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.

Genotypic and biological characteristics of non-identified strain of spotted fever group rickettsiae isolated in Crimea.

A strain of rickettsiae, designated Crimea-108, was isolated from ticks Dermacentor marginatus in the Crimea in 1977. Its immunobiological characteristics involve low pathogenicity for experimental animals, moderate infectivity for chick embryos, and antigenic relatedness to spotted fever group (SFG) rickettsiae (R. sibirica, R. conorii, R. akari), especially to R. sibirica. The genotypic characterization of the strain Crimea-108 was carried out in comparison with SFG and typhus group rickettsiae by using restriction fragment length polymorphism (RFLP) analysis and DNA-probe hybridization. The marked similarity was detected between DNA restriction patterns of the strains Crimea-108, R. sibirica and R. conorii, but each of them besides comigrating fragments had specific ones. Genotypic analysis of the strain Crimea-108, the SFG and typhus group rickettsiae by three independent DNA probes, based on R. prowazekii DNA, gave unique hybridization patterns for the Crimea-108 strain with all probes. The obtained data show that the Crimea-108 isolate does not belong to the species of R. sibirica, R. conorii, R. akari. The strain Crimea-108 is a novel strain of SFG rickettsiae for the Crimea region.

[Genotypic characteristics of Rickettsia sibirica strains]. / Genotipicheskaia kharakteristika shtammov Rickettsia sibirica.

Six Rickettsia sibirica strains isolated in Siberia and Far East (Primorje) from various sources (patient, ticks D. nuttali, D. silvarum, H. concinna) at different time (1940-1980) were studied by the RFLP and DNA probe hybridization techniques. All studied strains were found to have the identical profiles of migrating fragments in restrictograms got by using a set of endonucleases (EcoRI, PstI, PvuII, Bg1I, XbaI, HindIII, MspI) and similar zones of hybridization with a DNA probe derived from Rickettsia prowazekii DNA. The obtained data point to a close similarity between the genomes of investigated Rickettsia sibirica strains. Long-term isolation of the genetically similar Rickettsia sibirica strains testifies to their constant circulation, thus apparently determining the stability of epidemiologic manifestation of tick-borne typhus fever of Northern Asia in the central part of its area (Siberia, Far East).

[Biological properties of antibiotic-resistant mutants of the mildly pathogenic Rickettsia prowazekii E strain]. / Biologicheskie svoistva ustoichivykh k antibiotikam mutantov malopatogennogo shtamma E Rickettsia prowazekii.

The use of R. prowazekii strain E with low pathogenicity as live vaccine against exanthematous typhus is limited by its high specific reactogenicity, which is probably due to the reversion of the virulence of the strain. One of the approaches to the stabilization of the avirulent properties of strain E is obtaining its mutants with stable decreased pathogenic properties. The article presents the data on the infectious properties of R. prowazekii antibiotic-resistant strain E mutants obtained in earlier experiments, in respect of chick embryos and laboratory animals, as well as the capacity of this strain for producing immunity to challenge with R. prowazekii virulent strain in guinea pigs. The study has revealed that the erythromycin-resistant mutant of R. prowazekii strain E, induced by nitrosoguanidine (NG), has lower infective capacity for chick embryos, guinea pigs, cotton rats and white mice. The infective capacity of the NG-induced rifampicin-resistant and spontaneous erythromycin-resistant mutants of R. prowazekii strain E is similar to the infective capacity of the initial strain. The rifampicin-resistant and spontaneous erythromycin-resistant mutants of R. prowazekii strain E possess immunogenicity similar to that of the initial strain E, and the NG-induced erythromycin-resistant mutant possesses lower, but sufficiently pronounced immunogenicity despite its decreased infective capacity for guinea pigs.

[The protective activity of individual proteins and immunogenic fractions isolated from Rickettsia prowazekii]. / Issledovanie protektivnoi aktivnosti individual'nykh belkov i immunogennykh fraktsii, vydelennykh iz Rickettsia prowazekii.

The immunogenic properties and protective activity of basic protein I and tris-soluble antigens isolated from R. prowazekii were analyzed in comparison with those of chemical typhus vaccine on the model of anti-infectious immunity in guinea pigs. The analysis revealed that purified protein I has protective activity for guinea pigs, which is less pronounced in protein I from strain E with weak pathogenicity than protein I from strains EVir and Breinl. The activity and immunogenic properties of tris-soluble antigens, especially antigen I, are comparable with those of chemical typhus vaccines in the parameters under study.

[The need for a review of the procedure for the preventive vaccination of researchers working with Rickettsia prowazekii]. / Neobkhodimost' peresmotra taktiki profilakticheskoi vaktsinatsii sotrudnikov, rabotaiushchikh s Rickettsia prowazekii.

The analysis of the results of prolonged observations on the prophylactic immunization of employees working with R. prowazekii is presented. The necessity of the differentiated approach to the determination of the immunization schedule and the choice of vaccine is shown. The presence of specific antibodies (Ab) and the level of their titers have been found to be related to the degree of anti-infectious protection. The following characteristics indicate the presence of profound immunological transformation in vaccinees: complement-fixing Ab in titers 1:10 and more and/or immunofluorescent Ab in titers not below 1:180, Ab to protein in the hemagglutination test in titers not below 1:1000. These specific Ab and the level of their titers can be registered after the second injection of live combined typhus vaccine E and the third injection of chemical typhus vaccine. Cases of laboratory infection and their relationship to the character of immunization and the intensity of contacts with R. prowazekii virulent strains are discussed. Attention is drawn to the strict observance of professional safety rules.

[Infectivity and sensitivity to antibiotics of Rickettsia prowazekii mutants of the low-pathogenic strain E cultured in chick embryos]. / Infektsionnost' i chuvstvitel'nost' k antibiotikam mutantov Rickettsia prowazekii, malopatogennogo shtamma E, pri kul'tivirovanii v kurinykh émbrionakh.

Selection of mutants of a low pathogenic strain E of R. prowazekii is a trend in genetic investigation of this Rickettsia species and one of the approaches to stabilizing the strain avirulent properties with a purpose of using in vaccine prophylaxis of typhus. The mutants of R. prowazekii, strain E selected by the authors earlier were characterized with respect to their infective capacity for chick embryos (CE) and antibiotic sensitivity. It was found that the infective capacity for CE of the erythromycin resistant mutant induced by nitroso guanidine (EErrI) was by ID50 2-3 logarithms lower than that of the initial strain E. The infective capacity for CE of the rifampicin resistant mutant induced by nitroso guanidine (ERifrI) and the spontaneous erythromycin resistant mutant was similar to that of strain E. The ERifrI strain differed from the initial strain E by higher sensitivity to tetracycline and erythromycin and the EErrI strain differed from the initial strain E by higher sensitivity to tetracycline and rifampicin. It was shown that the biological properties of the nitroso guanidine-induced mutants resistant to rifampicin and erythromycin differed from those of the initial strain E and the properties of the spontaneous erythromycin resistant mutant were similar to those of the initial strain E.