, which fluoresce when they encounter the target DNA sequence. By manipulating the annealing temperature, and magnesium, probe and primer concentrations of the assays, the optimal conditions were determined. A panel of 22 strains of rickettsiae, 20 strains of orientiae and 19 species of non-rickettsial agents were used to determine the specificity of the assays. Plasmids encoding the target sequences were used to calculate the sensitivity of the assays. These real-time PCR assays were found to be capable of detecting rickettsial disease agents quickly and with great sensitivity and specificity.

Subject Categories:

  • Medicine and Medical Research
  • Microbiology

Distribution Statement:

APPROVED FOR PUBLIC RELEASE