Effects of Hydrophobic and Electrostatic Cell Surface Properties of Bacteria on Feeding Rates of Heterotrophic Nanoflagellates

The influence of cell surface hydrophobicity and electrostatic charge of bacteria on grazing rates of three common species of interception-feeding nanoflagellates was examined. The hydrophobicity of bacteria isolated from freshwater plankton was assessed by using two different methods (bacterial adhesion to hydrocarbon and hydrophobic interaction chromatography). The electrostatic charge of the cell surface (measured as zeta potential) was analyzed by microelectrophoresis. Bacterial ingestion rates were determined by enumerating bacteria in food vacuoles by immunofluorescence labelling via strain-specific antibodies. Feeding rates varied about twofold for each flagellate species but showed no significant dependence on prey hydrophobicity or surface charge. Further evidence was provided by an experiment involving flagellate grazing on complex bacterial communities in a two-stage continuous culture system. The hydrophobicity values of bacteria that survived protozoan grazing were variable, but the bacteria did not tend to become more hydrophilic. We concluded that variability in bacterial cell hydrophobicity and variability in surface charge do not severely affect uptake rates of suspended bacteria or food selection by interception-feeding flagellates.     

Effects of hydrophobic and electrostatic cell surface properties of bacteria on feeding rates of heterotrophic nanoflagellates

The influence of cell surface hydrophobicity and electrostatic charge of bacteria on grazing rates of three common species of interception-feeding nanoflagellates was examined. The hydrophobicity of bacteria isolated from freshwater plankton was assessed by using two different methods (bacterial adhesion to hydrocarbon and hydrophobic interaction chromatography). The electrostatic charge of the cell surface (measured as zeta potential) was analyzed by microelectrophoresis. Bacterial ingestion rates were determined by enumerating bacteria in food vacuoles by immunofluorescence labelling via strain-specific antibodies. Feeding rates varied about twofold for each flagellate species but showed no significant dependence on prey hydrophobicity or surface charge. Further evidence was provided by an experiment involving flagellate grazing on complex bacterial communities in a two-stage continuous culture system. The hydrophobicity values of bacteria that survived protozoan grazing were variable, but the bacteria did not tend to become more hydrophilic. We concluded that variability in bacterial cell hydrophobicity and variability in surface charge do not severely affect uptake rates of suspended bacteria or food selection by interception-feeding flagellates.     

Effect of bile on the lipid composition and surface properties of bifidobacteria

AIM: The changes produced on the bacterial surface of Bifidobacteria cells when they are grown in bile were compared with those provoked by bile added to bacteria grown in the absence of bile. METHODS AND RESULTS: The adhesive properties, the zeta potential and the lipid composition of Bifidobacterial strains, isolated from human faeces and grown in MRS medium, were determined. Bacteria grown in MRS with bile showed a loss of adherence and autoaggregation in correlation with a decrease in the surface hydrophobicity in comparison to those grown in MRS without bile, concomitant with the absence of two glycolipids, the increase of sugar content and minor changes in fatty acid composition. The surface changes caused by bile shock on bacteria grown in bile-free medium were much less pronounced and, in addition, no effect on the lipid composition was apparent. CONCLUSIONS: The comparison of the results indicates that bile action on surface properties is related to metabolic changes. SIGNIFICANCE AND IMPACT OF THE STUDY: Long-term exposure of bacteria to bile may cause metabolic changes affecting their adhesive properties irreversibly. This may be taken as a criterion to define the probiotic properties of different strains.     

