THESE Maryse Bonnin Jusserand - Université de Bourgogne

Résultats
(SA.BO 34), Italy (BR14/97) and the Languedoc Roussillon region (DIV7 A.3) located in the
south of France. The two other strains (DBA and DIV 5.74) were isolated from cider in the
Normandie region of France (Coton M. & Coton E.). So these strains are well representative
of the biodiversity of the O. oeni specie. Our results support the view of Marcobal et al. (13)
that the presence of the odc gene is a rare event in O. oeni. We confirm this point by our large
screening of O. oeni strains. Futhermore, according to these authors, the odc gene is acquired
via horizontal gene transfer and was found in a genetic environment containing several
transposases and phage-related proteins. In order to verify that the PCR products obtained
corresponded to the expected odc gene, the fragments were sequenced. Analysis of the
nucleotide sequences obtained revealed 97% identity within the sequence of the odc gene
from the strain O. oeni RM83 (GI 54260934), while ODC proteins sequences are 100%
identical to the RM83 ODC except for the two strains isolated from cider (Fig 3). Indeed we
can distinguish, from the proteins alignment, two kind of ODC depending on the habitat (wine
or cider) of O. oeni. Differences in sequence involved the two O. oeni strains isolated from
cider. Amino acid H31 is replaced by R and V448 is replaced by I in DBA and DIV 5.74
strains. D620 is replaced by N only in DBA. These changes don’t affect the catalytic domain
and the PLP binding domain of the enzyme as these changes are located in the N-terminal, C-
terminal domain and at the end of the “small” domain.
The odc gene was first identified in the strain RM83 and sequenced (13). Also this is the only
odc sequence from an O. oeni strain available since none of the sequenced genome carries this
gene.
Among the five odc + strains, the BR14/97 strain was chosen to clone and characterize the odc
gene because of its high capacity to produce a large amount of both putrescine and cadaverine
(7). This was confirmed by measuring biogenic amines in LAC medium by HPLC. O. oeni
BR14/97 strain produced 6,5 mg. L -1 putrescine and 4,6 mg. L -1 cadaverine, while SABO 34
produced 12,5 mg. L -1 putrescine and 6,0 mg. L -1 cadaverine and DIV7 A.3 produced 9,8
mg.L -1 putrescine and 4,3 mg.L -1 cadaverine. Futhermore BR14/97 is representative of the
five isolated since their odc sequences are highly similar (Fig 3). Indeed odc gene was
sequenced for all these strains. Corresponding accession numbers are as follow: FR751075
for DIV7A.3, FR751076 for DBA, FR751077 for DIV5.74, FR751078 for SA.BO 34 and
FR751079 for BR14/97.
112