Hi,
I've just finished my mini-assignment on biofilms and their utilisation
of glucose and phenol. Here are my methods and results and discussion.
Please feel free to give comments/suggestions/advice.
My objective is to see whether phenol or glucose is an inducer for
aggregate formation.
And lastly, to check if a higher aggregate count would give a lower
viable count.
Methods
Preparation of Special Bacterial Mixture (Primed Inoculum)
Sample ID Bacteria A6017 Z6011 Z6013 3 types 11 types 26 types
A5019ST B. cereus 96.2 ul
A6017 Ps. aeruginosa 2500ul 833.3ul 227.3 ul 96.2 ul
A6030 Achromobacter spp. 96.2 ul
B501 Ps. cepacia 96.2 ul
B509PT A. odorans 96.2 ul
B5011 B. primus 227.3 ul 96.2 ul
B5012ST B. primus 227.3 ul 96.2 ul
B5013 Acinetobacter spp. 96.2 ul
B6013 Ps. thorecen-25 96.2 ul
C505PT Ps. aeruginosa 96.2 ul
C5011ST Flav. odoratum 227.3 ul 96.2 ul
C5012ST B. sphaericus 96.2 ul
C5014 B. megaterium 96.2 ul
C601-1 Ps. malthophilia 96.2 ul
C601-2 Ps. malthophilia 96.2 ul
C602 Flav. odoratum 96.2 ul
C603 Ps. fluoresc/ putida 96.2 ul
C605 Flav. odoratum 227.3 ul 96.2 ul
C6011 Acinetobacter spp. 227.3 ul 96.2 ul
C6019 Aer. salmonicida 96.2 ul
Z507ST Sphmon. Paucimobilis 227.3 ul 96.2 ul
Z508SC A. anitratus 96.2 ul
Z6011 Flav. orzihabitans 2500ul 833.3ul 227.3 ul 96.2 ul
Z6013 Acinetobacter spp. 2500ul 833.3ul 227.3 ul 96.2 ul
Z6020PT B.brevis 227.3 ul 96.2 ul
Z6030PT Ps. aeruginosa 227.3 ul 96.2 ul
How to interpret the above table: 1 shaker flask contained 2500ul of Ps.
aeruginosa(A6017), Another flask contain 2500 ul Z6011 Flav.
orzihabitans, Another flask contain Flav. orzihabitans, Acinetobacter
spp., Ps. aeruginosa of 833.3 ul each and so on.
In addition, 2500ul of E.coli was inoculated into nutrient broth and a
control was included.
Discussion
Aggregation and cell count
--------------------------
Acinetobacter and Flavi oryzihabitans were that the aggregate count for
both were the highest. It appeared that these microcolony-colony forming
bacteria tend to have a trend whereby a high aggregate count will give
will give a low viable count.
A possible reason for that trend is that the daughter cells are enmeshed
within the microcolony together with the parent cells. The daughter cells
are probably unable to dislodge from the micro-colony due to attachment
of EPS (exopolysaccharide). Thereby giving one unusual colony on the
enumeration agar plate. Comments?
In constrast, Ps. aeruginosa was found to have a high bacteria growth
rate and low aggregate count. In this case, since no or very little
aggregates were formed, the daughter cells would exist as dispersed cells
in the suspension. Thus, viable count is higher. Comments?
Among the mixed cultures, 26 types of bacteria combination gave the best
performance in aggregate formation. Very little or no aggregates were
observed for 3 or 11 types of bacteria combination. The reason that the
mixture of 3 types and 11 types of bacteria did not form much aggregates
although they have Acinetobacter and Flavi. Oryzihabitans is not clear.
Comments?
Aggregates formed by pure cultures were more closely packed than mixed
cultures. Comments?
In general, the bacteria were constantly using glucose because their
concentration was constantly decreasing.
Phenol is not an inducer for aggregate formation.
------------------------------------------------
In a previous study, it was speculated that phenol was acting as an
inducer for aggregate formation in Acinetobacter, Flavi oryzihabitans and
the 26 types of bacteria. Acinetobacter and Flavi orzihabitans reduced
97% phenol readily after 23 hours. The other culture capable of reducing
97% phenol was the 26 bacteria after 95 hours.
The above 3 cultures also formed the most aggregates.
To verify the speculation, the experiment was repeated with the same 26
cultures but this time glucose was used instead. To our surprise,
aggregate formation was also seen in the same cultures as mentioned
above. This might imply that it is a natural ability for these bacteria
to form micro-colonies. Comments?
Conclusion
-----------
Throughout the study, it appeared that the aggregates formed by pure and
mixed cultures differed in arrangement. Possible reasons could be that in
mixed cultures, there were bacteria of different sizes and shapes,
different gram stain properties, surface charges, surface
hydrophobicity/hydrophilicity and some might have the ability to secrete
exopolysaccharide.
Comments?
Regards
Lee
silkwood at cyberway.com.sg
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