Polyhydroxybutirate increases survival after UVA exposure in Pseudomonas spp.

TRIBELLI PAULA; COSTA CRISTINA S.; BRITO MARIA GABRIELA; PEZZONI MM; LOPEZ NANCY I.

Congreso; XII Congreso Argentino de Microbiologia; 2017

Solar ultraviolet radiation (UVA; 315-400 nm) the major fraction of UVA radiation reaching the Earth?s surface represents one of the main environmental stress factors for bacteria. Lethal effects of UVA are due to oxidative damage of macromolecules by the action of reactive oxygen species (ROS). Antioxidative defenses and other strategies are a key factor for survival after UVA exposure. Pseudomonas species are ubiquitous microorganism with a wide variety of ecological niches and life styles. In this work we analyzed the survival after UVA exposure in P. extremaustralis, P. aeruginosa PAO1 and P. protegens Pf-5 in relation to polyhydroxyalkanoates (PHA) content. Most of Pseudomonas spp. produces medium chain length PHA while P. extremaustralis produces mainly polyhydroxybutyrate (PHB). Cultures were grown in LB medium with or without sodium octanoate as carbon source for PHA accumulation at 30°C and 200 rpm for 24 h. Cells were washed and adjusted to OD650nm 0.4 in saline solution. Each bacterial suspension was divided in two fractions. One of the fractions was irradiated from above at a fluence rate of 20 W m-2 at the level of the free surface of the suspension, while the other fraction was covered with a black plastic sheet (dark control). Aliquots were taken through time and plate counts were performed. In absence of PHA, P. aeruginosa PAO1 showed a higher UVA resistance than P. extremaustralis and P. protegens Pf-5. However, in LB cultures supplemented with sodium octanoate P. extremaustralis presented the highest survival after UVA exposure. Moreover, P. aeruginosa PAO1 carrying a PHB synthase gene showed an increase in survival in cultures supplemented with sodium octanoate, while a mutant strain of P. extremaustralis impaired of PHB accumulation presented similar survival notwithstanding the presence of sodium octanoate. To deeply study this phenotype PHB content was measured before and after UVA exposure and in dark control experiments. After 300 min of irradiation the PHB content drop in line with increased survival while in the control without irradiation the PHB content was similar to those observed at the initial time. Additionally, redox ratio was higher in cultures exposed to UVA, probably as consequence of PHB depolymerization. Our results showed that PHB accumulation increases the resistance to UVA in Pseudomonas species probably due to its depolymerization and the consequent NADH/NADPH availability, a key factor for stress resistance.