Isolation of four hydrocarbon effluent-degrading Bacillaceae species and evaluation of their ability to grow under high-temperature or high-salinity conditions

Four bacterial strains belonging to the family Bacillaceae were isolated from a polluted site and tested for their efficiency in degrading a refinery effluent highly polluted with hydrocarbons. Among 13 strains isolated, four were selected for their efficiency. Each of these four strains demonstrated a strong ability to grow as a single strain on a hydrocarbon effluent (HCE) as sole carbon source. In batch assays using clarified wastewater as diluent, the strains achieved high-percentage decreases in total hydrocarbon concentration within 18 days: 98% with Bacillus licheniformis STK08, 87% with Geobacillus stearothermophilus STM04, 80% with Lysinibacillus sphaericus STZ75 and 72% with Bacillus firmus STS84. The decreases were greater during the first three days of treatment, with 73, 66, 39 and 47% recorded for Bacillus licheniformis STK08, G. stearothermophilus STM04, L. sphaericus STZ75 and B. firmus STS84, respectively. Growth assays run under different conditions showed that B. licheniformis STK08 and G. stearothermophilus STM04 were able to grow at salinities of up to 120 g/L and at 55°C. Potential biosurfactant production tested using two methods namely modified drop collapse (MDC) and blue agar plate (BAP) demonstrated that the four Bacillaceae species are biosurfactant producers.Keywords: Hydrocarbons, biodegradation, pure culture, high salinity, high temperatureAfrican Journal of Biotechnology Vol. 12(14), pp. 1636-164

Bioconversion of ferulic acid to vanillic acid by Halomonas elongata isolated from table-olive fermentation

Halomonas elongata strain Mar (=CCUG 52759) isolated from table-olive fermentation is the first halophilic bacterium to be shown to transform ferulic acid to vanillic acid under hypersaline conditions. During growth on ferulic acid, this strain was capable of promoting the formation of a significant amount of vanillic acid and trace quantities of vanillin. The products were confirmed by high-performance liquid chromatography and gas chromatography-mass spectrometry analyses. Based on the different metabolites identified, an oxidative side chain degradation pathway of ferulic acid bioconversion to vanillic acid was suggested. Phylogenetic analysis of 16S rRNA gene revealed that this isolated strain Mar was identified as H. elongata. To increase the formation of vanillic acid, a resting cell method using H. elongata strain Mar was performed. The optimal yield of vanillic acid (86%) was obtained after a 6 h reaction using 5 mM of ferulic acid and 4 g of dry weight of cells L-1 pregrown on ferulic acid and harvested at the end of the exponential phase

Optimized conditions for the synthesis of vanillic acid under hypersaline conditions by Halomonas elongata DSM 2581(T) resting cells

During growth on ferulic acid, Halomonas elongata DSM 2581(T) was capable of promoting the formation of a significant amount of vanillic acid. The products were confirmed by high-performance liquid chromatography and gas chromatography mass-spectrometry analyses. To enhance the formation of vanillic acid and prevent its degradation, a resting-cell method using Halomonas elongata was developed. The growth state of the culture utilized for biomass production, the concentration of the biomass, the amount of ferulic acid that was treated and the reutilization of the biomass were optimized. The optimal yield of vanillic acid (82%) was obtained after a 10-h reaction using 10 mM ferulic acid and 5 g/l of cell pregrown on ferulic acid and harvested at the end of the exponential phase

Characterization of Desulfovibrio salinus sp. nov., a slightly halophilic sulfate-reducing bacterium isolated from a saline lake in Tunisia

