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Epidemiology, Glinical Aspects, Laboratory Diagnosis and Treatment of Rickettsial Diseases in the Mediterranean Area during COVID-19 Pandemic: a Review of the Literature

Andrea De Vito1, Nicholas Geremia1, Sabrina Maria Mameli1, Vito Fiore1, Pier Andrea Serra2-3, Gaia Rocchitta2-3, Susanna Nuvoli4, Angela Spanu4, Renato Lobrano4, Antonio Cossu4, Sergio Babudieri1 and Giordano Madeddu1-3.

1 Unit of Infectious Diseases, Department of Medical, Surgical and Experimental Sciences, University of Sassari, Sassari, Italy.
2 Department of Medical, Surgical, and Experimental Sciences, Section of Pharmacology, University of Sassari, V. le S. Pietro 43/B, 07100, Sassari, Italy.
3 Mediterranean Center for Disease Control, University of Sassari, Sassari, Italy.
4 Department of Medical, Surgical and Experimental Sciences, University of Sassari, 07100 Sassari, Italy.

Correspondence to: Giordano Madeddu, Professor. Dipartimento di Scienze Mediche, Chirurgiche e Sperimentali, S.C. di Malattie Infettive, Università degli Studi di Sassari, Viale San Pietro 43 07100 Sassari – Italy. Tel: +393403781734. Fax: +39.079.217620. E-mail: giordano@uniss.it

Published: September 1, 2020
Received: May 19, 2020
Accepted: August 4, 2020
Mediterr J Hematol Infect Dis 2020, 12(1): e2020056 DOI 10.4084/MJHID.2020.056

This is an Open Access article distributed under the terms of the Creative Commons Attribution License
(
https://creativecommons.org/licenses/by-nc/4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

The purpose of the present review is to give an update regarding the classification, epidemiology, clinical manifestation, diagnoses, and treatment of the Rickettsial diseases present in the Mediterranean area.
We performed a comprehensive search, through electronic databases (Pubmed – MEDLINE) and search engines (Google Scholar), of peer-reviewed publications (articles, reviews, and books).
The availability of new diagnostic tools, including Polymerase Chain Reaction and nucleotide sequencing has significantly modified the classification of intracellular bacteria, including the order Rickettsiales with more and more new Rickettsia species recognized as human pathogens. Furthermore, emerging Rickettsia species have been found in several countries and are often associated with unique clinical pictures that may challenge the physician in the early detection of the diseases.
Rickettsial infections include a wide spectrum of clinical presentations ranging from a benign to a potentially life treating disease that requires prompt recognition and proper management. Recently, due to the spread of SARS-CoV-2 infection, the differential diagnosis with COVID-19 is of crucial importance. The correct understanding of the clinical features, diagnostic tools, and proper treatment can assist clinicians in the management of Rickettsioses in the Mediterranean area.



Introduction

Human rickettsial diseases are a variety of different clinical zoonoses caused by the genus Rickettsia and Orientia (order Rickettsiales, family Rickettsiaceae) that comprises a small (0.3–0.5 by 0.8–2.0 mm), obligately, intracellular and gram-negative bacilli, within α-proteobacteria.[1–3]
The development of Polymerase Chain Reaction (PCR) and nucleotide sequencing for the study of 16S rRNA has significantly modified the taxonomic classification of bacteria, in particular, intracellular bacteria. Some species sequencing for the study of 16S rRNA has demonstrated the phenotypic and genotypic differences between some microorganisms and the genus Rickettsia. For example, this is the case of Orientia tsutsugamushi that was reclassified from the genus Rickettsia into a new genus Orientia. The order Rickettsiales currently comprised the genera Anaplasma, Ehrlichia, Neorickettsia, Orientia, Rickettsia, and Wolbachia.[4,5]
Rickettsial infections are transmitted to human hosts mostly through arthropod bites or arthropod faeces that infect scratching lesions. The most frequent vectors responsible for the transmission are ticks, which also act as reservoirs, but some infections are associated with lice, fleas, or mites.[2]
Rickettsia species are split into four pathological groups or clades, based on their phenotypic characteristics, vector hosts and phylogenetic organization, that include the ancestral group, the spotted fever group (SFG), the typhus group, and the transitional group. The SFG is the largest group, and it is composed of the most common rickettsiae, such as Rickettsia aeschlimannii, Rickettsia africae, Rickettsia conorii subsp. caspia, Rickettsia conorii subsp. conorii, Rickettsia conorii subsp. indica, Rickettsia conorii subsp. israelensis, Rickettsia massiliae, Rickettsia monacensis, Rickettsia raoultii, Rickettsia rickettsii, Rickettsia rioja, Rickettsia sibirica subsp. mongolitimonae, Rickettsia sibirica subsp. sibirica, and the Rickettsia slovaca. The typhus group is composed of Rickettsia prowazekii and Rickettsia typhi.[6] The ancestral group includes Rickettsia bellii and Rickettsia canadensi.[3] The transitional group includes species of clinical importance, as Rickettsia akari Rickettsia australis and the Rickettsia helvetica, phylogenetically similar to SFG species such as Rickettsia felis.[7] Several authors discuss the validity of this group; in fact, these species have no relevant differences with other SFG species, except for their phylogenetic position.[8]
Due to their adaptation from a free-living to an obligate intracellular life in eukaryotic cells, Rickettsia species modified and reduced their genome size progressively.[9,10] Unexpected property of the rickettsial genome is the presence of plasmids, the first described in obligate intracellular bacteria.[11] This discovery suggests possible exchanges of genetic material by conjugation, a mechanism that was previously considered to be absent in obligate intracellular bacteria.[9,11]
The transmission of the infection depends on the group. SFG is transmitted by the bite of an infected tick; whereas, organisms of typhus group are transmitted through inoculation via infected louse or flea faeces (Rickettsiae prowazekii and Rickettsia typhi, respectively) through a bite, wound or mucous membranes. Once inoculated into the skin, organisms are phagocytized by dendritic cells and transported via lymphatics to local lymph nodes where they replicate. Subsequently, the bacteria spread in the bloodstream and disseminate to infect the endothelium of the microcirculation, where the Rickettsiae can infect vascular endothelial cells of the small and medium-sized blood vessels. The damage of the endothelium and the subsequent endothelial dysfunction is followed by alteration in coagulation and the cytokine network. The endpoint of this pathogenetic results in a reduction in circulating peripheral CD4 T lymphocytes and perivascular infiltration by CD4 and CD8 T lymphocytes, B cells, and macrophages, causing a vasculitis.[12–14]

Epidemiology

There are several pathological Rickettsia species in Europe, and in the last years, new species and subspecies have been implicated as human pathogens, and new rickettsial syndromes have been described.[15]
Mediterranean spotted fever (MSF) caused by Rickettsia conorii subsp. conorii is the most frequent rickettsiosis in Europe. It is endemic in southern Europe, but sporadic cases have been reported in all the continents.[15,16] The first cases were first described in Tunisia in 1909 by Conor and Buch. The brown dog tick, Rhipicephalus sanguineus, is the vector and the potential reservoir of Rickettsia conorii subsp. conorii in the Mediterranean area.[15,17] Most MSF cases occur in summer when climatic conditions seem to be an essential factor in increasing the aggressiveness of Rhipicephalus sanguineus ticks to bite humans.[15–18]
Rickettsia conorii subsp. israelensis is the agent of Israeli spotted fever (ISF), which was first reported in 1946 in the Haifa Bay area, Israel.[17–20] In Europe and the Mediterranean region, the brown dog tick, Rhipicephalus sanguineus, is recognized to be the vector of Rickettsia conorii subsp. israelensis.[21] The geographic distribution of the disease appears to be spread more widely in the Mediterranean countries than previously thought. Cases have been reported in Italy, Portugal, Tunisia, and Libya.[22–25]
Other Rickettsia conorii subspecies reported in the Mediterranean area are Rickettsia conorii subsp. caspia and Rickettsia conorii subsp. indica. The first one is the agent of Astrakhan fever, endemic in the Astrakhan region, adjacent regions of the Caspian Sea, and described in Rhipicephalus sanguineus ticks in Kosovo and southern France.[16]
Rickettsia sibirica subsp. mongolitimonae, the microorganism that cause of lymphangitis-associated rickettsiosis (LAR), was isolated for the first time in China, from Hyalomma asiaticum ticks collected in Mongolia in 1991.[16] Rickettsia sibirica subsp. mongolitimonae was detected in Hyalomma anatolicum excavatumt ticks in Greece and Cyprus; in Rhipicephalus pusillus ticks in France, Portugal, and Spain; and in Rhipicephalus bursa ticks in Spain.[26–30] The first human infection with Rickettsia sibirica subsp. mongolitimonae was reported in France in 1996.[31] Rickettsia sibirica subsp. mongolitimonae is implicated in human pathogen in different countries, as France, Spain, Turkey, and Egypt.[32–35]
Rickettsia slovaca and Rickettsia raoultii are associated with a syndrome characterized by scalp eschars and neck lymphadenopathy following tick bites (SENLAT). These microorganisms have been found in Dermacentor marginatus and Dermacentor reticulatus ticks in a vast majority of European countries.[36–42] After MSF, SENLAT is the most prevalent tick-borne rickettsiosis in Europe. It has been reported in different countries, including Hungary, Spain, France, Germany, Italy, Bulgaria, and Portugal.[43–48] SENLAT occurs most frequently from March to May and from September to November, which corresponds to the periods of most considerable activity of Dermacentor adult ticks in Europe.[47–49]
Rickettsia helvetica is transmitted by Ixodes ricinus, which is the primary vector and the natural reservoir. However, human infection is rare, and it has been documented only in Austria, Denmark, France, Italy, Sweden, Slovakia, and Switzerland.[16,50,51]
Other rare rickettsial pathological species in Europe and the Mediterranean area are Rickettsia massiliae, Rickettsia monacensis, Rickettsia aeschlimannii, and Rickettsia sibirica subsp. sibirica.

