Limits...
pH-driven shifts in overall and transcriptionally active denitrifiers control gaseous product stoichiometry in growth experiments with extracted bacteria from soil.

Brenzinger K, Dörsch P, Braker G - Front Microbiol (2015)

Bottom Line: We found that denitrifier community composition, abundance and transcription changed throughout incubation concomitant with pH change in the medium, allowing for complete reduction of nitrate to N2 with little accumulation of intermediates.When exposed to pH 5.4, the denitrifier community was able to grow but reduced N2O to N2 only when near-neutral pH was reestablished by the alkalizing metabolic activity of an acid-tolerant part of the community.Denitrifiers of the nirS-type appeared to be severely suppressed by low pH and nirK-type and nosZ-containing denitrifiers showed strongly reduced transcriptional activity and growth, even after restoration of neutral pH.

View Article: PubMed Central - PubMed

Affiliation: Department of Biogeochemistry, Max Planck Institute for Terrestrial Microbiology Marburg, Germany.

ABSTRACT
Soil pH is a strong regulator for activity as well as for size and composition of denitrifier communities. Low pH not only lowers overall denitrification rates but also influences denitrification kinetics and gaseous product stoichiometry. N2O reductase is particularly sensitive to low pH which seems to impair its activity post-transcriptionally, leading to higher net N2O production. Little is known about how complex soil denitrifier communities respond to pH change and whether their ability to maintain denitrification over a wider pH range relies on phenotypic redundancy. In the present study, we followed the abundance and composition of an overall and transcriptionally active denitrifier community extracted from a farmed organic soil in Sweden (pH H2O = 7.1) when exposed to pH 5.4 and drifting back to pH 6.6. The soil was previously shown to retain much of its functioning (low N2O/N2 ratios) over a wide pH range, suggesting a high functional versatility of the underlying community. We found that denitrifier community composition, abundance and transcription changed throughout incubation concomitant with pH change in the medium, allowing for complete reduction of nitrate to N2 with little accumulation of intermediates. When exposed to pH 5.4, the denitrifier community was able to grow but reduced N2O to N2 only when near-neutral pH was reestablished by the alkalizing metabolic activity of an acid-tolerant part of the community. The genotypes proliferating under these conditions differed from those dominant in the control experiment run at neutral pH. Denitrifiers of the nirS-type appeared to be severely suppressed by low pH and nirK-type and nosZ-containing denitrifiers showed strongly reduced transcriptional activity and growth, even after restoration of neutral pH. Our study suggests that low pH episodes alter transcriptionally active populations which shape denitrifier communities and determine their gas kinetics.

No MeSH data available.


NMDS plots of denitrifier communities based on cDNA- and DNA-derived T-RFLP analysis of nirK, nirS, and nosZ from three pooled samples. Data points represent averaged results of three replicate T-RFLP analyses. Community similarity was calculated by using the statistical program R and the Bray–Curtis similarity measurement, which includes presence and relative abundance of T-RF. Clusters and arrows were inserted manually to highlight clustering and community development. Significant differences in the composition of denitrifier communities at given time points were determined by ANOSIM (P < 0.05). (A)nirK; (B)nirS; (C)nosZ.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4585170&req=5

Figure 5: NMDS plots of denitrifier communities based on cDNA- and DNA-derived T-RFLP analysis of nirK, nirS, and nosZ from three pooled samples. Data points represent averaged results of three replicate T-RFLP analyses. Community similarity was calculated by using the statistical program R and the Bray–Curtis similarity measurement, which includes presence and relative abundance of T-RF. Clusters and arrows were inserted manually to highlight clustering and community development. Significant differences in the composition of denitrifier communities at given time points were determined by ANOSIM (P < 0.05). (A)nirK; (B)nirS; (C)nosZ.

Mentions: Community composition data indicated selective transcriptional activity, followed by growth of only a few organisms (Figures S1A, S2A, S3A). Terminal restriction fragments (T-RFs) of 229 bp (representing nirK most closely related to nirK of Alcaligenes xylosoxidans) and of 37 bp length (38 bp in silico representing nosZ most closely related to nosZ of Pseudomonas denitrificans, Ps. stutzeri, and Ps. aeruginosa), (Table S5) which were of little abundance in or absent from the inocula, respectively, dominated the transcriptionally active nirK- and nosZ-containing denitrifier communities (Figures S1A, S3A). For nirS, a genotype most closely related to nirS of Ps. migulae (105-bp T-RF) was transcriptionally activated and proliferated that was not even detectable in the initial community (Figure S2A). Still, the composition of the transcriptionally active (TADC) and the overall denitrifier community (ODC) converged throughout the first 96 h of incubation as indicated by multi-dimensional scaling of T-RFs (Figures 5A–C; ANOSIM26–49h: P < 0.05; R between 0.423 and 0.873; ANOSIM70–96h: P>0.05; R between 0.142 and 0.275). The shifts in denitrifier community composition and the decrease in denitrifier diversity (Shannon index, Figures S1A–S3A) did not result in impairment of function, i.e., gaseous intermediates were efficiently taken up and reduced to N2 (Figures 1A,B). This suggests that it was not the microbial diversity per se that mediated the community's functioning, but the specific metabolic capacities of the dominating denitrifying taxa. Transcription of denitrification genes decreased after all nitrogen oxides were depleted (Figure 2A) and the number of transcripts relative to gene copies became very low (Figures 3A–C). Hence, the increase in diversity and shift in cDNA composition observed for nirK and nosZ-containing denitrifiers at 206 h was presumably the result of transcript degradation following starvation (Figures S1A, S3A).


pH-driven shifts in overall and transcriptionally active denitrifiers control gaseous product stoichiometry in growth experiments with extracted bacteria from soil.

