Gene diversity of CYP153A and AlkB alkane hydroxylases in oil‐degrading bacteria isolated from the Atlantic Ocean

Alkane hydroxylases, including the integral‐membrane non‐haem iron monooxygenase (AlkB) and cytochrome P450 CYP153 family, are key enzymes in bacterial alkane oxidation. Although both genes have been detected in a number of bacteria and environments, knowledge about the diversity of these genes in marine alkane‐degrading bacteria is still limited, especially in pelagic areas. In this report, 177 bacterial isolates, comprising 43 genera, were obtained from 18 oil‐degrading consortia enriched from surface seawater samples collected from the Atlantic Ocean. Many isolates were confirmed to be the first oil‐degraders in their affiliated genera including Brachybacterium, Idiomarina, Leifsonia, Martelella, Kordiimonas, Parvibaculum and Tistrella. Using degenerate PCR primers, alkB and CYP153A P450 genes were surveyed in these bacteria. In total, 82 P450 and 52 alkB gene fragments were obtained from 80 of the isolates. These isolates mainly belonged to Alcanivorax, Bacillus, Erythrobacter, Martelella, Parvibaculum and Salinisphaera, some of which were reported, for the first time, to encode alkane hydroxylases. Phylogenetic analysis showed that both genes were quite diverse and formed several clusters, most of which were generated from various Alcanivorax bacteria. Noticeably, some sequences, such as those from the Salinisphaera genus, were grouped into a distantly related novel cluster. Inspection of the linkage between gene and host revealed that alkB and P450 tend to coexist in Alcanivorax and Salinisphaera, while in all isolates of Parvibaculum, only P450 genes were found, but of multiple homologues. Multiple homologues of alkB mostly cooccurred in Alcanivorax isolates. Conversely, distantly related isolates contained similar or even identical sequences. In summary, various oil‐degrading bacteria, which harboured diverse P450 and alkB genes, were found in the surface water of Atlantic Ocean. Our results help to show the diversity of P450 and alkB genes in prokaryotes, and to portray the geographic distribution of oil‐degrading bacteria in marine environments.     

Indigenous oil-degrading bacteria in crude oil-contaminated seawater of the Yellow sea,China

Indigenous oil-degrading bacteria play an important role in efficient remediation of polluted marine environments. In this study, we investigated the diversity and abundance of indigenous oil-degrading bacteria and functional genes in crude oil-contaminated seawater of the Dalian coast. The gene copy number bacterial 16S rRNA in total were determined to be about 10¹⁰ copies L^?1 in contaminated seawater and 10⁹ copies L^?1 in uncontaminated seawater. Bacteria of Alcanivorax, Marinobacter, Novosphingobium, Rhodococcus, and Pseudoalteromonas were found to be predominant oil-degrading bacteria in the polluted seawater in situ. In addition, bacteria belonging to Algoriphagus, Aestuariibacter, Celeribacter, Fabibacter, Zobellia, Tenacibaculum, Citreicella, Roseivirga, Winogradskyella, Thioclava, Polaribacter, and Pelagibaca were confirmed to be the first time as an oil-degrading bacterium. The indigenous functional enzymes, including AlkB or polycyclic aromatic hydrocarbons ring-hydroxylating dioxygenases α (PAH-RHDα) coding genes from Gram-positive (GP) and Gram-negative bacteria (GN), were revealed and quite diverse. About 10¹⁰ to 10¹¹ copies L^?1 for the expression of alkB genes were recovered and showed that the two-thirds of all the AlkB sequences were closely related to widely distributed Alcanivorax and Marinobacter isolates. About 10⁹ copies L^?1 seawater for the expression of RHDαGN genes in contaminated seawater and showed that almost all RHDαGN sequences were closely related to an uncultured bacterium; however, RHDαGP genes represented only about 10⁵ copies L^?1 seawater for the expression of genes in contaminated seawater, and the naphthalene dioxygenase sequences from Rhodococcus and Mycobacterium species were most abundant. Together, their data provide evidence that there exists an active aerobic microbial community indigenous to the coastal area of the Yellow sea that is capable of degrading petroleum hydrocarbons.     

