Changes in the surface potential of Lactobacillus acidophilus under freeze-thawing stress

The zeta potential of Lactobacillus acidophilus CRL 640, a measure of the net distribution of electrical charges on the bacterial surface, is a function of the glucose concentration in the growing media. With 2% glucose, cells in the stationary phase showed a zeta potential of -45 +/- 2 mV. With these cells, the zeta potential after freezing and thawing decreased to -32 +/- 2 mV and there was a decrease in viability. The changes in the surface potential correlated with damage to the cell surface as shown by electron microscopy. Freeze-thawed cells incubated in a rich medium recovered a zeta potential of -38 +/- 2 mV without cell growth. L. acidophilus CRL 640 showed the same value of surface potential as control cells when they were frozen and thawed in 2 M glycerol.     

The role of electro-osmosis in the electric-field-induced movement of charged macromolecules on the surfaces of cells.

The surfaces of most cells bear a net negative charge. The imposition of an electric field parallel to the surface of the cell should produce, therefore, an electro-osmotic flow of fluid towards the cathodal side of the cell. Our analysis of a simple model of the cell surface indicates that a negatively charged mobile macromolecule will be swept by this electro-osmotic flow of fluid to the cathodal side of the cell if its zeta potential, zeta 1, is less negative than the zeta potential of the cell surface, zeta 2. Conversely, if zeta 2 is less negative than zeta 1, the negatively charged macromolecule will accumulate at the anodal side of the cell. Our experimental results demonstrate that concanavalin A (Con A) receptors on embryonic muscle cells normally accumulate at the cathodal side of the cell, but that they can be induced to accumulate at the anodal side of the cell by preincubating the myotubes either with neuraminidase, a treatment that removes negatively charged sialic acid residues, or with the lipid diI, a treatment that adds positive charges to the surface of the cell. Addition of the negatively charged lipid monosialoganglioside (GM1), on the other hand, enhances the accumulation of Con A receptors at the cathodal side of the cell.     

Experimental and functional changes in transmembrane potential and zeta potential of single cultured cells

Experiments were performed on single cells to investigate the relations between the total bioelectrical potential difference (PD) across the cell membrane (so-called transmembrane potential) and the net negative surface charge of the cell (zeta potential). The experiments were carried out on FL-cells, leucocytes and ovarian tumour cells. The PD was measured electrophysiologically by means of intracellular glass microelectrodes; the surface charge or the zeta potential was determined using cell electrophoresis. Both measuring methods are critically discussed.Under different conditions (hypothermia, hyperthermia, mitotic blocking agent, cell cycle), the transmembrane potential and zeta potential showed changes in an identical direction and often the response of transmembrane potential was found to be quicker and more intensive than that of the zeta potential. In other experiments (e.g. changing the extracellular Cl^? ion concentration) the reactions of both potentials showed no coincidence. Depending on the type of functionally or experimentally borne changes on the cytoplasmatic membrane, either both potentials or only one of them may be altered.     

Determination of zeta potential in Planctomycetes and its application in heavy metals toxicity assessment

Zeta potential of Planctomycetes was evaluated under different environmental conditions and correlated to cell viability. Phylogenetically distinct strains of the Planctomycetes presented different negative zeta potential values. More negative values were associated with Rhodopirellula spp. and related to the great amount of fimbriae in these species. Milli-Q water was chosen as the best dispersion media to perform the measurements. Zeta potential increased with ionic strength and varied with pH. In the physiological range of pH 5.0-9, zeta potential remained low and Rhodopirellula sp. strain LF2 cells were viable. Out of this range, zeta potential increased significantly and viability decreased. The effect on zeta potential of arsenic, cadmium, chromium, copper, lead, nickel, and zinc was assessed in Rhodopirellula sp. strain LF2. Zeta potential increased with increasing toxicity of the heavy metals in a dose-response way. This result was confirmed by the results observed for Rhodopirellula baltica strain SH1 under copper toxicity. Lead was the most toxic metal and zinc was the least toxic as observed by zeta potential and viability. The results support a correlation between zeta potential and cell viability which seem to indicate the possibility to use it as a viability predictor for the effects of heavy metals toxicity.     

