Chemical and physicochemical properties of Escherichia coli: variations among three strains and influence of culture conditions

Co/lords and Surfaces B: Biomterfaces. 0927-7765/94/$07.00 10 1994 -

2 ( 1994) 47-56 Elsevier Sctence B V. All rtghts reserved

47

Chemical and physicochemical properties of Escherichia coli: variations among three strains and influence of culture conditions H. Latrachea, N. Mozesb,*, C. Pelletier”, P. Bourlioux” “Laboratoire de Microbiologic, Facult& de Pharmacie, UniversitP Paris XI, Rue Jean Baptiste ClPment 5, 92296 Chdten~~-~~~l~br~, France ‘U?~it~ de Chim~e des l~lterf~ces~ U~~~l~ersit~Cotholique de ~ouvffi?z, PIace Croix du Sud I/18, 1348 Lol~v~i?~-la-Neuve, Belgium (Received

7 April 1993; accepted

10 September

1993)

Abstract The surface proper&s of three Esckericha coli strains (AL52. 382 and HBlOl) were Investigated: the surface chemrcal composition by X-ray photoelectron spectroscopy (XPS), the surface electrtcal properties by microelectrophorests and the surface hydrophobtctty by partntonmg between two aqueous phases The surface structures were also analyzed: fimbriae by the hemagglutmation test and lipopolysacchartde (LPS) profiles by electrophoresis on polyacrylamtde gel. The surface chemtcal compostnon depended on the compositton of the culture medtum and on the mode of culture. in liquid nutrrtive medium or on nutrtttve agar. Vartattons between the surface chemtcal composition of the three strains were also observed. It seemed that the chemrcal composttron could be related to the surface structures. The LPS profile of AL52 was the only one that indicated the presence of long polysacchartde chains; this was consistent with XPS analysts which showed that the surface of this stram was richer in hydroxide funcnons than the two other strains. There were more type 1 fimbriae (proteuuc appendages) on the 382 than on the LIB101 stram, and the surface mtrogen concentratton detected by XPS was indeed higher for 382 m compartson wtth the HBlOl strain. The chemtcal composttton could be related to the eiectrophorettc mobtlity whtch increased as the phosphate surface concentration increased. However, there was no coherent relation between any parameter of the surface chemtcal composttron and the surface hydrophobrctty Key words Cell surface; E. colt; Electrophoretic spectroscopy

mobthty:

Ftmbrtae;

Introduction Urinary tract infections (UTIs) are most often caused by Escherichia coli [l-4]. Most of the nosocomial UTIs are related to the presence of a catheter [ 5-71. Adhesion of E. coli onto the catheter is thought to be the first step in this kind of pathogenic infection [ 61. The outermost surface of the bacterial cell plays a crucial role in the *Corresponding

author

SSDI 0927-7765t93

)01089-A

Ltpopolysacchartde;

Surface hydrophobtctty;

X-ray photoelectron

adherence. Recent studies [ 8- 111 have indicated that the adhesion of many bacteria to solid surfaces depends on physicochemical surface properties, such as electrical potential and hydrophobicity. These properties are conferred upon the cell surface by its chemistry. A knowledge of the elemental, molecular and structural composition and of the physicochemical properties of the cell surface is required in order to understand fully the process of adhesion. The relationship between the physicochemical surface properties and the chemical and

H Latruchr et ul /Collards Surjbces B Bwmterfuces 2

48

structural including

composition of various microorganisms, bacteria and yeast, has been described

[ 12-171. The aims

of this work

surface properties

of various

are (i) to analyze E. coli strains

the

and to

minimum

medium

which was Na,HP0,.2H,O, 0.5 mM

0.02 mM FeS0,.7H,O pH was 7. Bacteria steps: preculture 22 h. To promote

chemical

bacteria

of the cell surface.

