Evaluation of Paraspinal Musculature in Small Breed Dogs with and without Atlantoaxial Instability Using Computed Tomography.

OBJECTIVE  The aim of this study was to evaluate differences in paraspinal musculature between dogs with and without atlantoaxial instability (AAI) using computed tomography scans. STUDY DESIGN  Retrospective multicentre study evaluating transverse reconstructed computed tomography scans of 83 small breed dogs (34 with and 49 without AAI) for the cross-sectional paraspinal musculature area at three levels (Occiput/C1, mid-C1, mid-C2). Ratio of moments, dorsal-to-ventral muscle-area ratios (d-v-ratio) and ratios of the dorsal and ventral musculature to C2 height (d-C2-ratio and v-C2-ratio) were evaluated for differences between groups using multivariate analysis of variance (p < 0.05) taking the head-neck position into account. RESULTS  Dogs with AAI showed a significantly lower d-v-ratio at levels 2 and 3, d-C2-ratio at level 2 and ratio of moments at all levels. When head-neck positions were analysed separately, ratio of moments was significantly lower in affected dogs at level 1 and 2. Also lower was d-C2-ratio at level 2, but only in flexed positioning. The head-neck position had a significant influence on ratio of moments and d-v-ratio at all three levels and on d-C2-ratio at level 1. CONCLUSION  Significant changes in muscle area were observed only for the hypaxial muscles at the C1 level, indicating a limited role of muscular adaption in AAI patients. Our results confirm an altered ratio of moments in dogs with AAI. The head-neck position has a significant impact and should be taken into account when evaluating spinal musculature

Domain analysis of lipoprotein LppQ in Mycoplasma mycoides subsp. mycoides SC

The lipoprotein LppQ is the most prominent antigen of Mycoplasma mycoides subsp. mycoides small colony type (SC) during infection of cattle. This pathogen causes contagious bovine pleuropneumonia (CBPP), a devastating disease of considerable socio-economic importance in many countries worldwide. The dominant antigenicity and high specificity for M. mycoides subsp. mycoides SC of lipoprotein LppQ have been exploited for serological diagnosis and for epidemiological investigations of CBPP. Scanning electron microscopy and immunogold labelling were used to provide ultrastructural evidence that LppQ is located to the cell membrane at the outer surface of M. mycoides subsp. mycoides SC. The selectivity and specificity of this method were demonstrated through discriminating localization of extracellular (i.e., in the zone of contact with host cells) vs. integral membrane domains of LppQ. Thus, our findings support the suggestion that the accessible N-terminal domain of LppQ is surface exposed and such surface localization may be implicated in the pathogenesis of CBPP

The IPCC’s reductive Common Era temperature history

J.E. acknowledges support by the Gutenberg Research College, J.E. M.T. and U.B. by the project AdAgrif (CZ.02.01.01/00/22_008/0004635) and ERC (AdG 882727), J.E.S. by the US NSF (OISE-1743738, AGS-2101214 and AGS-2303352), K.J.A. by the US NSF (AGS-1803946 and AGS-2102993), K.A. by the ARC (FT200100102), R.D. by the US NSF (OPP-2112314, OPP-2124885, and AGS-2102759), S.G and M. Stoffel by the SNSF (Sinergia CRSII5_183571), F.C.L. by the SRC (grant no. 2018-01272), Marianne and Marcus Wallenberg Foundation (grant no. MMW 2022-0114) and Swedish Collegium for Advanced Study (Pro Futura Scientia XIII Fellow), LS by the German Research Foundation (SCHN 1645/1-1), M. Sigl by the ERC (CoG 820047), and R.W. by the NSF-NERC (NE/W007223/1).Common Era temperature variability has been a prominent component in Intergovernmental Panel on Climate Change reports over the last several decades and was twice featured in their Summary for Policymakers. A single reconstruction of mean Northern Hemisphere temperature variability was first highlighted in the 2001 Summary for Policymakers, despite other estimates that existed at the time. Subsequent reports assessed many large-scale temperature reconstructions, but the entirety of Common Era temperature history in the most recent Sixth Assessment Report of the Intergovernmental Panel on Climate Change was restricted to a single estimate of mean annual global temperatures. We argue that this focus on a single reconstruction is an insufficient summary of our understanding of temperature variability over the Common Era. We provide a complementary perspective by offering an alternative assessment of the state of our understanding in high-resolution paleoclimatology for the Common Era and call for future reports to present a more accurate and comprehensive assessment of our knowledge about this important period of human and climate history.Peer reviewe

Contemporary multicenter outcomes of continent cutaneous ileocecocystoplasty in the adult population over a 10-year period: A Neurogenic Bladder Research Group study

