Tryptose phosphate broth improves<i>Rickettsia felis</i>replication in mammalian cells: 1

Saisongkorh, Watcharee; Karkouri, Khalid El; Patrice, Jean-Yves; Bernard, Annick; Rolain, Jean‐Marc; Raoult, Didier

doi:10.1111/j.1574-695x.2011.00882.x

Cited by 8 publications

(7 citation statements)

References 16 publications

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“…Supplementation of cytosine in cells enhanced Ehrlichia replication, which further implicates the use of the salvage pathway through cytidine. The enhanced growth of another rickettsia, Rickettsia felis, from pyrimidines in tryptose phosphate broth is also consistent with the involvement of mitochondrial enzymes in obligate intracellular bacterial growth (50). Moreover, cytidine is the least abundant nucleoside in cells (32).…”

Section: Discussionmentioning

confidence: 53%

Identification of Critical Host Mitochondrion-Associated Genes during Ehrlichia chaffeensis Infections

et al. 2012

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bEhrlichia chaffeensis is an obligate intracellular bacterium that causes human monocytic ehrlichiosis (HME). To determine what host components are important for bacterial replication, we performed microarray analysis on Drosophila melanogaster S2 cells by comparing host gene transcript levels between permissive and nonpermissive conditions for E. chaffeensis growth. Five-hundred twenty-seven genes had increased transcript levels unique to permissive growth conditions 24 h postinfection. We screened adult flies that were mutants for several of the "permissive" genes for the ability to support Ehrlichia replication. Three additional D. melanogaster fly lines with putative mutations in pyrimidine metabolism were also tested. Ten fly lines carrying mutations in the genes CG6479, separation anxiety, chitinase 11, CG6364 (Uck2), CG6543 (Echs1), withered (whd), CG15881 (Ccdc58), CG14806 (Apop1), CG11875 (Nup37), and dumpy (dp) had increased resistance to infection with Ehrlichia. Analysis of RNA by quantitative real-time reverse transcription-PCR (qRT-PCR) confirmed that the bacterial load was decreased in these mutant flies compared to wild-type infected control flies. Seven of these genes (san, Cht11, Uck2, Echs1, whd, Ccdc58, and Apop1) encoded proteins that had mitochondrial functions or could be associated with proteins with mitochondrial functions. Treatment of THP-1 cells with double-stranded RNA to silence the human UCK2 gene indicates that the disruption of the uridine-cytidine kinase affects E. chaffeensis replication in human macrophages. Experiments with cyclopentenyl cytosine (CPEC), a CTP synthetase inhibitor and cytosine, suggest that the nucleotide salvage pathway is essential for E. chaffeensis replication and that it may be important for the provision of CTP, uridine, and cytidine nucleotides. Ehrlichia chaffeensis is the causative agent of human monocytic ehrlichiosis (HME). There were 1,429 cases of HME in 2010 and 2011 (14). This represents a significant increase in the incidence of the disease since 2003 and qualifies HME as an emerging infectious disease (25). In addition to being reported in the United States, HME has also been documented in Africa, Europe, China, and Brazil (9,16,42,67). E. chaffeensis is an obligate intracellular bacterium. However, little is known about the parasitized-host requirements for bacterial replication.Drosophila melanogaster has been used to study a variety of intracellular pathogens. In particular, it has been successfully manipulated for the identification of genes involved in host-pathogen interactions. These pathogens include Listeria monocytogenes (1, 2), Chlamydia trachomatis (20), Mycobacterium marinum (1, 18, 30, 48), Francisella tularensis (51, 64), and the protozoan parasite Plasmodium gallinaceum (8,53).We previously demonstrated that E. chaffeensis is capable of infecting, completing its life cycle, and maintaining its pathogenicity in both Drosophila S2 cells (39) and adult flies (40). We have also identified growth conditions that were nonpermissiv...

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Section: Discussionmentioning

confidence: 53%

Identification of Critical Host Mitochondrion-Associated Genes during Ehrlichia chaffeensis Infections

et al. 2012

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“…A confluent monolayer of the Xenopus laevis cell line (XTC-2 cells) in Leibovitz's L-15 medium supplemented with tryptose phosphate buffer (5%) and fetal bovine serum (4%; Life Technologies) was inoculated with a Rickettsia species, as described by Saisongkorh et al (2012). For each studied Rickettsia species, about 25 × 10 4 –5 × 10 5 bacteria were inoculated in X. laevis cells (one 150 cm 2 flask each, containing 25 ml of fresh medium) and incubated at 28°C for 5 days.…”

