The genome sequence of Rickettsia felis identifies the first putative conjugative plasmid in an obligate intracellular parasite.
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ABSTRACT:
SUBMITTER: Ogata H
PROVIDER: S-EPMC1166351 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:The genus Rickettsia (Alphaproteobacteria, Rickettsiales, Rickettsiaceae) is comprised of obligate intracellular parasites, with virulent species of interest both as causes of emerging infectious diseases and for their potential deployment as bioterrorism agents. Currently, there are no effective commercially available vaccines, with treatment limited primarily to tetracycline antibiotics, although others (e.g. josamycin, ciprofloxacin, chloramphenicol, and azithromycin) are also effective. Much of the recent research geared toward understanding mechanisms underlying rickettsial pathogenicity has centered on characterization of secreted proteins that directly engage eukaryotic cells. Herein, we review all aspects of the Rickettsia secretome, including six secretion systems, 19 characterized secretory proteins, and potential moonlighting proteins identified on surfaces of multiple Rickettsia species. Employing bioinformatics and phylogenomics, we present novel structural and functional insight on each secretion system. Unexpectedly, our investigation revealed that the majority of characterized secretory proteins have not been assigned to their cognate secretion pathways. Furthermore, for most secretion pathways, the requisite signal sequences mediating translocation are poorly understood. As a blueprint for all known routes of protein translocation into host cells, this resource will assist research aimed at uniting characterized secreted proteins with their apposite secretion pathways. Furthermore, our work will help in the identification of novel secreted proteins involved in rickettsial 'life on the inside'.
| S-EPMC4344940 | biostudies-literature
Project description:BackgroundSince its first detection, characterization of R. felis has been a matter of debate, mostly due to the contamination of an initial R. felis culture by R. typhi. However, the first stable culture of R. felis allowed its precise phenotypic and genotypic characterization, and demonstrated that this species belonged to the spotted fever group rickettsiae. Later, its genome sequence revealed the presence of two forms of the same plasmid, physically confirmed by biological data. In a recent article, Gillespie et al. (PLoS One. 2007;2(3):e266.) used a bioinformatic approach to refute the presence of the second plasmid form, and proposed the creation of a specific phylogenetic group for R. felis.Methodology/principal findingsIn the present report, we, and five independent international laboratories confirmed unambiguously by PCR the presence of two plasmid forms in R. felis strain URRWXCal(2) (T), but observed that the plasmid content of this species, from none to 2 plasmid forms, may depend on the culture passage history of the studied strain. We also demonstrated that R. felis does not cultivate in Vero cells at 37 degrees C but generates plaques at 30 degrees C. Finally, using a phylogenetic study based on 667 concatenated core genes, we demonstrated the position of R. felis within the spotted fever group.SignificanceWe demonstrated that R. felis, which unambiguously belongs to the spotted fever group rickettsiae, may contain up to two plasmid forms but this plasmid content is unstable.
