Azospirillum brasilense viable cells enumeration using propidium monoazide-quantitative PCR.
Elisandra Triches da Cunha, Ana Marina Pedrolo, Franciele Paludo, Mirella Crhistine Scariot, Ana Carolina Maisonnave Arisi
Author Information
Elisandra Triches da Cunha: Laboratory of Molecular Biology, Food Science and Technology Department, Federal University of Santa Catarina, Florianópolis, SC, Brazil.
Ana Marina Pedrolo: Laboratory of Molecular Biology, Food Science and Technology Department, Federal University of Santa Catarina, Florianópolis, SC, Brazil.
Franciele Paludo: Laboratory of Molecular Biology, Food Science and Technology Department, Federal University of Santa Catarina, Florianópolis, SC, Brazil.
Mirella Crhistine Scariot: Laboratory of Molecular Biology, Food Science and Technology Department, Federal University of Santa Catarina, Florianópolis, SC, Brazil.
Ana Carolina Maisonnave Arisi: Laboratory of Molecular Biology, Food Science and Technology Department, Federal University of Santa Catarina, Florianópolis, SC, Brazil. ana.arisi@ufsc.br.
Azospirillum brasilense is a plant growth promoting bacteria used as an inoculant in diverse crops. Accurate analytical methods are required to enumerate viable cells in inoculant formulations or in planta. We developed a quantitative polymerase chain reaction (qPCR) assay associated to propidium monoazide (PMA) to evaluate the cell viability of A. brasilense in inoculant and in maize roots. A. brasilense was grown in culture medium and was exposed to 50 ℃. Maize roots were grown in vitro and harvested 7 days after inoculation. Quantification was performed by qPCR, PMA-qPCR, and plate counting. Standard curves efficiency values ranged from 85 to 99%. The limit of detection was 10 CFU per gram of fresh root. Enumeration obtained in maize roots by qPCR where higher than enumeration by PMA-qPCR and by plate counting. PMA-qPCR assay was efficient in quantifying inoculant viable cells and provides reliable results in a quickly and accurately way compared to culture-dependent methods.
