Light microscopic investigations on lysozyme- and penicillin-induced morphological changes in Erysipelothrix rhusiopathiae and on propagation of its protoplast type L-form.

Although lysozyme and penicillin are different in their molecular action on cell wall murein they produce similar morphological changes in Erysipelothrix rhusiopathiae grown on agar media. 2,000--5,000 micrograms/ml lysozyme and 0.1--2 IU/ml penicillin induce filament formation. Filaments are able to divide in rods, which shows that only cross wall formation and separation are inhibited. Higher doses of lysozyme (10,000 micrograms/ml) and penicillin (less than 1 IU/ml) inhibit cell wall synthesis and induce L-form growth. The propagation of this protoplast type L-form was investigated by microphotographic series in phase contrast microscope during L-form induction and in the stable L-form state. In both cases L-form cells propagate by formation and growth of small granular elements of about 0.2--0.6 micrometers in diameter, which spread in different directions in the agar medium. The multiplication process may be explained by the plasticity and flexibility of the L-form cell and its cytoplasmic membrane and by the structural and functional interaction between the "folded chromosome" and the surrounding cytoplasm.

Ultrastructural characterization of stable L-form cells from Erysipelothrix rhusiopathiae and of accompanying artifacts.

The stable L-form of Erysipelothrix rhusiopathiae is a typical protoplast type L-form. Cells are surrounded by a trilamellar cytoplasmic membrane only. They grow in form of aggregations in liquid media and their diameters vary between 0.1 and 2 micrometer. Always a large portion of cells undergoes lysis. It seems to be characteristic for L-form cultures of E. rhusiopathiae that always many artifact structures are formed. The artifacts are spherical particles with diameters of 0.1 micrometer to more than 3 micrometer. They can be differentiated from L-form cells only by electron microscopy. The artifacts consist of electron dense amorphous material and their surface is irregular without a clear boundary line. Obviously, these artifacts are produced from protein components of the medium and from cytoplasmatic components of the lysing L-form cells.

[Serum complement level (C'H5O) during treatment of rheumatoid arthritis patients]. / Nivo na serumniia komplement (C'H5O) pri lechenie na bolni s revmatoiden artrit

Decreased total complement serum level (C'H50) was established in 72 per cent of the 78 patients examined, associated most likely with the developing processes of antigen-antibody reaction and the severe course of the illness. The applied treatment with different antirheumatic remedies, according to their mechanism of action, determined the respective changes in the complement level. Indomethacinum and chinoline derivatives induce complement values elevation, whereas the corticosteroids, salicylate and pyrazolone preparations lead to the complement values normalization.

Adhesion and phagocytosis of Staphylococcus aureus L-forms.

Electron microscopical investigations on in vitro and in vivo interactions of normal Staphylococcus aureus cells with rat peritoneal macrophages showed that these bacteria were rapidly endocytosed and digested even in the absence of specific antibodies. In contrast to the parental strains oxacilin-induced and stable variante lacking a cell wall (L-forms) were ingested without subsequent formation of phagolysomes and digestive vacuoles. The intracytoplasmic L-form bodies retained their characteristic ultrastructure, i.e. no visible alterations occurred. Some morphological aspects of the L-forms and their persistence in macrophages 7 days after intraperitoneal administration of L-form to rats, suggest the possibility of their intracellular survival.

Persistence and isolation of L cycle forms of Erysipelothrix rhusiopathiae in experimental infection.

In experimental infection of rats induced with teh S-form of Erysipelothrix rhusiopathiae the pathogen was found to persist for a long time (up to fifteen months according to the author's personal observations). Under the effect of the specific and unspecific defense of the animal's organism the causative agent suffered a number of morphologic changes. In the course of the first days of infection the R-form was isolated, and thirty days later up to the fifteenth month forms of the L cycle were recovered from the parenchymal organs, blood, and joints. An electron microscopic study was carried out on the structure of the isolated L-forms. The long-term persistence of the agent in its different morphologic forms may well account for the chronic course of the infection in rats.