| S-EPMC2384004 | biostudies-literature
Project description:Rickettsia conorii are obligate intracellular bacteria responsible for the Mediterranean spotted fever. Their adaptation to highly divergent environments ranging from the arthropod vector to the human beings is essential to survival and virulence. Through a combination of total RNA purification and random prokaryotic cDNA amplification, we successfully analyzed transcription profiles by microarrays starting from 100 ng of R. conorii RNA. Real-time quantitative PCR results performed on unpurified total RNA was consistent with microarray measurements. Here, from the selected targets spotted on our microarray, we showed that in R. conorii, nutrient stress is accompanied by over-expression of the virB operon. Regulation of these genes could be mediated by the stringent response through ppGpp, consistent with the observed up-regulation of spoT and gmk. Exposure of R. conorii to a nutrient stress paradoxically reduced the expression of both GroEL and its transcriptional regulator RpoH. This shows that the over-expression of this chaperonin is a part of the natural life of intracellular growing rickettsiae. Our findings also evidenced that, unexpectedly, many atypical sequences, including small-size split genes, ORFans genes and highly conserved intergenic regions contains expressed sequences that are differentially regulated. The notable deficiency of transcriptional regulators within R. conorii genome does not reflect its incapability to undergo transcriptional changes but rather the existence of an alternative adaptation strategy. Keywords: Global transcriptional analysis
2006-07-06 | GSE5201 | GEO
Project description:BackgroundRickettsia felis is a recently described flea-borne spotted fever group Rickettsia that is an emerging human pathogen. Although there is information on the organism from around the world, there is no information on the organism in China.MethodsWe used a commercial ELISA to detect antibodies reactive against R. felis in blood samples and developed a PCR to detect the gltA of the organism in blood samples and external parasites.ResultsWe found reactive antibodies in people (16%; 28/180), dogs (47%; 128/271) and cats (21%; 19/90) and positive PCRs with DNA from people (0.1%; 1/822), dogs (0.8%; 8/1,059), mice (10%; 1/10), ticks (Rhipicephalus sanguineus; 10%; 15/146), lice (Linognathus setosus; 16%; 6/37), fleas (Ctenocephalides felis felis; 95%; 57/60) and mosquitoes (Anopheles sinensis, Culex pipiens pallens; 6%; 25/428), but not from cats (0/135) or canine fecal swabs (0/43).ConclusionsThis is the first report of R. felis in China where there is serological and/ or PCR evidence of the organism in previously reported [people, dogs, cats, ticks (Rhipicephalus sanguineus), fleas (Ctenocephalides felis felis) and mosquitoes (Anopheles sinensis, Culex pipiens pallens)] and novel species [mice and lice (Linognathus setosus)].
| S-EPMC4297373 | biostudies-literature
Project description:Genetic analysis of Rickettsia prowazekii has been hindered by the lack of selectable markers and efficient mechanisms for generating rickettsial gene knockouts. We have addressed these problems by adapting a gene that codes for rifampin resistance for expression in R. prowazekii and by incorporating this selection into a transposon mutagenesis system suitable for generating rickettsial gene knockouts. The arr-2 gene codes for an enzyme that ADP-ribosylates rifampin, thereby destroying its antibacterial activity. Based on the published sequence, this gene was synthesized by PCR with overlapping primers that contained rickettsial codon usage base changes. This R. prowazekii-adapted arr-2 gene (Rparr-2) was placed downstream of the strong rickettsial rpsL promoter (rpsL(P)), and the entire construct was inserted into the Epicentre EZ::TN transposome system. A purified transposon containing rpsL(P)-Rparr-2 was combined with transposase, and the resulting DNA-protein complex (transposome) was electroporated into competent rickettsiae. Following selection with rifampin, rickettsiae with transposon insertions in the genome were identified by PCR and Southern blotting and the insertion sites were determined by rescue cloning and inverse PCR. Multiple insertions into widely spaced areas of the R. prowazekii genome were identified. Three insertions were identified within gene coding sequences. Transposomes provide a mechanism for generating random insertional mutations in R. prowazekii, thereby identifying nonessential rickettsial genes.
| S-EPMC404435 | biostudies-literature
Project description:BackgroundCompleted genome sequences are rapidly increasing for Rickettsia, obligate intracellular alpha-proteobacteria responsible for various human diseases, including epidemic typhus and Rocky Mountain spotted fever. In light of phylogeny, the establishment of orthologous groups (OGs) of open reading frames (ORFs) will distinguish the core rickettsial genes and other group specific genes (class 1 OGs or C1OGs) from those distributed indiscriminately throughout the rickettsial tree (class 2 OG or C2OGs).Methodology/principal findingsWe present 1823 representative (no gene duplications) and 259 non-representative (at least one gene duplication) rickettsial OGs. While the highly reductive (approximately 1.2 MB) Rickettsia genomes range in predicted ORFs from 872 to 1512, a core of 752 OGs was identified, depicting the essential Rickettsia genes. Unsurprisingly, this core lacks many metabolic genes, reflecting the dependence on host resources for growth and survival. Additionally, we bolster our recent reclassification of Rickettsia by identifying OGs that define the AG (ancestral group), TG (typhus group), TRG (transitional group), and SFG (spotted fever group) rickettsiae. OGs for insect-associated species, tick-associated species and species that harbor plasmids were also predicted. Through superimposition of all OGs over robust phylogeny estimation, we discern between C1OGs and C2OGs, the latter depicting genes either decaying from the conserved C1OGs or acquired laterally. Finally, scrutiny of non-representative OGs revealed high levels of split genes versus gene duplications, with both phenomena confounding gene orthology assignment. Interestingly, non-representative OGs, as well as OGs comprised of several gene families typically involved in microbial pathogenicity and/or the acquisition of virulence factors, fall predominantly within C2OG distributions.Conclusion/significanceCollectively, we determined the relative conservation and distribution of 14354 predicted ORFs from 10 rickettsial genomes across robust phylogeny estimation. The data, available at PATRIC (PathoSystems Resource Integration Center), provide novel information for unwinding the intricacies associated with Rickettsia pathogenesis, expanding the range of potential diagnostic, vaccine and therapeutic targets.
