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1. Climate change is causing shifts in the distribution of many species and populations inhabiting mountain tops are particularly vulnerable to these threats because they are constrained in altitudinal shifts. Apennines are a relatively narrow and low mountain chain located in Southern Europe, which hosts many isolated populations of mountain species. The butterfly Erebia pandrose was recorded for the last time in the Apennines in 1977, on the top of a single massif (Monti della Laga). 2. We confirmed the presence of a small, isolated population of E. pandrose in the Apennines, at a distance of more than 400¿km to any other known populations. Then, we examined the cytochrome c oxidase subunit 1 mitochondrial DNA marker of this species across the Palaearctic area and estimated the potential decline over the Alps and the Apennines due to future climatic changes. 3. The Apennine population represents an endemic lineage characterised by eight mutations over the 658¿bp analysed (1.2%). In the Alps and Apennines, this species has shifted uphill more than 3 m per year since the end of the 19th century and more than 22 m per year since 1995. Species distribution models suggested that these mountain populations will experience a generalised loss of climatic suitability, which, according to our projections, could lead to the extinction of the Apennine population in a few decades. 4. Erebia pandrose has the potential to become a flagship species for advertising the risk of losing unique fractions of genetic diversity for mountain species., funded by the project: ‘Ricerca e conservazione sui lepidotteri diurni di sei Parchi Nazionali dell'Appennino centro-settentrionale’. Support for this research was also provided by ‘La Caixa’ Foundation (ID 100010434) to M.M. (grant LCF/BQ/DR20/11790020), by the Academy of Finland to V.D. (Academy Research Fellow, decision no. 328895) and by project PID2019-107078GB-I00/AEI/10.13039/501100011033 to R.V., 1 Introduction 2 Materials and methods 2.1 Data collection 2.2 COI analyses 2.3 Species distribution modelling 2.4 Elevational shifts of E. pandrose records through time 3 Results 3.1 COI structure 3.2 Climate change effects 4 Discussion 4.1 The population of Monte Gorzano 4.2 What is the future for E. pandrose on the Apennines? 4.3 Erebia pandrose in the context of preserving Apennine butterflies 5 Conclusions Acknowledgements Conflict of interest

We here characterize the response to the extremely low-frequency (ELF) magnetic field (MF, 50 Hz, 1 mT) of SH-SY5Y human neuroblastoma cells, cultured in a three-dimensional (3D) Alvetex® scaffold compared to conventional two-dimensional (2D) monolayers. We proved that the growing phenotype of proliferating SH-SY5Y cells is not affected by the culturing conditions, as morphology, cell cycle distribution, proliferation/differentiation gene expression of 3D-cultures overlap what reported in 2D plates. In response to 72-h exposure to 50-Hz MF, we demonstrated that no proliferation change and apoptosis activation occur in both 2D and 3D cultures. Consistently, no modulation of Ki67, MYCN, CCDN1, and Nestin, of invasiveness and neo-angiogenesis-controlling genes (HIF-1α, VEGF, and PDGF) and of microRNA epigenetic signature (miR-21-5p, miR-222-3p and miR-133b) is driven by ELF exposure. Conversely, intracellular glutathione content and SOD1 expression are exclusively impaired in 3D-culture cells in response to the MF, whereas no change of such redox modulators is observed in SH-SY5Y cells if grown on 2D monolayers. Moreover, ELF-MF synergizes with the differentiating agents to stimulate neuroblastoma differentiation into a dopaminergic (DA) phenotype in the 3D-scaffold culture only, as growth arrest and induction of p21, TH, DAT, and GAP43 are reported in ELF-exposed SH-SY5Y cells exclusively if grown on 3D scaffolds. As overall, our findings prove that 3D culture is a more reliable experimental model for studying SH-SY5Y response to ELF-MF if compared to 2D conventional monolayer, and put the bases for promoting 3D systems in future studies addressing the interaction between electromagnetic fields and biological systems.

Background: The familial nonmedullary thyroid cancer (FNMTC) is suspected to be a Mendelian condition in up to 3–8% of thyroid cancers. The susceptibility chromosomal loci and genes of 95% of FNMTC cases remain to be characterized. The inheritance of FNMTC appears to be autosomal dominant with incomplete penetrance and variable expressivity. The finding of the causative gene of FNMTC and the identification of patients at risk that need genetic testing were our aim. Methods: We analyzed by whole-exome sequencing patients and non-affected relatives of five families with at least two family members affected by papillary thyroid cancer, selecting for new or extremely rare variants with predicted pathogenic value. Results: A family showed, in all three affected members, a new loss-of-function variant (frameshift deletion) in BROX gene at 1q41 that was absent from all internal and external databases. In a second family with three affected relatives, we found an additional new BROX variant. The smaller families presented no variants in BROX or in the other causative genes studied. Conclusions: BROX could be a new causative gene for FNMTC. Variants in BROX may result in the haploinsufficiency of a key gene involved in the morphogenesis of MVBs, in the endosomal sorting of cargo proteins, and in EGFR. Functional studies are needed to support this result. The thorough genomic analysis by NGS in all families with three or more affected members should become a routine approach to obtain a comprehensive genetic view and find confirmative second cases.