Clinical Manifestation

Rickettsiosis is a rare disease: the incidence is around 1 case per 100.000 people by year, but it has been increasing during the last years, probably due to better diagnostic techniques.[15]
In Europe, the most important diseases are three: Mediterranean Spotted Fever (MSF), Lymphangitis-associated rickettsioses (LAR), and scalp eschar and neck lymphadenopathy (SENLAT).[19] The other significant disease caused by Rickettsia rickettsii is the Rocky Mountain Spotted Fever (RMSF), but no cases have been reported in Europe to date.[19]
Apart from these three pathologies, there are other minor forms caused by different pathogens.
Mediterranean spotted fever. MSF, caused by Rickettsia conorii, is the most common rickettsial-disease in Europe, where the highest incidence is during summer.[52] Not all people who come into contact with this bacterium develop the disease. A Spanish study, indeed, shows that 4-8% of the population carry antibodies against Rickettsia but without a previous clinical history of MSF.[53]
The most common symptoms are fever (93-98%), myalgia (64-75%), headache (48-65%), and asthenia (27%). The maculopapular rash is present in 85-94% of the patients; the tache noir has been noticed in 58-64% of the patients. The classic triad, fever, maculopapular rash, and inoculation eschar, is present in 40-50% of the patients.[54–56]
In most cases, MSF is a self-limiting disease but sometimes could be life-threatening. It was estimated that about 5-10% of MSF cases could be severe. Cases of severe respiratory distress syndrome,[57] cardiovascular symptoms (coronary ectasia,[58] myocarditis,[59,60] vasculitis[61]) ocular symptoms,[62–65] neurological symptoms[66,67] (sensorineural hearing lost,[68,69] polyneuropathy,[70,71] encephalitis,[72,73] meningitis.[74,75]), acute pancreatitis,[76] splenic rupture,[77] acute renal failure,[73] hemophagocytic syndrome[78–80] and arthritis[81,82] have been reported.
The most frequent hematological and biochemical modifications are thrombocytopenia, leukocyte count abnormalities, elevated hepatic enzyme levels and an increase of c-reactive protein.[54,83]
Mortality was around 1-3%[84] before the antimicrobial drug era. Thus, it has been considered a benign illness. In some recent studies, MSF appears to be more severe than it has been thought. Mortality rates were 5.4% in France, 3.6% in Portugal, 3.2% in Algeria, 0.8% in Spain and 0.36% in Italy.[52,54,84–86]
Risk factors for severe MSF include advanced age, immunodeficiency, chronic alcoholism, G6PDH deficiency, diabetes, prior prescription of an inappropriate antimicrobial drug, or delay in treatment.[84,85]
Scalp eschar and neck lymphadenopathy after a tick bite. SENLAT[87] syndrome is also known as TIBOLA[88] (tick-borne lymphadenopathy) or DEBONEL[44] (Dermacentor-borne necrotic erythema and lymphadenopathy), and it is caused by Rickettsia slovaca and Rickettsia raoultii[19] but also by other bacteria such as Bartonella henselae.[87] This disease is developed mostly during spring and autumn.[49]
The clinical description of SENLAT includes asthenia, headache, painful adenopathies (especially to the neck's lymph nodes), and a painful scalp eschar surrounded by a perilesional erythematous halo. Low fever, rash, and face edema have also been reported less frequently.[45,87,89] No malignant or fatal cases have been described in the literature. After the therapy, alopecia could potentially last for several months, with persistent asthenia.[89]
Lymphangitis-associated rickettsioses. LAR is caused by Rickettsia sibirica subsp. mongolitimonae. Just a few cases have been reported in Europe. In particular, until 2013, only 24 cases have been reported in the Mediterranean area.[19]
The typical period of this disease is spring. Commons symptoms include fever, headache, an eschar (frequently more than one) on the site of inoculation, and lymphangitis, which starts from the eschar and reaches an enlarged lymph node. The difference between LAR and the other two diseases are the period of occurrence (spring), and the presence of lymphangitis and multiple eschars. [90]
Until now, no deaths have been reported in patients that have been infected by Rickettsia. However, some severe cases have been reported, in particular: a retinal vasculitis,[91] sepsis with disseminated intravascular coagulation,[92] myopericarditis[93] and a septic shock.[94,95]
Mediterranean spotted fever-like. Other Rickettsiae in Europe could infect humans; most of them cause a disease very similar to MSF. For example, Rickettsia conorii subsp. caspia causes an illness called "Astrakhan fever". This disease is typical of the Caspian Sea area, but some cases have also been reported in France.[96] Astrakhan fever diverges from MSF in the percentage of patients who present with an eschar (only 20%) and because it could cause thrombocytopenia and bleed.[97]
Another similar disease is the Israeli spotted Fever (ISF), caused by Rickettsia conorii subsp. israelensis. In Europe, this bacterium has been found only in Portugal and in Italy. The symptoms are quite similar to MSF except for the presence of gastrointestinal symptoms in half of the patients. The main difference is the malignity; indeed, the mortality is higher (more than 25%).[24,40,56,98,99]
Other Rickettsiae who could cause a MSF-like illness are Rickettsia monacensis,[100,101] Rickettsia massiliae,[102,103] Rickettsia aeschlimannii,[104,105] and Rickettsia helvetica which could be malignant.[51,106,107]
Differential Diagnosis with other infectious diseases including COVID-19. Clinically, the patients with MSF present the classic triad, fever, tache noir, and maculopapular rash in 40-50% of cases. In the absence of this typical clinical picture, the diagnosis could be challenging.
A small percentage of patients could present only the tache noir, which is generally pathognomonic of rickettsial diseases. However, clinical cases in which the tache noir was present in other zoonoses have been reported in the literature.[108–110]
The presence of fever without other signs is, probably, the most difficult challenge for clinicians because it is the expression of many diseases, both infective (bacterial, viral, fungal, and parasitic) and not infective. In these patients, a proper anamnesis, laboratory findings, and radiological features are mandatory to permit the correct diagnosis. Blood cultures should be collected at the fever peak to exclude a bacterial or fungal infection. Furthermore, in the area where SARS-CoV-2 is circulating in the population, the nasopharyngeal swab, together with acute phase serology, is recommended to rule it out. Indeed, the common symptom in patients with COronaVIrus Disease (COVID-19) is the fever.[111,112] The other symptoms that these two diseases have in common are headache, asthenia, and myalgia. The associations of dysgeusia, anosmia, and gastrointestinal symptoms could suggest the diagnosis of COVID-19.[113–115]
The maculopapular rash is an expression of several diseases.[116] In these cases, clinicians should pay attention to the distribution, the pattern, and the relationship between the localization at the start of it and other clinical signs, especially the fever. Although respiratory symptoms are the most frequent in COVID-19, skin involvement should always be considered. Galván Casas C et al.[117] described the most common cutaneous pattern, and Magro et al.[118] demonstrated how SARS-CoV-2 is associated with microvascular damage and thrombosis.
Moreover, different cutaneous vasculitis-like patterns correlated with COVID-19 or SARS-CoV-2 therapy have been described.[119,120]

Diagnosis

Nowadays, the majority of reference laboratories in developed countries can provide quick identification of rickettsial pathogens thanks to molecular and serological assays. In many cases, the diagnosis could be made by the clinical manifestation, but the laboratory tests are necessary at the support of it.
The choice of the most appropriate diagnostic technique requires consideration of the suspected pathogen, the timing of symptoms onset, and the type of sample available for testing.[121]
Serological tests remain essential diagnostic tools,[122] but Rickettsiae can be isolated from or detected in clinical specimens. The diagnostic tools available include serologic assays, molecular testing, cultures, immunochemistry, and Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF).[123]
The diagnostic technique could be divided into two groups:
1)    Diagnostic techniques used as routine.
2)    Less common diagnostic techniques.

Diagnostic techniques used as routine (Table 1)


Table 1 Table 1. Tests used as a routine.

Serologic tests:
Indirect immunofluorescence antibody assay (IFA) is a widely accepted serologic test for the detection of rickettsial infection.[124,125] It is considered the most sensitive and specific method among serological assays.[126] IFA consists of rickettsial antigens fixed on a slide and detected by specific antibodies present in the patient's serum, which can be identified by a fluorescein-labeled conjugate. Serum of patients with clinical manifestation of disease must be collected on the day of the admission and 2-4 weeks after illness onset.[127] IFA assays are highly sensitive at detecting antibodies after 2-3 weeks after illness onset, and their results are best interpreted if serum samples collected in acute and convalescent phases are tested at the same time.[128] Most laboratories test for IgG antibodies because IgM antibodies reactive with Rickettsia rickettsii are frequently detected in patients with no other supportive evidence of a recent rickettsial infection. Therefore, the detection of IgM during the acute phase should not be considered diagnostic for an ongoing illness as there could be cross-reactivity with other species and persistence of IgM beyond acute status.[129,130]
The enzyme-linked immunosorbent assay (ELISA) detects the binding of specific antibodies to antigens in a serum sample. When secondary anti-human antibodies conjugated with an enzyme are bound to antibodies from a serum sample and subjected to a substrate, an enzymatic reaction will be measurable in a positive specimen.[131] ELISA is sensitive, reproducible, and allows the differentiation of IgG and IgM antibodies. The results are more sensitive than IFA for the detection of low antibodies level, such as during late convalescence.[121] ELISA has the advantage, compared to IFA, of eliminating the subjective evaluation since the absorbance of the enzyme reaction is measured with a spectrophotometer. The inhibition ELISA has been used only for the diagnosis of scrub typhus and seems to be more sensitive than IFA in the early phase of the disease.[132]
Molecular diagnostic methods:
These assays are more appropriate than serology in the diagnosis of acute infection; a sample collected early at disease onset, before the development of antibodies, is more likely to produce a positive result in PCR assays. When antibody production has increased to detectable levels, bacteria are rarely found in the bloodstream or at the inoculation site. Furthermore, if antibiotic treatment has been initiated, the sensitivity of PCR assays decreases for the same reason.[133,134]
The most used method is nucleic acid amplification tests (NAATs), such as PCR, which has acquired increasing importance over the past few years. The quick response allows a prompt diagnosis without the need to wait for seroconversion or cell culture's growth time, which can take from 10 to 30 days.[123] Amplification of species-specific DNA by PCR provides a useful method for the differentiation between the several Rickettsia spp. and to gain knowledge about the genomic differences within the genus.[124]
The conventional PCR format, due to a large number of PCR products, is more prone to contamination. For this reason, a single-use primer PCR has been introduced.[135] Another molecular method is real-time PCR that offers the advantage of speed, reproducibility, quantitative capability, and reduced risk of contamination compared with conventional PCR assays.[136]
Several clinical samples are suitable for PCR amplification: skin biopsy, eschar, swab, or CSF. Peripheral blood and serum could also be used, but PCR on these samples has a lower sensitivity compared to skin samples or eschar collected on the bite site.[137,138]
PCR detection of Rickettsia rickettsii in the blood is possible. Still, its sensitivity is lower because of the small numbers of rickettsiae in the blood in the first stages of the disease.[139] For this reason, during the acute phase, it is better to use the SFG tissue specimen.[116,140] Doxycycline treatment decreases the sensitivity of PCR; therefore, obtaining blood before starting antibiotic therapy is recommended to minimize false-negative results.[116]

Less common diagnostic technique (Table 2)


Table 2 Table 2. Less common diagnostic technique.

