Targeted lipidomic profiling of West Nile virus infection unveils alterations of sphingolipid metabolism in a mouse model and human patients

West Nile virus (WNV) is a neurotropic flavivirus transmitted by the bites of infected mosquitoes. Severe forms of West Nile disease (WND) can curse with meningitis, encephalitis, or acute flaccid paralysis. A better understanding of the physiopathology associated with disease progression is mandatory to find biomarkers and effective therapies. In this scenario, blood derivatives (plasma and serum) constitute the more commonly used biofluids due to their ease of collection and high value for diagnostic purposes. Therefore, the potential impact of this virus in the circulating lipidome was addressed by combining the analysis of samples from experimentally infected mice and naturally WND patients. Our results unveil dynamic alterations in the lipidome that define specific metabolic fingerprints of different infection stages. Concomitant with neuroinvasion in mice, the lipid landscape was dominated by a metabolic reprogramming that resulted in significant elevations of circulating sphingolipids (ceramides, dihydroceramides and dihydrosphingomyelins), along with some phosphatidylethanolamines and triacylglycerols. Remarkably, patients suffering from WND also displayed an elevation of ceramides, dihydroceramides, and lactosylceramides in their sera. The dysregulation of sphingolipid metabolism by WNV may provide new therapeutic opportunities and supports the potential of certain lipids as novel peripheral biomarkers of WND progression

Pharmacological Elevation of Cellular Dihydrosphingomyelin Provides a Novel Antiviral Strategy against West Nile Virus Infection

The flavivirus life cycle is strictly dependent on cellular lipid metabolism. Polyphenols like gallic acid and its derivatives are promising lead compounds for new therapeutic agents as they can exert multiple pharmacological activities, including the alteration of lipid metabolism. The evaluation of our collection of polyphenols against West Nile virus (WNV), a representative medically relevant flavivirus, led to the identification of N,N'-(dodecane-1,12-diyl)bis(3,4,5-trihydroxybenzamide) and its 2,3,4-trihydroxybenzamide regioisomer as selective antivirals with low cytotoxicity and high antiviral activity (half-maximal effective concentrations [EC50s] of 2.2 and 0.24 μM, respectively, in Vero cells; EC50s of 2.2 and 1.9 μM, respectively, in SH-SY5Y cells). These polyphenols also inhibited the multiplication of other flaviviruses, namely, Usutu, dengue, and Zika viruses, exhibiting lower antiviral or negligible antiviral activity against other RNA viruses. The mechanism underlying their antiviral activity against WNV involved the alteration of sphingolipid metabolism. These compounds inhibited ceramide desaturase (Des1), promoting the accumulation of dihydrosphingomyelin (dhSM), a minor component of cellular sphingolipids with important roles in membrane properties. The addition of exogenous dhSM or Des1 blockage by using the reference inhibitor GT-11 {N-[(1R,2S)-2-hydroxy-1-hydroxymethyl-2-(2-tridecyl-1-cyclopropenyl)ethyl]octanamide} confirmed the involvement of this pathway in WNV infection. These results unveil the potential of novel antiviral strategies based on the modulation of the cellular levels of dhSM and Des1 activity for the control of flavivirus infection.We thank Theodore C. Pierson (National Institutes of Health, USA) for the subgenomic replicon of WNV. This work was supported by the Spanish Ministry of Science and Innovation AEI/10.13039/501100011033 under grants PID2019-105117RR-C21 (to M.A.M.-A.), PID2019-105117RR-C22 (to M.-J.P.-P.), and PID2020-119195RJ-I00 (to N.J.d.O.) and by the AECSIC under grant PIE-201980E100 (to M.-J.P.-P. and A.S.-F.). This research work was also funded by the European Commission-NextGenerationEU (regulation EU 2020/2094) through CSIC’s Global Health Platform (PTI Salud Global). P.M.-C. was supported by an FPI fellowship (PRE2020-093374) from AEI/10.13039/501100011033. The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.Peer reviewe

Glycolytic shift during West Nile virus infection provides new therapeutic opportunities

Background Viral rewiring of host bioenergetics and immunometabolism may provide novel targets for therapeu‑ tic interventions against viral infections. Here, we have explored the effect on bioenergetics during the infection with the mosquito‑borne flavivirus West Nile virus (WNV), a medically relevant neurotropic pathogen causing out‑ breaks of meningitis and encephalitis worldwide. Results A systematic literature search and meta‑analysis pointed to a misbalance of glucose homeostasis in the cen‑ tral nervous system of WNV patients. Real‑time bioenergetic analyses confirmed upregulation of aerobic glycolysis and a reduction of mitochondrial oxidative phosphorylation during viral replication in cultured cells. Transcriptom‑ ics analyses in neural tissues from experimentally infected mice unveiled a glycolytic shift including the upregula‑ tion of hexokinases 2 and 3 (Hk2 and Hk3) and pyruvate dehydrogenase kinase 4 (Pdk4). Treatment of infected mice with the Hk inhibitor, 2‑deoxy‑D‑glucose, or the Pdk4 inhibitor, dichloroacetate, alleviated WNV‑induced neuroinflammation. Conclusions These results highlight the importance of host energetic metabolism and specifically glycolysis in WNV infection in vivo. This study provides proof of concept for the druggability of the glycolytic pathway for the future development of therapies to combat WNV pathology.This study was supported by the Spanish Ministry of Science and Innovation AEI/10.13039/501100011033 under Grants PID2019‑105117RR‑C21 (to MAMA) and PID2019‑105117RR‑C22 (to MJPP), PID2020‑119195RJ‑I00 (to NJO), by Synergistic Projects Community of Madrid under grant NUTRISION‑CM/Y2020/ BIO‑6350 (to ARM), and by the European Commission—NextGenerationEU through CSIC’s Global Health Platform (PTI Salud Global). PMC was sup‑ ported by an FPI fellowship from AEI/10.13039/501100011033 under Grant PRE2020‑093374. The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publicationPeer reviewe

