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PubMed2005

Comparative study of overlapping genes in bacteria, with special reference to Rickettsia prowazekii and Rickettsia conorii.

Overlapping genes have been proposed as a means of achieving genome reduction by compressing the maximum amount of information in limited sequence space. In this report, comparative analyses of the overlapping genes of genomes of nine bacteria with different lifestyles were performed. The results clearly suggest that overlapping genes may be a result of evolutionary pressure to minimize genome size. The genomes of two closely related obligatory intracellular parasites - Rickettsia prowazekii and Rickettsia conorii - were investigated further. Detailed analyses of these two genomes revealed that mutations at the ends of coding regions and elimination of intergenic DNA are the main forces that determine overlapping of genes.

Biological Evolution↗
PubMed1994

Nucleotide sequence of polymerase chain reaction product amplified from Rickettsia japonica DNA using Rickettsia rickettsii 190-kilodalton surface antigen gene primers.

The PCR product amplified from Rickettsia japonica with the primer pair Rr 190.70p and Rr 190.602n of R. rickettsii 190-kDa antigen gene was cloned into M13mp19 RF DNA at the Eco RI site and sequenced by chemiluminescent DNA sequencing. The sequence revealed a molecular size of 533 base pairs (bp). The primer-flanking region of 491 bp, an open reading frame, was compared with the corresponding region of R. rickettsii, demonstrating 35 nucleotide substitutions in R. japonica. The sequence of primer portions in R. japonica DNA was also analyzed, revealing one nucleotide substitution in the Rr 190.70p and two in the Rr 190.602n portion. The homology in the overall sequence of PCR-amplified regions between R. japonica and R. rickettsii was 93% in nucleotide and 85% in putative amino acid structure. The sequence contains no cleavage site for the restriction endonuclease AfaI but two PstI sites giving three fragments of 121, 159, and 253 bp, which differentiated R. japonica from other spotted fever group rickettsiae in addition to R. rickettsii. The cleavage sites for endonucleases AluI, HinfI, and MunI that disappeared or appeared in the sequence by nucleotide substitution differentiated R. japonica from others, as did PstI. The estimation of molecular size of DNA fragments on polyacrylamide gel electrophoresis is discussed.

Antigens, Bacterial↗
PubMed1993

Determination of genome size and restriction pattern polymorphism of Rickettsia prowazekii and Rickettsia typhi by pulsed field gel electrophoresis.

Pulsed field gel electrophoresis (PFGE) of SmaI, MluI and SalI digested DNA was used to estimate genome size and perform restriction fragment length polymorphism analysis for Rickettsia prowazekii and Rickettsia typhi. We concluded that the genome of R. prowazekii and R. typhi consisted of a single chromosomal DNA. The total length of DNA of R. prowazekii was 1,106 +/- 54 kb and of R. typhi was 1,133 +/- 44 kb. It was possible to differentiate two strains of R. prowazekii, Breinl and EVir, by PFGE analysis after SalI digestion. Restriction fragment length polymorphism analysis did not reveal intraspecies differences between three human isolates and one Xenopsilla cheopis isolate of R. typhi.

DNA Restriction Enzymes↗
PubMed1989

In vitro susceptibilities of Rickettsia rickettsii and Rickettsia conorii to roxithromycin and pristinamycin.

In vitro susceptibilities of Rickettsia rickettsii and Rickettsia conorii to roxithromycin, pristinamycin, and the pristinamycin compounds, P1 and P2, were determined by a dye uptake assay and a plaque assay. The MICs were 1 microgram/ml for roxithromycin, 2 micrograms/ml for pristinamycin, greater than 256 micrograms/ml for P1, and 2 micrograms/ml for P2. Compounds P1 and P2 did not share synergetic activity. The toxicity of each compound was determined by a dye uptake assay. Toxic concentrations were 128 micrograms/ml for roxithromycin, 32 micrograms/ml for pristinamycin, greater than 256 micrograms/ml for P1, and 32 micrograms/ml for P2. Roxithromycin and pristinamycin could be useful in the treatment of Rocky Mountain spotted fever and Mediterranean spotted fever.

Animals↗
PubMed1978

Biochemical characteristics of typhus group rickettsiae with special attention to the Rickettsia prowazekii strains isolated from flying squirrels.