Identities of Epilithic Hydrocarbon-Utilizing Diazotrophic Bacteria from the Arabian Gulf Coasts, and Their Potential for Oil Bioremediation Without Nitrogen Supplementation

Gravel particles from four sites along the Arabian Gulf coast in autumn, winter, and spring were naturally colonized with microbial consortia containing between 7 and 400 × 10² cm⁻² of cultivable oil-utilizing bacteria. The 16S rRNA gene sequences of 70 representatives of oil-utilizing bacteria revealed that they were predominantly affiliated with the Gammaproteobacteria and the Actinobacteria. The Gammaproteobacteria comprised among others, the genera Pseudomonas, Pseudoalteromonas, Shewanella, Marinobacter, Psychrobacter, Idiomarina, Alcanivorax, Cobetia, and others. Actinobacteria comprised the genera Dietzia, Kocuria, Isoptericola, Rhodococcus, Microbacterium, and others. In autumn, Firmicutes members were isolated from bay and nonbay stations while Alphaproteobacteria were detected only during winter from Anjefa bay station. Fingerprinting by denaturing gradient gel electrophoresis of amplified 16S rRNA genes of whole microbial consortia confirmed the culture-based bacterial diversities in the various epilithons in various sites and seasons. Most of the representative oil-utilizing bacteria isolated from the epilithons were diazotrophic and could attenuate oil also in nitrogen-rich (7.9–62%) and nitrogen-free (4–54%) cultures, which, makes the microbial consortia suitable for oil bioremediation in situ, without need for nitrogen supplementation. This was confirmed in bench-scale experiments in which unfertilized oily seawater was bioremediated by epilithon-coated gravel particles.     

Hydrocarbon-degrading bacteria and the bacterial community response in gulf of Mexico beach sands impacted by the deepwater horizon oil spill

A significant portion of oil from the recent Deepwater Horizon (DH) oil spill in the Gulf of Mexico was transported to the shoreline, where it may have severe ecological and economic consequences. The objectives of this study were (i) to identify and characterize predominant oil-degrading taxa that may be used as model hydrocarbon degraders or as microbial indicators of contamination and (ii) to characterize the in situ response of indigenous bacterial communities to oil contamination in beach ecosystems. This study was conducted at municipal Pensacola Beach, FL, where chemical analysis revealed weathered oil petroleum hydrocarbon (C₈ to C₄₀) concentrations ranging from 3.1 to 4,500 mg kg⁻¹ in beach sands. A total of 24 bacterial strains from 14 genera were isolated from oiled beach sands and confirmed as oil-degrading microorganisms. Isolated bacterial strains were primarily Gammaproteobacteria, including representatives of genera with known oil degraders (Alcanivorax, Marinobacter, Pseudomonas, and Acinetobacter). Sequence libraries generated from oiled sands revealed phylotypes that showed high sequence identity (up to 99%) to rRNA gene sequences from the oil-degrading bacterial isolates. The abundance of bacterial SSU rRNA gene sequences was ∼10-fold higher in oiled (0.44 × 10⁷ to 10.2 × 10⁷ copies g⁻¹) versus clean (0.024 × 10⁷ to 1.4 × 10⁷ copies g⁻¹) sand. Community analysis revealed a distinct response to oil contamination, and SSU rRNA gene abundance derived from the genus Alcanivorax showed the largest increase in relative abundance in contaminated samples. We conclude that oil contamination from the DH spill had a profound impact on the abundance and community composition of indigenous bacteria in Gulf beach sands, and our evidence points to members of the Gammaproteobacteria (Alcanivorax, Marinobacter) and Alphaproteobacteria (Rhodobacteraceae) as key players in oil degradation there.     

