Crystal structures of Schistosoma mansoni histone deacetylase 8 reveal a novel binding site for allosteric inhibitors.

Parasitic diseases cause significant global morbidity and mortality particularly in the poorest regions of the world. Schistosomiasis, one of the most widespread neglected tropical diseases, affects more than 200 million people worldwide. Histone deacetylase (HDAC) inhibitors are prominent epigenetic drugs that are being investigated in the treatment of several diseases, including cancers and parasitic diseases. Schistosoma mansoni HDAC8 (SmHDAC8) is highly expressed in all life cycle stages of the parasite, and selective inhibition is required in order to avoid undesirable off-target effects in the host. Herein, by X-ray crystal structures of SmHDAC8-inhibitor complexes, biochemical and phenotypic studies, we found two schistosomicidal spiroindoline derivatives binding a novel site, next to Trp198, on the enzyme surface. We determined that by acting on this site, either by mutation of the Trp198 or by compound binding, a decrease in the activity of the enzyme is achieved. Remarkably, this allosteric site differs from the human counterpart; rather, it is conserved in all Schistosoma species, as well as Rhabidoptera and Trematoda classes, thus paving the way for the design of HDAC8-selective allosteric inhibitors with improved properties.

Comparative metabolic profiling by 1H-NMR spectroscopy analysis reveals the adaptation of S. mansoni from its host to in vitro culture conditions: a pilot study with ex vivo and GSH-supplemented medium-cultured parasites.

Schistosomiasis is a neglected tropical disease caused by parasitic flatworms (blood fluke) of the genus Schistosoma. Parasites acquire most nutrients for their development and sustainment within the definitive host either by ingestion into the gut or across the body surface. Over the years, the best conditions for long-term maintenance of parasites in vitro have been thoroughly established. In our hands, 1H-NMR spectroscopy represents a powerful tool to characterize the metabolic changes in S. mansoni in response to culturing condition perturbations. In order to compare the metabolic fingerprint of ex vivo and parasites cultured in vitro with or without the supplement of reduced glutathione, we conducted a pilot study applying the 1H-NMR spectroscopy-based metabolomics. We obtained new insight into specific metabolic pathways modulated under these different experimental conditions.

(+)-(R)- and (-)-(S)-Perhexiline maleate: Enantioselective synthesis and functional studies on Schistosoma mansoni larval and adult stages.

Schistosomiasis is a neglected tropical disease mainly affecting the poorest tropical and subtropical areas of the world with the impressive number of roughly 200 million infections per year. Schistosomes are blood trematode flukes of the genus Schistosoma causing symptoms in humans and animals. Organ morbidity is caused by the accumulation of parasite eggs and subsequent development of fibrosis. If left untreated, schistosomiasis can result in substantial morbidity and even mortality. Praziquantel (PZQ) is the most effective and widely used compound for the treatment of the disease, in prevention and control programs in the last 30 years. Unfortunately, it has no effect on juvenile immature schistosomes and cannot prevent reinfection or interfere with the schistosome life cycle; moreover drug-resistance represents a serious threat. The search for an alternative or complementary treatment is urgent and drug repurposing could accelerate a solution. The anti-anginal drug perhexiline maleate (PHX) has been previously shown to be effective on larval, juvenile, and adult stages of S. mansoni and to impact egg production in vitro. Since PHX is a racemic mixture of R-(+)- and S-(-)-enantiomers, we designed and realized a stereoselective synthesis of both PHX enantiomers and developed an analytical procedure for the direct quantification of the enantiomeric excess also suitable for semipreparative separation of PHX enantiomers. We next investigated the impact of each enantiomer on viability of newly transformed schistosomula (NTS) and worm pairs of S. mansoni as well as on egg production and vitellarium morphology by in vitro studies. Our results indicate that the R-(+)-PHX is mainly driving the anti-schistosomal activity but that also the S-(-)-PHX possesses a significant activity towards S. mansoni in vitro.

