© Benaki Phytopathological Institute
Abstracts - 16th Hellenic Phytopathological Congress
17
to the above, all R
2
mutant strains exhibited
a low to moderate multi-drug resistance to
DMIs and to several other fungicide class-
es. Two different homologous genes,
cyp51A
and
cyp51B
, encoding C-14 alpha sterol dem-
ethylase (Cyp51) and an
mdr
gene encoding
an ATP-binding cassette protein were cloned
and characterized. Sequence comparison of
cyp51A
gene revealed an amino acid substitu-
tion from glycine (GGG) to
tryptophan
(TGG)
at position 54 (G54W) in two out of three of
R
1
mutant strains. Analysis of deduced amino
acid sequence of
cyp51B
showed that no mu-
tations were associated with DMI resistance.
Study of the transcription levels of
cyp51A
showed that this gene was over-expressed in
the third aflatoxigenic mutant strain. Neither
amino acid substitutions within nor overex-
pression of the
cyp51A
gene were found in
the R
2
mutant strains tested. Real-time PCR
analysis showed
high levels (up to 25-fold
higher) of the
mdr
transcript in all R
2
mutant
strains tested. This is the first report describ-
ing the existence of two
cyp51
genes and a
potential
mdr
gene coding for an ATP bind-
ing cassette protein in
A. parasiticus
. These
results also indicate that multiple biochem-
ical mechanisms, including target-site modi-
fication due to mutation of the
cyp51A
gene,
cyp51A
overexpression, and the function of
an ABC transporter protein, are responsible
for DMI-resistance in
A. parasiticus
. Our find-
ings suggest that
A. parasiticus
has the ge-
netic and biochemical potential to give rise
to highly aflatoxigenic DMI-resistant isolates
in the field.
Identification and determination of antibiotic susceptibility of cultured
enterobacterial flora from leafy vegetables in Cyprus
I. A
NASTASIOU
, E. S
AVVA
and D. T
SALTAS
Department of Agricultural Sciences, Biotechnology and Food Science, Cyprus
University of Technology, 3603 Limassol, Cyprus
In the last few years, the consumption of
raw leafy vegetables has increased due to
their well-documented nutritional value.
The contamination of these crops by mi-
crobes originating from soil, manure and
handler microflora is of increasing interest
from a public health perspective. In addi-
tion to the microbial load, the antibiotic re-
sistance of these microorganisms as a result
of antibiotic usage in agriculture and oth-
er related fields it is of increasing concern.
The aim of this study was to identify and
enumerate the enterobacterial microflo-
ra on samples of leafy vegetables (lettuce,
rucola, spinach, purslane) in Cyprus, and to
evaluate the levels of resistance of these mi-
croorganisms to the antibiotics ampicillin,
cefotaxime, gentamicin and vancomycin.
The results showed high levels of total mi-
croflora and
Enterobacteriaceae
contamina-
tion, within the ranges of 6.31-7.85 and 5.86-
6.7 log cfu/g respectively. More than 95% of
the total microflora and 87% of
Enterobac-
teriaceae
identified were resistant to ampi-
cillin. The rates of resistance to cefotaxime
(33.41%) and gentamicin (17.35%) for
Enter-
obacteriaceae
were lower, but for the total
microflora resistance to the latter antibiot-
ics was higher (83.33% and 76.58% respec-
tively). Rucola and spinach produced pos-
itive results for
Escherichia coli,
with rucola
being the only sample found to be contam-
inated with isolates resistant to cefotaxime
(0.43 log cfu/g). The samples also had similar
population numbers for yeasts and moulds
(4.04-5.67 log cfu/g). The current study
shows that leafy vegetables can be a signif-
icant source of pathogenic microorganisms,
including antibiotic resistant isolates.
Effect of Alternaria leaf-spot on the content of antioxidant compounds in
infected kiwi orhards
