Retinitis por Rickettsia conorii, una infección emergente en el sureste de la península ibérica / Rickettsia conorii retinitis: an emerging infection in the southeast of the Iberian Peninsula

CASO CLÍNICO: Se presenta un caso de un paciente de 60 años con pérdida de agudeza visual de ambos ojos tras un cuadro de fiebre y erupción cutánea con afectación palmo plantar. Tras una exploración completa y las pruebas complementarias pertinentes se llegó al diagnóstico de retinitis en el contexto de infección por Rickettsia conorii. Se expone la evolución tras el tratamiento con doxiciclina y prednisona a las seis semanas con una mejora anatómica y funcional significativa. La rickettsiosis es una zoonosis emergente que puede presentar afectación ocular. Ésta suele ser una retinitis multifocal que afecta al polo posterior con un desprendimiento seroso macular y vitritis. La sospecha clínica necesitará de una confirmación serológica para el diagnóstico definitivo. El tratamiento con antibióticos y corticoesteroides ha demostrado ser efectivo. Habrá que tenerla en cuenta en áreas endémicas mediterráneas y en periodo estival, donde el riesgo es mucho mayor

CASE REPORT: A case is presented of a 60-year-old patient with loss of visual acuity in both eyes after fever and skin rash with palmoplantar involvement. After a complete examination and relevant complementary tests, the diagnosis of retinitis was made in the context of Rickettsia conorii infection. The evolution after treatment with doxycycline and prednisone at six weeks with significant anatomical and functional improvement is presented. Rickettsiosis is an emerging zoonosis that can present with ocular involvement. This is usually a multifocal retinitis affecting posterior pole with macular serous detachment and vitritis. Clinical suspicion will require serological confirmation for a definitive diagnosis. Treatment with antibiotics and corticosteroids has been shown to be effective. It should be taken into account in Mediterranean endemic areas, and in in the summer period, where the risk is much higher

Enfermedad del injerto contra el receptor tras infección por Rickettsia conorii inducida por la picadura de una garrapata / Graft-versus-host disease after an infection by Rickettsia conorii induced by a tick's bite

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Prospective Cohort Study of Single-Day Doxycycline Therapy for Mediterranean Spotted Fever.

The objective of this study is to evaluate the results of single-day doxycycline therapy for Mediterranean spotted fever (MSF). This is a prospective cohort study of cases with confirmed MSF treated with the single-day doxycycline regimen in a teaching hospital from 1990 to 2015. Patients received two oral doses of 200 mg of doxycycline for 1 day. The outcomes evaluated were the time interval between the start of treatment and apyrexia, the time interval between the start of treatment and disappearance of other symptoms, and the adverse reactions to treatment and death. The study included 158 subjects, 18 of whom (11.4%) had a severe form of MSF and 31 (19.6%) were >65 years. The interval between onset of symptoms and start of treatment was 4.31 ± 1.54 days. All patients recovered uneventfully. Fever disappeared 2.55 ± 1.14 days after the start of treatment. The remaining symptoms (headache, arthromyalgia) disappeared 3.63 ± 1.35 days after the start of treatment. Only one patient had a delay in reaching apyrexia (8 days). The fever disappeared somewhat later in severe cases (median, 3 days; interquartile range [IQR], 2 to 4 days) than in nonsevere cases (median, 2 days; IQR, 2 to 3 days). Likewise, the remaining symptoms disappeared later in severe cases (median, 5 days; IQR, 4 to 6 days) than in nonsevere cases (median, 3 days; IQR, 3 to 4 days). The outcome was similar in both elderly and nonelderly patients. Eight patients had mild adverse effects possibly related to treatment. The results of the study confirm that single-day doxycycline therapy is an effective and well-tolerated treatment for MSF, including elderly patients and severe cases.

Shell-vial culture, coupled with real-time PCR, applied to Rickettsia conorii and Rickettsia massiliae-Bar29 detection, improving the diagnosis of the Mediterranean spotted fever.

Rickettsia conorii and Rickettsia massiliae-Bar29 are related to Mediterranean spotted fever (MSF). They are intracellular microorganisms. The Shell-vial culture assay (SV) improved Rickettsia culture but it still has some limitations: blood usually contains low amount of microorganisms and the samples that contain the highest amount of them are non-sterile. The objectives of this study were to optimize SV culture conditions and monitoring methods and to establish antibiotic concentrations useful for non-sterile samples. 12 SVs were inoculated with each microorganism, incubated at different temperatures and monitored by classical methods and real-time PCR. R. conorii was detected by all methods at all temperatures since 7th day of incubation. R. massiliae-Bar29 was firstly observed at 28°C. Real-time PCR allowed to detected it 2-7 days earlier (depend on temperature) than classical methods. Antibiotics concentration needed for the isolation of these Rickettsia species from non-sterile samples was determined inoculating SV with R. conorii, R. massiliae-Bar29, biopsy or tick, incubating them with different dilutions of antibiotics and monitoring them weekly. To sum up, if a MSF diagnosis is suspected, SV should be incubated at both 28°C and 32°C for 1-3 weeks and monitored by a sensitive real-time PCR. If the sample is non-sterile the panel of antibiotics tested can be added.

