Screening of plasmids in the bacteria of the genus Bacillus with antilysozyme activity

In the study of the microbiocenosis of the distal section of the patients' large intestine Bacillus strains with antilysozyme activity (ALA) were isolated. In B. cereus strain 26 with pronounced expression of antilysozyme factor the plasmid sized approximately 100 kb was detected. The transformation of the isolated plasmid in cells of B. cereus non-plasmid strain IP5832 the localization of genes encoding ALA and resistance to kanamycin was determined. The production of ALA factor in the recombinant clone of B. cereus strain IP5832 corresponded to the clinical isolate of B. cereus 26. The replicon of the detected plasmid could be used for the determination of the coding sequence of the antilysozyme sign of bacilli. Genetic determinants of antilysozyme factors and kanamycin resistance may be used for the construction of vector systems of cloning in bacilli.     

The antilysozyme activity of meningococci

The antilysozyme activity of 184 meningococcal strains was studied. Such activity was found in all strains within the range 1-25 micrograms/ml, which was due to the heterogeneity of bacterial population. Strains varying in the total level of their lysozyme activity differed in the population structure by this sign. Preparations inhibiting antilysozyme activity could be differentiated according to the character of their action by the method of the population analysis. In the process of phagocytosis the selection of clones with a high level of antilysozyme activity was found to occur. The antilysozyme factor, heat resistant protein with a molecular weight of 10,000-30,000 D, was sensitive to trypsin.     

The role of the antilysozyme activity of Shigella flexneri as a factor controlling the degree of its resistance to the lytic action of bacteriophages

The level of the antilysozyme activity of S. flexneri in ensuring the high level of their phage resistance has been studied. The realization of the phage protective effect of antilysozyme activity has been noted to occur due to disturbances in the lysis of infected bacteria by phage-synthetized lysozyme-like enzyme. The direct relationship between the level of the lysozyme production of bacteriophages and their capacity for overcoming the antilysozyme protection of the host bacterium has been shown.     

Antilysozyme activity of anaerobic bacteria from fecal microflora in man

For the first time anaerobic bacteria of the fecal microflora in man have been found be capable of inactivating lysozyme. The presence of this antilysozyme sign has been noted in both Gram-positive anaerobes (Bifidobacterium, Propionibacterium, Eubacterium, Actinomyces israelii) and in Gram-negative anaerobes (Bacteroids, Prevotella melaninogenica). The expression of antilysozyme activity in the anaerobes under study has been determined. The possible biological role of this sign of the indigenous intestinal microflora has been discussed.     

Lysozyme-antilysozyme interactions in protozoa-bacteria communities (a model Tetrahymena-Escherichia community)]

Lysozyme and antilysozyme activities present in a wide range of microorganisms determine the so-called lysozyme-antilysozyme system of hydrobionts, which greatly contribute to the formation of aquatic biocenoses. However, the mechanism of the functioning of this system in natural freshwater communities remains obscure. The experimental investigation of lysozyme-antilysozyme interactions in a model Tetrahymena--Escherichia community showed that the antilysozyme activity of Escherichia coli leads to incomplete phagocytosis, thus enhancing bacterial survival in a mixed culture with infusoria. The selection and reproduction of bacterial cells resistant to grazing by infusoria determine the character of host-parasite interactions and allow bacteria to survive. It was demonstrated that the antilysozyme activity of microorganisms, which is responsible for bacterial persistency in natural biocenoses, is involved in the maintenance of protozoa-bacteria communities in bodies of water.     

Method of determining the antilysozyme activity of microorganisms

The method for the determination of the antilysozyme activity of microorganisms is described. The method consists in the cultivation of the strains under study in a lysozyme-containing medium, and the effect of lysozyme inactivation is determined from the growth of Micrococcus luteu S indicator strain adjacent to active strains. The quantitative evaluation of this property is presented. The study of 1 296 strains belonging to 9 genera has disclosed that antilysozyme activity occurs most frequently among Gram-negative bacteria.     

Investigation of Lysozyme–Antilysozyme Interactions in a Model Tetrahymena–EscherichiaCommunity

Lysozyme and antilysozyme activities present in a wide range of microorganisms determine the so-called lysozyme–antilysozyme system of hydrobionts, which greatly contribute to the formation of aquatic biocenoses. However, the mechanism of the functioning of this system in natural freshwater communities remains obscure. The experimental investigation of lysozyme–antilysozyme interactions in a model Tetrahymena–Escherichiacommunity showed that the antilysozyme activity of Escherichia colileads to incomplete phagocytosis, thus enhancing bacterial survival in a mixed culture with infusoria. The selection and reproduction of bacterial cells resistant to grazing by infusoria determine the character of host–parasite interactions and allow bacteria to survive. It was demonstrated that the antilysozyme activity of microorganisms, which is responsible for bacterial persistency in natural biocenoses, is involved in the maintenance of protozoa–bacteria communities in bodies of water.     

