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Watercress (Nasturtium officinale) is an aquatic dicotyledonous vegetable belonging to
Brassicaceae (Aiton 1812). Watercress was grown in an aquaponic system on fired clay ball
medium at the Aquaponic Research Station of the University of Debrecen, in the city of
Debrecen (Hungary). During January 2020, 3-month-old plants showed symptoms in aquaponic
cultivation. A visual survey showed 30% of plants with symptoms. Leaves and stems withered
and showed white cotton-like mycelium. Mycelia from infected plants were placed on potato
dextrose agar (PDA) and incubated at 25°C for 7 days. Single hyphal tips were transferred to
produce a pure culture. All 10 fungal isolates showed similar morphological characteristics on
PDA. Colonies consisted of white mycelia after 3 days, and globoid to irregular and black 2.5
to 7 (average, 3) mm (n = 100 from 10 plates) sclerotia formed 10 days later, which are the
typical morphological features of Sclerotinia sclerotiorum (Mordue et al. 1976). Molecular
identification was performed with one of the 10 isolates (Scl_B). Mycelia were grown in 250 ml
of potato dextrose broth in a rotary shaker at 175 rpm at 24°C for 6 days. DNA was extracted
from mycelium using a NucleoSpin plant II (Macherey-Nagel, Germany) according to the
manufacturer’s protocol. Polymerase chain reaction (PCR) amplification (Kim et al. 2014) was
performed with primers ITS1/ITS4 for the internal transcribed spacer (ITS) region (White et al.
1990) on a Primus 96 thermal cycler (MWG Biotech, Germany). Specific PCR was performed
with primers SSasprF/SSasprR (Abd-Elmagid et al. 2013). PCR products were sequenced by
Microsynth Austria GmbH. NCBI BLAST analysis of the 440-bp ITS sequence (GenBank
MW012403.1) showed 100% identity with the sequence of S. sclerotiorum (MT177267.1, etc.).
The 170-bp specific gene sequence (GenBank MW959042.1) had a 100% similarity to
hypothetical proteins (GenBank MK028159.1), with a 99.4% similarity to a portion of the S.
sclerotiorum aspartyl protease gene (AF271387.1). Pathogenicity tests were carried out by
inoculating surface-disinfested, 30-day-old watercress plants in plastic pots (15 × 15 × 12 cm).
In three repeated experiments 90 watercress plants were measured. Fifteen plants (one plant
per pot) were planted into the five-times-autoclaved substrate (Biorgmix: pH 6.1 ± 0.5%; N,
1.5%; P2O5, 0.7%; K2O, 0.5%; organic matter content, 50%) and inoculated by 10 wheat kernels
that were colonized by S. sclerotiorum (Scl_B) (Garibaldi et al. 2019). Fifteen plants were
planted into the substrate with 10 noninoculated kernels as a control. Plants were kept in an
MLR-352 climatic test chamber (PHCbi, Japan) at 21 ± 1°C with a 12-h light/dark cycle. On the
first day of the experiment, complex nutrient solution (Tek-Land: N, 5%; P2O5, 5%; K2O, 5%;
B, 0.01%; Cu, 0.01%; Mn, 0.02%; Mo, 0.002%; Zn, 0.016%) was used, followed by autoclaved
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Brassicaceae (Aiton 1812). Watercress was grown in an aquaponic system on fired clay ball
medium at the Aquaponic Research Station of the University of Debrecen, in the city of
Debrecen (Hungary). During January 2020, 3-month-old plants showed symptoms in aquaponic
cultivation. A visual survey showed 30% of plants with symptoms. Leaves and stems withered
and showed white cotton-like mycelium. Mycelia from infected plants were placed on potato
dextrose agar (PDA) and incubated at 25°C for 7 days. Single hyphal tips were transferred to
produce a pure culture. All 10 fungal isolates showed similar morphological characteristics on
PDA. Colonies consisted of white mycelia after 3 days, and globoid to irregular and black 2.5
to 7 (average, 3) mm (n = 100 from 10 plates) sclerotia formed 10 days later, which are the
typical morphological features of Sclerotinia sclerotiorum (Mordue et al. 1976). Molecular
identification was performed with one of the 10 isolates (Scl_B). Mycelia were grown in 250 ml
of potato dextrose broth in a rotary shaker at 175 rpm at 24°C for 6 days. DNA was extracted
from mycelium using a NucleoSpin plant II (Macherey-Nagel, Germany) according to the
manufacturer’s protocol. Polymerase chain reaction (PCR) amplification (Kim et al. 2014) was
performed with primers ITS1/ITS4 for the internal transcribed spacer (ITS) region (White et al.
1990) on a Primus 96 thermal cycler (MWG Biotech, Germany). Specific PCR was performed
with primers SSasprF/SSasprR (Abd-Elmagid et al. 2013). PCR products were sequenced by
Microsynth Austria GmbH. NCBI BLAST analysis of the 440-bp ITS sequence (GenBank
MW012403.1) showed 100% identity with the sequence of S. sclerotiorum (MT177267.1, etc.).
The 170-bp specific gene sequence (GenBank MW959042.1) had a 100% similarity to
hypothetical proteins (GenBank MK028159.1), with a 99.4% similarity to a portion of the S.
sclerotiorum aspartyl protease gene (AF271387.1). Pathogenicity tests were carried out by
inoculating surface-disinfested, 30-day-old watercress plants in plastic pots (15 × 15 × 12 cm).
In three repeated experiments 90 watercress plants were measured. Fifteen plants (one plant
per pot) were planted into the five-times-autoclaved substrate (Biorgmix: pH 6.1 ± 0.5%; N,
1.5%; P2O5, 0.7%; K2O, 0.5%; organic matter content, 50%) and inoculated by 10 wheat kernels
that were colonized by S. sclerotiorum (Scl_B) (Garibaldi et al. 2019). Fifteen plants were
planted into the substrate with 10 noninoculated kernels as a control. Plants were kept in an
MLR-352 climatic test chamber (PHCbi, Japan) at 21 ± 1°C with a 12-h light/dark cycle. On the
first day of the experiment, complex nutrient solution (Tek-Land: N, 5%; P2O5, 5%; K2O, 5%;
B, 0.01%; Cu, 0.01%; Mn, 0.02%; Mo, 0.002%; Zn, 0.016%) was used, followed by autoclaved
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