Targeted metabolomics identifies accurate CSF metabolite biomarkers for the differentiation between COVID-19 with neurological involvement and CNS infections with neurotropic viral pathogens.

BACKGROUND: COVID-19 is primarily considered a respiratory tract infection, but it can also affect the central nervous system (CNS), which can result in long-term sequelae. In contrast to CNS infections by classic neurotropic viruses, SARS-CoV-2 is usually not detected in cerebrospinal fluid (CSF) from patients with COVID-19 with neurological involvement (neuro-COVID), suggesting fundamental differences in pathogenesis. METHODS: To assess differences in CNS metabolism in neuro-COVID compared to CNS infections with classic neurotropic viruses, we applied a targeted metabolomic analysis of 630 metabolites to CSF from patients with (i) COVID-19 with neurological involvement [n = 16, comprising acute (n = 13) and post-COVID-19 (n = 3)], (ii) viral meningitis, encephalitis, or myelitis (n = 10) due to herpes simplex virus (n = 2), varicella zoster virus (n = 6), enterovirus (n = 1) and tick-borne encephalitis virus (n = 1), and (iii) aseptic neuroinflammation (meningitis, encephalitis, or myelitis) of unknown etiology (n = 21) as additional disease controls. RESULTS: Standard CSF parameters indicated absent or low neuroinflammation in neuro-COVID. Indeed, CSF cell count was low in neuro-COVID (median 1 cell/µL, range 0-12) and discriminated it accurately from viral CNS infections (AUC = 0.99) and aseptic neuroinflammation (AUC = 0.98). 32 CSF metabolites passed quality assessment and were included in the analysis. Concentrations of differentially abundant (fold change ≥|1.5|, FDR ≤ 0.05) metabolites were both higher (9 and 5 metabolites) and lower (2 metabolites) in neuro-COVID than in the other two groups. Concentrations of citrulline, ceramide (d18:1/18:0), and methionine were most significantly elevated in neuro-COVID. Remarkably, triglyceride TG(20:1_32:3) was much lower (mean fold change = 0.09 and 0.11) in neuro-COVID than in all viral CNS infections and most aseptic neuroinflammation samples, identifying it as highly accurate biomarker with AUC = 1 and 0.93, respectively. Across all samples, TG(20:1_32:3) concentration correlated only moderately with CSF cell count (ρ = 0.65), protein concentration (ρ = 0.64), and Q-albumin (ρ = 0.48), suggesting that its low levels in neuro-COVID CSF are only partially explained by less pronounced neuroinflammation. CONCLUSIONS: The results suggest that CNS metabolite responses in neuro-COVID differ fundamentally from viral CNS infections and aseptic neuroinflammation and may be used to discover accurate diagnostic biomarkers in CSF and to gain insights into differences in pathophysiology between neuro-COVID, viral CNS infections and aseptic neuroinflammation.

Modulation of the Serum Metabolome by the Short-Chain Fatty Acid Propionate: Potential Implications for Its Cholesterol-Lowering Effect.

(1) Background: Dyslipidemia represents a major risk factor for atherosclerosis-driven cardiovascular disease. Emerging evidence suggests a close relationship between cholesterol metabolism and gut microbiota. Recently, we demonstrated that the short-chain fatty acid (SCFA) propionate (PA) reduces serum cholesterol levels through an immunomodulatory mechanism. Here, we investigated the effects of oral PA supplementation on the human serum metabolome and analyzed changes in the serum metabolome in relation to the cholesterol-lowering properties of PA. (2) Methods: The serum metabolome of patients supplemented with either placebo or propionate orally for 8 weeks was assessed using a combination of flow injection analysis-tandem (FIA-MS/MS) as well as liquid chromatography (LC-MS/MS) and mass spectrometry using a targeted metabolomics kit (MxP®Quant 500 kit: BIOCRATES Life Sciences AG, Innsbruck, Austria). A total of 431 metabolites were employed for further investigation in this study. (3) Results: We observed a significant increase in distinct bile acids (GCDCA: fold change = 1.41, DCA: fold change = 1.39, GUDCA: fold change = 1.51) following PA supplementation over the study period, with the secondary bile acid DCA displaying a significant negative correlation with the serum cholesterol levels. (4) Conclusions: Oral supplementation with PA modulates the serum metabolome with a particular impact on the circulatory bile acid profile. Since cholesterol and bile acid metabolism are interconnected, the elevation of the secondary bile acid DCA may contribute to the cholesterol-lowering effect of PA.

Effects of a biotechnologically produced Pleurotus sapidus mycelium on gut microbiome, liver transcriptome and plasma metabolome of broilers.

