Dataset Information

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

ABSTRACT:

Background

The biofilm of a methanol-fed, marine denitrification system is composed of a multi-species microbial community, among which Hyphomicrobium nitrativorans and Methylophaga nitratireducenticrescens are 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 of H. nitrativorans strain NL23 and M. nitratireducenticrescens in the biofilm cultures. Here, we report the impact of these changes on the dynamics of the overall microbial community of the denitrifying biofilm.

Methods

The 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 of H. nitrativorans strain NL23 and M. nitratireducenticrescens strain GP59.

Results

High proportions of M. nitratireducenticrescens occurred in the biofilm cultures. H. nitrativorans strain 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 to Marinobacter spp. and Paracoccus spp. 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 the Stappia spp. were metabolically active in the biofilm cultures.

Conclusions

These 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 of M. nitratireducenticrescens strain GP59 and H. nitrativorans strain 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.

SUBMITTER: Villemur R

PROVIDER: S-EPMC6697039 | biostudies-literature | 2019

REPOSITORIES: biostudies-literature

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Publications

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

Villemur Richard R Payette Geneviève G Geoffroy Valérie V Mauffrey Florian F Martineau Christine C

PeerJ 20190813

<h4>Background</h4>The biofilm of a methanol-fed, marine denitrification system is composed of a multi-species microbial community, among which <i>Hyphomicrobium nitrativorans</i> and <i>Methylophaga nitratireducenticrescens</i> are 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 previous ...[more]

PMID: 31423359

Similar Datasets

Project description:BackgroundThe biofilm of a methanol-fed denitrification system that treated a marine effluent is composed of multi-species microorganisms, among which Hyphomicrobium nitrativorans strain NL23 and Methylophaga nitratireducenticrescens strain JAM1 are the principal bacteria involved in the denitrifying activities. Here, we report the capacity of the denitrifying biofilm to sustain environmental changes, and the impact of these changes on the co-occurrence of H. nitrativorans and M. nitratireducenticrescens.MethodsIn a first set of assays, the original biofilm (OB) was cultivated in an artificial seawater (ASW) medium under anoxic conditions to colonize new carriers. The new formed biofilm was then subjected to short exposures (1-5 days) of a range of NaCl, methanol, nitrate (NO3 -) and nitrite (NO2 -) concentrations, and to different pHs and temperatures. In a second set of assays, the OB was cultivated in ASW medium for five weeks with (i) a range of NaCl concentrations, (ii) four combinations of NO3 -/methanol concentrations and temperatures, (iii) NO2 -, and (iv) under oxic conditions. Finally, the OB was cultivated for five weeks in the commercial Instant Ocean (IO) seawater. The growth of the biofilm and the dynamics of NO3 - and NO2 - were determined. The levels of M. nitratireducenticrescens and H. nitrativorans were measured by qPCR.ResultsIn the first set of assays, the biofilm cultures had the capacity to sustain denitrifying activities in most of the tested conditions. Inhibition occurred when they were exposed to high pH (10) or to high methanol concentration (1.5%). In the second set of assays, the highest specific denitrification rates occurred with the biofilm cultures cultivated at 64.3 mM NO3 - and 0.45% methanol, and at 30 °C. Poor biofilm development occurred with the biofilm cultures cultivated at 5% and 8% NaCl. In all biofilm cultures cultivated in ASW at 2.75% NaCl, H. nitrativorans strain NL23 decreased by three orders of magnitude in concentrations compared to that found in OB. This decrease coincided with the increase of the same magnitude of a subpopulation of M. nitratireducenticrescens (strain GP59 as representative). In the biofilm cultures cultivated at low NaCl concentrations (0% to 1.0%), persistence of H. nitrativorans strain NL23 was observed, with the gradual increase in concentrations of M. nitratireducenticrescens strain GP59. High levels of H. nitrativorans strain NL23 were found in the IO biofilm cultures. The concentrations of M. nitratireducenticrescens strain JAM1 were lower in most of the biofilms cultures than in OB.ConclusionsThese results demonstrate the plasticity of the marine methylotrophic denitrifying biofilm in adapting to different environmental changes. The NaCl concentration is a crucial factor in the dynamics of H. nitrativorans strain NL23, for which growth was impaired above 1% NaCl in the ASW-based biofilm cultures in favor of M. nitratireducenticrescens strain GP59.

Project description:A denitrifying microbial consortium was enriched in an anoxically operated, methanol-fed sequencing batch reactor (SBR) fed with a mineral salts medium containing methanol as the sole carbon source and nitrate as the electron acceptor. The SBR was inoculated with sludge from a biological nutrient removal activated sludge plant exhibiting good denitrification. The SBR denitrification rate improved from less than 0.02 mg of NO(3)(-)-N mg of mixed-liquor volatile suspended solids (MLVSS)(-1) h(-1) to a steady-state value of 0.06 mg of NO(3)(-)-N mg of MLVSS(-1) h(-1) over a 7-month operational period. At this time, the enriched microbial community was subjected to stable-isotope probing (SIP) with [(13)C]methanol to biomark the DNA of the denitrifiers. The extracted [(13)C]DNA and [(12)C]DNA from the SIP experiment were separately subjected to full-cycle rRNA analysis. The dominant 16S rRNA gene phylotype (group A clones) in the [(13)C]DNA clone library was closely related to those of the obligate methylotrophs Methylobacillus and Methylophilus in the order Methylophilales of the Betaproteobacteria (96 to 97% sequence identities), while the most abundant clone groups in the [(12)C]DNA clone library mostly belonged to the family Saprospiraceae in the Bacteroidetes phylum. Oligonucleotide probes for use in fluorescence in situ hybridization (FISH) were designed to specifically target the group A clones and Methylophilales (probes DEN67 and MET1216, respectively) and the Saprospiraceae clones (probe SAP553). Application of these probes to the SBR biomass over the enrichment period demonstrated a strong correlation between the level of SBR denitrification and relative abundance of DEN67-targeted bacteria in the SBR community. By contrast, there was no correlation between the denitrification rate and the relative abundances of the well-known denitrifying genera Hyphomicrobium and Paracoccus or the Saprospiraceae clones visualized by FISH in the SBR biomass. FISH combined with microautoradiography independently confirmed that the DEN67-targeted cells were the dominant bacterial group capable of anoxic [(14)C]methanol uptake in the enriched biomass. The well-known denitrification lag period in the methanol-fed SBR was shown to coincide with a lag phase in growth of the DEN67-targeted denitrifying population. We conclude that Methylophilales bacteria are the dominant denitrifiers in our SBR system and likely are important denitrifiers in full-scale methanol-fed denitrifying sludges.

Dataset Information

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

Background

Methods

Results

Conclusions

Publications

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

Similar Datasets

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