The ability of flea species on the common vole from mountainous Dagestan to transmit and preserve the causative agent of plague

The infection ability and terms of preservation of plague microbe in fleas of common vole from Dagestan high-mountain plague focus (Frontopsylla causasica, Megabothris turbidus, Ctenophthalmus intermedius and Amphipsylla rossica) was studied experimentally. Block formation and transmission of the infection by F. caucasica and M. turbidus was observed. The transmission of plague microbe by Ct. intermedius fleas was first carried out. These species of fleas preserve the infection for a long time. Experiments with fleas of A. rossica yielded negative results.     

Contrasted patterns of selection on MHC-linked microsatellites in natural populations of the Malagasy plague reservoir

Plague (Yersinia pestis infection) is a highly virulent rodent disease that persists in many natural ecosystems. The black rat (Rattus rattus) is the main host involved in the plague focus of the central highlands of Madagascar. Black rat populations from this area are highly resistant to plague, whereas those from areas in which the disease is absent (low altitude zones of Madagascar) are susceptible. Various lines of evidence suggest a role for the Major Histocompatibility Complex (MHC) in plague resistance. We therefore used the MHC region as a candidate for detecting signatures of plague-mediated selection in Malagasy black rats, by comparing population genetic structures for five MHC-linked microsatellites and neutral markers in two sampling designs. We first compared four pairs of populations, each pair including one population from the plague focus and one from the disease-free zone. Plague-mediated selection was expected to result in greater genetic differentiation between the two zones than expected under neutrality and this was observed for one MHC-class I-linked locus (D20Img2). For this marker as well as for four other MHC-linked loci, a geographic pattern of genetic structure was found at local scale within the plague focus. This pattern would be expected if plague selection pressures were spatially variable. Finally, another MHC-class I-linked locus (D20Rat21) showed evidences of balancing selection, but it seems more likely that this selection would be related to unknown pathogens more widely distributed in Madagascar than plague.     

Susceptibility to Yersinia pestis in the northern grasshopper mouse (Onychomys leucogaster)

The laboratory-born progeny from two geographically distant populations of northern grasshopper mice (Onychomys leucogaster) were challenged with Yersinia pestis to determine their relative susceptibilities to plague. One of the O. leucogaster populations was associated with a known epizootic focus of the disease and was found to be nearly 2,000 times more resistant to mortality than were members of another population from an area historically free of plague. The ecology and omnivorous behavior of O. leucogaster appears to promote strong selection for resistance to plague in areas where they are naturally exposed.     

Effect of the digestive process on the survival of the plague agent in Xenopsylla gerbilli Minax fleas

Hystological investigations of experimentally infected X. gerbilli minax have shown that at the early stages of the digestion, while in the intestine compact clots of non-digested blood, are preserved, proceeds the reproduction of the plague agent. With the decay of the alimentary clot it changes into the dying off of the microbe population and after the ceasing of digestion the accumulation of bacteria renews again. Simultaneously with the changes in the number of microbes their morphology varies. At the initial stage of blood digestion bacilli typical for the agent are dominant. With the decay of the alimentary clot they are replaced by ovoid and spheroid forms. After the ceasing of the digestion cycle the microbe mass is represented by small coccobacteria. During the decrease in the plague agent abundance bacteria are absent mostly often from the midgut and posterior parts of the alimentary canal. Most favourable conditions for their preservation exist in this period in the proventriculus and in adjoining to it oesophagus part.     

Antibiotic sensitivity of plague microbe strains from foreign countries]

The method of serial dilutions on the Hottinger agar was applied to comparative assay of antibiotic sensitivity in 50 strains of the plague microbe isolated abroad and in 5 strains isolated in the plague focus in the Central Caucasus. The antibiotics used in the assay were the following: streptomycin, gentamicin, doxycycline, monomycin, kanamycin, tetracycline, erythromycin, ristomycin, lincomycin and polymyxin M. Irrespective of the origin, all the isolates were resistant to erythromycin, lincomycin and polymyxin M. The levels of the sensitivity to the other antibiotics were different. The data serve as a ground for the statement that there is no tendency to development of antibiotic resistance in the plague microbe in patients treated with high doses of the antibiotics and mainly streptomycin. Along with streptomycin, such antibiotics as gentamicin, tetracycline, doxycycline and kanamycin are useful in the therapy of plague and require further investigation.     

