Obtaining complete and canonical ammonia-oxidizing bacteria through specific labeling and cell sorting

Area of Science:

• Environmental Microbiology
• Biogeochemical Cycles
• Nitrogen Cycling

Background:

• Excess anthropogenic nitrogen input causes environmental issues.
• Nitrification is a key nitrogen cycle process performed by diverse microorganisms.
• Traditional isolation methods are hindered by the difficult growth of nitrifying bacteria.

Purpose of the Study:

• To develop a workflow for targeted enrichment and isolation of ammonia-oxidizing bacteria.
• To overcome limitations of classical cultivation techniques for nitrifier isolation.
• To enable in-depth physiological characterization of ammonia oxidizers.

Main Methods:

• Function-specific in vivo fluorescent labeling combined with cell sorting.
• Optimized floc disruption and labeling for enhanced cell recovery and signal intensity.
• Cultivation in ammonium-containing media, screening for nitrite/nitrate production, and molecular characterization (16S rRNA, amoA PCR, FISH).

Main Results:

• Successfully isolated axenic and enriched Nitrosomonas cultures.
• Obtained seven comammox Nitrospira enrichment cultures.
• Retrieved cultures from bioreactors, aquaculture biofilters, and agricultural soil.

Conclusions:

• The presented workflow enables rapid and targeted isolation of ammonia oxidizers from complex environmental samples.
• This method facilitates further physiological studies of these important microorganisms.
• Improved understanding of the nitrogen cycle can aid in mitigating environmental pollution.