Adventitious rooting might be induced in recalcitrant woody genotypes by infection with Agrobacterium
rhizogenes, and, in some cases, might also require exogenous auxin. The objective of the present study was to
determine how agrobacteria trigger root formation in the stem of a recalcitrant woody microcutting, which
cytological events result from the combined presence of infection and exogenous auxin, and which types of roots
are induced by infection. Microcuttings of a recalcitrant walnut (Juglans regia), infected or not with A. rhizogenes
strain 1855, were cultured with either indolebutyric acid (IBA), IAA, or without exogenous hormones, to induce
rhizogenesis. They were cytohistologically and ultrastructurally investigated at various times in culture. Southern
blot and PCR analyses were performed to verify the frequency of transgenic, chimeric and bacterium-containing
roots. The infection was sufficient per se to stimulate rhizogenesis. Rooting on the infected cuttings was enhanced
by exogenous IBA, which accelerated and increased root meristemoid formation, in comparison with without-hormone treatment. Meristemoids were organized both directly by the cambial cells and indirectly by the callus,
and showed a pluricellular origin. Inter and intracellular bacteria were observed in the stem throughout the culture
period (30 d). They were preferentially present in the vessels, and mainly in those showing polyphenol deposition.
In the infected IAA-treated cultures, a high level of secondary xylem formation occurred instead of rhizogenesis.
Nontransformed roots were preferentially produced by the infected cuttings treated with the auxins. Bacterium-containing and chimeric roots were produced by infected cuttings independently of the treatment. Thus, in a
recalcitrant walnut, nontransformed root meristemoids are stimulated by combining infection and exogenous
indolebutyric acid. Furthermore, the persistence of bacteria in the stem during the culture and the pluricellular
origin of the meristemoids explain the presence of the bacterium-containing and chimeric roots.