Substrate Utilization Profiles of Bacterial Strains in Plankton from the River Warnow, a Humic and Eutrophic River in North Germany

Bacteria are very important degraders of organic substances in aquatic environments. Despite their influential role in the carbon (and many other element) cycle(s), the specific genetic identity of active bacteria is mostly unknown, although contributing phylogenetic groups had been investigated. Moreover, the degree to which phenotypic potential (i.e., utilization of environmentally relevant carbon substrates) is related to the genomic identity of bacteria or bacterial groups is unclear. The present study compared the genomic fingerprints of 27 bacterial isolates from the humic River Warnow with their ability to utilize 14 environmentally relevant substrates. Acetate was the only substrate utilized by all bacterial strains. Only 60% of the strains respired glucose, but this substrate always stimulated the highest bacterial activity (respiration and growth). Two isolates, both closely related to the same Pseudomonas sp., also had very similar substrate utilization patterns. However, similar substrate utilization profiles commonly belonged to genetically different strains (e.g., the substrate profile of Janthinobacterium lividum OW6/RT-3 and Flavobacterium sp. OW3/15-5 differed by only three substrates). Substrate consumption was sometimes totally different for genetically related isolates. Thus, the genomic profiles of bacterial strains were not congruent with their different substrate utilization profiles. Additionally, changes in pre-incubation conditions strongly influenced substrate utilization. Therefore, it is problematic to infer substrate utilization and especially microbial dissolved organic matter transformation in aquatic systems from bacterial molecular taxonomy.     

Bacterial migration along solid surfaces.

An in vitro system was developed to study the migration of uropathogenic Escherichia coli strains. In this system an aqueous agar gel is placed against a solid surface, allowing the bacteria to migrate along the gel/solid surface interface. Bacterial strains as well as solid surfaces were characterized by means of water contact angle and zeta potential measurements. When glass was used as the solid surface, significantly different migration times for the strains investigated were observed. Relationships among the observed migration times of six strains, their contact angles, and their zeta potentials were found. Relatively hydrophobic strains exhibited migration times shorter than those of hydrophilic strains. For highly negatively charged strains shorter migration times were found than were found for less negatively charged strains. When the fastest-migrating strain with respect to glass was allowed to migrate along solid surfaces differing in hydrophobicity and charge, no differences in migration times were found. Our findings indicate that strategies to prevent catheter-associated bacteriuria should be based on inhibition of bacterial growth rather than on modifying the physicochemical character of the catheter surface.     

Bacterial migration along solid surfaces.

An in vitro system was developed to study the migration of uropathogenic Escherichia coli strains. In this system an aqueous agar gel is placed against a solid surface, allowing the bacteria to migrate along the gel/solid surface interface. Bacterial strains as well as solid surfaces were characterized by means of water contact angle and zeta potential measurements. When glass was used as the solid surface, significantly different migration times for the strains investigated were observed. Relationships among the observed migration times of six strains, their contact angles, and their zeta potentials were found. Relatively hydrophobic strains exhibited migration times shorter than those of hydrophilic strains. For highly negatively charged strains shorter migration times were found than were found for less negatively charged strains. When the fastest-migrating strain with respect to glass was allowed to migrate along solid surfaces differing in hydrophobicity and charge, no differences in migration times were found. Our findings indicate that strategies to prevent catheter-associated bacteriuria should be based on inhibition of bacterial growth rather than on modifying the physicochemical character of the catheter surface.     

Effect of direct electric current on the cell surface properties of phenol-degrading bacteria

The change in cell surface properties in the presence of electric currents is of critical concern when the potential to manipulate bacterial movement with electric fields is evaluated. In this study, the effects of different direct electric currents on the cell surface properties involved in bacterial adhesion were investigated by using a mixed phenol-degrading bacterial culture in the exponential growth phase. The traits investigated were surface hydrophobicity (measured by adherence to n-octane), net surface electrostatic charge (determined by measurement of the zeta potential), and the cell surface shape and polymers (determined by scanning electron microscope analysis). The results showed that a lower current (less than 20 mA) induced no significant changes in the surface properties of phenol-degrading bacteria, that an electric current of 20 mA could increase the surface hydrophobicity and flatten the cell shape, and that a higher current (40 mA) could increase the surface extracellular substances and the net negative surface electrostatic charge. The results also revealed that the electric current effects on cell hydrophobicity varied with the suspending medium. We suggest that an electric current greater than 20 mA is not suitable for use in manipulation of the movement of the phenol-degrading bacteria, although such a current might favor the electrophoretic movement of the bacterial species.     

