Plant-based oil-sorbents harbor native microbial communities effective in spilled oil-bioremediation under nitrogen starvation and heavy metal-stresses.

Cultivation on selective media revealed that the oil-sorbents, wheat straw, corncobs and sugarcane bagasse harbor hydrocarbonoclastic, diazotrophic and heavy metal-resistant microorganisms. Nitrogen-free media containing 1.0% crude oil lost between 32.2 and 37.5% of this oil, after 8 months when they have been inoculated with such microorganism-loaded sorbents. The used wheat straw, corncobs and sugarcane bagasse samples, 1.0 g each, absorbed respectively, 1.9, 1.1 and 2.5 g oil samples, and lost 24.3-39.2% of these amounts, after they had been incubated for 8 months. Total genomic DNA's from culture media and sorbents revealed various nitrogenase-coding nifH-genes. Pure hydrocarbonoclastic microbial isolates tolerated certain concentrations of, Hg2+, Cd2+, Pb2+, AsO43- and AsO33-. Some of those isolates even grew excellently with up to 1000 ppm of Pb2+ and 36,000 ppm of AsO43- also in the presence of oil. Tested strains removed the tested heavy metals, Hg2+, Cd2+ and Pb2+ from the media and thus, reduced their toxicity against the hydrocarbon-degraders. It was concluded that plant-based sorbents, not only remove oil physically, but also harbor microbial communities effective in spilled oil-bioremediation under multiple stresses. Although each community consisted of one to three species only, the consortia which reached in numbers millions of CFU ml-1 enrich the oily media with fixed nitrogen, and remove heavy metals which otherwise inhibit the oil-degrading microorganisms.

Bioremediation of Atmospheric Hydrocarbons via Bacteria Naturally Associated with Leaves of Higher Plants.

Bacteria associated with leaves of sixteen cultivated and wild plant species from all over Kuwait were analyzed by a culture-independent approach. This technique depended on partial sequencing of 16S rDNA regions in total genomic DNA from the bacterial consortia and comparing the resulting sequences with those in the GenBank database. To release bacterial cells from leaves, tough methods such as sonication co-released too much leaf chloroplasts whose DNA interfered with the bacterial DNA. A more satisfactory bacterial release with a minimum of chloroplast co-release was done by gently rubbing the leaf surfaces with soft tooth brushes in phosphate buffer. The leaves of all plant species harbored on their surfaces bacterial communities predominated by hydrocarbonoclastic (hydrocarbon-utilizing) bacterial genera. Leaves of 6 representative plants brought about in the laboratory effective removal of volatile hydrocarbons in sealed microcosms. Each individual plant species had a unique bacterial community structure. Collectively, the phyllospheric microflora on the studied plants comprised the genera Flavobacterium, Halomonas, Arthrobacter, Marinobacter, Neisseria, Ralstonia, Ochrobactrum. Exiguobacterium, Planomicrobium, Propionibacterium, Kocuria, Rhodococcus and Stenotrophomonas. This community structure was dramatically different from the structure we determined earlier for the same plants using the culture-dependent approach, although in both cases, hydrocarbonoclastic bacteria were frequent.

Moderately thermophilic, hydrocarbonoclastic bacterial communities in Kuwaiti desert soil: enhanced activity via Ca(2+) and dipicolinic acid amendment.

Pristine and oil-contaminated desert soil samples from Kuwait harbored between 10 and 100 cells g(-1) of hydrocarbonoclastic bacteria capable of growth at 50 °C. Enrichment by incubation of moistened soils for 6 months at 50 °C raised those numbers to the magnitude of 10(3) cells g(-1). Most of these organisms were moderately thermophilic and belonged to the genus Bacillus; they grew at 40-50 °C better than at 30 °C. Species belonging to the genera Amycolatopsis, Chelativorans, Isoptericola, Nocardia, Aeribacillus, Aneurinibacillus, Brevibacillus, Geobacillus, Kocuria, Marinobacter and Paenibacillus were also found. This microbial diversity indicates a good potential for hydrocarbon removal in soil at high temperature. Analysis of the same desert soil samples by a culture-independent method (combined, DGGE and 16S rDNA sequencing) revealed dramatically different lists of microorganisms, many of which had been recorded as hydrocarbonoclastic. Many species were more frequent in the oil contaminated than in the pristine soil samples, which may reflect their hydrocarbonoclastic activity in situ. The growth and hydrocarbon consumption potential of all tested isolates were dramatically enhanced by amendment of the cultures with Ca(2+) (up to 2.5 M CaSO4). This enhanced effect was even amplified when in addition 8 % w/v dipicolinic acid was amended. These novel findings are useful in suggesting biotechnologies for waste hydrocarbon remediation at moderately high temperature.

