A unique bacteriohopanetetrol stereoisomer of marine anammox

Anaerobic ammonium oxidation (anammox) is a major process of bioavailable nitrogen removal from marine systems. Previously, a bacteriohopanetetrol (BHT) isomer, with unknown stereochemistry, eluting later than BHT using high performance liquid chromatography (HPLC), was detected in ‘Ca. Scalindua profunda’ and proposed as a biomarker for anammox in marine paleo-environments. However, the utility of this BHT isomer as an anammox biomarker is hindered by the fact that four other, non-anammox bacteria are also known to produce a late-eluting BHT stereoisomer. The stereochemistry in Acetobacter pasteurianus, Komagataeibacter xylinus and Frankia sp. was known to be 17β, 21β(H), 22R, 32R, 33R, 34R (BHT-34R). The stereochemistry of the late-eluting BHT in Methylocella palustris was unknown. To determine if marine anammox bacteria produce a unique BHT isomer, we studied the BHT distributions and stereochemistry of known BHT isomer producers and of previously unscreened marine (‘Ca. Scalindua brodeae’) and freshwater (‘Ca. Brocadia sp.’) anammox bacteria using HPLC and gas chromatographic (GC) analysis of acetylated BHTs and ultra high performance liquid chromatography (UHPLC)-high resolution mass spectrometry (HRMS) analysis of non-acetylated BHTs. The 34R stereochemistry was confirmed for the BHT isomers in Ca. Brocadia sp. and Methylocella palustris. However, ‘Ca. Scalindua sp.’ synthesise a stereochemically distinct BHT isomer, with still unconfirmed stereochemistry (BHT-x). Only GC analysis of acetylated BHT and UHPLC analysis of non-acetylated BHT distinguished between late-eluting BHT isomers. Acetylated BHT-x and BHT-34R co-elute by HPLC. As BHT-x is currently only known to be produced by ‘Ca. Scalindua spp.’, it may be a biomarker for marine anammox

Dark carbon fixation in the Arabian Sea oxygen minimum zone contributes to sedimentary organic carbon (SOM)

In response to rising CO2concentrations and increasing global sea surface temperatures,oxygen minimum zones (OMZ), or“dead zones”, are expected to expand. OMZs are fueled by highprimary productivity, resulting in enhanced biological oxygen demand at depth, subsequent oxygen depletion, and attenuation of remineralization. This results in the deposition of organic carbon‐rich sediments. Carbon drawdown is estimated by biogeochemical models; however, a major process is ignored: carbon fixation in the mid‐and lower water column. Here, we show that chemoautotrophic carbon fixation is important in the Arabian Sea OMZ; and manifests in a13C‐depleted signature of sedimentary organic carbon. We determined theδ13C values of Corg deposited in close spatial proximity but over a steepbottom‐water oxygen gradient, and theδ13C composition of biomarkers of chemoautotrophic bacteriacapable of anaerobic ammonia oxidation (anammox). Isotope mixing models show that detritus fromanammox bacteria or other chemoautotrophs likely forms a substantial part of the organic matter depositedwithin the Arabian Sea OMZ (~17%), implying that the contribution of chemoautotrophs to settling organicmatter is exported to the sediment. This has implications for the evaluation of past, and future, OMZs:biogeochemical models that operate on the assumption that all sinking organic matter is photosynthetically derived, without new addition of carbon, could significantly underestimate the extent of remineralization. Oxygen demand in oxygen minimum zones could thus be higher than projections suggest, leading to a more intense expansion of OMZs than expected

Dark CO2 fixation into phospholipid-derived fatty acids by the cold-water coral associated sponge Hymedesmia (Stylopus) coriacea (Tisler Reef, NE Skagerrak)

