Under development

Diagnosis of rickettsioses in clinical settings is a challenge due to the large range of pathogens that cause similar symptoms, and further confounded by the lack of reliable tests to confirm diagnosis. Syndromic diagnosis can be problematic due to the non-specific symptoms of the diseases, which can often lead to delays in diagnosis. The most widely used methods for diagnosing rickettsial diseases comprise of culture, nucleic acid amplification (NAAT), and serological methods such as rapid diagnostic tests (RDTs), indirect immunofluorescence assays (IFA), and enzyme-linked immunosorbent assays (ELISA). Laboratory diagnosis of rickettsial diseases relies heavily on serological methods, however, antibody evidence of infection may not be present until the second week of illness onset; this often leads to negative results in the acute phase of infection. Antigen detection in skin, eschar, or tissue biopsies and other molecular methods can be advantageous for acute phase diagnosis. Although, clinical settings and resource limitations need to be taken into consideration.

References
https://www.scielo.sa.cr/scielo.php?script=sci_arttext&pid=S0001-60022013000400005

An Update on the Laboratory Diagnosis of Rickettsia spp. Infection

Antibiotics: Treatment, antimicrobial susceptibility and resistance for rickettsial infections

Rickettsiae are intracellular bacteria insensitive to beta-lactam drugs. The cause for this has long been imputed to an absence of peptidoglycan and lipopolysaccharide (LPS) in their cell walls. Protein-synthesis inhibiting antibiotics are therefore preferred and doxycycline is considered the treatment of choice for all rickettsial infections. The standard regimen is 100mg twice daily for seven days. To reach maintenance levels more rapidly, an initial dose of 200mg can be administered. Importantly, recent findings of peptidoglycan-like structure in Orientia tsutsugamushi , accompanied by in vitro susceptibility to some cell-wall targeting antibiotics, seem to question this view so it is possible that other antibiotics (e.g. phosphomycin) might be used as treatment in the future[1]. The following paragraphs focus on the treatment of scrub typhus, murine typhus and the spotted fevers. The evidence base is poor and there is a need for randomised clinical trials, especially amongst children and for severe disease. There have been no trials of adjunctive, non-antimicrobial, treatment of rickettsial diseases.

Scrub typhus
For the treatment of scrub typhus, some alternatives to doxycycline have good evidence of efficacy: chloramphenicol, tetracycline, azithromycin and rifampicin[2-4]. Other drugs that have been successfully used but for which there is limited evidence are telithromycin and roxithromycin, though in a small case series from northern Thailand the latter seemed to be less effective than doxycycline or chloramphenicol [5, 6]. Some authors have advocated the use of quinolones. However, the clinical response seems slower when compared to doxycycline and a quinolone resistance gene (GyrA) has been found in O. tsutsugamushi, so this class of antibiotics should be avoided [7-9]. As a general rule, doxycycline should be considered the first choice; if it is contraindicated, azithromycin can be given. In case of failure to respond to doxycycline or azithromycin, chloramphenicol or rifampicin can be used but the scrub typhus diagnosis and/or the quality of the antibiotics used should be questioned. The evidence base for treating severe scrub typhus is meagre with no randomized clinical trials; nasogastric or intravenous doxycycline or chloramphenicol are generally used. Intravenous azithromycin is also used but many countries do not have intravenous azithromycin or doxycycline available.

Doxycycline resistance in scrub typhus
Starting from the 1990s, patients with scrub typhus poorly responsive to doxycycline have been reported, especially from northern Thailand. From two cases, O. tsutsugamushi isolates were obtained. They showed reduced response to doxycycline in a rodent model and in cell culture compared to the reference strain[10, 11]. It is not clear if such findings are related to increased disease severity, true antibiotic resistance, high strain virulence, or a combination of those[12, 13]. Because the reports of doxycycline resistance and the evidence behind this theory are limited, if scrub typhus is suspected, doxycycline should remain the treatment of choice.

Murine typhus
The treatment of choice for murine typhus is doxycycline. Although azithromycin and chloramphenicol are occasionally recommended as alternatives, a recent RCT comparing doxycycline to azithromycin showed a worse performance of the latter[14]. Similarly, a retrospective study hinted that doxycycline might superior to both chloramphenicol and ciprofloxacin alone[15]. Doxycycline should therefore be preferred. Murine typhus The treatment of choice for murine typhus is doxycycline. Although azithromycin and chloramphenicol are occasionally recommended as alternatives, a recent RCT comparing doxycycline to azithromycin showed a worse performance of the latter[14]. Similarly, a retrospective study hinted that doxycycline might superior to both chloramphenicol and ciprofloxacin alone[15]. Doxycycline should therefore be preferred.