The surface chemical composition was determined by X-ray photoelectron spectroscopy (XPS )_ a surface-specific method used mainly in material sciences. This technique is based on irradiating the sample by an X-ray beam, analyzing the kinetic energy of the ejected photoelectrons and calculating their binding energy in the source atom. Each peak of the recorded spectrum is characteristic of a given element. The position and the shape of the peaks are influenced by the chemical bonds and the oxidation state of the analyzed atom. The method thus provides elemental and functional information. Owing to inelastic scattering, the electrons collected originate from the outermost molecular layers and the analytical information thus concerns a thin layer (3-10 nm) at the surface [IS]. The use of XPS for analysis of microbial cells has been developed in the last 10 years [ 191. Materials and methods Bacteria Three E. coli strains were used: (i) HBlOl, a K-12 strain, non-pathogenic and phenotypically positive for mannose-recognizing adhesins; (ii) 382; (iii) AL52. The last two strains were isolated from patients with urinary tract infections. The strain 382 was phenotypically positive for mannoserecognizing adhesins, while AL52 was not. Of the three strains, only AL52 was genotypically positive for pap and sfa homologous DNA. Each strain was grown either in (liquid) Luria Bertani medium (LLB) or on (solid) Luria Bertani agar (SLB). For testing the effect of medium composition, the strain 382 was grown also in (liquid)

22 mM 9 mM

MgS0,.7H,O,

test the effect of culture conditions upon these properties, and (ii) to examine the relationships between the physicochemical properties and the composition

(LMin),

the

composition of 48 mM KHlPO,,

NaCl,

19 mM

0.07 mM

NH,Cl,

CaC1,.2H,O,

and 28 mM D-glucose: were cultured

the

at 37’ C in two

during 18 h and culture during the development of fimbriae, the

were cultured

X-U?: photorlect~on

( 1994 ) 47-M

without

agitation

[20].

sprctroscopJ

The surface chemical composition was analyzed by XPS. Bacteria were harvested after 22 h of growth (late stationary phase). collected by centrifugation (3500g x 10 min), suspended in distilled water and washed twice by successive resuspensions and centrifugations. The pellet of the last centrifugation was transferred to a glass vial, frozen in liquid nitrogen and kept at -80’C until freeze drying (Lyovac GT4 Thermovac TM, Leybold Heraeus). The samples were placed on the precooled (-50°C) shelf of the lyophilizer, evacuation was started and when the pressure reached 60 Pa, the temperature was raised to - 1O’C; lyophilization lasted about 18 h. The powder of freeze-dried cells was mounted in a stainless steel trough and pressed to obtain a macroscopically smooth surface. The analyses were carried out in an SSX-100 spectrometer (model 206) of Surface Science Instruments, interfaced with a Hewlett-Packard 9000/3 10 computer allowing instrument control, data accumulation and data treatment. The X-rays were generated from a monochromatized aluminum anode, the pressure in the chamber during analysis was about 10e6 Pa, the flood gun energy was 6 eV, and the pass energy of the analyzer 50 eV. Resolution, determined on a gold standard, (FWHM Au 4f7,2) was 1.0 eV. The size of the analyzed area was about 0.5 mm2 or 1.4 mm’. The order of peak analysis was Cls, 01s Nls, P2p, K2p, Nals, S2p, Cls. The duplication of Cls registration at the end provided an estimate of sample degradation under X-ray irradiation during the period of data accumulation

(about

3.5 h). The

H. Latrache et ul.JCollolds Surfaces B. Blointerfaces 2 ( 1994 ) 47-56

binding energies were calibrated with respect to the c-(C,H) component of the Cls peak set at 284.8 eV. Atom fractions were calculated from the peak areas normalized after non-linear background subtraction, and with the sensitivity factors supplied by the spectrometer manufacturer. Major complex peaks were decomposed using a leastsquares best fitting routine with a Gaussian/Lorenzian ratio of 85/15, employing literature information concerning component binding energies and fixing the full width at half maximum height at a constant value for all the components of a given peak. Samples of silica (quartz Sikron SF800, or Silica 27620298 from Prolabo) were prepared in parallel with the bacterial samples (centrifugation, freezing, lyophilization) and analyzed by XPS in order to evaluate the amount of contaminating carbonaceous compounds. Microelectrophoresis

