Microb Ecol (1988) 15:229-237

MICROBIAL ECOLOGY Q Springer-VerlagNew York Inc. 1988

Bacterial Desorption from Food Container and Food Processing Surfaces Sharron M c E l d o w n e y ~,2,** and Madilyn Fletcher t,* 'Department of Biological Sciences, Warwick University, Coventry CV4 7AL, England; and 2Campden Food Preservation Research Association, Chi00ing Camodcn. Glos GL55 6LD, England Abstract.

The desorption o f S t a p h y l o c o c c u s aureus, A c i n e t o b a c t e r calcoaceticus, and a c o r y n e f o r m f r o m the surfaces o f materials used for m a n -

ufacturing food containers (glass, tin plate, and polypropylene) or postprocess canning factory c o n v e y o r belts (stainless steel and nylon) was investigated. T h e effect o f time, pH, temperature, and adsorbed organic layers on desorption was studied: S. aureus did not detach from the substrata at any p H investigated (between p H 5 and 9). A. calcoaceticus and the c o r y n e f o r m in some cases detached, depending upon p H and substratum c o m p o s i t i o n . The degree o f bacterial d e t a c h m e n t from the substrata was not related to bacterial respiration at experimental p H values. Bacterial desorption was not affected by t e m p e r a t u r e (4-30~ nor by an adsorbed layer o f peptone and yeast extract on the substrata. T h e results indicate that b~icterial desorption, hence bacterial r e m o v a l during cleaning or their transfer via liquids flowing o v e r colonized surfaces, is likely to vary with the surface composition and the bacterial species colonizing the surfaces.

Introduction T h e d e v e l o p m e n t o f bacterial biofilms on solid surfaces is a d y n a m i c process involving the initial a t t a c h m e n t o f bacteria, growth o f attached ceils on the surface, and the release o f bacteria from the surface back into the liquid phase [4]. This release o f bacteria occurs when unattached daughter cells are produced through attached cell replication, when attached bacteria are sloughed off due to shear forces [4, 6] or surfactant treatment [25], and when bacteria desorb from the surface, apparently because o f localized changes in physicochemical conditions or cell surface properties. Little is k n o w n about the mechanisms o f bacterial detachment; however, the factors that influence attachment, hence the attractive forces between the bacterium and surface, might also be expected to influence d e t a c h m e n t to some extent. Bacterial a t t a c h m e n t is affected by variations in species [16, 22, 27], * Present address: Center of Marine Biotechnology,University of Maryland, 600 E. Lombard St., Baltimore, Maryland 21202, USA. ** Present address: School of Biotechnology, Polytechnic of Central London, 115 New Cavendish St., London WlM 8JS, England.

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substratum composition [ 12, 16, 22, 26], and environmental parameters such as pH [ 18, 25], temperature [19, 26], and the presence of organic macromolecules either adsorbed on the substratum [9] or dissolved in the liquid phase [15, 16]. Bacterial biofilms are significant in a number of industrial situations, including food processing plants where they may be a source of bacteria responsible for food spoilage and poisoning. Repeated cleaning and disinfection of food processing surfaces is necessary, as reinfection of such surfaces occurs regularly. By understanding the factors that control bacterial release and desorption from biofilms on surfaces, it may be possible to develop improved cleaning techniques or control environmental f a c t o r s , s o that reinfection of food processing surfaces is minimized. The aim of this study was to investigate the desorption of selected bacteria from solid surfaces, and to determine the influence of pH, temperature, and adsorbed organic molecules on their desorption. The principal aim of the study was to understand the significance of bacterial desorption in the food industry, with a specific interest in the significance of surface-attached bacteria in food spoilage due to food container leakage. This occurs, for example, when can contents are contaminated with bacteria that come from can or conveyor belt surfaces and pass into the can through temporary or permanent leaks, such as minor faults in can seams. Thus, the bacteria used in this study were isolates from a canning factory and the surfaces were materials used to manufacture food containers and postprocess conveyor belts. However, the results also have implications for bacterial detachment from surfaces in a wide range of natural and man-made environments.

