Abstract
Advances in microbial ecology research are more often than not limited by the capabilities of available methodologies. Aerobic autotrophic nitrification is one of the most important and well studied microbiological processes in terrestrial and aquatic ecosystems. We have developed and validated a microbial diagnostic microarray based on the ammonia-monooxygenase subunit A (amoA) gene, enabling the in-depth analysis of the community structure of bacterial and archaeal ammonia oxidisers. The amoA microarray has been successfully applied to analyse nitrifier diversity in marine, estuarine, soil and wastewater treatment plant environments. The microarray has moderate costs for labour and consumables and enables the analysis of hundreds of environmental DNA or RNA samples per week per person. The array has been thoroughly validated with a range of individual and complex targets (amoA clones and environmental samples, respectively), combined with parallel analysis using traditional sequencing methods. The moderate cost and high throughput of the microarray makes it possible to adequately address broader questions of the ecology of microbial ammonia oxidation requiring high sample numbers and high resolution of the community composition.
Highlights
The global nitrogen cycle is central to maintaining life on earth and maintaining this key geochemical process has recently been identified as one of the major threats to maintaining the Earth’s environment in a habitable state [1]
The importance of nitrification varies with the environment in which it occurs. It is an essential process for the removal of inorganic nitrogen from marine and freshwater ecosystems as well as during wastewater treatment processes, whilst in terrestrial systems, nitrification plays a major role in the removal of ammonia from soils and in doing so, increases the agricultural need for nitrogen fertilisation
It cannot be excluded that members of the ammonia oxidisers (AOA) and attributed exclusively to bacterial nitrifiers (AOB) represented by divergent ammonia monooxygenase phylotypes may be capable of utilising compounds such as acetate or methane [33]
Summary
The global nitrogen cycle is central to maintaining life on earth and maintaining this key geochemical process has recently been identified as one of the major threats to maintaining the Earth’s environment in a habitable state [1]. It cannot be excluded that members of the AOA and AOB represented by divergent ammonia monooxygenase (amoA) phylotypes may be capable of utilising compounds such as acetate or methane [33] Whilst many of these organisms are not currently cultivated, studying their occurrence in different environments may help to describe their ecology. To overcome the limitation associated with cultivation, molecular methods have been broadly used to assess the diversity of nitrifiers in various environments These methods have allowed nitrifiers to be quantified and characterized by analysis of functional marker genes involved in the nitrification process. We report on the development and thorough validation of an amoA-based short oligonucleotide microarray and associated methodology for the affordable, fast, high-throughput, in-depth analysis of the community structure of aerobic autotrophic ammonia oxidising bacteria and archaea from a wide range (marine, estuarine, soil and wastewater treatment plant) of environments
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