Biomedical subjects
Publications and source records attributed to Pierre-Edouard Fournier.
Over the last 20 years, advances in molecular techniques have greatly facilitated the identification of the members of the Rickettsiales, and numerous new species and diseases have been described. In this paper, we review taxonomic rules and appropriate approaches to valid naming of rickettsial species and the diseases they cause.
The analyses of genome sequences from Rickettsia conorii and R. prowazekii have allowed the identification of five genes encoding autotransporter proteins, including ompA and four genes annotated in the R. prowazekii genome as "surface cell antigen" (sca) genes. Of these, ompA and sca5 (ompB) are known to encode membrane-exposed antigenic proteins playing a major role in the host's immune response, and sca4 encodes a truncated autotransporter protein. In order to study further the phylogeny of the genus Rickettsia, we attempted amplification and sequencing of the sca2 genes from the 20 currently validated Rickettsia species. Sixteen species exhibited a complete sca2 gene, ranging from 2,727 bp in R. bellii to 5,580 bp in R. rhipicephali. R. helvetica and R. canadensis had a split gene, and R. prowazekii and R. typhi had remnant fragments of sca2. We also identified in R. akari, R. prowazekii, and R. typhi a duplication of the sca2 gene. The phylogenetic trees inferred from both the nucleotide and protein sequences of sca2 showed four clusters of rickettsiae, that is, the R. rickettsii, R. massiliae, R. akari, and typhus groups, which were supported by significant bootstrap values.
A potentially broad-range PCR system for gltA gene amplification has been designed. A nearly full (992 bp) sequence of the gltA gene was amplified in two steps. DNA was extracted from organs (spleen and liver) of four mostly spread species of rodents: Apodemus agrarius and Apodemus peninsulae mice; Clethrionomys rufocanus and Microtus fortis voles. All amplicons have been sequenced. The incidence of Bartonella species in rodents was 73%, 60%, 60%, and 83% for the above-mentioned rodents, respectively. Several genotypes have been identified, including two gltA genotypes close to B. taylorii, four gltA genotypes close to B. grahamii, and one genotype of a potentially new species. Several Bartonella species were found in both mice species, although each vole species harbored a single species of Bartonella. High incidence of Bartonella infection was found in all studied species of small rodents in the Russian Far East. Several amplified genotypes probably belong to local strains of B. taylorii and B. grahamii, and one genotype represents a possible new species.
The rickettsioses--zoonotic bacterial infections transmitted to humans by arthropods--were for many years considered to be oddities in travel medicine. During the previous 2 decades, however, reports of >450 travel-associated cases have been published worldwide, the vast majority being murine typhus caused by Rickettsia typhi, Mediterranean spotted fever caused by Rickettsia conorii, African tick bite fever caused by Rickettsia africae, and scrub typhus caused by Orientia tsutsugamushi. Most patients present with a benign febrile illness accompanied by headache, myalgia, and cutaneous eruptions, but severe complications and fatalities are occasionally seen. Current microbiological tests include culture, polymerase chain reaction, and serological analysis, of which only the latter method is widely available. Tetracyclines are the drugs of first choice and should be prescribed whenever a case of rickettsiosis is suspected. Preventive measures rely on minimizing the risk of arthropod bites when traveling in areas of endemicity.
BACKGROUND: Tick-borne rickettsioses are of emerging importance in today's travel medicine but have until recently received little attention. We describe the current knowledge of tick-borne rickettsioses as they relate to international travel, their microbiological diagnosis, treatment, possible prevention, and future prospects. METHODS: Literature-based review and personal observations. RESULTS: During the last decade, some 400 cases of tick-borne rickettsioses have been reported in international travellers, the vast majority being African tick bite fever caused by Rickettsia africae and Mediterranean spotted fever caused by Rickettsia conorii. Only a minority of infected travellers can recall a preceding tick bite. Most patients present with a mild-to-moderately severe flu-like illness typically accompanied by a cutaneous rash and an inoculation eschar at the site of the tick bite, but potentially life-threatening disease with disseminated vaculitis is occasionally seen. Definite microbiological confirmation of tick-borne rickettsioses by isolation or antigen detection is only available at reference laboratories and diagnosis must in most cases rely on clinical and epidemiological data supported by serology. Doxycycline is the recommended treatment for tick-borne rickettsioses and prevention is based on personal protective measures against tick bites when travelling in endemic areas. CONCLUSION: Tick-borne rickettsiosis should be suspected in febrile returnees from endemic areas, especially in cases with skin eruptions. Travellers to endemic areas should be encouraged to use personal protective measures against tick bites.