Optical tweezers as a new biomedical tool to measure zeta potential of stored red blood cells

During storage, red blood cells (RBCs) for transfusion purposes suffer progressive deterioration. Sialylated glycoproteins of the RBC membrane are responsible for a negatively charged surface which creates a repulsive electrical zeta potential. These charges help prevent the interaction between RBCs and other cells, and especially among each RBCs. Reports in the literature have stated that RBCs sialylated glycoproteins can be sensitive to enzymes released by leukocyte degranulation. Thus, the aim of this study was, by using an optical tweezers as a biomedical tool, to measure the zeta potential in standard RBCs units and in leukocyte reduced RBC units (collected in CPD-SAGM) during storage. Optical tweezers is a sensitive tool that uses light for measuring cell biophysical properties which are important for clinical and research purposes. This is the first study to analyze RBCs membrane charges during storage. In addition, we herein also measured the elasticity of RBCs also collected in CPD-SAGM. In conclusion, the zeta potential decreased 42% and cells were 134% less deformable at the end of storage. The zeta potential from leukodepleted units had a similar profile when compared to units stored without leukoreduction, indicating that leukocyte lyses were not responsible for the zeta potential decay. Flow cytometry measurements of reactive oxygen species suggested that this decay is due to membrane oxidative damages. These results show that measurements of zeta potentials provide new insights about RBCs storage lesion for transfusion purposes.     

Extracellular polymeric substances responsible for bacterial adhesion onto solid surface

The influence of extracellular polymeric substances (EPS) on bacterial cell adhesion onto solid surfaces was investigated using 27 heterotrophic bacterial strains isolated from a wastewater treatment reactor. Cell adhesion onto glass beads was carried out by the packed-bed method and the results were discussed in terms of the amount of each EPS component produced and cell surface characteristics such as zeta potential and hydrophobicity. Protein and polysaccharides accounted for 75-89% of the EPS composition, indicating that they are the major EPS components. Among the polysaccharides, the amounts of hexose, hexosamine and ketose were relatively high in EPS-rich strains. For EPS-poor strains, the efficiency of cell adhesion onto glass beads increased as the absolute values of zeta potential decreased, suggesting that electrostatic interaction suppresses cell adhesion efficiency. On the other hand, the amounts of hexose and pentose exhibited good correlations with cell adhesiveness for EPS-rich strains, indicating that polymeric interaction due to the EPS covering on the cell surface promoted cell adhesion. It was concluded that, if the EPS amount is relatively small, cell adhesion onto solid surfaces is inhibited by electrostatic interaction, and if it is relatively large, cell adhesion is enhanced by polymeric interaction.     

Architectural changes in the TCR:CD3 complex induced by MHC:peptide ligation

A hallmark of T cell activation is the ligation-induced down-modulation of the TCR:CD3 complex. However, little is known about the molecular events that drive this process. The CD3 zeta-chain has been shown to play a unique role in regulating the assembly, transport, and cell surface expression of the TCR:CD3 complex. In this study we have investigated the relationship between CD3zeta and the TCRalphabetaCD3epsilondeltagamma complex after ligation by MHC:peptide complexes. Our results show that there is a significant increase in free surface CD3zeta, which is not associated with the TCR:CD3 complex, after T cell stimulation. This may reflect dissociation of CD3zeta from the TCRalphabetaCD3epsilondeltagamma complex or transport of intracellular CD3zeta directly to the cell surface. We also show that MHC:peptide ligation also results in exposure of the TCR-associated CD3zeta NH2 terminus, which is ordinarily buried in the complex. These observations appears to be dependent on Src family protein tyrosine kinases, which are known to be critical for efficient T cell activation. These data suggest a mechanism by which ligated TCR may be differentiated from unligated TCR and selectively down-modulated.     

Divalent cation effects on membrane bending in heated erythrocytes

The thermal fragmentation of human erythrocytes involves either surface wave growth and membrane externalization at the cell rim or membrane internalization at the cell dimple. In symmetrical monovalent electrolytes an increase in membrane internalization at the cell dimple correlates with the decrease in zeta potential arising from surface charge (sialic acid residue) depletion. The influence of divalent cations on thermal fragmentation is examined in this work. The erythrocyte zeta potential decreased when divalent cations replaced some Na⁺ in the cell-suspending phase. The incidence of membrane internalization increased in rank order Ca²⁺>Ba²⁺>Mg²⁺Sr²⁺. Calcium continued to influence the thermal fragmentation of cells highly depleted of sialic acid, suggesting that the ion also interacted with membrane sites other than sialic acid. The divalent cation influence on cell fragmentation was shown to be greater than that due to zeta potential decrease alone. This conclusion was supported by the observation that the divalent cation-induced changes in zeta potential showed much less cation specificity than did the changes induced in the thermal fragmentation pattern. The result implies that the specificity of the divalent cation effects was due to interactions within the erythrocyte shear layer. The possibility that the interaction is with membrane lipids is examined.     