International audienceA novel slightly halophilic sulfate-reducing bacterium, designated strain P1BSR T , was isolated from water of a saline lake in Tunisia. Strain P1BSR T had motile (single polar flagellum), Gram-negative, rod-shaped, non-spore-forming cells, occurring singly or in pairs. Strain P1BSR T grew at temperatures between 15 and 45 C (optimum 40 C), and in a pH range between 6 and 8.5 (optimum pH 6.7). The strain required NaCl for growth (1 % w/v), and tolerated high NaCl concentration (up to 12 % w/v) with an optimum of 3 % (w/v). Sulfate, thiosulfate and sulfite served as terminal electron acceptors, but not elemental sulfur, fumarate, nitrate and nitrite. Strain P1BSR T utilized lactate, pyruvate, formate, D-fructose and glycerol as carbon and energy sources. The main cellular fatty acid was C 16 : 0 (50.8 %). The genomic DNA G+C content was 47.7 mol%. Phylogenetic analysis of 16S rRNA gene sequence similarity indicated that strain P1BSR T was affiliated to the genus Desulfovibrio, with the type strains Desulfovibrio salexigens (96.51 %), Desulfovibrio zosterae (95.68 %), Desulfovibrio hydrothermalis (94.81 %) and Desulfovibrio ferrireducens (94.73 %) as its closest phylogenetic relatives. On the basis of genotypic, phenotypic and phylogenetic characteristics, it is proposed to assign strain P1BSR T to a novel species of the genus Desulfovibrio, Desulfovibrio salinus sp. nov. The type strain is P1BSR T (=DSM 101510 T =JCM 31065 T). Sulfate-reducing prokaryotes (SRPs) are mainly chemohe-terotrophs, both Bacteria and Archaea (250 species of 65 genera), that can use sulfate as a terminal electron acceptor in their energy metabolism [1, 2]. Owing to their broad metabolic capacities, sulfate-reducing bacteria (SRB) are important in the mineralization of organic matter in anoxic marine sediments. Dissimilatory sulfate reduction has been observed in various hypersaline environments such as salt-erns, the Dead Sea and the Great Salt Lake [3]. However, most of the halophilic SRPs isolated so far are marine or slightly halophilic microorganisms [with optimum salinity ranging from 1 to 4 % (w/v) NaCl] belonging to several gen-era of SRPs, including Desulfovibrio, Desulfonatronovibrio and Desulfonatronobacter species [3-6]. The first SRB species belonging to the genus Desulfovibrio isolated from a hypersaline environment was Desulfovibrio salexigens, which did not grow at NaCl concentrations higher than 12 % (w/v) [7]. Cord-Ruwish [8] then isolated several strains of SRBs from hypersaline oilfield water containing about 10 % (w/v) NaCl. One isolate grew slowly up to 27 % (w/v) NaCl but has not been described in more detail since. One year later, Trüper and Galinski [9] isolated a few SRB strains from hot brines in the Red Sea that were similar to Desulfovibrio halophilus, a moderately halophilic sulfate reducer isolated by Caumette et al. [10] from the hypersa-line Solar Lake in Sinai. The latter isolate grew in salinity ranging from 3 to 18 % (w/v), and optimally at 6-7 % (w/v) NaCl. However, since 1991, no novel SRB species belonging to the genus Desulfovibrio has been characterized that grows in salt concentrations above 10 % NaCl (w/v). No pure culture of extreme halophilic SRB (i.e. growing under saturating salt conditions) has been isolated until now. In this study, we report the isolation and characterization of a mesophilic, slightly halophilic SRB isolated from water samples of a Tunisian saline lake that is able to grow in up to 12 % NaCl (w/v), but grows optimally at 3 % (w/v). This SRB isolate is proposed to represent a novel species of the genus Desulfovibrio. Water samples were collected from a continental saline lake located in the middle-east of Tunisia and transported to the laboratory at ambient temperature. Bacteria wer

Thermanaeromonas burensis sp nov., a thermophilic anaerobe isolated from a subterranean clay environment

A strictly anaerobic, thermophilic and halotolerant strain, designated IA106(T), was isolated from the seepage water collected in a metal biocorrosion test at a depth of 490 m, in a 130-160 m thick, subterranean Callovo-Oxfordian clay formation (158-152 million years old) in northern France. This geological formation has been selected as the potential host rock for the French high-level nuclear waste repository. Cells of strain IA106(T) stained Gram-positive and were non-motile, spore-forming, straight rods (0.5 x 2-6 mu m). The five major fatty acids were C-16:0 (15.9 %), C-18:0 (15.4 %), iso-C-17:1 I and/or anteiso-C-17:1 B(14.8 %), iso-C-17:0 (14.7 %) and iso-C-15:0 (13.0 %). Growth was observed at temperatures ranging from 55 to 70 degrees C and at pH 5.5-9. The salinity range for growth was 0-20 g NaCl 1(-1). Yeast extract was required for growth. Strain IA106(T) was able to grow on lactate and various sugars in the presence of thiosulfate as electron acceptor. Sulfate, sulfite, elemental sulfur, fumarate, nitrate and nitrite were not reduced. The DNA G+C content was 60.2 mol%. 16S rRNA gene sequence analysis indicated that strain IA106(T) belonged to the family Thermoanaerobacteraceae, class Clostridia, phylum Firmicutes, and was most closely related to Thermanaeromonas toyohensis DSM 14490(T) (95.16 % 16S rRNA gene sequence similarity). On the basis of 16S rRNA gene sequence comparisons and physiological characteristics, strain IA106(T) represents a novel species of the genus Thermanaeromonas, for which the name Thermanaeromonas burensis sp. nov. is proposed. The type strain is IA106(T) (=DSM 26576(T)=JCM 18718(T)

Mobilisporobacter senegalensis gen. nov., sp nov., an anaerobic bacterium isolated from tropical shea cake

