Improvement of the salt aggregation test to study bacterial cell-surface hydrophobicity

Abstract An improved salt aggregation test (improved SAT) was developed to sensitize the determination of bacterial cell-surface hydrophobicity. One drop of a fresh bacterial suspension standardized to an A ^1cm₅₄₀ of 20 (equivalent to 5 × 10⁹ cfu/ml), and one drop each of ammonium sulphate solutions stained with methylene blue, were mixed on a white hydrophobic paper card using toothpicks. The bacterial suspensions, methylene blue stock solutions and the ammonium sulphate solutions (0.01–4.0 M) were made in 0.02 M sodium phosphate buffer, pH 6.8. Bacterial aggregations were read immediately after mixing the salt/bacterial suspensions while the card was gently rocked. Readings were also confirmed the next day on dried preparations. The results proved independent of reading time and mixture conditions (wet or dry preparations). The improved SAT technique is very rapid and sensitive, the reaction is easily read with the naked eye, and the paper cards can be stored for documentation of aggregation patterns after drying. In the improved SAT, the Staphylococcus cells of different species aggregated in 5 ways: tiny, medial, flaky granular, particulated and macrofilamentous forms; Salmonella strains aggregated in flaky granular, particulated and macrofilamentous forms.     

Insight into heterogeneity in cell-surface hydrophobicity and ability to degrade hydrocarbons among cells of two hydrocarbon-degrading bacterial populations

The sequential bacterial adherence to hydrocarbons (BATH) of successive generations of hydrophobic fractions of Paenibacillus sp. R0032A and Burkholderia cepacia gave rise to bacterial populations of increasing cell-surface hydrophobicity. Thus, hydrophobicity of the first generation (H1) was less than that of the second generation (H2), which was less than that of the third generation (H3). Beyond H3, the hydrophobic populations became less stable and tended to lyse in hexadecane after violent (vortex) agitation, resulting in an apparent decline in BATH value. The exhaustively fractionated aqueous-phase population (L) was very hydrophilic. The overall cell-surface distribution of the population was L < parental strain < H1 < H2 < H3. The ability to degrade crude oil, hexadecane, or phenanthrene matched the degree of cell-surface hydrophobicity: L < P < H1 < H2 < H3. Thus, in natural populations of hydrocarbon-degrading Paenibacillus sp. R0032A and B. cepacia, there is a heterogeneity in the hydrophobic surface characteriistics that affects the ability of cells to use various hydrocarbon substrates.     

High-temperature hydrocarbon degradation by Bacillus stearothermophilus from oil-polluted Kuwaiti desert

Kuwaiti desert samples contaminated with crude oil contained Bacillus stearothermophilus strains capable of growth on crude oil as a sole source of carbon and energy, obligately at high temperature. No thermophilic oil utilizers were present in water samples collected from the Arabian Gulf. Most of the desert strains had an optimum temperature of 60°C and grew best on pentadecane (C₁₅), hexadecane (C₁₆) and heptadecane (C₁₇). n-Alkanes with shorter and longer chains, n-alkenes, and aromatic hydrocarbons were less readily utilized. Correspondence to: N. A. Sorkhoh     

Changes in bacterial communities from anaerobic digesters during petroleum hydrocarbon degradation

Anaerobic biodegradation of petroleum hydrocarbons (PHC) to methane has been recognized to occur in oil reservoirs and contaminated surface sites alike. This process could be employed efficiently for the treatment of contaminated materials, including petrochemical wastes and PHC-contaminated soil, since no external electron acceptor is required. Moreover, the controlled production of methane in digestion plants, similarly to the anaerobic digestion (AD) of energy crops or organic residues, would enable for energy recovery from these wastes. At present, little is known about the bacterial communities involved in and responsible for hydrocarbon fermentation, the initial step in PHC conversion to methane. In the present study, the fate of two different methanogenic communities derived from the AD of wastewater (WWT) and of biowaste, mixed with PHC-contaminated soil (SWT), was monitored during incubation with PHC using denaturing gradient gel electrophoresis (DGGE) of 16S rDNA genes amplified with Bacteria-specific primers. During 11 months of incubation, slight but significant degradation of PHC occurred in both sludges and distinct bacterial communities were developing. In both sludges, Bacteroidetes were found. In addition, in WWT, the bacterial community was found to be dominated by Synergistetes and Proteobacteria, while Firmicutes and unidentified members were abundant in SWT. These results indicate that bacterial communities from anaerobic digesters can adapt to and degrade petroleum hydrocarbons. The decontamination of PHC-containing waste via fermentative treatment appears possible.     

