Steelmaking slags are prospective base materials for seaweed beds, resulting from a continuous process of biofouling, starting from biofilm formation and leading to growing algae. While focusing on biofilm formation, we investigated specific features of steelmaking slags when utilized as a base for seaweed beds by comparing the bacterial communities in marine biofilms between steelmaking slags and artificially produced ones. Genomic DNA was extracted from the biofilms collected on days 3 and 7, and partial 16S rRNA libraries were generated and sequenced by second-generation next-generation sequencing. The read sequences were analyzed using QIIME 2™, then heatmaps and non-metric multidimensional scaling based on the Bray–Curtis dissimilarity index in the R program. Rhodobacteraceae and Flavobacteriaceae were the most dominant family members in all samples on both days 3 and 7. However, Mariprofundus, comprising iron-oxidative bacteria, was predominantly detected in the samples of steelmaking slags on day 7. This suggested that the growth of Mariprofundus was dependent on Fe(II) ion concentration and that steelmaking slags eluted Fe(II) ions more easily than artificial slags. In contrast, Sulfurovaceae, sulfur-oxidizing bacteria, were dominantly present in all samples on day 3, but decreased by day 7, regardless of the sulfur content. It was supposed that engine oil-derived sulfur compounds strongly influenced Sulfurovaceae growth, whereas slag-derived sulfur compounds did not. Heatmap analysis indicated that the submersion period significantly influenced the bacterial communities, regardless of the differences in the main slag content ratios. Summarizing these results, the elution characteristics of steelmaking slags have the potential to influence the formation of marine biofilms, and this formation is significantly influenced by environmental conditions. Full article

In ocean and coastal waters, the formation of biofilms on artificial matters often causes intractable phenomena such as the deterioration of surface functions and corrosion, resulting in significant economic damage. Thus, methods for inhibiting biofilm formation are in high demand, and many new anti-biofilm products are being designed on a daily basis. However, practical and safe assays for evaluating anti-biofilm formation have not yet been established. In this study, we developed a more practical and safer biofilm formation test system composed of a loop-type laboratory biofilm reactor (LBR) and HIG FST4 1, a Halomonas strain derived from ballast seawater, in comparison with a slowly rotating test-tube culture (TTC) test. To evaluate biofilm formation in an LBR and TTC, three materials (pure iron, pure aluminum, and soda lime glass) were tested, and Raman spectroscopic analysis was used for the identification and relative quantification of the biofilm contents. Regardless of the test method, biofilm formation progressed in the order of soda lime glass < pure aluminum < pure iron. The Raman peaks showed that the LBR test samples tended to remove polysaccharides compared to the TTC test samples and that the proportion of proteins and lipids in the LBR test samples was much higher than that of the TTC test samples. These results show that the combination of HIG FST4 1 and LBR is suitable for biofilm formation in a practical marine environment. Full article

The evaluation of biofilm formation is important, given the ubiquity and problematic nature of biofilms in industrial and medical settings, as well as in everyday life. Basically, biofilms are formed on substrates. Therefore, it is essential to consider the properties of the substrates during biofilm evaluation. The common dye staining method to evaluate biofilm formation requires a short evaluation time and enables the evaluation of a large area of the sample. Furthermore, it can be easily determined visually, and quantitative evaluation is possible by quantifying color adsorption. Meanwhile, the dye staining method has the problem of adsorption even on substrate surfaces where no biofilm has formed. Therefore, in this study, we focused on Ag⁺ reduction reaction to devise a novel biofilm evaluation method. Ag⁺ is highly reductive and selectively reacts with organic substances, such as saccharides, aldehydes, and proteins contained in biofilms, depositing as metallic Ag. First, to simply evaluate biofilm formation, we used a glass substrate as a smooth, transparent, and versatile oxide material. We observed that the amount of Ag deposited on the substrate was increased proportionally to the amount of biofilm formed under light irradiation. Upon comparing the Ag deposition behavior and adsorption behavior of crystal violet, we discovered that for short immersion times in AgNO₃ solution, Ag deposition was insufficient to evaluate the amount of biofilm formation. This result suggests that the Ag reduction reaction is more insensitive than the crystal violet adsorption behavior. The results of the Ag deposition reaction for 24 h showed a similar trend to the crystal violet dye adsorption behavior. However, quantitative biofilm evaluation using the proposed method was difficult because of the Ag⁺ exchange with the alkali metal ions contained in the glass substrate. We addressed this issue by using the basic solution obtained by adding an ammonia solution to aqueous AgNO₃. This can cause Ag⁺ to selectively react with the biofilm, thus enabling a more accurate quantitative evaluation. The optimum was determined at a ratio of distilled water to aqueous ammonia solution of 97:3 by weight. This biofilm was also evaluated for materials other than ceramics (glass substrate): organic material (polyethylene) and metal material (pure iron). In the case of polyethylene, a suitable response and evaluation of biofilm formation was successfully achieved using this method. Meanwhile, in the case of pure iron, a significantly large lumpy deposit of Ag was observed. The likely reason is that Ag precipitation occurred along with the elution of iron ions because of the difference in ionization tendency. It could be concluded that the detection of biofilm formation using this method was effective to evaluate biofilm formation on materials, in which the reduction reaction of [Ag(NH₃)₂]⁺ does not occur. Thus, a simple and relatively quantitative evaluation of biofilms formed on substrates is possible using this method. Full article

The impact of steroid hormones, and particularly estradiol, on human microbiota could be recognized as a substantial part of human-microbiota interactions. However, an area that remains poorly investigated is that of the skin and vaginal microbial communities and biofilms, which contain non-pathogenic bacteria of phyla Firmicutes and Actinobacteria, especially probiotic bacteria of the genus Lactobacillus and the widespread, safe skin genus, Micrococcus. Experiments with Lactobacillus paracasei AK508 and Micrococcus luteus C01 biofilms on PTFE cubes showed dose-dependent effects of estradiol at concentrations of 0.22 nM and 22 nM. The hormone mostly inhibits L. paracasei growth and stimulates M. luteus. The presented studies of colony-forming unit (CFU) amountsand cell aggregation in biofilms on glass fiber filters showed the same general tendencies. Estradiol generally increased the aggregation of cells in monospecies communities and potentially changed the synthesis of antibacterial metabolites in L. paracasei. The balance between two bacteria in mixed-species biofilms depended on the initial adhesion stage, and when this stage was reduced, micrococci were more resistant to the antagonistic action of L. paracasei. Moreover, in mixed-species biofilms, the effect of estradiol on lactobacilli altered from inhibition to stimulation, potentially due to the presence of M. luteus. At the same time, ethanol as a solvent for estradiol at the concentration 0.6% acted mostly as an antagonist of the hormone and had an opposite effect on bacteria; nevertheless, the overlapping of ethanol and estradiol effects was shown to be minimal. The data obtained prove the complexity of microbial interactions and the regulatory effect of estradiol on commensal bacteria biofilms. Full article