The effects of root surface charge and nitrogen forms on the adsorption of aluminum ions by the roots of rice with different aluminum tolerances

Aim

Our objectives were to compare effects of root charge properties on Al adsorption by the roots of rice that differed in Al-tolerance, and to examine effects of different nitrogen forms on charge properties of rice roots and Al adsorption.

Methods

Streaming potential and chemical methods were used to measure root zeta potential and investigate Al chemical forms adsorbed on the roots of rice obtained from solution culture experiments.

Results

Rice roots of the Al-sensitive variety Yangdao-6 carried greater negative charge than the Al-tolerant variety Wuyunjing-7, which meant the roots of Yangdao-6 adsorbed more exchangeable and complexed Al. When both rice varieties were grown in NH₄ ⁺-containing nutrient solutions, there were less functional groups and lower negative surface charge on their roots, which reduced Al adsorption compared to the rice grown in NO₃ ^? containing nutrient solutions. The decline in nutrient solution pH due to NH₄ ⁺ uptake by rice roots was responsible for the reduced numbers of functional groups and the lower negative surface charge on the roots compared to the rice grown in NO₃ ^? containing solutions.

Conclusions

Integrated root surface charge, as expressed by zeta potential, played an important role in Al adsorption by the roots of rice with different Al-tolerance.

     

Effect of Direct Electric Current on the Cell Surface Properties of Phenol-Degrading Bacteria

The change in cell surface properties in the presence of electric currents is of critical concern when the potential to manipulate bacterial movement with electric fields is evaluated. In this study, the effects of different direct electric currents on the cell surface properties involved in bacterial adhesion were investigated by using a mixed phenol-degrading bacterial culture in the exponential growth phase. The traits investigated were surface hydrophobicity (measured by adherence to n-octane), net surface electrostatic charge (determined by measurement of the zeta potential), and the cell surface shape and polymers (determined by scanning electron microscope analysis). The results showed that a lower current (less than 20 mA) induced no significant changes in the surface properties of phenol-degrading bacteria, that an electric current of 20 mA could increase the surface hydrophobicity and flatten the cell shape, and that a higher current (40 mA) could increase the surface extracellular substances and the net negative surface electrostatic charge. The results also revealed that the electric current effects on cell hydrophobicity varied with the suspending medium. We suggest that an electric current greater than 20 mA is not suitable for use in manipulation of the movement of the phenol-degrading bacteria, although such a current might favor the electrophoretic movement of the bacterial species.     

Hydrophobic and electrostatic cell surface properties of Cryptosporidium parvum.

Microbial adhesion to hydrocarbons and microelectrophoresis were investigated in order to characterize the surface properties of Cryptosporidium parvum. Oocysts exhibited low removal rates by octane (only 20% on average), suggesting that the Cryptosporidium sp. does not demonstrate marked hydrophobic properties. A zeta potential close to -25 mV at pH 6 to 6.5 in deionized water was observed for the parasite. Measurements of hydrophobicity and zeta potential were performed as a function of pH and ionic strength or conductivity. Hydrophobicity maxima were observed at extreme pH values, with 40% of adhesion of oocysts to octane. It also appeared that ionic strength (estimated by conductivity) could influence the hydrophobic properties of oocysts. Cryptosporidium oocysts showed a pH-dependent surface charge, with zeta potentials becoming less negative as pH was reduced, starting at -35 mV for alkaline pH and reaching 0 at isoelectric points for pH 2.5. On the other hand, variation of surface charge with respect to conductivity of the suspension tested in this work was quite small. The knowledge of hydrophobic properties and surface charge of the parasite provides information useful in, for example, the choice of various flocculation treatments, membrane filters, and cleaning agents in connection with oocyst recovery.     

Effect of fluorochromes on bacterial surface properties and interaction with granular media.