Biofilms constructed for the removal of hydrocarbon pollutants from hypersaline liquids.

Hydrocarbonoclastic biofilms were established on sterile glass plates vertically submerged for 1 month in a hypersaline soil/water suspension containing 0.3% crude oil. The culture-dependent analysis of the microbial community in those biofilms revealed hydrocarbonoclastic species in the magnitude of 10(3) cells cm(-2). Those species belonged to the halophilic bacterial genera Marinobacter, Halomonas, Dietzia, Bacillus, Arhodomonas, Aeromonas and Kocuria as well as to the haloarchaeal genera Haloferax and Halobacterium. Those organisms were not evenly distributed over the biofilm surface area. The culture-independent analysis revealed a different community composition, which was based on four uncultured and four cultured taxa. Depending on the culture conditions and the sort of chemical amendments, the biofilms succeeded in removing in 2 weeks up to about 60-70% of crude oil, pure n-hexadecane and pure phenanthrene in hypersaline pond water samples. The amendment with KCl, MgSO4 and a vitamin mixture composed of thiamin, pyridoxine, vitamin B12, biotin, riboflavin and folic acid was most effective.

Bioremediation of hydrocarbons contaminating sewage effluent using man-made biofilms: effects of some variables.

Biofilm samples were established on glass slides by submerging them in oil-free and oil-containing sewage effluent for a month. In batch cultures, such biofilms were effective in removing crude oil, pure n-hexadecane, and pure phenanthrene contaminating sewage effluent. The amounts of the removed hydrocarbons increased with increasing biofilm surface area exposed to the effluent. On the other hand, addition of the reducing agent thioglycollate dramatically inhibited the hydrocarbon bioremediation potential of the biofilms. The same biofilm samples removed contaminating hydrocarbons effectively in three successive batch bioremediation cycles but started to become less effective in the cycles thereafter, apparently due to mechanical biofilm loss during successive transfers. As major hydrocarbonoclastic bacteria, the biofilms harbored species belonging to the genera Pseudomonas, Microvirga, Zavarzinia, Mycobacterium, Microbacterium, Stenotrophomonas, Gordonia, Bosea, Sphingobium, Brachybacterium, and others. The nitrogen fixer Azospirillum brasilense and the microalga Ochromonas distigma were also present; they seemed to enrich the biofilms, with nitrogenous compounds and molecular oxygen, respectively, which are known to enhance microbiological hydrocarbon degradation. It was concluded that man-made biofilms based upon sewage microflora are promising tools for bioremediation of hydrocarbons contaminating sewage effluent.

Hydrocarbonoclastic biofilms based on sewage microorganisms and their application in hydrocarbon removal in liquid wastes.

