Articles
DEVELOPMENT OF AMBIVALENT MEDIA TO DETECT BACTERIAL CONTAMINATION OF GLOBE ARTICHOKE MICROPROPAGATION MEDIA
Article number
983_29
Pages
215 – 220
Language
English
Abstract
In industrial micropropagation laboratories culture substrate microbial contamination is a problem.
Visible contamination can be easily recognised and eliminated.
In the case of not detectable contamination, however, there is a high risk of transmitting and spreading the microorganisms vertically or horizontally in the laboratory.
Thus, it is essential to have strategies for establishing and maintaining aseptic tissue cultures.
To ensure that in vitro cultures of the globe artichoke Life are aseptic, we developed a strategy to eliminate a persistent, substrate contaminant bacterium, associated with substrate turbidity that is almost invisible over time, which apparently did not compromise plantlet development, but was resistant to common sanitization measures.
Ambivalent media were developed to favour the growth of the plantlets as well as bacteria, so that to make visible their presence.
Latently contaminated globe artichoke explants, propagated on standard MS media with BA (0.1 mg/L), were transplanted to standard media modified with the addition of combinations of microbial growth stimulants (yeast extract, malt extract, peptone, casamino acids, proteose peptone 3). The plantlets were subsequently transplanted on the same media three times, about every 15 days.
The bacterium, identified as a strain of Bacillus nealsonii or Bacillus benzoevorans, developed visible colonies on all the media, even those with the lowest concentrations of growth promoters.
The results of bacterial growth tests on the substrate variants indicate that the modified MS medium, with addition of (0.02%) yeast extract, (0.05%) casamino acids and (0.05%) proteose peptone 3, promotes the growth of the contaminant bacterium, as well as the growth of Life globe artichoke plantlets in vitro.
The highest concentrations of growth stimulators were shown to be phytotoxic.
Our results show that by using an appropriate ambivalent medium, it is possible to highlight the presence of contaminant bacteria in globe artichoke culture media and facilitate an accurate selection of aseptic jars especially in the early stages following the transplantation of the mother plant tissues.
Visible contamination can be easily recognised and eliminated.
In the case of not detectable contamination, however, there is a high risk of transmitting and spreading the microorganisms vertically or horizontally in the laboratory.
Thus, it is essential to have strategies for establishing and maintaining aseptic tissue cultures.
To ensure that in vitro cultures of the globe artichoke Life are aseptic, we developed a strategy to eliminate a persistent, substrate contaminant bacterium, associated with substrate turbidity that is almost invisible over time, which apparently did not compromise plantlet development, but was resistant to common sanitization measures.
Ambivalent media were developed to favour the growth of the plantlets as well as bacteria, so that to make visible their presence.
Latently contaminated globe artichoke explants, propagated on standard MS media with BA (0.1 mg/L), were transplanted to standard media modified with the addition of combinations of microbial growth stimulants (yeast extract, malt extract, peptone, casamino acids, proteose peptone 3). The plantlets were subsequently transplanted on the same media three times, about every 15 days.
The bacterium, identified as a strain of Bacillus nealsonii or Bacillus benzoevorans, developed visible colonies on all the media, even those with the lowest concentrations of growth promoters.
The results of bacterial growth tests on the substrate variants indicate that the modified MS medium, with addition of (0.02%) yeast extract, (0.05%) casamino acids and (0.05%) proteose peptone 3, promotes the growth of the contaminant bacterium, as well as the growth of Life globe artichoke plantlets in vitro.
The highest concentrations of growth stimulators were shown to be phytotoxic.
Our results show that by using an appropriate ambivalent medium, it is possible to highlight the presence of contaminant bacteria in globe artichoke culture media and facilitate an accurate selection of aseptic jars especially in the early stages following the transplantation of the mother plant tissues.
Authors
O. Navacchi, G. Zuccherelli, U. Mazzucchi
Keywords
Bacillus nealsonii, Bacillus benzoevorans, Cynara cardunculus var. scolymus, bacterial contamination in vitro, early detection
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