Microbes within an infection impact neighbors\\\\\\\' pathogenicity. This study aimed to address in vitro virulence activity of Pseudomonas aeruginosa under the binary interaction with Acinetobacter baumannii or Enterococcus faecium, co-isolated from two chronic wound infections. The biofilm formation of Pseudomonas was enhanced 1.5- and 1.4-fold when it was simultaneously cultured with Acinetobacter and Enterococcus, respectively. Pseudomonas motility was increased by 1.9- and 1.5-fold (swimming), 3.6- and 1.9-fold (swarming), and 1.5- and 1.5-fold (twitching) in the dual cultures with Acinetobacter and Enterococcus, respectively. The synergistic hemolysis activity of Pseudomonas was observed with the heat-killed Acinetobacter and Enterococcus cells. The minimum inhibitory concentration of ciprofloxacin against Pseudomonas was increased from (μg mL-1) 25 to 400 in the individual and mixed cultures, respectively. The pyocyanin production by Pseudomonas in the single and mixed cultures with Acinetobacter and Enterococcus was (μg/mL) 1.8, 2.3, and 2.9, respectively. The expression of lasI, rhlI, and pqsR genes was up-regulated by 1.0-, 1.9-, and 16.3-fold, and 4.9-, 1.0-, and 9.3-fold when Pseudomonas was incubated with Acinetobacter and Enterococcus, respectively. Considering the entire community instead of a single pathogen may lead to a more effective therapeutic design for persistent infections caused by Pseudomonas.

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