Shell Vial:
This method requires a large number of bacteria and specific cell lines to proliferate, such as Vero E6 cells, human embryogenic lung fibroblast, and the promyelocytic HL-60 leukemia cell line (the most widely used cell line for growing A. phagocytophilum).[141] Specimens for cell cultures should be collected before starting antibiotic treatment and should not be frozen.[121] To identify the cultivated small intracellular Rickettsiae, the laboratories should label bacteria by fluorescent antibodies or staining with the Gimenez method.
The low success rate and the complexity of this method do not permit the routinely use of this methodic.[142]
Serologic methods:
The Weil-Felix test, based on the detection of immune-response to different Proteus antigens that cross respond with Rickettsia[125] should not be considered a first-line testing method anymore, even if it remains an option developing countries. It allows the detection of IgM antibodies 5-10 days after clinical manifestations.
Western blot assay (WBA) was demonstrated to be more sensitive than IFA for the detection of early antibodies in Rickettsia spp. Nevertheless, it is generally more expensive and technically challenging to perform than other serological methods.[143] Furthermore, Rickettsia cultures are required. For these reasons, its use is limited to only a few reference laboratories.[144]
The line, or dot, blot immunoassay, may be particularly useful for screening the many antigens that might be considered for patients with nonspecific or atypical clinical presentation. This test can be regarded as valuable only as a first-line test for the rapid diagnosis of acute cases in areas with high prevalence.[121]
The microagglutination test could be divided into two different methods, which included the indirect hemagglutination test and the Latex agglutination method. The first one is specific for the detection of IgG and IgM for all Rickettsiae.[143]
The Latex agglutination permitted the directed detection of the R. conorii, R. prowazekii, R. rickettsia, R. typhi, and infections. This method has a high sensitivity, but it is not routinely used for the high cost.[126,145]
Micro immunofluorescence (MIF) assay is similar to IFA except that wells are spotted with multiple rickettsial antigens for simultaneous detection. The negative aspect of this method is cross-reactivity, and its costs.[123]
Complement Fixation (CF) test permitted the identification detection of antibodies specific for rickettsiae. It is peculiar, but it has shown a reduced sensitivity, especially during the early stage of the disease. For this reason, it is only used for seroepidemiological studies.[126]
Indirect immunoperoxidase assay (IPA). The procedure is the same as IFA, but it used the peroxidase instead of fluorescein. It needs a specific instrument and trained personal. For this reason, it is not commonly used.[143]
Other tests:
Circulating endothelial cells (CECs) method allows the detection of R. conorii in circulating endothelial cells isolated from whole blood by using immunomagnetic beads coated with an endothelial cell-specific monoclonal antibody.[127] The sensitivity is about 50%, and it is not influenced by previous antibiotic treatment. Furthermore, the CECs level detected correlates with the severity of the infection, so it can be considered a prognostic indicator.[146]
Immunohistochemistry (IHC) permits the Rickettsia's detection directly from biopsy specimens, but it could only be used during the acute phase and only if there is a rash or tache noir.[123,143]
The most recent diagnostic tool is the matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF). This technique has been using with promise application for the Tick-borne infections inside the arthropods.[123] The future role of this new method could be applied to help the clinical decision. The identification of Rickettsiae inside the vector[147] or in the hemolymph[148] is showing great potential but remained a niche method.[123]
Biosensors emerging technology allows the fast detection of Rickettsia-induced immune response. For example, the OmpA antigen, an outer membrane protein present in the R. rickettsia, the agent responsible for the spotted fever, allows the detection of anti-OmpA human IgG. This is possible through an amperometric immune-sensor by using a synthetic peptide, obtained from the H6PGA4 R. rickettsia protein, homologous to OmpA.[149]


Figure 1 Figure 1. Samples that can be obtained from patients and the diagnostic techniques that could be performed on them. Green: tests commonly used; Yellow: tests used less frequently or used in the past; Grey: tests used only for research studies. 


Treatment

Rickettsiae spp. are obligate intracellular bacteria; therefore, the standard treatment is based on tetracyclines or chloramphenicol. The gold standard therapy is indeed represented by doxycycline 100 mg per os twice daily x 7 days in adults and 2.2 mg/kg of body weight per dose twice daily, orally or intravenously.[140,150] It has been demonstrated, in several studies, that doxycycline shortens the course of MSF and induces a rapid remission of symptoms. The problem is that tetracycline should be avoided in childhood, during pregnancy,[151,152] in patients who are allergic to it, and in those who have a G6PDH deficiency. An alternative to doxycycline is chloramphenicol. It should be administered at a dosage of 50 mg/Kg/day in four doses for seven days.[56] Since 2000, chloramphenicol was used only for patients suffering from allergy, those having adverse effects to doxycycline or if fever persisted for more than five days or in those that relapsed after the therapy with tetracycline. However, chloramphenicol should also be avoided during pregnancy (grey baby syndrome), and because of the various adverse effects (aplastic anemia, bone marrow suppression), it is not recommended in children. Furthermore, in a randomized study on 415 children, the chloramphenicol group had a longer hospitalization.[56]
For this reason, in pregnant women and children, the first choice is a macrolide. Different randomized studies have shown the macrolides' non-inferiority. In particular, Meloni et al., Bella et all and Cascio et al., in their randomized studies, have demonstrated the non-inferiority, respectively, of azithromycin, josamycin, and clarithromycin vs. doxycycline.[153–155] On the contrary, Munoz-Espin et al. have shown that erythromycin is less effective than doxycycline.[156]
Studies in vitro have tested the efficacy of fluoroquinolones against Rickettsiae spp., showing encouraging results.[150,157–159] Furthermore, randomized studies have shown that there is no difference between tetracycline and fluoroquinolones.[160–162] However, other studies found that fluoroquinolones are associated with increased MSF severity and the worst outcome.[163] Ciprofloxacin has been shown to have a deleterious effect on Rickettsia conorii-infected cells.[164]
Rickettsiae spp. showed to be susceptible also to rifampicin,[165] but in 1991 a small trial showed its inferiority in comparison with doxycycline.[166]
Even trimethoprim-sulfamethoxazole has been considered as a possible therapeutic option, but in vitro and in vivo studies have demonstrated that it is not active against Rickettsia spp.[150,167]
In the presence of unspecific symptoms during the spring to summer months, starting azithromycin seems reasonable given the ongoing COVID-19 epidemic in Mediterranean countries.[164]