Materials and Methods To isolate strains from a food processing plant, sealed tin cans containing water were passed down the processing line, sterilized by autoclaving, cooled, and allowed to pass down the conveyor belt to the packing area, along with the food cans being processed in the plant. The tin cans were then taken off the line, and sections of tin plate were aseptically cut from the cans and used to inoculate continuous cultures (at 25~ for enrichment of factory isolates. The medium contained 0.1% (w/v) bacteriological peptone and 0.07% (w/v) yeast extract (PYE) in phosphate buffer (5.4 g liter-t K2HPO4), pH 7.4. Organisms were isolated on nutrient agar from continuous culture after 7 days at a dilution rate (D) of 0.1 hour-L Isolates were also obtained by swabbing the postprocess conveyor belt surfaces and using the swabs to inoculate nutrient agar; incubation for 7 days at 25~ followed. Isolates were identified by Gram stain, catalase reaction [ 13], oxidase reaction [ 11 ], eoagulase reaction [ 13], and by Analytical Profile Index (APt Laboratory Products Ltd., Basingstoke, England). Acinetobacter calcoaceticus, a Staphylococcus sp., and a coryneform were isolated from both tin plate and conveyor belt surfaces and were used in this study, along with a Staphylococcus aureus isolated from a nose swab, which was used for comparison. The organisms were grown to late exponential/early stationary phase in 0.1% (w/v) bacteriological peptone and 0.07% (w/v) yeast extract (PYE) at pH 7.4 and 25~ on a rotary incubator at 150 rpm.

Attachment~Detachment Substrata Materials used to manufacture food containers and postprocess canning conveyor belts were used for surfaces. The surfaces representing container materials were glass coverslips (13 m m diameter;

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231

Chance Propper Ltd., England), 16 m m diameter discs of Lamipac (polypropylene; Metal Box Ltd., Wantage, England), and 16 mm diameter discs of tin plate (Metal Box'Ltd.). Conveyor belt surfaces were represented by 13 m m diameter discs of Hyfax (nylon) and 13 m m diameter discs of stainless steel, finish type 2B (both Metal Box Ltd.). The substrata were cleaned by sonication for two 15 min intervals at 100 W on a Kenny sonicator (Jencons Scientific Ltd., England), and the glass was also autoclaved (121~ 15 min) to simulate the heating during food processing. This was not a rigorous cleaning procedure, and the high energy glass and metal surfaces still had adsorbed organic material, which caused the surfaces to be comparatively hydrophobic. This was demonstrated by the relatively high contact angle values produced by drops o f distilled water placed on the surfaces, as described in McEldowney and Fletcher [16]. Contact angle values (95% confidence limits) for glass, Lamipac, tin plate, Hyfax, and stainless steel were 36.7 (5.2), 80.3 (l), 81.3 (0.9), 78.8 (1.7), and 69.4 (3.3), respectively. However, chemically clean surfaces were not desired, because that would be unlike the factory situation. To investigate the influence of adsorbed substrate on desorption, glass, nylon, and stainless steel were "conditioned" with adsorbed PYE by immersing sterile (autoclaved 15 min, 12 I~ sonicated (above) substrata in PYE broth and incubating at 25~ for 24 hours in an orbital shaking incubator (150 rpm). The surfaces were rinsed thoroughly with deionized distilled water and allowed to airdry.

Tritium Radiolabeling of Bacteria The number of attached bacteria was determined by labeling the bacteria with 3H before allowing attachment and then by measuring the amount of 3H-label associated with the surfaces using a scintillation counter. The method for radiolabeling bacteria and calibrating 3H counts per rain (cpm) to numbers of attached cells was based on methods previously described [ 16, 22]. Briefly, bacteria were harvested by centrifugation at 11,000 av. g (4~ after growth in PYE (above). The pellet was resuspended in 20 ml phosphate buffer (above) to a concentration of approximately 10 ~~ cells m1-1. The bacteria were provided with 1 #Ci L-[4,5-3H]leucine ml -~ (specific activity 46 Ci mmol-l; Amersham) and incubated at 25~ for 1 hour to allow assimilation of labeled substrate. The cells were washed three times in 20 ml phosphate buffer to remove nonassimilated labeled substrate, collected by centrifugation (11,000 av. g), and resuspended in a further 20 ml of phosphate buffer, before adjustment to the final experimental concentration. Any leakage of label was checked for as described in Fletcher [8], but repeated used of leucine in our laboratory has demonstrated that this substrate tends to be fixed stably with little leakage in the short term. A calibration curve relating cpm to attached cell numbers was then prepared [22]. Cpms of bacteria can be considerably reduced through/3-absorption by the substrata when the cells are attached to surfaces [22]. Thus, to prepare the calibrations, the bacteria had to be in contact with the relevant surfaces. Radiolabeled cells of each organism were resuspended in phosphate buffer to ten different cell concentrations ranging from 2-4 x 107 to 2-4 x 109 cells ml -~. Samples (100 ~1) of each dilution were placed on duplicate samples of each surface and allowed to dry (-40~ Individual surfaces were counted for tritium [16, 22], and quenching was corrected by the H number system [22]. A calibration curve for each surface was prepared by plotting tritium counts against the known bacterial cells numbers.