Many cold-water sponges harbour microorganisms of which the role in the sponge host remains enigmatic. Here, we show a transfer of fixed inorganic carbon by sponge-associated microbes to its host, the cold-water coral encrusting sponge Hymedesmia (Stylopus) coriacea. Sponge were collected at approx. 100 m depth and incubated for 1.5–2.5 days with 13C labelled dissolved inorganic carbon (DIC) as tracer. Total DIC fixation rates ranged from 0.03–0.11 mmol C × mmol Csponge × d−1. 13C-tracer was recovered in bacterial-specific (i.e. short and branched) and sponge-specific (very long-chained) phospholipid-derived fatty acids (PLFA's), but was not incorporated into archaeal lipids. 13C-incorporation in biomarkers such as C16:1w7c and C18:1w7c indicated that nitrifying and/or sulphur-oxidizing bacteria (chemoautotrophs) were likely active in the sponge. Trophic transfer of microbially-fixed carbon to the sponge host was confirmed by recovery of label in very long chain fatty acids (VLCFA's) including C26:2 and C26:3. Tracer accumulation into several VLCFA's continued after removal of 13C-DIC, while tracer in most bacteria-specific PLFA's declined, indicating a transfer and elongation of bacterial-specific PLFA's to sponge-specific PLFA's. This implies that PLFA precursors released from chemo- as well as heterotrophic microbes in sponges contributed to the synthesis of VLCFA's, identifying sponge-associated bacteria as symbionts of the sponge

Archaeal and bacterial glycerol dialkyl glycerol tetraether (GDGT) lipids in environmental samples by high temperature-gas chromatography with flame ionisation and time-of-flight mass spectrometry detection

Archaeal isoprenoidal glycerol dibiphytanyl glycerol tetraether lipids (iGDGTs) and their non-isoprenoidal branched bacterial analogues (brGDGTs) have widespread applications in biogeochemistry and paleothermometry. Analysis of GDGTs usually involves separation using high performance liquid chromatography, typically coupled via atmospheric pressure chemical ionisation to mass spectrometric detection in selected ion-monitoring mode (HPLC–APCI-MS). However, reliable determination of ratios and, in particular, quantification by this technique, can be challenging due to differences in ionisation efficiencies of the various compounds. Quantification of GDGTs also relies on external calibration of the relative response to an internal standard with authenticated GDGTs, which are often not readily accessible. Here, we tested the suitability of high temperature gas chromatography with flame ionisation detection (HTGC-FID) for the determination of concentrations and tetraether lipid-based ratios in marine and terrestrial samples. For this, we identified GDGTs in environmental samples using HTGC coupled to time-of-flight mass spectrometry (HTGC–MS). Using a purified GDGT standard, we show we can quantify GDGT-0 in environmental samples by GC-FID. Some GDGT-based ratios measured by HTGC-FID exhibited a linear correlation (1:1) with ratios derived from HPLC–MS and weight-based ratios of mixtures of purified standards. However, ratios relying on minor isomers, such as TEX86 and MBT/CBT have many unresolved challenges for determination by HTGC. Detection limits were higher than for HPLC–MS. However, the advantages of employing HTGC-based methods include: (1) the independence from MS tuning-related differences in ionisation energies; (2) the potential for direct comparison with other, non-GDGT based biomarkers; and (3) a more complete insight into biomarker distributions in environmental samples by the extension of the temperature range. Quantitative elution of GDGTs from a HTGC column as demonstrated herein, will also enable their analysis by compound-specific isotope ratio mass spectrometry

A comprehensive and comparative phenotypic analysis of the collaborative founder strains identifies new and known phenotypes

The collaborative cross (CC) is a large panel of mouse-inbred lines derived from eight founder strains (NOD/ShiLtJ, NZO/HILtJ, A/J, C57BL/6J, 129S1/SvImJ, CAST/EiJ, PWK/PhJ, and WSB/EiJ). Here, we performed a comprehensive and comparative phenotyping screening to identify phenotypic differences and similarities between the eight founder strains. In total, more than 300 parameters including allergy, behavior, cardiovascular, clinical blood chemistry, dysmorphology, bone and cartilage, energy metabolism, eye and vision, immunology, lung function, neurology, nociception, and pathology were analyzed;in most traits from sixteen females and sixteen males. We identified over 270 parameters that were significantly different between strains. This study highlights the value of the founder and CC strains for phenotype-genotype associations of many genetic traits that are highly relevant to human diseases. All data described here are publicly available from the mouse phenome database for analyses and downloads

A large scale hearing loss screen reveals an extensive unexplored genetic landscape for auditory dysfunction

The developmental and physiological complexity of the auditory system is likely reflected in the underlying set of genes involved in auditory function. In humans, over 150 non-syndromic loci have been identified, and there are more than 400 human genetic syndromes with a hearing loss component. Over 100 non-syndromic hearing loss genes have been identified in mouse and human, but we remain ignorant of the full extent of the genetic landscape involved in auditory dysfunction. As part of the International Mouse Phenotyping Consortium, we undertook a hearing loss screen in a cohort of 3006 mouse knockout strains. In total, we identify 67 candidate hearing loss genes. We detect known hearing loss genes, but the vast majority, 52, of the candidate genes were novel. Our analysis reveals a large and unexplored genetic landscape involved with auditory function

A comprehensive and comparative phenotypic analysis of the collaborative founder strains identifies new and known phenotypes.

