Differential utilization of pyrene as the sole source of carbon by Bacillus subtilis and Pseudomonas aeruginosa strains: role of biosurfactants in enhancing bioavailability

AIMS: Our goal is to compare the efficiency of utilization of pyrene as the sole source of carbon for growth and energy by two nonactinomycetous groups of bacteria viz., Bacillus subtilis DM-04 and Pseudomonas aeruginosa mucoid (M) and nonmucoid (NM) strains, isolated from a petroleum-contaminated soil sample of north-east India. METHODS AND RESULTS: Bacillus subtilis DM-04 and P. aeruginosa M and NM bacterial strains were capable of secreting biosurfactant in the culture medium while growing on pyrene and their pyrene utilizing efficiency was demonstrated by correlating the bacterial growth in the presence of pyrene as the sole source of carbon along with a concomitant decrease in pyrene content from the culture medium with respect to time. The biosurfactant secreted by the respective bacterial strains enhanced the apparent solubility of pyrene by factors of 5-7 and influenced the bacterial cell surface hydrophobicity resulting in higher uptake and utilization of pyrene by bacteria. The growth of B. subtilis DM-04 and P. aeruginosa M and NM strains at the expense of pyrene after 96 h showed an assimilation of about 48.0 +/- 1.1% (mean +/- SD) and 32.0 +/- 0.6% (mean +/- SD) of pyrene carbon, respectively, showing differences in metabolism of pyrene by these bacterial strains. CONCLUSIONS: Bacillus subtilis DM-04 strain exhibited higher utilization and cellular assimilation of pyrene compared with P. aeruginosa M and NM strains. Further, the biosurfactants produced by the bacteria under study are capable of enhancing the solubility of pyrene in aqueous media and can influence the cell surface hydrophobicity of the biosurfactant-producing strains that results in a higher uptake of pyrene. SIGNIFICANCE AND IMPACT OF THE STUDY: It may be suggested that the bacteria used in this study are suitable candidates for practical field application for effective in situ bioremediation of pyrene-contaminated sites.     

Naphthalene degradation and biosurfactant activity by Bacillus cereus 28BN

Biosurfactant activity and naphthalene degradation by a new strain identified as Bacillus cereus 28BN were studied. The strain grew well and produced effective biosurfactants in the presence of n-alkanes, naphthalene, crude oil and vegetable oils. The biosurfactants were detected by the surface tension lowering of the medium, thin layer chromatography and infrared spectra analysis. With (2%) naphthalene as the sole carbon source, high levels of rhamnolipids at a concentration of 2.3 g 1(-1) were determined in the stationary growth. After 20 d of incubation 72 +/- 4% of the initial naphthalene was degraded. This is the first report for a Bacillus cereus rhamnolipid producing strain that utilized naphthalene under aerobic conditions. The strain looks promising for application in environmental technologies.     

Biosurfactant production using molasses and whey under thermophilic conditions

Biosurfactant production was studied by Bacillus licheniformis K51, B. subtilis 20B, B. subtilis R1 and Bacillus strain HS3 using molasses or cheese whey as a sole source of nutrition at 45 degrees C. The isolates were able to grow and produce biosurfactant under shaking as well as static conditions. Maximum biosurfactant production was achieved with molasses at 5.0-7.0% (w/v). The biosurfactant retained its surface-active properties after incubation at 80 degrees C at a wide range of pH values and salt concentrations for nine days. Oil displacement experiments in sand pack columns with crude oil showed 25-33% recovery of residual oil.     

