Quantification of the 15 major human bile acids and their precursor 7α-hydroxy-4-cholesten-3-one in serum by liquid chromatography-tandem mass spectrometry

Bile acids are increasingly gaining attention since they were discovered to be activators of the transcription factor farnesoid X receptor (FXR) in addition to their well-established role in dietary lipid emulsification. Moreover, the differential activation potency of bile acids on FXR, which is due to structural variation of the ligands, generates the need for new analytical tools that are sensitive and specific enough to quantify the individual species of this complex class of compounds. Because bile acids undergo enterohepatic circulation, the additional assessment of a bile acid precursor as a marker for bile acid biosynthesis is used to differentiate between newly synthesised bile acids and bile acids reabsorbed from the intestine. This paper describes two new methods using liquid chromatography-tandem mass spectrometry (LC-MS/MS) for the quantification of the major unconjugated bile acids in human serum (cholic acid, chenodeoxycholic acid, deoxycholic acid, lithocholic acid and ursodeoxycholic acid) with their glycine- and taurine-conjugates as well as their precursor 7α-hydroxy-4-cholesten-3-one (C4). Intra- and inter-day variation was less than 12% and accuracy was between 84% and 102% for all analytes. Extraction recovery was between 78% and 100% for the bile acids whereas it was 62% for C4 and limit of quantification values ranged from 2nmol/l to 50nmol/l for all compounds. These two methods have the practical advantage of requiring low sample volume (100μl serum for each method) and identical eluents, stationary phase as well as ionisation technique, so that they can be used in a combined way. Moreover, they provide information on the composition of the bile acid pool on one hand and on the relative amount of newly synthesised bile acids on the other, which taken together, gives new insights in the investigation of bile acid metabolism

Pest categorisation of Citrus leprosis viruses

The EFSA Panel on Plant Health performed a pest categorisation of the Citrus leprosis viruses for theEU territory and identifiedfive distinct viruses,Citrus leprosis virus C(CiLV-C),Citrus leprosis virus C2(CiLV-C2),Hibiscus green spot virus 2(HGSV-2), the Citrus strain ofOrchidfleck virus(OFV) andCitrusleprosis virus N sensu novo(CiLV-N) as causing this severe disease, most significantly in sweet orangeand mandarin. These viruses have in common that they do not cause systemic infections in their hostsand that they all are transmitted byBrevipalpusspp. mites (likely but not confirmed for HGSV-2). Mitesrepresent the most important means of virus spread, while plants for planting of Citrus are onlyconsidered of minor significance. These well characterised viruses occur in South and Central America.Leprosis is currently regulated in directive 2000/29 EC and, together with its associated viruses, hasnever been recorded in the EU. Allfive viruses have the potential to enter into, establish in and spreadwithin the EU territory, with plants for planting of non-regulated hosts, fruits of Citrus and hitch-hikingof viruliferous mites identified as the most significant pathways. Given the severity of the leprosisdisease, the introduction and spread of the various viruses would have negative consequences on theEU citrus industry, the magnitude of which is difficult to evaluate given the uncertainties affecting theBrevipalpusspp. vectors (identity, distribution, density, transmission specificity and efficiency). Overall,leprosis and itsfive associated viruses meet all the criteria evaluated by EFSA to qualify as Unionquarantine pests, but do not fulfil those of being present in the EU or of plants for planting being themain spread mechanism to qualify as Union regulated non-quarantine pests. The main uncertaintiesaffecting this categorisation concern theBrevipalpusspp. mite vectors