Due to the increasing number of studies reporting the detection of antimicrobial-resistant isolates of Listeria monocytogenes, we sought to determine the antimicrobial susceptibility of L. monocytogenes isolates collected in Italy and find potential correlations to their serotypes and multilocus sequence types (MLST). The antimicrobial susceptibility of 317 L. monocytogenes isolates collected from food, humans, and the environment from 1998 to 2009 was assessed by minimum inhibitory concentration (MIC). Serotyping and MLST was also performed on all isolates. Potential correlations among antimicrobial resistance profiles, serotyping, and MLST were statistically evaluated. Twenty-four percent of L. monocytogenes isolates were resistant to oxacillin, 28.7% intermediate to clindamycin, and 24.3% to ciprofloxacin. The majority of isolates with elevated MIC to oxacillin was of environmental origin and belonged to serotype 4b/4e and ST2. Isolates with intermediate MIC values to clindamycin and ciprofloxacin were mostly of food and human origin and belonged to serotype 4b/4e and ST9. Regarding the time frame of isolate collection, comparing the last 3 years (2007-2009) to previous years (1998-2006), an increase was observed in the percentage of resistant and intermediate isolates per year. This trend strongly suggests the need for increasing attention on the prevalence of antimicrobial resistance in L. monocytogenes in Italy. To predict future resistance trends, the monitoring of clinical intermediate resistance might represent a useful tool especially for antibiotics associated to multiple-step mechanisms of acquired resistance. A specific focus should be addressed to antimicrobial-resistant isolates of serotype 4b, repeatedly associated with food-borne outbreaks.

Background The serovar Typhimurium (4, [5],12:i:1,2), is the most frequently isolated serovar in case of salmonellosis in pigs in Europe and its monophasic variant (4, [5],12:i:-) has been increasingly responsible for Salmonella outbreaks in humans. A total of 25,215 samples were collected, during the years 2002–2017, from 1359 pig farms located in Northern Italy. Samples were collected from different material sources including fecal samples, rectal swabs, gut content and different organs. Results Salmonella was isolated in 15.80% of samples and, among the isolates, 733 were typed as Salmonella Typhimurium (ST) or its monophasic variant (MST). Over time, there was an increase of isolation of MST which outnumbered ST. Most of the strains were isolated in animals during the weaning stage and the growing – fattening period whereas the clinical cases were mainly present in young pigs after weaning. Conclusions This study confirms the presence of ST and MST in pig farms although, considering the total of isolated serotypes, with lower percentages than previously reported. In the last few years, ST has increasingly been replaced by MST suggesting that MST has a competitive advantage over ST, probably due to its different antigenicity and pathogenicity which renders the infection stealthier to recognize and control.

EFSA received a mandate from the European Commission to assess the effectiveness of some of the control measures against diseases included in the Category A list according to Regulation (EU) 2016/429 on transmissible animal diseases (‘Animal Health Law’). This opinion belongs to a series of opinions where these control measures will be assessed, with this opinion covering the assessment of control measures for Newcastle disease (ND). In this opinion, EFSA and the AHAW Panel of experts review the effectiveness of: (i) clinical and laboratory sampling procedures, (ii) monitoring period and (iii) the minimum radius of the protection and surveillance zone, and the minimum length of time the measures should be applied in these zones. The general methodology used for this series of opinions has been published elsewhere. Several scenarios for which these control measures had to be assessed were designed and agreed prior to the start of the assessment. The monitoring period (21 days) was assessed as effective in non-vaccinated chicken and turkey flocks, although large uncertainty remains surrounding the effectiveness of this period in vaccinated galliform flocks and flocks of other bird species. It was also concluded that the protection (3 km radius) and the surveillance (10 km radius) zones contain 99% of the infections from an infectious establishment. Recommendations provided for each of the scenarios assessed aim to support the European Commission in the drafting of further pieces of legislation, as well as for plausible ad hoc requests in relation to ND. info:eu-repo/semantics/publishedVersion

Environmental risk assessment (ERA) is an important component of risk analysis for plant pests and invasive alien species (IAS), and a standardized and consistent methodology has recently been developed for evaluating their impact on ecosystem services and biodiversity. This paper presents the application of this innovative methodology for ERA to Agrilus planipennis, the emerald ash borer, which causes significant mortality to Fraxinus (ash) species in forests and urban areas of North America (here: USA and Canada, excluding Mexico) and Russia. The methodology follows a retrospective analysis and summarizes information and observations in invaded areas in North America and Russia. Uncertainty distributions were elicited to define quantitatively a general pattern of the environmental impact in terms of reduction in ecosystem provisioning, supporting and regulating services, and biodiversity components. The environmental impacts of A. planipennis are time- and context-dependent, therefore two time horizons of 5 and 20 years after introduction and two ecosystems (urban and forest) were considered. This case study shows that the quantitative assessment of environmental impacts for IAS is both possible and helpful for decision-makers and risk managers who have to balance control costs against potential impacts of IAS.