Importance of 3-hydroxy fatty acid composition of lipopeptides for biosurfactant activity

Biosurfactant production may be an economic approach to improving oil recovery. To obtain candidates most suitable for oil recovery, 207 strains, mostly belonging to the genus Bacillus, were tested for growth and biosurfactant production in medium with 5% NaCl under aerobic and anaerobic conditions. All strains grew aerobically with 5% NaCl, and 147 strains produced a biosurfactant. Thirty-five strains grew anaerobically with 5% NaCl, and two produced a biosurfactant. In order to relate structural differences to activity, eight lipopeptide biosurfactants with different specific activities produced by various Bacillus species were purified by a new protocol. The amino acid compositions of the eight lipopeptides were the same (Glu/Gln:Asp/Asn:Val:Leu, 1:1:1:4), but the fatty acid compositions differed. Multiple regression analysis showed that the specific biosurfactant activity depended on the ratios of both iso to normal even-numbered fatty acids and anteiso to iso odd-numbered fatty acids. A multiple regression model accurately predicted the specific biosurfactant activities of four newly purified biosurfactants (r2= 0.91). The fatty acid composition of the biosurfactant produced by Bacillus subtilis subsp. subtilis strain T89-42 was altered by the addition of branched-chain amino acids to the growth medium. The specific activities of biosurfactants produced in cultures with different amino acid additions were accurately predicted by the multiple regression model derived from the fatty acid compositions (r2= 0.95). Our work shows that many strains of Bacillus mojavensis and Bacillus subtilis produce biosurfactants and that the fatty acid composition is important for biosurfactant activity.     

In situ biosurfactant production by Bacillus strains injected into a limestone petroleum reservoir

Biosurfactant-mediated oil recovery may be an economic approach for recovery of significant amounts of oil entrapped in reservoirs, but evidence that biosurfactants can be produced in situ at concentrations needed to mobilize oil is lacking. We tested whether two Bacillus strains that produce lipopeptide biosurfactants can metabolize and produce their biosurfactants in an oil reservoir. Five wells that produce from the same Viola limestone formation were used. Two wells received an inoculum (a mixture of Bacillus strain RS-1 and Bacillus subtilis subsp. spizizenii NRRL B-23049) and nutrients (glucose, sodium nitrate, and trace metals), two wells received just nutrients, and one well received only formation water. Results showed in situ metabolism and biosurfactant production. The average concentration of lipopeptide biosurfactant in the produced fluids of the inoculated wells was about 90 mg/liter. This concentration is approximately nine times the minimum concentration required to mobilize entrapped oil from sandstone cores. Carbon dioxide, acetate, lactate, ethanol, and 2,3-butanediol were detected in the produced fluids of the inoculated wells. Only CO(2) and ethanol were detected in the produced fluids of the nutrient-only-treated wells. Microbiological and molecular data showed that the microorganisms injected into the formation were retrieved in the produced fluids of the inoculated wells. We provide essential data for modeling microbial oil recovery processes in situ, including growth rates (0.06 +/- 0.01 h(-1)), carbon balances (107% +/- 34%), biosurfactant production rates (0.02 +/- 0.001 h(-1)), and biosurfactant yields (0.015 +/- 0.001 mol biosurfactant/mol glucose). The data demonstrate the technical feasibility of microbial processes for oil recovery.     

Production of biosurfactant and antifungal compound by fermented food isolate Bacillus subtilis 20B

A biosurfactant producing strain, Bacillus subtilis 20B, was isolated from fermented food in India. The strain also showed inhibition of various fungi in in-vitro experiments on Potato Dextrose Agar medium. It was capable of growth at temperature 55 degrees C and salts up to 7%. It utilized different sugars, alcohols, hydrocarbons and oil as a carbon source, with preference for sugars. In glucose based minimal medium it produced biosurfactant which reduced surface tension to 29.5 mN/m, interfacial tension to 4.5 mN/m and gave stable emulsion with crude oil and n-hexadecane. The biosurfactant activity was stable at high temperature, a wide range of pH and salt concentrations for five days. Oil displacement experiments using biosurfactant containing broth in sand pack columns with crude oil showed 30.22% recovery. The possible application of organism as biocontrol agent and use of biosurfactant in microbial enhanced oil recovery (MEOR) is discussed.     

