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Crosstalk between Trypanosoma brucei and the tsetse fly

Oberle, Michael. Crosstalk between Trypanosoma brucei and the tsetse fly. 2011, PhD Thesis, University of Basel, Faculty of Science.

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Official URL: http://edoc.unibas.ch/diss/DissB_9474

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Abstract

Trypanosoma brucei cause the fatal disease sleeping sickness in humans and the morbid
disease nagana in animals. About 36 sub-Saharan African countries are affected by these
diseases. The parasites are transmitted by tsetse flies (Glossina spp.) exclusively where they
colonise the alimentary tract and the salivary glands. The trypanosomes establish first in the
midgut as procyclic forms from where they colonise then the proventriculus (that connects the
mid- with the foregut) and migrate later as epimastigote forms into the salivary glands via the
foregut and proboscis. In the salivary glands epimastigote forms attach to the epithelium and
give rise to the mammalian infective forms, the metacyclics. During transmission through the
fly, trypanosomes are frequently severely reduced when they invade a new compartment.
Trypanosomes either recover and develop an infection or fail to establish an infection and are
eliminated by the tsetse fly’s defence. This complex interaction between vector and parasite
shows that both counterparts specifically regulate genes.
In this thesis we wanted to shed light into this complex crosstalk in three projects:
We established a model to analyse how the severe reductions during the life cycle influence
the diversity of trypanosomes. Short variable DNA sequences were integrated into the
trypanosome’s genome to establish an artificial diversity. These transfected trypanosomes
were cyclically transmitted through flies and mice. Tag DNA was isolated from infected flies
and/ or mice and identified by sequencing. This allowed us to monitor diversity of the
trypanosomes throughout their life cycle. We found that diversity was moderately reduced in
the tsetse fly’s midgut but that migration into the salivary glands decreases the diversity. This
decrease is mainly due to a shift in relative frequency which leads to a very uneven
distribution of the tags. The diversity constantly decreased during mouse infection due to the
constant gain of trypanosomes bearing the dominant tag. Surprisingly, the number of different
tags was not reduced during the whole life cycle of the trypanosomes.
The two anti-microbial peptides (AMPs), attacin and defensin, of tsetse flies were reported to
play an important role in eliminating trypanosomes in the midgut. The mRNA of these AMPs
was shown to be up-regulated upon trypanosome infection and it was hypothesised that
procyclins might specifically induce its activation. We wanted to test this with different
trypanosome strains as well with trypanosomes with incomplete or deleted procyclin coats.
Tsetse flies were infected and mRNA isolated after various times of trypanosome exposition.
None of the flies showed an up-regulated level of attacin and defensin mRNA. This result is
in strong contradiction to some publications dealing with AMP regulation in infected tsetse
flies. The tsetse flies, from the colony in Bratislava (Slovakia), show a high level of attacin
and defensin mRNA in teneral flies (what not all G. m. morsitans do), show a midgut
infection rate of about 50% (which is high compared to the infection rate in other
laboratories), and are infected sometimes with the salivary gland hypertrophy virus (SGHV).
It is very possible that attacin and defensin are not always up-regulated and that its activation
is dependent on tsetse colony and origin.
During the establishment in the midgut trypanosomes express procyclins, a stage specific
surface protein coat that was suggested to protect against proteolytic enzymes or to be
important to direct the parasite in the host. To test this hypothesis all procyclin genes were
deleted and tsetse fly infection experiments were carried out. Interestingly, the null-mutant
(Δprocyclin) was able to infect the midgut comparable to wild type trypanosomes, disclosing
that procyclins are not needed for the establishment in the midgut and that probably free
glycosylphosphatidylinositol (GPI) anchors, which are loaded with procyclins in wild type
trypanosomes, overtook their function. Surprisingly, Δprocyclin was able to infect the salivary
glands even though at very low rates, which reflects difficulties of trypanosomes to re-load
the free GPIs with epimastigote specific surface proteins (e.g. BARP) for efficient migration.
In competition, Δprocyclin was completely overgrown by wild type trypanosomes in the
tsetse midgut, reflecting the selective advantage of a procyclin coat.
Advisors:Brun, Reto
Committee Members:Roditi, Isabel and Abbeele, Jakke van den
Faculties and Departments:09 Associated Institutions > Swiss Tropical and Public Health Institute (Swiss TPH) > Department of Medical Parasitology and Infection Biology > Parasite Chemotherapy (Mäser)
Item Type:Thesis
Thesis no:9474
Bibsysno:Link to catalogue
Number of Pages:124 S.
Language:English
Identification Number:
Last Modified:30 Jun 2016 10:41
Deposited On:10 Jun 2011 09:07

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