The electrical properties of the bacteria were characterized by measuring their electrophoret~~ mobility. The cells were washed twice with 0.9% NaCl solution, and treated with 1% form01 solution during 20 min at room temperature, to eliminate the bacterial motility. The form01 was removed by centrifugation and the bacteria were suspended in distilled water. A portion of this suspension was diluted in 10F3 M KNO,. The pH was adjusted by HNO, or KOH and the electrophoretic mobility was determined with a zetameter Zm77 (Zetameter Incorporation, New York). Two-phase

partitioning

The surface hydrophobicity of the bacteria was evaluated by following their partitioning between two aqueous phases of different surface tensions. The system [21] consisted of a mixture of 4% (w/w) poly(ethylene glycol) 6000 (PEG), and 5% (w/w) dextran in 0.023 M phosphate buffer pH 6.8 and 0.123 M NaCl [22,23], equilibrated overnight in a separating funnel at 4°C. The bottom phase

49

(rich in dextran) and the top phase (rich in PEG) were then collected and stored separately. The top phase has lower surface energy [ 241. The bacteria were washed twice and suspended in phosphate buffer saline (PBS), pH 7.2; a 0.5 ml portion of this suspension was added to a mixture of 2 ml of the PEG-rich phase and 2 ml of the dextran-rich phase in a test tube; the material was mixed and the phases were allowed to separate at 4°C during 30 min followed by 30 min at room temperature. After separation, the number of bacteria in the top phase (lower surface tension) was estimated turbidimetrically. The interfacial electric potential is very low according to Reitherman et al. [22]; therefore the partition of cells between the two phases depends probably upon surface characteristics other than charge, which might be referred to as cell surface hydrophobi~ity [253. The hydrophobicity was expressed as the ratio (in per cent) between the amount of cells in the top phase and the total amount of cells in the test. Electrophoyet~c

analysis of ~ipopolysaec~ayi~es

(LPSS)

The harvested bacteria were washed twice in PBS, and suspended to an optical absorbance of 0.5 at 525 nm. A portion of this suspension (1.5 ml) was centrifuged, the cells in the pellet were then lysed in 0.05 ml of SDS buffer (2% SDS, 4% 2-mercaptoethanol, 10% glycerol, 0.002 Bromophenol Blue in 1 M Tris-HCl buffer, pH 6.8) at 100°C during 10 min, then treated with proteinase K (0.5 mg ml-l; Sigma) at 60” C during 60 min [26]. The lysate was submitted to electrophoresis on a 14% polyacrylamide gel. The LPS profiles were revealed by staining with silver nitrate as described by Fomsgaard et al. [27]. He~agg~~tinat~o~ test

Mannose-sensitive hemagglutination was used to determine the presence of type 1 fimbriae on the bacterial surface. Twice washed bacteria were resuspended in PBS at concentration of f09

0, N) were similar for all the samples (62~69%, 22228%. 6-9% respectively) with the exception of

cell mlK’ and serial twofold dilutions were prepared from this suspension; in parallel, a washed 3% (v/v) suspension

of guinea

pig erythrocytes

PBS) was prepared. The hemagglutination tested by mixing equal volumes of bacterial erythrocyte

suspensions

at

4. C during

96well (U-shaped) microdilution agglutination titer was then expressed

as the

highest

dilution

the strain

(in

m nitrogen,

plates. The hemdetermined and of the

bacterial suspension that resulted in positive agglutination; thus a high hemagglutlnation means a high amount of fimbriae.

medium

compared

with

the

which

and poorer

other

samples.

ences in phosphorus

concentration

could be noted

between

HBlOl>383

> AL52 in both

the strains:

solid and liquid cultures with LB medium. Low concentrations of sulfur (about 0.1%) were observed. Only cells from liquid cultures of the 382 strain grown on minimal medium and of the HBlOl mutant grown on LB had a potassium surface concentration significantly above the detection limit. Sodium was found in all samples, with slightly higher concentrations on the surface of the samples with low potassium concentration.

hemtiter

Results

All the XPS analyses were performed in duplicate (two independent cultures for each condition). The reproducibility was satisfactory: variations between duplicates were in the range of 1% for carbon. 5% for oxygen, up to 10% for nitrogen and phosphorus, and up to 94% for the minor elements. The silica controls showed that contamination was minimal (mean value for contaminating carbon was 5.6%). No serious degradation due to irradiation was noted. Elemrntul compositiorl Table 1 summarizes the results

culture

richer in oxygen

Phosphorus was found on all samples at a concentration of about 1%: small but systematic differ-