AG5 is a potent non-steroidal anti-inflammatory and immune regulator that preserves innate immunity [Oral]

III Jornadas Científicas PTI+ Salud Global, 20-22 noviembre 2023, Madrid[Background] An archetypal anti-inflammatory compound against cytokine storm would inhibit it without suppressing the innate immune response. [Methods] AG5, an anti-inflammatory compound, has been developed as synthetic derivative of andrographolide, which is highly absorbable and presents low toxicity. [Result]. We found that the mechanism of action of AG5 is through the inhibition of caspase-1. interestingly, we show with in vitro generated human monocyte derived dendritic cells that AG5 preserves innate immune response. AG5 minimizes inflammatory response in a mouse model of lipopolysaccharide (LPS)-induced lung injury and exhibits in vivo anti-inflammatory efficacy in the SARS-CoV-2-infected mouse model. Furthermore, AG5 showed interesting antiviral activity against SARS-CoV-2 in humanized mice. [Conclusions] AG5 opens up a new class of anti-inflammatories, since contrary to NSAIDs, AG5 is able to inhibit the cytokine storm, like dexamethasone, but, unlike corticosteroids, preserves adequately the innate immunity. This is critical at the early stages of any naïve infection, but particularly in SARS-CoV-2 infectionsN

AG5 is a potent non-steroidal anti-inflammatory and immune regulator that preserves innate immunity

12 pages, 5 figures.-- This is an open access article under the CC BY-NC-ND licenseAn archetypal anti-inflammatory compound against cytokine storm would inhibit it without suppressing the innate immune response. AG5, an anti-inflammatory compound, has been developed as synthetic derivative of andrographolide, which is highly absorbable and presents low toxicity. We found that the mechanism of action of AG5 is through the inhibition of caspase-1. Interestingly, we show with in vitro generated human monocyte derived dendritic cells that AG5 preserves innate immune response. AG5 minimizes inflammatory response in a mouse model of lipopolysaccharide (LPS)-induced lung injury and exhibits in vivo anti-inflammatory efficacy in the SARS-CoV-2-infected mouse model. AG5 opens up a new class of anti-inflammatories, since contrary to NSAIDs, AG5 is able to inhibit the cytokine storm, like dexamethasone, but, unlike corticosteroids, preserves adequately the innate immunity. This is critical at the early stages of any naïve infection, but particularly in SARS-CoV-2 infections. Furthermore, AG5 showed interesting antiviral activity against SARS-CoV-2 in humanized miceThis work has been supported by NextGenerationEU Recovery and Resilience Facility (RRF) through the PTI+ Global Health Platform of Spanish National Research Council, grants SGL2103023 (PBA), SGL2103053 (MMA) and SGL2103015 (MM); by Spanish National Research Council through the program “Ayudas extraodinarias a proyectos de investigacion en el marco de las medidas urgentes extraodinarias para hacer frente al impacto económico y social del COVID-19”, grants CSIC-COV19-093 (PBA) and CSIC-COV19-117 (MM); by Generalitat Valenciana through the program “Ayudas urgentes para proyectos de investigación, desarrollo tecnológico e innovación (I+D+i) por la COVID-19”, grant GVA-COVID19/2021/059 (PBA); by the Conference of Rectors of the Spanish Universities, Spanish National Research Council and Banco Santander through the FONDO SUPERA COVID-19, grant CAPriCORn (JSM, JMB); by Severo Ochoa center of excellence program (grant CEX2021-001230-S) (PBA)Peer reviewe

