253 Synthesis and discussion ON “Ca. Methanaperedens” METABOLIC RESILIENCE AND ADAPTATION (CHAPTER 3 TO 7) “Ca. Methanoperedens” as a model for top-down research This PhD dissertation employed a top-down methodological approach, leveraging the ecological importance of “Ca. Methanoperedens” across diverse natural ecosystems (Chapter 7) as anaerobic methanotrophs. This approach was made possible due to the availability of two highly enriched cultures: “Ca. Methanoperedens BLZ2” (as N-DAMO) and “Ca. Methanoperedens Vercelli”, discussed in Chapters 5 and 6, respectively. Furthermore, “Ca. Methanoperedens” was studied in both stable and dynamic cultures, allowing for the exploration of its role alongside microaerophilic methanotrophs in (micro)anoxic and denitrifying environments (Chapter 3). It was also examined in potential engineered ecosystem consortia involving anammox and N-DAMO processes (Chapter 4). Metabolic flexibility of “Ca. Methanoperedens” via extrachromosomal elements (ECEs) Borgs are described to harbor key metabolic and stress-related genes, including MHCs and some also the methyl-coenzyme A reductase (Al-Shayeb et al., 2022). In Chapters 6 and 7, we examine the presence of novel ECEs, Borgs, which have been associated with environmental “Ca. Methanoperedens” species. Specifically, in Chapter 6, we discuss Borgs in the context of osmolyte production, including N-acetyl-beta-l-lysine, which may help “Ca. Methanoperedens” spp. cope with increased salinities. We hypothesize that the genes responsible for the production of N-acetyl-beta-l-lysine may have been absorbed by the Borgs from “Ca. Methanoperedens” spp. This is an intriguing observation, as it suggests the potential loss of key genes for salinity adaptation from the host genome to the Borgs. Furthermore, we propose that the genes kamA and ablB, involved in N-acetylbeta-l-lysine production, may have been acquired through horizontal gene transfer (HGT) from the Firmicutes group. In Chapter 6, we also briefly describe the response of the plasmid harbored in “Ca. Methanoperedens Vercelli” to salinity stress, noting the downregulation of genes associated with cell motility (e.g., FlaI and type IV secretion systems). 8
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