Transmembrane helical interactions: zeta chain dimerization and functional association with the T cell antigen receptor.

Members of the zeta family of receptor subunits (zeta, eta and gamma) are structurally related proteins found as components of the T cell antigen receptor (TCR) and certain Fc receptors. These proteins share the ability to form disulfide-linked dimers with themselves and with other members of the family. Comparison of the amino acid sequences of zeta and gamma reveals a significant degree of homology, which is highest within their membrane-spanning domains. Analysis of their transmembrane sequences on a helical wheel projection suggests that all of the identical amino acids are clustered on one face of a potential alpha-helix. This face contains the only cysteine residue within zeta, suggesting that this conserved region may function to mediate dimerization. Indeed, replacing the transmembrane domain of the Tac antigen (alpha chain of the interleukin-2 receptor) by that of the zeta chain resulted in the formation of disulfide-linked dimers of Tac. The conserved aspartic acid residue found in the zeta and gamma transmembrane sequences was found to play a role in disulfide linkage. Replacing the aspartic acid with a lysine but not with an alanine or valine residue allowed formation of disulfide-linked dimers. The ability of the aspartic acid residue to support dimerization was dependent upon its position within the helix. Thus, these observations indicate that residues within the zeta transmembrane domain play a critical role in the formation of disulfide-linked dimers. Expression of zeta mutants in zeta-deficient T cells revealed that the zeta transmembrane domain is also responsible for reconstituting transport of functional TCR complexes to the cell surface and differentiated the requirements for disulfide-linked dimerization per se from assembly of the TCR complex.     

aPKC-PAR complex dysfunction and tight junction disassembly in renal epithelial cells during ATP depletion

Renal ischemia and in vitro ATP depletion result in disruption of the epithelial tight junction barrier, which is accompanied by breakdown of plasma membrane polarity. Tight junction formation is regulated by evolutionarily conserved complexes, including that of atypical protein kinase C (aPKC), Par3, and Par6. The aPKC signaling complex is activated by Rac and regulated by protein phosphorylation and associations with other tight junction regulatory proteins, for example, mLgl. In this study, we examined the role of aPKC signaling complex during ATP depletion and recovery in Madin-Darby canine kidney cells. ATP depletion reduced Rac GTPase activity and induced Par3, aPKC, and mLgl-1 redistribution from sites of cell-cell contact, which was restored following recovery from ATP depletion. Zonula occludens (ZO)-1 and Par3 phosphorylation was reduced and association of aPKC with its substrates Par3 and mLgl-1 was stabilized in ATP-depleted Madin-Darby canine kidney cells. ATP depletion also induced a stable association of Par3 with Tiam-1, a Rac GTPase exchange factor, which explains how aPKC and Rac activities were suppressed. Experimental inhibition of aPKC during recovery from ATP depletion interfered with reassembly of ZO-1 and Par3 at cell junctions. These data indicate that aPKC signaling is impaired during ATP depletion, participates in tight junction disassembly during cell injury and is important for tight junction reassembly during recovery. ischemia; atypical PKC; Par3; zonula occludens-1; mLgl-1     

Surface properties of calcium phosphate particles for self setting bone cements

Calcium phosphate cements (CPC), consist of multicomponent powder mixtures of calcium orthophosphates with grain sizes in the region of 1-20 microm. Due to the small particle sizes surface properties as the zeta potential and adsorption processes play a significant role during manufacturing and application. In the context of this work zeta potentials of different calcium phosphates, like dicalcium phosphate anhydride (DPCA) tetracalcium phosphate (TTCP) and hydroxyapatite were measured in various organic/aqueous media with different pH values. The results show a strong dependency of the zeta potential on the kind of suspension medium used associated with different milling properties. The addition of sodium phosphate leads to a pH value dependent stabilization of the particles in the liquid phase; the zeta potential of the surface increases from about -15 to -18 mV in water and from -35 to -45 mV in 0.05 mol/l sodium phosphate solution. Besides the interaction of particles with various antibiotics was determined on the basis of the zeta potential of the surface. The substances partly cause a tremendous change of the surface load. This is accompanied by a change of the rheological properties of the cement paste, the morphology of the hardened cement matrix and a significant deterioration of the application-relevant properties as setting time or mechanical strength.     

