Summary:
Infections caused by parasites of the genus Leishmania, lead to a set of
pathological conditions described by the term leishmaniasis. A number of
species of this parasite can infect several classes of mammals, including
humans and the main clinical forms of the disease are dermal, mucocutaneous and
visceral, the latter can be fatal. The species L. infantum and L. donovani are
categorized as those that cause the visceral form of the disease. The
geographical distribution of leishmaniasis refers to 98 contries worldwide and
350 million people are located in high risk areas, with 12 million of them
being infected. Leishmania parasites are of unicellular form and include
various developmental stages while their life cycle is formed between two types
of hosts, the phlebotomine sand fly and the mammal. The main target cells of
the parasite in mammals are macrophages and dendritic cells that are included
in the first line of defense of the immune system, while successful
immunological host response is based on cell-mediated responses that are
activated by mechanisms acquired as the TH1 response. To date, the treatment of
the disease is mainly based on chemotherapeutic agents while preparing an
effective and safe vaccine is imperative with many research efforts along these
lines. This research project studied antigenicity of the amino portion of the
molecule elongation factor 2 (EF-2, LdeEF-2: 1-357 a.a) of the parasite L.
infantum and the genomic expression of cell subsets of the host, to determine
whether it can be used as a candidate molecule for the development of a vaccine
with the use of dendritic cells as antigen delivery vehicles. All methodologies
utilized included culturing transformed bacteria expressing said molecule,
affinity chromatography, recovery of sensitive BALB / c mice as experimental
animals, the eukaryotic cell culture, flow cytometry, the assay of the in vitro
antigen presentation and the real time quantitative reverse transcriptase
polymerase chain reaction (qRT-PCR). The results confirmed the antigenicity of
the molecule, but the immune response of test animals occurred as a mixed TH1 /
TH2 with the ratio being directed to the acquired TH2 response. Therefore it
was found that this part of the molecule is not appropriate in terms of
utilization as a vaccine. However, this research effort can be a springboard
for further exploration of appropriate antigenic epitopes on this molecule,
which leads to the desired immune response, both by using in silico and by
using in vivo and in vitro approaches, in combination with studies of other
molecules of the parasite, so to design the most effective vaccine to treat the
disease. This research work was part of the interdisciplinary postgraduate
specialization program entitled: Clinical Biochemistry and Molecular
Diagnostics at the National University of Athens, while its preparation took
place in the laboratories of Hellenic Pasteur Institute supervised by the
program KRIPIS.
Keywords:
Antigenicity, Parasite, Gene, Leishmania, Kalaazar