Biodegradation of phenanthrene by<Emphasis Type="Italic"> Pseudomonas</Emphasis> sp. strain PP2: novel metabolic pathway,role of biosurfactant and cell surface hydrophobicity in hydrocarbon assimilation

Pseudomonas sp. strain PP2 isolated in our laboratory efficiently metabolizes phenanthrene at 0.3% concentration as the sole source of carbon and energy. The metabolic pathways for the degradation of phenanthrene, benzoate and p-hydroxybenzoate were elucidated by identifying metabolites, biotransformation studies, oxygen uptake by whole cells on probable metabolic intermediates, and monitoring enzyme activities in cell-free extracts. The results obtained suggest that phenanthrene degradation is initiated by double hydroxylation resulting in the formation of 3,4-dihydroxyphenanthrene. The diol was finally oxidized to 2-hydroxymuconic semialdehyde. Detection of 1-hydroxy-2-naphthoic acid, alpha-naphthol, 1,2-dihydroxy naphthalene, and salicylate in the spent medium by thin layer chromatography; the presence of 1,2-dihydroxynaphthalene dioxygenase, salicylaldehyde dehydrogenase and catechol-2,3-dioxygenase activity in the extract; O(2) uptake by cells on alpha-naphthol, 1,2-dihydroxynaphthalene, salicylaldehyde, salicylate and catechol; and no O(2) uptake on o-phthalate and 3,4-dihydroxybenzoate supports the novel route of metabolism of phenanthrene via 1-hydroxy-2-naphthoic acid --> [alpha-naphthol] --> 1,2-dihydroxy naphthalene --> salicylate --> catechol. The strain degrades benzoate via catechol and cis,cis-muconic acid, and p-hydroxybenzoate via 3,4-dihydroxybenzoate and 3-carboxy- cis,cis-muconic acid. Interestingly, the culture failed to grow on naphthalene. When grown on either hydrocarbon or dextrose, the culture showed good extracellular biosurfactant production. Growth-dependent changes in the cell surface hydrophobicity, and emulsification activity experiments suggest that: (1) production of biosurfactant was constitutive and growth-associated, (2) production was higher when cells were grown on phenanthrene as compared to dextrose and benzoate, (3) hydrocarbon-grown cells were more hydrophobic and showed higher affinity towards both aromatic and aliphatic hydrocarbons compared to dextrose-grown cells, and (4) mid-log-phase cells were significantly (2-fold) more hydrophobic than stationary phase cells. Based on these results, we hypothesize that growth-associated extracellular biosurfactant production and modulation of cell surface hydrophobicity plays an important role in hydrocarbon assimilation/uptake in Pseudomonas sp. strain PP2.     

Effect of a Pseudomonas rhamnolipid biosurfactant on cell hydrophobicity and biodegradation of octadecane.

In this study, the effect of a purified rhamnolipid biosurfactant on the hydrophobicity of octadecane-degrading cells was investigated to determine whether differences in rates of octadecane biodegradation resulting from the addition of rhamnolipid to four strains of Pseudomonas aeruginosa could be related to measured differences in hydrophobicity. Cell hydrophobicity was determined by a modified bacterial adherence to hydrocarbon (BATH) assay. Bacterial adherence to hydrocarbon quantitates the preference of cell surfaces for the aqueous phase or the aqueous-hexadecane interface in a two-phase system of water and hexadecane. On the basis of octadecane biodegradation in the absence of rhamnolipid, the four bacterial strains were divided into two groups: the fast degraders (ATCC 15442 and ATCC 27853), which had high cell hydrophobicities (74 and 55% adherence to hexadecane, respectively), and the slow degraders (ATCC 9027 and NRRL 3198), which had low cell hydrophobicities (27 and 40%, respectively). Although in all cases rhamnolipid increased the aqueous dispersion of octadecane at least 10(4)-fold, at low rhamnolipid concentrations (0.6 mM), biodegradation by all four strains was initially inhibited for at least 100 h relative to controls. At high rhamnolipid concentrations (6 mM), biodegradation by the fast degraders was slightly inhibited relative to controls, but the biodegradation by the slow degraders was enhanced relative to controls. Measurement of cell hydrophobicity showed that rhamnolipids increased the cell hydrophobicity of the slow degraders but had no effect on the cell hydrophobicity of the fast degraders. The rate at which the cells became hydrophobic was found to depend on the rhamnolipid concentration and was directly related to the rate of octadecane biodegradation.(ABSTRACT TRUNCATED AT 250 WORDS)     