Dynamic changes in the structure of microbial communities in Baltic Sea coastal seawater microcosms modified by crude oil,shale oil or diesel fuel

The coastal waters of the Baltic Sea are constantly threatened by oil spills, due to the extensive transportation of oil products across the sea. To characterise the hydrocarbon-degrading bacterial community of this marine area, microcosm experiments on diesel fuel, crude oil and shale oil were performed. Analysis of these microcosms, using alkane monooxygenase (alkB) and 16S rRNA marker genes in PCR-DGGE experiments, demonstrated that substrate type and concentration strongly influence species composition and the occurrence of alkB genes in respective oil degrading bacterial communities. Gammaproteobacteria (particularly the genus Pseudomonas) and Alphaproteobacteria were dominant in all microcosms treated with oils. All alkB genes carried by bacterial isolates (40 strains), and 8 of the 11 major DGGE bands from the microcosms, had more than 95% sequence identity with the alkB genes of Pseudomonas fluorescens. However, the closest relatives of the majority of sequences (54 sequences from 79) of the alkB gene library from initially collected seawater DNA were Actinobacteria. alkB gene expression, induced by hexadecane, was recorded in isolated bacterial strains. Thus, complementary culture dependent and independent methods provided a more accurate picture about the complex seawater microbial communities of the Baltic Sea.     

Predominant culturable crude oil-degrading bacteria in the coast of Kuwait

Total of 272 crude oil-degrading bacteria were isolated from seven locations along the coast of Kuwait. The analysis of the 16S rDNA sequences of isolated bacteria revealed the predominance of six bacterial genera: Pseudomonas, Bacillus, Staphylococcus, Acinetobacter, Kocuria and Micrococcus. Investigation of the factors associated with bacterial predominance revealed that, dominant culturable crude oil-degrading bacteria were better crude oil utilizers than the less frequently occurring isolates. Bacterial predominance was also influenced by the ability of bacteria to adapt to the level of organic content available. Predominant culturable bacteria constituted 89.7–54.2% of the total crude oil-degrading bacterial communities. Using 16S-RFLP analyses to assess the diversity of the dominant crude oil-degrading bacterial genera, four phylotypes of Pseudomonas sp. and seven phylotypes of Bacillus sp. were determined. This suggested high degree of diversity of crude oil-degrading bacterial population at the strain level, but low diversity at the genus level.     

Bacterial degradation of styrene in waste gases using a peat filter

A biofiltration process was developed for styrene-containing off-gases using peat as filter material. The average styrene reduction ratio after 190 days of operation was 70% (max. 98%) and the mean styrene elimination capacity was 12 g m⁻³ h⁻¹ (max. 30 g m⁻³ h⁻¹). Efficient styrene degradation required addition of nutrients to the peat, adjustment of the pH to a neutral level and efficient control of the humidity. Maintenance of the water balance was easier in a down-flow than in an up-flow process, the former consequently resulting in much better filtration efficiency. The optimum operation temperature was around 23 °C, but the styrene removal was still satisfactory at 12 °C. Seven different bacterial isolates belonging to the genera Tsukamurella, Pseudomonas, Sphingomonas, Xanthomonas and an unidentified genus in the γ group of the Proteobacteria isolated from the microflora of active peat filter material were capable of styrene degradation. The isolates differed in their capacity to decompose styrene to carbon dioxide and assimilate it to biomass. No toxic intermediate degradation products of styrene were detected in the filter outlet gas or in growing cultures of isolated bacteria. The use of these isolates in industrial biofilters is beneficial at low styrene concentrations and is safe from both the environmental and public health points of view. Received: 30 May 1997 / Received revision: 22 August 1997 / Accepted: 25 August 1997     

Aerobic heterotrophic bacteria and petroleum-utilizing bacteria from cow dung and poultry manure

