In vitro arachidonic acid metabolism by polymicrobial biofilms of Candida albicans and Pseudomonas aeruginosa

[摘要] English: Interkingdom interactions between microorganisms facilitate the growth and survival of microbial communities, and the understanding of these interactions can be beneficial to mankind. These interactions may also be detrimental to human health, with the combination of virulence factors promoting the survival of microbial populations during infection. A relevant model for this, is the interaction between Candida albicans and Pseudomonas aeruginosa. These two microorganisms are frequently found together at infection sites. During infection, C. albicans and P. aeruginosa elicit the release of arachidonic acid (AA), utilized by the host to form immunomodulatory compounds termed eicosanoids. Among these is prostaglandins, for example, prostaglandin E2 (PGE2). Prostaglandin E2 can inhibit Th1 and promote Th2 responses. In combination to other eicosanoids, they modulate inflammation in hosts, ultimately affecting the ability of the host to clear infection. Candida albicans and P. aeruginosa are also able to produce immunomodulatory eicosanoids from exogenous AA. This study confirms the production of PGE2 by C. albicans monomicrobial biofilms, together with the production of PGF2α and 15-hydroxyeicosatetraenoic acid (15-HETE) with the use of enzyme-linked immunosorbent assay (ELISA) and LC-MS/MS for PGE2 confirmation. The production of these eicosanoids is also reported here by P. aeruginosa monomicrobial biofilms. This study is the first to identify authentic PGE2 production by P. aeruginosa biofilms. In addition, polymicrobial biofilms were shown to produce significantly more eicosanoids that monomicrobial counterparts, possibly contributing to the increased morbidity during co-infection by these pathogens.Although the pathways and enzymes involved in eicosanoid production by mammalian systems have been well studied, the production of eicosanoids by microorganisms requires much research. This is due to the fact that microorganisms frequently don't possess homologs to mammalian enzymes responsible for eicosanoid production. Therefore, inhibitors previously identified to inhibit C. albicans PGE2 production, were evaluated in terms of their effect on eicosanoid production by monomicrobial and polymicrobial biofilms of C. albicans and P. aeruginosa. The inhibitors used are acetylsalicylic acid (ASA, a cyclooxygenase inhibitor), ammonium tetrathiomolybdate (ATM, inhibition of copper-dependant enzymes) and nordihydroguaiaretic acid (NDGA, a potent antioxidant inhibiting various enzyme classes). A possible 'shift in eicosanoid production by C. albicans is seen in the presence of ASA as well as ATM. This phenomenon is also seen for P. aeruginosa in the presence of ATM. Interestingly, ASA increased eicosanoid production by P. aeruginosa. The anti-oxidant NDGA decreased eicosanoid production by monomicrobial, as well as polymicrobial biofilms. Different profiles for eicosanoid production obtained between monomicrobial and polymicrobial biofilms in the presence of ASA and ATM were observed suggest the complex interaction of C. albicans and P. aeruginosa in terms of eicosanoid production. In addition, the inhibitors caused dramatic alterations in polymicrobial biofilm morphology. Interestingly, although these inhibitors did not affect C. albicans metabolic activity or biofilm biomass, ASA caused a significant increase in P. aeruginosa metabolic activity. In addition, P. aeruginosa metabolic activity was significantly inhibited by NDGA. The possible clinical relevance of these findings warrant further investigation, as the use of inhibitors, such as the ones used in the present study, could possibly affect virulence and the ability of hosts to clear infection. This study also evaluated the role of a secretable 15-lipoxygenase produced by P. aeruginosa capable of converting AA to 15-HETE, although further research and methodology is needed to elucidate its role.This study is the first to investigate the production of eicosanoids by polymicrobial biofilms of C. albicans and P. aeruginosa. The increased production of these eicosanoids compared to monomicrobial counterparts, suggest that the microbially produced eicosanoids may possibly play a role in pathogen-pathogen interaction, as well as host-pathogen interaction. This may ultimately affect the ability of the host to clear infection. In addition, with the use of inhibitors, the possible involvement of various enzymatic groups can be speculated during polymicrobial eicosanoid production. Further research into this interaction may provide valuable insight into polymicrobial eicosanoid production and may contribute to possible therapeutic intervention strategies during monomicrobial and polymicrobial infection.

[发布机构] University of the Free State