[Effect of the pesticide carbaryl (sevin) on the course of experimental Erysipelothrix rhusiopathiae infection in rats]. / Vliianie na pestitsida karbaril (sevin) vurkhu khoda na eksperimentalna infektsiia u plukhove s Erysipelothrix rhusiopathiae.

Rats treated with carbaril (sevin) at the rate of 2 to 5 mg were infected via the feed with Erysipelothrix rhusiopathiae in the course of 30 days. Studied were the clinical picture, some paraclinical indices, the mortality rate, and the morphologic and histopathologic changes in the parenchymal organs. It was found that rats that were given carbaril for a long time in minimum amounts and were then subjected to infection developed a characteristic clinical picture and showed a high mortality rate (nearly twice as high as that with the control animals). In hemocultures the causative agent was isolated over a longer period than in the case with the controls. Both the gross and the histopathologic findings were more strongly manifested in the test (carbaril-treated) rats.

Histoautoradiographic studies of 3H-thymidine labelled Erysipelothrix rhusiopathiae introduced into albino mice.

S-forms of E. rhusiopathiae were cultivated in a 3H-thimidine-containing medium. A suspension washed three times in succession was intravenously injected into albino mice. At certain intervals liver, spleen and kidney were examinated by routine autoradiogrphy. Intact labelled bacteria were found as early as 5 min after administration in the blood capillaries, around cells of the RES and in some nuclei of hepatocytes. Later they are detected in the cytoplasm of micro- and macrophages, around megacaryocytes and in some of their nuclei. After 6 h the labelled population sharply decreases due to a very active reproduction and mortality of the initial population. The localisation of bacteria in liver nuclei and the spreading of S-forms of E. rhusiopathiae in comparison with the avirulent L-forms therefrom are discussed.

Histoautoradiographic studies on the fate of 3H-thymidine labelled L-forms of E. rhusiopathiae in albino mice.

L-forms of Erysipelotrix rhusiopathiae labelled with 3H-thymidine were intravenously administered to albino mice. Autoradiographic studies of the liver, kidney and spleen were undertaken at periods ranging from 2 minutes to 15 days. On the second minute following the administration of the radioactive material whole labelled microorganisms and chains of silver grains were recovered in the examined organs. Up to the 15th minute labels were observed also in the cells of the RES. Following the 30th minute the silver grains were positioned at a characteristic site in the Golgi region of the hepatocytes. At the same time in the kidney they were localized in the glomerular space and in the lumen of the renal tubules, whereas in the spleen - mainly around the megakaryocytes. By the 15th day labelling gradually diminished, single silver grains being found over some nuclei of megakaryocytes, liver and kidney parenchymal cells. The present study throws light over some aspects of the interrelationship between the micro- and macroorganism concerning the mechanisms of desintegration, elimination and the uptaking of labelled microbial DNA.

Effect of experimentally induced trichinellosis on the infection with Erysipelothrix rhusiopathiae in rats.

Infection with Erysipelothrix rhusiopathiae in rats infested 20 days earlier with Trichinella spiralis developed more slowly, the clinical and pathoanatomic changes in the joints were expressed to a less extend, and the mortality rate was lower. The erythrocyte sedimentation rate, the precipitin formation and the phagocytic activity of the macrophages did not considerably change. Experiments carried out to elucidate this fact did not reveal any antigenic or antagonistic relationships between parasite and bacterium nor any protective effect of the host's serum. The inhibitory influence of corticosteroids on the defence forces was not completely manifested in the rats infested. This fact might be explained by interrelations depending both on the cycle of helminth development and on the non-specific immunological reactivity of the organism, the latter being stimulated by the helminth invasion.

Experimental pyelonephritis induced by L-forms of Proteus mirabilis in rats.

Pyelonephritis was produced in 75 white rats by applying a 24-hour ligature on the ureter and by intravenous injection of a suspension of 5X10(9) cells of a stable Proteus mirabilis L-form. Pyelonephritis was proved microscopically in 44% of the animals. The revertants of the introduced L-form play an etiological role in the development of the infection. The stability of the pathohistological findings makes this model suitable for the purpose of experimental chemotherapy.