Small nuclear RNA (snRNA) is a class of molecules involved in the processing of pre-mRNA and in regulatory cell processes. snRNAs are always associated with a set of specific proteins. The complexes are referred to as small nuclear ribonucleoproteins, and spliceosome U RNAs are their most common snRNA components. The repetitive sequences of U snDNAs have been cytogenetically mapped in several species of Arthropoda, fishes, and mammals; however, their distribution remains unknown in amphibians. Here, we show results of FISH mapping of U2 snDNA repetitive sequences in species of the amphibian genus Leptodactylus to reveal the distribution patterns of this sequence in their karyotypes. The probe hybridized in the metacentric chromosome pair 6 in Leptodactylus fuscus, L. gracilis, L. latrans, L. chaquensis, L. petersii, L. podicipinus, and L. brevipes. A different pattern was observed in L. labyrinthicus with hybridization signals in 4 chromosome pairs. The same localization of U2 gene sequences in most of the species analyzed suggests a relatively conserved pattern and a similarity of the chromosome 6 among these species of Leptodactylus.

Background Genotypic variability in M. hyopneumoniae has been reported within and among herds. However, information regarding VNTR types within single lung lobes is lacking. The objective of his study was to analyse M. hyopneumoniae infections and their association with VNTR types and lung lesions at the lobe level. Lungs from 300 pigs from 10 farms experiencing an enzootic pneumonia outbreak were collected and scored. M. hyopneumoniae was detected by real-time PCR and genotyped by MLVA assay in all samples. Results The results showed genotypic variability within single pigs and among lung lobes. At the lobe level, infection with one VNTR type (SN infection) was dominant. Lobes with lesion scores > 0 were associated with positive results for real-time PCR. At the lobe level, no relationship was observed between infections with more than one genotype (MX infections) and the proportion of Mycoplasma-like lesions. Lesion-free lobes presented a higher proportion of MX infections than lobes scored > 0. M. hyopneumoniae was detected more frequently in the right lobe of the lung (p M. hyopneumoniae genotype analyses, including lung lobes with no visible lesions. Conclusion The results did not show a significant association between the number of detected genotypes and the severity of the lesions at the lung lobe level, but revealed the unexpected detection of M. hyopneumoniae genotypes in lesion-free lobes. These results imply that a representative sampling of all lobes may lead to an accurate identification of the VNTR-type distribution. Further studies including factors that can affect pathogenetic evolution of this bacterium could shed light on the complexity of the relationship between genotypes and the lung lesions magnitude.

International audience; Scientific communication is facilitated by a data-driven, scientifically sound taxonomy that considers the end-user's needs and established successful practice. Previously (Geiser et al. 2013; Phytopathology 103:400-408. 2013), the Fusarium community voiced near unanimous support for a concept of Fusarium that represented a clade comprising all agriculturally and clinically important Fusarium species, including the F. solani Species Complex (FSSC). Subsequently, this concept was challenged by one research group (Lombard et al. 2015 Studies in Mycology 80: 189-245) who proposed dividing Fusarium into seven genera, including the FSSC as the genus Neocosmospora, with subsequent justification based on claims that the Geiser et al. (2013) concept of Fusarium is polyphyletic (Sandoval-Denis et al. 2018; Persoonia 41:109-129). Here we test this claim, and provide a phylogeny based on exonic nucleotide sequences of 19 orthologous protein-coding genes that strongly support the monophyly of Fusarium including the FSSC. We reassert the practical and scientific argument in support of a Fusarium that includes the FSSC and several other basal lineages, consistent with the longstanding use of this name among plant pathologists, medical mycologists, quarantine officials, regulatory agencies, students and researchers with a stake in its taxonomy. In recognition of this monophyly, 40 species recently described as Neocosmospora were recombined in Fusarium, and nine others were renamed Fusarium. Here the global Fusarium community voices strong support for the inclusion of the FSSC in Fusarium, as it remains the best scientific, nomenclatural and practical taxonomic option available.