PKC zeta participates in activation of inflammatory response induced by enteropathogenic E. coli

We showed previously that enteropathogenic Escherichia coli (EPEC) infection of intestinal epithelial cells induces inflammation by activating NF-B and upregulating IL-8 expression. We also reported that extracellular signal-regulated kinases (ERKs) participate in EPEC-induced NF-B activation but that other signaling molecules such as PKC may be involved. The aim of this study was to determine whether PKC is activated by EPEC and to investigate whether it also plays a role in EPEC-associated inflammation. EPEC infection induced the translocation of PKC from the cytosol to the membrane and its activation as determined by kinase activity assays. Inhibition of PKC by the pharmacological inhibitor rottlerin, the inhibitory myristoylated PKC pseudosubstrate (MYR-PKC-PS), or transient expression of a nonfunctional PKC significantly suppressed EPEC-induced IB phosphorylation. Although PKC can activate ERK, MYR-PKC-PS had no effect on EPEC-induced stimulation of this pathway, suggesting that they are independent events. PKC can regulate NF-B activation by interacting with and activating IB kinase (IKK). Coimmunoprecipitation studies showed that the association of PKC and IKK increased threefold 60 min after infection. Kinase activity assays using immunoprecipitated PKC-IKK complexes from infected intestinal epithelial cells and recombinant IB as a substrate showed a 2.5-fold increase in IB phosphorylation. PKC can also regulate NF-B by serine phosphorylation of the p65 subunit. Serine phosphorylation of p65 was increased after EPEC infection but could not be consistently attenuated by MYR-PKC-PS, suggesting that other signaling events may be involved in this particular arm of NF-B regulation. We speculate that EPEC infection of intestinal epithelial cells activates several signaling pathways including PKC and ERK that lead to NF-B activation, thus ensuring the proinflammatory response. inflammation; enteropathogenic Escherichia coli; nuclear factor-B; protein kinase C; IB kinase; extracellular signal-regulated kinase     

Effect of surface potential on Rb+ uptake in yeast: The effect of pH

The apparent K_m of Rb⁺ uptake and the zeta potential of yeast cells are appreciably affected by changes in the pH, variation of the concentration of the buffer cation Tris⁺ and addition of Ca²⁺ to the suspending medium. Irrespective of the way in which the zeta potential is affected, a direct relationship between the apparent K_m of the Rb⁺ uptake and the zeta potential is observed. A reduction of 8 mV in the zeta potential is accompanied by a 20-fold increase in the apparent K_m, which illustrates that electrostatic effects in ion uptake cannot be ignored. Measured zeta potentials are, to a good approximation, linearly related to surface potentials evaluated from a kinetic analysis of the Rb⁺ uptake. This shows the practical use of the zeta potential as a measure of the surface potential in studies of electrostatic effects in ion uptake by yeast. It is concluded that Tris⁺ and the aikaline earth cations inhibit the Rb⁺ uptake in yeast exclusively via a reduction in the surface potential. Protons, in addition, exert a competitive inhibition.     

核酸载体lipopolyplex 的制备及细胞毒性研究

目的：为降低聚阳离子基因载体polyplex 的正电荷和毒性,在其表面构建中性磷脂膜制备lipopolyplex,并测定lipopolyplex 对小鼠结肠癌细胞CT26 和人乳腺癌细胞MCF-7 的细胞毒性。方法：采用PEI25KDa与DNA 复合制备polyplex,在polyplex 体系 中加入中性脂质体和SADGE 制备lipopolyplex。采用琼脂糖凝胶电泳考察lipopolyplex 对质粒DNA的包裹能力;采用激光粒度 仪和zeta 电位分析仪测定lipopolyplex 的粒径与zeta 电位;采用透射电镜观察lipopolyplex 的形态;采用CCK-8 试剂盒考察 lipopolyplex 对CT26和MCF-7 的细胞毒性。结果：琼脂糖凝胶电泳显示lipopolyplex 可以完全包裹质粒DNA;lipopolyplex 的粒 径在200 nm 左右,电位在-20 mV 左右;透射电镜下为较为规则的球状颗粒;lipopolyplex 在CT26 和MCF-7 细胞中的毒性明显 低于聚阳离子基因载体polyplex。结论：在polyplex 表面成功构建中性磷脂膜制备的lipopolyplex,可以完全的包裹DNA 并且细 胞毒性明显低于polyplex,在基因输送载体领域具有潜在应用价值。     

