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D Raoult

Publications and source records attributed to D Raoult.

At least 73 records · Page 4Linked to original sources
PubMed1994

Phagolysosomal alkalinization and intracellular killing of Staphylococcus aureus by amikacin.

The aminoglycosides are ineffective against intracellular Staphylococcus aureus, which resides within lysosomes, despite a strong extracellular bactericidal activity. Since they are slowly concentrated within lysosomes it was hypothesized that acidity within these cell compartments might impair antibiotic activity. The bactericidal activity of amikacin alone and combined with lysosomotropic alkalinizing agents (LAA), which can alkalinize acidic cell compartments, was evaluated. Before antibiotic challenge, some cells were preincubated with amikacin for 3 days to allow intracellular accumulation of drug. No intracellular killing activity was shown when non-preincubated cells were used. Conversely, with preincubated cells, S. aureus was killed within 4 h when LAA were incorporated into the incubation media but not with amikacin alone. Enhanced antimicrobial activity correlated with increase in lysosomal pH. Intracellular accumulation of amikacin was not changed by LAA. These results provide evidence that acidic pH within lysosomes impairs amikacin in killing S. aureus.

Amikacin↗
PubMed1994

Taxonomic position of the rickettsiae: current knowledge.

The term rickettsiae initially encompassed all intracellular bacteria. Early rickettsial taxonomy was based on a comparison of a few phenotypic characteristics and recently, molecular studies brought new bases for rickettsial taxonomy. All rickettsial species studied so far belong to the alpha and gamma groups of the Proteobacteria. Ehrlichiae complex groups Cowdria ruminantium, Anaplasma marginale and Wolbachia pipientis and the related parthenogenesis and cytoplasmic incompatibility bacteria, whereas Rochalimaea species group with Bartonella bacilliformis. Rickettsia tsutsugamushi may form an independent lineage, whereas molecular data allow to regroup serologically defined typhus and spotted fever group rickettsiae. The true scale of Rickettsia and Coxiella genera remain to be determined.

Alphaproteobacteria↗
PubMed1994

Q fever serology: cutoff determination for microimmunofluorescence.

Q fever, a worldwide zoonosis caused by Coxiella burnetii, lacks clinical specificity and may present as acute or chronic disease. Because of this polymorphism, serological confirmation is necessary to assess the diagnosis. Although microimmunofluorescence is our reference technique, the cutoff titers that are currently used to make a diagnosis of active or chronic Q fever were determined years ago with limited series of patients and sera. We determined the titers of immunoglobulin G (IgG), IgM, and IgA against both phases (I and II) of Coxiella burnetii. Rheumatoid factor was removed before testing IgM and IgA. We report here the various cutoff titers and the kinetics of antibody development from 2,218 first serum samples of patients, among whom 208 suffered from acute Q fever and 53 had chronic Q fever. In active Q fever, we have defined a low cutoff (phase II IgG titer < or = 100) below which the diagnosis cannot be made and would need further confirmation and confirmed a high cutoff (phase II IgG titer > or = 200 and phase II IgM titer > or = 50) over which the diagnosis can be made. For chronic Q fever diagnosis, phase I IgA titers are not contributive despite previous works claiming their usefulness; a phase I IgG titer of > or = 800 is highly predictive (98%) and sensitive (100%). We have also studied the possibility of rejecting or evoking the diagnosis of chronic Q fever by phase II IgG and IgA titers. This method is useful when phase I testing is not available, but the sensitivity remains low (57%).

Acute Disease↗
PubMed1994

Serological response of patients suffering from primary and recrudescent typhus: comparison of complement fixation reaction, Weil-Felix test, microimmunofluorescence, and immunoblotting.

Microimmunofluorescence and Western immunoblotting were compared with the classical complement fixation reaction and the Weil-Felix test to study the serological responses of patients to Rickettsia prowazekii and both Proteus vulgaris OX19 and OX2 during primary and recrudescent typhus infections. The serological response to R. prowazekii was found to be similar during primary and recrudescent typhus, and all sera examined contained antibodies to the same R. prowazekii cell structures. Immunoglobulin G (IgG) and IgM were found to be the dominant anti-R. prowazekii immunoglobulins in all sera tested and were found to be directed against the 100-kDa protein and the lipopolysaccharide. IgA antibodies, when present, were mainly against the 100-kDa protein. For P. vulgaris, IgG antibodies recognized the proteins and lipopolysaccharides of both OX19 and OX2 serotypes; IgM antibodies were directed against the P. vulgaris OX2 lipopolysaccharide. In addition, donor blood sera, which were negative by microimmunofluorescence, were found to contain IgG immunoglobulins reacting with R. prowazekii protein antigens of 135, 60, and 47 kDa by western immunoblotting.

