The Bacterial Cell and Attachment

The different bacterial Isolates attached differently to the glass

coversllps with no relationship between the generic classification and

attachment. Similar results were reported in the literature

(Zvyagintsev, 1979) and these different attachment abilities must be due

to the bacteria varying in other factors such as cell surface

characteristics or In physiological activity.

Xotlle and non-motlle Isolates attached to the surfaces in these

experiments. However, It would be wrong to conclude that motility Is not

Important In the attachment process. In situations, within

macroenvironments of flowing systems, the Importance of motility to

attachment Is questionable, however, when considering microenvironments

the presence of a hydrodynamic boundary layer could Increase the

significance of motility In attachment with motility posslblly Involved

In carrying the bacteria against the direction of flow to allow

attachment to the surface (Caldwell, 1984). Piette (1990) has suggested

that motility Indirectly helps bacterial attachment to solid surfaces by

Increasing the number of bacteria reaching the surface In a given time.

This phenomenon is Important if the surface is beneficial to the

bacterium's growth, e.g. If the surface supplies nutrients for bacterial

Jtotility can also result in the bacterium arriving perpendicular to

the surface which has been shown to be Important in the initial

attachment process (Lawrence, 1987a). Motility can also be Important

once the bacterium has arrived at a surface. After the bacterium has

become reversibly attached, it must overcome the repulsion barriers

present at the surface to become irreversibly attached (Marshall, 1974j

Fletcher, 1984) and it is possible that motility could aid the bacterium

in this. Reports have suggested that attachment is an active process

facilitated by motility and therefore an increase in attachment could be

obtained with bacterial motility (Lawrence, 1987a). It could also be due

to motile bacteria coming randomly into contact with the surface more

often than non-motile bacteria. There could also be other factors

involved indicated by the fact that non-motile cells can also attach to

the surfaces like their motile counterparts (Lawrence, 1987b>.

There have also been reports of bacteria attaching to solid surfaces

by their polar flagella (Meadows, 1971) with the flagellum being the

initial point of contact between the bacterium and the solid surface.

This interaction could be due to the diameter of the flagellum being

smaller than that of the bacterium and possibly allowing a lowering of

the repulsion barrier between the bacterium and the solid surface

(Rogers, 1979). This initial attachment with the flagellum may not last,

and for irreversable attachment, the surface of the bacterium will have

to come into contact with the solid surface (SJoblad, 1982).

As the bacterial surface Itself acts as a point of contact between

the bacterium and the solid surface, the different cell surface

characteristics of the bacterium could be Important in the attachment

levels displayed by bacteria can be attributed to their different cell

surface characteristics (McEldowney, 1986). In these studies, different

attachment levels were obtained due to the surface components reacting

differently at a physlochenlcal level with the solid surface (Chapter

4 ) .

Physiological activity can also be an important bacterial factor to

consider in attachment studies. The effect of this activity on

attachment can again vary with the bacterial species involved and it is

also possible the physiological activity can influence other important

attachment factors such as cell surface characteristics. The

physiological activity of a bacterium can itself be affected by factors

such as growth conditions (Fletcher 1984). In all the attachment

experiments, the cells were grown to early stationary phase, this was

necessary as slight changes in the growth stage lead to significant

changes in bacterial attachment (Fletcher, 1977; Zvyagintsev, 1973;

Xinato, 1979). Feldner (1983), demonstrated that bacteria in different

growth stages were affected differently by proteins present on the

attachment surface. The attachment of early log phase or stationary

phase cultures was more inhibited by proteins on the surface than other

growth phases. Stationary phase was chosen in these attachment

experiments as a constant state, although in practice some species of

bacteria may have attached better or worse in other growth phases.

As the aim of subsequent experiments was to investigate attachment

under different growth conditions such as carbon source, a brief survey

of bacterial attachment in different carbon sources was carried out to

select bacteria for these future experiments. The results (Table 3.1),

sources as was expected from the literature (McEldowney, 1986).

Caldwell (1986), used different growth conditions to select for bacteria

which could attach to surfaces. This procedure was not used in these

studies as the bacteria isolated did not vary significantly in

attachment abilities. The bacteria selected for future use were chosen

as their attachment abilities to surfaces varied significantly. The

bacterial morphology on attachment to surfaces was also studied.

Bacterial cells have been shown to decrease in size during attachment

(Caldwell, 1966; Marshall, 1980) or elongate after attachment (Lawrence,

1987b). Bacteria which were seen to vary in cell morphology on

attachment were not selected for subsequent experiments as these

morphology changes could hinder bacterial identification after

attachment.

3.5.2 Detachment

The detachment of bacteria from a surface will be influenced by the

bacterial species present on the surface, the surface itself and the

surfactant used (Scheraga, 1979). During these studies, the detachment

rates were not changed by varying the pH or electrolyte concentration

used. This was thought to be due to a strong bonding between the

bacteria and the surface. As bacteria which are strongly attached are

hard to detach, numerous methods of detachment have been tried.

Antimetabolites have been shown to decrease the attachment of a

Vibrio species to surfaces. This process usually resulted in the non-

attached bacteria being non-viable, therefore, the numbers of these non-

attached bacteria obtained would be hard to determine (Paul, 1984). iron-

viable non-attached bacteria were also obtained when taurolln, an non-

bacteria to surfaces. When taurolln was used in sub-minimal inhibitory

concentrations, the adherence of E.coll and Canidlda albicans was

reduced or prevented. To influence the attachment of a Staphylccnccns

species, much higher concentrations of taurolln were required. This

process resulted in the bacterial cells being unable to complete cell

division, losing their fimbriae and becoming elongated. Due to the non-

attached cells being non-viable and their morphology changing on

detachment, this process again made the numbers of non-attached cells

hard to determine. The above processes only influenced bacterial

attachment before attachment occurs, therefore, were not suited to

detach bacteria when they had already attached.

Golllnge (1985), suggested using monoclonal antibodies to detach

bacteria from surfaces. Studies Indicated that the attachment of

Bordetella pertussis was influenced by monoclonal antibodies, however,

the detachment of these bacteria by these antibodies was not reported.

Degradatlve enzymes such as pronase and trypsin detached some bacteria

from surfaces, the proportion of detachment obtained however, again

depended on the surfaces and the species used in the experiments (Corpe,

1974b; Fletcher, 1980; Danielsson, 1977). The attachment of

Streptococcus faecium to surfaces was seen to be reversed by the

surfactant Tween-80 (Orstavik, 1977). Surfactants are amphiphilic

molecules composed of a hydrophobic portion and a charged or polar

portion. They include anionic and cationic detergents and non-ionic

types. These surfactants have been used by other researchers in

detachment experiments with varying degrees of success (Ball, 1986).

These surfactants were used in these detachment experiments as at

Problems can 6tlll arise when these surfactants are used, because

cell lysis has been reported to be Involved in detaching cells from

surfaces (Corps, 1974b). Many external factors such as the length of

time of the attachment or detachment (Fletcher, 1977) and the

temperature at which the procedure Is performed (Berger, 1966) have been

shown to influence attachment and possibly detachment.