A critical role for PKC zeta in endothelin-1-induced uterine contractions at the end of pregnancy

We have previously shown that protein kinase C (PKC) and/or PKC are necessary for endothelin-1 (ET-1)-induced human myometrial contraction at the end of pregnancy (Eude I, Paris P, Cabrol D, Ferré F, and Breuiller-Fouché M. Biol Reprod 63: 1567–1573, 2000). Here, we report that the selective inhibitor of PKC isoform, Rottlerin, does not prevent ET-1-induced contractions, whereas LY-294002, a phosphatidylinositol (PI) 3-kinase inhibitor, affects the contractile response. This study characterized the in vitro contractile response of cultured human pregnant myometrial cells to ET-1 known to induce in vitro contractions of intact uterine smooth muscle strips. Cultured myometrial cells incorporated into collagen lattices have the capacity to reduce the size of these lattices, referred to as lattice contraction. Neither the selective conventional PKC isoform inhibitor, Gö-6976, or rottlerin affected myometrial cell-mediated gel contraction by ET-1, whereas this effect was blocked by LY-294002. We found that treatment of myometrial cell lattices with an inhibitory peptide specific for PKC or with an antisense against PKC resulted in a significant loss of ET-1-induced contraction. Evidence is also presented by using confocal microscopy that ET-1 induced translocation of PKC to a structure coincident with the actin-rich microfilaments of the cytoskeleton. We have shown that PKC has a role in the actin organization in ET-1-stimulated cells. Accordingly, our results suggest that PKC plays a role in myometrial contraction in pregnant women. protein kinase C; uterine smooth muscle; parturition     

Zeta potential as a diagnostic tool to evaluate the biomass electrostatic adhesion during ion-exchange expanded bed application

Expanded bed adsorption is an integrative technology in downstream processing allowing the direct capture of target proteins from biomass (cells or cell debris) containing feedstocks. Potential adhesion of biomass on the surface of adsorbent, however, may hamper the application of this technique. Since the electrostatic forces dominate the interactions between biomass and adsorbent, the concept of zeta potential was introduced to characterize the biomass/adsorbent electrostatic interactions during expanded bed application. The criterion of zeta potential evaluation proposed in the previous paper (Biotechnol Bioeng, 83(2):149-157, 2003) was verified further with the experimental validation. The zeta potential of intact cells and homogenates of four microorganisms (Escherichia coli, Bacillus subtilis, Pichia pastoris, and S. cerevisiae) were measured under varying pH and salt concentration, and two ion-exchange adsorbents (Streamline DEAE and Streamline QXL) were investigated. The biomass transmission index (BTI) from the biomass pulse response experiments was used as the indicator of biomass adhesion in expanded bed. Combining the influences from zeta potential of adsorbent (zeta(a)), zeta potential of biomass (zeta(b)) and biomass size (d), a good relationship was established between the zeta potential parameter (-zeta(a)zeta(b)d) and BTI for all experimental conditions. The threshold value of parameter (-zeta(a)zeta(b)d) can be defined as 120 mV2 microm for BTI above 0.9. This means that the systems with (-zeta(a)zeta(b)d) < 120 show neglectable electrostatic bio-adhesion, and would have a considerable probability of forming stable expanded beds in a biomass suspension under the particular experimental conditions.     

Effect of polyamines on the electrokinetic properties of red blood cells.

At the physiological pH 7.4, the zeta potential of the normal red blood cell in 1.5% glycine buffer was found to be ?52 mv, whereas that of sickling erythrocytes is ?45 mv. Addition of spermidine to normal red blood cells reduced the zeta potential by approximately 20 mv. In sickling red blood cells, where the polyamine content is determined to be 5 to 6 times greater than in the normal erythrocyte, addition of spermidine reduced the zeta potential by only 5 mv, indicating that little more polyamine binding occurs. The polyamine content of whole blood taken from 24 patients having sickle cell anemia was found to be more than ten times that of whole blood from normal donors. Binding of polyamines to the normal red blood cell was analyzed from the surface charge potential variation as a function of polyamine concentration and the apparent binding constant determined to be 130 d1/g. The difference in the electrokinetic properties of normal and sickling red blood cells in this system may be attributed, in part, to a variation in the polyamine content of the two types of erythrocytes.     