Recent seroepidemiological studies and examinations of Ixodes ricinus ticks in Europe have demonstrated the presence of an emerging tick-borne infection with Rickettsia helvetica. We conducted a serosurvey in 168 Danish patients seropositive for borreliosis reflecting their exposure to I. ricinus ticks. A total of 21 patients (12.5%) had positive antibody titres to R. helvetica including 4 cases of seroconversion. None of the samples were positive for antibodies to Ehrlichia. We conclude that in humans exposed to I. ricinus ticks in Denmark the risk of acquiring rickettsial infection is for the first time demonstrated. In the same region of Denmark we collected 570 I. ricinus ticks from various sources, and examinations by PCR for Rickettsia were performed. Positive reactions were obtained in 23 ticks (4%), and R. helvetica was identified in all 13 of those for which sequencing was performed.
In testing paired serum samples from 40 consecutive cases of African tick bite fever, we detected diagnostic antibodies against spotted fever group rickettsiae in 45% of the patients by immunofluorescence assay (IFA) and in 100% of the patients by Western blotting (WB) (P < 0.01). A specific diagnosis of Rickettsia africae infection could be established in 15% of the patients by IFA and in 73% of the patients by a combination of WB and cross-adsorption assays (P < 0.01).
The availability of the Tropheryma whipplei genome offers the putative possibility of choosing logical DNA targets. We applied a PCR assay (targeting repeated sequences of T. whipplei) to samples from patients with Whipple's disease and to those from members of a control group. When compared to the results seen with regular PCR, the sensitivity of repeat PCR was significantly enhanced (P = 0.02) without alteration of its specificity.
By use of the nearly perfectly colinear genomes of Rickettsia conorii and Rickettsia prowazekii, we compared the usefulness of three types of sequences for typing of R. conorii isolates: (i) 5 variable coding genes comprising the 16S ribosomal DNA, gltA, ompB, and sca4 (gene D) genes, which are present in both genomes, and the ompA gene, which is degraded in R. prowazekii; (ii) 28 genes degraded in R. conorii but intact in R. prowazekii, including 23 split and 5 remnant genes; and (iii) 27 conserved and 25 variable intergenic spacers. The 4 conserved and 23 split genes as well as the 27 conserved intergenic spacers each had identical sequences in 34 human and 5 tick isolates of R. conorii. Analysis of the ompA sequences identified three genotypes of R. conorii. The variable intergenic spacers were significantly more variable than conserved genes, split genes, remnant genes, and conserved spacers (P < 10(-2) in all cases). Four of the variable intergenic spacers (dksA-xerC, mppA-purC, rpmE-tRNA(fMet), and tRNA(Gly)-tRNA(Tyr)) had highly variable sequences; when they were combined for typing, multispacer typing (MST) identified 27 different genotypes in the 39 R. conorii isolates. Two batches from the same R. conorii strain, Malish (Seven), with different culture passage histories were found to exhibit the same MST type. MST was more discriminatory for strain genotyping than multiple gene sequencing (P < 10(-2)). Phylogenetic analysis based on MST sequences was concordant with the geographic origins of R. conorii isolates. Our study supports the usefulness of MST for strain genotyping. This tool may be useful for tracing a strain and identifying its source during outbreaks, including those resulting from bioterrorism.
We developed a new molecular tool based on rpoB gene (encoding the beta subunit of RNA polymerase) sequencing to identify streptococci. We first sequenced the complete rpoB gene for Streptococcus anginosus, S. equinus, and Abiotrophia defectiva. Sequences were aligned with these of S. pyogenes, S. agalactiae, and S. pneumoniae available in GenBank. Using an in-house analysis program (SVARAP), we identified a 740-bp variable region surrounded by conserved, 20-bp zones and, by using these conserved zones as PCR primer targets, we amplified and sequenced this variable region in an additional 30 Streptococcus, Enterococcus, Gemella, Granulicatella, and Abiotrophia species. This region exhibited 71.2 to 99.3% interspecies homology. We therefore applied our identification system by PCR amplification and sequencing to a collection of 102 streptococci and 60 bacterial isolates belonging to other genera. Amplicons were obtained in streptococci and Bacillus cereus, and sequencing allowed us to make a correct identification of streptococci. Molecular signatures were determined for the discrimination of closely related species within the S. pneumoniae-S. oralis-S. mitis group and the S. agalactiae-S. difficile group. These signatures allowed us to design a S. pneumoniae-specific PCR and sequencing primer pair.
We report that eight patients from France, Italy, and Thailand had serological evidence of Rickettsia helvetica infection. The infection presented as a mild disease in the warm season and was associated with fever, headache, and myalgia but not with a cutaneous rash. R. helvetica should be suspected in patients with unexplained fever, especially following a bite from an Ixodes sp. tick.
Using PCR, we screened 411 ticks from four genera collected in Russia and Kazakhstan for the presence of rickettsiae and ehrlichiae. In Russia, we detected "Rickettsia heilongjiangensis," Rickettsia sp. strain RpA4, and Ehrlichia muris. In Kazakhstan, we detected Rickettsia sp. strain RpA4 and a rickettsia closely related to Rickettsia aeschlimannii. These agents should be considered in a differential diagnosis of tick-borne infections in these areas.