AIMS: Evidence is sparse on the long-term outcomes of continent cutaneous ileocecocystoplasty (CCIC). We hypothesized that obesity, laparoscopic/robotic approach, and concomitant surgeries would affect morbidity after CCIC and aimed to evaluate the outcomes of CCIC in adults in a multicenter contemporary study. METHODS: We retrospectively reviewed the charts of adult patients from sites in the Neurogenic Bladder Research Group undergoing CCIC (2007-2017) who had at least 6 months of follow-up. We evaluated patient demographics, surgical details, 90-day complications, and follow-up surgeries. the Mann-Whitney U test was used to compare continuous variables and χ² and Fisher\u27s Exact tests were used to compare categorical variables. RESULTS: We included 114 patients with a median age of 41 years. The median postoperative length of stay was 8 days. At 3 months postoperatively, major complications occurred in 18 (15.8%), and 24 patients (21.1%) were readmitted. During a median follow-up of 40 months, 48 patients (42.1%) underwent 80 additional related surgeries. Twenty-three patients (20.2%) underwent at least one channel revision, most often due to obstruction (15, 13.2%) or incontinence (4, 3.5%). Of the channel revisions, 10 (8.8%) were major and 14 (12.3%) were minor. Eleven patients (9.6%) abandoned the catheterizable channel during the follow-up period. Obesity and laparoscopic/robotic surgical approach did not affect outcomes, though concomitant surgery was associated with a higher rate of follow-up surgeries. CONCLUSIONS: In this contemporary multicenter series evaluating CCIC, we found that the short-term major complication rate was low, but many patients require follow-up surgeries, mostly related to the catheterizable channel

Elongase Reactions as Control Points in Long-Chain Polyunsaturated Fatty Acid Synthesis

Extent: 9p.Background: Δ6-Desaturase (Fads2) is widely regarded as rate-limiting in the conversion of dietary α-linolenic acid (18:3n-3; ALA) to the long-chain omega-3 polyunsaturated fatty acid docosahexaenoic acid (22:6n-3; DHA). However, increasing dietary ALA or the direct Fads2 product, stearidonic acid (18:4n-3; SDA), increases tissue levels of eicosapentaenoic acid (20:5n-3; EPA) and docosapentaenoic acid (22:5n-3; DPA), but not DHA. These observations suggest that one or more control points must exist beyond ALA metabolism by Fads2. One possible control point is a second reaction involving Fads2 itself, since this enzyme catalyses desaturation of 24:5n-3 to 24:6n-3, as well as ALA to SDA. However, metabolism of EPA and DPA both require elongation reactions. This study examined the activities of two elongase enzymes as well as the second reaction of Fads2 in order to concentrate on the metabolism of EPA to DHA. Methodology/Principal Findings: The substrate selectivities, competitive substrate interactions and dose response curves of the rat elongases, Elovl2 and Elovl5 were determined after expression of the enzymes in yeast. The competitive substrate interactions for rat Fads2 were also examined. Rat Elovl2 was active with C20 and C22 polyunsaturated fatty acids and this single enzyme catalysed the sequential elongation reactions of EPA→DPA→24:5n-3. The second reaction DPA→24:5n-3 appeared to be saturated at substrate concentrations not saturating for the first reaction EPA→DPA. ALA dose-dependently inhibited Fads2 conversion of 24:5n-3 to 24:6n-3. Conclusions: The competition between ALA and 24:5n-3 for Fads2 may explain the decrease in DHA levels observed after certain intakes of dietary ALA have been exceeded. In addition, the apparent saturation of the second Elovl2 reaction, DPA→24:5n-3, provides further explanations for the accumulation of DPA when ALA, SDA or EPA is provided in the diet. This study suggests that Elovl2 will be critical in understanding if DHA synthesis can be increased by dietary means.Melissa K. Gregory, Robert A. Gibson, Rebecca J. Cook-Johnson, Leslie G. Cleland and Michael J. Jame

High-confidence glycosome proteome for procyclic form <em>Trypanosoma brucei</em> by epitope-tag organelle enrichment and SILAC proteomics

The glycosome of the pathogenic African trypanosome Trypanosoma brucei is a specialized peroxisome that contains most of the enzymes of glycolysis and several other metabolic and catabolic pathways. The contents and transporters of this membrane-bounded organelle are of considerable interest as potential drug targets. Here we use epitope tagging, magnetic bead enrichment, and SILAC quantitative proteomics to determine a high-confidence glycosome proteome for the procyclic life cycle stage of the parasite using isotope ratios to discriminate glycosomal from mitochondrial and other contaminating proteins. The data confirm the presence of several previously demonstrated and suggested pathways in the organelle and identify previously unanticipated activities, such as protein phosphatases. The implications of the findings are discussed

An anchored chromosome-scale genome assembly of spinach improves annotation and reveals extensive gene rearrangements in euasterids.