Investigating the Antiparasitic Potential of the Marine Sesquiterpene Avarone, Its Reduced form Avarol, and the Novel Semisynthetic Thiazinoquinone Analogue Thiazoavarone.

The chemical analysis of the sponge Dysidea avara afforded the known sesquiterpene quinone avarone, along with its reduced form avarol. To further explore the role of the thiazinoquinone scaffold as an antiplasmodial, antileishmanial and antischistosomal agent, we converted the quinone avarone into the thiazinoquinone derivative thiazoavarone. The semisynthetic compound, as well as the natural metabolites avarone and avarol, were pharmacologically investigated in order to assess their antiparasitic properties against sexual and asexual stages of Plasmodium falciparum, larval and adult developmental stages of Schistosoma mansoni (eggs included), and also against promastigotes and amastigotes of Leishmania infantum and Leishmania tropica. Furthermore, in depth computational studies including density functional theory (DFT) calculations were performed. A toxic semiquinone radical species which can be produced starting both from quinone- and hydroquinone-based compounds could mediate the anti-parasitic effects of the tested compounds.

Detection of selected trace elements in yogurt components.

The objective of this study was to determine the concentrations of Cu, Cd, Pb, Mn, Cr, Co, Ni, Zn, and Hg in the white and fruit parts of commercially available yogurts (n = 30) from Nitra markets (Slovak Republic). The results were correlated to determine their relationships. Three yogurt fruit flavors were chosen and tested, strawberry (n = 10), blueberry (n = 10), and cherry (n = 10). The elements were analyzed using atomic absorption spectrophotometry. Higher concentrations of toxic elements, such as Cd and Pb, were found in the fruit parts of the yogurt, and in some cases, the tolerable limit was exceeded. The white part of the yogurt was not contaminated by toxic elements. White yogurt is a good source of nutrients for humans, but the fruit part in yogurt requires detailed monitoring and improvements in the processing techniques.

A LC-MS-MS method to detect recombinant bovine somatotropin misuse in buffalos.

Recombinant bovine somatotropin (rbST) is a peptide hormone used to increase milk yield in cows and buffalos. In Europe, its use has been banned. However, rbST is sometimes illegally included in zootechnical practices for profit purposes, undermining the fair trade and the law prescriptions. For this reason, efficient and reliable analytical techniques are required to contrast rbST misuse. A few LC-MS-MS methods have been developed to detect, in cow serum, methyonil-rbST, one of the two main rbST forms available on the market. The other form, which is widespread, is identical to the most abundant variant of bovine somatotropin (bST) and differs from the buffalo somatotropin for one amino acid in the N-terminus. For this reason, it is technically possible to distinguish both rbST forms in serum of buffalos. In this work, we describe a novel LC-MS-MS-based method, capable to quantify, with a high sensitivity and selectivity, the methyonil-rbST and the other bST-identical recombinant form in buffalo serum, previously purified using a solid-phase extraction procedure. The method was internally validated and used to analyse 152 serum samples, collected from eight buffalos administered with rbST for a period of 3 months, according to conventional protocols. The obtained results confirmed the suitability of the method in the detection of illegal hormonal treatments. Graphical abstract ᅟ.

Levels of Essential and Trace Elements in Mozzarella Available on the Slovak Market with the Estimation of Consumer Exposure.

The aim of this study was to determinate the content of some elements in a specific dairy product, mozzarella, in a particular area of western Slovakia and to evaluate the estimation of the risk to the consumers based on the contribution to the provisional tolerable weekly intake. The consumption of mozzarella can contribute to the intake of important elements in the diet, such as calcium and magnesium, along with others. The contents of some toxic and trace elements were low and have not exceeded the permitted limit. In addition, the contribution to PTWI was found to be very low, which means that the consumption of mozzarella possesses no risk to humans. It is concluded that the data obtained in this study can help as a valuable addition to methodological and scientific material in the field of food safety of dairy products and their positive impact on human health.