Adult↗
PubMed1994

Serologic diagnosis of human monocytic ehrlichiosis by immunoblot analysis.

Human monocytic ehrlichiosis is caused by Ehrlichia chaffeensis, an intracellular bacterium probably transmitted by the tick Amblyomma americanum in the United States. Despite its lack of specificity in discriminating among infections by closely related Ehrlichia spp., immunofluorescence assay (IFA) is the most frequently used serological diagnostic method. To improve the specificity of the serological diagnosis, we compared antigenic profile of E. canis and E. chaffeensis antigen with homologous and heterologous sera, searching for the specificity of the presence of low-molecular-weight proteins. Western immunoblot analysis of IFA-positive human sera revealed 27- and 29-kDa proteins which are not found in E. canis IFA-positive sera from dogs. IFA-positive sera from dogs revealed a low-molecular-weight group of proteins (20 to 28 kDa) which were not found in human E. chaffeensis-positive sera except for a weak band at 22 kDa. The presence o antibodies directed against the 27- and 29-kDa proteins on Western blots is specific for E. chaffeensis infection, and we suggest that the Western blot might complete IFA in cases with low positive predictive value.

Animals↗
PubMed1994

Cytopathic effect, plaque formation, and lysis of Ehrlichia chaffeensis grown on continuous cell lines.

Ehrlichiae are strict intracellular bacterial pathogens that parasitize leukocytes or other blood cells. Only six agents of the tribe Ehrlichieae, namely, Cowdria ruminantium, Neorickettsia helminthoeca, Ehrlichia risticii, Ehrlichia sennetsu, Ehrlichia canis, and Ehrlichia chaffeensis, have been adapted to growth in continuous cell lines. E. chaffeensis, the agent of human ehrlichiosis, has been cultured only in a cell line of canine origin. We adapted purified cell-free E. chaffeensis for growth in human embryonic lung (HEL) fibroblasts (HEL 299), green monkey kidney cells (Vero), and a human cervical epithelioid carcinoma (HeLa) cell line. We observed a cytopathic effect with both Vero cells and HEL cells and plaque formation with cellular lysis when infected Vero cells were cultured in agar. Human fibroblasts are already commonly used for the isolation of viruses, coexiellae, and rickettsiae. Furthermore, the capability of these cells to support the growth of ehrlichiae suggests that they may be useful for primary isolation of ehrlichiae as well. The cytopathic effect produced in Vero or HEL cells offers a very helpful indicator of the infection. Plaque formation in Vero cells is a new phenomenon not yet reported for ehrlichiae and will allow the titration of inocula and clonal purification of this bacterium.

Adaptation, Physiological↗
PubMed1994

Differentiation among spotted fever group rickettsiae species by analysis of restriction fragment length polymorphism of PCR-amplified DNA.

Restriction fragment length polymorphism (RFLP) analysis of PCR-amplified genes was used to study spotted fever group (SFG) rickettsiae, extending the previous work of Regnery et al. (R.L. Regnery, C.L. Spruill, and B.D. Plikaytis, J. Bacteriol. 173:1576-1589, 1991). Twenty-six strains of SFG rickettsia were studied, including several recognized species which have never been studied (R. parkeri, R. helvetica, and R. japonica) as well as strains which are not currently classified. Two previously used primer pairs derived from the R. prowazekii citrate syntase gene and the R. rickettsii 190-kDa protein antigen gene were studied, as were primer pairs obtained from the R. rickettsii 120-kDa protein antigen gene. By using three amplifications and three enzyme digestions, it was possible to differentiate between almost all of the known SFG rickettsia species and to differentiate between several strains of the R. conorii complex. Two human pathogens, "R. africae" and the Israeli tick typhus rickettsia, were first separated by using BG-12 pair primer amplification and then RsaI restriction endonuclease digestion. The proposed simplified model of identification may be useful in studying the geographical distributions of SFG rickettsiae.

Animals↗
PubMed1994

Isolation and characterization by immunofluorescence, sodium dodecyl sulfate-polyacrylamide gel electrophoresis, western blot, restriction fragment length polymorphism-PCR, 16S rRNA gene sequencing, and pulsed-field gel electrophoresis of Rochalimaea quintana from a patient with bacillary angiomatosis.