Spinach (Spinacia oleracea L.) is a member of the Caryophyllales family, a basal eudicot asterid that consists of sugar beet (Beta vulgaris L. subsp. vulgaris), quinoa (Chenopodium quinoa Willd.), and amaranth (Amaranthus hypochondriacus L.). With the introduction of baby leaf types, spinach has become a staple food in many homes. Production issues focus on yield, nitrogen-use efficiency and resistance to downy mildew (Peronospora effusa). Although genomes are available for the above species, a chromosome-level assembly exists only for quinoa, allowing for proper annotation and structural analyses to enhance crop improvement. We independently assembled and annotated genomes of the cultivar Viroflay using short-read strategy (Illumina) and long-read strategies (Pacific Biosciences) to develop a chromosome-level, genetically anchored assembly for spinach. Scaffold N50 for the Illumina assembly was 389 kb, whereas that for Pacific BioSciences was 4.43 Mb, representing 911 Mb (93% of the genome) in 221 scaffolds, 80% of which are anchored and oriented on a sequence-based genetic map, also described within this work. The two assemblies were 99.5% collinear. Independent annotation of the two assemblies with the same comprehensive transcriptome dataset show that the quality of the assembly directly affects the annotation with significantly more genes predicted (26,862 vs. 34,877) in the long-read assembly. Analysis of resistance genes confirms a bias in resistant gene motifs more typical of monocots. Evolutionary analysis indicates that Spinacia is a paleohexaploid with a whole-genome triplication followed by extensive gene rearrangements identified in this work. Diversity analysis of 75 lines indicate that variation in genes is ample for hypothesis-driven, genomic-assisted breeding enabled by this work

The small compound inhibitor K22 displays broad antiviral activity against different members of the family Flaviviridae and offers potential as pan-viral inhibitor.

The virus family Flaviviridae encompasses several viruses, including (re)emerging viruses which cause widespread morbidity and mortality throughout the world. Members of this virus family are positive-strand RNA viruses and replicate their genome in close association with reorganized intracellular host cell membrane compartments. This evolutionarily conserved strategy facilitates efficient viral genome replication and contributes to evasion from host cell cytosolic defense mechanisms. We have previously described the identification of a small-compound inhibitor, K22, which exerts a potent antiviral activity against a broad range of coronaviruses by targeting membrane-bound viral RNA replication. To analyze the antiviral spectrum of this inhibitor, we assessed the inhibitory potential of K22 against several members of the Flaviviridae family, including the reemerging Zika virus (ZIKV). We show that ZIKV is strongly affected by K22. Time-of-addition experiments revealed that K22 acts during a postentry phase of the ZIKV life cycle, and combination regimens of K22 together with ribavirin (RBV) or interferon alpha (IFN-α) further increased the extent of viral inhibition. Ultrastructural electron microscopy studies revealed severe alterations of ZIKV-induced intracellular replication compartments upon infection of K22-treated cells. Importantly, the antiviral activity of K22 was demonstrated against several other members of the Flaviviridae family. It is tempting to speculate that K22 exerts its broad antiviral activity against several positive-strand RNA viruses via a similar mechanism and thereby represents an attractive candidate for development as a panviral inhibitor

Deletion of transketolase triggers a stringent metabolic response in promastigotes and loss of virulence in amastigotes of Leishmania mexicana

Transketolase (TKT) is part of the non-oxidative branch of the pentose phosphate pathway (PPP). Here we describe the impact of removing this enzyme from the pathogenic protozoan Leishmania mexicana. Whereas the deletion had no obvious effect on cultured promastigote forms of the parasite, the Δtkt cells were not infective to mice. Δtkt promastigotes were more susceptible to oxidative stress and various leishmanicidal drugs than wild-type, and metabolomics analysis revealed profound changes to metabolism in these cells. In addition to changes consistent with those directly related to the role of TKT in the PPP, central carbon metabolism was substantially decreased, the cells consumed significantly less glucose, flux through glycolysis diminished, and production of the main end products of metabolism was decreased. Only minor changes in RNA abundance from genes encoding enzymes in central carbon metabolism, however, were detected although fructose-1,6-bisphosphate aldolase activity was decreased two-fold in the knock-out cell line. We also showed that the dual localisation of TKT between cytosol and glycosomes is determined by the C-terminus of the enzyme and by engineering different variants of the enzyme we could alter its sub-cellular localisation. However, no effect on the overall flux of glucose was noted irrespective of whether the enzyme was found uniquely in either compartment, or in both

Untargeted Metabolomics Reveals a Lack Of Synergy between Nifurtimox and Eflornithine against Trypanosoma brucei

A non-targeted metabolomics-based approach is presented that enables the study of pathways in response to drug action with the aim of defining the mode of action of trypanocides. Eflornithine, a polyamine pathway inhibitor, and nifurtimox, whose mode of action involves its metabolic activation, are currently used in combination as first line treatment against stage 2, CNS-involved, human African trypanosomiasis (HAT). Drug action was assessed using an LC-MS based non-targeted metabolomics approach. Eflornithine revealed the expected changes to the polyamine pathway as well as several unexpected changes that point to pathways and metabolites not previously described in bloodstream form trypanosomes, including a lack of arginase activity and N-acetylated ornithine and putrescine. Nifurtimox was shown to be converted to a trinitrile metabolite indicative of metabolic activation, as well as inducing changes in levels of metabolites involved in carbohydrate and nucleotide metabolism. However, eflornithine and nifurtimox failed to synergise anti-trypanosomal activity in vitro, and the metabolomic changes associated with the combination are the sum of those found in each monotherapy with no indication of additional effects. The study reveals how untargeted metabolomics can yield rapid information on drug targets that could be adapted to any pharmacological situation