Adsorption of dirhamnolipid on four microorganisms and the effect on cell surface hydrophobicity

In this study, adsorption of dirhamnolipid biosurfactant on a Gram-negative Pseudomonas aeruginosa, two Gram-positive Bacillus subtilis, and a yeast, Candida lipolytica, was investigated, and the causality between the adsorption and change of cell surface hydrophobicity was discussed. The adsorption was not only specific to the microorganisms but also depended on the physiological status of the cells. Components of the biosurfactant with different rhamnosyl number or aliphatic chain length also exhibited slight difference in adsorption manner. The adsorption indeed caused the cell surface hydrophobicity to change regularly; however, the changes depended on both the concentrations of rhamnolipid solutions applied and the adsorbent physiological conditions. Orientation of rhamnolipid monomers on cell surface and micelle deposition are supposed to be the basic means of adsorption to change cell hydrophobicity at low and high rhamnolipid concentrations, respectively. This study proposed the possibility to modify cell surface hydrophobicity with biosurfactant of low concentrations, which may be of importance in in situ soil remediation. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Production and characterization of a glycolipid biosurfactant from Bacillus megaterium using economically cheaper sources

Criteria selected for screening of biosurfactant production by Bacillus megaterium were hemolytic assay, bacterial cell hydrophobicity and the drop-collapse test. The data on hemolytic activity, bacterial cell adherence with crude oil and the drop-collapse test confirmed the biosurfactant-producing ability of the strain. Accordingly, the strain was cultured at different temperatures, pH values, salinity and substrate (crude oil) concentration in mineral salt medium to establish the optimum culture conditions, and it was shown that 38°C, 2.0% of substrate concentration, pH 8.0 and 30‰ of salt concentration were optimal for maximum growth and biosurfactant production. Laboratory scale biosurfactant production in a fermentor was done with crude oil and cheaper carbon sources like waste motor lubricant oil and peanut oil cake, and the highest biosurfactant production was found with peanut oil cake. Characterization of partially purified biosurfactant inferred that it was a glycolipid with emulsification potential of waste motor lubricant oil, crude oil, peanut oil, diesel, kerosene, naphthalene, anthracene and xylene.     

Rhamnolipid production by Pseudomonas aeruginosa engineered with the Vitreoscilla hemoglobin gene

The potential of Pseudomonas aeruginosa expressing the Vitreoscilla hemoglobin gene (vgb) for rhamnolipid production was studied. P. aeruginosa (NRRL B-771) and its transposon mediated vgb transferred recombinant strain, PaJC, were used in the research. The optimization of rhamnolipid production was carried out in the different conditions of cultivation (agitation rate, the composition of culture medium and temperature) in a time-course manner. The nutrient source, especially the carbon type, had a dramatic effect on rhamnolipid production. The PaJC strain and the wild type cells of P. aeruginosa started producing biosurfactant at the stationary phase and its concentration reached maximum at 24 h (838 mg/l(-1)) and at 72 h (751 mg l(-1)) of the incubation respectively. Rhamnolipid production was optimal in batch cultures when the temperature and agitation rate were controlled at 30 degrees C and 100 rpm. It reached 8373 mg l(-1) when the PaJC cells were grown in 1.0% glucose supplemented minimal media. Genetic engineering of biosurfactant producing strains with vgb may be an effective method to increase its production.     

