This study explored the pathways of ingress of Acidovorax citrulli, the causal agent of bacterial fruit blotch of cucurbits, into watermelon seeds. Up until 7days post-inoculation (DPI), a significantly higher percentage of watermelon seeds was infected with A.citrulli when the bacteria were applied (c. 1x10(6)colony-forming units) to stigmas versus ovary pericarps of female flowers. Immunofluorescence microscopy showed that, with stigma inoculation, A.citrulli colonized style and ovary tissues by 1 DPI, and the bacteria co-localized with pollen germ tubes in these tissues. With ovary pericarp inoculation, A.citrulli cells penetrated the epicarp and mesocarp tissues by 1 DPI but did not reach endocarp until 4 DPI. Finally, manual pollination followed by stigma inoculation led to >53% A.citrulli-infected seed lots, while A.citrulli was not detected in seeds/ovules generated by stigma inoculation without pollination (chemically induced parthenocarpy). These results show that stigma inoculation results in faster colonization of watermelon ovules by A.citrulli than pericarp inoculation, even though there is no difference in the levels of infection in mature seeds. The data also indicate that pollen germ tubes play an important role in A.citrulli ingress into watermelon seeds via stigmas.