Characteristics of Klebsiella pneumoniae plasmid bearing the markers of drug resistance and antilysozyme activity

Electrophoretic study of the profile of plasmid DNA in agarose gel has shown the presence of a plasmid with a molecular weight of 55-60 MD in K. pneumoniae strains possessing antilysozyme activity. Plasmid pAlz60 of K. pneumoniae 22-110, isolated from the blood of a septicemia patient, is a fi- type conjugative plasmid. This plasmid is transferred to recipient strains of different species of enterobacteria with a frequency of 1 X 10(-5) to 1 X 10(-7). Simultaneously with the transfer of the plasmid, recipient cells inherit the antilysozyme markers and resistance to a number of drugs. The discovered plasmid has one restriction site for each of endonucleases EcoRI and XhoI and 16-20 sites for restrictases KpNI, BglII and Hind III.     

The antilysozyme activity of yeasts in the genus Candida

The fungi of the genus Candida isolated from patients with oral mucosa candidiasis and from candidiasis carriers have been studied for their antilysozyme activity. These fungi (Candida albicans, C. tropicalis, C. krusei, C. quilliermondii) are antilysozyme-active. A high antilysozyme activity of the fungi isolated from patients with oral mucosa candidiasis permits supposing that the presence of this trait may be one of the factors of microorganism pathogenicity. The effective antimycotic therapy (clotrimasole, sanguirhitrin) decreases the antilysozyme activity of fungi of the genus Candida.     

Enzymatic and immunochemical properties of lysozyme—Restriction of recognition of lysozyme antigenic determinants in pigs

Pigs immunized with lysozyme responded by producing only nonprecipitating antibody throughout the immunization period. Fig antilysozyme antibodies were found to be resistant to papain fragmentation, only 33% of the antibodies were fragmented with papain. From the binding of fluorescein labeled or ¹⁴C-labeled lysozyme to antilysozyme antibodies it was concluded that the antibodies elicited in pigs recognized only two antigenic determinants of lysozyme. These results were confirmed from the binding of Fab fragments to ¹⁴C-lysozyme. Fab fragments prepared from precipitating rabbit antilysozyme antibody bound ¹⁴C-lysozyme at a molar ratio of Fab/lysozyme = 3. Therefore nonprecipitating antibodies are the outcome of recognition of only two antigenic determinants on lysozyme and inability to form a lattice structure when antibody and antigen interact. This work emphasizes the limitations of using antibodies as a biological reagent for delineating the antigenic determinants on proteins.     

Characterization of antilysozyme and anticomplement signs in brucellae during experimental infection

The dependence of the course of experimental infection on the initial level of antilysozyme activity of Brucella clones was studied using 40 noninbred guinea pigs. As revealed in this study, the antilysozyme sign of brucellae influenced the pathogenesis of brucellosis, facilitating the prolonged survival of brucellae in the host body. The results thus obtained may be used for the prognostication of the course of Brucella infection.     

Persistent properties of protozoa-associated bacteria

The species structure and persistent properties (antilysozyme and antihistone activity) of bacteria forming associations with protozoa is revealed. Among them, 68.9% of the isolates were enterobacteria, the remaining organisms belonged to the families Aeromonas, Alcaligenes, Pseudomonas, Vibrio, etc. Within the family Enterobacteriaceae bacteria of the Escherichia group prevailed. 50.4% of the isolates were found to have antilysozyme activity and 97%--antihistone activity. The level of persistent properties in the representatives of allochthonous microflora was higher than that in the representatives of autochthonous microflora. In addition to antilysozyme activity antihistone activity was noted in protozoa-associated bacteria, which could be of importance for the formation of symbiotic links in natural associations. These data may be used in sanitary and hygienic practice for microecological monitoring of the environment.     

Circadian rhythms of antibacterial resistance, coagulase and antilysozyme activity of Staphylococcus aureus

AIM: To determine daily dynamics of antibacterial resistance as well as antilysozyme and coagulase activity of S. aureus strains. MATERIALS AND METHODS: On an example of clinical strains of S.aureus isolated from patients with surgical infections daily dynamics of biological characteristics of staphylococci was studied. After 12 hours of incubation strains were tested for coagulase activity by standard method (test tube method), antilysozyme activity by photometric method, and antibacterial resistance by method of serial dilutions in agar. Tests were repeated each 3-hours during a day. RESULTS: Variation of levels of studied biological characteristics of staphylococci during a day was revealed. Structures of coagulase and antilysozyme circadian rhythms had some differences in different S. aureus strains. Alongside with it, similarity in temporal expression of such biological characteristics of staphylococci as antibacterial resistance and antilysozyme activity was noted. CONCLUSION: Obtained data open prospect to use biorhythmological approach in study of biological characteristics of microorganisms during evaluation of their mechanisms of adaptation to changing environmental conditions. Chronobiological approach allows to reveal periods of maximal expression of S. aureus characteristics that could be used for increasing of effectiveness of antibacterial treatment by the choice of optimal time for administration of antibiotic.     