Detachment Assays Radiolabeled ceils ofA. calcoaceticus, S. aureus, or the coryneform were resuspended to concentrations of i-3 x 109 ml -~ in phosphate buffer (pH 7). Replicate (x6) 6 ml samples were added to 25 ml screw-cap bottles containing either cleaned nylon, stainless steel, glass, polypropylene, or tin plate, or PYE-conditioned glass, nylon, or stainless steel surfaces. The surfaces were held vertically in the bottles in silicon rubber tubing rings (14 m m internal diameter, 5-10 mm long). The cells were allowed to attach for 2 hours at 25~ pH 7. Loosely attached and suspended cells were then removed by passing 100 ml deionized distilled water through the bottle at a flow rate

232

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of 200 ml min -~ , controlled by a constant head. The following experimental protocols were then followed:

Attachment control. Each replicate surface was placed in a scintillation vial, dried at 40~

and counted for 3H after addition of 10 ml scintillation cocktail (2:1 [w/v] toluene : Triton X-100 and 0.6% [w/v] butyl PBD; Fisons).

Relationship between detachment and time. Replicate glass or nylon surfaces (• 6) with attached coryneform or S. aureus, respectively, were placed in sterile, clean 25 ml bottles containing 6 ml phosphate buffer at pH 7 and incubated at 25~ for 5, 15, 30, 60, 90, or 120 min to allow bacterial detachment. The surfaces were rinsed with 100 ml deionized distilled water (as above) before drying and determining aH cpms.

Relationship between detachment and pH. The substrata were transferred to sterile, clean 25 ml bottles containing 6 ml phosphate buffer at pH 5, 6, 7, 8, or 9, and incubated a~ 25~ for 2 hourg. The surfaces were rinsed and dried (above), and 3H cpms per surface were measured.

Relationship between detachment and temperature, Surfaces were placed in clean, sterile 25 ml bottles containing 6 ml phosphate buffer (pH 7) and incubated at either 4, 12, 20, 25, or 30~ for 2 hours before rinsing (above) and determining 3H cpms on dried surfaces.

PYE-conditioned surfaces and detachment. Replicate ( x 6) conditioned substrata with attached A. calcoaceticus or S. aureus were placed in 25 ml bottles containing 6 ml phosphate buffer(pH 6) and incubated at 25*C for 2 hours to allow detachment. Tritium counts were estimated for each rinsed and dried surface. The results were recorded as cells attached cm -2, and to facilitate comparison of results, data were normalized by conversion to an index of detachment (Id), which was the ratio of the n u m b e r of cells remaining attached to the test substratum divided by t h e number of bacteria attached to the control substratum. Id values of 1 were recorded when 95% confidence limits of the mean (n = 6) of the test and control surfaces overlapped; values < 1 represented detachment. All experiments were repeated at least once.

Bacterial Oxygen Consumption Organisms were harvested by centrifugation (11,000 av. g) and resuspended i n phosphate buffer at pH 5, 6, 7, 8, or 9 to a concentration of 2-3 • 109 cells ml -~. Three-ml samples were placed in an oxygen electrode chamber (RMR Brothers), maintained at 25~ and equilibrated before adding 100 tzl o f P Y E substrate and determining oxygen consumption. The results were expressed as nmoles oxygen concentration in air-saturated buffer suspension of 268 nmol ml-I at 25"C.