1 h in

factor

AL52 on solid

seemed to be slightly

was and

Fulzctmzul composition Decomposition of complex XPS peaks supplies information regarding the chemical function in which a given element is involved. Figure 1 shows representative Cls, 01s and Nls spectra. The component of the Cls peak at a binding energy of 284.8 eV was attributed to carbon bound only to carbon or hydrogen, C-(C-H); the one at a binding energy of 286.2 eV was attributed to carbon singly bound to oxygen or nitrogen. C-(0,N). like in alcohol or amine, and the component at 287.9 eV to carbon doubly bound to oxygen, C=O. as in

of the surface

elemental composition of the different samples. The surface concentrations of major elements (C,

carbonyl.

Table 1 Elemental composltlon of the surface of three E. co11 strams cultivated mean of at least two analyses of cells from Independent cultures)

carboxyiate,

under different conditions

amide

etc. The 01s

peak

(atom fraction (‘%) ewcludmg hydrogen:

Stram

Culture

C

0

N

P

Ii --

Na

s

AL52 HBlOl 382 AL52 HBlOl 382 382

SLB SLB SLB LT.3 LLB LLB LMm

62.5 66 4 62.7 65.3 64.3 6X.9 63.3

31.0 ‘5.4 76.7 ‘7.6 16.9 ‘2.4 27.7

5.3 6.3 8.7 5.9 6.5 9.1 6.7

07 13 1.0 09 15 1.0 1.5

< dl” 0.08
0.43 0.40 0.74 0.27 013 0 51 0. I 5


“dl. detectIon

limit (0 05%‘)

H. Latrache

et ul.iCollmds

Surfaces B Biomterfuc~es -7 ( 1994 ) 47--56

sition for all the samples. The C-( C,H) component was clearly higher than the CJO,N) component in most of the samples except the strain AL52 on SLB. The oxygen was found mainly in hydroxide or acetal functions in all samples. It is interesting to note in this respect that the variations in the total surface oxygen concentration were mainly due to variations in hydroxide or acetal function: the sample richest in surface 0, had also highest -OH or CO-C (which was three times more than the other 0 component); the poorest sample in 0 had lowest -QH (the value of which was the same as the other 0 component). The protonated nitrogen constituted generally about 10% of the total nitrogen detected.

x

5 E Q) .’ 5 8 594

532

51

5AO

D$erences 402

400

398

Binding energy (eV) Fig. 1. XPS spectra

of HBlOl

cells: a. Cls; b. 01s; c. Nls.

component at a binding energy of 532.6 eV was attributed to hydroxide, C-OH. and acetal or hemiacetal, C-C-C functions; that at 531.2 eV to doubly bound oxygen, Q=C. The main Nls component at 399.9 eV was due to non-protonated nitrogen, like amine or amide, and the 401.6 eV component to protonated nitrogen, e.g. ammonium ion. Table 2 summarizes Table 2 Functtonal hydrogen;

the results of peak decompo-

betweet

the struins

The AL52 strain differed from the two others, when cultivated on solid culture medium. in having higher surface oxygen, lower nitrogen, lower phosphorus. and undetectable sulfur. The high surface oxygen concentration of AL52 was mainly due to an increase in the QH or CO-C component and the lower nitrogen concentration was due to a decrease in the non-protonated nitrogen. The strain 382 had slightly higher surface nitrogen (non-protonated) than the other two strains in both solid and liquid cultures. Injuence

of growth conditions

Comparison of the surface composition of bacteria grown on solid medium with those of cells

composttion of the surface of three E. CO/I strams cultwated under mean of at least two analyses of cells from independent cultures)

