Rickettsial Serology

Rickettsial diseases are caused by obligate intracellular bacteria transmitted by arthropods.

They include the spotted fever group (Rickettsia rickettsii, R. conorii, R. africae, R. japonica), the typhus group (R. prowazekii in epidemic louse borne typhus, R. typhi in murine typhus), scrub typhus (Orientia tsutsugamushi), ehrlichiosis and anaplasmosis (Ehrlichia chaffeensis, Anaplasma phagocytophilum).

The reference serological test is the indirect immunofluorescence assay (IFA) for IgM and IgG against group specific antigens. Other methods are ELISA, immunoperoxidase and the obsolete Weil-Felix test. PCR of whole blood (first week), skin biopsy of an eschar or rash and swabs of an eschar is highly specific and detects infection before seroconversion.

These infections occur worldwide with regional patterns: African tick bite fever in sub-Saharan Africa (R. africae, mild, with eschars), Mediterranean spotted fever in southern Europe and North Africa, scrub typhus in the Asia-Pacific "tsutsugamushi triangle" (about 1 million cases each year), Rocky Mountain spotted fever in the Americas (fatal in 20 to 30% if untreated),

louse borne typhus in cold, crowded and conflict settings (Burundi, Ethiopia, Rwanda), and murine typhus in ports and cities with rats.

Disease presents with fever, headache, myalgia, rash, eschar and regional lymphadenopathy. Severe disease includes encephalitis, pneumonitis, hepatitis, renal failure and shock.

Indications. Acute febrile illness with rash, eschar or tick or mite exposure after travel or field work. Fever with headache and myalgia in endemic regions where malaria and dengue tests are negative. Returned traveller from safari, trekking or rural areas with fever and eschar. Louse borne typhus outbreak investigation in refugee camps and prisons.

Fever with thrombocytopenia, raised liver enzymes and leucopenia (ehrlichiosis and anaplasmosis). Pyrexia of unknown origin in endemic zones.

Specimen. Acute and convalescent serum 1 to 2 mL (collected at presentation and 14 to 21 days later). Whole blood in EDTA for PCR within the first week. Skin biopsy from rash or eschar, or an eschar swab, for PCR and immunohistochemistry.

Reference values (assay dependent).

• IFA IgG: negative under 1 in 64 (some laboratories 1 in 128). IFA IgM: negative under 1 in 32.
• Rickettsial DNA PCR: not detected.

Interpretation.

• A fourfold or greater rise in IFA IgG between acute and convalescent sera taken 2 to 4 weeks apart is the diagnostic criterion.
• A single IgG titre of 1 in 64 to 1 in 128 or higher with compatible illness supports the diagnosis in non-endemic areas. In endemic areas high background titres (up to 1 in 256 or more for scrub typhus) mean paired sera are needed.
• IgM appears from days 5 to 7 of illness and IgG from days 7 to 10, and both are usually negative at presentation. A negative first sample does not exclude infection.
• Group cross-reactivity within spotted fever group and within typhus group means species level identification needs cross-adsorption or PCR.
• PCR of whole blood is positive in the first days when antibody is absent (sensitivity about 50 to 70% early in the disease, higher for eschar swabs and skin biopsies).
• Weil-Felix test (Proteus OX19, OX2 and OXK agglutination) has poor sensitivity and specificity and is obsolete, though it is still used in resource limited settings.
• Scrub typhus is also diagnosed by IgM ELISA and rapid immunochromatographic tests, with background positivity in endemic areas.
• Laboratory features: thrombocytopenia, leucopenia or normal white count, raised transaminases, hyponatraemia, raised creatine kinase and mild renal impairment.

Critical values and findings needing immediate action.

• Fever with rash, headache and tick or mite exposure: start doxycycline immediately on clinical suspicion, without waiting for serology, since delay beyond day 5 increases mortality (Rocky Mountain spotted fever is fatal in 20 to 30% untreated).
• Altered consciousness, seizures, shock, respiratory failure or oliguria: severe rickettsial disease, intensive care and doxycycline, often intravenous.
• Louse borne typhus outbreak: delousing, isolation, single dose doxycycline for contacts and public health notification.
• Pregnancy with suspected rickettsial infection: doxycycline is used for severe disease because the risk of untreated infection exceeds the risk of drug exposure.
• Fever with eschar in a returning traveller: urgent infectious disease review.

Interferences and pitfalls.

• Serology is retrospective and cannot direct initial treatment. Treatment starts on clinical suspicion.
• Antibodies are absent in the first week, and early doxycycline can blunt the antibody response.
• Cross-reactions occur with other rickettsiae and, for IgM, with other infections (dengue, leptospirosis, Q fever, Epstein-Barr virus, autoimmune disease).
• Titre cut offs vary by region, with high baseline prevalence in endemic populations, and a single titre is not reliable.
• Paired sera are rarely obtained, so many cases remain unconfirmed.
• PCR sensitivity from blood is moderate and eschar samples are better, so a negative result does not exclude infection.
• Differential diagnosis includes malaria, dengue, typhoid, leptospirosis, chikungunya, meningococcaemia, measles, secondary syphilis, drug rash and viral haemorrhagic fevers.
• Ehrlichiosis and anaplasmosis are diagnosed by PCR, blood smear (morulae in leucocytes) and paired serology.
• Chloramphenicol is the alternative when doxycycline cannot be used, and beta lactams, macrolides (other than for scrub typhus in pregnancy) and aminoglycosides are ineffective.

Special groups.

• Children: doxycycline is used in children of any age for rickettsial disease, as short courses cause negligible staining of teeth.
• Pregnancy: azithromycin is an option for mild scrub typhus and Mediterranean spotted fever.
• Older adults and alcoholism: higher mortality.
• Travellers: African tick bite fever is the commonest rickettsiosis in travellers returning from southern Africa, often with multiple eschars.
• Regions: scrub typhus is a leading cause of undifferentiated fever in South and Southeast Asia, and louse borne typhus occurs in highland Africa in poor, overcrowded conditions.

Follow up tests. Paired IFA IgG and IgM, PCR of blood, eschar swab or biopsy, full blood count, liver and renal function, creatine kinase, sodium, coagulation screen, malaria smear or rapid test, dengue NS1 and serology, leptospirosis MAT, blood cultures, CSF analysis in neurological disease, chest radiograph, and public health notification.