Applicability of the 16S-23S rDNA internal spacer for PCR detection of the phytostimulatory PGPR inoculant Azospirillum lipoferum CRT1 in field soil.
OBJECTIVE
To assess the applicability of the 16S-23S rDNA internal spacer regions (ISR) as targets for PCR detection of Azospirillum ssp. and the phytostimulatory plant growth-promoting rhizobacteria seed inoculant Azospirillum lipoferum CRT1 in soil.


RESULTS
Primer sets were designed after sequence analysis of the ISR of A. lipoferum CRT1 and Azospirillum brasilense Sp245. The primers fAZO/rAZO targeting the Azospirillum genus successfully yielded PCR amplicons (400-550 bp) from Azospirillum strains but also from certain non-Azospirillum strains in vitro, therefore they were not appropriate to monitor indigenous Azospirillum soil populations. The primers fCRT1/rCRT1 targeting A. lipoferum CRT1 generated a single 249-bp PCR product but could also amplify other strains from the same species. However, with DNA extracts from the rhizosphere of field-grown maize, both fAZO/rAZO and fCRT1/rCRT1 primer sets could be used to evidence strain CRT1 in inoculated plants by nested PCR, after a first ISR amplification with universal ribosomal primers. In soil, a 7-log dynamic range of detection (10(2)-10(8) CFU g(-1) soil) was obtained.


CONCLUSIONS
The PCR primers targeting 16S-23S rDNA ISR sequences enabled detection of the inoculant A. lipoferum CRT1 in field soil.


CONCLUSIONS
Convenient methods to monitor Azospirillum phytostimulators in the soil are lacking. The PCR protocols designed based on ISR sequences will be useful for detection of the crop inoculant A. lipoferum CRT1 under field conditions.
Baudoin
E
E
IRD, UMR 113, LSTM, Campus International de Baillarguet, TA-A82/J, 34398 Montpellier cedex5, France.
Couillerot
O
O
Spaepen
S
S
Moënne-Loccoz
Y
Y
Nazaret
S
S
D042081:Azospirillum lipoferum / Q000235:genetics* / Q000302:isolation & purification*; D001483:Base Sequence; D021903:DNA, Ribosomal Spacer / Q000235:genetics*; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000379:methods*; D012988:Soil Microbiology
D021903:DNA, Ribosomal Spacer