Guidelines for the Detection ofRickettsiaspp.

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Abstract

The genus Rickettsia (Rickettsiales: Rickettsiaceae) includes Gram-negative, small, obligate intracellular, nonmotile, pleomorphic coccobacilli bacteria transmitted by arthropods. Some of them cause human and probably also animal disease (life threatening in some patients). In these guidelines, we give clinical practice advices (microscopy, serology, molecular tools, and culture) for the microbiological study of these microorganisms in clinical samples. Since in our environment rickettsioses are mainly transmitted by ticks, practical information for the identification of these arthropods and for the study of Rickettsia infections in ticks has also been added.

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Most cited references 38

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Genotypic identification of rickettsiae and estimation of intraspecies sequence divergence for portions of two rickettsial genes.

R Regnery, C L Spruill, B D Plikaytis (1991)

DNA sequences from specific genes, amplified by the polymerase chain reaction technique, were used as substrata for nonisotopic restriction endonuclease fragment length polymorphism differentiation of rickettsial species and genotypes. The products amplified using a single pair of oligonucleotide primers (derived from a rickettsial citrate synthase gene sequence) and cleaved with restriction endonucleases were used to differentiate almost all recognized species of rickettsiae. A second set of primers was used for differentiation of all recognized species of closely related spotted fever group rickettsiae. The procedure circumvents many technical obstacles previously associated with identification of rickettsial species. Multiple amplified DNA digest patterns were used to estimate the intraspecies nucleotide sequence divergence for the genes coding for rickettsial citrate synthase and a large antigen-coding gene of the spotted fever group rickettsiae. The estimated relationships deduced from these genotypic data correlate reasonably well with established rickettsial taxonomic schemes.

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Current and past strategies for bacterial culture in clinical microbiology.

Jean-Christophe Lagier, Sophie Edouard, Isabelle Pagnier … (2015)

A pure bacterial culture remains essential for the study of its virulence, its antibiotic susceptibility, and its genome sequence in order to facilitate the understanding and treatment of caused diseases. The first culture conditions empirically varied incubation time, nutrients, atmosphere, and temperature; culture was then gradually abandoned in favor of molecular methods. The rebirth of culture in clinical microbiology was prompted by microbiologists specializing in intracellular bacteria. The shell vial procedure allowed the culture of new species of Rickettsia. The design of axenic media for growing fastidious bacteria such as Tropheryma whipplei and Coxiella burnetii and the ability of amoebal coculture to discover new bacteria constituted major advances. Strong efforts associating optimized culture media, detection methods, and a microaerophilic atmosphere allowed a dramatic decrease of the time of Mycobacterium tuberculosis culture. The use of a new versatile medium allowed an extension of the repertoire of archaea. Finally, to optimize the culture of anaerobes in routine bacteriology laboratories, the addition of antioxidants in culture media under an aerobic atmosphere allowed the growth of strictly anaerobic species. Nevertheless, among usual bacterial pathogens, the development of axenic media for the culture of Treponema pallidum or Mycobacterium leprae remains an important challenge that the patience and innovations of cultivators will enable them to overcome.

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Phylogenetic analysis of members of the genus Rickettsia using the gene encoding the outer-membrane protein rOmpB (ompB).

Ana V. Diez Roux, D Raoult (2000)

To confirm the phylogenetic analysis previously inferred by comparison of the citrate synthase and rOmpA gene sequences (gitA and ompA, respectively), the rOmpB gene (ompB) of 24 strains of the genus Rickettsia was amplified and sequenced. rOmpB is an outer-membrane protein of high molecular mass, the presence of which can be demonstrated in most rickettsiae by immunological cross-reactivity in Western blots. No PCR amplification was obtained with Rickettsia bellii or Rickettsia canadensis. For the other rickettsiae, phylogenetic analysis was inferred from the comparison of both the gene and derived protein sequences by using parsimony, maximum-likelihood and neighbour-joining methods which gave the same organization. All nodes were well supported (>86% bootstrap values), except in the cluster including Rickettsia africae strain S and Rickettsia parkeri, and this analysis confirmed the previously established phylogeny obtained from combining results from gltA and ompA. Based on phylogenetic data, the current classification of the genus Rickettsia is inappropriate, specifically its division into two groups, typhus and spotted fever. Integration of phenotypic, genotypic and phylogenetic data will contribute to the definition of a polyphasic taxonomy as has been done for other bacterial genera.