Food safety culture in food companies: evaluation of the perception of food safety culture in three Tuscan food companies.

The Commission Regulation (EU) No. 2021/382 (European Commission, 2021), amending the Regulation (EC) No. 852/2004 (European Commission, 2004), introduced the obligation for companies to establish and maintain a food safety culture (FSC). The methodology to evaluate, implement, and enhance the level of FSC is up to the individual companies. This study aimed to investigate the perception of FSC among the employees of 3 Tuscan medium-sized enterprises in the food sector, producing cured meat (A), dairy products (B), and frozen fish products (C). The survey was conducted through the development and administration of a questionnaire based on a 5 points Likert scale, referring to different aspects of FSC, organized in 6 sections with 5-6 statements each and subjected to a percentage of employees between 76 and 85%, classified also by the length of service (≤3 and >3 years). For all the companies, the minimum median and mode value for scores obtained by the different sections was 4, and the minimum median and mode value for the single statement was 3 (A, B; except for a bimodal value 2-4) and 4 (C). The section awareness and perception of risk showed the highest mean scores in all companies. As for the length of service, senior employees gave lower scores than junior ones in all sections in B and 3 sections in C. Overall, the results of the questionnaires showed a good perception of FSC, even though it was possible to identify some partial weaknesses.

Activated retinal glia mediated axon regeneration in experimental glaucoma.

Glaucoma, a leading cause of blindness, is a neurodegenerative disease characterized by progressive loss of retinal ganglion cell axons in the optic nerve and their cell bodies in the retina. Reactive retinal glial changes have been observed in glaucoma but the role of such glial changes in the pathogenesis of the condition remains unclear. In the present study we found that retinal ganglion cells in an experimental animal model of glaucoma have an increased axon regenerative potential. Regeneration of adult rat retinal ganglion cell axons after optic nerve crush was significantly increased in vivo when combined with intraocular pressure-induced experimental glaucoma. This enhanced axon regeneration response was correlated with a significant increase in activation of glial fibrillary acidic protein+retinal glia. Using a dissociated retinal ganglion cell culture model we showed that reducing the number of activated retinal glia with a glial specific toxin, α-Aminoadipic acid, significantly reduced the growth potential of retinal ganglion cells from glaucomatous rat eyes, suggesting that activated retinal glia mediate, at least in part, the growth promoting effect. This was shown to be mediated by both membrane-bound and soluble glial-derived factors. Neurotrophin and ciliary neurotrophic/leukemia inhibitory factor blockers did not affect the regenerative potential, excluding these growth factors as principal mediators of the enhanced growth response occurring in glaucomatous retinal cultures. These observations are the first to reveal that retinal ganglion cells from glaucomatous rat eyes have an enhanced regenerative capacity. Furthermore, our results suggest that activated retinal glia mediate at least part of this response. Further work to understand and enhance the regeneration-promoting effect of activated retinal glia is required to determine if this approach could be useful as part of a therapeutic strategy to encourage optic nerve regeneration in glaucoma.

Gas-phase ion migration spectrum analysis of the volatile flavors of large yellow croaker oil after different storage periods.

