Denitrifying and nitrous oxide reducing genotypes

Denitrification is a biogeochemical process of major importance for nitrogen loss from ecosystems. This four-step pathway is modular, as organisms can have different subsets and variants of the genes involved in each step. The last step is the only known biological sink of nitrous oxide, a potent greenhouse gas and ozone depleting substance. The aim of this thesis was to assess whether specific environmental conditions favour certain denitrifying and nitrous oxide reducing genotypes. The effects of nitrogen, carbon and oxygen availabilities, as well as habitat type and diversity were examined in studies of denitrifying and nitrous oxide reducing microorganisms in pure cultures, enrichment cultures and natural communities from coastal sediments. By utilizing molecular techniques and directly targeting functional genes encoding for nitrite and nitrous oxide reductases, this work explores the link between genetic potential and functionality of denitrifying and nitrous oxide reducing microbial communities. Microorganisms harbouring genes for complete denitrification dominated in coastal marine sediments, irrespective of oxygen regime and habitat type, which suggests they have an important role not only for nitrogen removal, but also nitrous oxide reduction in coastal ecosystems. However, oxygen affected the nitrous oxide reducing communities. The results indicate niche differentiation between nitrous oxide reducers in relation to oxic/anoxic conditions, as specific lineages within the nitrous oxide reductase gene phylogeny were favoured by certain oxygen regimes. In enrichment cultures, complete denitrifiers were competitive nitrous oxide reducers and could outcompete organisms only capable of nitrous oxide reduction when subjected to carbon and nitrous oxide limitation. For denitrifiers, functional difference between the genes nirS and nirK encoding two structurally different nitrite reductases within the same organism was observed, corroborating that closely related and almost identical genotypes differ in their denitrification activity. Furthermore, primers targeting nitrite reducing communities harbouring either nirS or nirK were re-evaluated and clade-specific primers are suggested. Finally, a framework for primer evaluation using metagenomes is suggested. These results highlight the importance of accounting for the genetic potential of denitrifying and nitrous oxide reducing communities to better understand overall ecosystem constraints and how environmental factors might control this potential