Denitrifying Activity and Homologous Enzyme Analysis of Alcanivorax dieselolei Strain N1203

Abstract

An Alcanivorax dieselolei strain, termed strain N1203, was isolated from the consortia of ammonia-oxidizing bacteria (AOB) combined with denitrifying bacteria from our previous study and was shown to have ability to reduce nitrate to nitrite to either nitrous oxide or molecular nitrogen. Analysis of 16S rRNA gene sequences established strain N1203 as a member of the species Alcanivorax dieselolei. In addition, the ability of strain N1203 to utilize various organic substrates as the sole carbon source, supplemented with carbohydrates, amino acids, and n-alkane compounds, was investigated, and this strain was found to have a narrow substrate range of growth such as grycerol, succinate, ethanol and n-alkane hydrocarbon. Furthermore, N1203's stepwise denitrifying activity, utilizing succinate and hexadecane as sole carbon sources, was measured. Gene fragments of nirK and qnorB genes, which are involved in denitrifying activities, were obtained, cloned and sequenced. Phylogenetic analysis for these two genes showed that both the nirK and qnorB sequences, although found in separate branches within clusters, formed subclusters branching from uncultured environmental clones. This demonstrated the typical uniqueness of these genes from any cultivated denitrifiers. Thus, strain N1203 is novel type of denitrifying bacteria that demonstrated denitrifying activities when cultivated using succinate as the sole carbon source.