The large yellow croaker, a species of fish found in the northwestern Pacific, is favored by consumers because of its prevalence in saltwater bodies, golden yellow abdomen, high calcium content, high protein, high fat content, and a flavor that originates from its lipids and volatile components. Volatile organic compounds significantly affect the aroma of food. In this work, electronic nose and headspace gas chromatography-ion mobility spectrometry were applied to analyze the flavor differences in fish oil durations. Through electronic nose system analysis, sensors W1C, W3S, W6S, and W2S directly affected fish oil flavor, and their flavor components were different. Gas chromatography-ion mobility spectrometry identified 26 volatile components (19 aldehydes, 3 ketones, 2 alcohols, 1 furan, and 1 olefin). (E,E)-2,4-hexadienal (D), (E,E)-2,4-hexadienal (M), 2,4-heptadienal (M), (E)-2-octenal, 2-propanone, 2-heptanone (M), 3-pentanone (D), and 1-octen-3-ol were the key flavor components of the fish oil. In conclusion, the combination of GC-IMS and PCA can identify the differences in flavor changes of large yellow croaker oil during 0-120 days storage. After 60 days storage, the types and signals of 2-propanone, 2-heptanone (M) components increase significantly. When 120 days storage, at this time, (E,E)-2,4-hexadienal (D), (E,E)-2,4-hexadienal (M), 2,4-heptadienal (M), (E)-2-octenal,(E)-2-octenal significantly. It has become the main flavor substance of fish oil. In summary, as the storage period increases, the components increase, and the oxidizing substances will increase, resulting in the deterioration of fish oil.

Effects of various logistics packaging on the quality and microbial variation of bigeye tuna (Thunnus obesus).

Bigeye tuna (Thunnus obesus) is an economically valuable ocean fish species. It is susceptible to contamination during storage and transportation. Having proper transportation packaging and stable temperature during transportation are critical to prevent quality deterioration. However, the influence of packaging on retaining freshness in transit remains unknown. Here, the impact of different transportation packaging on the quality and microbiological variation of bigeye tuna during the logistics process was investigated by measuring physical-chemical indexes and microbial diversity. It turned out that aluminum foil paper (AFP) group had minimum temperature fluctuation, exhibited preferable water retaining capacity and color protection effect. AFP packaging could efficiently prevent TVB-N increase and microbial growth. After 40 h, the TVB-N value was 21.28 mg/100 g and microbial total plate count was 3.53 lg CFU/g, which was within the acceptable range. Temperature fluctuations and packaging materials had a major effect on the microbial community structure of bigeye tuna. Chitinophagaceae, Acinetobacter, and Knoellia were dominant in the AFP group, while Pseudomonas, Acinetobacter, and Macrococcus were dominant in the expanded polystyrene foam (EPSF) and European logistics (EUL) groups. AFP packaging could effectively slow down the growth and reproduction of Pseudomonas, restraining the growth of microorganisms and preserve the quality of bigeye tuna. This study provides insights into understanding the effects of packaging material on maintaining quality during logistics transportation.

Somatotropic gene response to recombinant growth hormone treatment in buffalo leucocytes.

The use of recombinant bovine growth hormone (rbGH) to increase milk yield in cows is banned in some countries. In others, where it is authorised, it has triggered harsh debates on labelling of dairy products. If many studies have been performed on bovines, there is a lack of information on buffaloes, which are sometimes treated with rbGH and re-present an important economical resource for dairy products in some countries. Analytical methods with legal value for surveillance of rbGH treatments do not yet exist. Research on gene expression biomarkers is one of the most promising approaches to this purpose. For this reason, we treated five buffaloes for 10 weeks with a sustained-release formulation of rbGH and analysed the response of 20 somatotropic axis genes in leucocytes by real-time polymerase chain reaction. Overall changes in gene expression levels were of low magnitude and sometimes affected by the 'time' factor. Only the IGFBP-1 gene showed a significant under-expression (about two-fold; p <0.001) in treated animals. Taken together, these results give evidence that expression analysis of the somatotropic axis genes in leucocytes is little helpful for discrimination of rbGH-treated buffaloes, but do not exclude that another array of genes could provide useful patterns of variation.

Hormone variations in serum and milk of buffaloes (Bubalus bubalis) as potential indicators of treatment with recombinant bovine somatotropin.

