Quantitative and specific detection of the biocontrol agent, Serratia plymuthica, in plant extracts using a real-time TaqMan(A (R)) assay

TY - JOUR

T1 - Quantitative and specific detection of the biocontrol agent, Serratia plymuthica, in plant extracts using a real-time TaqMan(A (R)) assay

AU - Czajkowski, R.L.

AU - Van der Wolf, J.M.

N1 - Reporting year: 2012 Metis note: 5379; WAG; ME

PY - 2012

Y1 - 2012

N2 - A Serratia plymuthica-specific TaqManA (R) assay was designed based on the consensus nucleotide sequence from the 3'- end of the luxS gene present in all S. plymuthica strains tested. The specificity of the assay was demonstrated by testing 21 Serratia spp. strains and 30 isolates belonging to various species that can potentially coexist with S. plymuthica in the same environment. Positive reactions in the TaqManA (R) assay were observed only for S. plymuthica isolates and not for other bacteria. The TaqManA (R) assay could detect down to 1.95 ng of S. plymuthica DNA, down to 5 bacterial cells per reaction (100 cfu ml(-1)) in vitro, down to 50 bacterial cells per reaction (1,000 cfu ml(-1)) in spiked potato root extracts and down to 5 bacterial cells per reaction (100 cfu ml(-1)) in spiked potato haulm extracts. We used this assay to quantify S. plymuthica A30 cells in potato and tomato haulms and roots grown from S. plymuthica A30-inoculated potato seed tubers and tomato seeds. The results were comparable with the spread-plating of plant extracts on a newly developed S. plymuthica A30 selective medium (CVTR2Arif). The TaqManA (R) assay can be used to quantify S. plymuthica isolates in different ecosystems and in complex substrates.

AB - A Serratia plymuthica-specific TaqManA (R) assay was designed based on the consensus nucleotide sequence from the 3'- end of the luxS gene present in all S. plymuthica strains tested. The specificity of the assay was demonstrated by testing 21 Serratia spp. strains and 30 isolates belonging to various species that can potentially coexist with S. plymuthica in the same environment. Positive reactions in the TaqManA (R) assay were observed only for S. plymuthica isolates and not for other bacteria. The TaqManA (R) assay could detect down to 1.95 ng of S. plymuthica DNA, down to 5 bacterial cells per reaction (100 cfu ml(-1)) in vitro, down to 50 bacterial cells per reaction (1,000 cfu ml(-1)) in spiked potato root extracts and down to 5 bacterial cells per reaction (100 cfu ml(-1)) in spiked potato haulm extracts. We used this assay to quantify S. plymuthica A30 cells in potato and tomato haulms and roots grown from S. plymuthica A30-inoculated potato seed tubers and tomato seeds. The results were comparable with the spread-plating of plant extracts on a newly developed S. plymuthica A30 selective medium (CVTR2Arif). The TaqManA (R) assay can be used to quantify S. plymuthica isolates in different ecosystems and in complex substrates.

KW - national

U2 - 10.1007/s13353-012-0106-0

DO - 10.1007/s13353-012-0106-0

M3 - Article

SN - 1234-1983

VL - 53

SP - 457

EP - 467

JO - Journal of Applied Genetics

JF - Journal of Applied Genetics

IS - 4

ER -