As rickettsioses may be severe diseases and Rickettsia prowazekii is a potential agent of bioterrorism, highly efficient diagnostic techniques are required to detect rickettsiae in patients. We developed a nested PCR assay using single-use primers targeting single-use gene fragments present in the genomes of both Rickettsia conorii and R. prowazekii. We used this "suicide" PCR with DNA from 103 skin biopsy specimens from patients who definitely had a rickettiosis, 109 skin biopsy specimens from patients who possibly had a rickettsiosis, and 50 skin biopsy specimens from patients with nonrickettsial diseases. The suicide PCR detected "R. conorii conorii" in 38 biopsy specimens, R. africae in 28 biopsy specimens, R. slovaca in 12 biopsy specimens, "R. sibirica mongolotimonae" in 5 biopsy specimens, R. aeschlimannii in 2 biopsy specimens, and "R. conorii caspia" and "R. sibirica sibirica" in 1 biopsy specimen each. The technique had a specificity of 100% and a sensitivity of 68%. It was 2.2 times more sensitive than culture (P < 10(-2)) and 1.5 times more sensitive than regular PCR (P < 10(-2)). The efficacy of the suicide PCR was reduced by antibiotic therapy prior to biopsy (P < 10(-2)) and was increased when it was performed with eschar biopsy specimens (P = 0.03). We propose the use of the suicide PCR as a sensitive, specific, and versatile technique for improving the diagnosis of rickettsioses, especially when it is used on eschar biopsy specimens taken prior to antibiotic therapy.
An acute tick-borne rickettsiosis caused by Rickettsia heilongjiangensis was diagnosed in 13 patients from the Russian Far East in 2002. We amplified and sequenced four portions of three rickettsial genes from the patients' skin biopsy results and blood samples and showed that the amplified rickettsial genes belong to R. heilongjiangensis, which was recently isolated from Dermacentor sylvarum ticks in nearby regions of China. This rickettsia, belonging to subgroup of R. japonica, was previously suggested to be pathogenic for humans on the basis of serologic findings. We tested serum samples with different rickettsial antigens from 11 patients and confirmed increasing titers of immunoglobulin (Ig) G and IgM to spotted fever group rickettsiae, including R. heilongjiangensis. Clinical and epidemiologic data on these patients show that this disease is similar to other tick-borne rickettsioses.
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Three pandemics have been attributed to plague in the last 1,500 years. Yersinia pestis caused the third, and its DNA was found in human remains from the second. The Antiqua biovar of Y. pestis may have caused the first pandemic; the other two biovars, Medievalis and Orientalis, may have caused the second and third pandemics, respectively. To test this hypothesis, we designed an original genotyping system based on intergenic spacer sequencing called multiple spacer typing (MST). We found that MST differentiated every biovar in a collection of 36 Y. pestis isolates representative of the three biovars. When MST was applied to dental pulp collected from remains of eight persons who likely died in the first and second pandemics, this system identified original sequences that matched those of Y. pestis Orientalis. These data indicate that Y. pestis caused cases of Justinian plague. The two historical plague pandemics were likely caused by Orientalis-like strains.
To estimate the incidence of, identify risk factors for, and describe the clinical presentation of travel-associated African tick bite fever (ATBF), a rapidly emerging disease in travel medicine, we prospectively studied a cohort of 940 travelers to rural sub-Equatorial Africa. Diagnosis was based on suicide polymerase chain reaction and the detection of specific antibodies to Rickettia africae in serum samples by multiple-antigen microimmunofluorescence assay, Western blotting, and cross-adsorption assays. Thirty-eight travelers, 4.0% of the cohort and 26.6% of those reporting flulike symptoms, had ATBF diagnosed. More than 80% of the patients had fever, headache, and/or myalgia, whereas specific clinical features such as inoculation eschars, lymphadenitis, cutaneous rash, and aphthous stomatitis were seen in < or = 50% of patients. Game hunting, travel to southern Africa, and travel during November through April were found to be independent risk factors. Our study suggests that ATBF is not uncommon in travelers to rural sub-Saharan Africa and that many cases have a nonspecific presentation.
Information regarding the inflammatory response in African tick-bite fever (ATBF), an emerging spotted-fever-group rickettsiosis, in international travelers to sub-Saharan Africa, is scarce. Plasma/serum levels of von Willebrand factor (vWF), soluble (s) E-selectin, tumor necrosis factor-alpha, interleukin (IL)-6, interferon-gamma, IL-10, IL-13, IL-8, RANTES, macrophage inflammatory protein-1alpha, and C-reactive protein were studied, at both first presentation and follow-up, in 15 patients with travel-associated ATBF and in 14 healthy travelers who served as control subjects. Our main and novel findings are the following: (1) patients with ATBF had increased levels of vWF and sE-selectin, with a subsequent decrease at follow-up; (2) with the exception of IFN-gamma, levels of cytokines and chemokines were also increased in these patients at the first presentation; and (3) IL-10 and IL-13 tended to increase during follow-up, whereas most of the inflammatory cytokines decreased. The induction of these mediators and the balance between them may be critical both for the regulation of inflammation and for protective immunity in ATBF.