dtfierent

condtttons

(atom

fractton

(4’) evcludmg

Stram

Culture

C-(C.H I 284.8 eV

C-(O,N 1 286.2 eV

I=0 287 9 eV

Q=C 5312 eV

+H. C-Q-C 532.6 eV

Non-prot-N 399.9 eV

Prot-Nb 401.6 eV

AL52 HBlOl 382 AL52 HBlOl 382 382

SLB SLB SLB LLB LLB LLB LMin

24.6 38 7 30.2 32 1 36.5 38 5 332

26.9 19.5 218 24.2 20.1 18 5 21.1

11.0 8.2 10.7 9.0 86 9.9 90

8.2 95 10.5 68 10.6 10 2 10.6

22 8 15.9 16.2 20.8 16.2 12.2 172

4.8 5.7 8.2 5.4 5x 8.5 59

0.53 0.63 0.50 0.52 0 72 0.65 0.80

“Non-protonated mtrogen. bProtonated mtrogen.

cultivated

in liquid

systematic

difference:

medium

revealed

for the three

only

strains

one there

seemed to be more sodium on the surface of ceils from solid cultures. For the AL52 and 382 strains the C4C.H ) component of the C 1s peak was much lower on cells from solid medium culture

cells. For each strain

concentrations

than

the nitrogen

were very similar

on hquld surface

in the two cul-

ture modes. The effect of medium composition on the surface composition of the bacteria was tested by analyzing the strain 382 cultivated in two different liquid media, LLB and LMm. In the former the cell surface was poorer in hydroxide or acetal, potassium and phosphorus, and richer in nonprotonated nitrogen and sodium.

Table 3 summarizes the physicochemical ties of the three strains.

2

Strain

(electrokmetlc

Type of culture

propertIes

8

6

PH Fig 2 Elecrophoretlc moblhty of E CYJI grnun m hquld LB as a function of pH A. HBIOI. 0, 381. II. AL57 Bars represent two values obtamed wtth two mdependent cultures

the strain HBlOl was the most negatively charged, and AL52 the least charged. The EPMs of bacteria from solid cultures differed from those of cells from liquid cultures (Table 3). The composition of the culture medium (tested for the strain 382) had no influence on the EPM at pH 7 (results not shown).

proper-

The variation of the electrophoretic mobility (EPM) as a function of pH for the three strains cultivated in LLB is presented in Fig. 2. The isoelectric points were 3.0. 2.0 and 3.5 for AL52. HBlOl and 382 respectively, The EPMs became more negative with an increasing pH. At pH 5-9 Table 3 Physlcochemlcal properties cultwated m LB

4

Surface hydrophobicit?

The differences in hydrophobicity among the strains were evident only in the liquid cultures {Table 3 ): AL.52 cells were hydrophobic~ HBlOl

and surface

hydrophoblclty)

IEP”

and hemagglutmation

titer of three E CO/I strams

Proportion m the PEG phase’

Hemagglutlnatlon titer

(C) AL52 HBlOl 382 AL52 HE101 382

Sohd Sohd Sohd Llqwd Ltquid I_lquld

nd nd nd 2.0 20 3.0

- 2.50 -3.44 -3.46 -198 -4.39 -2 92

(0.09) (006) (002) 10.20) (005) (0.36)

45 (31 43 (4) 30 (3) 64 12) 19(I) 31 12)

nd. not determmed. “Isoelectric pornt. bElectrophoretx mob&y at pIi 7, mean values of two rephcate experzments with sample dewatzon m parentheses ‘Mean values of three replicate expernnents: the dewatlon between extreme values 1s given In parentheaes

nd

1 8 nd 16 32

H. Latruche

et a/.iCollolds

were hydrophilic intermediate.

Surfaces B Biomterfuces

and 382 could

2 ( 1994 ) 47--M

be considered

as

53

no other type of fimbriae. The type 1 fimbriae are known to be surface proteinic appendages which mediate

LPS projles Figure 3 shows

the LPS

profiles

of the three

and AL52 had long polysaccharidic chains. No major modification in the LPS profiles as a function of type of culture could be detected. Presence ofjmbrine The results of the hemagglutination test are included in Table 3. The strain 382 showed higher titers of hemagglutination than HBlOl. This means that 382 had more fimbriae than HBlOl. It is interesting to note that both strains were more fimbriated when grown in liquid medium than on agar. Discussion Relation between surface chemical composition and structures

