Rickettsia (Spotted Fever Group and Typhus Group)

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Background
This page provides routine testing information for spotted fever group (SFG) and typhus group (TG) rickettsioses at Public Health Ontario (PHO).

The etiological agent of SFG rickettsioses include many rickettsiae species such Rickettsia rickettsii (Rocky Mountain spotted fever), Rickettsia africae (African tick bite fever), Rickettsia akari (rickettsialpox), Rickettsia conorii (Mediterranean spotted fever), Rickettsia australis (Queensland tick typhus), Rickettsia felis (flea rickettsiosis), Rickettsia honei (Flinders Island spotted fever), Rickettsia philipii (Pacific coast tick fever), Rickettsia japonica (Japanese spotted fever), Rickettsia sibirica (Siberian tick typhus), and Rickettsia parkeri.

The etiological agent of TG rickettsioses includes Rickettsia typhi (murine typhus) and Rickettsia prowazekii (epidemic louse-borne typhus).

For testing of Orientia tsutsugamushi (scrub typhus), refer to the following PHO webpage:

Orientia Tsutsugamushi

Updates

As of September 27, 2024, additional clarification regarding testing criteria, performance, and limitations have been added, as well as instructions for PCR testing.

* Copie d'impression non contrôlée. Valide uniquement le jour de l'impression : 21 nov. 2024.

Testing Indications

Testing is indicated for individuals with a history of exposure to relevant arthropods in endemic areas and clinical presentation compatible with a suspected rickettsial infection.

Acceptance/Rejection Criteria

Requests for testing will only be accepted if the following information is provided:

Clinical history with symptom onset within the last 30 days
Exposure to endemic region and/or relevant arthropod

Specimen Collection and Handling

Specimen Requirements

Test Requested	Required Requisition(s)	Specimen Type	Minimum Volume	Collection Kit
Rickettsia or rickettsiosis serology or antibody	General Test Requisition	Serum	5.0 ml whole blood or 1.0 ml serum	Serum separator tube (SST)
Rickettsia or rickettsiosis PCR or molecular	General Test Requisition	Whole blood	1.0 ml whole blood	EDTA tube
Rickettsia or rickettsiosis PCR or molecular	General Test Requisition	Tissue biopsy (eschar or skin lesion)	N/A	Empty sterile container
Rickettsia or rickettsiosis PCR or molecular	General Test Requisition	Skin swab (under eschar)	N/A	Empty sterile container

Submission and Collection Notes

Complete all fields of the General Test Requisition form including:

Clinical history (including if antibiotics administered)
Symptom onset date
Travel and arthropod exposure history

Label the specimen container(s) with the patient’s first and last name, date of collection, and one other unique identifier such as the patient’s date of birth or Health Card Number. Failure to provide this information may result in rejection or testing delay.

Timing of Specimen Collection

For serology, collecting both an acute serum (collected early after the onset of symptoms) and a convalescent serum (collected either 2-3 weeks later for most Rickettsia species or a minimum of 4 weeks later for Rickettsia africae) may be required for laboratory confirmation of infection.

For PCR, samples should ideally be collected prior to the initiation of antibiotic therapy, but treatment should never be withheld, if required based on the patient’s clinical status.

Limitations

Grossly haemolysed, lipemic, or contaminated specimens are unsuitable for testing.

Storage and Transport

Centrifuge serum if using SST. To prevent erroneous results due to the presence of fibrin, ensure that complete clot formation has taken place prior to centrifugation of samples.
Blood samples should be stored at 2-8°C following collection and shipped on ice packs to PHO’s laboratory as soon as possible.
Tissue samples should be stored frozen following collection and shipped on dry ice to PHO’s laboratory as soon as possible.
All clinical specimens must be shipped in accordance with the Transportation of Dangerous Goods Act.

Requisitions and Kit Ordering

Test Frequency and Turnaround Time (TAT)

Rickettsia serology is performed once per week at PHO’s laboratory. Turnaround time for serology is up to 10 days from receipt at PHO’s laboratory.

Rickettsia PCR is forwarded to the National Microbiology Laboratory (NML) in Winnipeg. Turnaround time for PCR is up to 21 calendar days from receipt at PHO’s laboratory.

Test Methods

Method: Rickettsia serology is performed at PHO using the commercial Focus Diagnostics Rickettsia Indirect Immunofluorescence Antibody (IFA) IgG assay. This duplex assay provides semi-quantitative detection of IgG antibodies to either SFG rickettsiae (based on a R. rickettsii antigen) or TG rickettsiae (based on a R. typhi antigen).