References   

  1. Brenner, Krieg SJ. Bergey's Manual of Systematic Bacteriology - Vol 2: The Proteobacteria Part A - Introductory Essays. Springer-Verlag New York Inc. 2005;332. https://doi.org/10.1007/0-387-29298-5
  2. Raoult D, Roux V. Rickettsioses as paradigms of new or emerging infectious diseases. Clin Microbiol Rev. 1997;10(4):694-719. https://doi.org/10.1128/CMR.10.4.694 PMid:9336669 PMCid:PMC172941
  3. Stothard DR, Clark JB, Fuerst PA. Ancestral Divergence of Rickettsia bellii from the Spotted Fever and Typhus Groups of Rickettsia and Antiquity of the Genus Rickettsia. Int J Syst Bacteriol. 1994;44(4):798-804. https://doi.org/10.1099/00207713-44-4-798 PMid:7981106
  4. Tamura A, Ohashi N, Urakami H, Miyamura S. Classification of Rickettsia tsutsugamushi in a new genus, Orientia gen. nov., as Orientia tsutsugamushi comb. nov. Int J Syst Bacteriol. 1995;45(3):589-91. https://doi.org/10.1099/00207713-45-3-589 PMid:8590688
  5. Dumler JS, Barbet AF, Bekker CPJ, Dasch GA, Palmer GH, Ray SC, Rikihisa Y, Rurangirwa FR. Reorganization of genera in the families Rickettsiaceae and Anaplasmataceae in the order Rickettsiales: Unification of some species of Ehrlichia with Anaplasma, Cowdria with Ehrlichia and Ehrlichia with Neorickettsia, descriptions of six new species combi. Int J Syst Evol Microbiol. 2001;51(6):2145-65. https://doi.org/10.1099/00207713-51-6-2145 PMid:11760958
  6. Diop A, Raoult D, Fournier P-E. Paradoxical evolution of rickettsial genomes. Ticks Tick Borne Dis. 2019;10(2):462-9. https://doi.org/10.1016/j.ttbdis.2018.11.007 PMid:30448253
  7. Gillespie JJ, Williams K, Shukla M, Snyder EE, Nordberg EK, Ceraul SM, Dharmanolía C, Rainey D, Soneja J, Shallom JM, Vishnubhat ND, Wattam R, Purkayastha A, Czar M, Crasta O, Setubal JC, Azad AF, Sobral BS. Rickettsia phylogenomics: Unwinding the intricacies of obligate intracellular life. PLoS One. 2008;3(4):e2018. https://doi.org/10.1371/journal.pone.0002018 PMid:19194535 PMCid:PMC2635572
  8. Shpynov SN, Fournier P-E, Pozdnichenko NN, Gumenuk AS, Skiba AA. New approaches in the systematics of rickettsiae. New Microbes New Infect. 2018;23:93-102. https://doi.org/10.1016/j.nmni.2018.02.012 PMid:29692912 PMCid:PMC5913362
  9. Georgiades K, Raoult D. Genomes of the most dangerous epidemic bacteria have a virulence repertoire characterized by fewer genes but more toxin-antitoxin modules. PLoS One. 2011;6(3):e17962. https://doi.org/10.1371/journal.pone.0017962 PMid:21437250 PMCid:PMC3060909
  10. Merhej V, Royer-Carenzi M, Pontarotti P, Raoult D. Massive comparative genomic analysis reveals convergent evolution of specialized bacteria. Biol Direct. 2009;4(1):13. https://doi.org/10.1186/1745-6150-4-13 PMid:19361336 PMCid:PMC2688493
  11. Ogata H, Renesto P, Audic S, Robert C, Blanc G, Fournier P-E, Parinello H, Claverie J-M, Raoult D. The Genome Sequence of Rickettsia felis Identifies the First Putative Conjugative Plasmid in an Obligate Intracellular Parasite. Moran N, editor. PLoS Biol. 2005;3(8):e248. https://doi.org/10.1371/journal.pbio.0030248 PMid:15984913 PMCid:PMC1166351
  12. E. Rydkina, D.J. Silverman SKS. Similarities and Differences in Host Cell Signaling following Infection with Different Rickettsia Species. Ann N Y Acad Sci. 2005;1063(1):203-6. https://doi.org/10.1196/annals.1355.030 PMid:16481515
  13. Feng H, Popov VL, Yuoh G, Walker DH. Role of T lymphocyte subsets in immunity to spotted fever group Rickettsiae. J Immunol. 1997;158(11):5314-20.
  14. Mansueto P, Vitale G, Cascio A, Seidita A, Pepe I, Carroccio A, Di Rosa S, Rini GB, Cillari E, Walker DH. New insight into immunity and immunopathology of Rickettsial diseases. Clin Dev Immunol. 2012;2012:1-26. https://doi.org/10.1155/2012/967852 PMid:21912565 PMCid:PMC3170826
  15. Portillo A, Santibáñez S, García-Álvarez L, Palomar AM, Oteo JA. Rickettsioses in Europe. Microbes Infect. 2015;17(11-12):834-8. https://doi.org/10.1016/j.micinf.2015.09.009 PMid:26384814
  16. Parola P, Paddock CD, Raoult D. Tick-borne rickettsioses around the world: Emerging diseases challenging old concepts. Clin Microbiol Rev. 2005;18(4):719-56. https://doi.org/10.1128/CMR.18.4.719-756.2005 PMid:16223955 PMCid:PMC1265907
  17. Parola P, Socolovschi C, Raoult D. Deciphering the Relationships between Rickettsia conorii conorii and Rhipicephalus sanguineus in the Ecology and Epidemiology of Mediterranean Spotted Fever. Ann N Y Acad Sci. 2009;1166(1):49-54. https://doi.org/10.1111/j.1749-6632.2009.04518.x PMid:19538263  
  18. Parola P, Socolovschi C, Jeanjean L, Bitam I, Fournier PE, Sotto A, Labauge P, Raoult D. Warmer weather linked to tick attack and emergence of severe Rickettsioses. PLoS Negl Trop Dis. 2008;2(11):e338. https://doi.org/10.1371/journal.pntd.0000338 PMid:19015724 PMCid:PMC2581602
  19. Parola P, Paddock CD, Socolovschi C, Labruna MB, Mediannikov O, Kernif T, Abdad MY, Stenos J, Bitam I, Fournier PE, Raoult D. Update on tick-borne rickettsioses around the world: A geographic approach. Clin Microbiol Rev. 2013;26(4):657-702. https://doi.org/10.1128/CMR.00032-13 PMid:24092850 PMCid:PMC3811236  
  20. Sentausa E, El Karkouri K, Robert C, Raoult D, Fournier PE. Genome sequence of Rickettsia conorii subsp. israelensis, the agent of israeli spotted fever. J Bacteriol. 2012;194(18):5130-1. https://doi.org/10.1128/JB.01118-12 PMid:22933760 PMCid:PMC3430316  
  21. Zemtsova G, Killmaster LF, Mumcuoglu KY, Levin ML. Co-feeding as a route for transmission of Rickettsia conorii israelensis between Rhipicephalus sanguineus ticks. Exp Appl Acarol. 2010;52(4):383-92. https://doi.org/10.1007/s10493-010-9375-7 PMid:20589416
  22. Boillat N, Genton B, D'Acremont V, Raoult D, Greub G. Fatal case of Israeli spotted fever after Mediterranean cruise. Emerg Infect Dis. 2008;14(12):1944-6. https://doi.org/10.3201/eid1412.070641 PMid:19046528 PMCid:PMC2634608
  23. R. De Sousa, N. Ismail, S. Dória‐Nóbrega, P. Costa, T. Abreu, A. França, M. Amaro, P. Proença, P. Brito, J. Poças, T. Ramos, G. Cristina, G. Pombo, L. Vitorino, J. Torgal FBD. The Presence of Eschars, but Not Greater Severity, in Portuguese Patients Infected with Israeli Spotted Fever. Ann N Y Acad Sci. 2005;1063(1):197-202. https://doi.org/10.1196/annals.1355.032 PMid:16481514
  24. Giammanco GM, Vitale G, Mansueto S, Capra G, Caleca MP, Ammatuna P. Presence of Rickettsia conorii subsp. israelensis, the causative agent of Israeli spotted fever, in Sicily, Italy, ascertained in a retrospective study. J Clin Microbiol. 2005;43(12):6027-31. https://doi.org/10.1128/JCM.43.12.6027-6031.2005 PMid:16333093 PMCid:PMC1317185
  25. Znazen A, Hammami B, Lahiani D, Jemaa M Ben, Hammami A. Israeli spotted fever, Tunisia. Emerg Infect Dis. 2011;17(7):1328-30. https://doi.org/10.3201/eid1707.101648 PMid:21762610 PMCid:PMC3381377 
  26. Chochlakis D, Ioannou I, Sandalakis V, Dimitriou T, Kassinis N, Papadopoulos B, Tselentis Y, Psaroulaki A. Spotted Fever Group Rickettsiae in Ticks in Cyprus. Microb Ecol. 2012;63(2):314-23. https://doi.org/10.1007/s00248-011-9926-4 PMid:21833539  
  27. Toledo Á, Olmeda AS, Escudero R, Jado I, Valcárcel F, Casado-Nistal MA, Rodríguez-Vargas M, Gil H, Anda P. Tick-borne zoonotic bacteria in ticks collected from central Spain. Am J Trop Med Hyg. 2009;81(1):67-74. https://doi.org/10.4269/ajtmh.2009.81.67 PMid:19556569
  28. Edouard S, Parola P, Socolovschi C, Davoust B, la Scola B, Raoult D. Clustered cases of rickettsia sibirica mongolitimonae infection, France. Emerg Infect Dis. 2013;19(2):337-8. https://doi.org/10.3201/eid1902.120863 PMid:23460995 PMCid:PMC3559049
  29. Psaroulaki A, Germanakis A, Gikas A, Scoulica E, Tselentis Y. Simultaneous detection of "Rickettsia mongolotimonae" in a patient and in a tick in Greece. J Clin Microbiol. 2005;43(7):3558-9. https://doi.org/10.1128/JCM.43.7.3558-3559.2005 PMid:16000506 PMCid:PMC1169122  
  30. De Sousa R, Barata C, Vitorino L, Santos-Silva M, Carrapato C, Torgal J, Walker D, Bacellar F. Rickettsia sibirica isolation from a patient and detection in ticks, Portugal. Emerg Infect Dis. 2006;12(7):1103-8. https://doi.org/10.3201/eid1207.051494 PMid:16836827 PMCid:PMC3291052
  31. Raoult D, Brouqui P, Roux V. A new spotted-fever-group rickettsiosis [20]. Lancet. 1996;348(9024):412. https://doi.org/10.1016/S0140-6736(05)65037-4 
  32. Angelakis E, Richet H, Raoult D. Rickettsia sibirica mongolitimonae infection, France, 2010-2014. Emerg Infect Dis. 2016;22(5):880-2. https://doi.org/10.3201/eid2205.141989 PMid:27088367 PMCid:PMC4861502
  33. Kuscu F, Orkun O, Ulu A, Kurtaran B, Komur S, Seza Inal A, Erdogan D, Tasova Y, Aksu HSZ. Rickettsia sibirica mongolitimonae infection, Turkey, 2016. Emerg Infect Dis. 2017;23(7):1214-6. https://doi.org/10.3201/eid2307.170188 PMid:28628458 PMCid:PMC5512508  
  34. Ramos JM, Jado I, Padilla S, Masiá M, Anda P, Gutiérrez F. Human infection with rickettsia sibirica mongolitimonae, Spain, 2007-2011. Emerg Infect Dis. 2013;19(2):267-9. https://doi.org/10.3201/eid1902.111706 PMid:23343524 PMCid:PMC3559030