Rochalimaea quintana was isolated from the blood of a French human immunodeficiency virus-infected patient with bacillary angiomatosis. The isolate showed the typical growth characteristics of Rochalimaea species and was inert when typical biochemical testing was used. The purpose of the present work was to characterize and compare this new isolate with reference strains of R. quintana, Rochalimaea vinsonii, and Rochalimaea henselae by using immunofluorescence, sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), Western blot (immunoblot), restriction fragment length polymorphism-PCR of the citrate synthase gene, 16S rRNA gene sequencing, and pulsed-field gel electrophoresis. SDS-PAGE, Western blot, restriction fragment length polymorphism-PCR with TaqI enzyme, and 16S rRNA gene sequencing could differentiate the three Rochalimaea species and allowed characterization of the French isolate as R. quintana. However, identification of the Rochalimaea isolate to the species level was more easily obtained by immunofluorescence with specific murine antisera. Pulsed-field gel electrophoresis allowed differentiation of the French R. quintana isolate from R. quintana Fuller and may serve as an epidemiological tool.

AIDS-Related Opportunistic Infections↗
PubMed1994

Chronic Q fever in hemodialysis patients.

We describe two cases of chronic Q fever in hemodialysis patients (HD). In the first case, we discovered chronic Q fever when looking for the cause of an unexplained fever. In the second case, Q fever was diagnosed in a patient who complained of an unexplained shoulder arthritis. To our knowledge these are the first reported cases of chronic Q fever in HD.

Adult↗
PubMed1994

Isolation and identification of a rickettsial strain related to Rickettsia massiliae in Greek ticks.

Adult ticks were collected in a rural area of central Greece in order to isolate and identify rickettsiae. A hemolymph test using Gimenez staining was used for detection, while simultaneous isolation was performed using the shell-vial technique. Serologic, antigenic, and genomic characterization of the isolates was achieved by microimmunofluorescence, sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and Western blotting, the polymerase chain reaction and restriction fragment length polymorphism (PCR-RFLP), and pulsed-field gel electrophoresis (PFGE), respectively. Although none of the 242 collected ticks was positive by the hemolymph test, one rickettsial isolate, designated GS, was obtained by the shell-vial technique. This isolate originated from a female Rhipicephalus sanguineus. Microimmunofluorescence serologic typing by the method of Philip and others demonstrated that GS belongs to the same serotype as the recently isolated Rickettsia massiliae (Mtu1). Protein analysis by SDS-PAGE and immunoblotting by Western blot revealed similar profiles between the two rickettsiae. Using Alu I, Rsa I, and Pst I restriction endonucleases in PCR-RFLP analysis, GS and R. massiliae were found to possess identical restriction sites. However, PFGE showed differences when the two genomes were digested with Bss HII and Sma I restriction endonucleases, in spite of their equal size. In conclusion, the first rickettsial isolation in Greece was found to be antigenically identical and genotypically close to the French isolate R. massiliae, despite small differences showed by PFGE.

Animals↗
PubMed1994

Identification of rickettsiae from ticks collected in the Central African Republic using the polymerase chain reaction.

Spotted fever rickettsiosis have been identified on the African continent since their historical description in 1909. However, only Rickettsia conorii and R. africae have been described in Africa, and the current techniques for the detection of rickettsiae in ticks are difficult to apply in large field studies. We report here a preliminary study using genomic amplification by the polymerase chain reaction followed by restriction fragment length polymorphism (PCR-RFLP) analysis directly on 310 crushed ticks (Rhipicephalus, Amblyomma, and Haemaphysalis species) collected in 1985 in the Central African Republic. Among 310 specimen tested, 21.6% were positive. The rate of infection ranged from 0% to 64.3%, depending on the tick species. Based on PCR-RFLP, five different rickettsiae profiles were found: R. conorii and R. africae, previously known in Africa, R. rhipicephali, which has never been described in Africa, and two isolates identical to R. massiliae and Mtu5, previously obtained from Rh. turanicus in southern France. This work shows that PCR-RFLP is a powerful tool to study tick collections, and that it is applicable to samples from developing countries. Further work is needed to confirm the identification of the rickettsiae found in this work, using traditional identification procedures.

Animals↗
PubMed1994

Identification of spotted fever group rickettsiae isolated from Dermacentor marginatus and Ixodes ricinus ticks collected in Switzerland.