Genetic factors play an important role in the susceptibility to pancreatic cancer (PC). However, established loci explain a small proportion of genetic heritability for PC; therefore, more progress is needed to find the missing ones. We aimed at identifying single nucleotide polymorphisms (SNPs) affecting PC risk through effects on micro-RNA (miRNA) function. We searched in silico the genome for SNPs in miRNA seed sequences or 3 prime untranslated regions (3'UTRs) of miRNA target genes. Genome-wide association data of PC cases and controls from the Pancreatic Cancer Cohort (PanScan) Consortium and the Pancreatic Cancer Case–Control (PanC4) Consortium were re-analyzed for discovery, and genotyping data from two additional consortia (PanGenEU and PANDoRA) were used for replication, for a total of 14,062 cases and 11,261 controls. None of the SNPs reached genome-wide significance in the meta-analysis, but for three of them the associations were in the same direction in all the study populations and showed lower value of p in the meta-analyses than in the discovery phase. Specifically, rs7985480 was consistently associated with PC risk (OR = 1.12, 95% CI 1.07–1.17, p = 3.03 × 10−6 in the meta-analysis). This SNP is in linkage disequilibrium (LD) with rs2274048, which modulates binding of various miRNAs to the 3'UTR of UCHL3, a gene involved in PC progression. In conclusion, our results expand the knowledge of the genetic PC risk through miRNA-related SNPs and show the usefulness of functional prioritization to identify genetic polymorphisms associated with PC risk.

EFSA received a mandate from the European Commission to assess the effectiveness of some of the control measures against diseases included in the Category A list according to Regulation (EU) 2016/429 on transmissible animal diseases ('Animal Health Law'). This opinion belongs to a series of opinions where these control measures will be assessed, with this opinion covering the assessment of control measures for Classical swine fever (CSF). In this opinion, EFSA and the AHAW Panel of experts review the effectiveness of: (i) clinical and laboratory sampling procedures, (ii) monitoring period and (iii) the minimum radii of the protection and surveillance zones, and the minimum length of time the measures should be applied in these zones. The general methodology used for this series of opinions has been published elsewhere; nonetheless, details of the model used for answering these questions are presented in this opinion as well as the transmission kernels used for the assessment of the minimum radius of the protection and surveillance zones. Several scenarios for which these control measures had to be assessed were designed and agreed prior to the start of the assessment. Here, several recommendations are given on how to increase the effectiveness of some of the sampling procedures. Based on the average length of the period between virus introduction and the reporting of a CSF suspicion, the monitoring period was assessed as non-effective. In a similar way, it was recommended that the length of the measures in the protection and surveillance zones were increased from 15 to 25 days in the protection zone and from 30 to 40 days in the surveillance zone. Finally, the analysis of existing Kernels for CSF suggested that the radius of the protection and the surveillance zones comprise 99% of the infections from an affected establishment if transmission occurred. Recommendations provided for each of the scenarios assessed aim to support the European Commission in the drafting of further pieces of legislation, as well as for plausible ad hoc requests in relation to CSF. (C) 2021 European Food Safety Authority. EFSA Journal published by John Wiley and Sons Ltd on behalf of European Food Safety Authority.

DSBs are harmful lesions produced through endogenous metabolism or by exogenous agents such as ionizing radiation, that can trigger genomic rearrangements. We have recently shown that exposure to 2 Gy of X-rays has opposite effects on the induction of Shh-dependent MB in NHEJ- and HR-deficient Ptch1+/− mice. In the current study we provide a comprehensive link on the role of HR/NHEJ at low doses (0.042 and 0.25 Gy) from the early molecular changes through DNA damage processing, up to the late consequences of their inactivation on tumorigenesis. Our data indicate a prominent role for HR in genome stability, by preventing spontaneous and radiation-induced oncogenic damage in neural precursors of the cerebellum, the cell of origin of MB. Instead, loss of DNA-PKcs function increased DSBs and apoptosis in neural precursors of the developing cerebellum, leading to killing of tumor initiating cells, and suppression of MB tumorigenesis in DNA-PKcs-/-/Ptch1+/− mice. Pathway analysis demonstrates that DNA-PKcs genetic inactivation confers a remarkable radiation hypersensitivity, as even extremely low radiation doses may deregulate many DDR genes, also triggering p53 pathway activation and cell cycle arrest. Finally, by showing that DNA-PKcs inhibition by NU7441 radiosensitizes human MB cells, our in vitro findings suggest the inclusion of MB in the list of tumors beneficiating from the combination of radiotherapy and DNA-PKcs targeting, holding promise for clinical translation.

As doenças causadas por helmintos e protozoários transmitidas pela alface (Lactuca sativa) possuem grande impacto em saúde pública, sendo consideradas importantes zoonoses. Por isso é importante cuidar da higiene dessa hortaliça previamente ao consumo. O objetivo deste estudo foi avaliar a contaminação por parasitos de importância na saúde pública em amostras de alfaces produzidas e comercializadas na cidade de Campos Novos, Santa Catarina. O estudo foi realizado em duas etapas. Na primeira etapa foram coletadas amostras de alface de supermercados e fruteiras associadas a câmara de dirigentes lojistas da cidade (CDL). Essas alfaces, segundo a descrição da embalagem, eram produzidas de forma hidropônica. Na segunda etapa foram coletadas amostras de alface de escolas municipais e estaduais. As técnicas parasitológicas utilizadas foram Willis Mollay, Dennis Stones e Swanson, Sheater e Faust. Todas as 26 amostras demonstraram ausência de parasitas, sugerido baixo ou nulo grau de contaminação por esses agentes nas hortaliças consumidas no município. Todos os locais onde foram coletadas amostras apresentavam qualidade de alimentos e higiene adequados. É recomendado sempre apresentar os riscos de contaminação à população e salientar a importância da higiene e profilaxia.