Randomized Trial of Clarithromycin for Mediterranean Spotted Fever.

The classic antibiotic treatment for Mediterranean spotted fever (MSF) is based on tetracyclines or chloramphenicol, but chloramphenicol's bone marrow toxicity makes tetracyclines the treatment of choice. However, it is convenient to have alternatives available for patients who are allergic to tetracyclines, pregnant women, and children <8 years old. We conducted a randomized clinical trial to compare clarithromycin with doxycycline or josamycin in the treatment of MSF. Forty patients were evaluated (23 male; mean age, 39.87 years); 13 patients were aged <14 years. Seventeen patients received clarithromycin, and 23 received doxycycline or josamycin. The interval between the onset of symptoms and the start of treatment was 4.04 ± 1.70 days in the clarithromycin group versus 4.11 ± 1.60 days in the doxycycline/josamycin group (P = not significant [NS]). Time to the disappearance of fever after treatment was 2.67 ± 1.55 days in the clarithromycin group versus 2.22 ± 1.35 days in the doxycycline/josamycin (P = NS). The symptoms had disappeared at 4.70 ± 2.25 days in the clarithromycin group versus at 4.75 ± 3.08 days in the doxycycline/josamycin (P = NS). There were no adverse reactions to treatment or relapses in either group. In conclusion, clarithromycin is a good alternative to doxycycline or josamycin in the treatment of MSF.

A case of Mediterranean spotted fever associated with severe respiratory distress syndrome.

Mediterranean spotted fever (MSF) is usually a mild endemic rickettsial disease occurring in southern Croatia. We have reported the clinical and epidemiological characteristics of an acute MSF case associated with severe respiratory distress syndrome and hemodynamical instability. The patient recovered completely after antimicrobial treatment. Indirect immunofluorescence assay (FOCUS Diagnostics Inc.) was performed to detect IgM and IgG antibodies to Rickettsia conorii. A significant increase of both IgM and IgG antibody titres found in paired acute- and convalescent-phase serum confirmed the diagnosis of acute MSF.

Host-directed antimicrobial drugs with broad-spectrum efficacy against intracellular bacterial pathogens.

We sought a new approach to treating infections by intracellular bacteria, namely, by altering host cell functions that support their growth. We screened a library of 640 Food and Drug Administration (FDA)-approved compounds for agents that render THP-1 cells resistant to infection by four intracellular pathogens. We identified numerous drugs that are not antibiotics but were highly effective in inhibiting intracellular bacterial growth with limited toxicity to host cells. These compounds are likely to target three kinds of host functions: (i) G protein-coupled receptors, (ii) intracellular calcium signals, and (iii) membrane cholesterol distribution. The compounds that targeted G protein receptor signaling and calcium fluxes broadly inhibited Coxiella burnetii, Legionella pneumophila, Brucella abortus, and Rickettsia conorii, while those directed against cholesterol traffic strongly attenuated the intracellular growth of C. burnetii and L. pneumophila. These pathways probably support intracellular pathogen growth so that drugs that perturb them may be therapeutic candidates. Combining host- and pathogen-directed treatments is a strategy to decrease the emergence of drug-resistant intracellular bacterial pathogens. Importance: Although antibiotic treatment is often successful, it is becoming clear that alternatives to conventional pathogen-directed therapy must be developed in the face of increasing antibiotic resistance. Moreover, the costs and timing associated with the development of novel antimicrobials make repurposed FDA-approved drugs attractive host-targeted therapeutics. This paper describes a novel approach of identifying such host-targeted therapeutics against intracellular bacterial pathogens. We identified several FDA-approved drugs that inhibit the growth of intracellular bacteria, thereby implicating host intracellular pathways presumably utilized by bacteria during infection.

Deleterious effect of ciprofloxacin on Rickettsia conorii-infected cells is linked to toxin-antitoxin module up-regulation.

