H2O-1 strain (AMS H2O-1) were necessary to evaluate its potential use in the petroleum industry. Therefore, this study presents the taxonomic affiliation of Bacillus sp. H2O-1,

the structure of AMS H2O-1 and its effects on sulfate reducing bacteria cells. Danusertib Furthermore, the surface free energy and the hydrophilic or hydrophobic characteristics of different surfaces conditioned with the antimicrobial substance produced by strain H2O-1 were determined and compared to surfaces treated with a surfactin produced by B. subtilis ATCC 21332. Methods Microorganisms The antimicrobial substance producer strain Bacillus sp. H2O-1 was originally isolated from an oil reservoir in Brazil and previously described by Korenblum et al. [11]. This strain was grown in Luria-Bertani broth (LB), pH 7.0-7.2, containing 10 g of tryptone, 5 g of yeast extract and 5 g of NaCl

per liter of distilled water. The strain Desulfovibrio alaskensis NCIMB 13491 was used as a sulfate reducing bacteria indicator (AMS H2O-1 sensitive) and was grown at 30°C in Postgate E medium [27] purged with a N2 flux to achieve anaerobiosis. Bacillus subtilis ATCC 21332 was used to produce surfactin as described by Nitschke [28]. Taxonomic affiliation The bacterial strain H2O-1 was characterized by using the kit API 50CH (Apparéils et Prócédes d′Identification – bioMérieux sa, Lyon, France) as described by the manufacturer. In addition, the 16S rRNA gene was amplified by PCR from H2O-1 genomic DNA

using the universal primers 27f (5’-AGAGTTTGATCCTGGCTCAG-3’) and 1492r (5’-GGTTACCTTGTTACGACTT-3’). S63845 manufacturer DNA was extracted from Bacillus sp. H2O-1 grown overnight at 30°C in LB broth using the ZR Fungal/Bacterial DNA MiniPrepTM kit (ZYMO Research, Irvine, CA, USA) according to the manufacturer’s instructions. The full 16S rRNA gene sequencing (GenBank accession number AMN-107 JX575798) was carried out by the Macrogen Genomic Division, South Korea, using ABI PRISM Big Dye Terminator Cycle Sequencing technology (Applied BioSystems, Foster city, CA, USA). The sequence obtained was compared also with 16S rRNA gene sequences of closely related type strains using RDP database (http://​rdp.​cme.​msu.​edu/​). Alignment and phylogenetic tree construction were performed using the Tree Builder tool from RDP website. Isolation and purification of the lipopeptide The Bacillus sp. H2O-1 was cultured in LB broth at 30°C for three days and then harvested by centrifugation at 12,500 x g for 30 min. The supernatant was adjusted to pH 2.0 with concentrated HCl and allowed to stand overnight at 4°C. The precipitate was then dissolved in 0.4 M HCl and extracted with chloroform-methanol (2:1 v/v) [29]. The mixture was shaken vigorously and then left static for phase separation. The organic phase was concentrated at reduced pressure at 40°C, yielding the crude extract containing the lipopeptide. The AMS H2O-1 lipopeptide extract was applied to a silica gel 60 column chromatography (particle size 0.