Simple, efficient, and safe tagging methods are desired in short-term microbial transport studies such as in the study of filtration systems for water and wastewater treatment. Suitability of selected fluorochromes as bacterial tagging agents in transport studies was evaluated on the basis of stability of stained cells and the effect of staining on bacterial surface characteristics and interaction with granular media. Surface properties were characterized by zeta potential and microbial adhesion to hydrocarbons. The effect of staining on interactions between bacteria and porous media was evaluated in terms of removal of bacteria in batch adsorption tests using sand coated with aluminum hydroxide to enhance adsorption. The DNA-specific fluorochrome 4',6-diamidino-2-phenylindole (DAPI) had generally negligible effects on bacterial surface properties and interaction with sand, as indicated in batch adsorption tests using pure cultures (Escherichia coli or Acinetobacter sp.) and wastewater bacteria. Cells stained with DAPI were stable for 48 h at 4 or 20 degrees C. Other nucleic acid fluorochromes tested had different but significant effects on bacterial cells and produced less stable fluorescence. Since transport through porous media is modulated by surface properties, it may be concluded based on these results that the choice of fluorochromes is critical in microbial transport studies. DAPI appeared to be a promising tagging agent. Time dependence of fluorescence of stained cells may limit the use of fluorochrome-tagged cells in long-term transport studies.     

Retention in Treated Wastewater Affects Survival and Deposition of Staphylococcus aureus and Escherichia coli in Sand Columns

The fate and transport of pathogenic bacteria from wastewater treatment facilities in the Earth''s subsurface have attracted extensive concern over recent decades, while the impact of treated-wastewater chemistry on bacterial viability and transport behavior remains unclear. The influence of retention time in effluent from a full-scale municipal wastewater treatment plant on the survival and deposition of Staphylococcus aureus and Escherichia coli strains in sand columns was investigated in this paper. In comparison to the bacteria cultivated in nutrient-rich growth media, retention in treated wastewater significantly reduced the viability of all strains. Bacterial surface properties, e.g., zeta potential, hydrophobicity, and surface charges, varied dramatically in treated wastewater, though no universal trend was found for different strains. Retention in treated wastewater effluent resulted in changes in bacterial deposition in sand columns. Longer retention periods in treated wastewater decreased bacterial deposition rates for the strains evaluated and elevated the transport potential in sand columns. We suggest that the wastewater quality should be taken into account in estimating the fate of pathogenic bacteria discharged from wastewater treatment facilities and the risks they pose in the aquatic environment.     

Surface charge influences enterococcal prevalence in mixed-species biofilms

AIM: To study the influence of 15 microbial isolates on the prevalence of charge-heterogeneous and charge-homogeneous Enterococcus faecalis strains, all isolated from biliary stents, in mixed-species biofilms. METHODS AND RESULTS: Six Enterococcus faecalis strains were paired with 15 other microbial isolates to form mixed-species biofilms in a microtitre plate assay. Charge-heterogeneous Enterococcus faecalis strains display two subpopulations with different surface charges, expressed as a biomodal zeta potential distribution. It was found that, in general, the prevalence of the charge-heterogeneous, biofilm forming Enterococcus faecalis was reduced in mixed-species biofilms. The prevalence of charge-homogeneous, nonbiofilm-forming Enterococcus faecalis strains was increased only in the presence of Citrobacter freundii BS5126, Stenotrophomonas malthophilia BS937, and Candida lusitaniae BS8256, all of which introduced sizeable charge heterogeneity in the mixed microbial population. CONCLUSIONS: Charge-homogeneous Enterococcus faecalis strains are stimulated to form biofilm only by the presence of another microbial species with a considerably less negative zeta potential, thereby creating a charge-heterogeneous microbial population. SIGNIFICANCE AND IMPACT OF THE STUDY: Enterococcus faecalis is one of the predominant species isolated from mixed-species biofilms in clogged biliary stents. The current study shows how charge-homogeneous Enterococcus faecalis strains form more biofilm in the presence of other microbial species.     