Pest categorisation of Tatter leaf virus

The EFSA Panelon Plant Health performed a pest categorisation of Citrus tatter leaf virus (CTLV) for the EU territory. This virus is the causal agent of tatter leaf and graft incompatibility in trifoliate orange (Poncirustrifoliata) and its hybrids. CTLV is now recognised as a synonym of Apple stem grooving virus (ASGV), the type Capillovirus species, for which efficient diagnostics are available. There are no known ASGV vectors. The virus is reported in citrus from many countries. In the EU, while ASGV is widely present on apple and pear, it has never been reported on citrus. Since the citrus plants for planting pathway is closed by existing legislation, the main pathway for entry is plants for planting of other host species. In the EU, the high prevalence of ASGV in non-citrus hosts, but its absence in citrus ones suggests that interspecific host transfers are rare. However, there are high uncertainties on the importance and specifics of such host change events. No limits to the establishment of ASGV are identified and spread is likely through the vegetative propagation and trade of infected hosts. Infection of sensitive citrus rootstocks leads to stunted growth and decline of the entire plant a few years after grafting. The rootstocks that are now widely used to prevent citrus tristeza decline are the most affected. Among the criteria evaluated by EFSA for an organism to qualify as a Union quarantine pest, ASGV does not meet the criterion of being absent from or under official control in the EU territory. ASGV satisfies all the criteria evaluated by EFSA to qualify as a Union regulated non-quarantine pest. The main uncertainties concern the possible unreported presence of ASGV in citrus in the EU, the existence and efficiency of interspecific host transfers and the existence of ASGV natural spread

Pest categorisation of naturally‐spreading psorosis

The EFSA Panelon Plant Health performed a pest categorisation of naturally-spreading psorosis of citrus for the European Union. Naturally-spreading psorosis is poorly defined, because the status of both the disease and its causal agent(s) is uncertain. However, Citrus psorosis virus (CPsV) is a well- characterised Ophiovirus that is systematically associated with the psorosis disease and therefore considered to be its causal agent. Efficient diagnostics are available for CPsV. It is present in at least three EU MS. Naturally-spreading psorosis is currently regulated by Directive 2000/29/EC, while CPsV is not explicitly mentioned in this Directive. CPsV has the potential to enter, establish and spread in the EU territory. However, the main pathway for entry is closed by the existing legislation so that entry is only possible through minor alternative pathways. Plants for planting are the major means of spread while there are uncertainties on the existence and efficiency of a natural spread mechanism. CPsV introduction and spread in the EU would have negative consequences on the EU citrus industry. Of the criteria evaluated by EFSA to qualify as a Union quarantine pest or as a Union regulated non-quarantine pest (RNQP), Naturally-spreading psorosis does not meet the criterion of being a well characterised pest or disease. As it is not explicitly mentioned in the legislation, it is unclear whether CPsV meets the criterion of being currently regulated or under official control. It meets, however, all the RNQP criteria. The key uncertainties of this categorisation concern: (1) the causal role of CPsV in the psorosis disease as well as elements of its biology and epidemiology, (2) the exact nature of the Naturally-spreading psorosis syndrome and the identity of its causal agent and, consequently, (3) whether CPsV should be considered as being covered by the current legislation

Pest categorisation of ‘Blight and blight-like’ diseases of citrus

The EFSA Panel on Plant Health performed a pest categorisation of ‘ Blight and blight-like ’ for the EU territory. Blight is a major disease of citrus. Similar ‘ blight-like ’ diseases are also known (e.g. declinio, declinamiento) and are addressed simultaneously with Blight in the present categorisation. The causal agent(s) remain(s) unknown and the potential role of a recently identi fi ed citrus endogenous pararetrovirus (Citrus Blight-associated pararetrovirus, CBaPRV) remains to be established. Transmissibility and ability to produce consistent (although poorly speci fi c) symptoms have been demonstrated and a combination of indirect approaches is used, with limits, for diagnosis. There are large uncertainties on the biology of the causal agent(s) and on the epidemiology of the disease, including the transmission mechanism(s) responsible for the observed fi eld spread. Blight has been reported from North, Central and South America, Africa and Oceania but is not known to occur in the EU. It is listed in Annex IIA of Directive 2000/29EC. It has the potential to enter, establish and spread in the EU territory. The main entry pathway (citrus plants for planting) is closed by existing legislation and entry is only possible on minor pathways (such as illegal import). Blight is a severe disease and a negative impact is expected should it be introduced in the EU, but the magnitude of this negative impact is very dif fi cult to estimate. ‘ Blight and blight like ’ satis fi es all criteria evaluated by EFSA to qualify as a Union quarantine pest. It does not meet the criterion of being present in the EU to qualify as a Union regulated non-quarantine pest (RNQP). Since the identity of the causal agent(s) of the Blight and blight-like disease(s) and the existence and ef fi ciency of natural spread mechanism(s) remain unknown, large uncertainties affect all aspects of the present pest categorisation