Brenzinger K, Dörsch P, Braker G - Front Microbiol (2015)

NMDS plots of denitrifier communities based on cDNA- and DNA-derived T-RFLP analysis of nirK, nirS, and nosZ from three pooled samples. Data points represent averaged results of three replicate T-RFLP analyses. Community similarity was calculated by using the statistical program R and the Bray–Curtis similarity measurement, which includes presence and relative abundance of T-RF. Clusters and arrows were inserted manually to highlight clustering and community development. Significant differences in the composition of denitrifier communities at given time points were determined by ANOSIM (P < 0.05). (A)nirK; (B)nirS; (C)nosZ.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4585170&req=5

Figure 5: NMDS plots of denitrifier communities based on cDNA- and DNA-derived T-RFLP analysis of nirK, nirS, and nosZ from three pooled samples. Data points represent averaged results of three replicate T-RFLP analyses. Community similarity was calculated by using the statistical program R and the Bray–Curtis similarity measurement, which includes presence and relative abundance of T-RF. Clusters and arrows were inserted manually to highlight clustering and community development. Significant differences in the composition of denitrifier communities at given time points were determined by ANOSIM (P < 0.05). (A)nirK; (B)nirS; (C)nosZ.
Mentions: Community composition data indicated selective transcriptional activity, followed by growth of only a few organisms (Figures S1A, S2A, S3A). Terminal restriction fragments (T-RFs) of 229 bp (representing nirK most closely related to nirK of Alcaligenes xylosoxidans) and of 37 bp length (38 bp in silico representing nosZ most closely related to nosZ of Pseudomonas denitrificans, Ps. stutzeri, and Ps. aeruginosa), (Table S5) which were of little abundance in or absent from the inocula, respectively, dominated the transcriptionally active nirK- and nosZ-containing denitrifier communities (Figures S1A, S3A). For nirS, a genotype most closely related to nirS of Ps. migulae (105-bp T-RF) was transcriptionally activated and proliferated that was not even detectable in the initial community (Figure S2A). Still, the composition of the transcriptionally active (TADC) and the overall denitrifier community (ODC) converged throughout the first 96 h of incubation as indicated by multi-dimensional scaling of T-RFs (Figures 5A–C; ANOSIM26–49h: P < 0.05; R between 0.423 and 0.873; ANOSIM70–96h: P>0.05; R between 0.142 and 0.275). The shifts in denitrifier community composition and the decrease in denitrifier diversity (Shannon index, Figures S1A–S3A) did not result in impairment of function, i.e., gaseous intermediates were efficiently taken up and reduced to N2 (Figures 1A,B). This suggests that it was not the microbial diversity per se that mediated the community's functioning, but the specific metabolic capacities of the dominating denitrifying taxa. Transcription of denitrification genes decreased after all nitrogen oxides were depleted (Figure 2A) and the number of transcripts relative to gene copies became very low (Figures 3A–C). Hence, the increase in diversity and shift in cDNA composition observed for nirK and nosZ-containing denitrifiers at 206 h was presumably the result of transcript degradation following starvation (Figures S1A, S3A).

Bottom Line: We found that denitrifier community composition, abundance and transcription changed throughout incubation concomitant with pH change in the medium, allowing for complete reduction of nitrate to N2 with little accumulation of intermediates.When exposed to pH 5.4, the denitrifier community was able to grow but reduced N2O to N2 only when near-neutral pH was reestablished by the alkalizing metabolic activity of an acid-tolerant part of the community.Denitrifiers of the nirS-type appeared to be severely suppressed by low pH and nirK-type and nosZ-containing denitrifiers showed strongly reduced transcriptional activity and growth, even after restoration of neutral pH.

View Article: PubMed Central - PubMed

Affiliation: Department of Biogeochemistry, Max Planck Institute for Terrestrial Microbiology Marburg, Germany.

ABSTRACT
Soil pH is a strong regulator for activity as well as for size and composition of denitrifier communities. Low pH not only lowers overall denitrification rates but also influences denitrification kinetics and gaseous product stoichiometry. N2O reductase is particularly sensitive to low pH which seems to impair its activity post-transcriptionally, leading to higher net N2O production. Little is known about how complex soil denitrifier communities respond to pH change and whether their ability to maintain denitrification over a wider pH range relies on phenotypic redundancy. In the present study, we followed the abundance and composition of an overall and transcriptionally active denitrifier community extracted from a farmed organic soil in Sweden (pH H2O = 7.1) when exposed to pH 5.4 and drifting back to pH 6.6. The soil was previously shown to retain much of its functioning (low N2O/N2 ratios) over a wide pH range, suggesting a high functional versatility of the underlying community. We found that denitrifier community composition, abundance and transcription changed throughout incubation concomitant with pH change in the medium, allowing for complete reduction of nitrate to N2 with little accumulation of intermediates. When exposed to pH 5.4, the denitrifier community was able to grow but reduced N2O to N2 only when near-neutral pH was reestablished by the alkalizing metabolic activity of an acid-tolerant part of the community. The genotypes proliferating under these conditions differed from those dominant in the control experiment run at neutral pH. Denitrifiers of the nirS-type appeared to be severely suppressed by low pH and nirK-type and nosZ-containing denitrifiers showed strongly reduced transcriptional activity and growth, even after restoration of neutral pH. Our study suggests that low pH episodes alter transcriptionally active populations which shape denitrifier communities and determine their gas kinetics.

No MeSH data available.