Submerged cultivation using low-value agro-industrial side streams allows large-scale and efficient production of fungal mycelia, which has a high nutritional value. As the dietary properties of fungal mycelia in poultry are largely unknown, the present study aimed to investigate the effect of feeding a Pleurotus sapidus (PSA) mycelium as a feed supplement on growth performance, composition of the cecal microbiota and several physiological traits including gut integrity, nutrient digestibility, liver lipids, liver transcriptome and plasma metabolome in broilers. 72 males, 1-day-old Cobb 500 broilers were randomly assigned to 3 different groups and fed 3 different adequate diets containing either 0% (PSA-0), 2.5% (PSA-2.5) and 5% (PSA-5.0) P. sapidus mycelium in a 3-phase feeding system for 35 d. Each group consisted of 6 cages (replicates) with 4 broilers/cage. Body weight gain, feed intake and feed:gain ratio and apparent ileal digestibility of crude protein, ether extract and amino acids were not different between groups. Metagenomic analysis of the cecal microbiota revealed no differences between groups, except that one α-diversity metric (Shannon index) and the abundance of 2 low-abundance bacterial taxa (Clostridia UCG 014, Eubacteriales) differed between groups (P < 0.05). Concentrations of total and individual short-chain fatty acids in the cecal digesta and concentrations of plasma lipopolysaccharide and mRNA levels of proinflammatory genes, tight-junction proteins, and mucins in the cecum mucosa did not differ between groups. None of the plasma metabolites analyzed using targeted-metabolomics differed across the groups. Hepatic transcript profiling revealed a total of 144 transcripts to be differentially expressed between group PSA-5.0 and group PSA-0 but none of these genes was regulated greater 2-fold. Considering either the lack of effects or the very weak effects of feeding the P. sapidus mycelium in the broilers it can be concluded that inclusion of a sustainably produced fungal mycelium in broiler diets at the expense of other feed components has no negative consequences on broilers´ performance and metabolism.

Effect of replacing soybean meal with Hermetia illucens meal on cecal microbiota, liver transcriptome, and plasma metabolome of broilers.

Despite the existence of a number of studies investigating the effect of insect meal on the growth performance of broilers, knowledge about the metabolic effects of insect meal in broilers is still scarce. Thus, the present study investigated the effect of partial replacement of soybean meal with Hermetia illucens (HI) larvae meal on the liver transcriptome, the plasma metabolome, and the cecal microbiota in broilers. For the study, 72 male one-day-old Cobb 500 broilers were divided into three groups and fed 3 different diets with either 0% (HI0), 7.5% (HI7.5), or 15% (HI15) defatted HI meal for 35 d. Each group consisted of 6 cages (replicates) with 4 broilers/cage. While body weight (BW) gain, feed intake, and feed:gain ratio did not differ between groups, breast muscle weight, carcass yield, and apparent ileal digestibility (AID) of 5 amino acids were higher in group HI15 than in group HI0 (P < 0.05). Indicators of α-diversity (Chao1 and Observed) in the cecal digesta were higher in groups HI15 and HI7.5 than in group HI0 (P < 0.05). The abundance of 5 families and 18 genera, all of which belonged to the Firmicutes phylum, in the cecal digesta differed among groups (P < 0.05). Concentrations of butyric acid, valeric acid, and isobutyric acid in the cecal digesta were lower in group HI15 than in the other 2 groups (P < 0.05), whereas those of total and other short-chain fatty acids were not different between groups. Liver transcriptomics revealed a total of 70 and 61 differentially expressed transcripts between groups HI15 vs. HI0 and between groups HI7.5 vs. HI0, respectively, (P < 0.05). Targeted metabolomics identified 138 metabolites, most of which were triglyceride species, being different between the 3 groups (FDR < 0.05). According to this study, dietary inclusion of HI larvae meal has no detrimental impact but increases breast muscle weight and carcass weight in broilers suggesting that HI larvae meal can be recommended as a sustainable alternative protein source for broilers.

Tryptophan degradation as a systems phenomenon in inflammation - an analysis across 13 chronic inflammatory diseases.

BACKGROUND: Chronic inflammatory diseases (CIDs) are systems disorders that affect diverse organs including the intestine, joints and skin. The essential amino acid tryptophan (Trp) can be broken down to various bioactive derivatives important for immune regulation. Increased Trp catabolism has been observed in some CIDs, so we aimed to characterise the specificity and extent of Trp degradation as a systems phenomenon across CIDs. METHODS: We used high performance liquid chromatography and targeted mass spectrometry to assess the serum and stool levels of Trp and Trp derivatives. Our retrospective study incorporates both cross-sectional and longitudinal components, as we have included a healthy population as a reference and there are also multiple observations per patient over time. FINDINGS: We found reduced serum Trp levels across the majority of CIDs, and a prevailing negative relationship between Trp and systemic inflammatory marker C-reactive protein (CRP). Notably, serum Trp was low in several CIDs even in the absence of measurable systemic inflammation. Increases in the kynurenine-to-Trp ratio (Kyn:Trp) suggest that these changes result from increased degradation along the kynurenine pathway. INTERPRETATION: Increases in Kyn:Trp indicate the kynurenine pathway as a major route for CID-related Trp metabolism disruption and the specificity of the network changes indicates excessive Trp degradation relative to other proteogenic amino acids. Our results suggest that increased Trp catabolism is a common metabolic occurrence in CIDs that may directly affect systemic immunity. FUNDING: This work was supported by the DFG Cluster of Excellence 2167 "Precision medicine in chronic inflammation" (KA, SSchr, PR, BH, SWa), the BMBF (e:Med Juniorverbund "Try-IBD" 01ZX1915A and 01ZX2215, the e:Med Network iTREAT 01ZX2202A, and GUIDE-IBD 031L0188A), EKFS (2020_EKCS.11, KA), DFG RU5042 (PR, KA), and Innovative Medicines Initiative 2 Joint Undertakings ("Taxonomy, Treatments, Targets and Remission", 831434, "ImmUniverse", 853995, "BIOMAP", 821511).