  35. Socolovschi C, Barbarot S, Lefebvre M, Parola P, Raoult D. Rickettsia sibirica mongolitimonae in traveler from Egypt. Emerg Infect Dis. 2010;16(9):1495-6. https://doi.org/10.3201/eid1609.100258 PMid:20735946 PMCid:PMC3294977  
  36. Fernandez-Soto P, Perez-Sanchez R, Alamo-Sanz R E-GA. Spotted Fever Group Rickettsiae in Ticks Feeding on Humans in Northwestern Spain: Is Rickettsia conorii Vanishing? Ann N Y Acad Sci. 2006;1078(1):331-3. https://doi.org/10.1196/annals.1374.063 PMid:17114733
  37. Špitalská E, Štefanidesová K, Kocianová E, Boldiš V. Rickettsia slovaca and Rickettsia raoultii in Dermacentor marginatus and Dermacentor reticulatus ticks from Slovak Republic. Exp Appl Acarol. 2012;57(2):189-97. https://doi.org/10.1007/s10493-012-9539-8 PMid:22392435
  38. Dobler G, Wölfel R. Typhus and other rickettsioses - Emerging infections in Germany. Dtsch Arztebl. 2009;106(20):348-54. https://doi.org/10.3238/arztebl.2009.0348 PMid:19547738 PMCid:PMC2689634
  39. Milhano N, Carvalho IL de, Alves AS, Arroube S, Soares J, Rodriguez P, Carolino M, Núncio MS, Piesman J, de Sousa R. Coinfections of Rickettsia slovaca and Rickettsia helvetica with Borrelia lusitaniae in ticks collected in a Safari Park, Portugal. Ticks Tick Borne Dis. 2010;1(4):172-7. https://doi.org/10.1016/j.ttbdis.2010.09.003 PMid:21771525
  40. Chisu V, Masala G, Foxi C, Socolovschi C, Raoult D, Parola P. Rickettsia conorii israelensis in Rhipicephalus sanguineus ticks, Sardinia, Italy. Ticks Tick Borne Dis. 2014;5(4):446-8. https://doi.org/10.1016/j.ttbdis.2014.02.003 PMid:24852264
  41. Kachrimanidou M, Souliou E, Pavlidou V, Antoniadis A, Papa A. First detection of Rickettsia slovaca in Greece. Exp Appl Acarol. 2010;50(1):93-6. https://doi.org/10.1007/s10493-009-9283-x PMid:19554462
  42. Chmielewski T, Podsiadly E, Karbowiak G, Tylewska-Wierzbanowska S. Rickettsia spp. In ticks, Poland. Emerg Infect Dis. 2009;15(3):486-8. https://doi.org/10.3201/eid1503.080711 PMid:19239772 PMCid:PMC2681112
  43. Lakos A. Tick-borne lymphadenopathy - a new rickettsial disease? Lancet. 1997;350(9083):1006. https://doi.org/10.1016/S0140-6736(05)64072-X
  44. Ibarra V, Oteo J., Portillo A, Santibanez S, Blanco J., Metola L, Eiros J., Perez-Martinez L SM. Rickettsia slovaca Infection: DEBONEL/TIBOLA. Ann N Y Acad Sci. 2006;1078(1):206-14. https://doi.org/10.1196/annals.1374.040 PMid:17114711
  45. Selmi M, Bertolotti L, Tomassone L, Mannelli A. Rickettsia slovaca in Dermacentor marginatus and tick-borne lymphadenopathy, Tuscany, Italy. Emerg Infect Dis. 2008;14(5):817-20. https://doi.org/10.3201/eid1405.070976 PMid:18439371 PMCid:PMC2600248
  46. Komitova R, Lakos A, Aleksandrov A, Christova I, Murdjeva M. A case of tick-transmitted lymphadenopathy in Bulgaria associated with Rickettsia slovaca. Scand J Infect Dis. 2003;35(3):213. https://doi.org/10.1080/0036554021000027016 PMid:12751725  
  47. Gouriet F, Rolain JM, Raoult D. Rickettsia slovaca infection, France. Emerg Infect Dis. 2006;12(3):521-3. https://doi.org/10.3201/eid1203.050911 PMid:16710981 PMCid:PMC3293430
  48. Rieg S, Schmoldt S, Theilacker C, de With K, Wölfel S, Kern W V, Dobler G. Tick-borne lymphadenopathy (TIBOLA) acquired in Southwestern Germany. BMC Infect Dis. 2011;11(1):167. https://doi.org/10.1186/1471-2334-11-167 PMid:21663601 PMCid:PMC3128054  
  49. Parola P, Rovery C, Rolain JM, Brouqui P, Davoust B, Raoult D. Rickettsia slovaca and R. raoultii in Tick-borne Rickettsioses. Emerg Infect Dis. 2009;15(7):1105-8. https://doi.org/10.3201/eid1507.081449 PMid:19624931 PMCid:PMC2744242
  50. Sekeyova Z, Subramanian G, Mediannikov O, Diaz MQ, Nyitray A, Blaskovicova H, Raoult D. Evaluation of clinical specimens for Rickettsia, Bartonella, Borrelia, Coxiella, Anaplasma, Franciscella and Diplorickettsia positivity using serological and molecular biology methods. FEMS Immunol Med Microbiol. 2012;64(1):82-91. https://doi.org/10.1111/j.1574-695X.2011.00907.x PMid:22098390  
  51. Nilsson K, Elfving K, Påhlson C. Rickettsia helvetica in patient with meningitis, Sweden, 2006. Emerg Infect Dis. 2010;16(3):490-2. https://doi.org/10.3201/eid1603.090184 PMid:20202426 PMCid:PMC3322002  
  52. Gomez-Barroso D, Vescio MF, Bella A, Ciervo A, Busani L, Rizzo C, Rezza G, Pezzotti P. Mediterranean spotted fever rickettsiosis in Italy, 2001-2015: Spatio-temporal distribution based on hospitalization records. Ticks Tick Borne Dis. 2019;10(1):43-50. https://doi.org/10.1016/j.ttbdis.2018.09.001 PMid:30197269
  53. Bernabeu-Wittel M, Del Toro MD, Nogueras MM, Muniain MA, Cardeñosa N, Márquez FJ, Segura F, Pachón J. Seroepidemiological study of Rickettsia felis, Rickettsia typhi, and Rickettsia conorii infection among the population of southern Spain. Eur J Clin Microbiol Infect Dis. 2006;25(6):375-81. https://doi.org/10.1007/s10096-006-0147-6 PMid:16767485  
  54. Crespo P, Seixas D, Marques N, Oliveira J, da Cunha S, Meliço-Silvestre A. Mediterranean spotted fever: case series of 24 years (1989-2012). Springerplus. 2015;4(1):272. https://doi.org/10.1186/s40064-015-1042-3 PMid:26090319 PMCid:PMC4469589
  55. Madeddu G, Fiore V, Mancini F, Caddeo A, Ciervo A, Babudieri S, Masala G, Bagella P, Nunnari G, Rezza G, Mura MS. Mediterranean spotted fever-like illness in Sardinia, Italy: a clinical and microbiological study. Infection. 2016;44(6):733-8. https://doi.org/10.1007/s15010-016-0921-z PMid:27380385  
  56. Colomba C, Saporito L, Polara VF, Rubino R, Titone L. Mediterranean spotted fever: clinical and laboratory characteristics of 415 Sicilian children. BMC Infect Dis. 2006;6(1):60. https://doi.org/10.1186/1471-2334-6-60 PMid:16553943 PMCid:PMC1435909  
  57. Dželalija B, Punda-Polić V, Medić A, Mraović B, Šimurina T. A case of Mediterranean spotted fever associated with severe respiratory distress syndrome. Microbes Infect. 2015;17(11-12):870-3. https://doi.org/10.1016/j.micinf.2015.08.012 PMid:26344605
  58. Cascio A, Maggio MC, Cardella F, Zangara V, Accomando S, Costa A, Iaria C, Mansueto P, Giordano S. Coronary involvement in Mediterranean spotted fever. New Microbiol. 2011;34(4):421-4.  
  59. Cascio A, Colomba C, Siracusa L, Trizzino M, Gioè C, Giammanco A. Myocarditis in Mediterranean spotted fever: a case report and a review of the literature. JMM Case Reports. 2016;3(4):e005039. https://doi.org/10.1099/jmmcr.0.005039 PMid:28348768 PMCid:PMC5330236  
  60. Ben Mansour N, Barakett N, Hajlaoui N, Haggui A, Filali T, Dahmen R, Fehri W, Haouala H. [Acute myocarditis complicating Mediterranean spotted fever. A case report]. Ann Cardiol Angeiol (Paris). 2014;63(1):55-7. https://doi.org/10.1016/j.ancard.2011.05.003 PMid:21664598
  61. Pennell D, Grundy HC JM. Mediterranean spotted fever presenting as acute leucocytoclastic vasculitis. Lancet. 1988;331(8599):1393-4. https://doi.org/10.1016/S0140-6736(88)92202-7  
  62. Leone S, De Marco M, Ghirga P, Nicastri E, Lazzari R, Narciso P. Retinopathy in Rickettsia conorii infection: Case report in an immunocompetent host. Infection. 2008;36(4):384-6. https://doi.org/10.1007/s15010-007-6291-9 PMid:18084718
  63. Alió J, Ruiz-Beltran R, Herrero-Herrero JI, Hernandez E, Guinaldo V, Millan A. Retinal Manifestations of Mediterranean Spotted Fever. Ophthalmologica. 1987;195(1):31-7. https://doi.org/10.1159/000309777 PMid:3658334
  64. Beselga D, Campos A, Castro M, Mendes S, Campos J, Neves A, Violante L, Sousa JPC. A rare case of retinal artery occlusion in the context of mediterranean spotted fever. Case Rep Ophthalmol. 2014;5(1):22-7. https://doi.org/10.1159/000358248 PMid:24596555 PMCid:PMC3934608
  65. Khairallah M, Ladjimi A, Chakroun M, Messaoud R, Yahia S Ben, Zaouali S, Romdhane F Ben, Bouzouaia N. Posterior segment manifestations of Rickettsia conorii infection. Ophthalmology. 2004;111(3):529-34. https://doi.org/10.1016/j.ophtha.2003.04.012 PMid:15019331
  66. Kularatne SAM, Weerakoon KGAD, Rajapakse RPVJ, Madagedara SC, Nanayakkara D, Premaratna R. A case series of spotted fever rickettsiosis with neurological manifestations in Sri Lanka. Int J Infect Dis. 2012;16(7):514-7. https://doi.org/10.1016/j.ijid.2012.02.016 PMid:22541336
  67. Tzavella K, Hatzizisis IS, Vakali A, Mandraveli K, Zioutas D, Alexiou-Daniel S. Severe case of Mediterranean spotted fever in Greece with predominantly neurological features. J Med Microbiol. 2006;55(3):341-3. https://doi.org/10.1099/jmm.0.46337-0 PMid:16476800
  68. Rossio R, Conalbi V, Castagna V, Recalcati S, Torri A, Coen M, Cassulini LR, Peyvandi F. Mediterranean spotted fever and hearing impairment: A rare complication. Int J Infect Dis. 2015;35:34-6. https://doi.org/10.1016/j.ijid.2015.04.005 PMid:25892247
  69. Tsiachris D, Deutsch M, Vassilopoulos D, Zafiropoulou R, Archimandritis AJ. Sensorineural hearing loss complicating severe rickettsial diseases: Report of two cases. J Infect. 2008;56(1):74-6. https://doi.org/10.1016/j.jinf.2007.10.002 PMid:18023483
  70. Popivanova N, Hristova D, Hadjipetrova E. Guillain‐Barré Polyneuropathy Associated with Mediterranean Spotted Fever: Case Report. Clin Infect Dis. 1998;27(6):1549. https://doi.org/10.1086/517751 PMid:9868689