Cytokinin together with MdoBRR1, MdoBRR8 and MdoBRR10 genes participate in the downregulation of MdoDAM1, contributing to the transition from endo- to ecodormancy in apple buds. The final step of cytokinin (CK) signaling pathway culminates in the activation of type-B response regulators (BRRs), important transcriptional factors in the modulation of CK-responsive genes. In this study, we performed a genome-wide analysis aiming to identify apple BRR family members and understand their involvement in bud dormancy control. The investigation identified ten MdoBRR protein-coding genes. A higher expression of three MdoBRR (MdoBRR1, MdoBRR9 and MdoBRR10) was observed in dormant buds in comparison to other developmental stages. Interestingly, in ecodormant buds these three MdoBRR genes were upregulated in a CK-dependent manner. Transcription profiles, determined during dormancy cycle under field and artificially controlled conditions, revealed that MdoBRR1 and MdoBRR8 played important roles in the transition from endo- to ecodormancy, probably mediated by endogenous CK stimuli. The expression of MdoBRR7, MdoBRR9, and MdoBRR10 was induced in ecodormant buds exposed to warm temperatures, indicating a putative role in growth resumption after chilling requirement fulfillment. Contrasting expression patternsin vivo between MdoBRRs and MdoDAM1, an essential dormancy establishment regulator, were observed during dormancy cycle and in CK-treated buds. Thereafter, in vivo transactivation assays showed that CK stimuli combined with transient overexpression of MdoBRR1, MdoBRR8, and MdoBRR10 resulted in downregulation of the reporter gene gusA driven by the MdoDAM1 promoter. These pieces of evidences point to the integration of CK-triggered responses through MdoBRRs that are able to downregulate MdoDAM1, contributing to dormancy release in apple.

We assessed the causes of polyserositis in pigs, categorized by causative agents and ages of animals affected. In a 3-y study, 246 pigs from 80 different farms with recurrent problems of polyserositis, in a high-density breeding area, were submitted for autopsy; 154 pigs with typical fibrinous serosal lesions were sampled for further bacterial and viral investigation. The most common gross lesions were pleuritis and pericarditis (141 of 154; 92%). The animals most affected were weaned pigs (139 of 154; 90%). Haemophilus parasuis and Mycoplasma hyorhinis were the most common bacteria detected and were present at the same rate (85 of 154; 55%). Other bacteria isolated were Streptococcus sp. (44 of 154; 29%), Pasteurella multocida (21 of 154; 14%), Escherichia coli (19 of 154; 12%), Actinobacillus pleuropneumoniae (7 of 154; 5%), and Trueperella pyogenes (4 of 154; 3%). Porcine reproductive and respiratory syndrome virus (PRRSV; 119 of 154; 77%) predominated among the viruses detected, followed, with lesser prevalence, by porcine circovirus 2 (40 of 154; 26%) and swine influenza A virus (19 of 154; 12%). Bacterial coinfection and coinfection of bacteria and viruses were common (128 of 154; 83%). A strong positive correlation was found between coinfection by H. parasuis and M. hyorhinis and also by H. parasuis with PRRSV.

Interferon-α (IFNα), a type I interferon, is expressed in the islets of type 1 diabetic individuals, and its expression and signaling are regulated by T1D genetic risk variants and viral infections associated with T1D. We presently characterize human beta cell responses to IFNα by combining ATAC-seq, RNA-seq and proteomics assays. The initial response to IFNα is characterized by chromatin remodeling, followed by changes in transcriptional and translational regulation. IFNα induces changes in alternative splicing (AS) and first exon usage, increasing the diversity of transcripts expressed by the beta cells. This, combined with changes observed on protein modification/degradation, ER stress and MHC class I, may expand antigens presented by beta cells to the immune system. Beta cells also up-regulate the checkpoint proteins PDL1 and HLA-E that may exert a protective role against the autoimmune assault. Data mining of the present multi-omics analysis identifies two compound classes that antagonize IFNα effects on human beta cells., The cytokine IFNα is expressed in the islets of individuals with type 1 diabetes and contributes to local inflammation and destruction of beta cells. Here, the authors provide a global multiomics view of IFNα-induced changes in human beta cells at the level of chromatin, mRNA and protein expression.

