The character of PA3235 virulence factors of Pseudomonas aeruginosa PAO1 – a preliminary study

Abstract

Introduction: Many virulence factors of Pseudomonas aeruginosa PAO1 are regulated by temperature and host conditions upon infection. Based on microarray data, the PA3235 gene is one of the upregulated genes during cell growth at 37 °C. Until now, no information about its role in PAO1 pathogenicity. Methods: The PAO1∆PA3235 strain was constructed by overlapping polymerase chain reaction (PCR) method and through biparental mating. The deletion was confirmed by PCR and restriction analyses. The virulence factors of both P. aeruginosa PAO1 wild type and ∆PA3235 mutant strains were examined, which consisted of the amount of pyocyanin and pyoverdine, swarming, swimming, and twitching motility, biofilm formation, 3-oxo-dodecanoyl-homoserine lactone concentration, and growth curve profile. Data were analyzed using Student’s t-tests to determine differences between treatments. P-value < 0.05 were considered significant. Results: The ∆PA3235 strain was successfully constructed. At 37 ºC, the mutant produced less pyocyanin (p-value 0.0004), pyoverdine (p-value 0.0009), and swarm area than the wild-type. The dendrites pattern of both strains was similar. The mutant and parental strains showed no differences in swimming and twitching motility when incubated at 22 ºC and 37 ºC. The mutant produced more biofilm compared to the wild-type strain (p-value 0.0013). The AHL was higher in the mutant than in wild type strain (p-value 0.0095) after 24 h incubation. Both the wild type and mutant strains exhibited similar growth patterns in LB broth. The mutant colonies also showed the same morphology as the wild type on the LB plate (not shown here). Conclusion: The deletion of the PA3235 gene from the Pseudomonas aeruginosa genome caused some changes in virulence factors production, as the bacterium grew at body temperature 37°C. We predicted that the PA3235 gene might function to transport molecules involved in the early infection of this bacterium to humans.