In this study, enumeration and identification of total aerobic heterotrophic bacteria and petroleum-utilizing bacteria as well as the degradative potential of petroleum-utilizing bacterial isolates were carried out. The average counts of total aerobic heterotrophic bacteria in cow dung and poultry manure were 74.25 × 10⁵ c.f.u. g⁻¹ and 138.75 × 10⁵ c.f.u. g⁻¹ respectively. Acinetobacter sp, Bacillus sp, Pseudomonas sp, and Serratia spp. occurred as aerobic heterotrophs in both cow dung and poultry manure. However, Alcaligenes spp. occurred only in cow dung while, Flavobacterium sp, Klebsiella sp, Micrococcus sp, and Nocardia spp. occurred only in poultry manure as aerobic heterotrophs. The average counts of petroleum-utilizing bacteria in cow dung and poultry manure were 9.25 × 10⁵ c.f.u. g⁻¹ and 17.25 × 10⁵ c.f.u. g⁻¹ respectively. Pseudomonas spp. occurred as petroleum utilizer in both cow dung and poultry manure. However, Bacillus spp. occurred only in cow dung while Acinetobacter spp. and Micrococcus spp. occurred only in poultry manure as petroleum utilizers. Relative abundance of petroleum utilizers in total aerobic heterotrophs ranged from 6.38% to 20.00% for cow dung and from 9.38% to 17.29% for poultry manure. Introduction of pure cultures of petroleum-utilizing bacteria from cow dung and poultry manure into sterile oil-polluted soil revealed oil degradation in one week period.     

Phylogenetic and Physiological Diversity of Bacteria Isolated from Puruogangri Ice Core

The microbial abundance, the percentage of viable bacteria, and the diversity of bacterial isolates from different regions of a 83.45-m ice core from the Puruogangri glacier on the Tibetan Plateau (China) have been investigated. Small subunit 16S rRNA sequences and phylogenetic relationships have been studied for 108 bacterial isolates recovered under aerobic growth conditions from different regions of the ice core. The genomic fingerprints based on ERIC (enterobacterial repetitive intergenic consensus)-polymerase chain reaction and physiological heterogeneity of the closely evolutionary related bacterial strains isolated from different ice core depths were analyzed as well. The results showed that the total microbial cell, percentages of live cells, and the bacterial CFU ranged from 10⁴ to 10⁵ cell ml⁻¹ (Mean, 9.47 × 10⁴; SD, 5.7 × 10⁴, n = 20), 25–81%, and 0–760 cfu ml⁻¹, respectively. The majority of the isolates had 16S rRNA sequences similar to previously determined sequences, ranging from 92 to 99% identical to database sequences. Based on their 16S rRNA sequences, 42.6% of the isolates were high-G + C-content (HGC) gram-positive bacteria, 35.2% were low-G + C (LGC) gram-positive bacteria, 16.6% were Proteobacteria, and 5.6% were CFB group. There were clear differences in the depth distribution of the bacterial isolates. The isolates tested exhibited unique phenotypic properties and high genetic heterogeneity, which showed no clear correlation with depths of bacterial isolation. This layered distribution and high heterogeneity of bacterial isolates presumably reflect the diverse bacterial sources and the differences in bacteria inhabiting the glacier’s surface under different past climate conditions.     

Antibacterial Activity of Marine Culturable Bacteria Collected from a Global Sampling of Ocean Surface Waters and Surface Swabs of Marine Organisms

The purpose of the present study was to isolate marine culturable bacteria with antibacterial activity and hence a potential biotechnological use. Seawater samples (244) and 309 swab samples from biotic or abiotic surfaces were collected on a global Danish marine research expedition (Galathea 3). Total cell counts at the seawater surface were 5 × 10⁵ to 10⁶ cells/ml, of which 0.1–0.2% were culturable on dilute marine agar (20°C). Three percent of the colonies cultured from seawater inhibited Vibrio anguillarum, whereas a significantly higher proportion (13%) of colonies from inert or biotic surfaces was inhibitory. It was not possible to relate a specific kind of eukaryotic surface or a specific geographic location to a general high occurrence of antagonistic bacteria. Five hundred and nineteen strains representing all samples and geographic locations were identified on the basis of partial 16S rRNA gene sequence homology and belonged to three major groups: Vibrionaceae (309 strains), Pseudoalteromonas spp. (128 strains), and the Roseobacter clade (29 strains). Of the latter, 25 strains were identified as Ruegeria mobilis or pelagia. When re-testing against V. anguillarum, only 409 (79%) retained some level of inhibitory activity. Many strains, especially Pseudoalteromonas spp. and Ruegeria spp., also inhibited Staphylococcus aureus. The most pronounced antibacterial strains were pigmented Pseudoalteromonas strains and Ruegeria spp. The inhibitory, pigmented Pseudoalteromonas were predominantly isolated in warmer waters from swabs of live or inert surfaces. Ruegeria strains were isolated from all ocean areas except for Arctic and Antarctic waters and inhibitory activity caused by production of tropodithietic acid.     