OBJECTIVES: To confirm and better understand the deleterious effect of fluoroquinolones reported during Rickettsia conorii infection in humans. METHODS: We used a new plaque assay to test the effect of ciprofloxacin on cells infected by R. conorii. Controls were mock-treated infected cells and infected cells treated with doxycycline. We used real-time quantitative RT-PCR to quantify vapC and vapB transcripts in cells infected by R. conorii treated with ciprofloxacin or mock treated. RESULTS: By plaque assay, at baseline (0 h) there is no difference in lytic area between cells treated with ciprofloxacin (whatever concentration used) and controls. Ciprofloxacin at 0.5 and 50 mg/L induced a significant increase in lytic areas compared with the control at 2 h, 4 h, 6 h (P<0.0001) and 24 h (P<0.0001 and P=0.035, respectively) when not induced with doxycycline. By real-time quantitative RT-PCR, ciprofloxacin was found to cause an up-regulation of toxin-antitoxin (TA) module transcription. Indeed, the mRNA levels of vapC and vapB were significantly increased at 2 h and 4 h in cells treated with 50 mg/L ciprofloxacin (not with 0.5 mg/L ciprofloxacin) compared with control levels (fold change >2.9). CONCLUSIONS: These in vitro findings correlated with our previous clinical findings: fluoroquinolones have a deleterious effect during R. conorii infection, not found with doxycycline. We speculate that the toxic effect of fluoroquinolones on R. conorii-infected cells is mediated by the overexpression of TA, possibly followed by their release into the host cytoplasm as described in Rickettsia felis.

Statins limit Rickettsia conorii infection in cells.

Recent data suggest that statins may have a beneficial effect during sepsis. In this study, we tested the effect of lovastatin and pravastatin on the cellular culture of Rickettsia conorii using a quantitative plaque assay model associated with an original image analysis algorithm. Statins added at the time of infection did not modify plaque formation, whereas pre-incubation with statins for 48h resulted in a significant 30-68% plaque reduction, depending on the tested compounds and doses. These preliminary findings raise the hypothesis that statins may prevent or moderate rickettsial disease in exposed people.

Nitric oxide as a mediator of increased microvascular permeability during acute rickettsioses.

Rickettsiae primarily infect the microvascular endothelium, leading to changes in microvascular permeability that result in potentially severe pulmonary and cerebral edema. The mechanisms responsible for these changes are not well understood. One potential mechanism of increased vascular permeability is the anti-rickettsial nitric oxide response described by Walker and colleagues. We hypothesized that anti-rickettsial levels of nitric oxide adversely affects microvascular permeability in vitro. To this end we sought to describe the effects of exogenous nitric oxide on the proliferation of intracellular rickettsiae while monitoring the transendothelial electrical resistance as a measure of endothelial barrier integrity. It was determined that the addition of the NO-donor DETA NONOate at certain levels results in a dose-dependent change in electrical resistance across the monolayer while effectively limiting the number of intracellular rickettsiae in human microvascular endothelial cells. The data presented support the idea that nitric oxide produced by infected endothelial cells may be contributing to the changes in vascular permeability that occur during acute rickettsioses. Future experiments aim to elaborate on these results in a model that more clearly depicts the in vivo response as well as to describe the changes that occur with respect to interendothelial junctions.

Effect of antibody on the rickettsia-host cell interaction.

A recent study demonstrated that polyclonal antibodies to Rickettsia conorii and monoclonal antibodies to outer membrane proteins A (OmpA) and B (OmpB) provided effective, Fc-dependent, passive immunity, even in severe combined immunodeficient mice with an established infection. In order to determine the mechanism of protection, mouse endothelial and macrophage-like cell lines were infected with R. conorii that had been exposed to polyclonal antibodies, monoclonal antibodies to OmpA or OmpB, Fab fragments of the polyclonal antibodies, or normal serum or that were left untreated. At 0 h, Fc-dependent antibody enhancement of R. conorii adherence to endothelial and macrophage-like cell lines was inhibited by the presence of normal serum, suggesting Fc receptor-mediated adherence of opsonized rickettsiae. At 3 h, the opsonized rickettsiae had been internalized. After 72 h, inhibited survival of rickettsiae exposed to polyclonal antibodies or monoclonal antibodies to OmpA or OmpB was evident compared with growth of untreated and normal serum-treated and polyclonal Fab antibody-treated R. conorii. Polyclonal antibodies and an anti-OmpB monoclonal antibody inhibited the escape of R. conorii from the phagosome, resulting in intraphagolysosomal rickettsial death. At 48 h of infection, rickettsicidal activity of macrophages by opsonized rickettsiae was inhibited by NG-monomethyl-L-arginine, superoxide dismutase, mannitol, or supplemental L-tryptophan, and endothelial rickettsicidal activity against opsonized rickettsiae was inhibited by NG-monomethyl-L-arginine, superoxide dismutase, catalase, or supplemental L-tryptophan. Thus, Fc-dependent antibodies protected against R. conorii infection of endothelium and macrophages by opsonization that inhibited phagosomal escape and resulted in phagolysosomal killing mediated by nitric oxide, reactive oxygen intermediates, and L-tryptophan starvation.