The secretome of Agaricus bisporus: Temporal dynamics of plant polysaccharides and lignin degradation

Despite substantial lignocellulose conversion during mycelial growth, previous transcriptome and proteome studies have not yet revealed how secretomes from the edible mushroom Agaricus bisporus develop and whether they modify lignin models in vitro. To clarify these aspects, A. bisporus secretomes collected throughout a 15-day industrial substrate production and from axenic lab-cultures were subjected to proteomics, and tested on polysaccharides and lignin models. Secretomes (day 6-15) comprised A. bisporus endo-acting and substituent-removing glycoside hydrolases, whereas β-xylosidase and glucosidase activities gradually decreased. Laccases appeared from day 6 onwards. From day 10 onwards, many oxidoreductases were found, with numerous multicopper oxidases (MCO), aryl alcohol oxidases (AAO), glyoxal oxidases (GLOX), a manganese peroxidase (MnP), and unspecific peroxygenases (UPO). Secretomes modified dimeric lignin models, thereby catalyzing syringylglycerol-β-guaiacyl ether (SBG) cleavage, guaiacylglycerol-β-guaiacyl ether (GBG) polymerization, and non-phenolic veratrylglycerol-β-guaiacyl ether (VBG) oxidation. We explored A. bisporus secretomes and insights obtained can help to better understand biomass valorization

From 13C-lignin to 13C-mycelium: Agaricus bisporus uses polymeric lignin as a carbon source

Plant biomass conversion by saprotrophic fungi plays a pivotal role in terrestrial carbon (C) cycling. The general consensus is that fungi metabolize carbohydrates, while lignin is only degraded and mineralized to CO2. Recent research, however, demonstrated fungal conversion of 13C-monoaromatic compounds into proteinogenic amino acids. To unambiguously prove that polymeric lignin is not merely degraded, but also metabolized, carefully isolated 13C-labeled lignin served as substrate for Agaricus bisporus, the world's most consumed mushroom. The fungus formed a dense mycelial network, secreted lignin-active enzymes, depolymerized, and removed lignin. With a lignin carbon use efficiency of 0.14 (g/g) and fungal biomass enrichment in 13C, we demonstrate that A. bisporus assimilated and further metabolized lignin when offered as C-source. Amino acids were high in 13C-enrichment, while fungal-derived carbohydrates, fatty acids, and ergosterol showed traces of 13C. These results hint at lignin conversion via aromatic ring-cleaved intermediates to central metabolites, underlining lignin's metabolic value for fungi

Warmth and competence perceptions of key protagonists are associated with containment measures during the COVID-19 pandemic: Evidence from 35 countries

It is crucial to understand why people comply with measures to contain viruses and their effects during pandemics. We provide evidence from 35 countries (Ntotal = 12,553) from 6 continents during the COVID-19 pandemic (between 2021 and 2022) obtained via cross-sectional surveys that the social perception of key protagonists on two basic dimensions—warmth and competence—plays a crucial role in shaping pandemic-related behaviors. Firstly, when asked in an open question format, heads of state, physicians, and protest movements were universally identified as key protagonists across countries. Secondly, multiple-group confirmatory factor analyses revealed that warmth and competence perceptions of these and other protagonists differed significantly within and between countries. Thirdly, internal meta-analyses showed that warmth and competence perceptions of heads of state, physicians, and protest movements were associated with support and opposition intentions, containment and prevention behaviors, as well as vaccination uptake. Our results have important implications for designing effective interventions to motivate desirable health outcomes and coping with future health crises and other global challenges.publishedVersio