Recombinant bovine somatotropin (rbST) is used to increase milk yield in cows, but it has been forbidden in some countries and in the EU. However, rbST misuse represents a concern in both bovine and buffalo dairy production. A number of studies on rbST treatment have been performed on bovines, but there are few data on buffaloes. In this study, we treated eight lactating buffaloes with biweekly injections of a slow-release formulation of rbST, for five cycles of administration, and analysed total ST and insulin-like growth factor 1 (IGF-1) variations in serum and IGF-1 in milk. The aim was to assess their power as potential indicators of rbST-treatment. Blood was collected on days 2, 5, 9 and 14 of each cycle, and milk on days 2, 9 and 14 of cycles 2 and 5. Results showed an extraordinary increase in ST levels on day 2 in treated animals, followed by a rapid decrease over the following days, while a significant increase in IGF-1 was observed both in serum and in milk throughout most of each cycle. These results suggest that serum ST levels are a good indicator of treatment. However, the rapid decrease after the peak limits the useful period of sample collection.

Edible Insects and Sustainable Development Goals.

The insect sector can become an important component of sustainable circular agriculture by closing nutrient and energy cycles, fostering food security, and minimising climate change and biodiversity loss, thereby contributing to SDGs. The high levels of the interaction of the insect sector with the SDGs is clearly illustrated inside the review, analysing all of the SDGs that can have direct and indirect effects on insects. Mapping the interactions between the SDGs goals and insect sector offers a starting point, from which it could be possible to define practical next steps for better insect policy.

Selecting reference genes in the white blood cells of buffalos treated with recombinant growth hormone.

In this study, we assessed the expression stability of eight genes (GADPH, ACTB, 18S, YWHAZ, SDHA, HMBS, SF3A1, and EEF1A) in the white blood cells of lactating buffalos and their possible use as reference genes for studies on growth hormone (GH)-treated animals. All of the genes showed acceptable stability according to the threshold values suggested by some of the software that was used to analyze them, although the differences between the most stable (SF3A1 and ACTB) and the least stable (18S) were considerable. GH treatment did not influence their expression levels.

Luminescence-Based, Low- and Medium-Throughput Assays for Drug Screening in Schistosoma mansoni Larval Stage.

Schistosomiasis is one of the major parasitic diseases with more than  200 million people infected worldwide every year. Praziquantel is the drug of choice against the schistosomiasis although the use of a single drug to treat such a large amount of infected people appears particularly worrisome. For this reason, the search of new schistosomicidal compounds is viewed as an urgent goal and a number of screening campaigns have been carried out in the past years. The larval stage of Schistosoma (schistosomula) has been widely used in order to identify new compounds against the parasite. Here we describe detailed practical procedures for a luminescence-based assay proven to be highly effective for the selection of schistosomicidal compounds on small and medium-high scale. The assay is based on the quantitation of the parasite ATP, a good indicator of metabolically active cells, as measure of schistosomula viability. This assay is fast and reproducible, and it is suitable either for manual or for semiautomated screenings.

Risks and critical issues related to the discovery on the market of unauthorized live alien species on the Italian territory: Chinese crab (Eriocheir sinensis).

Eriocheir sinensis, Chinese Crab or Chinese Mitten Crab is a catadromous species belonging to the Varunidae family, native to river and estuarine areas of North and South East China and Korea. At European level, E. sinensis is widespread in the main water basins of Central and Northern Europe and, since 2016, it has been included in the list of invasive species important for the European Union and subjected to confinement and eradication measures which include the prohibition of collection, transit and placing on the market of live specimens (Regulation (EC) N° 1143/2014). The Chinese Crab can represent a significant danger for the local ecosystem and for the native biota as well as contributing to the appearance of hydrogeological instability phenomena resulting from the intense excavation and erosion of the riverbanks. The first finding of 5 kg of live specimens of Eriocheir sinensis was recorded in the official control by the UFS (Functional Simple Unit) veterinary public health and food safety of the ASL Toscana Centro at an ethnic catering establishment. The specimens were subjected to seizure, photographed, identified morphologically, and subjected to euthanasia and destruction in accordance with the European requirements for welfare and management of animal by-products. From the sanitary point of view, the dangers associated with the consumption of this crab are mainly biological and chemical therefore, risk communication is fundamental, not only at the level of the competent authorities in the sector, but also for the food business operators.