小黑麦对石油污染盐碱土壤细菌群落与石油烃降解的影响

为了研究小黑麦对石油污染盐碱土壤中的细菌群落与石油烃降解率的影响,采用高通量测序技术,设置0 g/kg,1 g/kg和5 g/kg三个石油浓度,以未种植小黑麦的土壤作为对照,对6组不同处理的盐碱土壤样品的细菌群落结构及其多样性进行测定,并分析土壤中的石油烃降解率。结果表明:在土壤石油浓度为1 g/kg和5 g/kg时,小黑麦根际土壤中的石油烃降解率相较对照组分别提高了36.67%和33.20%。从6个土壤样品中分别获得21398—27899条测序序列。在石油污染土壤中,小黑麦根际土壤的细菌群落多样性和丰度均大于对照组的土壤。同时,在"门","纲","属"的分类水平上,小黑麦根际土壤细菌群落中的一些根际细菌的相对丰度增加了,主要包括变形菌门(Proteobacteria)、酸杆菌门(Acidobacteria)、γ-变形菌纲(Gammaproteobacteria)、烷烃降解菌科-未命名菌属(Alcanivoracaceae_norank)、黄单胞菌属(Xanthomonas)、亚硝化单胞菌-不可培养菌属(Nitrosomonadaceae_unculture)等。有一些相对丰度增加的根际细菌是以石油及石油分解物为碳源的微生物。本研究证明种植小黑麦改变了石油污染盐碱土壤根际土壤细菌群落结构组成和多样性,促进了降解石油微生物群落的构建,显著提高了盐碱土壤石油污染的降解效果。研究结果为石油污染盐碱土壤的植物修复奠定了理论基础。     

Root exudates modify bacterial diversity of phenanthrene degraders in PAH-polluted soil but not phenanthrene degradation rates

To determine whether the diversity of phenanthrene‐degrading bacteria in an aged polycyclic aromatic hydrocarbon (PAH) contaminated soil is affected by the addition of plant root exudates, DNA stable isotope probing (SIP) was used. Microcosms of soil with and without addition of ryegrass exudates and with ¹³C‐labelled phenanthrene (PHE) were monitored over 12 days. PHE degradation was slightly delayed in the presence of added exudate after 4 days of incubation. After 12 days, 68% of added PHE disappeared both with and without exudate. Carbon balance using isotopic analyses indicated that a part of the ¹³C‐PHE was not totally mineralized as ¹³CO₂ but unidentified ¹³C‐compounds (i.e. ¹³C‐PHE or ¹³C‐labelled metabolites) were trapped into the soil matrix. Temporal thermal gradient gel electrophoresis (TTGE) analyses of 16S rRNA genes were performed on recovered ¹³C‐enriched DNA fractions. 16S rRNA gene banding showed the impact of root exudates on diversity of PHE‐degrading bacteria. With PHE as a fresh sole carbon source, Pseudoxanthomonas sp. and Microbacterium sp. were the major PHE degraders, while in the presence of exudates, Pseudomonas sp. and Arthrobacter sp. were favoured. These two different PHE‐degrading bacterial populations were also distinguished through detection of PAH‐ring hydroxylating dioxygenase (PAH‐RHD_α) genes by real‐time PCR. Root exudates favoured the development of a higher diversity of bacteria and increased the abundance of bacteria containing known PAH‐RHD_α genes.     