The antilysozyme activity of Mycobacterium tuberculosis L forms

The method for the detection of antilysozyme activity (ALA) in M. tuberculosis L forms was developed. The level of ALA in M. tuberculosis L forms isolated from patients with different clinical forms of the disease varied within 1-5 micrograms. M. tuberculosis L forms with the ALA level > 4 micrograms were isolated from patients with the progressing course of the disease. The method for the prognostication of the course of the tuberculous process in the lungs by the results of the antilysozyme test was proposed.     

Persistence factors of Francisella tularensis

The study of the persistence potential of 64 F. tularensis strains isolated from different sources was carried out. The wide spread of the antilysozyme, antilactoferrin and anticomplementory activities of F. tularensis were detected. F. tularensis, isolated from ticks and water, were characterized by the highest level of the expression of antilysozyme activity, while anticomplementory and antilactoferrin activities of the infective agents were characteristic of those microorganisms which were isolated from rodents and their excrements.     

Effect of vaginal Lactobacillus metabolites on biological properties of Staphylococcus epidermidis and Escherichia coli

The influence of supernatants and cell extracts of vaginal lactic acid bacilli on the growth, catalase, antilysozyme and anticomplementary activity of S. epidermidis and E. coli was studied. Changes observed in the persistence characteristics of these microorganisms were found to develop in different directions: in S. epidermidis a decrease in antilysozyme activity was mainly observed, while in E. coli the suppression of anticomplementary activity was registered. The modifying influence of lactic acid flora, revealed in this investigation, is regarded as one of the mechanisms facilitating the formation of microbiocenosis and ensuring colonization resistance.     

Effect of gastrointestinal tract secretions on the anti-lysozyme activity of enterobacteria

The effect of the duodenal contents and its components on the antilysozyme activity (ALA) of enterobacteria was studied. Under the action of native bile and the preparation of dried bile the stimulation of the expression of ALA factor in 86.6-93.3% of enterobacteria was observed. Pancreatin increased the level of the secretion of ALA factor by 50% in comparison with the control. The preparation of egg lysozyme produced no changes in the frequency and level of the expression of ALA factor. The study revealed the dose-dependant character of the action of duodenal contents: in the native form, as well as at concentrations of 50% and 25%, duodenal contents stimulated the expression of the ALA factor of enterobacteria; at concentrations of 12.5% and lower, its effect could be stimulating, inhibitory or neutral.     

Antibiotic resistance of coryneform bacteria isolated from the reproductive tract of women

The species composition, antilysozyme activity and antibiotic resistance of coryneform bacteria, isolated from the reproductive tract of women with microecological disturbances, were studied. Sixty six women without microecological disturbances and 102 female patients with microecological disturbances in the reproductive tract were examined. The study showed that Corynebacterium minutissimum, C. amycolatum, C. group JK, C. bovis and C. pseudodiphtheriticum prevailed in the healthy women. In the patients with microecological disturbances in the reproductive tract C. vitarumen, C. matruchotii, C. striatum, C. renale and C. urealyticum were detected in addition to the above species. The average antilysozyme activity of the coryneform bacteria isolated from the healthy women was 1.32 +/- 0.47. In the patients with microecological disturbances in the reproductive tract it was 1.84 +/- 0.38. The in vitro susceptibility of the coryneform bacteria to antimicrobial agents was determined. High prevalence of resistance to beta-lactams (oxacillin and penicillin), erythromycin and co-trimoxazole was detected. Thus, the species variety and the antilysozyme activity of the coryneform bacteria in the reproductive tract of the women with microecological disturbances were found to be higher. The high prevalence of resistance to oxacillin, penicillin, erythromycin and co-trimoxazole in the coryneform bacteria isolated from the patients with the microecological disturbances did not differ from that in the healthy women.     

Meningococcal plasmids

In Neisseria meningitidis strains isolated from patients with meningococcal infection and from carriers, the presence of plasmids has been detected by the modified method of H. C. Bernboim and J. Doly. The strains have been studied with respect to their pathogenicity and sensitivity to antibacterial substances. No correlation between the presence of plasmids and pathogenicity factors, such as antilysozyme activity, the presence of hyaluronidase and the positive reaction to Congo red, resistance to penicillin, serogroup and the sources of isolation, has been established.