Yu, U., Zhao, Y.-Q., Zhao, B., Ren, S. and Guo, Y.-D. 2011. Influencing factors and structural characterization of hyperhydricity of in vitro regeneration in Brassica oleracea var. italica. Can. J. Plant Sci. 91: 159-165. This study examines factors that affect the occurrence of hyperhydric tissue in in vitro cultures of Brassica oleracea variety italica. The anatomy of normal and hyperhydric leaves of plantlets regenerated from the hypocotyls was compared using scanning electron microscopy and transmission electron microscopy. In hyperhydric leaves palisade tissue was absent and the spongy mesophyll displayed large, unorganized intercellular spaces. Hyperhydric leaves had abnormal stomata with deformed guard cells. Significant ultrastructural differences were observed between chloroplasts in normal and hyperhydric leaves. The effects of zeatin, indoleacetic acid, silver nitrate and sucrose on the formation of hyperhydric shoots were studied. Zeatin was the most important factor, followed by sucrose concentration, AgNO₃ and indoleacetic acid. The process of hyperhydricity was found to be reversed by increasing the agar concentration and eliminating NH₄NO₃ from the macro-elements in the MS medium. This is the first report of hyperhydricity in Brassica oleracea, and our study gives a better understanding of the factors that influence hyperhydricity during in vitro regeneration in Brassica crops.