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Identifier |
000441534 |
Title |
Συγκριτική μελέτη στελεχών Aeromonas veronii ως νοσογόνου παράγοντα σε εκτρεφόμενα λαβράκια. Ανάπτυξη αυτεμβολίων για την πρόληψη της ασθένειας |
Alternative Title |
Comparative study on Aeromonas veronii isolates as disease agent in farmed European seabass. Development of autogenus vaccines for the prevention of the disease |
Author
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Σμυρλή, Μαρία Ν
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Thesis advisor
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Παυλίδης, Μιχαήλ
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Reviewer
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Καθάριος, Παντελής
Αθανασάκη, Ειρήνη
Καλαντίδης, Κρίτων
Μπακόπουλος, Βασίλειος
Ντότσικα, Ελένη
Χαμηλός, Γεώργιος
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Abstract |
The object of the study was a bacterial disease affecting farmed European seabass
caused by Aeromonas veronii bv. sobria. The disease affects farmed sea bass weighing>
50 g on fish farms in the Aegean Sea, mainly in the west. It occurs in temperature > 21-
22oC, while the outbreaks occur mainly in the summer months. The cumulative
mortality can reach up to 80% of the farmed population. The current study concerns
an emerging pathology for seabass fish farming and was initially considered a local,
individual case. The pathology and epidemiological features of the disease were
studied. Pathogen diversity was studied with molecular (16S rRNA and gyrB) and
biochemical (Biolog and API20E) tools. As part of the work, a genus-based detection
protocol was developed. The infectivity of the pathogen was tested in vivo, with
experimental infections in healthy seabass and zebrafish and in vitro, on a blood agar
medium. The genomes of representative strains were sequenced, and the phylogenetic
relationships and the antigenic nature and diversity of the pathogen were studied. Τhe
principles of reverse vaccinology were applied to detect conserved membrane
proteins–antigens for the development of the widely used Aeromonas veronii vaccine.
Finally, a bacterial vaccine (water and oil-based) was developed. The efficacy of the
vaccine was tested by challenge tests on the vaccinated fish, by measuring the titer of
antibodies (ELISA) in the blood serum and by the expression of selected genes related
to the immune system via qPCR. Three phenotypes of the pathogen were identified
which are distinguished in terms of motility and pigment production and correspond to
different regions of the Aegean Sea. Strains from different regions were almost (ppsA)
indistinguishable with the studied housekeeping genes used as genetic markers. The
symptoms of the disease were reproduced in artificial infections in seabass, confirming
the pathogenicity of A. veronii. The use of zebrafish for the study of bacterial infectivity
in Greek fish farming species is encouraged by the results since infections induced
mortality in zebrafish too. Genomic analyses showed high similarity between A. veronii
strains of seabass in different regions of the Aegean Sea. However, their differentiation
was possible by single nucleotide polymorphisms corresponding to the phenotypic
groups mentioned above and the different geographical origins of the strains (East
/West Aegean). The outer membrane antigenic proteins also showed a high degree of
similarity at the amino acid sequence, but differences were found in the S-layer protein,
maltoporin LamB, and O-antigen, suggesting the use of a multivalent vaccine for
Aegean seabass culture. The injectable bivalent adjuvanted (NS+PDB+Montanide)
vaccine offered protection to adult seabass against bath challenge with strain PDB (RPS
= 62.5%). The aqueous bivalent (NS + PDB) vaccine administered by immersion in
seabass fry also provided protection against strain PDB (RPS = 89.4%). The antibody
titer for NS and PDB antigens was stable for at least 60 days after injection vaccination
with bivalent adjuvanted (NS+PDB+Montanide) vaccine. Gene expression of MHCII-β,
IgM and especially TCR-β genes in the head kidney tissue increased up to day 15 after
vaccination by injection with aqueous and adjuvanted injectable vaccine with strain NS,
while cellular immunity (CD4) appears to be activated on day 30 only in the group
vaccinated with the adjuvanted preparation. The results produced here encourage the
use of autogenous vaccines for the control of A. veronii disease in farmed European
seabass. Boost vaccinations and/or increased antigen doses are proposed in the future
to increase the vaccine efficacy.
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Language |
Greek |
Subject |
Aegean sea |
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Αιγαίο πέλαγος |
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Αυτεμβόλια |
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Λαβράκι |
Issue date |
2021-07-29 |
Collection
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School/Department--School of Sciences and Engineering--Department of Biology--Doctoral theses
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Type of Work--Doctoral theses
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Permanent Link |
https://elocus.lib.uoc.gr//dlib/8/b/a/metadata-dlib-1627893637-780765-30530.tkl
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Views |
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