Among

the three strains

only 382 and HBlOl

12

to D-mannose

containing

struc-

tures found on erythrocytes, buccal and uroepithelial cells. The titer of agglutination with erythrocytes

strains: HBlOl had very little polysaccharidic chains, 382 had some short polysaccharidic chains,

surface

binding

345678

described

in this work,

had type 1 fimbriae;

they had

9

Fig. 3. Silver-stained PAGE profiles of proteinase K treated lysate preparations of E colr. Lanes: 1. HBlOl (LLB); 2. HBlOl (SLB). 3, 382 (LMin), 4. 382 (SLB); 5, 382 (LLB); 6. AL52 ( LMm). 7, AL52 (SLB); 8, AL52 (LLB); 9. a smooth-type LPS preparation from E. CO/I 055 B5 (for compatxon).

was higher indicating

for the strain that 382 contained

382 than

for HBlOl,

more type 1 fimbriae.

Indeed, a higher surface nitrogen concentration was detected by XPS analysis for the strain 382 than for HBlOl. Glucose-rich media hinder expression of some fimbriae. including type 1 fimbriae [28]. Eshdat et al. [29] showed that the static culture in Luria Bertani broth favors the synthesis of type 1 fimbriae. Unfortunately, the hemagglutination test was not performed on cells from minimal medium in the present study. The higher surface nitrogen concentration found by XPS for the strain 382 cultivated in LLB compared with cells cultivated in LMin may suggest that there are more fimbriae on the bacteria from LLB; one could assume therefore that the observations of DarfeuilleMichaud and Joly 1281 and of Eshdat et al. [29] are valid here. A relation between the presence of fibrils and surface nitrogen concentration detected by XPS has been reported by van der Mei et al. [30]. They have shown that a progressive loss of proteinous fibrillar surface antigens from Streptococcus salitlarius was concurrent with a decrease in the N/C surface concentration ratio. Comparison of the LPS profiles of the three strains indicated that AL52 was rich in long polysaccharidic chains while the polysaccharides of the two others strains contained short (382) or undetected (HBlOl) chains. This is consistent with the higher proportion of hydroxide and acetal groups observed by XPS analysis for the AL52. However. the clear distinction between the LPS profiles of 382 and HBlOl could not be reflected in XPS data. Magnusson et al. [31] have shown, by using the partitioning method, that the LPSs play an important role in determining the physicochemical properties of bacterial surfaces, and the polysaccharide side-chains increase the hydrophobicity of

H Lutrache et trl.:‘Collods

54

bacteria. One may suggest, longer polysaccharidic chains

therefore, that the of the LPSs on the

impossible Cls

to distinguish

components

AL52 strain may explain the higher hydrophobicity

Therefore

of this strain

surface charge

Surface

compared

chemical

to the two others.

composition

B Bmnterfacrs

( lY94

2

the carboxylic

from other

by XPS on microbial

the contribution

samples.

of such function

may not be excluded.

) 47-56

to the

However,

it

seems that deprotonated carboxyl does not play a major role in development of the surface charge.

and ph~~sicochemicnl

properties

Figure 4 shows

Sttrtues

Mozes et al. [ 171 have already mentioned that the ionization of carboxyl groups is expected to be the electrophoretic

mobility

of

weak owing to the influence

of dissociated

neigh-

the different samples at pH 7 as a function of the surface phosphate concentration determined by

boring phosphate groups. The idea that phosphate is the main, if not the sole, source of the surface

XPS. It seems that as the phosphate concentration increased, the surface became more negative. Phosphate groups (in a form of phosphodiester) are constituents of phospholipids and LPS in the walls of Gram-negative bacteria. Deprotonation of these groups confers a negative charge on the surface. The predominant role of phosphate in determining the surface charge of microorganisms has been recognized by several authors [33-353. Mozes et al. [35] showed that the EPM at pH 4 for several yeasts and bacteria strains has a tendency to become more negative as the phosphate surface concentration increases, but beyond a certain phosphate level the EPM does not change any more. It is sometimes considered that carboxyl groups play also a role in determining the negative charge of the cell surface. Unfortunately, it is

negative charge in our system may be supported by Fig. 5. It shows a 1:l ratio between the sum of the surface concentrations of the three cations ( Naf, K+. NH:), associated with the cell surface probably as counterions, and the surface concentration of phosphate which is a constitutive component of the cell wall. The surface hydrophobicity, estimated by partition in a two-phase system, could not be related to any parameter of the surface chemical composition, determined by XPS. This is contrary to previous reports by Mozes et al. [ 173 and van der Mei et al. [30] who have shown that there is a relation between the surface oxygen concentration and the water contact angle or surface energy