Special stomatal distribution in Sabina vulgaris in relation to its survival in a desert environment

In the semi-desert environment of the Mu Us Sandland, the vegetation is composed chiefly of shrubs and semi-shrubs, coverage normally amounting to 30-40%. However, an exception can be found in the community of Sabina vulgaris Antoine, an evergreen shrub, which tends to grow so densely that it covers the sand dunes completely. Previous research has indicated that the high density of Sabina is parallelled by the very low transpiration rate. Based on anatomical and scanning electronic microscopic observations, this paper also points out that two structural features should play an important role in the potential mechanism of water conservation in this plant. First, a low stomatal density in S. vulgaris can lead to a high stomatal resistance and low transpiration. Second, the morphological feature of S. vulgaris leaves being tightly pressed to the stems might be a structural feature of water conservation, because in this way the leaf side bearing more (and mostly larger) stomatal pores could be protected from direct high radiation, while the side with fewer (and mostly smaller) pores (the outer side) is exposed to the dry desert air.     

Putative degraders of low-density polyethylene-derived compounds are ubiquitous members of plastic-associated bacterial communities in the marine environment

It remains unknown whether and to what extent marine prokaryotic communities are capable of degrading plastic in the ocean. To address this knowledge gap, we combined enrichment experiments employing low-density polyethylene (LDPE) as the sole carbon source with a comparison of bacterial communities on plastic debris in the Pacific, the North Atlantic and the northern Adriatic Sea. A total of 35 operational taxonomic units (OTUs) were enriched in the LDPE-laboratory incubations after 1 year, of which 20 were present with relative abundances > 0.5% in at least one plastic sample collected from the environment. From these, OTUs classified as Cognatiyoonia, Psychrobacter, Roseovarius and Roseobacter were found in the communities of plastics collected at all oceanic sites. Additionally, OTUs classified as Roseobacter, Pseudophaeobacter, Phaeobacter, Marinovum and Cognatiyoonia, also enriched in the LDPE-laboratory incubations, were enriched on LDPE communities compared to the ones associated to glass and polypropylene in in-situ incubations in the northern Adriatic Sea after 1 month of incubation. Some of these enriched OTUs were also related to known alkane and hydrocarbon degraders. Collectively, these results demonstrate that there are prokaryotes capable of surviving with LDPE as the sole carbon source living on plastics in relatively high abundances in different water masses of the global ocean.     

Adhesion of bacillus spores in relation to hydrophobicity

The adhesion of spores of five different Bacillus species to solid surfaces of different hydrophobicity was evaluated. The spore surface hydrophobicity was measured using hydrophobic interaction chromatography (HIC). A large variation in hydrophobicity was found among the spores of the different species tested. The degree of adhesion of spores to the solid surfaces was consistent with the results obtained using the HIC method. The most hydrophobic spores, according to the HIC method, adhered in a much larger extent to the hydrophobic surfaces. Furthermore, spores generally adhered to a greater extent to hydrophobic and hydrophilic surfaces than did the vegetative cells.     

Adhesion of bacillus spores in relation to hydrophobicity

R önner , U., H usmark , U. & H enriksson , A. 1990. Adhesion of bacillus spores in relation to hydrophobicity. Journal of Applied Bacteriology 69 , 550–556.
The adhesion of spores of five different Bacillus species to solid surfaces of different hydrophobicity was evaluated. The spore surface hydrophobicity was measured using hydrophobic interaction chromatography (HIC). A large variation in hydrophobicity was found among the spores of the different species tested. The degree of adhesion of spores to the solid surfaces was consistent with the results obtained using the HIC method. The most hydrophobic spores, according to the HIC method, adhered in a much larger extent to the hydrophobic surfaces. Furthermore, spores generally adhered to a greater extent to hydrophobic and hydrophilic surfaces than did the vegetative cells.     