The Effect of Rhamnolipid Biosurfactant Produced by <Emphasis Type="Italic">Pseudomonas fluorescens</Emphasis> on Model Bacterial Strains and Isolates from Industrial Wastewater

In this study, the effect of rhamnolipid biosurfactant produced by Pseudomonas fluorescens on bacterial strains, laboratory strains, and isolates from industrial wastewater was investigated. It was shown that biosurfactant, depending on the concentration, has a neutral or detrimental effect on the growth and protein release of model Gram (+) strain Bacillus subtilis 168. The growth and protein release of model Gram (−) strain Pseudomonas aeruginosa 1390 was not influenced by the presence of biosurfactant in the medium. Rhamnolipid biosurfactant at the used concentrations supported the growth of some slow growing on hexadecane bacterial isolates, members of the microbial community. Changes in cell surface hydrophobicity and permeability of some Gram (+) and Gram (−) isolates in the presence of rhamnolipid biosurfactant were followed in experiments in vitro. It was found that bacterial cells treated with biosurfactant became more or less hydrophobic than untreated cells depending on individual characteristics and abilities of the strains. For all treated strains, an increase in the amount of released protein was observed with increasing the amount of biosurfactant, probably due to increased cell permeability as a result of changes in the organization of cell surface structures. The results obtained could contribute to clarify the relationships between members of the microbial community as well as suggest the efficiency of surface properties of rhamnolipid biosurfactant from Pseudomonas fluorescens making it potentially applicable in bioremediation of hydrocarbon-polluted environments.     

Surfactant activity of a naphthalene degrading Bacillus pumilus strain isolated from oil sludge

We studied the growth, biosurfactant activities and petroleum hydrocarbon compounds utilisation of strain 28-11 isolated from a solid waste oil. The isolate was identified as Bacillus pumilus. It grew well in the presence of 0.1% (w/v) of crude oil and naphthalene under aerobic conditions and utilised these substances as carbon and energy source. The capacity of strain 28-11 to emulsify crude oil and its ability to remove hydrocarbons looks promising for its application in environmental technologies.     

一株石油烃降解菌的细胞疏水性及其乳化性质

【目的】从新疆油田石油污染土壤中分离到一株在25 °C条件下利用烃类产生生物表面活性剂的菌株红球菌(Rhodococcus sp.) HL-6, 对其菌体细胞疏水性及所产表面活性剂进行研究。【方法】通过细胞粘附性、表面张力及乳化活性测定对菌株所产表面活性剂进行性质研究。【结果】菌株HL-6在亲水性和疏水性基质中均能产生生物表面活性剂, 在疏水性基质中可以将培养液表面张力由初始的62.487 mN/m降到30.667 mN/m, 培养液在pH 6?9及NaCl浓度1%?5%范围内乳化效果良好, 在4 °C到55 °C范围内乳化效果均为100%, 菌株对柴油的耐受能力很高, 在30%柴油浓度下依然生长良好并且有44%的乳化活性。【结论】HL-6菌株的细胞表面具有很强的疏水性, 这有助于菌体细胞对烃类的摄取。该菌株能够利用烃类基质生产生物表面活性剂, 可以明显降低培养液表面张力并且对石油烃具有良好的乳化作用。说明菌株HL-6能够适应海洋滩涂石油污染的环境, 并可用于严重石油污染区域的生物修复。     

Production of biosurfactant using different hydrocarbons by Pseudomonas aeruginosa EBN-8 mutant

The present investigation dealt with the use of previously isolated and studied gamma-ray mutant strain Pseudomonas aeruginosa EBN-8 for the production of biosurfactant by using different hydrocarbon substrates viz. n-hexadecane, paraffin oil and kerosene oil, provided in minimal medium, as the sole carbon and energy sources. The batch experiments were conducted in 250 mL Erlenmeyer flasks, containing 50 mL minimal salt media supplemented with 1% (w/v) hydrocarbon substrate, inoculated by EBN-8 and incubated at 37 degrees C and 100 rpm in an orbital shaker. The sampling was done on 24 h basis for 10 d. The surface tension of cell-free culture broth decreased from 53 to 29 mN/m after 3 and 4 d of incubation when the carbon sources were paraffin oil and n-hexadecane, respectively. The largest reduction in interfacial tension from 26 to 0.4 mN/m was observed with n-hexadecane, while critical micelle dilution was obtained as 50 x CMC for paraffin oil as carbon source. When grown on n-hexadecane and paraffin oil, the EBN-8 mutant strain gave 4.1 and 6.3 g of the rhamnolipids/L, respectively. These surface-active substances subsequently allowed the hydrocarbon substrates to disperse readily as emulsion in aqueous phase.     

