Download ANALYSE OF THE MOLECULAR EVOLUTION OF THE ZOONOTIC

ANALYSIS OF THE MOLECULAR EVOLUTION OF THE
ZOONOTIC AGENT Bartonella henselae BY SEQUENCING NINE
GENES
Alix Boulouis
The cat-scratch disease, which is one of the most common diseases in the world
transmissible from animals to humans, is caused by a bacterium, Bartonella henselae. This
bacterium infects 30-60% of domestic cats in the US. It can be transmitted to humans by either a cat
scratch or a cat bite. In the case of the cat scratch, the mechanism is quite easy to understand: the cat
flea (Ctenocephalides felis), which is the transmission agent between cats, makes the cat scratch
herself which contamines her claws. The presence of B. henselae in humans is very difficult to
estimate, as the symptoms are at the same time varying and generally benign for patients with
operational immune systems. B. henselae infects vessel and red blood cells and can cause cancerlike growth of the vessel cells in immunocompromised patients such as those who are HIV-positive.
These bacteria can be isolated from cat and human infections, and a quite large panel of
different strains is now available for laboratory experiments. The total genetic information of one of
them, Houston-1, has recently been identified, and a DNA microarray experiment has been carried
out on several strains to compare them with the sequenced Houston-1 genome. In the microarray
experiment small pieces of DNA from the Houston-1 strain (probes) were anchored on a plate, and
fragments of the genomic DNA from the strains to be analyzed were added to this: if this genomic
DNA contained the same genes as were present in the array, they bound to them, which could be
measured with biochemistry techniques. A microarray analysis allows one to see the differences in
the gene content among the strains, and thus enables one to estimate relations between the strains: it
is more probable that the same change has occurred once in the common ancestor of two strains
than twice independently in both strains. It is not a good evaluation of the molecular evolution of
the genomes, though. Single changes in the DNA of genes can happen during the evolution, having
no influence on the functionality of the gene product, but staying witnesses of the evolution and the
oldness of the strain.
The measurement of single substitutions in chosen genes, by amplification of the genes and
sequencing, is a good complement to the microarray for the analysis of the bacteria evolution. I
carried out this sequencing experiment on nine common genes of the same strains used for the
microarray. The sequencing of the nine genes allowed differentiating several versions of the genes
with very few differences for each gene (except one for which there was no diversity), and the
combination of the alleles for each strain allowed me to define the sequence type of the strain. I
found five different sequence types among these strains. The clustering of the strains was in general
in agreement with the one observed in the microarray, with the exception of one strain having good
similarities with a second strain in the microarray, whereas they were different with the sequence
analysis, which remains unexplained.
Degree project in biology, University of Uppsala, spring 2004
Examensarbete i biologi, 10p
Departments of Biology Education and Molecular Evolution, EBC, Uppsala University
Supervisor: Siv Andersson