  71. Caroleo S, Longo C, Pirritano D, Nisticò R, Valentino P, Iocco M, Santangelo E, Amantea B. A case of acute quadriplegia complicating Mediterranean spotted fever. Clin Neurol Neurosurg. 2007;109(5):463-5. https://doi.org/10.1016/j.clineuro.2007.02.007 PMid:17382465
  72. Duque V, Ventura C, Seixas D, Barai A, Mendonça N, Martins J, Da Cunha S, Meliço-Silvestre A. Mediterranean spotted fever and encephalitis: A case report and review of the literature. J Infect Chemother. 2012;18(1):105-8. https://doi.org/10.1007/s10156-011-0295-1 PMid:21879306  
  73. Aliaga L, Sánchez-Blázquez P, Rodríguez-Granger J, Sampedro A, Orozco M, Pastor J. Mediterranean spotted fever with encephalitis. J Med Microbiol. 2009;58(4):521-5. https://doi.org/10.1099/jmm.0.004465-0 PMid:19273650  
  74. Silpapojakul K, Ukkachoke C, Krisanapan S, Silpapojakul K. Rickettsial Meningitis and Encephalitis. Arch Intern Med. 1991;151(9):1753-7. https://doi.org/10.1001/archinte.1991.00400090051010 PMid:1888241  
  75. Salva I, De Sousa R, Gouveia C. Rickettsial meningitis. BMJ Case Rep. 2014;bcr2013203283. https://doi.org/10.1136/bcr-2013-203283 PMid:24614778 PMCid:PMC3962967
  76. Rombola F. Mediterranean spotted fever presenting as an acute pancreatitis. Acta Gastroenterol Belg. 2011;74(1):91-2.
  77. Schmulewitz L, Moumile K, De Serre NPM, Poirée S, Gouin E, Mechaï F, Cocard V, Mamzer-Bruneel MF, Abachin E, Berche P, Lortholary O, Lecuit M. Splenic rupture and malignant Mediterranean spotted fever. Emerg Infect Dis. 2008;14(6):995-7. https://doi.org/10.3201/eid1406.071295 PMid:18507929 PMCid:PMC2600289  
  78. Cascio A, Pernice LM, Barberi G, Delfino D, Biondo C, Beninati C, Mancuso G, Rodriguez-Morales AJ, Iaria C. Secondary hemophagocytic lymphohistiocytosis in zoonoses. A systematic review. Eur Rev Med Pharmacol Sci. 2012;16(10):1324-37.
  79. Premaratna R, Williams HSA, Chandrasena TGAN, Rajapakse RPVJ, Kularatna SAM, de Silva HJ. Unusual pancytopenia secondary to haemophagocytosis syndrome in rickettsioses. Trans R Soc Trop Med Hyg. 2009;103(9):961-3. https://doi.org/10.1016/j.trstmh.2009.04.003 PMid:19446860
  80. Cascio A, Giordano S, Dones P, Venezia S, Iaria C, Ziino O. Haemophagocytic syndrome and rickettsial diseases. J Med Microbiol. 2011;60(4):537-42. https://doi.org/10.1099/jmm.0.025833-0 PMid:21163825
  81. Pedro-Botet J, Auguet T, Pallás O, Gimeno JL. Arthritis in Mediterranean spotted fever. Infection. 1991;19(5):346-7. https://doi.org/10.1007/BF01645364 PMid:1800374
  82. Aragón A CA. Arthritis in mediterranean spotted fever. an immune complex mediated synovitis. Rheumatology. 1993;32(7):642-3. https://doi.org/10.1093/rheumatology/32.7.642 PMid:8339144
  83. Brouqui P, Bacellar F, Baranton G, Birtles RJ, Bjoërsdorff A, Blanco JR, Caruso G, Cinco M, Fournier PE, Francavilla E, Jensenius M, Kazar J, Laferl H, Lakos A, Lotric Furlan S, Maurin M, Oteo JA, Parola P, Perez-Eid C, Peter O, Postic D, Raoult D, Tellez A, Tselentis Y, Wilske B. Guidelines for the diagnosis of tick-borne bacterial diseases in Europe. Clin Microbiol Infect. 2004;10(12):1108-32. https://doi.org/10.1111/j.1469-0691.2004.01019.x PMid:15606643
  84. Rovery C, Brouqui P, Raoult D. Questions on Mediterranean spotted fever a century after its discovery. Emerg Infect Dis. 2008;14(9):1360-7. https://doi.org/10.3201/eid1409.071133 PMid:18760001 PMCid:PMC2603122
  85. de Sousa R, Nóbrega SD, Bacellar F, Torgal J. Mediterranean spotted fever in Portugal: risk factors for fatal outcome in 105 hospitalized patients. Ann N Y Acad Sci. 2003; https://doi.org/10.1111/j.1749-6632.2003.tb07378.x PMid:12860641
  86. Herrador Z, Fernandez-Martinez A, Gomez-Barroso D, León I, Vieira C, Muro A, Benito A. Mediterranean spotted fever in Spain, 1997-2014: Epidemiological situation based on hospitalization records. PLoS One. 2017;12(3):e0174745. https://doi.org/10.1371/journal.pone.0174745 PMid:28355307 PMCid:PMC5371374
  87. Angelakis E, Pulcini C, Waton J, Imbert P, Socolovschi C, Edouard S, Dellamonica P, Raoult D. Scalp Eschar and Neck Lymphadenopathy Caused by Bartonella henselae after Tick Bite . Clin Infect Dis. 2010;50(4):549-51. https://doi.org/10.1086/650172 PMid:20070235
  88. Lakos A. Tick-borne lymphadenopathy (TIBOLA). Wien Klin Wochenschr. 2002;114(13-14):648-54.  
  89. Raoult D, Lakos A, Fenollar F, Beytout J, Brouqui P, Fournier P. Spotless Rickettsiosis Caused by Rickettsia slovaca and Associated with Dermacentor Ticks . Clin Infect Dis. 2002;34(10):1331-6. https://doi.org/10.1086/340100 PMid:11981728  
  90. Fournier P-E, Gouriet F, Brouqui P, Lucht F, Raoult D. Lymphangitis-Associated Rickettsiosis, a New Rickettsiosis Caused by Rickettsia sibirica mongolotimonae: Seven New Cases and Review of the Literature. Clin Infect Dis. 2005;40(10):1435-44. https://doi.org/10.1086/429625 PMid:15844066
  91. Caron J, Rolain JM, Mura F, Guillot B, Raoult D, Bessis D. Rickettsia sibirica subsp. mongolitimonae infection and retinal vasculitis. Emerg Infect Dis. 2008;14(4):683-4. https://doi.org/10.3201/eid1404.070859 PMid:18394301 PMCid:PMC2570939
  92. Gaillard, D.E., Socolovschi, C., Fourcade, C., Lavigne, J., Raoult, D., & Sotto A. A case of severe sepsis with disseminated intravascular coagulation during Rickettsia sibirica mongolitimonae infection. Med Mal Infect. 2015;45(1-2):56-9. https://doi.org/10.1016/j.medmal.2014.10.005 PMid:25455075
  93. Revilla-Martí P, Cecilio-Irazola Á, Gayán-Ordás J, Sanjoaquín-Conde I, Linares-Vicente JA, Oteo JA. Acute myopericarditis associated with tickborne Rickettsia sibirica mongolitimonae. Emerg Infect Dis. 2017;23(12):2091-3. https://doi.org/10.3201/eid2312.170293 PMid:29148392 PMCid:PMC5708254
  94. Carlo P Di, Trizzino M, Giarratano A, Giammanco A, Montalto F, Raineri M, Dones F, Bonura C. Real-time PCR for early diagnosis of Rickettsia conorii and prompt management in patients with septic shock and multiple organ failure: two case reports. INFECT DIS TROP MED. 2015;1(2):e100.
  95. Ibarra V, Portillo A, Palomar AM, Sanz MM, Metola L, Blanco JR, Oteo JA. Septic shock in a patient infected with Rickettsia sibirica mongolitimonae, Spain. Clin Microbiol Infect. 2012;18(8):W283-E285. https://doi.org/10.1111/j.1469-0691.2012.03887.x PMid:22548679
  96. Renvoisé A, Delaunay P, Blanchouin E, Cannavo I, Cua E, Socolovschi C, Parola P, Raoult D. Urban family cluster of spotted fever rickettsiosis linked to Rhipicephalus sanguineus infected with Rickettsia conorii subsp. caspia and Rickettsia massiliae. Ticks Tick Borne Dis. 2012;3(5-6):389-92. https://doi.org/10.1016/j.ttbdis.2012.10.008 PMid:23140893
  97. Tarasevich I V., Makarova VA, Fetisova NF, Stepanov A V., Miskarova ED, Raoult D. Studies of a "new" rickettsiosis "Astrakhan" spotted fever. Eur J Epidemiol. 1991;7(3):294-8. https://doi.org/10.1007/BF00145681 PMid:1884783  
  98. Amaro M, Bacellar F, França A. Report of eight cases of fatal and severe Mediterranean spotted fever in Portugal. Ann N Y Acad Sci. 2003;990:331-43. https://doi.org/10.1111/j.1749-6632.2003.tb07384.x PMid:12860647
  99. Colomba C, Trizzino M, Giammanco A, Bonura C, Di Bona D, Tolomeo M, Cascio A. Israeli Spotted Fever in Sicily. Description of two cases and minireview. Int J Infect Dis. 2017;61:7-12. https://doi.org/10.1016/j.ijid.2017.04.003 PMid:28408252
  100. Madeddu G, Mancini F, Caddeo A, Ciervo A, Babudieri S, Maida I, Fiori ML, Rezza G, Mura MS. Rickettsia monacensis as Cause of Mediterranean Spotted Fever-like Illness, Italy. Emerg Infect Dis. 2012;18(4):702-4. https://doi.org/10.3201/eid1804.111583 PMid:22469314 PMCid:PMC3309684
  101. Jado I, Oteo JA, Aldámiz M, Gil H, Escudero R, Ibarra V, Portu J, Portillo A, Lezaun MJ, García-Amil C, Rodríguez-Moreno I, Anda P. Rickettsia monacensis and human disease, Spain. Emerg Infect Dis. 2007;13(9):1405-7. https://doi.org/10.3201/eid1309.060186 PMid:18252123 PMCid:PMC2857266
  102. García-García JC, Portillo A, Núñez MJ, Santibáñez S, Castro B, Oteo JA. Case report: A patient from Argentina infected with Rickettsia massiliae. Am J Trop Med Hyg. 2010;82(4):691-2. https://doi.org/10.4269/ajtmh.2010.09-0662 PMid:20348520 PMCid:PMC2844561  
  103. Fernández De Mera IG, Zivkovic Z, Bolaños M, Carranza C, Pérez-Arellano JL, Gutiérrez C, De La Fuente J. Rickettsia massiliae in the Canary Islands. Emerg Infect Dis. 2009;15(11):1869-70. https://doi.org/10.3201/eid1511.090681 PMid:22531111 PMCid:PMC2857243  
  104. A. Tosoni, A. Mirijello, A. Ciervo, F. Mancini, G. Rezza, F. Damiano, R. Cauda, A. Gasbarrini, G. Addolorato, On Behalf Of The Internal, Medicine Sepsis Study Group. Human Rickettsia aeschlimannii infection: first case with acute hepatitis and review of the literature. Eur Rev Med Pharmacol Sci. 2016;20:2630-3.
  105. Mura A, Masala G, Tola S, Satta G, Fois F, Piras P, Rolain JM, Raoult D, Parola P. First direct detection of rickettsial pathogens and a new rickettsia, "Candidatus Rickettsia barbariae", in ticks from Sardinia, Italy. Clin Microbiol Infect. 2008;14(11):1028-33. https://doi.org/10.1111/j.1469-0691.2008.02082.x PMid:19040474  