The deficiency in micronutrients is a public health problem, principally in lower-middle-income countries. Vitamin A (VA) is considered a micronutrient fundamental to the maintenance and development of different tissues in the organism. Therefore, it is an essential micronutrient in the human diet. In these terms, goat milk is the leading food consumed to provide nutritional support in innumerous lower-middle-income countries. Here our work aimed to produce goat cheese studying strategies to promote the retention of VA. Our experiment design also explores the use of the salting process to evaluate the levels of VA retention. The level of VA in goat cheese was determined using LC-MS/MS analysis. Additionally, the redox status of the goat cheese in terms of lipid peroxidation and protein oxidation was determined. The texture analysis was also evaluated to verify if the redox status and salting process influence the texture profile. The results showed that the salting process during goat cheese production improves the retention of VA in goat cheese. Moreover, the salting process also is related to alterations in the status redox of the goat cheese and texture parameters. Therefore, our results show that goat cheese production can be an alternative to produced dairy derivates with recognized concentrations of VA for human nutrition.

Demethylase inhibitors (DMIs) also referred to as azoles or triazoles are currently the main fungicides used for controlling Fusarium diseases and associated toxins in cereals. DMIs also represent an important class of fungicides used in the medical domain. The level of sensitivity of a set of F. graminearum strains (n = 23), collected over the period 1994–2010 in Luxembourg, Germany, Canada, USA, Italy and Belgium against three DMIs (cyproconazole, propiconazole, tebuconazole) used in agriculture and one DMI used in medicine (tioconazole) was assessed using a microplate test. Median molar EC50 values varied 113-fold among DMIs and on average 11-fold within DMIs with cyproconazole and tebuconazole being the least and the most effective ones, respectively. The EC50 values of the two DMIs registered for use against Fusarium species on cereals (propiconazole and tebuconazole) were significantly correlated (r = 0.597**), while no evidence for cross-resistance was obtained for other fungicide combinations. Haplotypes for CYP51A and CYP51C were defined based on snps determining amino acid variations in the two genes. EC50 values of strains with the CYP51A haplotype A0 and the CYP51C haplotype D1 varied greatly for the agricultural DMIs tebuconazole, propiconazole and cyproconazole, but not for the medical DMI tioconazole. None of the mutations and snps that were previously reported to be associated with resistance towards propiconazole was unambiguously related with resistance to tioconazole, because the mutations and snps were found in strains with low as well as with high EC50 values. Our results show that (1) DMI sensitivity of F. graminearum mycelium has been largely stable between 1994 and 2010, (2) effects of snps on sensitivity towards one DMI detected in one set of strains cannot be extrapolated to other DMIs and sets of strains and (3) F. graminearum strains responded differently to DMIs used in agriculture and to a representative of a medical DMI with no evidence for cross-resistance.

Made available in DSpace on 2020-12-12T01:13:54Z (GMT). No. of bitstreams: 0 Previous issue date: 2020-02-01 Cytogenetic data showed a variation in diploid chromosome number in the genus Hyphessobrycon ranging from 2n = 46 to 52, and studies involving repetitive DNA sequences are scarce in representatives of this genus. The purpose of this paper was the chromosomal mapping of repetitive sequences (rDNA, histone genes, U snDNA and microsatellites) and investigation of the amplification of 5S rDNA clusters in the Hyphessobrycon eques genome. Two H. eques populations displayed 2n = 52 chromosomes, with the acrocentric pair No. 24 bearing Ag-NORs corresponding with CMA3+/DAPI−. FISH with a 18S rDNA probe identified the NORs on the short (p) arms of the acrocentric pairs Nos. 22 and 24. The 5S rDNA probe visualized signals on almost all chromosomes in genomes of individuals from both populations (40 signals); FISH with H3 histone probe identified two chromosome pairs, with the pericentromeric location of signals; FISH with a U2 snDNA probe identified one chromosome pair bearing signals, on the interstitial chromosomal region. The mononucleotide (A), dinucleotide (CA) and tetranucleotide (GATA) repeats were observed on the centromeric/pericentromeric and/or terminal positions of all chromosomes, while the trinucleotide (CAG) repeat showed signals on few chromosomes. Molecular analysis of 5S rDNA and non-transcribed spacers (NTS) showed microsatellites (GATA and A repeats) and a fragment of retrotransposon (SINE3/5S-Sauria) inside the sequences. This study expanded the available cytogenetic data for H. eques and demonstrated to the dispersion of the 5S rDNA sequences on almost all chromosomes. Centro de Ciências Biológicas e da Saúde Laboratório de Citogenética Universidade Estadual do Oeste do Paraná (UNIOESTE), Rua Universitária, 2069 Universidade Estadual de Mato Grosso do Sul (UEMS) Unidade de Mundo Novo, BR 163, Km 20.2 Instituto de Biociências Departamento de Biologia Laboratório de Citogenética Universidade Estadual Paulista “Júlio de Mesquita Filho” (UNESP), Av. 24A, 1515 Instituto de Biociências Departamento de Biologia Laboratório de Citogenética Universidade Estadual Paulista “Júlio de Mesquita Filho” (UNESP), Av. 24A, 1515

