VOLUM OMAGIAL - Facultatea de Ştiinţe ale Naturii şi Ştiinţe Agricole

Aurelia Manuela Moldoveanu, Ioan Ardelean / Ovidius University Annals, Biology-Ecology Series 14: 147-156 (2010)
minute. Afterwards, they were abundantly washed
twice with osmosis water in order to eliminate the
excess of coloring matter [27].
The sample investigation was done by means of
the Hund microscope, cell counting being done using
an ocular grid, calibrated according to the standard
procedure [28]; cells within 20 microscopic fields
per each sample were counted, according to the
standard counting procedures for the surface bacteria
[29]. Thus, the values of cellular density are
expressed on the graphs represented by the mean of
the 80 microscopic fields per each sample.
3. Results and Discussions
3.1 The chemical analysis of water
There are differences between the two types of
culture media used for the generation of bacterial
biofilms on the hydrophile surface of glass slides and
in order to emphasize their existence we analyzed the
seawater samples in the Chemistry Laboratory within
the “Grigore Antipa” Marine Research Institute of
Constanta (Table 1).
The chemical analysis of seawater emphasized
the existence of differences among the chemical
parameters: salinity, pH, concentration of inorganic
substances between the two types of seawater used.
The littoral seawater has normal parameters also
registered in previous years [30], but the aquarium
seawater has values well over the normal limit with
an increase of over 10g/l of salinity and a decrease of
pH from 8.12 (normal value for littoral seawater) to
6.56 units for the aquarium water, almost two units
less than the initial value. The concentration of
inorganic substances is well above the normal one for
seawater. The concentration of nitrates is three times
higher compared to the normal value, while the
concentration of polyphosphates is over 84 times
higher.
The existence of these differences between the
two used culture media can cause changes in the
formation manner of bacterial biofilms in liquid
medium, as well as the temporal dynamic of their
formation. The bacterial biofilms formed are an
assemblage of surface-associated microbial cells that
149
is enclosed in an extracellular polymeric substance
matrix.
3.2 The formation of biofilms under the
influence of temperature
Figure one shows the values of cellular density
obtained after the modification of the temperature
factor for the biofilms formed on the hydrophile
surface of glass slides and collected from the
containers with littoral seawater kept at a constant
temperature of 18ºC and 6 ºC, respectively.
The data analysis emphasized the existence of
successive stages for the formation of biofilms. Thus,
in the case of the biofilms formed at 18 ºC, one hour
after the slides immersion the cellular density is
12∙10 2 cel/mm 2 . This value doubles eight hours later
to 25 ∙10 2 cel/mm 2 and increases progressively to a
tendency to triple the cellular density to 37 ∙ 10 2
cel/mm 2 11 hours later. After 12 hours, during which
the slides were left over night, the following day the
cellular density reaches the value of 45∙10 2 cel/mm 2
24 hours after immersion. The value increases
progressively to 60∙10 2 cel/mm 2 36 hours after
immersion
For the seawater in the containers kept at 6 ºC in
the refrigerator, there is a progressive increase from 5
∙10 2 cel/mm 2 only one hour after immersion and a
doubling of this value seven hours later to 10 ∙10 2
cel/mm 2 , as well as tripling to 15∙10 2 cel/mm 2 eight
hours later. The following day, after 12 hours, the
cellular density was 41∙10 2 cel/mm 2 and increased
progressively to 54∙10 2 cel/mm 2 .
The progression of cellular density growth is
over 2.3 for the biofilms formed at 18 ºC during the
first 12 hours and below 1.2 after 24 hours. Also, in
the case of the biofilms formed in containers kept at 6
ºC, the progression is over 1.9 during the first 12
hours and below 1.1 after 24 hours. On the first day,
after 12 hours, there is a difference of approx. 9∙10 2
cel/mm 2 between the two progressions of density
growth, depending on temperature. 24 hours later, the
difference is below 8∙10 2 cel/mm 2 and 36 hours later
it rises to 10∙10 2 cel/mm 2 .
A number of experiments regarding bacterial
adhesion were accomplished on different types of
surfaces (copper, PVC and polybuten) by Rogers [31]
at different temperatures (20 ºC, 40 ºC, 50 ºC and 60