To further characterize the immune response elicited by two live Newcastle disease vaccines, humoral, cellular and mucosal immunity was evaluated after oculo-nasal vaccination of day-old chickens. The preferential replication sites for each vaccine strain were investigated by screening different tissues using quantitative real-time reverse transcription-polymerase chain reaction (QRRT-PCR). The interference of maternally derived antibody with vaccination was also considered in conventional layer chickens. In SPF chickens, similar humoral immune-response was measured in blood and tears but a differential profile of cell-mediated immunity was observed according to the vaccine strain. The lung-associated humoral immunity was higher with the tracheotropic strain while the enterotropic vaccine induced a more important specific immunity in the digestive tract. The presence of maternally derived antibody in conventional layer chickens limited, if not completely abrogated, their immune responses to vaccination. This study increases our understanding of the protective immune response against Newcastle disease virus (NDV) and provides new useful informations for the development and evaluation of new types of vaccines.