Six strains of Rickettsia prowazekii, two derived from human infections and four isolated from flying squirrels, two strains of R. typhi, and the single available strain of R. canada, were characterized by several biochemical procedures. The electrophoretic patterns on polyacrylamide gels of rickettsial proteins solubilized by sodium dodecyl sulfate revealed several species differences, but strains of the same species appeared to have identical patterns. Cytoplasmic fractions of the rickettsiae were examined for enzymatic activities and for polyacrylamide gel isoelectric focusing patterns. Some species differences were encountered in the activities or ratios of activities of glutamate-oxaloacetate transaminase, glutamate dehydrogenase, and malate dehydrogenase. When polyacrylamide gels were stained for malate dehydrogenase after electrophoresis, a single band became apparent with single extracts or mixtures of two strains of R. prowazekii, but two bands were seen with mixtures of a strain of R. prowazekii and one of R. typhi. The isoelectric focusing patterns of the soluble proteins revealed numerous species differences, especially between R. canada and the other two species, and a few differences among the strains of R. prowazekii. The patterns of the two human strains, Breinl and E(R), differed in at least one location, and both differed from the flying squirrel strains in the displacement of one band. One of the flying squirrel strains, GvF-16, contained a protein band not seen in the other five strains. Despite these minor differences, a striking similarity was revealed by all the biochemical tests performed between the R. prowazekii strains of human and flying squirrel origin.

Animals↗
PubMed2003

Molecular and functional analysis of the lepB gene, encoding a type I signal peptidase from Rickettsia rickettsii and Rickettsia typhi.

The type I signal peptidase lepB genes from Rickettsia rickettsii and Rickettsia typhi, the etiologic agents of Rocky Mountain spotted fever and murine typhus, respectively, were cloned and characterized. Sequence analysis of the cloned lepB genes from R. rickettsii and R. typhi shows open reading frames of 801 and 795 nucleotides, respectively. Alignment analysis of the deduced amino acid sequences reveals the presence of highly conserved motifs that are important for the catalytic activity of bacterial type I signal peptidase. Reverse transcription-PCR and Northern blot analysis demonstrated that the lepB gene of R. rickettsii is cotranscribed in a polycistronic message with the putative nuoF (encoding NADH dehydrogenase I chain F), secF (encoding protein export membrane protein), and rnc (encoding RNase III) genes in a secF-nuoF-lepB-rnc cluster. The cloned lepB genes from R. rickettsii and R. typhi have been demonstrated to possess signal peptidase I activity in Escherichia coli preprotein processing in vivo by complementation assay.

Amino Acid Sequence↗
PubMed2006

Evidence of infection in humans with Rickettsia typhi and Rickettsia felis in Catalonia in the Northeast of Spain.

Murine typhus is a cause of fever of intermediate duration in the south of Spain, where antibodies against Rickettsia typhi and Rickettsia felis were observed in humans. This study presents the first report from the northeast of Spain. Human serum samples were tested by serological test. R. typhi and R. felis seroprevalences were 8.8% and 3.2%, respectively.

Age Distribution↗
PubMed2006

Proposal to create subspecies of Rickettsia sibirica and an emended description of Rickettsia sibirica.

The Rickettsia sibirica species is composed of isolates that are genotypically close but can be classified within two distinct serotypes, that is, R. sibirica sensu stricto and R. sibirica mongolitimonae (incorrectly named R. mongolotimonae). We investigated the possibility of classifying rickettsiae closely related to R. sibirica as R. sibirica subspecies, as proposed by the ad hoc Committee on Reconciliation of Approaches to Bacterial Systematics. For this, we first estimated the genotypic variability by using multilocus sequence typing (MLST), including the sequencing of five genes, and multispacer typing (MST) using three intergenic spacers, of five isolates and three tick amplicons of R. sibirica sensu stricto and six isolates of R. sibirica mongolotimonae. Then, we selected a representative of each MLST genotype and used mouse serotyping to estimate their degree of taxonomic relatedness. Among the 14 isolates or tick amplicons studied, 2 MLST genotypes were identified: (i) the R. sibirica sensu stricto type; and (ii) the R. sibirica mongolitimonae type. Representatives of the two MLST types were classified within three MST types and into two serotypes. Therefore, as isolates within the R. sibirica species are genotypically homogeneous but show MST genotypic, serotypic, and epidemio-clinical dissimilarities, we propose to modify the nomenclature of the R. sibirica species through the creation of subspecies. We propose the names R. sibirica subsp. sibirica subsp. nov. (type strain = 2-4-6, ATCC VR-541(T)), and R. sibirica subsp. mongolitimonae subsp. nov. (type strain = HA-91, ATCC VR-1526(T)). The description of R. sibirica is emended to accommodate the two subspecies.