International audienceA Gram-stain positive, endospore-forming, strictly anaerobic bacterium, designated strain Gal1(T), was isolated from shea cake, a waste material from the production of shea butter, originating from Saraya, Senegal. The cells were rod-shaped, slightly curved, and motile with peritrichous flagella. The strain was oxidase-negative and catalase-negative. Growth was observed at temperatures ranging from 15 to 45 degrees C (optimum 30 degrees C) and at pH 6.5-9.3 (optimum pH 7.8). The salinity range for growth was 0-3.5 % NaCl (optimum 1 %). Yeast extract was required for growth. Strain Gal1(T) fermented various carbohydrates such as mannose, mannitol, arabinose, cellobiose, fructose, glucose, maltose, sucrose, trehalose and lactose and the major end-products were ethanol and acetate. The only major cellular fatty acid was C16 : 0 (19.6 %). The DNA base G+ C content of strain Gal1(T) was 33.8 mol%. Analysis of the 16S rRNA gene sequence of the isolate indicated that this strain was related to Mobilitalea sibirica DSM 26468(T) with 94.27 % similarity, Clostridium populeti ATTC 35295(T) with 93.94 % similarity, and Clostridium aminovalericum DSM 1283(T) and Anaerosporobacter mobilis DSM 15930(T) with 93.63 % similarity. On the basis of phenotypic characteristics, phylogenetic analysis and the results of biochemical and physiological tests, strain Gal1(T) was clearly distinguished from closely related genera, and strain Gal1(T) can be assigned to a novel species of a new genus for which the name Mobilisporobacter senegalensis gen. nov., sp. nov. is proposed. The type strain is Gal1(T) (5DSM 26537(T) 5JCM 18753(T))

Anaerosalibacter bizertensis gen. nov., sp. nov., a new halotolerant bacterium isolated from sludge.

International audienceA strictly anaerobic, halotolerant and thermotolerant strain designated C5BELT was isolated from storage tanks holding wastes generated by the recycling of discarded motor oils located in north Tunisia. Cells of strain C5BELT stained Gram-positive, were motile by laterally inserted flagella, straight, and spore-forming. Their two major fatty acids were iso-C15:0 (60.1 %) and iso-C15:0 dimethyl acetal (22.0 %). Growth was observed at temperatures ranging from 25 to 55 °C (optimum 40 °C) and at pH 6 to 9 (optimum 7.5). The salinity range for growth was 0-100 g l-1 of NaCl (optimum 5 g l-1). Yeast extract is required for growth. Strain C5BELT was heterotrophic, able to use glucose, pyruvate, succinate, yeast extract, bio-trypcase and peptone, but was unable to grow on casamino acids. Sulfate, thiosulfate, sulfite, elemental sulfur, fumarate, nitrate and nitrite were not reduced. The DNA G + C content was 31.1 mol%. 16S rRNA gene sequence analysis indicated that strain C5BELT was a member of the family Clostridiaceae, class Clostridia, phylum Firmicutes and was most closely related to Sporanaerobacter acetigenes DSM 13106T (92.4 % similarity). On the basis of 16S rRNA gene sequence comparisons and physiological characteristics, strain C5BELT can be classified as a new species of a new genus, for which the name Anaerosalibacter bizertensis gen. nov., sp. nov. is hereby proposed. The type strain is C5BELT (= DSM 23801 = JCM 17239)

Desulfovibrio senegalensis sp. nov., a mesophilic sulfate reducer isolated from marine sediment

International audienc

Thermospira aquatica gen. nov., sp. nov., a novel thermophilic spirochete isolated from a Tunisian hot spring, and description of the novel family Thermospiraceae

International audienceA novel thermophilic, anaerobic bacterium, strain F1F22 T , was isolated from hot spring water collected in northern Tunisia. The cells were non-motile, Gram-negative and helical with hooked ends, 0.5×10–32 µm in size. Growth of the strain was observed at 45–70 °C (optimum, 55 °C), in 0.0–1.0 % (w/v) NaCl (optimum without NaCl) and at pH 6.5–8.5 (optimum, pH 7.5). Yeast extract was required for growth, and the strain grew on glucose, sucrose and maltose. The major fatty acids were C 16:0 (40.2 %), iso-C 16: 0 (30.2 %) and C 16 :0 DMA (14.5 %). The genome consisted of a circular chromosome (2.5 Mb) containing 2672 predicted protein-encoding genes with a G+C content of 43.15 mol %. Based on a comparative 16S rRNA gene sequence analysis, strain F1F22 T formed a deeply branching lineage within the phylum Spirochaetota , class Spirochaetia , order Brevinematales , and had only low sequence similarity to other species of the phylum (lower than 83 %). Genome-based analysis of average nucleotide identity and digital DNA–DNA hybridization of strain F1F22 T with Treponema caldarium DSM 7334 T , Brevinema andersonii ATCC 43811 T and Spirochaeta thermophila DSM 6578 T showed values between 63.26 and 63.52 %, and between 20 and 25 %. Hence, we propose strain F1F22 T as a representative of a novel family ( Thermospiraceae fam. nov.), genus and species of Brevinematales : Thermospira aquatica gen. nov., sp. nov. (type strain F1F22 T =JCM 31314 T =DSM 101182 T )