Partitioning of temporal activity among desert lizards in relation to prey availability and temperature

Partitioning of activity time within ecological communities potentially reduces interspecific competition and increases the number of species that can coexist. We investigated temporal activity in a highly diverse lizard assemblage in the Simpson Desert, central Australia, to determine the degree of partitioning that occurs. Three periods were defined, daytime (sunrise to sunset), early night (sunset to midnight) and late night (midnight to sunrise), and live captures of lizards were tallied for each period during two sampling months (September and November 2007). We also quantified the activity times of potential invertebrate prey and measured ambient temperatures during the different time periods to investigate any associations between these factors and lizard activity. Some 77% of captures of 13 lizard species were made by day, with Ctenotus pantherinus, Egernia inornata (Scincidae) and Nephrurus levis (Gekkonidae) the only species showing extended nocturnal activity. Activity of both species of skink was recorded at temperatures 4°C lower than those for agamid and varanid lizards early in the night, and at temperatures as low as 18–20°C. Surface‐active invertebrates differed in composition between time periods and were less abundant during the late night period in the drier of the two sample months (September), but were distributed equally over time in the other month. Termites were active in subterranean galleries at night in September and mostly by day in November, but available at all times on surface/subsurface baits. We conclude that activity is distributed unevenly within this lizard assemblage, with partitioning facilitated by the ready availability of invertebrate prey and by lizards having relatively broad temperature tolerances that, in some cases, permit opportunistic exploitation of resources beyond usual times of activity.     

Molecular characterization of bacterial populations in petroleum-contaminated groundwater discharged from underground crude oil storage cavities

Petroleum-contaminated groundwater discharged from underground crude oil storage cavities (cavity groundwater) harbored more than 10(6) microorganisms ml(-1), a density 100 times higher than the densities in groundwater around the cavities (control groundwater). To characterize bacterial populations growing in the cavity groundwater, 46 PCR-amplified almost full-length 16S ribosomal DNA (rDNA) fragments were cloned and sequenced, and 28 different sequences were obtained. All of the sequences were affiliated with the Proteobacteria; 25 sequences (43 clones) were affiliated with the epsilon subclass, 2 were affiliated with the beta subclass, and 1 was affiliated with the delta subclass. Two major clusters (designated clusters 1 and 2) were found for the epsilon subclass proteobacterial clones; cluster 1 (25 clones) was most closely related to Thiomicrospira denitrificans (88% identical in nucleotide sequence), while cluster 2 (11 clones) was closely related to Arcobacter spp. Denaturing gradient gel electrophoresis (DGGE) of PCR-amplified partial 16S rDNA fragments showed that one band was detected most strongly in cavity groundwater profiles independent of storage oil type and season. The sequence of this major band was identical to the sequences of most of the cluster 1 clones. Fluorescence in situ hybridization (FISH) indicated that the cluster 1 population accounted for 12 to 24% of the total bacterial population. This phylotype was not detected in the control groundwater by DGGE and FISH analyses. These results indicate that the novel members of the epsilon subclass of the Proteobacteria grow as major populations in the petroleum-contaminated cavity groundwater.     

Surfactant-induced bacterial community changes correlated with increased polycyclic aromatic hydrocarbon degradation in contaminated soil

Bioremediation as a method for removing polycyclic aromatic hydrocarbons (PAHs) from contaminated environments has been criticized for poor removal of potentially carcinogenic but less bioavailable high molecular weight (HMW) compounds. As a partial remedy to this constraint, we studied surfactant addition at sub-micellar concentrations to contaminated soil to enhance the biodegradation of PAHs remaining after conventional aerobic bioremediation. We demonstrated increased removal of four- and five-ring PAHs using two nonionic surfactants, polyoxyethylene(4)lauryl ether (Brij 30) and polyoxyethylene sorbitol hexaoleate (POESH), and analyzed bacterial community shifts associated with those conditions. Eight groups of abundant bacteria were implicated as potentially being involved in increased HMW PAH removal. A group of unclassified Alphaproteobacteria and members of the Phenylobacterium genus in particular showed significantly increased relative abundance in the two conditions exhibiting increased PAH removal. Other implicated groups included members of the Sediminibacterium, Terrimonas, Acidovorax, and Luteimonas genera, as well as uncharacterized organisms within the families Chitinophagaceae and Bradyrhizobiaceae. Targeted isolation identified a subset of the community likely using the surfactants as a growth substrate, but few of the isolates exhibited PAH-degradation capability. Isolates recovered from the Acidovorax and uncharacterized Bradyrhizobiaceae groups suggest the abundance of those groups may have been attributable to growth on surfactants. Understanding the specific bacteria responsible for HMW PAH removal in natural and engineered systems and their response to stimuli such as surfactant amendment may improve bioremediation efficacy during treatment of contaminated environmental media.     