The spotted fevers
The spotted fever group rickettsiae comprise a vast and ever-increasing number of species but with meagre evidence for optimal therapy. Doxycycline is considered the treatment of choice for all of them. However, almost all research concerning treatment has focused on two species: Rickettsia rickettsii (causal agent of Rocky Mountain Spotted Fever, RMSF) and Rickettsia conorii (causal agent of Mediterranean Spotted Fever, MSF). For RMSF, antibiotics other than doxycycline have shown inferior results and, especially since the disease is severe compared to the other spotted fevers, should be avoided. In cases of severe allergy to doxycycline, the CDC proposes rapid desensitization procedures in an inpatient setting as an option for RMSF. If this is not possible, the only alternative treatment for RMSF, albeit with worse clinical outcomes, is chloramphenicol. For the treatment of MSF, clarithromycin, josamycin and azithromycin showed results comparable to doxycycline and the available evidence suggests that they be used as alternatives[16-18].

Contraindications to doxycycline treatment
Doxycycline is classified as a class D drug by the FDA (“potential benefits may warrant use of the drug in pregnant women despite potential risks”), so most authors recommend prescribing an alternative antibiotic in pregnancy. However, it must be borne in mind that the teratogenic potential of doxycycline has been inferred from the effects of tetracycline and there is hardly any evidence of direct teratogenic effects of doxycycline so the contraindication should be considered relative, especially in the presence of severe disease[19, 20]. Similarly, there have been concerns of tooth discoloration using tetracycline antibiotics in children less than 8 years old. When using doxycycline, recent reviews have not found any evidence of dental damage or any other serious toxicity so doxycycline should be considered a safe treatment option[21].

References
Atwal, S., et al., Evidence for a peptidoglycan-like structure in Orientia tsutsugamushi. Mol Microbiol, 2017. 105(3): p. 440-452.

Kim, Y.S., et al., A Comparative Trial of a Single Dose of Azithromycin versus Doxycycline for the Treatment of Mild Scrub Typhus. Clin Infect Dis, 2004. 39(November): p. 1329-35.

Diagnosis and identification of rickettsiae is challenging due to the non-specific clinical manifestations of the disease, combined with the shortcomings of currently available diagnostic modalities. Serological tests remain widely used in clinical settings and for epidemiological and sousveillance purposes. Molecular methods are often used for acute phase detection and species-specific identification.

Serology

Serological methods have been the mainstay for rickettsial disease diagnostics since the Weil-Felix test was developed. The Weil-Felix test is rarely used now given its poor sensitivity and specificity, and the advancements in serological assays. The current reference-standard for rickettsial diagnositcs is the indirect immunofluorescence assay (IFA). While the IFA has a high sensitivity and specificity, it is prone to subjectivity. An technique similar to the IFA is the immunoperoxidase assay (IIP), exhibiting comparable levels of sensitivity and specificity. The microimmunofluorescence assay (MIF) is a serological assay that allows for the detection of multiple rickettsial antigens, however it is often hindered by cross-reactivity. An alternative gold standard serological assay is the enzyme-linked immunosorbent assay (ELISA), which is widely used and better suited for low-resource settings.

Molecular

Molecular methods are more commonly used for direct detection of rickettsial species. Nucleic acid amplification tests (NAAT) and immunohistochemistry (IHC) assays allow for identification at the species level. NAAT has often been used to identify Rickettsia species directly from clinical samples. Many different probes and primers are used for the identification of rickettsial DNA, i.e. 16S rRNA gene, gltA (citrate synthase), ompA (outer membrane protein A), sca0, sca4, and sca5 (outer membrane proteins), HSP60, and 17-kDa gene. Conventional, nested, and real-time PCR techniques are all commonly used for detection of these genes. PCR demonstrates good sensitivity and specificity, and is more reliable during acute stages of illness.

Culture

Cell cultures are often performed for the isolation of rickettsiae. Culture methods are also often used as a confirmatory procedure in clinical settings.

References
An Update on the Laboratory Diagnosis of Rickettsia spp. Infection

Under development