Results E f f e c t o f T i m e on B a c t e r i a l D e t a c h m e n t The detachment of the coryneform from glass substrata increased with time at p H 7, b u t S. a u r e u s d i d n o t d e t a c h f r o m a n y l o n s u b s t r a t u m a f t e r 1 2 0 m i n a t p H 7 ( F i g . 1).

E f f e c t o f p H on B a c t e r i a l D e t a c h m e n t The desorption of bacteria from surfaces varied, depending on pH, species, a n d s u b s t r a t u m c h a r a c t e r i s t i c s ( T a b l e 1). T h e c o r y n e f o r m d e s o r b e d t o t h e g r e a t e s t e x t e n t , d e t a c h i n g i n m o s t e x p e r i m e n t a l c o n d i t i o n s , i n c o n t r a s t , kS. a u r e u s o n l y d e t a c h e d f r o m g l a s s a t p H 6 a n d 9. I n s o m e c a s e s , s u b s t r a t u m c h a r a c t e r i s t i c s i n f l u e n c e d b a c t e r i a l d e s o r p t i o n ; f o r e x a m p l e , A. calcoaceticus d i d n o t

Bacterial D e s o r p t i o n from Surfaces

,,, 9- t

q,d

233

301

,~oE u 20 O,,,o ..CO

or

Fig. 1. D e t a c h m e n t o f the c o r y n e f o r m from glass (O) a n d o f S. a u r e u s from nylon (0) at p H 7 with time. T h e error bars represent 95% confidence limits o f the m e a n .

10

3'0

6'0 9'0 Time ( r a i n )

Table 1.

Effect o f p H on the d e t a c h m e n t o f bacteria from surfaces I n d e x o f d e t a c h m e n t (Id) a

pH

Glass

Polypropylene

Coryneform

5 6 7 8 9

1 0.3 0.45 0,68 1

0.15 0;17 0.14 0.27 0.2t

0.49 1 0.45 1 1

S. a u r e u s

5

1

1

1

1

1

6 7 8 9

0.58 1 I 0.07

1 1 I 1

1 1 i 1

1 1 1 1

1 1 1 1

5 6 7 8 9

0.24 1 0.36 1 1

0.36 0.36 0,21 1 0.28

1 1 1 1 1

1 1 0.31 1 0.31

1 1 0.03 1 1

Bacterium

A. c a l c o a c e t i c u s

Tin plate

Nylon

Stainless steel

1 0.5 0.35 0.52 1

1 1 0.03 0.25 0.07

" I n d e x o f d e t a c h m e n t (Id) was calculated as n u m b e r o f cells rem a i n i n g . a t t a c h e d t o the test surface after the d e t a c h m e n t period/ n u m b e r o f bacteria a t t a c h e d to the control surface. Id values o f l were recorded for t r e a t m e n t s w h o s e 95% confidence limits o f the m e a n (n = 6) o v e r l a p p e d with the controls; values < 1 represent detachment

detach at any pH from tin plate but detached from polypropylene at every pH except pH 8. There was no relationship between the effect of pH on bacterial respiration and on bacterial detachment from the substrata (Tables 1, 2).

Effect of Temperature and P YE :conditioned Substrata on Bacterial Detachment There were n o variations in levels of detachment that corresponded with progressive changes in temperature (Table 3). Similarly, desorption of S. aureus

S. McEldowney and M. Fletcher

234 Table 2. tion

Effect of pH on bacterial oxygen consumpOxygen consumption (nmol Oz ml-t min -~) A.

pH

calcoaceticus

Coryneform

S. aureus

5 6 7 8 9

4.0 12.1 34.2 20.8 16.1

4.7 15.4 18.1 18.1 18.1

27.3 48.2 36.8 39.3 34.8

Table 3. Effect of temperature on bacterial detachment from stainless steel and nylon Index of detachment (I~)~ Temperature (~

Stainless steel CoryneS. form aureus

4 12 20 25 30

0.63 0.5 0.49 0.45 0.51

1 1 1 1 1

Nylon S. aureus

A. calcoaceticus

1 1 1 1 1

0.39 0.46 0.25 0.39 0.39

" See Table 1 from glass, nylon, and steel and o f A . c a l c o a c e t i c u s from nylon and steel was the same for both P Y E - c o n d i t i o n e d and u n c o n d i t i o n e d surfaces.

