Godovalov A P, Stepanov M S, Yakovlev M V, Kobzarenko E E, Batog K A
Acad. E.A. Wagner Perm State Medical University, Russian Federation.
Klin Lab Diagn. 2019;64(12):758-761. doi: 10.18821/0869-2084-2019-64-12-758-761.
Microorganisms are able to form biofilms on surfaces of biotic and abiotic nature. In turn, in human biotopes there are optimal conditions for the implementation of biofilm-forming activity. Moreover, in medical practice, polymeric materials are often used for drainage or prosthetics, which can also be successfully colonized by bacteria. However, in laboratory practice, the formation of biofilms is usually evaluated on glass or polystyrene. The purpose of the study is to evaluate the methodological features of studying the biofilm-forming activity of microorganisms on the surface of synthetic polymeric materials. We used strains of Staphylococcus aureus ATCC 25923, Escherichia coli K-12, Candida albicans ATCC 10231, as well as synthetic polymeric materials - DentLight Flow light-curing composite material (nano-hybrid fluid composite; Russia), glass ionomer chemical curing Fuji 1 (Japan), cement for temporary fixation of orthopedic constructions TempBond NE (USA), acrylic, polyurethane and polyvinyl chloride. The formation of biofilms in flat-bottomed ELISA plates in this study was considered as a control group. If the polymer belonged to cold curing materials, sterile flat-bottomed tablets were used, the bottom of which was filled with a thin layer of plastic. After hardening of the plastic, biofilms were formed in the tablets. In the second series of experiments, hot cured materials cut into equal parts 5×5×1 mm in size were placed in the wells of a plate and again used to determine biofilm formation with subsequent coloring. To extract the dye, the pieces were transferred to a new plate to exclude the amount of film biomass formed on the walls of the plate wells. In both cases, cultivation was carried out at 37° C for 24-48 hours. The biomass of the film was stained with fuchsin. Statistical data processing was performed using t-Student criterion. For the threshold level of significance, the value p <0.05 was taken. It is established that the proposed options for determining biofilm forming ability are available and indicative. It was revealed that the same microorganisms have individual biofilm formation indicators for each polymer material. The light curing dental composite and polyvinyl chloride exhibit the more pronounced antiadhesive properties than cements and polyurethane. Up to date, most of the studies of biofilm formation have been carried out using glass or polystyrene, which, as a rule, are not used for the manufacture of prostheses, catheters, drains, etc., which makes it difficult to assess the true film-forming activity of microorganisms. The proposed methodological approaches, especially the second option for preparing testing samples, solve this problem. In general, the proposed approaches to testing biofilm-forming activity on polymers are very simple to implement and generally available. For an adequate study of the biofilms formation, it will be advisable to use polymer materials, directly used in medicine, rather than polystyrene tablets, the material of which is found exclusively in laboratory practice.
微生物能够在生物和非生物表面形成生物膜。相应地,在人体生物环境中存在着实现生物膜形成活性的最佳条件。此外,在医学实践中,聚合材料常被用于引流或修复,细菌也能够成功地在这些材料上定殖。然而,在实验室实践中,生物膜的形成通常是在玻璃或聚苯乙烯上进行评估的。本研究的目的是评估研究微生物在合成聚合材料表面生物膜形成活性的方法学特点。我们使用了金黄色葡萄球菌ATCC 25923、大肠杆菌K-12、白色念珠菌ATCC 10231菌株,以及合成聚合材料——DentLight Flow光固化复合材料(纳米混合流体复合材料;俄罗斯)、玻璃离子化学固化富士1型(日本)、用于骨科结构临时固定的TempBond NE水泥(美国)、丙烯酸、聚氨酯和聚氯乙烯。本研究中,将平底酶标板中生物膜的形成作为对照组。如果聚合物属于冷固化材料,则使用无菌平底片剂,其底部填充一薄层塑料。塑料硬化后,在片剂中形成生物膜。在第二系列实验中,将切成5×5×1mm大小等份的热固化材料放入板孔中,再次用于测定生物膜形成并随后进行染色。为了提取染料,将切片转移到新的板中,以排除在板孔壁上形成的膜生物量。在这两种情况下,培养均在37℃下进行24 - 48小时。用品红对膜的生物量进行染色。使用t检验标准进行统计数据处理。对于显著性阈值水平,取p <0.05的值。已确定所提出的测定生物膜形成能力的方法是可行且具有指示性的。结果表明,相同的微生物对每种聚合材料都有各自的生物膜形成指标。光固化牙科复合材料和聚氯乙烯比水泥和聚氨酯表现出更显著的抗粘附性能。迄今为止,大多数生物膜形成的研究都是使用玻璃或聚苯乙烯进行的,而玻璃或聚苯乙烯通常不用于制造假体、导管、引流管等,这使得难以评估微生物真正的成膜活性。所提出的方法学方法,尤其是制备测试样品的第二种方法,解决了这个问题。总体而言,所提出的测试聚合物上生物膜形成活性的方法实施起来非常简单且普遍适用。为了充分研究生物膜的形成,建议使用直接用于医学的聚合材料,而不是聚苯乙烯片剂,聚苯乙烯片剂的材料仅在实验室实践中使用。