2’01

030

0,8

1,2

116

2,o

P (%)

P(%) Fig. 4. Variation of the electrophoretic mobiltty function of the surface phosphate concentration. hnear correlation IS 0.768.

0.4

at pH 7 as a Coefficient of

Fig. 5. The relation between the surface concentration of cations (K+. Na’ and NH;) and surface phosphate concentration Coefficient of hnear correlation IS 0.614.

H. Latruche

et ul./Colloids Surf&es B. Bioirlterfaces 2 ( 1994) 47-56

respectively.

This discrepancy

specific method phobicity.

used to evaluate

It must be recalled

may be due to the the surface hydroin this context

that

there is no universal definition for the term “surface hydrophobicity” for microbial cells, and no consensus about a scale for its assessment [ 361. Three independent concluded

comparative that

studies

the parameters

various methods used to probe have quite often different physical

[ 37-391 provided

55 2 3 4 5 6

have by the

hydrophobicity meanings.

7 8

9

Conclusion XPS data provide global information on the chemical composition of the cell surface. This information can be roughly related to the presence of certain types of molecule or structure at the cell surface: the highest concentration of hydroxide and acetal groups is consistent with the presence of long polysaccharidic chains on AL52; the highest concentration of nitrogen is in harmony with an excess of proteinic appendages on 382. The chemical composition of the cell surface is also related to the electrical properties: more phosphate at the surface indicates a more negative value of the electrophoretic mobility. The surface composition and properties may vary as functions of growth conditions: composition of the culture medium; solid/liquid medium.

10

11

12

13

14

15 16 17 18

Acknowledgments The financial support of the French Ministry of Research and Technology (contract 88-0598) and of the Department of Education and Scientific Research of the French Community in Belgium (Concerted Action Physical Chemistry of Interfaces and Biotechnology) is gratefully acknowledged. The authors wish to thank Dr. A.J. Leonard for his help in data treatment. References 1

R. Gregor 470-487.

and J.D. Sobet,

Rev. Infect. Dis., 9(3)

( 1987)