The effect of electrostatic charge of food particles on capture efficiency by Oxyrrhis marina Dujardin (dinoflagellate)

Laboratory experiments were carried out to investigate the effect of food quality, measured as surface charge of the particles, on capture efficiency and ingestion rate by the heterotrophic dinoflagellate Oxyrrhis marina. Fluorescent particles in two size classes of around 1 and 4 microm and of 7 different qualities were offered to the flagellate: carbohydrate and albumin particles, the algae Synechocystis spec. and Chlorella spec., carboxylated microspheres, silicate particles and bacteria. Rates of particle uptake showed significant differences depending on particle size and quality, and ranged from 0 to 4 particles cell(-1) h(-1). Ingestion rates were up to 4 times higher for 4 pm particles than for 1 microm particles, which indicates strong size-selective feeding. Our main result is that the surface charge or zeta potential, of artificial particles, i.e. carboxylated microspheres (> or = -107 mV) and silicate particles, strongly differ from more natural and natural food (< or = -17 mV). For both size classes Oxyrrhis had ingestion rates up to 4 times higher for particles with less negative charge, such as albumin particles or algae. Thus, the zeta potential of the model food should be considered in experimental design. Particles with a zeta potential similar to that of natural food, e.g. albumin, seem to be the preferred model food.     

Role of the zeta chain in the expression of the T cell antigen receptor: genetic reconstitution studies.

The zeta (zeta) chain plays a central role in T cell antigen receptor assembly and signal transduction. From previous work in murine T cell hybridomas we have inferred that the zeta subunit is limiting in receptor assembly. Partial receptors made in excess of zeta are assembled in the endoplasmic reticulum, transported through the Golgi, but then rapidly and efficiently degraded in lysosomes. zeta would therefore seem to play a unique role in targeting receptors from the Golgi to the cell surface. To determine directly whether zeta limits receptor assembly we have reconstituted a zeta-deficient T cell line by transfection of the murine zeta cDNA. Transfection results in restoration of expression of surface T cell receptor. In addition, increasing zeta expression results in a commensurate increase in the survival of previously excess subunits. This is reflected in an increased surface expression of complete receptors. Finally, transfection of the zeta cDNA fails to produce detectable zeta-eta heterodimers. The implications of these findings with regard to receptor assembly, and the relationship between zeta and eta, are discussed.     

The interaction between saliva and Actinobacillus actinomycetemcomitans influenced by the Zeta potential

The adhesion of Actinobacillus actinomycetemcomitans is a virulence factor in the aetiology of periodontitis and is determined by physico-chemical properties, e.g. surface charge and hydrophobicity, of the bacterial cell surface. Although oral surfaces are constantly coated with saliva, few studies have dealt with the binding of A. actinomycetemcomitans with saliva. In this report, the charge properties of A. actinomycetemcomitans have been studied through measurement of the zeta potential and the saliva-bacteria interaction investigated at different pH-values.At physiological conditions the zeta potential was negative, varying from -11 to -26 mV, for two laboratory and two fresh isolates of A. actinomycetemcomitans. Under these conditions, binding of the low-molecular-weight salivary mucin, lactoferrin, and S-IgA was confirmed using salivary samples and purified salivary fractions in liquid-phase and in ELISA. The iso-electric points of the laboratory and fresh clinical isolates of A. actinomycetemcomitans were determined at pH 4.6 and 3.8, respectively. At pH below the iso-electric point, giving positive values of the zeta potential, additional salivary protein species bound to A. actinomycetemcomitans, including the high-molecular-weight salivary mucin (MG1) and agglutinin. Binding of the low-molecular-weight salivary mucin (MG2), lactoferrin, and S-IgA, was hardly affected by this change in zeta potential. A salivary coating formed on the bacterium at pH 7 reduced the zeta potential of the laboratory strain Y4 greatly and an iso-electric point for the bacterium could not be determined. Overall, the study suggests that upon changes in environmental pH additional salivary attachment sites on the micro-organism are exposed.