Background Clonal chromosome changes are often found in the bone marrow (BM) of patients with Shwachman-Diamond syndrome (SDS). The most frequent ones include an isochromosome of the long arm of chromosome 7, i (7)(q10), and an interstitial deletion of the long arm of chromosome 20, del (20)(q). These two imbalances are mechanisms of somatic genetic rescue. The literature offers few expression studies on SDS. Results We report the expression analysis of bone marrow (BM) cells of patients with SDS in relation to normal karyotype or to the presence of clonal chromosome anomalies: del (20)(q) (five cases), i (7)(q10) (one case), and other anomalies (two cases). The study was performed using the microarray technique considering the whole transcriptome (WT) and three gene subsets selected as relevant in BM functions. The expression patterns of nine healthy controls and SDS patients with or without chromosome anomalies in the bone marrow showed clear differences. Conclusions There is a significant difference between gene expression in the BM of SDS patients and healthy subjects, both at the WT level and in the selected gene sets. The deletion del (20)(q), with the EIF6 gene consistently lost, even in patients with the smallest losses of material, changes the transcription pattern: a low proportion of abnormal cells led to a pattern similar to SDS patients without acquired anomalies, whereas a high proportion yields a pattern similar to healthy subjects. Hence, the benign prognostic value of del (20)(q). The case of i (7)(q10) showed a transcription pattern similar to healthy subjects, paralleling the positive prognostic role of this anomaly as well.

The repetitive DNA content of fish sex chromosomes provides valuable insights into specificities and patterns of their genetic sex determination systems. In this study, we revealed the genomic satellite DNA (satDNA) content of Megaleporinuselongatus, a Neotropical fish species with Z1Z1Z2Z2/Z1W1Z2W2 multiple sex chromosomes, through high-throughput analysis and graph-based clustering, isolating 68 satDNA families. By physically mapping these sequences in female metaphases, we discovered 15 of the most abundant satDNAs clustered in its chromosomes, 9 of which were found exclusively in the highly heterochromatic W1. This heteromorphic sex chromosome showed the highest amount of satDNA accumulations in this species. The second most abundant family, MelSat02-26, shared FISH signals with the NOR-bearing pair in similar patterns and is linked to the multiple sex chromosome system. Our results demonstrate the diverse satDNA content in M. elongatus, especially in its heteromorphic sex chromosome. Additionally, we highlighted the different accumulation patterns and distribution of these sequences across species by physically mapping these satDNAs in other Anostomidae, Megaleporinusmacrocephalus and Leporinusfriderici (a species without differentiated sex chromosomes).

Most eukaryotic genomes contain substantial portions of repetitive DNA sequences. These are located primarily in highly compacted heterochromatin and, in many cases, are one of the most abundant components of the sex chromosomes. In this sense, the anuran Proceratophrys boiei represents an interesting model for analyses on repetitive sequences by means of cytogenetic techniques, since it has a karyotype with large blocks of heterochromatin and a ZZ/ZW sex chromosome system. The present study describes, for the first time, families of satellite DNA (satDNA) in the frog P. boiei. Its genome size was estimated at 1.6 Gb, of which 41% correspond to repetitive sequences, including satDNAs, rDNAs, transposable elements, and other elements characterized as non-repetitive. The satDNAs were mapped by FISH in the centromeric and pericentromeric regions of all chromosomes, suggesting a possible involvement of these sequences in centromere function. SatDNAs are also present in the W sex chromosome, occupying the entire heterochromatic area, indicating a probable contribution of this class of repetitive DNA to the differentiation of the sex chromosomes in this species. This study is a valuable contribution to the existing knowledge on repetitive sequences in amphibians. We show the presence of repetitive DNAs, especially satDNAs, in the genome of P. boiei that might be of relevance in genome organization and regulation, setting the stage for a deeper functional genome analysis of Proceratophrys.

Made available in DSpace on 2022-04-29T08:40:32Z (GMT). No. of bitstreams: 0 Previous issue date: 2021-01-01 Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) A study was conducted to evaluate the effects of including a xylanase + β-glucanase enzyme product in corn–soy-based diets on performance, caecal short-chain fatty acids and thermal profile, ileal digestibility, and intestinal kinetics of broiler chickens. A total of 744 male day-old chicks were randomly allotted to 24 floor pens and distributed in a 2 × 2 factorial arrangement with low or standard energy level × without or with 100 g of the enzyme per ton of feed. Enzyme supplementation improved bodyweight gain from 1 to 21 d (P < 0.05). An increased caecal concentration of acetic acid was observed when the enzyme was added to the low-energy diet (P < 0.05). The pH of the caecal content was reduced (P < 0.01), and the caecal temperature was increased by low-energy diets (P < 0.05). The apparent ileal digestibility of energy was improved by the enzyme (P < 0.01). The addition of the enzyme increased the mean retention time in the distal ileum (P < 0.05). In summary, the addition of a xylanase + β-glucanase enzyme product in corn-based diets increases the retention time of digesta in the distal ileum and the caecal acetic acid concentration, improves ileal digestibility of energy and performance from 1 to 21 d in broiler chickens fed corn–soy-based diets. Department of Breeding and Animal Nutrition College of Veterinary Medicine and Animal Science São Paulo State University, São Paulo Department of Breeding and Animal Nutrition College of Veterinary Medicine and Animal Science São Paulo State University, São Paulo FAPESP: 2015/26623-0