Attempts to establish hydrocarbonoclastic biofilms that could be applied in waste-hydrocarbon removal are still very rare. In this work, biofilms containing hydrocarbonoclastic bacteria were successfully established on glass slides by submerging them in oil-free and oil-containing sewage effluent for 1 month. Culture-dependent analysis of hydrocarbonoclastic bacterial communities in the biofilms revealed the occurrence of the genera Pseudomonas, Microvirga, Stenotrophomonas, Mycobacterium, Bosea, and Ancylobacter. Biofilms established in oil-containing effluent contained more hydrocarbonoclastic bacteria than those established in oil-free effluent, and both biofilms had dramatically different bacterial composition. Culture-independent analysis of the bacterial flora revealed a bacterial community structure totally different from that determined by the culture-dependent method. In microcosm experiments, these biofilms, when used as inocula, removed between 20% and 65% crude oil, n-hexadecane, and phenanthrene from the surrounding effluent in 2 weeks, depending on the biofilm type, the hydrocarbon identity, and the culture conditions. More of the hydrocarbons were removed by biofilms established in oil-containing effluent than by those established in oil-free effluent, and by cultures incubated in the light than by those incubated in the dark. Meanwhile, the bacterial numbers and diversities were enhanced in the biofilms that had been previously used in hydrocarbon bioremediation. These novel findings pave a new way for biofilm-based hydrocarbon bioremediation, both in sewage effluent and in other liquid wastes.

Bioremediation of oily hypersaline soil and water via potassium and magnesium amendment.

Ten hydrocarbonoclastic halobacterial species and 5 haloarchaeal species that had been isolated on a mineral medium with oil as the sole carbon source grew better and consumed more crude oil, as measured by gas-liquid chromatography, in media receiving between 0.50 and 0.75 mol/L KCl and between 1.50 and 2.25 mol/L MgSO4. Chemical analysis revealed that within a certain limit, the higher the KCl and MgSO4 concentrations in the medium, the more K⁺ and Mg²âº, respectively, was accumulated by cells of all the tested halobacteria and haloarchaea. Also, in experiments in which total natural microbial consortia in hypersaline soil and water samples were directly used as inocula, the consumption of hydrocarbons was enhanced in the presence of the above given concentrations of KCl and MgSO4. It was concluded that amendment with calculated concentrations of K⁺ and Mg²âº could be a promising practice for hydrocarbon bioremediation in hypersaline environments.

Oil-bioremediation potential of two hydrocarbonoclastic, diazotrophic Marinobacter strains from hypersaline areas along the Arabian Gulf coasts.

Two halophilic, hydrocarbonoclastics bacteria, Marinobacter sedimentarum and M. flavimaris, with diazotrophic potential occured in hypersaline waters and soils in southern and northern coasts of Kuwait. Their numbers were in the magnitude of 10(3) colony forming units g(-1). The ambient salinity in the hypersaline environments was between 3.2 and 3.5 M NaCl. The partial 16S rRNA gene sequences of the two strains showed, respectively, 99 and 100% similarities to the sequences in the GenBank. The two strains failed to grow in the absence of NaCl, exhibited best growth and hydrocarbon biodegradation in the presence of 1 to 1.5 M NaCl, and still grew and maintained their hydrocarbonoclastic activity at salinities up to 5 M NaCl. Both species utilized Tween 80, a wide range of individual aliphatic hydrocarbons (C9-C40) and the aromatics benzene, biphenyl, phenanthrene, anthracene and naphthalene as sole sources of carbon and energy. Experimental evidence was provided for their nitrogen-fixation potential. The two halophilic Marinobacter strains successfully mineralized crude oil in nutrient media as well as in hypersaline soil and water microcosms without the use of any nitrogen fertilizers.

Enhanced haloarchaeal oil removal in hypersaline environments via organic nitrogen fertilization and illumination.

Hypersaline soil and pond water samples were mixed with 3 % crude oil, some samples were autoclaved to serve as sterile controls; experimental samples were not sterilized. After 6-week incubation at 40 °C under light/dark cycles, the soil microflora consumed 66 %, and after 4 weeks the pond water microflora consumed 63 % of the crude oil. Soil samples treated with 3 % casaminoacids lost 89 % of their oil after 6 weeks and water samples lost 86 % after 4 weeks. Samples treated with casaminoacids and antibiotics that selectively inhibited bacteria, lost even more oil, up to 94 %. Soil-water mixtures incubated under continuous illumination lost double as much more oil than samples incubated in the dark. The soil-water mixture at time zero contained 1.3 × 10(4) CFU g(-1) of hydrocarbon-utilizing microorganisms which were affiliated to Halomonas aquamarina, Exiguobacterium aurantiacum, Haloferax sp., Salinococcus sp., Marinococcus sp. and Halomonas sp. After 6-week incubation with oil, these numbers were 8.7 × 10(7) CFU g(-1) and the Haloferax sp. proportion in the total microflora increased from 20 to 93 %. Experiments using the individual cultures and three other haloarchaea isolated earlier from the same site confirmed that casaminoacids and light enhanced their oil consumption potential in batch cultures.

