Monitoring viable cells of the biological control agent Lactobacillus plantarum PM411 in aerial plant surfaces by means of a strain-specific viability quantitative PCR method
Autor: | Anna Bonaterra, J. Francés, Jordi Cabrefiga, Emilio Montesinos, Esther Badosa, Núria Daranas |
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Přispěvatelé: | Ministerio de Economía y Competitividad (Espanya) |
Jazyk: | angličtina |
Rok vydání: | 2020 |
Předmět: |
0301 basic medicine
DNA Bacterial 030106 microbiology Population biological control Biology Real-Time Polymerase Chain Reaction Applied Microbiology and Biotechnology Fragaria Pyrus 03 medical and health sciences chemistry.chemical_compound viability-qPCR Plant Microbiology Species Specificity Molecular marker TaqMan Food science education Plants -- Diseases and pests DNA Primers Plant Diseases 2. Zero hunger education.field_of_study PEAR Microbial Viability Ecology Bacterial diseases of plants Plantes -- Malalties bacterianes Amplicon Plant Components Aerial Plantes -- Malalties i plagues biology.organism_classification Pests -- Control Plagues -- Control biològic chemistry Biological Control Agents Malus Nucleic acid Lactobacillus plantarum Bacteria Food Science Biotechnology |
Zdroj: | Applied and Environmental Microbiology © Applied and Environmental Microbiology, 2018, vol. 84, p. Articles publicats (D-EQATA) DUGiDocs – Universitat de Girona instname |
Popis: | A viability quantitative PCR (v-qPCR) assay was developed for the unambiguous detection and quantification of Lactobacillus plantarum PM411 viable cells in aerial plant surfaces. A 972-bp region of a PM411 predicted prophage with mosaic architecture enabled the identification of a PM411 strain-specific molecular marker. Three primer sets with different amplicon lengths (92, 188, and 317 bp) and one TaqMan probe were designed. All the qPCR assays showed good linearity over a 4-log range and good efficiencies but differed in sensitivity. The nucleic acid-binding dye PEMAX was used to selectively detect and enumerate viable bacteria by v-qPCR. The primer set amplifying a 188-bp DNA fragment was selected as the most suitable for v-qPCR. The performance of the method was assessed on apple blossoms, pear, strawberry, and kiwifruit leaves in potted plants under controlled environmental conditions, as well as pear and apple blossoms under field conditions, by comparing v-qPCR population estimations to those obtained by qPCR and specific plate counting on de Man-Rogosa-Sharpe (MRS)-rifampin. The population estimation did not differ significantly between methods when conditions were conducive to bacterial survival. However, under stressful conditions, differences between methods were observed due to cell death or viable-but-nonculturable state induction. While qPCR overestimated the population level, plate counting underestimated this value in comparison to v-qPCR. PM411 attained stable population levels of viable cells on the flower environment under high relative humidity. However, the unfavorable conditions on the leaf surface and the relatively dryness in the field caused an important decrease in the viable population. IMPORTANCE The v-qPCR method in combination with plate counting and qPCR is a powerful tool for studies of colonization and survival under field conditions, to improve formulations and delivery strategies of PM411, and to optimize the dose and timing of spray schedules. It is expected that PEMAX v-qPCR could also be developed for monitoring other strains on plant surfaces not only as biological control agents but also beneficial bacteria useful in the sustainable management of crop production. |
Databáze: | OpenAIRE |
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