Pathogenic action of the plague microbe on the flea Xenopsylla cheopis and the ultrastructure of the causative agent at various times of its stay in the vector

Pathology of gastro-intestinal tract of Xenopsylla cheopis fleas infected with plague microbe was determined by means of electron microscopy. Ultrastructure of plague microbe during different periods of its stay in the vector was studied.     

Doxycycline in the prevention of experimental plague induced by plague microbe variants]

A comparative study was performed on the efficacy of doxycycline in experimental plague infection induced in albino mice by strain 231 of the plague microbe and its variant 231 Fra- deprived of the ability to produce the fraction I antigen. It was shown that the LD50 for strain 231 during animal treatment with doxycycline was significantly higher than that for variant 231 Fra-. Prophylaxis of the plague infection caused by the Fra- forms of the plague microbe required significantly higher doses of doxycycline (ED50) than that of the infection caused by the Fra+ forms. The use of the daily maximum permissible doses of doxycycline (50 to 100 mg/kg a day) for 10 days in treatment of albino mice infected with the strain Fra- did not provide animal survival at the level higher than 60 to 70 per cent while the survival rate in the animals infected with the strain Fra+ of the plague microbe and treated according to the same scheme amounted to 90-100 per cent. The lower therapeutic efficacy of doxycycline in the treatment of the infection caused by the fractionless variant of the plague microbe should be considered in development of rational schemes for prophylaxis and treatment of plague.     

Results of using the serological method of epidemiological investigation of plague foci in the Muyunkum and Eastern Kyzylkum deserts in 1978-1982

Results of serological examination of 81,853 mammals from the Muyunkum and 26,303 from the Eastern Kyzylkum deserts were compared with results of a corresponding bacteriological examination for plague. In both foci and in all phases throughout the epizootic of plague, the serological method made it possible to detect infected animals 6-10 times more frequently in comparison with the bacteriological method. The big meriones mice, i.e., the basic carriers of plague in natural foci, were found to be infected considerably more (in percentage) frequently than the meridian meriones. Differences in the reactivity of meriones to the invasion of plague microbe were observed in both foci (the indices of titres in the antigen neutralization reaction were considerably higher in the Eastern Kyzylkums). The serological method of investigation is highly informative in evaluating both the present state of epizootic activity of a plague focus and its activity in the past.     

The epizootiological significance of flea accumulation in aggregations of the female long-tailed suslik in the Tuva plague focus

The formation of aggregations of related females is a peculiarity of the intrapopulational organization of long-tailed suslik that results in the accumulation of Citellophilus tesquorum, the main vector and keeper of the plague microbe in the Tuva plague nidus. The plague agent is adapted to the existence on the territory occupied by aggregations of females that manifests itself in the delay of the beginning and prolixity of block-formation periods in fleas. The transfer of the agent from one place to another takes place only at the high abundance of long-tailed suslik. When the number of the suslik is low the plague microbe circulates in the ranges of one or several neighbouring aggregations of females having no possibility for successful transfer to a new place.     

Ecological Aspects of Evolution of the Plague Microbe Yersinia pestis and the Genesis of Natural Foci

A new hypothesis of the origin of the plague microbe in the Mongolian bobak (Marmota sibirica Radde, 1862) populations in Central Asia during the Pleistocene is based on the ideas of its relative phylogenetic recency. The Late Pleistocene cooling, which induced a deep freezing of the grounds in southern Siberia, Mongolia, and Manchuria, is considered as an inducer of speciation. The main ecological factors of the plague microbe evolution include the species specific behavior of the Mongolian bobak as it prepared to hibernate related to its occurrence in arid petrophytic landscapes and the larval parasitism of the flea Oropsylla silantiewi Wagn., 1898 in winter. Genesis of the plague foci is divided into two periods: natural-historical and biosocial. During the first period, the primary natural foci in Eurasia were formed and, during the second period, synanthropic (rat) and secondary natural foci appeared with the participation of humans in Africa, The New World, and on some tropical islands.     