Discussion

Bacterial adhesion to solid surfaces is often a nonspecific adsorption process which involves a c o m b i n a t i o n o f long-range forces, e.g., electrostatic interactions and short-range interactions chemical b o n d s and h y d r o p h o b i c interactions, e.g., between the cell and substratum surfaces [15, 23]. Therefore, if desorption occurs subsequent to attachment, it is p r e s u m a b l y due to changes in, or dissipation of, the short-range attractive forces that p r o m o t e adhesion. Such changes could be brought a b o u t through physiologically induced m o d i fications in the cell surface or localized changes in physicochemical factors at the substratum-cell interface. In this study, the a m o u n t o f desorption differed considerably with the species and substratum composition. This is p r o b a b l y not surprising, as the variation in bacterial adhesion with species [16, 22, 28] a n d substratum c o m p o s i t i o n [7, 22, 26] is well d o c u m e n t e d . The difference in a t t a c h m e n t ability o f various species is determined, at least in part, by differences in cell surface composition. Bacterial surfaces, and accordingly their adhesiveness, vary with growth condition and growth rate [ 16], and similar differences have been induced by periods o f starvation [10]. The

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235

nature of the change is species dependent [ 16]. In the experimental conditions used in this study, i.e., incubation in the absence of nutrients, detachment may have been related to such species-specific changes in cell surface characteristics, resulting in a loss of adhesion and subsequent cell desorption. The desorption characteristics of bacteria from Dowex anion exchange resin also were found to vary with species [28]. The number of bacteria that become attached to a surface depends upon substratum characteristics, e.g., electrostatic charge and hydrophobicity [12, 26]. Accordingly, the desorption of bacteria from surfaces might also be expected to be influenced by such surface properties. Moreover, there is some evidence suggesting separate adhesion mechanisms to hydrophobic and hydrophilic substrata [9, 16, 20], and desorption from these substrata may also be different. It is not yet clear whether bacterial desorption is favored from any particular type of surface, but there is considerable evidence that the strength of both polymer adsorption and cell adhesion is dependent upon substratum surface tension [2]. Van Pelt et al. [26] found greater desorption o f Streptococcus sanguis from hydrophobic than hydrophilic surfaces. However, in this study, bacteria frequently detached from glass, a charged hydrophilic surface, with S. aureus detaching from only this surface. Similarly, it has been found that Bacillus cereus bound more strongly to hydrophobic surfaces [21]. However, in order to draw any conclusions from such comparisons, clean, well-characterized substrata must be used, as was not the case in this study. Solution pH has altered bacterial adhesion to solid surfaces [ 18, 25], and in this study, it also influenced bacterial desorption (Table 2). Changes in dissociation of charged bacterial surface groups [ 14, 21 ], or in the steric arrangement of cell surface molecules, may modify adhesive interactions between cell and substratum. Because of the differences in bacterial and substratum surface composition, pH-induced desorption might be expected to vary both with species and solid surface, as was found (Table 2). Other workers have found similar pH effects on desorption. For example, a marine pseudomonad detached from glass with large pH changes [5], but only Escherichia colL and not Aerom o n a s proteolytica nor Flavobacterium oceanosedimentum, desorbed from sediment [24]. Similarly, pH changes had little effect on the desorption of four freshwater strains, with only limited detachment of Enterobacter cloacae and a Flexibacter sp. from polystyrene surfaces [17]. In this study, S. aureus was unusual in that its adhesion was quite stable and resisted desorption with time. The persistence of its adhesion suggests either a comparatively strong adhesive interaction between cell and substratum or less cell surface variability associated with changes in pH. Similarly, S. aureus adsorbed on Dowex anion exchange resin was not completely desorbed by a NaC1 gradient solution, unlike B. cereus [28]. One of the ways changes in pH might affect desorption is through modifications of bacterial metabolic activity, resulting in physiologically induced changes in surface properties. However, since there was no relationship between the oxygen consumption of any species and its detachment at the range of pH values tested, it is unlikely that physiological activity had a direct influence on desorption. The relationship between physiological activity and adhesion is not understood. Although activity seems to promote adhesion in some cases

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s. McEldowney and M. Fletcher