Influence of culture heterogeneity in cell surface charge on adhesion and biofilm formation by Enterococcus faecalis

Biofilm formation is an increasing problem in medicine, due to the intrinsic resistance of microorganisms in the biofilm mode of growth against the host immune system and antimicrobial therapy. Adhesion is an important step in biofilm formation, influenced, among other factors, by the surface hydrophobicities and charges of both the substratum and the adhering microorganisms. Enterococcus faecalis strains generally display subpopulations with different surface charges, expressed as bimodal zeta potential distributions. Two-thirds of E. faecalis strains isolated from clogged biliary stents displayed such heterogeneity of surface charges in culture. In this study, the influence of this culture heterogeneity on initial adhesion and subsequent biofilm formation was investigated. Heterogeneous strains were retained in higher numbers on polystyrene than homogeneous strains. Also, biofilm formation was much more pronounced for heterogeneous strains than for homogeneous strains. In a population enriched to display only one subpopulation, fewer bacteria were retained than in its original heterogeneous culture. Also, the enriched subpopulation formed less biofilm than its original heterogeneous culture. The presence of ox bile during adhesion resulted in fewer retained bacteria, although heterogeneous strains were still retained in significantly higher numbers than were homogeneous strains, and, in general, the presence of ox bile reduced biofilm formation. The initial adhesion and biofilm formation were independent of the presence of the gene encoding the enterococcal surface protein (esp) or the expression of gelatinase (GelE). It is concluded that heterogeneity in cell surface charge represents an advantage for bacteria in the colonization of surfaces.     

Physicochemical characterization of Escherichia coli. A comparison with gram-positive bacteria.

Eight Escherichia coli strains were characterized by determining their adhesion to xylene, surface free energy, zeta potential, relative surface charge, and their chemical composition. The latter was done by applying X-ray photoelectron spectroscopy (XPS) and infrared spectroscopy (IR). No relationship between the adhesion to xylene and the water contact angles of these strains was found. Three strains had significantly lower surface free energies than the other strains. Surface free energies were either obtained from polar and dispersion parts or from Lifshitz-van der Waals and acid/base parts of the surface free energy. A correlation (r = 0.97) between the polar parts and the electron-donor contributions to the acid/base part of the surface free energy was found. The zeta potentials of all strains, measured as a function of pH (2-11), were negative. Depending on the zeta potential as a function of pH, three groups were recognized among the strains tested. A relationship (r = 0.84) was found between the acid/base component of the surface free energy and the zeta potential measured at pH = 7.4. There was no correlation between results of XPS and IR studies. Data from the literature of XPS and IR studies of the gram-positive staphylococci and streptococci were compared with data from the gram-negative E. coli used in this study. It appeared that in these three groups of bacteria, the polysaccharide content detected by IR corresponded well with the oxygen-to-carbon ratio detected by XPS.     

Nanofibrous microfiltration membrane based on cellulose nanowhiskers

A multilayered nanofibrous microfiltration (MF) membrane system with high flux, low pressure drop, and high retention capability against both bacteria and bacteriophages (a virus model) was developed by impregnating ultrafine cellulose nanowhiskers (diameter about 5 nm) into an electrospun polyacrylonitrile (PAN) nanofibrous scaffold (fiber diameter about 150 nm) supported by a poly(ethylene terephthalate) (PET) nonwoven substrate (fiber diameter about 20 μm). The cellulose nanowhiskers were anchored on the PAN nanofiber surface, forming a cross-linked nanostructured mesh with very high surface-to-volume ratio and a negatively charged surface. The mean pore size and pore size distribution of this MF system could be adjusted by the loading of cellulose nanowhiskers, where the resulting membrane not only possessed good mechanical properties but also high surface charge density confirmed by the conductivity titration and zeta potential measurements. The results indicated that a test cellulose nanowhisker-based MF membrane exhibited 16 times higher adsorption capacity against a positively charged dye over a commercial nitrocellulose-based MF membrane. This experimental membrane also showed full retention capability against bacteria, for example, E. coli and B. diminuta (log reduction value (LRV) larger than 6) and decent retention against bacteriophage MS2 (LRV larger than 2).     