Section: Methodsmentioning

confidence: 99%

Multi-omics Analysis Sheds Light on the Evolution and the Intracellular Lifestyle Strategies of Spotted Fever Group Rickettsia spp.

et al. 2017

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Arthropod-borne Rickettsia species are obligate intracellular bacteria which are pathogenic for humans. Within this genus, Rickettsia slovaca and Rickettsia conorii cause frequent and potentially severe infections, whereas Rickettsia raoultii and Rickettsia massiliae cause rare and milder infections. All four species belong to spotted fever group (SFG) rickettsiae. However, R. slovaca and R. raoultii cause scalp eschar and neck lymphadenopathy (SENLAT) and are mainly associated with Dermacentor ticks, whereas the other two species cause Mediterranean spotted fever (MSF) and are mainly transmitted by Rhipicephalus ticks. To identify the potential genes and protein profiles and to understand the evolutionary processes that could, comprehensively, relate to the differences in virulence and pathogenicity observed between these four species, we compared their genomes and proteomes. The virulent and milder agents displayed divergent phylogenomic evolution in two major clades, whereas either SENLAT or MSF disease suggests a discrete convergent evolution of one virulent and one milder agent, despite their distant genetic relatedness. Moreover, the two virulent species underwent strong reductive genomic evolution and protein structural variations, as well as a probable loss of plasmid(s), compared to the two milder species. However, an abundance of mobilome genes was observed only in the less pathogenic species. After infecting Xenopus laevis cells, the virulent agents displayed less up-regulated than down-regulated proteins, as well as less number of identified core proteins. Furthermore, their similar and distinct protein profiles did not contain some genes (e.g., ompA/B and rickA) known to be related to rickettsial adhesion, motility and/or virulence, but may include other putative virulence-, antivirulence-, and/or disease-related proteins. The identified evolutionary forces herein may have a strong impact on intracellular expressions and strategies in these rickettsiae, and that may contribute to the emergence of distinct virulence and diseases in humans. Thus, the current multi-omics data provide new insights into the evolution and fitness of SFG virulence and pathogenicity, and intracellular pathogenic bacteria.

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“…R. felis was cultured in a XTC-2 cell line at 28 °C in Leibovitz-15 media (GIBCO, Rockville, MD), supplemented with 10% foetal calf serum, 2 mM l -glutamine and 5% tryptose phosphate broth [13]. R. typhi was cultured in a L929 cell line in RPMI media (GIBCO, Rockville, MD), supplemented with 10% foetal calf serum and 2 mM l -glutamine.…”

Section: Methodsmentioning

confidence: 99%

Evidence of exposure to Rickettsia felis in Australian patients

et al. 2016

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Rickettsia felis is an emerging zoonosis, causing flea-borne spotted fever (FBSF). Serological diagnosis is typically confounded by cross-reactivity with typhus group rickettsiae and prior to the development of specific serological methods, cases of FBSF in Australia were misdiagnosed.Patient sera tested between August 2010 and December 2013 and known to be seropositive to R. typhi by immunofluorescence antibody testing (IFAT) were subsequently retested against R. felis using an R. felis-specific IFAT. Sera of 49 patients were of a sufficient quality to be included in re-analysis. A classification of FBSF and murine typhus (MT) was attributed to fourteen and seven patients respectively, based on a minimum four-fold higher antibody titre to R. felis than to R. typhi and vice versa. Twenty-eight patients were classified as indeterminate for either R. felis or R. typhi (antibody titres within two-fold of one another).Historically, it is likely that Australian patients clinically ill with FBSF were misdiagnosed. It is important that medical practitioners consider FBSF as part of their differential diagnoses, and obtain relevant history with regard to patient's exposure to domestic pets and their fleas. Australian microbiology diagnostic laboratories should include serological testing for R. felis as part of the diagnostic panel for febrile diseases. Veterinarians are encouraged to increase their awareness of this emerging zoonosis and advocate flea control in pets.

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Tryptose phosphate broth improvesRickettsia felisreplication in mammalian cells: 1

Cited by 8 publications

References 16 publications

Identification of Critical Host Mitochondrion-Associated Genes during Ehrlichia chaffeensis Infections

Identification of Critical Host Mitochondrion-Associated Genes during Ehrlichia chaffeensis Infections

Multi-omics Analysis Sheds Light on the Evolution and the Intracellular Lifestyle Strategies of Spotted Fever Group Rickettsia spp.

Evidence of exposure to Rickettsia felis in Australian patients

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