Relationship of the flea Citellophilus tesquorum altaicus (Siphonaptera: Ceratophyllidae) with the plague agent

Some aspects of relationships of the flea Citellophilus tesquorum altaicus and bacterium Yersinia pestis of two strains isolated from different parts of the Tuva natural plague focus were studied. Peculiarities of elimination and blood meal activity of fleas infected with two strains of the plague agent were not revealed. Differences in mortality and alimentary activity are considerably determined by the sex of insects. The ability of examined strains to form a proventriculus block was not identical in the strains examined. This ability was expressed higher in the strain I-3428, which originated from the same part of the natural focus as the insectarium flea culture, than in the strain I-3327. During the spring and first half of summer, the proventriculus block appeared more frequently in females. The increasing of the fraction of blocked individuals was observed in both sexes from spring to summer. As for the ability to transmit the plague agent, similar seasonal increasing was noted in males, but in females, the ability to inoculate the plague microbe was always maintained at the same level.     

The expression of Fc gamma receptors on the surface of macrophages activated by a vaccinal strain of Yersinia pestis

The expression of Fc gamma R on the surface of macrophages in the process of antiplague immunity formation is analyzed. The stud is performed on the alveolar and peritoneal macrophages obtained from intact and immunized guinea pigs in different periods after vaccination (the 1st, 7th, 14th and 21st day). It is established that during the formation of the antiplague immunity there occurs activation of macrophages which is accompanied by an increase of the Fc gamma R expression on the outer surface of the membrane both of peritoneal and alveolar macrophages and the pattern of response of these cells to the interaction with the vaccine strain of the plague microbe changes. The Fc gamma R expression heterogeneity of certain macrophage populations is revealed both in an intact and in immune organism as well as different pattern of the intact alveolar and peritoneal macrophage response during the interaction with the vaccine strain of the antiplague microbe. These differences are levelled in the process of the antiplague immunity formation.     

Concepts of macro- and microevolution as related to the problem of origin and global expansion of the plague pathogen Yersinia pestis

The relationship between macro- and microevolutionary processes is considered with reference to the ecological scenario of the origin of the plague pathogen and its subsequent natural and anthropogenic global expansion. The macroevolutionary transformation of the ancestral pseudotuberculosis microbe clone into the initial plague microbe Yersinia pestis tarbagani occurred in Central Asia at the end of the Late Pleistocene by a “vertical” Darwinian way in an inadaptive heterothermal continual intermediate environment—the Mongolian marmot Marmota sibirica—flea Oropsylla silantiewi system—via a sequence of unstable and currently extinct intermediate forms. Its natural geographic expansion on the “oil spot” principle in the postglacial time led to the microevolutionary formation of 20–30 hostal subspecies circulating in populations of the background species of burrowing rodents and pikas in arid areas of Eurasia. The intercontinental spread of the “marmot” and “rat” pathogen subspecies in the past few centuries has been exclusively anthropogenic, with the involvement of synanthropic (ship) rats.     

Iron assimilation by Yersinia pestis on iron-deficient media

The growth of plague bacteria may be limited by the level of iron concentration in the nutrient medium. The virulent strains of the plague microbe possess the more pronounced mechanism of iron assimilation as compared to the vaccine strain. The iron ions are extracted by the virulent and vaccine strains only under the cell surface contact with the iron-saturated transferrin. The iron-sorbing function is peculiar to the plague microbe cell walls which is pronounced more strongly in the virulent strains.     

Efficiency of Neopsylla abagaitui fleas as plague vectors in Transbaikalia

Experiments have shown that the block of proventriculus develops in 1.1 to 1.5% of individuals of the flea Neopsylla abagaitui infected with plague microbe. These insects transmit the agent during bloodsucking to different plague carriers (Citellus dauricus, C. undulatus, Meriones unguiculatus, Microtus gregalis). The plague microbe is preserved in fleas for 65 days (the observation period).     

Experimental evaluation of prospects for the use of beta-lactams in plague infection caused by pathogens with plasmid resistance to penicillins]

High therapeutic efficacies of ceftriaxone, ceftazidime, cefotaxime and azthreonam in the treatment of experimental plague induced by beta-lactamase-producing strains of the plague microbe containing R plasmids RP-1, R57b and R40a were shown to correlate with their in vitro antibacterial activities. The therapeutic efficacy of sulbactam/ampicillin was recorded in the treatment of plague induced by the strain containing R plasmids R57b and R40a (the treatment course of 7 days). However, it was lower when the infection was due to the strain containing plasmid RP-1 (beta-lactamase TEM-2). Cefoperazone was not active in the treatment of experimental plague induced by the strains containing plasmids RP-1 and R57b (beta-lactamases TEM-2 and OXA-3). Ceftriaxone versus the antibiotics tested was considered to be the drug of choice for the etiotropic therapy of plague induced not only by the type strains of the plague microbe but also by its variants with the plasmid pattern resistance to penicillins.     

