Influence of salinity on novel (moderately) thermophilic nitrite-oxidizing bacteria

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Nitrification, the biological oxidation of ammonia to nitrite and further to nitrate, is of environmental importance to avoid accumulation of harmful ammonia and nitrite, which can result in human and aquatic animal health risk. In WWTPs its functioning has to be maintained even under adverse conditions caused for example by elevated temperature or salinity. Especially the fastidious nitrite-oxidizing bacteria (NOB) that perform the second step of nitrification are not sufficiently investigated. From geothermal systems and warm sewage, Nitrospira and different Chloroflexi NOB were already cultivated, but metagenome data indicated the presence of a highly diverse community of putative NOB by specific sequences of the key enzyme, the nitrite oxidoreductase (NXR). In the first phase of this project, directed cultivation approaches led to the description of new (moderately) thermophilic species of Nitrospira and Nitrolancea. Additionally, several novel NOB candidates growing at elevated temperature (37-50°C) from diverse phyla (Ca. Dadabacteria, Actinobacteria, Chloroflexi, Acidobacteria, (Delta)-Proteobacteria, Nitrospirae) were selectively enriched and correlated with specific nxrA/B and 16S rRNA gene sequences as well as distinct morphotypes, but their 16S rRNA gene similarities to the next taxonomically described bacteria are rather low (80-94%). Molecular characterization of metagenome assembled genomes (MAGs) gave first insights into their carbon, nitrogen and oxygen metabolism including the finding of rubisco genes for some Chloroflexi and Actinobacteria, supporting their chemolithoautotrophic lifestyle and differentiating them from denitrifiers. In the past, new NOB isolates were obtained by various selecting factors (nitrite concentration, temperature, DO, pH), whereas in the second part of the project, salinity stress (especially nitrate) will be the main driver for niche separation in combination with high temperature. Preliminary results indicated high NaCl tolerances of Dehalococcoides-like NOB and novel Actinobacteria and reduced abundance of Nitrospira. It is the goal to highly enrich/isolate novel salt- and heat-tolerant NOB by cell sorting, to characterize them physiologically and to screen metagenomes for new metabolic pathways. RNA based gene expression and metatranscriptome data obtained under different growth conditions will verify activity of novel nxr genes and give insights into tolerance mechanisms. Most promising candidates will be tested for their ability to grow in bioreactor systems under salt and temperature stress for biotechnological applications. With regard to a warming world, there is a growing interest in nitrification under thermal conditions and it has to be ensured that agricultural nitrification inhibitors will still affect a shifting community of nitrifying microorganisms. Furthermore, thermophilic bacteria as survivors of early Earth are interesting with regard to evolutionary aspects.