  106. Nilsson K. Septicaemia with Rickettsia helvetica in a patient with acute febrile illness, rash and myasthenia. J Infect. 2009;58(1):79-82. https://doi.org/10.1016/j.jinf.2008.06.005 PMid:18649945
  107. Fournier PE, Grunnenberger F, Jaulhac B, Gastinger G, Raoult D. Evidence of Rickettsia helvetica infection in humans, eastern France. Emerg Infect Dis. 2000;6(4):389-92. https://doi.org/10.3201/eid0604.000412 PMid:10905974 PMCid:PMC2640907
  108. Fiore V, Mancini F, Ciervo A, Bagella P, Peruzzu F, Nunnari G, Deiana GA, Rezza G, Babudieri S, Madeddu G. Tache Noire in a Patient with Acute Q Fever. Med Princ Pract. 2018;27(1):92-4. https://doi.org/10.1159/000486573 PMid:29298443 PMCid:PMC5968302  
  109. Yang J, Liu Z, Niu Q, Liu J, Han R, Guan G, Hassan MA, Liu G, Luo J, Yin H. A novel zoonotic Anaplasma species is prevalent in small ruminants: potential public health implications. Parasites and Vectors. 2017;10(1). https://doi.org/10.1186/s13071-017-2182-9 PMid:28558749 PMCid:PMC5450374
  110. Eldin C, Mélenotte C, Mediannikov O, Ghigo E, Million M, Edouard S, Mege JL, Maurin M, Raoult D. From Q fever to Coxiella burnetii infection: A paradigm change. Clin Microbiol Rev. 2017;30(1):115-90. https://doi.org/10.1128/CMR.00045-16 PMid:27856520 PMCid:PMC5217791
  111. Siordia JA. Epidemiology and clinical features of COVID-19: A review of current literature. J Clin Virol. 2020;127:104357. https://doi.org/10.1016/j.jcv.2020.104357 PMid:32305884 PMCid:PMC7195311  
  112. Gao Y, Yan L, Huang Y, Liu F, Zhao Y, Cao L, Wang T, Sun Q, Ming Z, Zhang L, Ge J, Zheng L, Zhang Y, Wang H, Zhu Y, Zhu C, Hu T, Hua T, Zhang B, Yang X, Li J, Yang H, Liu Z, Xu W, Guddat LW, Wang Q, Lou Z, Rao Z. Structure of the RNA-dependent RNA polymerase from COVID-19 virus. Science (80- ). 2020;368(6492):779-82. https://doi.org/10.1126/science.abb7498 PMid:32277040 PMCid:PMC7164392
  113. Guan W, Ni Z, Hu Y, Liang W, Ou C, He J, Liu L, Shan H, Lei C, Hui DSC, Du B, Li L, Zeng G, Yuen K-Y, Chen R, Tang C, Wang T, Chen P, Xiang J, Li S, Wang J, Liang Z, Peng Y, Wei L, Liu Y, Hu Y, Peng P, Wang J, Liu J, Chen Z, Li G, Zheng Z, Qiu S, Luo J, Ye C, Zhu S, Zhong N. Clinical Characteristics of Coronavirus Disease 2019 in China. N Engl J Med. 2020;382(18):1708-20. https://doi.org/10.1056/NEJMoa2002032 PMid:32109013 PMCid:PMC7092819
  114. Vaira LA, Deiana G, Fois AG, Pirina P, Madeddu G, De Vito A, Babudieri S, Petrocelli M, Serra A, Bussu F, Ligas E, Salzano G, Riu G De. Objective evaluation of anosmia and ageusia in COVID‐19 patients: a single‐center experience on 72 cases. Head Neck. 2020;42(6):1252-8. https://doi.org/10.1002/hed.26204 PMid:32342566 PMCid:PMC7267244
  115. Vaira LA, Hopkins C, Salzano G, Petrocelli M, Melis A, Cucurullo M, Ferrari M, Gagliardini L, Pipolo C, Deiana G, Fiore V, De Vito A, Turra N, Canu S, Maglio A, Serra A, Bussu F, Madeddu G, Babudieri S, Giuseppe Fois A, Pirina P, Salzano FA, De Riu P, Biglioli F, De Riu G. Olfactory and gustatory function impairment in COVID‐19 patients: Italian objective multicenter‐study. Head Neck. 2020;hed.26269. https://doi.org/10.1002/hed.26269 PMid:32437022 PMCid:PMC7280583
  116. Chapman AS, Bakken JS, Folk SM, Paddock CD, Bloch KC, Krusell A, Sexton DJ, Buckingham SC, Marshall GS, Storch GA, Dasch GA, McQuiston JH, Swerdlow DL, Dumler SJ, Nicholson WL, Walker DH, Eremeeva ME, Ohl CA, Tickborne Rickettsial Diseases Working Group, CDC. Diagnosis and management of tick-borne rickettsial diseases: Rocky Mountain spotted fever, ehrlichioses, and anaplasmosis--United States: a practical guide for physicians and other health-care and public health professionals. MMWR Recomm reports Morb Mortal Wkly report Recomm reports. 2006;
  117. Galvan Casas C, Catala A, Carretero Hernandez G, Rodriguez-Jimenez P, Fernandez Nieto D, Rodriguez-Villa Lario A, Navarro Fernandez I, Ruiz-Villaverde R, Falkenhain D, Llamas Velasco M, Garcia-Gavin J, Baniandres O, Gonzalez-Cruz C, Morillas-Lahuerta V, Cubiro X, Figueras Nart I, Selda-Enriquez G, Romani J, Fusta-Novell X, Melian-Olivera A, Roncero Riesco M, Burgos-Blasco P, Sola Ortigosa J, Feito Rodriguez M, Garcia-Doval I, Galván Casas C, Català A, Carretero Hernández G, Rodríguez‐Jiménez P, Fernández Nieto D, Rodríguez‐Villa Lario A, Navarro Fernández I, Ruiz‐Villaverde R, Falkenhain D, Llamas Velasco M, García‐Gavín J, Baniandrés O, González‐Cruz C, Morillas‐Lahuerta V, Cubiró X, Figueras Nart I, Selda‐Enriquez G, Romaní J, Fustà‐Novell X, Melian‐Olivera A, Roncero Riesco M, Burgos‐Blasco P, Sola Ortigosa J, Feito Rodriguez M, García‐Doval I. Classification of the cutaneous manifestations of COVID-19: a rapid prospective nationwide consensus study in Spain with 375 cases. Br J Dermatol. 2020; https://doi.org/10.1111/bjd.19163 PMid:32348545 PMCid:PMC7267236  
  118. Magro C, Mulvey JJ, Berlin D, Nuovo G, Salvatore S, Harp J, Baxter-stoltzfus A. Complement associated microvascular injury and thrombosis in the pathogenesis of severe COVID-19 infection: A report of five cases. Transl Res. 2020;220:1-13. https://doi.org/10.1016/j.trsl.2020.04.007 PMid:32299776 PMCid:PMC7158248
  119. Li H, Zhou Y, Zhang M, Wang H, Zhao Q, Liu J. Updated approaches against SARS-CoV-2. Antimicrob Agents Chemother. 2020;64(6):e00483-20. https://doi.org/10.1128/AAC.00483-20 PMid:32205349 PMCid:PMC7269512
  120. Bouaziz J, Duong T, Jachiet M, Velter C, Lestang P, Cassius C, Arsouze A, Domergue Than Trong E, Bagot M, Begon E, Sulimovic L, Rybojad M. Vascular skin symptoms in COVID-19: a french observational study. J Eur Acad Dermatology Venereol. 2020;Accepted Author Manuscript. https://doi.org/10.1111/jdv.16544 PMid:32339344 PMCid:PMC7267662  
  121. La Scola B, Raoult D. Laboratory diagnosis of Rickettsioses: Current approaches to diagnosis of old and new Rickettsial diseases. J Clin Microbiol. 1997;35(11):2715-27. https://doi.org/10.1128/JCM.35.11.2715-2727.1997 PMid:9350721 PMCid:PMC230049
  122. Kováčová E, Kazár J. Rickettsial diseases and their serological diagnosis. Clin Lab. 2000;46(5-6):239-45.
  123. Abdad MY, Abdallah RA, Fournier PE, Stenos J, Vasoo S. A concise review of the epidemiology and diagnostics of rickettsioses: Rickettsia and orientia spp. J Clin Microbiol. 2018;56(8):e01728-17. https://doi.org/10.1128/JCM.01728-17 PMid:29769278 PMCid:PMC6062794
  124. Bouyer DH, Walker DH. Rickettsia and Orientia. In: Manual of Clinical Microbiology, 11th Edition. American Society of Microbiology; 2015. p. 1122-34. https://doi.org/10.1128/9781555817381.ch64
  125. Dumler JS, Reller ME. Ehrlichia, Anaplasma, and Related Intracellular Bacteria. In: Manual of Clinical Microbiology, 11th Edition. American Society of Microbiology; 2015. p. 1135-49. https://doi.org/10.1128/9781555817381.ch65
  126. Newhouse VF, Shepard CC, Redus MD, Tzianabos T, McDade JE. A comparison of the complement fixation, indirect fluorescent antibody, and microagglutination tests for the serological diagnosis of rickettsial diseases. Am J Trop Med Hyg. 1979;28(2):387-95. https://doi.org/10.4269/ajtmh.1979.28.387 PMid:378003
  127. Drancourt M, George F, Brouqui P, Sampol J, Raoult D. Diagnosis of mediterranean spotted fever by indirect immunofluorescence of rickettsia conorii in circulating endothelial cells isolated with monoclonal antibody-coated immunomagnetic beads. J Infect Dis. 1992;166(3):660-3. https://doi.org/10.1093/infdis/166.3.660 PMid:1500755
  128. Clements ML, Dumler JS, Fiset P, Wisseman CL, Snyder MJ, Levine MM. Serodiagnosis of Rocky Mountain spotted fever: Comparison of IgM and IgG enzyme-linked immunosorbent assays and indirect fluorescent antibody test. J Infect Dis. 1983;148(5):876-80.  https://doi.org/10.1093/infdis/148.5.876 PMid:6415180
  129. Walls JJ, Aguero-Rosenfeld M, Bakken JS, Goodman JL, Hossain D, Johnson RC, Dumler JS. Inter- and intralaboratory comparison of Ehrlichia equi and human granulocytic ehrlichiosis (HGE) agent strains for serodiagnosis of HGE by the immunofluorescent-antibody test. J Clin Microbiol. 1999;37(9):2968-73. https://doi.org/10.1128/JCM.37.9.2968-2973.1999 PMid:10449483 PMCid:PMC85424
  130. Brouqui P, Salvo E, Dumler JS, Raoult D. Diagnosis of granulocytic ehrlichiosis in humans by immunofluorescence assay. Clin Diagn Lab Immunol. 2001;8(1):199-202. https://doi.org/10.1128/CDLI.8.1.199-202.2001 PMid:11139221 PMCid:PMC96036
  131. Engvall E, Perlmann P. Enzyme-linked immunosorbent assay (ELISA) quantitative assay of immunoglobulin G. Immunochemistry. 1971;8(9):871-4. https://doi.org/10.1016/0019-2791(71)90454-X
  132. Furuya Y, Yamamoto S, Otu M, Yoshida Y, Ohashi N, Murata M, Kawabata N, Tamura A, Kawamura A. Use of monoclonal antibodies against Rickettsia tsutsugamushi Kawasaki for serodiagnosis by enzyme-linked immunosorbent assay. J Clin Microbiol. 1991;29(2):340-5. https://doi.org/10.1128/JCM.29.2.340-345.1991 PMid:1706729 PMCid:PMC269764
