Changes in the distribution of membrane lipids during growth of <i>Thermotoga maritima</i> at different temperatures: Indications for the potential mechanism of biosynthesis of ether-bound diabolic acid (membrane-spanning) lipids

Membrane-spanning lipids are present in a wide variety of archaea, but they are rarely in bacteria. Nevertheless, the (hyper)thermophilic members of the order Thermotogales harbor tetraester, tetraether, and mixed ether/ester membrane-spanning lipids mostly composed of core lipids derived from diabolic acids, C30, C32, and C34 dicarboxylic acids with two adjacent mid-chain methyl substituents. Lipid analysis of Thermotoga maritima across growth phases revealed a decrease of the relative abundance of fatty acids together with an increase of diabolic acids with independence of growth temperature. We also identified isomers of C30 and C32 diabolic acids, i.e., dicarboxylic acids with only one methyl group at C-15. Their distribution suggests they are products of the condensation reaction but are preferably produced when the length of the acyl chains is not optimal. Compared with growth at the optimal temperature of 80°C, an increase of glycerol ether-derived lipids was observed at 55°C. Our analysis only detected diabolic acid-containing intact polar lipids with phosphoglycerol (PG) head groups. Considering these findings, we hypothesize a biosynthetic pathway for the synthesis of membrane-spanning lipids based on PG polar lipid formation, suggesting that the protein catalyzing this process is a membrane protein. We also identified, by genomic and protein domainanalyses, a gene coding for a putative plasmalogen synthase homologue inT. maritima that is also present in other bacteria producing sn-1-alkyl ether lipids but not plasmalogens, suggesting it is involved in the conversion of the ester-to-ether bond in the diabolic acids bound in membrane-spanning lipids.</p

Transcriptome Analysis Reveals Cr(VI) Adaptation Mechanisms in <i>Klebsiella</i> sp. Strain AqSCr

Klebsiella sp. strain AqSCr, isolated from Cr(VI)-polluted groundwater, reduces Cr(VI) both aerobically and anaerobically and resists up 34 mM Cr(VI); this resistance is independent of the ChrA efflux transporter. In this study, we report the whole genome sequence and the transcriptional profile by RNA-Seq of strain AqSCr under Cr(VI)-adapted conditions and found 255 upregulated and 240 downregulated genes compared to controls without Cr(VI) supplementation. Genes differentially transcribed were mostly associated with oxidative stress response, DNA repair and replication, sulfur starvation response, envelope-osmotic stress response, fatty acid (FA) metabolism, ribosomal subunits, and energy metabolism. Among them, genesnot previously associated with chromium resistance, for example, cybB, encoding a putative superoxide oxidase (SOO), gltA2 , encoding an alternative citrate synthase, and des, encoding a FA desaturase, were upregulated. The sodA gene encoding a manganesesuperoxide dismutase was upregulated in the presence of Cr(VI), whereas sodB encoding an iron superoxide dismutase was downregulated. Cr(VI) resistance mechanisms in strain AqSCr seem to be orchestrated by the alternative sigma factors fecl, rpoE, and rpoS (all of them upregulated). Membrane lipid analysis of the Cr(IV)-adapted strain showed a lower proportion of unsaturated lipids with respect to the control, which we hypothesized could result from unsaturated lipid peroxidation followed by degradation, together with de novosynthesis mediated by the upregulated FA desaturase-encoding gene, des. This report helps to elucidate both Cr(VI) toxicity targets and global bacterial response to Cr(VI).</p

Demethylated hopanoids in ‘Ca. Methylomirabilis oxyfera’ as biomarkers for environmental nitrite-dependent methane oxidation

Hopanoids are lipids that are widespread in the bacterial domain and well established molecular biomarkers in modern and paleo environments. In particular, the occurrence of 13C-depleted 3-methylated hopanoids are characteristic of aerobic bacteria involved in methane oxidation. Previously the intra-aerobic methanotroph ‘Candidatus Methylomirabilis oxyfera’ (‘Ca. M. oxyfera’), which performs nitrite-dependent methane oxidation in anoxic environments, has been shown to synthesize bacteriohopanepolyols (BHPs) and their 3-methylated counterparts. However, since ‘Ca. M. oxyfera’ does not utilize methane as a carbon source, its biomass and lipids do not show the characteristic 13C-depletion. Therefore, the detection of ‘Ca. M. oxyfera’ in various environments is challenging, and still underexplored. Here, we re-investigated the hopanoid content of ‘Ca. M. oxyfera’ bacteria using enrichment cultures. We found the GC-amenable hopanoids of ‘Ca. M. oxyfera’ to be dominated by four demethylated hopanoids of which only one, 22,29,30-trisnorhopan-21-one, had been identified previously. The three novel hopanoids were tentatively identified as 22,29,30-trisnorhopan-21-ol, 3-methyl-22,29,30-trisnorhopan-21-one and 3-methyl-22,29,30-trisnorhopan-21-ol. These unique demethylated hopanoids are most likely biosynthesized directly by ‘Ca. M. oxyfera’ bacteria. Bioinformatical analysis of the ‘Ca. M. oxyfera’ genome revealed potential candidate genes responsible for the demethylation of hopanoids. For the sensitive detection of the four trisnorhopanoid biomarkers in environmental samples, a multiple reaction monitoring (MRM) method was developed and used to successfully detect the trisnorhopanoids in a peatland where the presence of ‘Ca. M. oxyfera’ had been confirmed previously by DNA-based analyses. These new biomarkers may be a novel tool to trace nitrite-dependent methane oxidation in various (past) environments

Demethylated hopanoids in ‘Ca. Methylomirabilis oxyfera’ as biomarkers for environmental nitrite-dependent methane oxidation

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