Endophytic Bacteria Associated with Growing Shoot Tips of Banana (Musa sp.) cv. Grand Naine and the Affinity of Endophytes to the Host

A cultivation-based assessment of endophytic bacteria present in deep-seated shoot tips of banana suckers was made with a view to generate information on the associated organisms, potential endophytic contaminants in tissue-cultured bananas and to assess if the endophytes shared a beneficial relationship with the host. Plating the tissue homogenate from the central core of suckers showed colony growth on nutrient agar from just 75% and 42% of the 12 stocks during May and November, respectively (average 58%; 6 × 10³ colony-forming units per gram), yielding diverse organisms belonging to firmicutes (Bacillus, Brevibacillus, Paenibacillus, Virgibacillus, Staphylococcus spp.), actinobacteria (Cellulomonas, Micrococcus, Corynebacterium, Kocuria spp.), α-proteobacteria (Paracoccus sp.), and γ-proteobacteria (Pseudomonas, Acinetobacter spp.). Each shoot tip showed one to three different organisms and no specific organism appeared common to different sucker tips. Tissue homogenate from shoot tips including the ones that did not yield culturable bacteria displayed abundant bacterial cells during microscopic examination suggesting that a high proportion of cells were in viable-but-nonculturable state, or their cultivation requirements were not met. Direct application of cultivation-independent approach to study endophytic bacterial community using bacterial 16S ribosomal RNA universal primers resulted in high interference from chloroplast and mitochondrial genome sequences. Dislodging the bacterial cells from shoot tips that did not show cultivable bacteria and incubating the tissue crush in dilute-nutrient broth led to the activation of four organisms (Klebsiella, Agrobacterium, Pseudacidovorax spp., and an unidentified isolate). The endophytic organisms in general showed better growth at 30–37 °C compared with 25 °C, and the growth of endophytes as well as pathogenic Erwinia carotovora were promoted with the supply of host tissue extract (HTE) while that of the isolates from nonplant sources were inhibited or unaffected by HTE, suggesting an affinity or dependence of the endophytes on the host and the prospect of an HTE-based assay for discriminating the nonendophytes from endophytes.     

Bioremoval of organic and inorganic sulphur from coal samples

The microbial ecology of different Spanish coal samples has been studied. Several bacteria have been isolated from enrichment cultures and characterised and their biodesulphurization abilities evaluated. Using morphological and physiological properties, different isolates have been related to species of the Xanthomonas, Pseudomonas, Chryseomonas and Moraxella genera. Some of the isolates, B(30)15 and T(30)10, gave important levels of organic desulphurization, close to 70%. Other isolates, B(30)7 and B(30)8, were able to remove inorganic sulphur with high efficiencies, over 67%. One of the isolates, B(30)10, metabolically related to Xanthomonas maltophila, was able to remove both organic and inorganic sulphur at neutral pH, with efficiencies of 69% and 68% respectively. The results obtained underline the potential use of some of these strains for industrial coal desulphurization processes. Received: 26 June 1998 / Received revised: 1 October 1998 / Accepted: 2 October 1998     

Arbuscular Mycorrhizal Fungi and Rhizospheric Bacteria Diversity Along an Altitudinal Gradient in South American Puna Grassland