(cf. [15]), it is not always necessary. M o r e o v e r , detached cells h a v e been f o u n d to be b o t h m o r e active [8] a n d less active [3] t h a n c o m p a r a b l e bacteria that r e m a i n e d attached. T e m p e r a t u r e h a d no influence on the degree o f desorption. H o w e v e r , t e m perature has b e e n s h o w n to affect the d e s o r p t i o n o f other bacteria, as b o v i n e t u r e e n bacteria were d e s o r b e d f r o m starch granules b y cooling after adhesion at 38~ [19]. Since, within limits, bacterial activity increases with increased t e m p e r a t u r e , the lack o f influence o f t e m p e r a t u r e on d e s o r p t i o n in this study adds further weight to the suggestion (above) that with these organisms, det a c h m e n t or the m a i n t e n a n c e o f a d h e s i o n is not a physiologically d e p e n d e n t process. T h e irreversible a d s o r p t i o n o f organic molecules, producing a conditioning film [ 1], on a solid s u b s t r a t u m results in a new set o f surface characteristics for interaction with the bacterium. Such conditioning layers can affect adhesion [1, 9] but did not a p p e a r to influence subsequent d e s o r p t i o n in this study. H o w e v e r , this does not necessarily indicate that conditioning films per se do not affect d e t a c h m e n t . T h e s e surfaces were not rigorously cleaned before the experiments, so that they w o u l d simulate those surfaces as f o u n d in a food factory situation. Consequently, they were not chemically clean and, in effect, were already coated with an organic conditioning film. T h e d e s o r p t i o n o f bacteria f r o m surfaces in food processing plants has i m p o r t a n t implications. First, the ease with which such bacteria desorb will influence their susceptibility to r e m o v a l b y chemical t r e a t m e n t s or shear forces. T h e results o f this study suggest that the efficacy o f cleaning t r e a t m e n t s m a y well v a r y with the colonizing o r g a n i s m s a n d the surface c o m p o s i t i o n . T h e y also suggest that p H t r e a t m e n t s m a y not be successful in r e m o v i n g bacteria. Second, where attached bacteria h a v e resisted cleaning, the s u b s e q u e n t desorption o f those cells o r their progeny will allow the cells to be spread b y liquids m o v i n g o v e r those surfaces a n d result in c o n t a m i n a t i o n o f o t h e r surfaces. T h e s e results d e m o n s t r a t e that with certain strains, limited d e s o r p t i o n readily occurs a n d p r o v i d e s a potential route for cross-infection o f food processing surfaces.

Acknowledgments. This work was supported by Project Grant no. 234 from the Ministry of Agriculture, Fisheries, and Food. We thank Pam Banks for outstanding technical assistance and Colin Dennis, Roy Thorpe, David Jonas, Peter Bean, and David Shapton for very helpful discussions. References 1. Baier RE (1981) Substrata influences on adhesion of microorganisms and their resultant new surface properties. In: Bitton G, Marshall KC (eds) Adsorption of microorganisms to surfaces. John Wiley and Sons, New York, pp 59-104 2. Baler RE (1982) Conditioning surfaces to suit the biomedical environment: recent progress. J Biomech Eng 104:257-271 3. Bright JJ, Fletcher M (1983) Amino acid assimilation and electron transport system activity in attached and free-living marine bacteria. Appl Environ Microbiol 45:818-825 4. Characklis WG (1984) Biofilm development: a process analysis. In: Marshall KC (ed) Microbial adhesion and aggregation. Springer-Verlag, Berlin, pp 137-157 5~ Corpe WA (1974) Detachment of marine periphytic bacteria from surfaces of glass slides. Dev Indust Microbiol 15:281-287