  133. Angelakis E, Munasinghe A, Yaddehige I, Liyanapathirana V, Thevanesam V, Bregliano A, Socolovschi C, Edouard S, Fournier PE, Raoult D, Parola P. Short report: Detection of rickettsioses and Q fever in Sri Lanka. Am J Trop Med Hyg. 2012;86(4):711-2. https://doi.org/10.4269/ajtmh.2012.11-0424 PMid:22492158 PMCid:PMC3403782  
  134. Edouard S, Bhengsri S, Dowell SF, Watt G, Parola P, Raoult D. Two human cases of Rickettsia felis infection, Thailand. Emerg Infect Dis. 2014;20(10):1780-1. https://doi.org/10.3201/eid2010.140905 PMid:25272251 PMCid:PMC4193185  
  135. Fournier PE, Raoult D. Suicide PCR on skin biopsy specimens for diagnosis of Rickettsioses. J Clin Microbiol. 2004;42(8):3428-34. https://doi.org/10.1128/JCM.42.8.3428-3434.2004 PMid:15297478 PMCid:PMC497613
  136. 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;29(5):433-9. https://doi.org/10.1097/QCO.0000000000000298 PMid:27429138 PMCid:PMC5029442
  137. Znazen A, Sellami H, Elleuch E, Hattab Z, Ben Sassi L, Khrouf F, Dammak H, Letaief A, Ben Jemaa M, Hammami A. Comparison of Two Quantitative Real Time PCR Assays for Rickettsia Detection in Patients from Tunisia. PLoS Negl Trop Dis. 2015;9(2):e0003487. https://doi.org/10.1371/journal.pntd.0003487 PMid:25706392 PMCid:PMC4338037  
  138. Kantsø B, Svendsen CB, Jørgensen CS, Krogfelt KA. Evaluation of serological tests for the diagnosis of rickettsiosis in Denmark. J Microbiol Methods. 2009;76(3):285-8. https://doi.org/10.1016/j.mimet.2008.12.012 PMid:19162092
  139. Kaplowitz LG, Lange J V., Fischer JJ, Walker DH. Correlation of Rickettsial Titers, Circulating Endotoxin, and Clinical Features in Rocky Mountain Spotted Fever. Arch Intern Med. 1983;143(6):1149-51. https://doi.org/10.1001/archinte.1983.00350060073012 PMid:6407418  
  140. Biggs HM, Behravesh CB, Bradley KK, Dahlgren FS, Drexler NA, Dumler JS, Folk SM, Kato CY, Lash RR, Levin ML, Massung RF, Nadelman RB, Nicholson WL, Paddock CD, Pritt BS, Traeger MS. Diagnosis and management of tick-borne rickettsial diseases: Rocky mountain spotted fever and other spotted fever group rickettsioses, ehrlichioses, and anaplasmosis - United States a practical guide for health care and public health professionals. MMWR Recomm Reports. 2016;65(2):1-44. https://doi.org/10.15585/mmwr.rr6502a1 PMid:27172113
  141. Paddock CD. Perspectivas sobre el diagnóstico de laboratorio de enfermedades rickettsiales en el siglo 21. Acta Médica Costarrciense. 2013;
  142. Wallménius K. Studies of Spotted Fever Rickettsia - Distribution, Detection, Diagnosis and Clinical Context. 2016. 1-78 p.  
  143. Putli Bai PS. Laboratory diagnosis of rickettsial infections. Pediatr Infect Dis. 2015;7(3):85-7. https://doi.org/10.1016/j.pid.2015.12.002
  144. Fenollar F, Fournier PE, Raoult D. Diagnostic strategy of rickettsioses and ehrlichioses. In: Rickettsial Diseases. CRC Press; 2007. p. 315-30. https://doi.org/10.3109/9781420019971.023  
  145. Anacker RL, Philip RN, Thomas LA, Casper EA. Indirect hemagglutination test for detection of antibody to Rickettsia rickettsii in sera from humans and common laboratory animals. J Clin Microbiol. 1979;10(5):677-84. https://doi.org/10.1128/JCM.10.5.677-684.1979 PMid:120875 PMCid:PMC273246
  146. George F, Brouqui P, Boffa MC, Mutin M, Drancourt M, Brisson C, Raoult D, Sampol J. Demonstration of Rickettsia conorii-induced endothelial injury in vivo by measuring circulating endothelial cells, thrombomodulin, and von Willebrand factor in patients with Mediterranean spotted fever. Blood. 1993;82(7):2109-16. https://doi.org/10.1182/blood.V82.7.2109.2109 PMid:7691249
  147. Yssouf A, Almeras L, Terras J, Socolovschi C, Raoult D, Parola P. Detection of Rickettsia spp in Ticks by MALDI-TOF MS. Walker DH, editor. PLoS Negl Trop Dis. 2015;9(2):e0003473. https://doi.org/10.1371/journal.pntd.0003473 PMid:25659152 PMCid:PMC4319929  
  148. Yssouf A, Almeras L, Berenger JM, Laroche M, Raoult D, Parola P. Identification of tick species and disseminate pathogen using hemolymph by MALDI-TOF MS. Ticks Tick Borne Dis. 2015;6(5):579-86. https://doi.org/10.1016/j.ttbdis.2015.04.013 PMid:26051210
  149. Prado IC, Chino META, dos Santos AL, Souza ALA, Pinho LG, Lemos ERS, De-Simone SG. Development of an electrochemical immunosensor for the diagnostic testing of spotted fever using synthetic peptides. Biosens Bioelectron. 2018;100:115-21. https://doi.org/10.1016/j.bios.2017.08.029 PMid:28886455
  150. Rolain JM, Maurin M, Vestris G, Raoult D. In vitro susceptibilities of 27 rickettsiae to 13 antimicrobials. Antimicrob Agents Chemother. 1998;42(7):1537-41. https://doi.org/10.1128/AAC.42.7.1537 PMid:9660979 PMCid:PMC105641
  151. Smilack JD. The tetracyclines. Mayo Clin Proc. 1999;74(7):727-9. https://doi.org/10.4065/74.7.727 PMid:10405705  
  152. Kline AH, Blattner RJ, Lunin M. Transplacental Effect of Tetracyclines on Teeth. JAMA J Am Med Assoc. 1964;188(2):178-80. https://doi.org/10.1001/jama.1964.03060280080021 PMid:14172262
  153. Meloni G, Meloni T. Azithromycin vs. doxycycline for Mediterranean spotted fever. Pediatr Infect Dis J. 1996;15(11):1042-4. https://doi.org/10.1097/00006454-199611000-00022 PMid:8933556  
  154. Bella F, Font B, Uriz S, Munoz T, Espejo E, Traveria J, Serrano JA, Segura F. Randomized trial of doxycycline versus josamycin for Mediterranean spotted fever. Antimicrob Agents Chemother. 1990;34(5):937-8. https://doi.org/10.1128/AAC.34.5.937 PMid:2193627 PMCid:PMC171727
  155. Cascio A, Colomba C, Di Rosa D, Salsa L, di Martino L, Titone L. Efficacy and Safety of Clarithromycin as Treatment for Mediterranean Spotted Fever in Children: A Randomized Controlled Trial. Clin Infect Dis. 2001;33(3):409-11. https://doi.org/10.1086/321864 PMid:11438914
  156. Muñoz-Espin T, López-Parés P, Espejo-Arenas E, Font-Creus B, Martinezvila I, Travería-Casanova J, Segura-Porta F, Bella-Cueto F. Erythromycin versus tetracycline for treatment of mediterranean spotted fever. Arch Dis Child. 1986;61(10):1027-9. https://doi.org/10.1136/adc.61.10.1027 PMid:3535687 PMCid:PMC1777969
  157. Raoult D, Roussellier P, Galicher V, Perez R, Tamalet J. In vitro susceptibility of Rickettsia conorii to ciprofloxacin as determined by suppressing lethality in chicken embryos and by plaque assay. Antimicrob Agents Chemother. 1986;29(3):424-5. https://doi.org/10.1128/AAC.29.3.424 PMid:2940972 PMCid:PMC180407  
  158. Raoult D, Roussellier P, Vestris G, Galicher V, Perez R, Tamalet J. Susceptibility of rickettsia conorii and r. Rickettsii to pefloxacin, in vitro and in ovo. J Antimicrob Chemother. 1987;19(3):303-5. https://doi.org/10.1093/jac/19.3.303 PMid:3106303
  159. Jabarit-Aldighieri N, Torres H, Raoult D. Susceptibility of Rickettsia conorii, R. rickettsii, and Coxiella burnetii to PD 127,391, PD 131,628, pefloxacin, ofloxacin, and ciprofloxacin. Antimicrob Agents Chemother. 1992;36(11):2529-32. https://doi.org/10.1128/AAC.36.11.2529 PMid:1336950 PMCid:PMC284367
  160. Ruiz Beltrán R, Herrero Herrero JI. Evaluation of ciprofloxacin and doxycycline in the treatment of mediterranean spotted fever. Eur J Clin Microbiol Infect Dis. 1992;11(5):427-31. https://doi.org/10.1007/BF01961857 PMid:1425713  
  161. Gudiol F, Pallares R, Carratala J, Bolao F, Ariza J, Rufi G, Viladrich PF. Randomized double-blind evaluation of ciprofloxacin and doxycycline for Mediterranean spotted fever. Antimicrob Agents Chemother. 1989;33(6):987-8. https://doi.org/10.1128/AAC.33.6.987 PMid:2669629 PMCid:PMC284272  
  162. Raoult D, Gallais H, De Micco P, Casanova P. Ciprofloxacin therapy for Mediterranean spotted fever. Antimicrob Agents Chemother. 1986;30(4):606-7. https://doi.org/10.1128/AAC.30.4.606 PMid:3789693 PMCid:PMC176489  
  163. Botelho-Nevers E, Rovery C, Richet H, Raoult D. Analysis of risk factors for malignant mediterranean spotted fever indicates that fluoroquinolone treatment has a deleterious effect. J Antimicrob Chemother. 2011;66(8):1821-30. https://doi.org/10.1093/jac/dkr218 PMid:21642652
  164. Botelho-nevers E, Edouard S, Leroy Q, Raoult D. Deleterious effect of ciprofloxacin on Rickettsia conorii-infected cells is linked to toxin-antitoxin module up-regulation. J Antimicrob Chemother. 2012;67(7):1677-82. https://doi.org/10.1093/jac/dks089 PMid:22467631  
  165. Raoult D. Antibiotic susceptibility of Rickettsia and treatment of rickettsioses. Eur J Epidemiol. 1989;5(4):432-5. https://doi.org/10.1007/BF00140135 PMid:2606171
  166. Bella F, Espejo E, Uriz S, Serrano JA, Alegre MD, Tort J. Randomized trial of 5-day rifampin versus 1-day doxycycline therapy for mediterranean spotted fever. J Infect Dis. 1991;164(2):433-4. https://doi.org/10.1093/infdis/164.2.433 PMid:1856496  
  167. Ruiz Beltran R, Herrero Herrero JI. Deleterious effect of trimethoprim-sulfamethoxazole in Mediterranean spotted fever. Antimicrob Agents Chemother. 1992;36(6):1342-3. https://doi.org/10.1128/AAC.36.6.1342 PMid:1416836 PMCid:PMC190344
  168. Castaneda MR. The antigenic relationship between bacillus proteus x-19 and rickettsiae: III. a study of the antigenic composition of the extracts of bacillus proteus x-19. J Exp Med. 1935;62(3):289-96. https://doi.org/10.1084/jem.62.3.289 PMid:19870415 PMCid:PMC2133285