Prospecção de micro-organismos e genes desenvolvidos com produção de biosurfactantes em solo de manguezal

Abstract

Biosiirfactants and bioemulsifíers are amphiphilic compounds which have antimicrobial properties and the ability of reducing surface tension, and stabilizing emulsions. These molecules have attracted major attention worldwide due to their great efficiency over chemical surfactants in the bioremediation of environments contaminated by oil and derivatives. The main objective of this study was to select bacteria able to produce biosurfàctants among 100 strains isolated from soil samples collected in a northeast Brazil mangrove. For bacteria isolation, soils were submitted to thermal treatment at 80 °C for 1 h, aüning to select only spore producing species. After that, the soils were cultivated in ATGE medium at 30 °C for 24h for spore germination and growth. This methodology intended to capture species of the genus Bacillus, which are known biosurfactant producers. Among the obtained isolates the selection of biosiirfactant producing strains was carried out through cultivation in Mineral broth at 30 °C for 48h, followed by supernatant analyses for superficial tension reduction and emulsification, to evaluate, respectively, production of biosurfactants and bioemulsifiers, and also antifüngal activity. Simultaneously, the isolates were screened for genes related with those activities by PCR using primers for surfactin and iturin. The results were submitted to cluster analysis in order to elect the most efficient strains. Five strains were selected as good biosurfactant producers: Jag248; Jag252; Jag263; Jag263.1 and Jagl732; while six strains were considered good bioemulsifiers: Jagl85; Jagl91; Jagl98; Jag21; Jag242; and Jag310. The strains Jag245, Jag249, Jag251 and Jag255 showed both properties. Interestingly, the supernatant of Jag248 presented demulsifying activity. The sjp gene, related to surfactin production, was detected only in Jag 15B, Jagl85 and Jag249. The isolates, Jagl85,Jag245,Jag248,Jag249,Jag255 and Jagl732 were tested for oil dispersion, and Jag248, Jag249 and Jag255 have showed the best results, with halos greater than 50% of the plate diameter. Among these strains, Jag248 was chosen for further assays, due to its great potential for applications in bioremediation. Jag248 was identified as Bacillus sp., and showed great effectiveness in soil and seawater oil clean up, being even better than the synthetic surfactants used for comparison. Jag248 tensoactive-molecule is probably an iturin, as demonstrated by its antifungal activity and also by the amplification of ituD and lpal4 by PCR. This molecule was partially purified by HPLC and presented two low values for the formation of micelles, being most effective than commercial surfactants. This is the first study relating iturine to a relevant activity of surface tension reduction. Our results highlight that mangrove soils represent a rich source of known and new surface active molecules.