125 Unraveling nitrogen, sulfur and carbon microbial bioreactor responses to stress methane monooxygenase (pmoABC) coupled to nitric oxide dismutation, with four putative nod genes present and transcribed. Additionally, nitrate, nitrite and nitric oxide reductase-encoding genes were present and transcribed: nxrABC, ammonium-forming cytoplasmic NADH-nitrite reductase nirBD genes, electrogenic cytochrome c-oxidizing (sNOR) nitric oxide reductase-encoding norSY genes, nonelectrogenic quinone-oxidizing (gNOR) nitric oxide reductase-encoding norGH genes, and norZ. All four genes encoding the two subunits of the putative nitric oxide dismutases NOD-1 and NOD-2 had some of the highest transcription (TPM ~ 7-60) when the community was being primed for the sulfide and NO toxicity experiment, receiving ~ 1-5% external NO in the reactor along with methane, ammonium and sulfide (T4), as well as pmoABC (TPM ~ 6-22) and the lanthanidedependent methanol dehydrogenase-encoding xoxF gene (TPM = 5.5 ± 3.5) (Supplementary Table 2). Although “Ca. Methylomirabilis tolerans” was still abundant after the sulfide and NO toxicity experiment (Figure 3), TPM values for all aforementioned functional genes decreased 10-100 fold from T4 to T5, indicating that the microorganism suffered a degree of inhibition (Supplementary Table 2). MAG 60 Thermoanaerobaculia 1, 88.3% complete with 1.5% contamination, had highest normalized genome coverage when the reactor was being primed for the sulfide and NO toxicity experiment (Figure 3, G4). Based on metatranscriptomic analyses, the most likely metabolism performed by this organism was heterotrophic denitrification (Figure 4). A nitrous oxide reductase nosZ gene was among the functional genes with highest transcription (T4, TPM = 0.21 ± 0.17), followed by narGHI and nrfAH. Interestingly, hydrogenase-2 and low affinity cytochrome c oxidase-encoding ctaCDEF genes were also transcribed, as well as all electron transport chain-encoding genes, including two RNF complexes (Supplementary Table 2 and 4). Finally, “Ca. Kuenenia stuttgartiensis”, represented by MAG 32, 95.6% complete with 0.6% contamination, was one of two microorganisms that persisted throughout all regular and experimental conditions (Figure 3). Interestingly, the transcription of hydrazine synthase hzsABC genes decreased 18-27x during the ammonium removal experiment (T1 to T2), and 2-5x during the sulfide and NO toxicity experiment (T4 to T5), suggesting that substrate deprivation was a stronger 4
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