Animals↗
PubMed1989

Seroprevalence of Rickettsia typhi and Rickettsia conorii infection among rodents and dogs in Egypt.

A serological survey of 1813 rodent and 549 dog sera, collected from 1979 to 1986 from animals in 16 Egyptian Governorates were tested for antibody to Rickettsia typhi and Rickettsia conorii by the indirect fluorescent antibody test. Only three of 82 (4%) sera from Rattus rattus collected near Aswan had antibody to R. conorii. The prevalence of R. typhi antibody in dog sera was only 0.4% (n = 549) while 25% (n = 547) of Rattus norvegicus and 11% (n = 1138) of R. rattus had measurable antibodies. Among the other rodents, antibody was demonstrated in only 2% (n = 45) of Arvicanthis spp., and 1% (n = 83) of Acomys spp. Collectively, rodents captured in the Nile Delta had a higher prevalence (mean 24% (n = 787] than those captured in the Nile Valley (mean 4% (n = 650]. Antibody to R. typhi was detected in rodents collected in all port cities: ismailiya, 13%; Port Said, 9%; Suez, 9%; Safaga, 16%; Quseir, 32% and Alexandria, 34%. These data showed evidence of R. typhi infection among rodents in widespread geographic localities of Egypt and suggested that infected rodents may be a source of human infections.

Animals↗
PubMed1994

Genotypic characterization of rickettsiae by DNA probes generated from Rickettsia prowazekii DNA.

Southern blot analysis of HindIII-cleaved rickettsial DNA was used for genotypic characterization of the typhus group (TG) species (R. prowazekii, R. typhi, R. canada) and a few species of the spotted fever group (SFG) rickettsiae (R. sibirica, R. conorii, R. akari). Four different DNA probes were employed. PBH11 and PBH13 probes were morphospecific HindIII fragments of R. prowazekii DNA. MW218 probe contained the gene for 51 K antigen and MW264 probe contained the citrate synthase gene of R. prowazekii. All the probes hybridized with the tested TG and SFG rickettsial DNAs, forming from 1 to 5 bands, but they did not with R. tsutsugamushi or C. burnetii DNAs. All the probes demonstrated specific hybridization patterns with TG species and R. akari. PBH11, PBH13 and MW264 probes clearly distinguished R. sibirica and R. conorii from the other tested rickettsiae, but not from each other. However, these two species differed slightly with MW218 probe. Several strains of each species were analyzed in this way and except for strains of R. conorii identical intraspecies patterns were obtained. These data lead us to consider the obtained hybridization patterns as criteria for genotypic identification.

Animals↗
PubMed2002

Kinetics of antibody responses in Rickettsia africae and Rickettsia conorii infections.

African tick-bite fever, caused by Rickettsia africae, is the most common tick-borne rickettsiosis in sub-Saharan Africa. Mediterranean spotted fever due to Rickettsia conorii also occurs in the region but is more prevalent in Mediterranean countries. Using microimmunofluorescence, we compared the development of immunoglobulin G (IgG) and IgM titers in 48 patients with African tick-bite fever and 48 patients with Mediterranean spotted fever. Doxycycline treatment within 7 days from the onset of disease significantly prevented the development of antibodies to R. africae. In patients with African tick-bite fever, the median times to seroconversion with IgG and IgM were 28 and 25 days, respectively, after the onset of symptoms. These were significantly longer by a median of 6 days for IgG and 9 days for IgM than the times for seroconversion in patients with Mediterranean spotted fever (P < 10(-2)). We recommend that sera collected 4 weeks after the onset of signs of patients with suspected African tick-bite fever should be used for the definitive serological diagnosis of R. africae infections.

Anti-Bacterial Agents↗
PubMed1997

Protein characterization of Australian spotted fever group rickettsiae and monoclonal antibody typing of Rickettsia honei.

Rickettsial proteins rOmp A and rOmp B exist in both Rickettsia australis and Rickettsia honei but differ in molecular weight and antigenicity; in addition, they produce distinct immunogenic responses and appear to be to conformationally dependent antigens. Species-specific monoclonal antibodies for other spotted fever group rickettsial species did not react with R. honei. A PCR product of the repeat region of the rOmp A gene from R. honei was amplified and calculated to contain 11 repeat units.