Heterologous production of Pseudomonas aeruginosa EMS1 biosurfactant in Pseudomonas putida

A new bacterial strain isolated from activated sludge, identified as Pseudomonas aeruginosa EMS1, produced a biosurfactant when grown on acidified soybean oil as the sole carbon source. An optimum biosurfactant production of 5 g/L was obtained with the following medium composition: 2% acidified soybean oil, 0.3% NH4NO3, 0.03% KH2PO4, 0.03% K2HPO4, 0.02% MgSO4.7H2O and 0.025% CaCl2.2H2O, with shaking at 200 rpm for an incubation period of 100 h at 30 degrees C. The production of the biosurfactant was found to be a function of cell growth, with maximum production occurring during the exponential phase. Hemolysis of erythrocytes and thin-layer chromatography studies revealed that the secreted biosurfactant was rhamnolipid. To overcome the complex environmental regulation with respect to rhamnolipid biosynthesis, and to replace the opportunistic pathogen P. aeruginosa with a safe industrial strain, attempts were made to achieve rhamnolipid production in a heterologous host, Pseudomonas putida, using molecular cloning of rhlAB rhamnosyltransferase genes with the rhlRI quorum sensing system, assuming that a functional rhamnosyltransferase would catalyze the formation of rhamnosyl-6-hydroxydecanoyl-6-hydroxydecanoate (mono-rhamnolipid) in P. putida. It was shown that rhamnolipid can be produced in the heterologous strain, P. putida, when provided with the rhamnosyltransferase genes.     

Biosurfactant-producing Bacillus are present in produced brines from Oklahoma oil reservoirs with a wide range of salinities

Nine wells producing from six different reservoirs with salinities ranging from 2.1% to 15.9% were surveyed for presence of surface-active compounds and biosurfactant-producing microbes. Degenerate primers were designed to detect the presence of the surfactin/lichenysin (srfA3/licA3) gene involved in lipopeptide biosurfactant production in members of Bacillus subtilis/licheniformis group and the rhlR gene involved in regulation of rhamnolipid production in pseudomonads. Polymerase chain reaction amplification, cloning, and sequencing confirmed the presence of the srfA3/licA3 genes in brines collected from all nine wells. The presence of B. subtilis/licheniformis strains was confirmed by sequencing two other genes commonly used for taxonomic identification of bacteria, gyrA (gyrase A) and the 16S rRNA gene. Neither rhlR nor 16S rRNA gene related to pseudomonads was detected in any of the brines. Intrinsic levels of surface-active compounds in brines were low or not detected, but biosurfactant production could be stimulated by nutrient addition. Supplementation with a known biosurfactant-producing Bacillus strain together with nutrients increased biosurfactant production. The genetic potential to produce lipopeptide biosurfactants (e.g., srfA3/licA3 gene) is prevalent, and nutrient addition stimulated biosurfactant production in brines from diverse reservoirs, suggesting that a biostimulation approach for biosurfactant-mediated oil recovery may be technically feasible.     

