Apical necrosis and premature fruit drop of walnut result in significant yield losses, while these symptoms are associated with different microorganisms. Brown apical necrosis (BAN) is caused by Fusarium and Alternaria species, although this symptomatology can also be caused by Xanthomonas arboricola pv. juglandis (Xaj), causal agent of walnut blight disease. The objectives of this study were to determine the prevalence of the main microorganisms causing apical necrosis in walnut orchards of south-central and southern Chile; evaluate the effects of single vs. multiple species inoculation of microorganisms involved in the expression of apical necrosis and fruit drop; and determine the developmental stage(s) in which walnut flower or fruit is most susceptible to infection by those microorganisms under field conditions. Results showed that Xaj was the most prevalent pathogen causing apical necrosis in walnut orchards, while the incidence of BAN fungal microorganisms associated with apical necrosis in walnuts fruits varied between orchards and seasons. Infection trials demonstrated that the three microorganisms, acting either individually or as a mixed infection, can cause symptoms and premature drop of inoculated fruits, while the combined infection resulted in more frequent infections than those caused by each fungal pathogen separately. Full female flowering (Ff2) and stigma wilting (Gf) were the developmental stages with the greatest susceptibility to these microrganisms, while copper sprays decreased disease incidence. These results suggest that apical necrosis is caused by a complex of microorganisms and that the infection depends on the phenological stage at which the infection occurs.