[Effect of long-term application of nitrogen fertilizer on the diversity of nitrifying genes (amoA and hao) in paddy soil].

The aim of this study was to determine the effect of long-term (16 years) application of nitrogen fertilizer on the diversity of nitrifying genes (amoA and hao) in paddy soil on the basis of long-term paddy field experimental station (started in 1990) located in Taoyuan, with the molecular approaches of PCR, constructing libraries and sequencing. The fertilizer was urea and no fertilizer was as control. The Shannon index showed that long-term application of nitrogen fertilizer made the diversity of amoA gene descend while no effect on the diversity of hao gene. The LIBSHUFF statistical analyses demonstrated that both amoA and hao libraries of CK and N treatments were significantly different from each other and the rarefaction curves of libraries failed to meet the plateaus indicating that there were lots kinds of genes haven't been detected. The results of blasting with GenBank and the phylogenetic tree showed that the amoA genes detected in our study had a similarity with the uncultured gene of amoA, which showed some similar to Nitrosospira. Otherwise, the hao genes cloned showed a relationship to the genes of cultured bacteria such as Silicibacteria, Nitrosospira and Methylococcus, and the hao genes found in the N treatment dominated in alpha-Proteobacteria. These results suggest that long-term fertilization of nitrogen had significant impacts on the diversity or community of amoA and hao genes.

Diversity of soluble methane monooxygenase-containing methanotrophs isolated from polluted environments.

Methanotrophs were enriched and isolated from polluted environments in Canada and Germany. Enrichments in low copper media were designed to specifically encourage growth of soluble methane monooxygenase (sMMO) containing organisms. The 10 isolates were characterized physiologically and genetically with one type I and nine type II methanotrophs being identified. Three key genes: 16S rRNA; pmoA and mmoX, encoding for the particulate and soluble methane monooxygenases respectively, were cloned from the isolates and sequenced. Phylogenetic analysis of these sequences identified strains, which were closely related to Methylococcus capsulatus, Methylocystis sp., Methylosinus sporium and Methylosinus trichosporium. Diversity of sMMO-containing methanotrophs detected in this and previous studies was rather narrow, both genetically and physiologically, suggesting possible constraints on genetic diversity of sMMO due to essential conservation of enzyme function.

[Stability of genetic markers in methane-oxidizing bacteria]. / Stabil'nost' geneticheskikh markerov u metanokisliaiushchikh bakterii.

A number of Methylococcus thermophilus 111p clones have been obtained which have acquired resistance to tetracycline. The stability of maintenance of marker resistance in these clones and also in already designed Methylomonas rubra 15sh mutants has been investigated. Chromosomal markers resistance to antibiotics or formaldehyde were maintained in the marked strains Methylococcus thermophilus 111p and Methylomonas rubra 15sh after storage in nonselective conditions. The markers of resistance to antibiotics, which were coded by plasmids (pAS8-121 and pULB113), were not always preserved in Methylomonas rubra and Methylococcus thermophilus. The stability of maintenance of chromosomal markers in the investigated methane oxidizing bacteria testifies to the fact that they can be used in laboratory and industrial practice for testing the marked bacteria on selective media. The collection of the marked bacteria-mutants Methylomonas rubra 15sh and Methylococcus thermophilus 111p has been created. These strains stably support the marker resistance to various antibiotics or formaldehyde in unselective conditions.