Predictable bacterial composition and hydrocarbon degradation in Arctic soils following diesel and nutrient disturbance

Increased exploration and exploitation of resources in the Arctic is leading to a higher risk of petroleum contamination. A number of Arctic microorganisms can use petroleum for growth-supporting carbon and energy, but traditional approaches for stimulating these microorganisms (for example, nutrient addition) have varied in effectiveness between sites. Consistent environmental controls on microbial community response to disturbance from petroleum contaminants and nutrient amendments across Arctic soils have not been identified, nor is it known whether specific taxa are universally associated with efficient bioremediation. In this study, we contaminated 18 Arctic soils with diesel and treated subsamples of each with monoammonium phosphate (MAP), which has successfully stimulated degradation in some contaminated Arctic soils. Bacterial community composition of uncontaminated, diesel-contaminated and diesel+MAP soils was assessed through multiplexed 16S (ribosomal RNA) rRNA gene sequencing on an Ion Torrent Personal Genome Machine, while hydrocarbon degradation was measured by gas chromatography analysis. Diversity of 16S rRNA gene sequences was reduced by diesel, and more so by the combination of diesel and MAP. Actinobacteria dominated uncontaminated soils with <10% organic matter, while Proteobacteria dominated higher-organic matter soils, and this pattern was exaggerated following disturbance. Degradation with and without MAP was predictable by initial bacterial diversity and the abundance of specific assemblages of Betaproteobacteria, respectively. High Betaproteobacteria abundance was positively correlated with high diesel degradation in MAP-treated soils, suggesting this may be an important group to stimulate. The predictability with which bacterial communities respond to these disturbances suggests that costly and time-consuming contaminated site assessments may not be necessary in the future.     

Effects of Pinus sylvestris root growth and mycorrhizosphere development on bacterial carbon source utilization and hydrocarbon oxidation in forest and petroleum-contaminated soils

The hypothesis that Pinus sylvestris L. root and mycorrhizosphere development positively influences bacterial community-linked carbon source utilization, and drives a concomitant reduction in mineral oil levels in a petroleum hydrocarbon- (PHC-) contaminated soil was confirmed in a forest ecosystem-based phytoremediation simulation. Seedlings were grown for 9 months in large petri dish microcosms containing either forest humus or humus amended with cores of PHC-contaminated soil. Except for increased root biomass in the humus/PHC treatment, there were no other significant treatment-related differences in plant growth and needle C and N status. Total cell and culturable bacterial (CFU) densities significantly increased in both rhizospheres and mycorrhizospheres that actively developed in the humus and PHC-contaminated soil. Mycorrhizospheres (mycorrhizas and extramatrical mycelium) supported the highest numbers of bacteria. Multivariate analyses of bacterial community carbon source utilization profiles (Biolog GN microplate) from different rhizosphere, mycorrhizosphere, and bulk soil compartments, involving principal component and correspondence analysis, highlighted three main niche-related groupings. The respective clusters identified contained bacterial communities from (i) unplanted bulk soils, (ii) planted bulk PHC and rhizospheres in PHC-contaminated soils, and (iii) planted bulk humus and rhizosphere/mycorrhizosphere-influenced humus, and mycorrhizosphere-influenced PHC contaminated soil. Correspondence analysis allowed further identification of amino acid preferences and increased carboxylic/organic acid preferences in rhizosphere and mycorrhizosphere compartments. Decreased levels of mineral oil (non-polar hydrocarbons) were detected in the PHC-contaminated soil colonized by pine roots and mycorrhizal fungi. These data further support our view that mycorrhizosphere development and function plays a central role in controlling associated bacterial communities and their degradative activities in lignin-rich forest humus and PHC-contaminated soils.     

