Резюме
Мінухін В.В., Кочнєва О.В., Граматюк С.М., Крячко С.С. Комбінована дія протимікробних речовин на консорціум C. albicans і S. aureus.
Консорціум C. albicans і S. aureus займає третє місце серед причин виникнення полімікробних захворювань та у 27% випадках ці патогени є причиною госпітальних інфекції В патогенезі інфекційних захворювань, близько 80% всіх мікробних інфекцій у людини протікають з утворенням біоплівок. У складі біоплівок мікроорганізми в 50-500 разів більш стійкі до дії дезинфікуючих речовин, антибактеріальних препаратів, бактеріофагів, антитіл і фагоцитиів. Залишається актуальним питання визначення нових комбінацій протимікробних засобів ефективних відносно консорціума C. albicans і S. aureus. В результаті проведеного дослідження було встановлено, що здатність до формування біоплівок ізолятів С. аlbicans і S. aureus різна та найбільші показники оптичної щільності виявились у біоплівок утворених асоціацією цих мікроорганізмів. Комбінація лікарських речовин тербінафін, еритроміцин та ефірна олія полину мали високу ефективність відносно косорціума С. аlbicans і S. aureus, МІК для планктонних клітин склала 2,6 мкг/мл, для біоплівок 6,2 мкг/мл.
Ключові слова: консорціум С. аlbicans і S. aureus, комбінована дія, протимікробні препарати.

Резюме
Минухин В.В., Кочнева А.В., Граматюк С.М., Крячко С.С. Комбинированное действие противомикробных веществ на консорциум C. albicans и S. aureus.
Консорциум C. albicans и S. aureus занимает третье место среди причин возникновения полимикробных заболеваний и в 27% случаев эти патогены являются причиной госпитальных инфекций. В патогенезе инфекционных заболеваний, около 80% всех микробных инфекций у человека протекают с образованием биопленок. В составе биопленок микроорганизмы в 50-500 раз более устойчивы к действию дезинфицирующих веществ, антибактериальных препаратов, бактериофагов, антител и фагоцитов. Остается актуальным вопрос определения новых комбинаций противомикробных средств эффективных относительно консорциума C. albicans и S. aureus. В результате проведенного исследования было установлено, что способность к формированию биопленок изолятов С. аlbicans и S. aureus разная и наибольшие показатели оптической плотности оказались у биопленок образованных ассоциацией этих микроорганизмов. Комбинация лекарственных веществ тербинафин, эритромицин и эфирное масло полыни имели высокую эффективность в отношении косорциума С. аlbicans и S. aureus, МПК для планктонных клеток составила 2,6 мкг / мл, для биопленок 6,2 мкг / мл.
Ключевые слова: консорциум С. аlbicans и S. aureus, комбинированное действие, противомикробные препараты.

Summary
Minuкhin V.V., Kochneva E.V., Gramatyuk S.N., Krychko S.S. Combined action of antimicrobial substances on consortium C. albicans and S. aureus.
Consortium S. aureus and C. albicans ranks third among causes of polymicrobial disease, and in 27% of cases, these pathogens cause nosocomial infection in the pathogenesis of infectious diseases, about 80% of all microbial infections in humans occur with the formation of biofilms. In the bacteria in biofilms 50-500 times more resistant to disinfectant, antibiotics, bacteriophages, and antibody phagocytes. The issues of identifying new combinations of antimicrobial products effective against consortium C. albicans and S. aureus. As a result of this study found that the ability to form biofilms and S.aureus isolates C.albicans different and the highest rates of optical density appeared in biofilms formed by the association of these microorganisms. Combination drugs terbinafine, erythromycin and essential oil of wormwood had relatively high efficiency consortium C. albicans and S. aureus, the minimum inhibitory concentration for planktonic cells was 2.6 micrograms / ml for biofilms 6.2 mg / ml.
Key words: consortium C. albicans and S. aureus, combined action, antimicrobial drugs.

Рецензент: д.мед.н. С.І. Похил

УДК 582.282.23:579.861.2:57.083.2:615.28

Харківський національний медичний університет

Харьковский национальный медицинский университет

проспект Ленина, 4, Харьков, 61022,Украина

Kharkiv National Medical University,

4 Lenin Avenue, Kharkiv,61022,Ukraine
 

falkone28@mail.ru

Topicality of the theme. To date, it remains questionable as to understand the existence of bacteria as social communities, which form numerous associations. The components of these associations are active together, forming a specifically organized and attached to the substrate biofilms. In biofilm bacteria combined complex intercellular connections that maintain "a sense of quorum" [1].

In the pathogenesis of infectious diseases, according to U.S. researchers, about 80% of microbial infections in humans occur with the formation of biofilms. 
As part of the microorganisms in biofilms 50-500 times more resistant to disinfectant agents, antibacterial agents, bacteriophages, and antibody fahotsytyiv [2]. 
Trying to survive in environments microorganisms have acquired the ability to form interspecies communities based on respect synergies. In the study of associations formed by S.aureus and C.albicans was found that yeast cells on their surface with specific receptors (Als3p), which detect and bind cells of Staphylococcus aureus. The ability of C.albicans hyphae-elements penetrate the epithelium allows cells to Staphylococcus aureus freely move into tissues [3].Such relationships between these microorganisms have a great influence on the development and course of infectious diseases that cause difficulty for diagnosis and treatment. 