Physicochemical characterization ofEscherichia coli

EightEscherichia coli strains were characterized by determining their adhesion to xylene, surface free energy, zeta potential, relative surface charge, and their chemical composition. The latter was done by applying X-ray photoelectron spectroscopy (XPS) and infrared spectroscopy (IR). No relationship between the adhesion to xylene and the water contact angles of these strains was found. Three strains had significantly lower surface free energies than the other strains. Surface free energies were either obtained from polar and dispersion parts or from Lifshitz-van der Waals and acid/base parts of the surface free energy. A correlation (r=0.97) between the polar parts and the electron-donor contributions to the acid/base part of the surface free energy was found. The zeta potentials of all strains, measured as a function of pH (2–11), were negative. Depending on the zeta potential as a function of pH, three groups were recognized among the strains tested. A relationship (r=0.84) was found between the acid/base component of the surface free energy and the zeta potential measured at pH=7.4. There was no correlation between results of XPS and IR studies. Data from the literature of XPS and IR studies of the gram-positive staphylococci and streptococci were compared with data from the gram-negativeE. coli used in this study. It appeared that in these three groups of bacteria, the polysaccharide content detected by IR corresponded well with the oxygen-to-carbon ratio detected by XPS.     

Effects of Inorganic Particles on Metabolism by a Periphytic Marine Bacterium

Measurements were made of adsorption of a periphytic marine bacterium, glucose, and glutamic acid to inorganic particles in seawater and defined bacterial growth medium. Measurements of the metabolism of bacteria were made in the presence and absence of particles by microcalorimetry and radiorespirometry. It was found that hydroxyapatite adsorbs glutamic acid, but not glucose, from the experimental medium. It was also found that hydroxyapatite adsorbs essentially all of the bacteria from the medium when the bacterial concentration is approximately 6 × 10⁵ bacteria per ml. If the bacterial concentration is approximately 6 × 10⁷, then only a small fraction of cells become attached. It was therefore possible to select bacterial concentrations and organic nutrients so that bacterial attachment, organic nutrient adsorption, or both would occur in different experiments. In this experimental system the metabolism by attached and nonattached bacteria of adsorbing and nonadsorbing organic nutrients was measured. The results show that bacterial activity in this model system was not enhanced by the particles, regardless of whether the bacteria, the organic nutrient, or both were associated with the surface. In fact, the respiratory activity of the attached bacteria was diminished in comparison with that of free bacteria.     

Relationship between Cell Surface Properties and Transport of Bacteria through Soil

A study was conducted to relate the properties of Enterobacter, Pseudomonas, Bacillus, Achromobacter, Flavobacterium, and Arthrobacter strains to their transport with water moving through soil. The bacteria differed markedly in their extent of transport; their hydrophobicity, as measured by adherence to n-octane and by hydrophobic-interaction chromatography; and their net surface electrostatic charge, as determined by electrostatic interaction chromatography and by measurements of the zeta potential. Transport of the 19 strains through Kendaia loam or their retention by this soil was not correlated with hydrophobicities or net surface charges of the cells or the presence of capsules. Among 10 strains tested, the presence of flagella was also not correlated with transport. Retention was statistically related to cell size, with bacteria shorter than 1.0 μm usually showing higher percentages of cells being transported through the soil. We suggest that more than one characteristic of bacterial cells determines whether the organisms are transported through soil with moving water.     

THE MODIFICATION OF ANTIBODIES BY FORMALDEHYDE

Certain strains of bacteria which have only minimal zeta potentials over a wide range of pH, and upon which surface deposits can be formed, afford a favorable means of studying certain chemical and physical properties of the surface deposits. Films of specific antibody-globulin upon these bacteria possess basic groups which can combine with formaldehyde. Combination of these groups with HCHO under the conditions of the present experiments shifts the isoelectric point of the sensitizing film toward the acid side by about 0.6 to 0.8 pH unit, and reduces the agglutinating tendency of the sensitizing film. Antibodies may be formalinized before combination with antigen without marked change in their specific combining affinities. The properties of the sensitizing films are similar whether formol treatment occurs before or after the antigen-antibody combination. The nature of the basic groups has been discussed.