MB 495 Lab Project
Can Tetracycline Resistance be Induced in Pseudomonas Fluorescens After Three
Weeks?
David S. Abuín & Tania Bembridge
Department of Microbiology, North Carolina State University
There was a comment made during an ICAAC podcast stating
that pseudomonas “liked” tetracycline. The comment
prompted this study on how much pseudomonas actually
“liked” tetracycline and when is resistance induced. In a
three week study we tested Pseudomonas fluorescens against
a variety of concentrations to see how P. fluorescens reacted
to this broad spectrum antibiotic.
One of the controls was using a concentration that P.
fluorescens would not grow in. 20.0µl/ml (0.42mM) was
enough to prevent any growth of the bacteria each week.
However, when we took the bacteria from a lower
concentration and added it to a higher concentration of
tetracycline, it showed resistance. The growth rate,
maximum growth, and lag time were mildly affected.
Fig 1. The chemical structure of
tetracycline. Tetracycline inhibits
protein synthesis within Pseudomonas
by blocking the binding of tRNA to the
mRNA ribosomal complex.
The experiments consisted of
growing P. fluorescens in five different concentrations of
tetracycline. These ranged from 0.50µl/ml to 20.0µl/ml. Each
week a sample was taken from the previous week and added
to fresh media and tetracycline. Low concentrations of
0.50µl/ml and 1.00µl/ml tetracycline did not strongly inhibit
the growth of P. fluorescens. The growth rate, maximum
growth (maximal OD), and lag time were affected but not
significant enough to show a marked change from a nontetracycline environment (data not shown). Here P.
fluorescens didn’t show much “like” for tetracycline as
overpower it.
The concentrations where tetracycline showed inhibitory
effects were 5.00µl/ml (0.10mM) and 10.0µl/ml (0.20mM).
The first week P. fluorescens struggled to grow in tetracycline
taking over 48 hours in one instance. After letting the
bacteria sit for a week then testing it again, it showed strong
resistance to the tetracycline. This trend continued through
the third week. Lag time was reduced, while maximum
growth and growth rate increased.
Fig 3. The result s of increasing the tetracycline concentration on
bacteria that is already displaying resistance.
The Psuedomonas genus has numerous genes for tetracycline
resistance and it appears after one week those genes are
activated.
A follow up experiment would be to check how long the
resistance genes are produced when tetracycline is absent.
References
1.
Goshe, Amy and Sasha Minium. Tetracycline Repressor
Complex. Ed. Daniel Barich. 2007. Biomolecules at Kenyon.
Kenyon College.
http://biology.kenyon.edu/BMB/Jmol2007/Tet/index2.html.
2.
Li, Xiaojing and Hua H. Wang. Tetracycline Resistance
Associated with Commensal Bacteria from Representative
Ready-to-Consume Deli and Restaurant Foods. 2010. Journal of
Food Protection. 73(10): 1841–1848.
3.
Mehta, Akul. Mechanism of Action of Tetracyclines. 27 May
2011. PharmaXchange.info.
http://pharmaxchange.info/press/2011/05/mechanism-ofaction-of-tetracyclines.
4.
Thaker, Maulik, Peter Spanogiannopoulos, and Gerard D.
Wright. The tetracycline resistome. 2010. Cellular and
Molecular Life Sciences. 67:419–431.
Fig 2. The growth of P. fluorescens in 10µl/ml (0.20mM) tetracycline
each week. The error bars so the maximum and minimum values for
the obtained results.
December 2011