Rabies in Spain. A Peculiarity in Eurasia

Spain accounts for some natural and political features which have influenced the situation of rabies, making it peculiar in the context of Europe. Dog rabies was eradicated from Spain in 1965 and the European vulpine epizootic was stopped by the French Oral Rabies Vaccination programs (ORVs) and it never crossed the Pyrenees this fact implying the first peculiarity. The possibility of the movement of infected animals from endemic territories in North Africa into the Spanish territories of Ceuta and Melilla by land, causing imported cases regularly is another peculiarity of rabies in Spain. Indeed, the existence of an intense maritime traffic from these endemic territories into the Spanish mainland is another peculiar feature which constitutes a permanent high risk for rabies introduction in Spain. European bat lyssavirus 1 is the most frequently lyssavirus found infecting bats in Spain, as occurs in most European countries. Eptesicus serotinus accounts for more than the 95% of infected bats declared in Europe and are considered the natural reservoir of EBLV-1. However, in the southern half of the Iberian Peninsula, E. serotinus is replaced by the sibling specie Eptesicus isabellinus, which has been found frequently infected by EBLV-1. E. isabellinus is present also in the other side of the Gibraltar Strait along a Mediterranean strip in Northern Africa. This constitutes another peculiarity of rabies in Spain. Finally, a new putative lyssavirus called Lleida Bat Lyssavirus (LLEBV) has been reported from a Miniopterus scheibersii bat from Catalonia. It still remains as another peculiarity of rabies in Spain.Patricia Mingo-Casas was part-funded by a Residencia de Estudiantes (CSIC) Postgraduate Fellowship from Spanish Ministry of Economy and Competitiveness (MINECO). This project was financially supported by the Spanish Ministry of Economy and Competitiveness (MINECO) project MPY 1321/14 and Virginia Sandonís was supported by a contract of the internal research program of the Instituto de Salud Carlos III.S

Dysregulation in host sphingolipid metabolism caused by West Nile virus infection as a potential disease progression biomarker and new therapeutic target

West Nile virus (WNV) is a neurotropic flavivirus transmitted by the bites of infected Culex sp mosquitoes and it is widely distributed across Europe. Severe forms of West Nile disease (WND) can cause meningitis, encephalitis or even death. In 2020, WNV caused an outbreak in southwest Spain that ended up with 70 human infections and 7 deaths. There are no vaccines or specific drugs to prevent and treat the symptoms of WND in humans, so the search for novel targets for therapeutic interventions is mandatory. Lipid metabolism is gaining interest as a potential target to combat flavivirus using a host-targeted approach.Peer reviewe

Lipid signatures of West Nile virus infection unveil alterations of sphingolipid metabolism providing novel biomarkers

AbstractWest Nile virus (WNV) is a neurotropic flavivirus transmitted by the bites of infected mosquitoes. Severe forms of West Nile disease (WND) can curse with meningitis, encephalitis or acute flaccid paralysis. A better understanding of the physiopathology associated with disease progression is mandatory to find biomarkers and effective therapies. In this scenario, blood derivatives (plasma and serum) constitute the more commonly used biofluids due to its ease of collection and high value for diagnostic purposes. Therefore, the potential impact of this virus in the circulating lipidome was addressed combining the analysis of samples from experimentally infected mice and naturally WND patients. Our results unveil dynamic alterations in the lipidome that define specific metabolic fingerprints of different infection stages. Concomitant with neuroinvasion in mice, the lipid landscape was dominated by a metabolic reprograming that resulted in significant elevations of circulating sphingolipids (ceramides, dihydroceramides and dihydrosphingomyelins), phosphatidylethanolamines and triacylglycerols. Remarkably, patients suffering from WND also displayed an elevation of ceramides, dihydroceramides, lactosylceramides and monoacylglycerols in their sera. The dysregulation of sphingolipid metabolism by WNV may provide new therapeutic opportunities and supports the potential of certain lipids as novel peripheral biomarkers of WND progression

First cases of European bat lyssavirus type 1 in Iberian serotine bats: Implications for the molecular epidemiology of bat rabies in Europe

<div><p>Previous studies have shown that EBLV-1 strains exclusively hosted by <i>Eptesicus isabellinus</i> bats in the Iberian Peninsula cluster in a specific monophyletic group that is related to the EBLV-1b lineage found in the rest of Europe. More recently, enhanced passive surveillance has allowed the detection of the first EBLV-1 strains associated to <i>Eptesicus serotinus</i> south of the Pyrenees. The aim of this study is the reconstruction of the EBLV-1 phylogeny and phylodynamics in the Iberian Peninsula in the context of the European continent. We have sequenced 23 EBLV-1 strains detected on nine <i>E</i>. <i>serotinus</i> and 14 <i>E</i>. <i>isabellinus</i>. Phylogenetic analyses were performed on the first 400-bp-5’ fragment of the Nucleoprotein (N) gene together with other 162 sequences from Europe. Besides, fragments of the variable region of the phosphoprotein (P) gene and the glycoprotein-polymerase (G-L) intergenic region were studied on Spanish samples. Phylogenies show that two of the new EBLV-1a strains from Iberian <i>E</i>. <i>serotinus</i> clustered together with French strains from the North of the Pyrenees, suggesting a recent expansion southwards of this subtype. The remaining seven Iberian strains from <i>E</i>. <i>serotinus</i> grouped, instead, within the cluster linked, so far, to <i>E</i>. <i>isabellinus</i>, indicating that spatial distribution prevails over species specificity in explaining rabies distribution and supporting interspecific transmission. The structure found within the Iberian Peninsula for EBLV-1b is in concordance with that described previously for <i>E</i>. <i>isabellinus</i>. Finally, we have found that the current EBLV-1 European strains could have emerged only 175 years ago according to our evolutionary dynamics analyses.</p></div