The collaborative cross (CC) is a large panel of mouse-inbred lines derived from eight founder strains (NOD/ShiLtJ, NZO/HILtJ, A/J, C57BL/6J, 129S1/SvImJ, CAST/EiJ, PWK/PhJ, and WSB/EiJ). Here, we performed a comprehensive and comparative phenotyping screening to identify phenotypic differences and similarities between the eight founder strains. In total, more than 300 parameters including allergy, behavior, cardiovascular, clinical blood chemistry, dysmorphology, bone and cartilage, energy metabolism, eye and vision, immunology, lung function, neurology, nociception, and pathology were analyzed; in most traits from sixteen females and sixteen males. We identified over 270 parameters that were significantly different between strains. This study highlights the value of the founder and CC strains for phenotype-genotype associations of many genetic traits that are highly relevant to human diseases. All data described here are publicly available from the mouse phenome database for analyses and downloads

Mutation in the mouse histone gene Hist2h3c1 leads to degeneration of the lens vesicle and severe microphthalmia

During an ENU (N-ethyl-N-nitrosourea) mutagenesis screen, we observed a dominant small-eye mutant mouse with viable homozygotes. A corresponding mutant line was established and referred to as Aey69 (abnormality of the eye #69). Comprehensive phenotyping of the homozygous Aey69 mutants in the German Mouse Clinic revealed only a subset of statistically significant alterations between wild types and homozygous mutants. The mutation causes microphthalmia without a lens but with retinal hyperproliferation. Linkage was demonstrated to mouse chromosome 3 between the markers D3Mit188 and D3Mit11. Sequencing revealed a 358 A- > C mutation (I1e120Leu) in the Hist2h3c1 gene and a 71 T- > C (Val24Ala) mutation in the Gja8 gene. Detailed analysis of eye development in the homozygous mutant mice documented a perturbed lens development starting -from the lens vesicle stage including decreasing expression of crystallins as well as of lens-specific transcription - factors like PITX3 and FOXE3. In contrast, we observed an early expression of retinal progenitor cells characterized by several markers including BRN3 (retinal ganglion cells) and OTX2 (cone photoreceptors). The changes in the retina at the early embryonic stages of E11.5-E15.5 happen in parallel with apoptotic processes in the lens at the respective stages. The excessive retinal hyperproliferation is characterized by an increased level of Ki67. The hyperproliferation, however, does not disrupt the differentiation and appearance of the principal retinal cell types at postnatal stages, even if the overgrowing retina covers finally the entire bulbus of the eye. Morpholino-mediated knock-down of the hist2h3ca1 gene in zebrafish leads to a specific perturbation of lens development. When injected into zebrafish zygotes, only the mutant mouse mRNA leads to severe malformations, ranging from cyclopia to severe microphthalmia. The wild-type Hist2h3c1 mRNA can rescue the morpholino-induced defects corroborating its specific function in lens development. Based upon these data, it is concluded that the ocular function of the Hist2h3c1 gene (encoding a canonical H3.2 variant) is conserved throughout evolution. Moreover, the data highlight also the importance of Hist2h3c1 in the coordinated formation of lens and retina during eye development

δ¹³C values of water column organic carbon in the Arabian Sea oxygen minimum zone

Freeze-dried punches from 0.7 µm GFF filters for suspended particulate organic matter were decalcified with 2N HCl, washed, freeze-dried, and subjected to analysis via a Flash EA 1112 Series (Thermo Scientific) analyser, coupled via a Conflo II interface to a Finnigan DELTA plus mass spectrometer as described by and Pitcher et al. (2011; doi:10.1038/ismej.2011.60). Standards for δ¹³C analysis were acetanilide and benzoic acid and samples were analysed in duplicate