19

20

21 22 23 24 25 26 27

R.J. James, Chn. Microbial. Rev., 4( 1) (1991) 80-128. M. Archambaud and L. Labigne. Bull. Inst. Pasteur, Paris. 87 ( 1989) 247-279. G. Chabanon and M. Archambaud. Med. Malad. Infect. No. speciale. ( 1987) 33340. A.J. Schaeffer, Ural. Chn North Am. 1314) 11982) 7355747. H L. Mobely, G.R Chipmdale. J.H. Tenny, R A Hull and J.W. Warren, J. Clin. Microbial, 25 (1987) 2253-2257. J.C Nickel, A.G. Gristma and J.W. Costerton, Can. J. Surg , 28 ( 1985) 50-52. N. Mozes. F. Marchal, M.P. Hermesse. J.L. van Haecht, L Reuhaux. A.J Leonard and PG. Rouxhet, Biotechnol. Bioeng., 30 (1987) 4399450. M.C.M. van Loosdrecht, K. Lyklema, W. Norde, G. Schraa and A.J.B. Zehder. Appl. Environ Microbial., 53 (1987) 189331897 M.C.M van Loosdrecht, K. Lyklema. W. Norde. G. Schraa and A J B. Zehder. Appl. Environ. Microbial. 53 ( 1987) 189991902. M.-N. Bellon-Fontaine. N Mozes, H C. van der Mei. J Sjollema. 0 Cerf, P.G. Rouxhet and H J Busscher, Cell Btophys., 17 (1990) 93-105 H C. van der Mel. M.J. Genet, A.J. Weerkamp, P.G. Rouxhet and H.J. Busscher, Arch. Oral Biol., 34 (1989) 8899694. H.C. van der Mei, P. Brokke, J. Dankert. J. FeuJen, P.G. Rouxhet and H.J Busscher, Appl. Environ. Mtcrobiol., 55 (1989) 280662814. H.J. Busscher. A.H. Weerkamp, H.C. van der Mei, D. van Steenberghe, M. Qmrynen, I.H. Pratt, M. Marechal and P.G. Rouxhet, Colloids Surfaces, 42 (1989) 345-353. D.E Amory, P.G Rouxhet and J.P. Dufour, J. Inst. Brew., 94 (1988) 79-84. D.E. Amory and P.G. Rouxhet, Blochim. Biophys. Acta, 938(1988)61-70. N. Mozes, A.J. Leonard and P.G. Rouxhet, Biochim. Biophys. Acta, 945 (1988) 324-334. P.G. Rouxhet and M.J. Genet. m N. Mozes, P.S. Handley, H.J. Busscher and P.G. Rouxhet (Eds.), Microbial Cell and Physico-chemical Surface analysis: Structural Methods, VCH, New York, 1991, Chapter 8, pp. 1733220. P.G Rouxhet. N Mazes. P.B. Dengis, Y.F. Dufrene, P.A. Germ and M.J. Genet. Colloids Surfaces B: Biomterfaces. 2 (1994) 347. R. Lock, C. Dahlgren, M. Linden, 0. Stendahl, A. Svensbergh and L. Ohman, Infect. Immun., 58 (1990) 37-42 P. Albertsson, J. Chromatogr., 159 (1978) 111-122. SD. Reitherman, S. Flanagan, H. Barndes, Biochim. Biophys. Acta, 297 (1973) 193-197. D. Fisher, Biochem. J., 196 (1981) l-10. P.A. Albertsson, Adv. Protein Chem., 24 (1970) 309-341. D.F. Gerson, Biochim. Biophys. Acta, 602 (1980) 269-280. P. Hitchcock and T.M. Brown, J. Bacterial.. 154 (1983) ‘699277. A. Fomsgaard, M.A. Freudenberg and C Galanos. J. Chn Microbial., 18 (1990) 262772631.

H. Latruche

56 28

29 30

31

32 33

A Darfeutlle-Michaud and B. Joly. Colloque Adheston d‘Etude Centre Pharmaceuttque, Bactertenne. P Bourhoux, Chatena}~-Malabr~, 1983. pp. 77-9.5 Y Eshdat. V. Speth and K. Jann, Infect Immun.. 34 (1981) 980-986. H.C van der Met, A.J Leonard, AH Weerkamp, P.G Rouxhet and H.J. Busscher, J. Bactertol, 170 (1988) 24622’466. K.E. Magnusson. 0. Stendahl, C. Tagesson. L Edebo and G. Johansson. Acta Pathol Mtcrobtol. Stand. Sect. B, 85 (1977)212-218 R Nerhof and W H Echols, Biochem Brophys. Acta. 318 (1973) 23-32 A.A Eddy and A.D Rudin, Proc. R. Sot. London, Ser B. 148 (1958) 419-432.

34 35 36

37 38 39

et ul./Colloids

Surfus

B Bmntwface,

2 ( 1994 ) 47-56

L E Hardham and A.M Janes, Btochem. Btophys Acta, 66 (1963) 2377249. N Mazes. D.E. Amory. A J Leonard and P.G Rouxhet. Collards Surfaces, 42 ( 1989 i 313. 323. H C van der Mel, M. Rosenberg and H J Busscher. m N Mozes. P.S. Handley, H.J. Busscher and P.G. Rouxhet (Eds.), Microbtal Cell Surface Analysts. Structural and Physico-chemtcal Methods, VCH. New York, 1991. Chapter 8, pp 265 287. J.K. Dtllon, J A Fuerst, A.C. Hayward and G.H.G. David. J. Mtcrobrol Methods, 6 ( 1986) 13-19 N. Mozes and P.G. Rouxhet, J. Mtcrobiol Methods, 6 (1987) 999112. H.C van der Met, A.H Weerkamp and H.J. Busscher, J Mrcrobtol. Methods, 6 ( 1987) 777-287