Antibodies, Bacterial↗
PubMed1997

Seroepidemiology of Rickettsia typhi, spotted fever group rickettsiae, and Coxiella burnetti infection in pregnant women from urban Tanzania.

Immunofluorescent antibody (IFA) testing was performed on sera drawn from 150 pregnant women in the port city of Dar es Salaam, Tanzania. Prevalence of antibodies to Rickettsia typhi was 28%, higher than in any of the 12 other African countries in which serosurveys using IFA testing have been performed. Seroprevalence of antibodies to spotted fever group rickettsiae antigens was 25.3%, comparable with that found in other sub-Saharan countries endemic for Amblyomma ticks. Only 4.7% of women were seropositive for Coxiella burnetii.

Adolescent↗
PubMed1985

Rickettsia-like organisms in ticks and antibodies to spotted fever-group rickettsiae in mammals from northern Mississippi.

Studies were conducted from April through August during 1974 and 1975 on 30 randomly selected trapping sites in Wall Doxey State Park, Holly Springs National Forest, and 1.6 km south of the park in Marshall County, Mississippi to provide information on the occurrence of ticks involved in the maintenance and transmission of spotted fever-group rickettsiae in nature. Of 365 animals (14 species) collected, 186 (51%) were parasitized by 4,169 ticks. Species of ticks collected included: Dermacentor variabilis; Amblyomma americanum; Rhipicephalus sanguineus; Haemaphysalis leporispalustris; and Ixodes texanus. Of 2,105 ticks examined, 116 (5.5%) were positive for rickettsia-like organisms.

Agglutination Tests↗
PubMed1977

Antigenic relations of Rickettsia prowazekii and Rickettsia canada, established in the study of sera of patients with Brill's disease.

Sera of patients with Brill's disease and of healthy persons with spotted fever in their past history were examined in the complement fixation reaction (CFR) to determine antigenic relations between R. prowazekii and R. canada. R. canada was found to have common antigenic determinants with R. prowazekii and R. mooseri. However, the antigenic determinants of R. canada differed from those of the mentioned rickettsiae. The titres of complement-fixing antibodies in the sera of patients with Brill's disease with the antigen of R. mooseri were lower than the titres with the homologous antigen within the range of 1-2 twofold dilutions of the serum. However, the oscillations of the titres with the antigen of R. canada in the study of the same sera were expressed in 1-5 twofold dilutions. In serological identification of canada rickettsiosis, antigens of rickettsiae of the spotted fever group should invariably be included in the investigation of the sera.

Antigens↗
PubMed1995

Cross-reactivity of Rickettsia japonica and Rickettsia typhi demonstrated by immunofluorescence and Western immunoblotting.

Cross-reactivity between Rickettsia japonica and R. typhi was observed by immunofluorescence tests using sera from patients with Oriental spotted fever (OSF), from whom the causative agent was isolated and identified as R. japonica. Western immunoblotting with these sera revealed that only the 120-kilodalton surface polypeptide, i.e., rickettsial outer membrane protein (rOmp) B, has a common antigenicity with the 105-kilodalton surface polypeptide of R. typhi. In some cases, antibodies specifically reactive with R. typhi were detected in acute-phase sera followed by a significant rise in titers, possibly because of an anamnestic response to a previous infection with an R. typhi-like agent; the sera retained reactivity to R. typhi even after absorption by a homologous strain. A lipopolysaccharide (LPS)-like antigen of R. typhi was found to be reactive with some sera of OSF patients. The ladder bands on Western immunoblot of rickettsial organisms were confirmed to be polysaccharide in nature, which was demonstrated by comparing them with the pattern of silver-stained gel of proteinase K-treated rickettsial specimens after sodium dodecyl sulfate-polyacrylamide gel electrophoresis.

Antibodies, Bacterial↗
PubMed2006

Detection of Rickettsia rickettsii and Rickettsia sp. in blood clots in 24 patients from different municipalities of the State of Sao Paulo, Brazil.

The authors detected Rickettsia genus organisms using shell vial and polymerase chain reaction (PCR)/sequencing analysis in blood clots in patients suspected of having Brazilian spotted fever (BSF). DNA was detected using PCR with three sets of primers to access the gltA, ompA, and ompB genes. Sequence analysis was carried out using an automatic sequencer with Bioedit software. Seventy-five percent of the culture samples were positive and all samples amplified rickettsial gene fragments. To date, 46% of the samples have been sequenced.

Blood Coagulation↗