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6. Costerton JW, Cheng K-J (1982) Microbe-microbe interactions at surfaces. In: Bums RG, Slater JH (eds) Experimental microbial ecology. Blackwell Scientific Publishers, Oxford, England, pp 275-290 7. Dexter SC (1977) Influence of substrate wettability on the formation of bacterial slime films on solid surfaces immersed in natural sea water. In: Romanovsky V (ed) Proc 4th Intl Cong on Marine Corrosion and Fouling, Centre de Recherches et d'Etudes Oceanographiques, Boulogne, France, pp 137-144 8. Fletcher M (1986) Measurement of glucose utilization by Pseudomonas fluorescens that are free living and that are attached to surfaces. Appl Environ Microbiol 52:672-676 9. Fletcher M, Marshall KC (1982) Bubble contact angle method for evaluating substratum interfacial characteristics and its relevance to bacterial attachment. Appl Environ Microbiol 44:184-192 10. Kjelleberg S, Hermansson M (1984) Starvation-induced effects on bacterial surface characteristics. Appl Environ Microbiol 84:497-503 11. Kovacs N (1956) Identification of Pseudomonas pyocyanea by the oxidase reaction. Nature 178:703-708 12. Ludwicka A, Jansen B, WadstriSm T, Pulverer G (1984) Attachment of staphylococci to various synthetic polymers. ZBI Bakt Hyg A 256:479-489 13. MacFaddin JF (1976) Biochemical tests for identification of medical bacteria. Williams and Wilkins Co, Baltimore 14. Marsha•• KC ( • 96 7) E•ectr•ph•retic pr•perties •f fast- and s••w.gr•wing species •f Rhiz•bium. Aust J Biol Sci 20:429-438 15. Marshall KC (1985) Mechanisms of bacterial adhesion at solid-water interfaces. In: Savage DC, Fletcher M (eds) Bacterial adhesion. Plenum Press, New York, pp 133-161 16. McEldowney S, Fletcher M (1986) Effect of growth conditions and surface characteristics of aquatic bacteria on their attachment to solid surfaces. J Gen Microbiol 132:513-523 17. McEldowney S, Fletcher M (1986) Variability in the influence of physicochemical factors affecting bacterial adhesion to polystyrene substrata. Appl Environ Microbiol 52:460-465 18. Minato H, Suto T (1976) Technique for fractionation of bacteria in rumen microbial ecosystem. I. Attachment of rumen bacteria to starch granules and elution of bacteria attached to them. J G e n Appl Microbiol 22:259-276 19. Minato H, Suto T (1979) Technique for fractionation of bacteria in rumen microbial ecosystem. III. Attachment of bacteria isolated from bovine rumen to starch granules in vitro and elution of bacteria attached therefrom. J Gen Appl Microbiol 25:71-93 20. Paul JH, Jeffray WH (1985) Evidence for separate adhesion mechanisms for hydrophilic and hydrophobic surfaces in Vibrio proteolytica. Appl Environ Microbiol 50:431-437 21. Poweli SM, Slater NKH (1982) Removal rates of bacterial cells from glass surfaces by fluid shear. Biotech Bioeng 24:2527-2537 22. Pringle JH, FletcherM (1983) Influence of substratum wettability on attachment offreshwater bacteria to solid surfaces. Appl Environ Microbiol 45:811-817 23. Rutter PR, Vincent B (1984) Physicochemical interactions of the substratum, microorganisms, and the fluid phase. In: Marshall KC (ed) Microbial adhesion and aggregation. Springer-Verlag, Berlin, pp 21-38 24. Scheraga M, Meskill M, Litchfield CD (1979) Analysis ofmethods for the quantitative recovery of bacteria sorbed onto marine sediments. In: Litchfield CD, Seigfried PL (eds) Methodology ofbiomass determinations and microbial activities in sediments. American Society for Testing & Materials Special Technical Publication 673, pp 21-39 25. Stanley PM (1983) Factors affecting the irreversible attachment of Pseudomonas aeruginosa to stainless steel. Can J Microbiol 29:1493-1499 26. Van Pelt AWJ, Weerkamp AH, Uyen MHWJC, Busscher HJ, de Jong HP, Arends J (1985) Adhesion of Streptococcus sanguis CH3 to polymers with different surface-free energies. Appl Environ Microbiol 49:1270-1275 27. Zvyaglntsev D G (1959) Adsorption of microorganisms by glass surfaces. Microrirlogy (USSR) (Eng trans) 28:104-108 28. Zvyagintsev DG, Guzev VS (1971) Concentration and separation of bacteria on Dowex anionite. Microbiology (USSR) (Eng trans) 40:139-143

Bacterial desorption from food container and food processing surfaces.

The desorption ofStaphylococcus aureus, Acinetobacter calcoaceticus, and a coryneform from the surfaces of materials used for manufacturing food conta...
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