When 155 ticks collected in different regions of Switzerland were tested by the hemolymph test, 10.3% were found to contain spotted fever group rickettsiae. Six rickettsial isolates were made from Dermacentor marginatus ticks and three were made from Ixodes ricinus ticks. The polymerase chain reaction followed by restriction fragment length polymorphism analysis showed that the Dermacentor ticks were infected with Rickettsia length polymorphism analysis showed that the Dermacentor ticks were infected with Rickettsia slovaca and the Ixodes ticks were infected with a spotted fever group rickettsia. Microimmunofluorescence serologic type, sodium dodecyl sulfate-polyacrylamide gel electrophoresis of proteins, and Western blot assay with polyclonal mouse antisera confirmed the results and determined that the Ixodes were infected with R. helvetica, the only previously described Swiss rickettsia. However, an additional new strain that could not be isolated was detected in one I. ricinus by hemolymph test and provisionally characterized by enzymatic restriction of its amplified DNA.

Animals↗
PubMed1994

Astrakhan fever rickettsiae: antigenic and genotypic analysis of isolates obtained from human and Rhipicephalus pumilio ticks.

Two spotted fever group rickettsia strains, A-108 and A-167, were isolated from the hemolymph of Rhipicephalus pumilio ticks collected in the Astrakhan region of Russia, which is area endemic for Astrakhan fever. These tick isolates were compared with a strain isolated from a patient suffering from Astrakhan fever and with reference spotted fever group rickettsiae strains. New tick isolates and the human strain were identical in their serologic, antigenic, and genetic characteristics by several methods: microimmunofluorescence, protein gel electrophoresis with immunoblotting, polymerase chain reaction followed by restriction endonuclease fragment length polymorphism analysis, and pulsed-field gel electrophoresis (PFGE). Astrakhan fever rickettsiae were found to be serologically and antigenically similar to Israeli spotted fever rickettsiae. Both of them probably belong to a single Rickettsia conorii pathotype complex. Only PFGE pattern analysis could clearly discriminate Astrakhan fever rickettsiae from other isolates.

Animals↗
PubMed1994

Further evidence for the efficacy of imidocarb dipropionate in the treatment of Ehrlichia canis infection.

Three dogs experimentally infected with Ehrlichia canis developed thrombocytopaenia and high antibody titres to E. canis in indirect fluorescent antibody tests. One dog also became leukopaenic. At Weeks 6 and 8 post-infection, the dogs were treated with imidocarb dipropionate (5 mg kg-1 subcutaneously) and a further dose was administered at Week 12 (5 mg kg-1 intramuscularly). Twelve weeks after the last treatment (post-treatment), all dogs had normal platelet counts which persisted for a further 10 weeks until the end of the experiment. The leukopaenia resolved 20 weeks post-treatment. Although antibody titres (< 1/5) to E. canis could not be detected prior to infection, titres of 1/2 560 to 1/5 120 developed by Week 6. By Week 8 post-treatment titres began to decline and by the end of the experiment were 5- to 6-fold serum dilutions lower (1/80 to 1/320). Sub-inoculation experiments 18 weeks post-treatment, failed to cause disease or stimulate antibody responses in susceptible dogs. Serology and sub-inoculation studies on 2 dogs experimentally infected with E. canis but not treated with imidocarb dipropionate, showed that these animals remained infected for the duration of the experiment. The results of these experiments confirm that imidocarb dipropionate is effective in the treatment of canine ehrlichiosis.

Animals↗
PubMed1994

Genomic identification of Rickettsia slovaca among spotted fever group rickettsia isolates from Dermacentor marginatus in Armenia.

Restriction fragment length polymorphism (RFLP) analysis of polymerase chain reaction (PCR) amplified genes was used for genomic identification of Armenian isolates of the Spotted fever group (SFG) rickettsiae with unclear taxonomic position. Analysis was performed by using one genus-specific primer pair derived from R. prowazekii citrate synthase gene and two species-specific primer pairs derived from R. rickettsii genes for 190 K and 120 K antigens following AluI, PstI and RsaI digestion of amplicons. All tested rickettsial SFG Armenian isolates from Dermacentor marginatus were identified as R. slovaca. The geographic distribution and genetic homogeneity of R. slovaca strains are discussed.

Animals↗
PubMed1994

Purification of rickettsial cultures contaminated by mycoplasmas.

An experimental biological model is proposed for purification of rickettsiae-infected cell cultures from mycoplasma contamination using intravenous mouse infection and subsequent passages on Vero cell monolayers. Mycoplasma-free rickettsial cultures were obtained in mouse brain or spleen suspension within 3 hrs or 3 days, respectively, after mouse inoculation. Ten strains of spotted fever group rickettsiae were purified from mycoplasmas by this procedure.

Animals↗