Rhizospheric soil samples were taken from Puna native grasses along an altitudinal gradient. Biodiversity of arbuscular mycorrhizal fungi (AMF) and associated bacteria was analyzed considering altitude and grasses photosynthetic pathways (metabolic type C₃, C₄). Cultivation-dependent approaches were applied to obtain further information about the phylogeny of the dominating cultivable aerobic–heterotrophic bacteria communities present in rhizospheric soil samples. In average, the bacterial count ranged between 1.30 × 10² and 8.66 × 10⁴ CFU g⁻¹ of dry weight of soil. Individual bacterial colonies of aerobic heterotrophic bacteria grown on R2A medium were morphologically grouped and identified as typical soil bacteria belonging to the genera Bacillus, Pseudomonas, and Arthrobacter. Ten AMF taxa were found: Acaulospora sp., A. laevis, A. spinosa, Gigaspora sp., Gi. ramisporophora, Glomus sp., Gl. aggregatum, Gl. ambisporum, Gl. sinuosum, and Scutellospora biornata. AMF diversity decreased with altitude.     

The effect of turbulent flow and surface roughness on biofilm formation in drinking water

There is considerable interest in both Europe and the USA in the effects of microbiological fouling on stainless steels in potable water. However, little is known about the formation and effects of biofilms, on stainless steel in potable water environments, particularly in turbulent flow regimes. Results are presented on the development of biofilms on stainless steel grades 304 and 316 after exposure to potable water at velocities of 0.32, 0.96 and 1.75 m s⁻¹. Cell counts on slides of stainless steel grades 304 and 316 with both 2B (smooth) and 2D (rough) finishes showed viable and total cell counts were higher at the higher flow rates of 0.96 and 1.75 m s⁻¹, compared to a flow rate of 0.32 m s⁻¹. Extracellular polysaccharide levels were not significantly different (P< 0.05) between each flow rate on all stainless steel surfaces studied. higher levels were found at the higher water velocities. the biofilm attached to stainless steel was comprised of a mixed bacterial flora including Acinetobacter sp, Pseudomonas spp, Methylobacterium sp, and Corynebacterium/Arthrobacter spp. Epifluorescence microscopy provided evidence of rod-shaped bacteria and the formation of stands, possibly of extracellular material attached to stainless steel at high flow rates but not at low flow rates. Received 04 February 1998/ Accepted in revised form 12 February 1999     

Isolation of alkane‐degrading bacteria from deep‐sea Mediterranean sediments

Aims: To isolate and identify alkane‐degrading bacteria from deep‐sea superficial sediments sampled at a north‐western Mediterranean station. Methods and Results: Sediments from the water/sediment interface at a 2400 m depth were sampled with a multicorer at the ANTARES site off the French Mediterranean coast and were promptly enriched with Maya crude oil as the sole source of carbon and energy. Alkane‐degrading bacteria belonging to the genera Alcanivorax, Pseudomonas, Marinobacter, Rhodococcus and Clavibacter‐like were isolated, indicating that the same groups were potentially involved in hydrocarbon biodegradation in deep sea as in coastal waters. Conclusions: These results confirm that members of Alcanivorax are important obligate alkane degraders in deep‐sea environments and coexist with other degrading bacteria inhabiting the deep‐subsurface sediment of the Mediterranean. Significance and Impact of the Study: The results suggest that the isolates obtained have potential applications in bioremediation strategies in deep‐sea environments and highlight the need to identify specific piezophilic hydrocarbon‐degrading bacteria (HCB) from these environments.     

Characterization of some fish and shrimp spoiling bacteria

The classification of some important groups of bacteria involved in fish and shrimp spoilage was studied. Trimethylamine is produced byPseudomonas putrefaciens, a “non-defined” group resemblingPs. putrefaciens, Photobacterium spp. and someMoraxella-like bacteria. Hypoxanthine is produced by the same groups of bacteria except the last named and also by the “typical shrimp spoilers” (presumptiveAlteromonas). Strong off-odours are produced on fresh fish byPs. putrefaciens, dextroseoxidativePseudomonas spp. (Groups I and 11 according to Shewan, Hobbs and Hodgkiss, 1960), the above mentioned “non-defined” group and by only some of the “typical shrimp spoilers”, whereasMoraxella-like bacteria andPhotobacterium spp. failed to produce strong odours. Strong off-odours are produced on boiled shrimp by the “typical shrimp spoilers” (presumptiveAltermonas),Ps. putrefaciens, the dextrose-oxidativePseudomonas spp. and the “non-defined” group;Moraxella-like bacteria produced less offensive odours or none, nor didPhotobacterium.     