[Immunosuppression in an experimental infection with the infectious rhinotracheitis virus]. / Imunosupresiia pri eksperimentalna infektsiia s virusa na zarazniia rinotrakheit.

The changes in the immune status of the calves infected with bovine herpes virus-1 inducing infectious rhinotracheitis were studied. The virus was introduced by intratracheal inoculation of 5 cm3 suspension with minimal concentration of 10(-7) CPU50 per 1 cm2 from the Cervena voda strain cultivated in calf kidney culture. An infection of moderate seriousness and characteristic clinical manifestations was caused. On the 2nd, 5th, 10th and 15th day after the infection lung lavage was carried out and samples were taken from the peripheral blood. The number, viability and differential distribution of the alveolar macrophages and phagocytic and microbicidal ability showed a tendency to decrease during the acute phase of the infection (the 2nd-5th day) followed by a slow restoration in the period of convalescence (the 15th day). Some parameters of the systemic immunity--number of the plaque-forming and the rosette-forming cells, differential and absolute number of leucocytes in the peripheral blood--also had similar dynamics. The changes in the local and in the systemic, in the cell and humoral immunity were connected to the pathogenesis of the infection and the immune suppressive properties of the agent. The rhinotracheitis infection is indicative of the character of changes in the immune status at herpes virus infections and, on the other hand, it can be used as a model for testing immunostimulators of recovery effect.

Immunological and biochemical investigations of the infectious bovine rhinotracheitis. III. Alterations in pulmonary surfactant lipids.

The effect of experimental bovine herpes virus (BHV) type I rhinotracheitis on the surfactant system phospholipids in calves was examined. A stimulated exocytosis of pulmonary surfactant phospholipids in the acute phase of the disease was documented biochemically and ultrastructurally. The data presented were assumed as an evidence of the involvement of pulmonary surfactant in lung defense.

Fatty acid composition of lipids of Escherichia coli W 1655 F+ and its stable protoplast type L-form.

The comparative fatty acid analysis of extractable and non-extractable lipids of Escherichia coli W 1655 F+ and its stable protoplast type L-form shows quantitative as well as qualitative differences. From 10 different fatty acids obtained 16:0, 17:0 and 18:0 are present at about the same quantities in the lipid fractions of the bacterial and L-form. The absence of larger amounts of 12:0, 14:0, and 14:beta OH fatty acids in non-extractable L-form lipids reflects the loss of the cell wall in L-form cells. 16:1 fatty acid was found in L-form lipids only. This qualitative difference and the 2-3 times higher content of 18:1 in L-form lipids and the 7 times lower content of cyc 19:0 in extractable lipids of the L-form may be interpreted as alterations characteristic for the changed composition of the cytoplasmic membrane in L-form cells.

[The immunostimulating action of suramin in infection with the infectious rhinotracheitis virus]. / Imunovuzstanovitelno deistvie na suramin pri infektsiia s virusa na zarazniia rinotrakheit.

The immunostimulating antiviral preparation suramin has a favourable effect on the process and outcome of an experimental infection with the virus of infectious rhinotracheitis (bovine herpes virus type 1, BHV1), thus reducing the gravity of the clinical course and increasing the survival rate. The therapeutic protective effect of the preparation is expressed in the prevention of virus-induced immunosuppression which is markedly strong during the acute phase of the infection process. Suramin restored the number, viability, relative share in the bronchial lavage population, phagocytic and microbicidal capacity of the alveolar macrophages--a major factor of the local defence mechanisms of the lung. In a similar way suramin influenced some parameters of the systemic immunity (number and differential count of polymorphonuclear phagocytes, plaque- and rosette-forming cells). The application of suramin in doses lower than those used for virus inhibition and its considerable action on different mechanisms of the immune system suggested that the preparation's therapeutic effect could be related mainly to its immunostimulating properties.