Dynamics of a methanol-fed marine denitrifying biofilm: 2—impact of environmental changes on the microbial community

Abstract

BackgroundThe biofilm of a methanol-fed, marine denitrification system is composed of a multi-species microbial community, among whichHyphomicrobium nitrativoransandMethylophaga nitratireducenticrescensare the principal bacteria involved in the denitrifying activities. To assess its resilience to environmental changes, the biofilm was cultivated in artificial seawater (ASW) under anoxic conditions and exposed to a range of specific environmental conditions. We previously reported the impact of these changes on the denitrifying activities and the co-occurrence ofH. nitrativoransstrain NL23 andM. nitratireducenticrescensin the biofilm cultures. Here, we report the impact of these changes on the dynamics of the overall microbial community of the denitrifying biofilm.MethodsThe original biofilm (OB) taken from the denitrification system was cultivated in ASW under anoxic conditions with a range of NaCl concentrations, and with four combinations of nitrate/methanol concentrations and temperatures. The OB was also cultivated in the commercial Instant Ocean seawater (IO). The bacterial diversity of the biofilm cultures and the OB was determined by 16S ribosomal RNA gene sequences. Culture approach was used to isolate other denitrifying bacteria from the biofilm cultures. The metatranscriptomes of selected biofilm cultures were derived, along with the transcriptomes of planktonic pure cultures ofH. nitrativoransstrain NL23 andM. nitratireducenticrescensstrain GP59.ResultsHigh proportions ofM. nitratireducenticrescensoccurred in the biofilm cultures.H. nitrativoransstrain NL23 was found in high proportion in the OB, but was absent in the biofilm cultures cultivated in the ASW medium at 2.75% NaCl. It was found however in low proportions in the biofilm cultures cultivated in the ASW medium at 0–1% NaCl and in the IO biofilm cultures. Denitrifying bacterial isolates affiliated toMarinobacterspp. andParacoccusspp. were isolated. Up regulation of the denitrification genes of strains GP59 and NL23 occurred in the biofilm cultures compared to the planktonic pure cultures. Denitrifying bacteria affiliated to theStappiaspp. were metabolically active in the biofilm cultures.ConclusionsThese results illustrate the dynamics of the microbial community in the denitrifying biofilm cultures in adapting to different environmental conditions. The NaCl concentration is an important factor affecting the microbial community in the biofilm cultures. Up regulation of the denitrification genes ofM. nitratireducenticrescensstrain GP59 andH. nitrativoransstrain NL23 in the biofilm cultures suggests different mechanisms of regulation of the denitrification pathway in the biofilm. Other denitrifying heterotrophic bacteria are present in low proportions, suggesting that the biofilm has the potential to adapt to heterotrophic, non-methylotrophic environments.