Hydrocarbon-utilizing microorganisms naturally associated with sawdust.

Sawdust, one of the materials used as sorbent for removing spilled oil from polluted environments was naturally colonized by hydrocarbon-utilizing fungi, 1×10(5)-2×10(5) colony forming units (CFU) g(-1), depending on the hydrocarbon substrate. This sorbent was initially free of hydrocarbon-utilizing bacteria. Incubating wet sawdust at 30°C resulted in gradually increasing the fungal counts to reach after 6months between 5×10(6) and 7×10(6)CFUg(-1), and the appearance of hydrocarbon-utilizing bacteria in numbers between 8×10(4) and 3×10(5)cellsg(-1). The fungi belonged to the genera Candida (32% of the total), Penicillium (21%), Aspergillus (15%), Rhizopus (12%), Cladosporium (9%), Mucor (7%) and Fusarium (4%). Based on their 16S rRNA gene sequences the bacteria were affiliated to Actinobacterium sp. (38%), Micrococcus luteus (30%), Rhodococcus erythropolis, (19%) and Rhodococcus opacus (13%). Individual pure fungal and bacterial isolates grew on a wide range of individual pure aliphatic (n-alkanes with chain lengths between C(9) and C(40)) and aromatic (benzene, biphenyl, anthracene, naphthalene and phenanthrene) hydrocarbons as sole sources of carbon and energy. Quantitative determinations revealed that all fungal and bacterial isolates could consume considerable proportions of crude oil, phenanthrene (an aromatic hydrocarbon) and n-hexadecane (an aliphatic hydrocarbon) in batch cultures. It was concluded that when sawdust is used as a sorbent, the associated microorganisms probably contribute to the bioremediation of oil and hydrocarbon pollutants in the environment.

Mercury resistance and volatilization by oil utilizing haloarchaea under hypersaline conditions.

The hydrocarbon utilizing haloarchaea, Haloferax (two strains), Halobacterium and Halococcus from a hypersaline coastal area of the Arabian Gulf, had the potential for resistance and volatilization of Hg(2+). Individual haloarchaea resisted up to between 100 and 200 ppm HgCl2 in hydrocarbon free media with salinities between 1 and 4 M NaCl, but only up to between 20 and 30 ppm in a mineral medium containing 3 M NaCl, with 0.5% (w/v) crude oil, as a sole source of carbon and energy. Halococcus and Halobacterium volatilized more mercury than Haloferax. The individual haloarchaea consumed more crude oil in the presence of 3 M NaCl than in the presence of 2 M NaCl. At both salinities, increasing the HgCl2 concentration in the medium from 0 to 20 ppm resulted in decreasing the oil consumption values by the individual haloarchaea. However, satisfactory oil consumption still occurred in the presence of 10 ppm HgCl2. It was concluded that haloarchaea with the combined potential for mercury resistance and volatilization and hydrocarbon consumption could be useful in removing toxic mercury forms effectively from oil free, mercury contaminated, hypersaline environments, and mercury and oil, albeit less effectively, from oily hypersaline environments.

Phytoremediation of mercury in pristine and crude oil contaminated soils: Contributions of rhizobacteria and their host plants to mercury removal.