Urban Consumer Trust and Food Certifications in China.

China has experienced frequent food safety incidents that have undermined consumer trust in the food supply chain. To overcome this problem, China requalified the legislative framework and adopted a comprehensive food certification system over the years. Here, we investigated the influences of food traceability and Chinese certifications (QS/SC-food quality safety market access/production system, hazard-free, green, and organic) on Chinese consumer trust of food safety for different types of products: fish, meat, milk, eggs, and rice. Data were collected through face-to-face surveys conducted in rural and urban Chinese areas. With a sample of 757 questionnaires, we ran a logit model. The results show consumers' uncertainty and skepticism of certifications guaranteeing food safety attributes, especially for animal-based products. We found that price is used as a cue of safety by Chinese consumers. Individuals with higher education seem less influenced by certifications and other cues included in the analysis. The findings demonstrate that Chinese policy makers should implement new strategies to enhance consumer food safety trust, and design policies by considering different categories (e.g., vegetables, meat, fish, etc.) of food.

Drug effects on metabolic profiles of Schistosoma mansoni adult male parasites detected by 1H-NMR spectroscopy.

Schistosomiasis is one of the most devastating neglected tropical parasitic diseases caused by trematodes of the genus Schistosoma. Praziquantel (PZQ) is today the only drug used in humans and animals for the treatment of schistosomiasis but unfortunately it is poorly effective on larval and juvenile stages of the parasite. Therefore, it is urgent the discovery of new drug targets and compounds. We have recently showed that the anti-anginal drug perhexiline maleate (PHX) is very active on multiple developmental stages of Schistosoma mansoni in vitro. It is well known that PHX impacts the lipid metabolism in mammals, but the final target on schistosomes still remains unknown. The aim of this study was to evaluate the ability of 1H nuclear magnetic resonance (NMR) spectroscopy in revealing metabolic perturbations due to PHX treatment of S. mansoni adult male worms. The effects of PHX were compared with the ones induced by vehicle and gambogic acid, in order to detect different metabolic profiles and specificity of the PHX action. Remarkably a list of metabolites associated to PHX-treatment was identified with enrichment in several connected metabolic pathways including also the Kennedy pathway mediating the glycerophospholipid metabolism. Our study represents the first 1H-NMR metabolomic approach to characterize the response of S. mansoni to drug treatment. The obtained "metabolic fingerprint" associated to PHX treatment could represent a strategy for displaying cellular metabolic changes for any given drug and to compare compounds targeting similar or distinct biochemical pathways.

Thiazinoquinones as New Promising Multistage Schistosomicidal Compounds Impacting Schistosoma mansoni and Egg Viability.

Schistosomiasis is the most significant neglected tropical parasitic disease caused by helminths in terms of morbidity and mortality caused by helminths. In this work, we present the antischistosomal activity against Schistosoma mansoni of a rationally selected small set of thiazinoquinone derivatives, some of which were previously found to be active against Plasmodium falciparum and others synthesized ad hoc. The effects on larvae, juvenile, and adult parasite viability as well as on egg production and development were investigated, resulting in the identification of new multistage antischistosomal hit compounds. The most promising compounds 6, 8, 13, and 14 with a LC50 value on schistosomula from ∼5 to ∼15 µM also induced complete death of juvenile (28 days old) and adult worm pairs (7 weeks old) and a detrimental effect on egg production and development in vitro. Structure-activity relationships (SARs) were analyzed by means of computational studies leading to the hypothesis of a redox-based mechanism of action with a one-electron reduction bioactivation step and the subsequent formation of a toxic semiquinone species, similarly to what was previously observed for the antiplasmodial activity. Our results also evidenced that the selective toxicity against mammalian cells or parasites as well as specific developmental stages of a parasite can be addressed by varying the nature of the introduced substituents.