Rickettsia PCR is performed at NML using a laboratory-developed real-time PCR specific for the 17kDa antigen gene and citrate synthase gene of the Rickettsia genus. If a sample is positive by real-time PCR at the genus level, conventional PCR and sequencing of the outer membrane protein A will be performed by NML to determine the exact Rickettsia species.¹

Performance: Single acute serology sensitivity is < 50% when collected within the first week of illness. Paired acute and convalescent serology demonstrating a 4-fold increase in titres has an estimated sensitivity of 85-100% and specificity of 91-100%. Skin tissue PCR sensitivity for SFG Rickettsia infections ranges from 55-79%, but is low for TG Rickettsia infections (5-6% only). Whole blood PCR sensitivity for SFG Rickettsia infections ranges from 24-56% with higher sensitivity in early or severe infections, and is low for TG Rickettsia infections (2-10% only). PCR specificity is often ≥ 98% and can provide species-level identification.^2,3,4,5

Limitations: Serology may be negative in the acute stage of illness, and does not distinguish active versus remote infection unless paired sera are collected demonstrating a 4-fold increase in titres. A positive SFG Rickettsia serology result cannot distinguish between SFG Rickettsia species. Similarly, a positive TG Rickettsia serology result cannot distinguish between TG Rickettsia species. Cross-reactivity between the SFG and TG rickettsioses can also occur, but titres from the cross-reactive group are generally lower than the titres of the group causing the patient’s infection. Cross-reactivity may also occur with other non-rickettsial infections.⁶

Interpretation

For Rickettsia serology:

SFG Rickettsia IgG by IFA Result	TG Rickettsia IgG by IFA Result	Interpretation
Non-reactive < 1:64	Non-reactive < 1:64	No serological evidence of SFG or TG rickettsial infection. A non-reactive test result does not exclude the diagnosis of rickettsial disease. If rickettsial disease is suspected, submit a new specimen for repeat serology.
Reactive ≥ 1:64	Non-reactive < 1:64	Serological evidence of SFG rickettsial infection/exposure. Results should be compared between paired samples with 4-fold or higher changes in titers indicating acute infection.
Non-reactive < 1:64	Reactive ≥ 1:64	Serological evidence of TG rickettsial infection/exposure. Results should be compared between paired samples with 4-fold or higher changes in titers indicating acute infection.
Reactive ≥ 1:64	Reactive ≥ 1:64	Serological evidence of SFG and/or TG rickettsial infection/exposure. Cross-reactivity between the groups can occur, but cross-reactive titers are generally lower than group-specific titers. Results should be compared between paired samples with 4-fold or higher changes in titers indicating acute infection.

For Rickettsia PCR:

Typhi (TG Rickettsia) PCR Result	SFG Rickettsia PCR Result	Rickettsia PCR Sequencing Result	Interpretation
Negative	Negative	-	No evidence of Rickettsia DNA. A negative result does not exclude the diagnosis of rickettsial disease.
Positive	N/A	Species Reported	<<Species detected>> DNA detected
N/A	Positive	Species reported	<<Species detected>> DNA detected

Reporting

Results are reported to the physician, authorized health care provider (General O. Reg 45/22, s.18) or submitter as indicated on the requisition.

References

Stenos J, Graves SR, Unsworth NB. A highly sensitive and specific real-time PCR assay for the detection of spotted fever and typhus group Rickettsiae. Am J Trop Med Hyg. 2005 Dec;73(6):1083-5.
Dumler JS, Taylor JP, Walker DH. Clinical and laboratory features of murine typhus in south Texas, 1980 through 1987. JAMA. 1991 Sep 11;266(10):1365-70.
Stewart AG, Stewart AGA. An Update on the Laboratory Diagnosis of Rickettsia spp. Infection. Pathogens. 2021 Oct 13;10(10):1319. doi: 10.3390/pathogens10101319.
Paris DH, Dumler JS. State of the art of diagnosis of rickettsial diseases: the use of blood specimens for diagnosis of scrub typhus, spotted fever group rickettsiosis, and murine typhus. Curr Opin Infect Dis. 2016 Oct;29(5):433-9. doi: 10.1097/QCO.0000000000000298.
Reller ME, Dumler JS. Optimization and Evaluation of a Multiplex Quantitative PCR Assay for Detection of Nucleic Acids in Human Blood Samples from Patients with Spotted Fever Rickettsiosis, Typhus Rickettsiosis, Scrub Typhus, Monocytic Ehrlichiosis, and Granulocytic Anaplasmosis. J Clin Microbiol. 2020 Aug 24;58(9):e01802-19. doi: 10.1128/JCM.01802-19
Wood H, Artsob H. Spotted fever group rickettsiae: a brief review and a Canadian perspective. Zoonoses Public Health. 2012 Sep;59 Suppl 2:65-79. doi: 10.1111/j.1863-2378.2012.01472.x.