A case of pleurisy associated with antibodies to Rickettsia conorii.

Rickettsia conorii is endemic in Mediterranean area. We describe an unusual sace of R. Conorii infection, which concerns a farmer with clinical, radiological and cytological findings of pleurisy without evidence of malignancy. An elevated antibody titre for R. Conorii was observed, using an indirect immunofluorescent antibody test. After treatment with Doxycycline, the patient presented a significant improvement of his clinical and radiological image and a four-fold decrease of the antibody titre for R. conorii.

Evaluation of antibiotic susceptibilities of three rickettsial species including Rickettsia felis by a quantitative PCR DNA assay.

Rickettsiae grow only intracellularly, and the antibiotic susceptibilities of these bacteria have been assessed by either plaque, dye uptake, or immunofluorescence assays, which are time-consuming. We used a quantitative PCR (with the LightCycler instrument) to assess the levels of inhibition of Rickettisa felis, R. conorii, and R. typhi DNA synthesis in the presence of various antibiotics. We established the kinetics of rickettsial DNA during growth and showed that R. conorii grows more quickly than R. typhi in cell culture, with maximum replication occurring after 5 and 7 days, respectively. The MICs of the antibiotics tested for R. conorii and R. typhi by the quantitative PCR assay were similar to those previously obtained by plaque and dye uptake assays. We found that R. felis is susceptible to doxycycline, rifampin, thiamphenicol, and fluoroquinolones but not to gentamicin, erythromycin, amoxicillin, or trimethoprim-sulfamethoxazole. The resistance of this new species to erythromycin is consistent with its current taxonomic position within the spotted fever group. We believe that quantitative PCR could be used in the future to simplify and shorten antibiotic susceptibility assays of other rickettsiae and other strict intracellular pathogens.

Clarithromycin versus azithromycin in the treatment of Mediterranean spotted fever in children: a randomized controlled trial.

We conducted an open-label randomized controlled trial to compare the efficacy and safety of clarithromycin (15/mg/kg/day in 2 divided doses for 7 days) with those of azithromycin (10 mg/kg/day in 1 dose for 3 days) in the treatment of children with Mediterranean spotted fever. Until now, there has not been a gold-standard therapy for this rickettsial disease in children. Eighty-seven children were randomized to receive 1 of the 2 drugs. The mean time to defervescence (+/- standard deviation) was 46.2+/-36.4 h in the clarithromycin group and 39.3+/-31.3 h in the azithromycin group. These differences were not statistically significant and both drugs were equally well-tolerated. Clarithromycin and azithromycin could be acceptable therapeutic alternatives to chloramphenicol and tetracyclines for children aged < or =8 years with Mediterranean spotted fever. Azithromycin, because it has a long half-life, offers the advantages of administration in a single daily dose and a shorter duration of therapy, which could increase compliance in children.

Rickettsial phospholipase A2 as a pathogenic mechanism in a model of cell injury by typhus and spotted fever group rickettsiae.

Phospholipase A2 activity by typhus group rickettsiae causes hemolysis in vitro. Rickettsial phospholipase A2 has been proposed to mediate entry into the host cell, escape from the phagosome, and cause injury to host cells by both typhus and spotted fever group rickettsiae. In a rickettsial contact-associated cytotoxicity model, the interaction of Rickettsia prowazekii or R. conorii with Vero cells caused temperature-dependent release of 51Cr from the cells. Treatment of rickettsiae, but not the cells, with a phospholipase A2 inhibitor (bromophenacyl bromide) or with antibody to king cobra venom inhibited cell injury. Rickettsial treatment with bromophenacyl bromide inhibited the release of free fatty acids from the host cell. Neither the inhibitor nor antivenom impaired rickettsial active transport of L-lysine. Thus, host cell injury was mediated by a rickettsial phospholipase A2-dependent mechanism.

In vitro activities of telithromycin (HMR 3647) against Rickettsia rickettsii, Rickettsia conorii, Rickettsia africae, Rickettsia typhi, Rickettsia prowazekii, Coxiella burnetii, Bartonella henselae, Bartonella quintana, Bartonella bacilliformis, and Ehrlichia chaffeensis.

In vitro activities of telithromycin compared to those of erythromycin against Rickettsia spp., Bartonella spp., Coxiella burnetii, and Ehrlichia chaffeensis were determined. Telithromycin was more active than erythromycin against Rickettsia, Bartonella, and Coxiella burnetii, with MICs of 0.5 microg/ml, 0.003 to 0.015 microg/ml, and 1 microg/ml, respectively, but was inactive against Ehrlichia chaffeensis.