Thymidine phosphorylase activity and its relationship to trimethoprim in initial strains and thymidine-, thymine-dependent and trimethoprim-resistant mutants of plague microbe]

The comparative study revealed thymidine phosphorylase activity in the initial strains of a plague microbe of the field variety and in thymidine-, thymine-dependent and trimethoprim-resistant mutants of the plague microbe of other varieties. The data fully conformed to the results of the microbiological investigation of the strains' ability to grow on the nutrient media with trimethoprim in the presence of thymine and thymidine. On the basis of these results it appeared possible to divide the initial and mutant strains of the plague microbe into four arbitrary groups: initial strains of the plague microbe of all the varieties except the field ones sensitive to trimethoprim under any temperature conditions of incubation on any medium with any supplements; initial strains of the plague microbe of the field variety resistant to trimethoprim at 28 degrees C in the presence of thymine or thymidine alone; Tmpr mutants whose resistance to trimethoprim at 28 degrees C did not depend on the presence of thymine or thymidine, purine and vitamins, but depended on the presence of these substances at a temperature of 37 degrees C.     

Sensitivity of different strains of the causative agent of plague to disinfectants]

Various strains of the plague microbe was found to have statistically significant differences in their sensitivity to phenol, formaldehyde and chloramine. These differences did not correlate with the origin, virulence and nutritional requirements of the plague strains. The sensitivity of the plague microbe to formaldehyde and phenol was found to be related to the permeability of its cell wall.     

Broader investigation into the external environment of the specific antigen of the infectious agent in epizootiological observation and study of the structure of natural foci of plague.

The possibilities of detection of the infectious agent in natural foci of plague can be expanded to some extent by using the method of search for the specific FI capsular antigen of the plague microbe in such objects of the external environment which have not yet been subjected to laboratory analysis and serological analysis in particular. Such objects include soil from the burrow and the substrate of the nest of the rodent as well as the fleas of rodents and regurgitations of birds of prey.The effectivity and suitability of examination of these objects using immuno-adsorption reactions with standard erythrocyte diagnostics (passive haemagglutination reaction, antibody neutralization reaction) were demonstrated experimentally and confirmed under field conditions in various geographical and ecological zones of the Caucasus. Thus in 1971, an earlier unknown natural focus of plague was found in the Mount Elbrus massif by means of serological analysis of the substrate of the nest and of the soil from the burrows of rodents. It can be expected that broadened serological search for the FI antigen of plague will prove useful in the study of the structure of natural foci of plague, in the reconnaissance or retrospective investigation of poorly explored territories and in the solution of some questions concerning the epizootiology of plague, such as the survival of the infectious agent in the inter-epizootic period, telluric plague, etc.     

Spheroplasts of plague microbe strains from the Transcaucasian uplands and their capacity for pesticin synthesis]

Spherical formations of the plague microbe strains from the Transcaucasian Upland, I plague microbe strain of the sandwort variety and I strain of the marmot variety were obtained under the effect of lithium chloride. They had the remains of the cell wall, were viable, sensitive to osmotic shock, preserved sensitivity to the specific bacteriophage and pesticins. All this was evident of isolation of the spheroplasts of the plague microbe. The spheroplasts showed a capacity for pesticin production. The pesticin synthesis by the spheroplasts of the plague causative agent from the Transcaucasian Upland increased with an increase in the content of lithium chloride in the medium. The largest inhibiiton zones were observed, when 0.7-0.8 per cent of lithium chloride were present in the medium. In the spheroplasts of the plague causative agent from the Mountain Altai (the marmot variety) the pesticin synthesis was inhibited with an increase in the content of lithium chloride in the medium. The activity spectrum of the pesticins of the spheroplasts of the plague causative agent from the Transcaucasian Upland and the spheroplasts of the strains of the marmot and sandwort varieties was broader than that of the rod-like forms of these strains. The indicator properties were found in the strains of the plague microbe of the marmot and sandwort varieties with respect to the pesticins of the spheroplasts of the sel-like producing organisms and organisms from the Transcaucasian Upland.