Genodiversity of dominant Rhizobium leguminosarum bv. Trifolii isolated from 11 types of soil in Serbia

Abstract

Rhizobium leguminosarum bv. trifolii is microsymbiont Trifolium pratense and Trifolium repens, which are very important legumes in Serbia. The natural nodulating population of those bacteria was collected and estimated biodiversity distribution by monitoring dominant genotypes of these bacteria. The population of Rhizobium leguminosarum bv. trifolii were collected from 50 marked locations of 11 types of soil in Serbia. 437 natural isolates, rescued from nodules of Trifolium repens or Trifolium pratense, were analyzed by phenotypic approach. We obtained 156 different isolates on the basis of differences in their IAR - intrinsic antibiotic resistance (five antibiotics) and HMT- heavy methal tolerance (five heavy metals). We investigated 56 dominant isolates with more than three differences in IAR-HMT patterns by REP-PCR and RAPD fingerprinting (AP10 and SPH 1 primers). The results showed genodiversity of dominant Rhizobium leguminosarum bv. trifolii field isolates and offered the possibility to assess their changes on marked locations during time and under different environmental conditions and geographical distribution.

Rhizobium leguminosarum bv. trifolii je mikrosimbiont Trifolium pratense i Trifolium repens, veoma značajnih leguminoza u Srbiji. Kolekcionisana je prirodna populacija ovih simbiotskih azotofiksatora i procenjen nivo biodiverziteta analizom dominantnih izolata ovih bakterija. Populacija Rhizobium leguminosarum bv. trifolii je prikupljena sa 50 markiranih lokacija sa 11 tipova zemljišta u Srbiji. 437 autohtonih izolata, izolovanih iz nodula Trifolium repens ili Trifolium pratense, analizirano je fenotpskim metodama. Dobijeno je 156 različitih izolata na osnovu razlika u osetljivosti na pet antibiotika (IAR - intrinsic antibiotic resistance) i tolerantnosti na pet teških metala (HMT- heavy methal tolerance). Utvrđeno je prisustvo 56 dominantnih izolata sa značajnim razlikama u IAR-HMT profilu i za reprezentativne izolate grupe urađen REP-PCR i RAPD fingerprinting (AP10 i SPH 1 prajmerima). Rezultati su pokazali visok stepen genodiverziteta dominantnih izolata Rhizobium leguminosarum bv. trifolii omogućili praćenje promena diverziteta i zastupljenosti na markiranim lokacijama tokom vremena i pri različitim uslovima životne sredine i geografske rasprostranjenosti.