The rhizospheric soils of three tested legume crops: broad beans (Vicia faba), beans (Phaseolus vulgaris) and pea (Pisum sativum), and two nonlegume crops: cucumber (Cucumis sativus) and tomato, (Lycopersicon esculentum) contained considerable numbers (the magnitude of 10(5)g(-1) soil) of bacteria with the combined potential for hydrocarbon-utilization and mercury-resistance. Sequencing of the 16S rRNA coding genes of rhizobacteria associated with broad beans revealed that they were affiliated to Citrobacter freundii, Enterobacter aerogenes, Exiquobacterium aurantiacum, Pseudomonas veronii, Micrococcus luteus, Brevibacillus brevis, Arthrobacter sp. and Flavobacterium psychrophilum. These rhizobacteria were also diazotrophic, i.e. capable of N(2) fixation, which makes them self-sufficient regarding their nitrogen nutrition and thus suitable remediation agents in nitrogen-poor soils, such as the oily desert soil. The crude oil attenuation potential of the individual rhizobacteria was inhibited by HgCl(2), but about 50% or more of this potential was still maintained in the presence of up to 40 mgl(-1) HgCl(2). Rhizobacteria-free plants removed amounts of mercury from the surrounding media almost equivalent to those removed by the rhizospheric bacterial consortia in the absence of the plants. It was concluded that both the collector plants and their rhizospheric bacterial consortia contributed equivalently to mercury removal from soil.

Biodegradation of crude oil and pure hydrocarbons by extreme halophilic archaea from hypersaline coasts of the Arabian Gulf.

Two extreme halophilic Haloferax strains and one strain each of Halobacterium and Halococcus were isolated from a hypersaline coastal area of the Arabian Gulf on a mineral salt medium with crude oil vapor as a sole source of carbon and energy. These archaea needed at least 1 M NaCl for growth in culture, and grew best in the presence of 4 M NaCl or more. Optimum growth temperatures lied between 40 and 45 degrees C. The four archaea were resistant to the antibiotics chloramphenicol, cycloheximide, nalidixic acid, penicillin, streptomycin and tetracycline. The strains could grow on a wide scope of aliphatic and aromatic (both mono-and polynuclear) hydrocarbons, as sole sources of carbon and energy. Quantitative measurements revealed that these extreme halophilic prokaryotes could biodegrade crude oil (13-47%, depending on the strain and medium salinity), n-octadecane (28-67%) and phenanthrene (13-30%) in culture after 3 weeks of incubation. The rates of biodegradation by all strains were enhanced with increasing NaCl concentration in the medium. Optimal concentration was 3 M NaCl, but even with 4 M NaCl the hydrocarbon-biodegradation rates were higher than with 1 and 2 M NaCl. It was concluded that these archaea could contribute to self-cleaning and bioremediation of oil-polluted hypersaline environments.

Oil phytoremediation potential of hypersaline coasts of the Arabian Gulf using rhizosphere technology.

The rhizosphere and phyllosphere of the halophyte Halonemum strobilaceum naturally inhabiting hypersaline coastal areas of the Arabian Gulf harbor up to 8.1 x 10(4)g(-1) and 3 x 10(2)g(-1), respectively, of extremely halophilic oil-utilizing microorganisms. Such organisms were 14- to 38-fold more frequent in the rhizosphere than in the plant-free soil. Frequent genera in the rhizosphere were affiliated to the archaea Halobacterium sp. and Halococcus sp., the firmicute Brevibacillus borstenlensis, and the proteobacteria Pseudoalteromonas ruthenica and Halomonas sinaensis. The phyllospheric microflora consisted of the dimorphic yeast Candida utilis and the two proteobacteria Ochrobactrum sp. and Desulfovibrio sp. Individual strains grew on a range of pure aliphatic and aromatic hydrocarbons, as sole sources of carbon and energy. All the strains, except C. utilis which could not tolerate salinities >2M NaCl, grew also in media with salinities ranging between 1 and 4M NaCl, with optimum growth between 1 and 2M NaCl. With the exception of the two archaeal genera, all isolates could grow in a nitrogen-free medium. The total rhizospheric and phyllospheric microbial consortia could attenuate crude oil in complete (nitrogen-containing) medium, but also equally well in a nitrogen-free medium. It was concluded that H. strobilaceum could be a valuable halophyte for phytoremediation of oil-polluted hypersaline environments via rhizosphere technology.

Oil-utilizing bacteria associated with fish from the Arabian Gulf.