| S-EPMC2635572 | biostudies-literature
Project description: Not available
| S-EPMC7301703 | biostudies-literature
Project description:The present study evaluated the rickettsial infection in a laboratory colony of cat fleas, Ctenocephalides felis felis (Bouche) in Brazil. All flea samples (30 eggs, 30 larvae, 30 cocoons, 30 males, and 30 females) tested by polymerase chain reaction (PCR) were shown to contain rickettsial DNA. PCR products, corresponding to the rickettsial gltA, htrA, ompA and ompB gene partial sequences were sequenced and showed to correspond to Rickettsia felis, indicating that the flea colony was 100% infected by R. felis. The immunofluorescence assay (IFA) showed the presence of R. felis-reactive antibodies in blood sera of 7 (87.5%) out of 8 cats that were regularly used to feed the flea colony. From 15 humans that used to work with the flea colony in the laboratory, 6 (40.0%) reacted positively to R. felis by IFA. Reactive feline and human sera showed low endpoint titers against R. felis, varying from 64 to 256. With the exception of one human serum, all R. felis-reactive sera were also reactive to Rickettsia rickettsii and/or Rickettsia parkeri antigens at similar titers to R. felis. The single human serum that was reactive solely to R. felis had an endpoint titer of 256, indicating that this person was infected by R. felis.
| S-EPMC3768654 | biostudies-literature
Project description:Termination of transcription is an important component of bacterial gene expression. However, little is known concerning this process in the obligate intracellular pathogen and model for reductive evolution, Rickettsia prowazekii. To assess transcriptional termination in this bacterium, transcripts of convergent gene pairs, some containing predicted intrinsic terminators, were analyzed. These analyses revealed that, rather than terminating at a specific site within the intervening region between the convergent genes, most of the transcripts demonstrated either a lack of termination within this region, which generated antisense RNA, or a putative non-site-specific termination that occurred throughout the intervening sequence. Transcripts terminating at predicted intrinsic terminators, as well as at a putative Rho-dependant terminator, were also examined and found to vary based on the rickettsial host environment. These results suggest that transcriptional termination, or lack thereof, plays a role in rickettsial gene regulation.
| S-EPMC3027695 | biostudies-literature
Project description:Conjugative plasmid transfer is the most important route for the spread of resistance and virulence genes among bacteria. Consequently, bacteria carrying conjugative plasmids are a substantial threat to human health, especially hospitalized patients. Whilst detailed information about the process has been obtained for Gram-negative type-4 secretion systems, little is known about the corresponding mechanisms in Gram-positive (G+) bacteria. The successful purification and crystallization of the putative transfer protein TraN from the G+ conjugative model plasmid pIP501 of Enterococcus faecalis are presented. Native crystals diffracted to 1.8?Å resolution on a synchrotron beamline. The crystals belonged to space group P2(1), with unit-cell parameters a=32.88, b=54.94, c=57.71?Å, ?=91.89° and two molecules per asymmetric unit.
| S-EPMC3515391 | biostudies-literature