Pollen in aeolian dust with relation to allergy and asthma in Kuwait

The studying of spatial and temporal distribution of pollen is essential, but rarely tackled in the region including Kuwait. Pollen grains and aeolian dust in the state of Kuwait were seasonally monitored from August 2009 to August 2011 to identify and correlate spatially and temporally pollen data with human health asthma and allergy data. The year 2010–2011 yielded a high in deposited dust and pollen compared to 2009–2010 by 33.2 and 9.9%, respectively. The spring (April–May) and the autumn (October–November) seasons marked the most top pollen distribution for Chenopodiaceae, Gramineae (Poaceae), Cyperaceae, Fabaceae, and Plantaginaceae. The pollen graphs for summer (July–August) and winter (January–February) seasons showed low pollen distribution due to the severe drought, dust storms, winter wind and rainfall. There are no correlations found between pollen deposition and the number of asthma patients. The annual dust deposited rate in Kuwait during 2010/2011 was 373 t km^?2 which represented a 33.2% increase compared with 2009/2010. The study has concluded that pollen of Malvaceae, Compositae and Chenopodiaceae is the most common triggers of allergy in Kuwait. Additionally, a positive correlation between the number of allergy patients and deposited dust was observed during March 2010 and 2011.     

Pheromones in relation to aggregation and reproduction in desert locusts

Abstract. Desert locusts, Schistocerca gregaria (Forskål) (Orthoptera: Acrididae), exhibit a population density-dependent phase polymorphism which includes the gradual change of many morphological, physiological and behavioural characteristics. Many volatiles associated with desert locusts have been identified recently and it is assumed that they are involved in pheromonal control of behaviour and development of locusts. Ovipositing females deposit with their egg pods several volatiles that appear to be attractive to other females resulting – possibly in combination with environmental factors – in an aggregated oviposition. Mature males release several volatiles, among them phenylacetonitrile, which are reported to accelerate sexual maturation in young males. Also, aggregation pheromone systems for hoppers and adults have been described. However, recent studies and publications shed a new light on the postulated effects of some of these volatiles. Gregarious behaviour can undoubtedly be induced by mechanical stimuli. Furthermore, the main component of the adult aggregation pheromone system, phenylacetonitrile, is found to be a repellent obviously not involved in aggregation. Comprehensive studies have demonstrated that phenylacetonitrile is used by mature gregarious males as a courtship inhibition pheromone to enhance mate guarding. Recent progress, contradictory results and perspectives in desert locust pheromone research related to reproduction are summarized and discussed in this paper.     

Role of hydrophobicity in bacterial adherence to carbon nanostructures and biofilm formation

The role of cell and surface hydrophobicity in the adherence of the waterborne bacterium Mycobacterium smegmatis to nanostructures and biofilm formation was investigated. Carbon nanostructures (CNs) were synthesized using a flame reactor and deposited on stainless steel grids and foils, and on silicon wafers that had different initial surface hydrophobicities. Surface hydrophobicity was measured as the contact angle of water droplets. The surfaces were incubated in suspensions of isogenic hydrophobic and hydrophilic strains of M. smegmatis and temporal measurements of the numbers of adherent cells were made. The hydrophobic, rough mutant of M. smegmatis adhered more readily and formed denser biofilms on all surfaces compared to its hydrophilic, smooth parent. Biofilm formation led to alterations in the hydrophobicity of the substratum surfaces, demonstrating that bacterial cells attached to CNs are capable of modifying the surface characteristics.