AIMS: The objectives were to count and identify the oil-utilizing bacteria associated with fish, and to study their hydrocarbon-degradation potential. METHODS AND RESULTS: The standard dilution-plate method using a medium with crude oil as a sole source of carbon and energy revealed that 10 different fish sorts from the Arabian Gulf and two from fish farms accommodated millions of oil-utilizing bacteria per square centimetre of fish surface and per gram of gills and guts. According to their 16S rRNA sequences, those bacteria were affiliated to Psychrobacter, Vibrio, Planococcus, Pseudomonas and Actinobacterium. Planktonic and benthic biomass samples from the Gulf were also rich in oil-utilizing bacteria, but with different composition. All isolates could grow on n-alkanes from C(8) to C(40) and three representative aromatics as individual sole sources of carbon and energy. Quantitative analysis of hydrocarbons by gas-liquid chromatography revealed that the biomass samples of the individual bacteria could consume crude oil, n-octadecane and phenanthrene in liquid media. CONCLUSIONS: The abundant oil-utilizing bacterial associated with fish have the potential for cleaning oily waters. SIGNIFICANCE AND IMPORTANCE OF THE STUDY: Aquatic fauna accommodates rich consortia of oil-utilizing bacteria.

Alkaliphilic and halophilic hydrocarbon-utilizing bacteria from Kuwaiti coasts of the Arabian Gulf.

Green animate materials from the intertidal zone of the Arabian Gulf coast accommodated more alkaliphilic and halophilic bacteria than inanimate materials. The alkaliphilic oil-utilizing bacteria, as identified by their 16S ribonucleic acid sequences, belonged to the following genera arranged in decreasing frequences: Marinobacter, Micrococcus, Dietzia, Bacillus, Oceanobacillus, and Citricoccus. The halophilic oil-utilizing bacteria belonged to the genera: Marinobacter, Georgenia, Microbacterium, Stappia, Bacillus, Isoptericola, and Cellulomonas. Most isolates could grow on a wide range of pure n-alkanes and aromatic compounds, as sole sources of carbon and energy. Quantitative gas liquid chromatographic analysis showed that individual isolates attenuated crude oil and representative pure hydrocarbons in culture. The optimum pH for most of the alkaliphilic genera was pH 10, and the optimum salinity for the halophiles ranged between 2.5 and 5% NaCl (w/v). It was concluded that as far as their microbial makeup is concerned, oily alkaline and saline intertidal areas of the Kuwaiti coasts have a self-cleaning potential.

Enhancing the growth of Vicia faba plants by microbial inoculation to improve their phytoremediation potential for oily desert areas.

Two experiments were conducted to investigate the effect of inoculating Vicia faba plants (broad beens) raised in clean and oily sand with nodule-forming rhizobia and plant-growth-promoting rhizobacteria (PGPR) on growth of these plants in sand and to test whether this can improve the phytoremediation potential of this crop for oily desert areas. It was found that crude oil in sand at concentrations < 1.0% (w/w) enhanced the plant heights, their fresh and dry weights, the total nodule weights per plant, and the nitrogen contents of shoots and fruits. Similar enhancing effects were recorded when roots of the young plants were inoculated with nodule bacteria alone, PGPR alone, or a mixture of one strain of nodule bacteria and one of the PGPR. Such plant growth effects were associated with a better phytoremediation potential of V. faba plants for oily sand. The total numbers of oil-utilizing bacteria increased in the rhizosphere and more hydrocarbons were eliminated in sand close to the roots. The nodule bacteria tested were two strains of Rhizobium leguminosarum and the PGPR were Pseudomonas aeruginosa and Serratia liquefaciens. The four strains were found to use crude oil, n-octadecane, and phenanthrene as sole sources of carbon and energy. It was concluded that coinoculation of V. faba plant roots in oily sand with nodule bacteria and PGPR enhances the phytoremediation potential of this plant for oily desert sand through improving plant growth and nitrogen fixation.

Hydrocarbon accumulation by picocyanobacteria from the Arabian Gulf.