Biofilm formation in an ice cream plant

The sites of biofilm formation in an ice cream plant were investigated by sampling both the production line and the environment. Experiments were carried out twice within a 20-day period. First, stainless steel coupons were fixed to surfaces adjacent to food contact surfaces, the floor drains and the doormat. They were taken for the analysis of biofilm at three different production stages. Then, biofilm forming bacteria were␣enumerated and also presence of Listeria monocytogenes was monitored. Biofilm forming isolates were selected on the basis of colony morphology and Gram’s reaction; Gram negative cocci and rod, Gram positive cocci and spore forming isolates were identified. Most of the biofilm formations were seen on the conveyor belt of a packaging machine 8 h after the beginning of the production, 6.5 × 10³ cfu cm⁻². Most of the Gram negative bacteria identified belong to Enterobacteriaceae family such as Proteus, Enterobacter, Citrobacter, Shigella, Escherichia, Edwardsiella. The other Gram negative microflora included Aeromonas, Plesiomonas, Moraxella, Pseudomonas or Alcaligenes spp. were also isolated. Gram positive microflora of the ice cream plant included Staphyloccus, Bacillus, Listeria and lactic acid bacteria such as Streptococcus, Leuconostoc or Pediococcus spp. The results from this study highlighted the problems of spread of pathogens like Listeria and Shigella and spoilage bacteria. In the development of cleaning and disinfection procedures in ice cream plants, an awareness of these biofilm-forming bacteria is essential for the ice cream plants.     

Isolation,gene detection and solvent tolerance of benzene,toluene and xylene degrading bacteria from nearshore surface water and Pacific Ocean sediment

BTX (benzene, toluene and xylene) degrading bacteria were isolated from Pacific Ocean sediment and nearshore surface water. In the seawater near a ferry dock, degrading bacteria of a relatively wide diversity were detected, including species of Pseudomonas, Rhodococcus, Exiguobacterium and Bacillus; while species of Bacillus only have been detected from the deep-sea sediment. Most of the isolates showed degradation to more than one compound. Generally better growth was obtained with p-xylene and ethylbenzene than with the other two. All the bacteria could tolerate and grow with the compounds at 5–20% (v/v). Both benzene and toluene degradation related genes had been successfully PCR cloned from the isolates of nearshore water, the detected benzene dioxygenase gene was identical among all the species and close to its soil counterpart. However, they were not detected in all the isolates from deep sea. Results in this report suggested that BTX degrading bacteria widely spread in marine environments and they might be of potentials in biotreatment of BTEX in saline environments.     

Investigation of alkane biodegradation using the microtiter plate method and correlation between biofilm formation, biosurfactant production and crude oil biodegradation

Among 25 crude oil-degrading bacteria isolated from a marine environment, four strains, which grew well on crude oil, were selected for more study. All the four isolated had maximum growth on 2.5% of crude oil and strain BC (Pseudomonas) could remove crude oil by 83%. The drop collapse method and microtiter assay show that this strain produces more biosurfactant, and its biofilm formation is higher compared to other strains. Bacterial adhesions to crude oil for strains CS-2 (Pseudomonas), BC, PG-5 (Rhodococcus) and H (Bacillus) were 30%, 46%, 10% and 1%, respectively. Therefore, strain H with a low production of biosurfactant and biofilm formation had showed the least growth on these compounds. PCR analysis of these four strains showed that all isolates had alk-B genes from group (III) alkane hydroxylase. All isolate strains could utilize cyclohexan, octane, hexadecane, octadecan and diesel fuel oil; however, the microtiter plate assay showed that strain BC had more growth, respiration and biofilm formation on octadecan.