That why Consortium S.aureus and C.albicans ranks third among causes of polymicrobial disease and in 27% of cases, these pathogens cause nosocomial infection [4]. According to researchers C.albicans ranks fourth among the etiological factors causing sepsis and catheter-associated infections, resulting in high mortality [5]. 
Important for the development of medicine is the study of the mechanisms of formation of microbial resistance to chemotherapeutic drugs. 
Integrated use of antimicrobial prebarativ is the standard treatment of bacterial infections. The optimal combination of chemotherapy drugs allows to extend the range of actions and enhance antibacterial effect, especially in mixed infections when the causative agents of microbial associations have different sensitivity. 
In many countries scientists conducted research to determine the combined effect of antimicrobials for infections that are caused by C.albicans association and S.aureus. [5], but to date range of chemotherapeutic agents regarding use of this consortium is insufficient. 
Experimental studies conducted in this field, aimed at exploring ways to the development of resistance of microorganisms to chemotherapeutic drugs and their interaction processes in interspecific associations, the ability to form biofilms formed C.albicans and S.aureus, factors of pathogenicity and virulence. However, it remains questionable identification of new combinations of antimicrobials effective according to C.albicans and S.aureus Consortium. 

Goal of research. Determine the combined effect of antimicrobial agents on the association of isolates and reference strains S.albicans and S.aureus.

Materials and Methods. In research we used 10 S.aureus strains isolated from patients with various purulent - inflammatory and 4 reference strains ATCC 25923 as a control group. Also studied 10 strains of C.albicans, which were isolated from sputum from patients with pneumonia and reference strain ATCC 885.

The ability of microorganisms to form biofilms was determined by optical schylnistyu longed wave at 545 nm [6]. The minimum inhibitory concentration (MIC) of drugs (terbinafine, erythromycin, essential oil of wormwood) in association S.aureus  and S.albicans determined by serial dilutions. The results were evaluated by optical density at longed wave 545 nm on analyzer LabLine-90. The data were processed by a package of Excel.

The results of research. In determining the ability to form biofilms isolates and reference strains of S.aureus and  S.albicans were obtained the following data (Table 1)

The indication of average optical density for biofilms S.albicans and S.aureus С.albicans і S.аureus

Note: * significant difference p <0.05. The results of 3 experiments.

Analyzing the data of the study it was found that the performance of optical schylnisti in biofilms formed by association S.albicans and S.aureus were higher than the biofilm formed by certain microorganisms. The average optical schylnist for S.aureus isolates formed biofilms was 0,0680 ± 0,007 od.OSch for reference strains 0,0450 ± 0,005 od.OSch. For biofilms formed by isolates and reference strains S.albicans optical parameters were schylnosti 0,0765 ± 0,008 and 0,0550 ± 0,006 od.OSch respectively. In determining the ability to form biofilms formed by association S.albicans and S.aureus optical density performance for isolates were to 0,0987 ± 0,007 od.OSch for reference strains 0,0776 ± 0,004 od.OSch.

In determining the MIC of drugs (terbinafine, erythromycin, essential oil of wormwood) in association S.aureus and S.albicans following results were obtained (tab.2)

The average optical density when determining the MIC

terbinafine, erythromycin and essential oil of wormwood for planktonic cells and S.albicans and S.aureus biofilms.

Note: * - significant difference of p <0.05. The results of 3 experiments.

In determining MICs of terbinafine for planktonic cells S.albicans was 4 mcg / ml for biofilms 32 mg / ml, the average optical density for planktonic cells were 0,0483 ± 0,005 units. OSCH and 0,0715 ± 0,007 units. OSCH for biofilms.

In determining MICs of terbinafine for planktonic and biofilm cells of S.aureus values ​​were 16 and 32 mg / ml, the average optical density of 0,0543 ± 0,015 units. OSCH and 0,0754 ± 0,01 m. OSCH respectively, indicating a relatively ineffective substances microorganisms.

In determining the action of erythromycin average optical density for planktonic cells and biofilms S.albicans was 0,0775 ± 0,006 and 0,0931 ± 0,008 units. OSCH, MIC was 10 and 10 mg / ml, respectively. For S.aureus average optical density was 0,0505 ± 0,005 and 0,0688 ± 0,008 units. OSCH, MIC was 1.25 and 2.5 mg / ml.

The average optical density in the determination of the essential oil of wormwood for planktonic cells and biofilms were S.albicans 0,0734 ± 0,005 and 0,0882 ± 0,006 units. OSCH, MICs of 0.8 and 1.6 mg / ml, respectively. MICs for S.aureus was 1.6 and 3.1 mg / ml, the average optical density of 0,0624 ± 0,008 and 0,0782 ± 0,01 m. OSCH respectively.

In determining the combined effect of drugs have been used by their half-concentration. MICs of terbinafine, erythromycin and essential oil of wormwood for association S.albicans and S.aureus were 2.6 mg / ml for planktonic cells and 6.2 mg / ml for biofilms, the average optical density was 0,0451 ± 0,007 and 0 0596 ± 0,008 units. OSCH respectively.

Conclusions: The result of the study it was found that the ability to form biofilms isolates for S.aureus and S.albicans different and largest values ​​of optical density appeared in biofilms formed by the association of these microorganisms. The coexistence of these microorganisms in the consortium enhances their virulence and pathogenicity factors that determine their polirezystentnist to chemotherapeutic drugs. A combination of drugs terbinafine, erythromycin and essential oil of wormwood had relatively high efficiency consortsium S.albicans and S.aureus, MICs for planktonic cells was 2.6 mcg / mL for biofilms 6.2 mg / ml.

A promising direction is to study the interaction of microorganisms in associations and mechanisms of development of resistance polymicrobial infections.
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