Transcription Analysis of
Tetracylcine Resistant Genes in
Chlamydia suis
Presented by
Erika K. VanDenBerg
Mentor
Dr. Dan Rockey
Department of Microbiology
What is Chlamydia?
• Chlamydia is a bacterium
• Obligate intracellular pathogen
Chlamydia trachomatis (C. trachomatis)
• #1 Sexually
transmitted disease
(STD) in U.S.
• 3 million Americans
become infected
yearly
• NO SYMPTOMS
• Causes sterility if untreated and can lead to
life-threatening problems
C. trachomatis
• #1 cause of preventable blindness worldwide
-500 million people suffer from trachoma
C. pneumoniae
• 10-20% of pneumonia worldwide
• Associated with1. Coronary atherosclerosis
2. Heart disease
• Chlamydia suis ( C. suis) found in all farmed
pigs
• C. suis has acquired tetracycline (tet)
resistance
• TET is a class of antibiotic, inexpensive, and
commonly used to treat chlamydial infections
• Over 50 yrs TET has been added to animal
feed in high doses evolving microbes to
acquire resistance to antibiotics
Significance of C. suis acquiring
tet-resistance
1.
First example of genes recombining into
Chlamydia or any other obligate
intracellular pathogen.
2.
Resistance could eventually occur in the
human pathogens.
C. suis tet-resistant genes tet R and tet C share its
operator sequence. Plasmids pSC101 and pRAS have
these genes as well.
Mechanism for plasmids pSC101 and pRAS is known.
•TET is present tet C is being transcribed
• TET is not present tet C is not transcribed
The induction of tet C was analyzed using
pSC101 (in E. coli), pRAS (in E. coli), and
tetracycline resistant C. suis by performing
Reverse Transcriptase Polymerase Chain
Reaction (RT-PCR) to analyze transcription in
the presence and absence of TET.
E. coli+pSC1011) 100 base pair Ladder
1 2 3 4 5 6 7 8 9 10 11
2) tet C expressed in presence of TET
3) tet C not expressed in absence of TET
4) Negative control (DNA) for tet C in
presence of TET
5) Negative control (DNA) for tet C in
absence of TET
6) Positive control for tet C
7) tet R expressed in presence of TET
8) tet R expressed in absence of TET
9) Negative control (DNA) for tet R in
presence of TET
E. coli+pRAS had
same results
10)Negative control (DNA) for tet R in
absence of TET
11) Positive control for tet R
E. coli+pSC1011) 100 base pair Ladder
2) tet C expressed in presence of TET
1 2 3 4 5 6 7 8 9 10 11
3) tet C not expressed in absence of
TET
4) Negative control (DNA) for tet C in
presence of TET
5) Negative control (DNA) for tet C in
absence of TET
6) Positive control for tet C
7) tet R expressed in presence of TET
8) tet R expressed in absence of TET
E. coli+pRAS had
same results
9) Negative control (DNA) for tet R in
presence of TET
10)Negative control (DNA) for tet R in
absence of TET
11) Positive control for tet R
1, 2) Positive control for presence of chlamydial RNA
C. suis R19
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15
3, 4) Negative control for presence of DNA in
chlamydial RNA
5) PCR of R19 gDNA
6) tet R expressed in absence of TET
7) tet R expressed in presence of TET
8) Negative control (DNA) for tet R in absence of
TET
9) Negative control (DNA) for tet R in presence of
TET
10) Positive control for tet R
11) tet C expressed in absence of TET
12) tet C expressed in presence of TET
13) Negative control (DNA) for tet C in absence of
TET
14) Negative control (DNA) for tet C in presence of
TET
15) Positive control for tet C
1, 2) Positive control for presence of chlamydial RNA
C. suis R19
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15
3, 4) Negative control for presence of DNA in
chlamydial RNA
5) PCR of R19 gDNA
6) tet R expressed in absence of TET
7) tet R expressed in presence of TET
8) Negative control (DNA) for tet R in absence of
TET
9) Negative control (DNA) for tet R in presence of
TET
10) Positive control for tet R
11) tet C expressed in absence of TET
12) tet C expressed in presence of TET
13) Negative control (DNA) for tet C in absence of
TET
14) Negative control (DNA) for tet C in presence of
TET
15) Positive control for tet C
ResultsIn E. coli+pSC101 and E. coli+pRAS tet
C is only expressed in the presence of
TET, where as, in C. suis tet C is
constitutive.
The two sequenced chlamydial strains of
C. suis showed that tet C and tet R had a 6 base
pair deletion in its operator region in
comparison to plasmids pSC101 and pRAS.
The two sequenced chlamydial strains of
C. suis showed that tet R had a truncation.
The
Rockey
Lab
is
currently
investigating whether or not these two factors
are the reason tet C is constitutive.
Acknowledgements to• Rockey Lab
• Dr. Dan Rockey
• Jae Dugan
• Dr. Kevin Ahern
• Howard Hughes Medical Institute