Characterization of biosurfactant produced by Pseudoxanthomonas sp. PNK-04 and its application in bioremediation

Biosurfactant producing bacterium was identified as Pseudoxanthomonas sp. PNK-04 based on morphological, physiological, biochemical tests and 16S rRNA gene sequencing. This strain was screened for biosurfactant production using different carbon sources by measuring the surface tension of the medium at different time intervals, and hemolytic activity. The produced biosurfactant was found to be a rhamnolipid based on the formation of dark blue haloes around the colonies in CTAB–methylene blue agar plates and the content of rhamnose sugar. The rhamnolipids produced by this bacterium were found to contain mono- and dirhamnose units linked to β-hydroxy alkonic acids containing 8–12 carbon atoms. This biosurfactant has high emulsifying activity when compared to chemical surfactants such as Tween-80 and Triton X-100 with respect to aliphatic and aromatic hydrocarbons. Further, the biosurfactant stimulates the degradation of 2-chlorobenzoic acid, 3-chlorobenzoic acid and 1-methyl naphthalene by Pseudoxanthomonas sp. PNK-04 probably by aiding in the uptake and increasing the solubility.     

Cell hydrophobicity of <Emphasis Type="Italic">Pseudomonas</Emphasis> spp. and <Emphasis Type="Italic">Bacillus</Emphasis> spp. bacteria and hydrocarbon biodegradation in the presence of Quillaya saponin

Biodegradation and hydrophobicity of Pseudomonas spp. and Bacillus spp. strains were tested at different concentrations of the biosurfactant Quillaya saponin. A model mixture of hydrocarbon (dodecane and hexadecane) was used for estimating the influence of surfactants on biodegradation. The bacterial adhesion to hydrocarbon method for determination of bacterial cell surface hydrophobicity was exploited. Among the tested bacterial strains the higher hydrophobicity was noticed for Pseudomonas aeruginosa TK. The hydrophobicity of this strain was 84%. The highest hydrocarbon biodegradation was observed for P. aeruginosa TK (49%) and Bacillus subtilis (35%) strains after 7 days of experiments. Generally the addition of Quillaya saponin increased hydrocarbon biodegradation remarkably. The optimal concentration proved to be 80 mg l⁻¹. The degree of hydrocarbon biodegradation was 75% for P. aeruginosa TK after the addition of saponin. However the most significant increase in biodegradation after addition of Quillaya saponin was in the case of P. aeruginosa 25 and Pseudomonas putida (the increase of biodegradation from 21 to 52% and from 31 to 66%, respectively). It is worth mentioning that decrease of hydrophobicity is correlated with the best biodegradation by P. aeruginosa strain. For the remaining strains, no significant hydrophobicity changes in relation to the system without surfactant were noticed.     

Crude biosurfactant from thermophilic Alcaligenes faecalis: Feasibility in petro-spill bioremediation

The thermophilic bacterium Alcaligenes faecalis isolated from the crude oil contaminated soil of Upper Assam, India. The isolated bacterium was first screened for the ability to produce biosurfactant. The strain growing at 42 °C could produce higher amount of biosurfactant in medium supplemented with 2% (v/v) diesel as sole source of carbon and energy. Biochemical characterizations including FT-IR and MS studies suggested the biosurfactant to be glycolipid. Tensiometric studies revealed that the biosurfactant produced by the bacterial strain could decrease the surface tension (??) at air-water interface from 71.6 to 32.3 mNm⁻¹ after 96 h of growth on hydrocarbon and possessed a low critical micelle concentration (CMC) value of approximately 38 mgl⁻¹, indicating high surface activity. The culture supernatant containing the biosurfactant was found to be functionally stable at varying pH (2-12), temperature (100 and 121 °C) and salinity (1-6% NaCl, w/v) conditions. Both the culture broth and the cell free supernatant exhibited high emulsifying activity against the different hydrocarbons and the crude oil components. The increase in cell surface hydrophobicity and glycolipid production by the strain suggested the existence of biosurfactant enhanced interfacial uptake of the hydrocarbons. Moreover, the partially purified biosurfactant exhibited antimicrobial activity by inhibiting the growth of several bacterial and fungal species. The strain represented a new class of biosurfactant producers and could be a potential candidate for the production of glycolipid biosurfactant which could be useful in a variety of biotechnological and industrial processes, particularly in the oil industry.