Non-sterilized fish waste containing fish bones was fermented using combined starter cultures of film-forming yeast (Candida ethanolica) and lactic acid bacteria (LAB; Lactobacillus casei and Lactobacillus rhamnosus) in order to obtain a liquefied fermented broth without spoiling. During the entire fermentation, the number of LAB cells was maintained at a high level (6 x 10(8)-5 x 10(7) cells/ml). Although the number of general bacteria was 10(6)cell/ml after adding non-sterilized fish biomass, its growth was suppressed to be 1-3 x 10(4) cells/ml. The entire biomass had completely liquefied and the fermented broth contained all 20 alpha-amino acids composed of protein and also various kinds of minerals in abundance. The weight of mice group fed the fermented broth content feed (sample feed) for 31 days significantly increased compared with that fed no broth feed (control feed) (2137 g vs 20.76 g (p < 0.05). No abnormal behavior and appearance were observed. All internal organs (the heart, the liver, the lung, the intestines, and the spleen) of both groups were confirmed to be normal by visual observation. In peripheral blood, the percentages of NK cells and CD8 + T cells of the mice in the sample feed group increased significantly relative to those in the control feed group (NK cells: 19% vs 11%, CD8 + T cells: 9% vs 5%, p < 0.05). In the spleen, the percentage of NK cells in the sample feed group also increased significantly compared to that in the control feed group (p < 0.05). The fermented fish biomass is expected to be effective for innate and adaptive immunity and thus fit for animal feed. (C) 2013, The Society for Biotechnology, Japan. All rights reserved.