Kombucha is usually obtained from the fermentation of black or green tea by a consortium of acetic acid bacteria and yeasts. In this study, kombucha was prepared from the same starter consortium using green and black teas as well as, for the first time, an infusion of rooibos leaves (Aspalathus linearis). Microbial diversity was analysed during fermentation both in the biofilm and in the corresponding kombuchas, using culture-dependent and -independent methods. Polyphenols, flavonoids, ethanol, and acids were quantified and anti-oxidant activities were monitored. All of the Kombuchas showed similarity in bacterial composition, with the dominance of Komagataeibacter spp. Beta diversity showed that the yeast community was significantly different among all tea substrates, between 7 and 14 days of fermentation and between biofilm and kombucha, indicating the influence of the substrate on the fermenting microbiota. Kombucha from rooibos has a low ethanol concentration (1.1 mg/mL), and a glucuronic acid amount that was comparable to black tea. Although antioxidant activity was higher in black and green kombucha compared to rooibos, the latter showed an important effect on the recovery of oxidative damage on fibroblast cell lines against oxidative stress. These results make rooibos leaves interesting for the preparation of a fermented beverage with health benefits.

About 95% of global hydrogen production is made by fossil fuels using different technologies which are all characterized by high energy consumption and high carbon emissions. Alternatively, more sustainable production methods, such as biological fermentation processes, are under study. Dark fermentation, also called acidogenesis, entails the transformation of a great variety of organic substances into a mixture of organic and inorganic products, as well as gases (H2 and CO2). In this study we tested an exhausted fermentation broth, derived after Clostridium fermentation for H2 production, as a biostimulant via foliar application in an intensive apple orchard. Two different doses were applied upon dilution of the broth in water (100 mL L−1 and 10 mL L−1), evaluating the main fruit quality parameters (fresh weight, fruit diameter, dry matter, firmness, soluble solid content, color lightness, DA index) in addition to macro- and micro-nutrients and heavy metals concentrations. Chemical characterization of the broth showed a high amount of low-MW polypeptides (Trp-Glu-Lys, Ile-Pro-Ile, Phe-Pro-Lys, His-Pro) and organic acids (formic acid, butyric acid, butanedioic acid); moreover, quantitative analyses of inorganic ions showed no heavy metal detection but high concentrations of nitrogen, phosphorus and potassium, compatible with use in agriculture. The fruit quality parameters showed significantly higher mean fruit weight compared to the untreated trees, as well as higher dry matter. No statistical differences were recorded among the treatments for fruit firmness, diameter and yield. Soluble solids content in both treatments were significantly lower than the controls, whereas the DA index mean values were higher in both treatments compared to the controls, indicating a delay in fruit ripening probably due to the high nitrogen broth concentration. Regarding the chemical analyses of fruits, no particular differences were found among the treatments, except for Fe, which showed a significantly higher amount upon treatment with the lower dose. As concerns leaves, no phytotoxic symptoms were detected in both treatments, making the described exhausted broth a candidate for its use as a plant biostimulant. Additional studies are needed to evaluate the ideal application dose, identify further action targets and implement appropriate strategies to concentrate the biostimulant active compounds.