AIMS: The objective of this work was to study picocyanobacteria in the Arabian Gulf water in relation to oil pollution. METHODS AND RESULTS: Epifluorescent microscopic counting showed that offshore water samples along the Kuwaiti coast of the Arabian Gulf were rich in picocyanobacteria which ranged in numbers between about 1 x 10(5) and 6 x 10(5) ml(-1). Most dominant was the genus Synechococcus; less dominant genera were Synechocystis, Pleurocapsa and Dermocarpella. All isolates grew well in an inorganic medium containing up to 0.1% crude oil (w/v) and could survive in the presence of up to 1% crude oil. Hydrocarbon analysis by gas liquid chromatography (GLC) showed that representative strains of the four genera had the potential for the accumulation of hydrocarbons (the aliphatic n-hexadecane, aromatic phenanthrene and crude oil hydrocarbons) from aqueous media. Electron microscopy showed that the cells of these strains appeared to store hydrocarbons in their inter thylakoid spaces. Analysis by GLC of constituent fatty acids of total lipids and individual lipid classes from representative picoplankton strains grown in the absence and presence of hydrocarbons showed, however, that the fatty acid patterns were not markedly affected by the hydrocabon substrates, meaning that the test strains could not oxidize the accumulated hydrocarbons. CONCLUSION: The Arabian Gulf is among the water bodies of the world richest in picocyanobacteria. These micro-organisms accumulate hydrocarbons from the water body, but do not biodegrade these compounds. It is assumed that hydrocarbon-utilizing bacteria that were always found associated with all picocyanobacteria in nature may carry out the biodegradation of these compounds. SIGNIFICANCE AND IMPORTANCE OF THE STUDY: The results shed light on the potential role of picocyanobacteria in controlling marine oil pollution.

Effects of lipids on n-alkane attenuation in media supporting oil-utilizing microorganisms from the oily Arabian Gulf coasts.

The Arabian Gulf is one of the most extensively oil-polluted areas of the world. The major objectives of this work were to study whether hydrocarbon-utilizing microorganisms indigenous to that area would readily accumulate added lipids, and whether this might affect their hydrocarbon consumption potential. Two prokaryotes, Arthrobacter nicotianae KCC B35 and the unidentified organisms KCC B6, as well as one eukaryote, Candida parapsilosis KCC Y1, were selected for this study. Biomass samples of the test organisms were incubated in an inorganic medium containing various concentrations of cholesterol, stearic acid, triolein or egg-phospholipids. The results revealed that all lipid classes were readily accumulated by the three test organisms. In addition, biomass samples were incubated for 6 h in an inorganic medium containing mixtures of individual lipid classes and either n-octadecane or n-docosane. The cells were removed and the residual alkanes in the medium were quantitatively recovered and analyzed by GLC. The results showed that out of the tested lipid classes only stearic acid exhibited a common stimulatory effect on the consumption of both n-alkanes by all test organisms. Other lipid classes were either inhibitory or had less pronounced effects than stearic acid.

Vitamin requirements of hydrocarbon-utilizing soil bacteria.

The numbers of oil-utilizing bacteria in several samples of clean and oil-polluted soils counted on vitamin-containing media were severalfold higher than the numbers counted on vitamin-free media. Colonies that grew on a medium containing a vitamin mixture were tested for growth on the same medium lacking any vitamins. More than 90% of the total colonies failed to grow. The remaining 10% grew, yet their growth was enhanced, when vitamins were added. The predominant oil-utilizing bacteria in one of the test desert soil samples were various strains of Cellulomonas flavigena and Rhodococcus erythropolis. Minor organisms belonged to the genera Pseudomonas, Bacillus and Arthrobacter. Two vitamin-requiring biovars of C. flavigena and R. erythropolis were selected for further study. Their growth on n-octadecane and phenanthrene as sole sources of carbon and energy as well as their potential for hydrocarbon consumption were enhanced by added vitamins, e.g. folic acid, pyridoxine, vitamin B12, biotin and others. In a field experiment, it was confirmed that vitamin fertilization of an oil-polluted sand sample enhanced the biodegradation of constituent hydrocarbons of that sample.