Abstract: Pine wilt disease has become a worldwide threat to pine forests and forest ecosystem. However, its mechanism has not been clearly illustrated, though most scientists working in the area believe that pine wood nematode (PWN) is the only pathogen of the disease. This study indicated the bacteria symbiotically associated with the nematode played an important part in the disease. A series of inoculation experiments were completed with a new developed experimental device, cold valve, by which the nematode and its associated bacteria were confined within the inoculated branch. Here this paper show that the pine wilt disease is a complex disease caused by a complex of PWN and its associated pathogenic bacteria. The results also indicated that without the movement of the two pathogens to other parts of the inoculated pine the axenic PWN the systemic toxins in the culture produced by the pathogenic bacterial strain resulted in wilt and death of the host. Based on the results of the study and reports in the mixed with the pathogenic bacterium, Pseudomonas fluorescens (GcM5-1A) or the cell free culture of the bacterium were able to cause the disease. The bacterium was reisolated from the dead host, inoculated with axenic PWN and the bacterial strain proving Koch’s postulates. The result’ from the inoculation of the cell free culture of the bacterial strain in indicated that it was the systemic toxins in the culture produced by the pathogenic bacterial strain that resulted in wilt and death of the host. Based on results of the study and previous reports of the authors on mutualistic symbiosis between PWN and its associated bacteria, the authors pointed out that pine wilt disease is the first experimentally confirmed plant disease induced by a mutualistic symbiotic nematode-bacterial complex.