Soil type-dependent responses to phenanthrene as revealed by determining the diversity and abundance of polycyclic aromatic hydrocarbon ring-hydroxylating dioxygenase genes by using a novel PCR detection system.
A novel PCR primer system that targets a wide range of polycyclic aromatic hydrocarbon ring-hydroxylating dioxygenase (PAH-RHD(alpha)) genes of both Gram-positive and Gram-negative bacteria was developed and used to study their abundance and diversity in two different soils in response to phenanthrene spiking. The specificities and target ranges of the primers predicted in silico were confirmed experimentally by cloning and sequencing of PAH-RHD(alpha) gene amplicons from soil DNA. Cloning and sequencing showed the dominance of phnAc genes in the contaminated Luvisol. In contrast, high diversity of PAH-RHD(alpha) genes of Gram-positive and Gram-negative bacteria was observed in the phenanthrene-spiked Cambisol. Quantitative real-time PCR based on the same primers revealed that 63 days after phenanthrene spiking, PAH-RHD(alpha) genes were 1 order of magnitude more abundant in the Luvisol than in the Cambisol, while they were not detected in both control soils. In conclusion, sequence analysis of the amplicons obtained confirmed the specificity of the novel primer system and revealed a soil type-dependent response of PAH-RHD(alpha) gene-carrying soil bacteria to phenanthrene spiking.
Ding
Guo-Chun
GC
Julius Kuhn-Institut, Messeweg, D-38104 Braunschweig, Germany.
Heuer
Holger
H
Zühlke
Sebastian
S
Spiteller
Michael
M
Pronk
Geertje Johanna
GJ
Heister
Katja
K
Kögel-Knabner
Ingrid
I
Smalla
Kornelia
K
D001419:Bacteria / Q000145:classification* / Q000235:genetics; D001426:Bacterial Proteins / Q000235:genetics*; D044822:Biodiversity; D003001:Cloning, Molecular; D016000:Cluster Analysis; D017931:DNA Primers / Q000235:genetics; D049308:Dioxygenases / Q000235:genetics*; D054892:Metagenome; D008969:Molecular Sequence Data; D010616:Phenanthrenes / Q000378:metabolism*; D010802:Phylogeny; D016133:Polymerase Chain Reaction / Q000379:methods; D017422:Sequence Analysis, DNA; D017385:Sequence Homology; D012988:Soil Microbiology
D001426:Bacterial Proteins; D017931:DNA Primers; D010616:Phenanthrenes; C031181:phenanthrene; D049308:Dioxygenases