Diversity of the alkB Genes of n-Alkane Biodegradation in Thermophilic Hydrocarbon-Oxidizing Bacteria of the Genera Geobacillus, Parageobacillus, and Aeribacillus

Abstract

Analysis of complete genomes of thermophilic bacteria of the genus Geobacillus, oxidizing n-alkanes and crude oil revealed the ladA gene and its homologues. In the genomes of some strains, the ladA gene was not detected, although they were capable of growth on n-alkanes. Cloning with degenerate primers has previously revealed eight homologues of the alkB gene (alkB-geo1–alkB-geo8) encoding alkane hydroxylases in Geobacillus. In the present work, investigation of the alkB genes of several new strains of thermophilic, hydrocarbon-oxidizing bacilli was carried out. In the clone libraries obtained using degenerate primers for the alkB genes from five strains of the genera Geobacillus, Parageobacillus, and Aeribacillus, mostly the universal homologous genes alkB-geo1 and alkB-geo4 were revealed. Additional PCR amplification with specific primers for each of the eight known alkB homologues revealed the universal homologous genes only in some of the studied strains. A correlation was found between the set of the alkB-geo3–alkB-geo6 homologus genes from Geobacillus subterraneus strain K amplified with specific primers and the previously revealed a set of these homologous genes transcripted at different cultivation conditions. This correlation may be due to accumulation of the copies of individual homologues under different cultivation conditions, which results in higher sensitivity of specific primers. The least common homologue, alkB-geo7, which was not detected in the relevant clone libraries, was found in two strains, indicating the possibility of development of more specific primers for amplification of this homologue in order to reveal hydrocarbon-oxidizing bacteria of related genera Geobacillus–Parageobacillus in molecular ecological research.