The Inhibitory Activity of Lactobacillus plantarum Supernatant against Enterobacteria, Campylobacter, and Tumor Cells.

Cell-free supernatant of Lactobacillus plantarum exhibit a strong antimicrobial effect against a number of pathogenic enterobacteria (E. coli, Shigella flexneri, Salmonella typhimurium, Proteus mirabilis, and Campylobacter jejuni). The degree of growth inhibition in broth culture reached a high level for all tested bacteria. The highest rates were noted for P. mirabilis (by 13 times) and the lowest for S. flexneri (by 5 times) and C. jejuni (by 4.5 times). Significant antiproliferative effect of the supernatant on cells of tumor-derived epithelial cell lines was shown. The highest degree of inhibition (by 22 times) was observed for HT-29 cells (colon carcinoma). Thus, inclusion of probiotics in traditional treatment schemes can increase the effectiveness of antibacterial and antitumor drug therapy.

Molecular Genetic Characterization of Streptococcus pyogenes Strains Isolated from Patients with Various Manifestations of Streptococcal Infection.

In 82 clinical strains of Streptococcus pyogenes (group A streptococci) isolated from patients with various manifestations of streptococcal infection, emm-typing revealed 27 emm-types (n=77) with a predominance of emm-89 (n=15; 18%), emm-75 (n=9; 11%), and emm-1 (n=6; 7%); types emm-3, emm-12, and emm-58 (n=4; 5% each) were found with almost equal frequency; other types were less common. The superantigen genes speC, speG, speH, speI, speJ, speK, speL, speM, smeZ, and SSA were identified in S. pyogenes strains using multiprimer PCR; the genes of the superantigen SpeA and cysteine proteinase SpeB were detected using real-time PCR. All the studied S. pyogenes strains contained superantigen genes, and 98% of the strains had several (from 2 to 7) genes. The number of variants of these sets reached 37; 2% of the strains contained only one superantigen gene. The distribution frequencies of superantigen genes in the studied strains were: speA - 43%; speC - 38%; speG - 93%; speH - 13%; speI - 6%; speJ - 24%; speK - 13%; speL and speM - 11% each; smeZ - 98%; SSA - 15%. All studied S. pyogenes strains contained the speB gene. Our studies have demonstrated that the sets of superantigen genes of group A streptococci are characterized by pronounced diversity to some extent associated with emm-type.

Kinetics of Serum Cytokine Profile in Mice after Injection of Supernatants of Group A Streptococci Culture.

We analyzed cytokine profile in the sera of CBA mice in 1, 5, and 24 h after intraperitoneal injection of supernatants of broth cultures of group A Streptococcus types 1M and 3M and Dochez NY5 type 10M strain. The increase of the cytokine content was observed in response to supernatants of all three types, but the highest values were recorded after injection of supernatant of strain Dochez-NY5. The level of IL-2 increased most drastically (by 51 times) and the level of IL-5 increased by 8.9 times in comparison with the control. The level of IL-2 also increased after injection of supernatants of type 1M and type 3M, but to a lesser extent (by 5 and 2.3 times, respectively). The content of proinflammatory cytokines IL-1ß, TNFα, and IFNγ in mouse sera increased to a lesser extent than IL-2 after administration of all three supernatants.

Effects of Miramistin and Phosprenil on Microbial Biofilms.

Effects of Miramistin and Phosprenil on biofilms of S. pyogenes, S. aureus, E. coli, L. acidophilus, and L. plantarum were studied. Significant differences in the effects of these substances on mature biofilms of microorganisms and the process of their formation were observed. Miramistin had significant inhibiting effects on the forming of biofilms and on the formed biofilms of all studied microorganisms. Treatment with Miramistin inhibited biofilm formation by 2-3 times compared to the control. This effect was found already after using of Miramistin in the low doses (3.12 µg/ml). Inhibition of the growth of a formed biofilm was observed only after treatment with Miramistin in the high doses (25-50 µg/ml). Phosprenil in the high doses (15-30 mg/ml) inhibited the forming of biofilms, especially the biofilms of S. pyogenes and L. plantarum (by 3-4.5 times). Treatment of formed biofilms with the agent in doses of 6.0 and 0.6 mg/ml was associated with pronounced stimulation of its growth in S. pyogenes, S. aureus, and L. acidophilus.

Interleukin-6 in Sera of Mice Inoculated with Group A Streptococcus and Cultural Supernatants.

Dynamics of IL-6 level was studied in blood serum of CBA mice receiving intraperitoneal injections of killed and live culture of group A Streptococcus and its supernatants. It was found that administration of killed culture was followed by a significant rise in IL-6 level (by 4.7 times in 1 h and by 9.3 times in 5 h in comparison with the control). By 24 h, cytokine content was below the control. The highest levels of IL-6 were found after treatment with supernatants of Streptococcus cultures (by 10.5 times in 1 h and by 14.9 times in 5 h, in comparison with the control). Administration of live culture was accompanied by an increase in IL-6 concentration by 3.2 times in 3 h. In this experimental series, the maximum level of IL-6 was found in 48 h (by 5.2 times), and then it gradually decreased below the control. Different dynamics of changes in IL-6 level after administration of killed and live cultures of group A Streptococcus may suggests that they activate different signal pathways.

Changed Serum Cytokine Profile in Mice in Response to Streptococcus A Culture.

Comparative analysis of serum cytokine profiles of CBA mice was carried out 1, 5, 24, and 48 h after intraperitoneal injection of killed culture of different streptococcus A types. The production of cytokines in response to different streptococcus types varied. The highest level was recorded in response to types 1M and 3T+M, more often detected in invasive streptococcal infection. The highest levels of IL-2 were recorded in response to 1M (47-fold increase in comparison with the control) and 3T+M streptococcus types (more than 10-fold increase). Injections of these types also led to an increase of IFN-γ level (15.6 and 11.3 times, respectively). The level of TNF-α increased less (3.6 times in response to 3T+M and 2.6 times in response to 1M type). The levels of IL-5, IL-10, and IL-12 increased 2-3-fold. Injections of 1T and 5M types led to just a 2-fold increase of cytokine levels. These data indicated induction of the immune response trend by mainly Th1 or mixed Th1/Th2 pattern in response to group A streptococcus antigens.

[Biofilm formation by group A Streptococci of various types and study of antibiotics effect on this process].

AIM: Evaluation of the ability to form biofilms by various M, T and MT-types of group A streptococci (GAS), as well as study of the effect of various antibiotics on biofilm formation. MATERIALS AND METHOD: 43 strains of various M and T type GAS were studied. The cultures were grown in Todd-Hewitt broth with the addition of 0.5% yeast extract. Comparative evaluation of the ability to form biofilm was carried out using photometry. Benzylpenicillin, oxacillin, cepha- losporin, cefuroxime and ceftriaxone antibiotics were used at various concentrations. RESULTS: GAS differ significantly by their ability to form biofilms. The highest ability was noted in 8 strains--2M, 9M, 12M, 13M, 19M, 30M, 36M-types and 6MT type. Simultaneous introduction of GAS cultures and antibiotics into the culture well, except for ceftriaxone, resulted in growth inhibition of both plankton cells and biofilms. CONCLUSION: The ability of GAS to form biofilm depends on streptococci serotype. During simultaneous introduction of GAS with antibiotics into the well, the biofilm does not form.