Kaposi's sarcoma-associated herpesvirus (KSHV) can cause several human cancers, including primary effusion lymphoma (PEL), which frequently occur in immunocompromised patients. KSHV-infected patients often suffer from polymicrobial infections caused by opportunistic bacterial pathogens. Therefore, it is crucial to understand how these coinfecting microorganisms or their secreted metabolites may affect KSHV infection and the pathogenesis of virus-associated malignancies. Quorum sensing (QS), a cell density-based intercellular communication system, employs extracellular diffusible signaling molecules to regulate bacterial virulence mechanisms in a wide range of bacterial pathogens, such as Pseudomonas aeruginosa , which is one of the most common opportunistic microorganisms found in immunocompromised individuals. In this study, we evaluated and compared the influence on PEL growth and the host/viral interactome of the major QS signaling molecules [ N -(3-oxododecanoyl)- l -homoserine lactone (OdDHL), N -butyrylhomoserine lactone (BHL), and 2-heptyl-3-hydroxy-4-quinolone (PQS)] in conditioned medium from wild-type (wt) and QS mutant laboratory strains as well as clinical isolates of P. aeruginosa . Our data indicate that P. aeruginosa coinfection may facilitate virus dissemination and establishment of new infection and further promote tumor development through effectively inducing viral lytic gene expression by its QS systems. IMPORTANCE Currently, most studies about KSHV infection and/or virus-associated malignancies depend on pure culture systems or immunodeficient animal models. However, the real situation should be much more complicated in KSHV-infected immunocompromised patients due to frequent polymicrobial infections. It is important to understand the interaction of KSHV and coinfecting microorganisms, especially opportunistic bacterial pathogens. Here we report for the first time that P. aeruginosa and its quorum-sensing signaling molecules display a complicated impact on KSHV-associated lymphoma growth as well as the intracellular host/viral gene expression profile. Our data imply that targeting of coinfecting pathogens is probably necessary during treatment of virus-associated malignancies in these immunocompromised patients.