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Introduction: Production of strawberries in greenhouses and polytunnels is gaining popularity worldwide. This study investigated the effect of reuse of coir and peat, two substrates commonly adapted to soilless strawberry production, as well as stand-alone wood fiber from Norway spruce, a promising substrate candidate. Methods: The experiment was performed in a polytunnel at NIBIO Apelsvoll, Norway, and evaluated both virgin substrates, as well as spent materials that were used in one or two years. Yield, berry quality and plant architecture of the strawberry cultivar 'Malling Centenary' were registered. In addition, chemical and physical properties of virgin and reused substrates were investigated. Results: While plants grown in peat and wood fiber had highest yield in the first year of production, the berry yield was slightly reduced when these substrates were utilized for the second and third time. However, yield was comparable to the yield level attained in new and reused coir. Interestingly, berries grown in wood fiber had a tendency to a higher sugar accumulation. This substrate also produced the highest plants. Stand-alone wood fiber was the substrate with the highest accumulation of nitrogen during the three consecutive production cycles. All three investigated materials revealed a trend for decreased potassium accumulation. Wood fiber is characterized by the highest percentage of cellulose, however after three years of production the cellulose content was reducedto the same levels as for coir and peat. Discussion: Implementation of wood fiber as a growing medium, as well as general practice of substrate reuse can be therefore an achievable strategy for more sustainable berry production. [ABSTRACT FROM AUTHOR]