Probiotic bacteria inclusion on fruit pulp is an exceptional alternative for the production and innovation of functional foods. Therefore, the present work aims to study the drying of Caja (Spondias mombin) pulp added with Bifidobacterium animalis ssp. lactis using a spouted bed. An experimental design comprising 11 experiments was used on the drying, varying the inlet temperature (50 ºC, 60 ºC, and 70 ºC), the maltodextrin concentration (2.5, 5.0, and 7.5% wt) and inulin concentration (2.5, 5.0, and 7.5% wt). The dependent variables studied in our work were the process yield and the number of viable cells. The liquid fruit pulp presented physicochemical conditions for bacteria growth and high content of bioactive compounds. Results revealed that lower inlet temperature in combination with lower concentrations of maltodextrin and inulin resulted in a powder with a higher number of viable cells. Even though temperature degradation is a critical factor for product quality while using drying processes, it was considered that the process was efficient because of the high cell viability, which is the critical factor to consider the product as probiotic.

In this opinion the antimicrobial-resistant bacteria responsible for transmissible diseases that constitute a threat to dog and cat health have been assessed. The assessment has been performed following a methodology based on information collected via an extensive literature review and expert judgement. Details of the methodology used for this assessment are explained in a separate opinion. A global state of play of antimicrobial resistant Staphylococcus pseudintermedius, Staphylococcus aureus, Staphylococcus schleiferi, Escherichia coli, Proteus mirabilis, Klebsiella spp., Enterobacter spp., Pseudomonas aeruginosa, Clostridium perfringens, Clostridioides difficile, Enterococcus faecalis and Enterococcus faecium has been provided. Among those bacteria, EFSA identified S. pseudintermedius, E. coli and P. aeruginosa with > 90% certainty as the most relevant antimicrobial resistant bacteria in the EU based on the available evidence. The animal health impact of these most relevant bacteria, as well as their eligibility for being listed and categorised within the animal health law framework will be assessed in separate scientific opinions. (C) 2021 European Food Safety Authority. EFSA Journal published by John Wiley and Sons Ltd on behalf of European Food Safety Authority.

The European epidemic monophasic variant of Salmonella enterica serovar Typhimurium (S. 1,4,[5],12:i:-) characterized by the multi locus sequence type ST34 and the antimicrobial resistance ASSuT profile has become one of the most common serovars in Europe (EU) and the United States (US). In this study, we reconstructed the time-scaled phylogeny and evolution of this Salmonella in Europe. The epidemic S. 1,4,[5],12:i:- ST34 emerged in the 1980s by an acquisition of the Salmonella Genomic Island (SGI)-4 at the 3′ end of the phenylalanine phe tRNA locus conferring resistance to copper and arsenic toxicity. Subsequent integration of the Tn21 transposon into the fljAB locus gave resistance to mercury toxicity and several classes of antibiotics used in food-producing animals (ASSuT profile). The second step of the evolution occurred in the 1990s, with the integration of mTmV and mTmV-like prophages carrying the perC and/or sopE genes involved in the ability to reduce nitrates in intestinal contents and facilitate the disruption of the junctions of the host intestinal epithelial cells. Heavy metals are largely used as food supplements or pesticide for cultivation of seeds intended for animal feed so the expansion of the epidemic S. 1,4,[5],12:i:- ST34 was strongly related to the multiple-heavy metal resistance acquired by transposons, integrative and conjugative elements and facilitated by the escape until 2011 from the regulatory actions applied in the control of S. Typhimurium in Europe. The genomic plasticity of the epidemic S. 1,4,[5],12:i:- was demonstrated in our study by the analysis of the plasmidome. We were able to identify plasmids harboring genes mediating resistance to phenicols, colistin, and fluoroquinolone and also describe for the first time in six of the analyzed genomes the presence of two plasmids (pERR1744967-1 and pERR2174855-2) previously described only in strains of enterotoxigenic Escherichia coli and E. fergusonii.

This opinion assesses the risk posed by different matrices to introduce African swine fever virus (ASFV) to non‐affected regions of the EU. Matrices assessed are feed materials, enrichment/bedding materials and empty live pigs transport vehicles returning from affected areas. Although the risk from feed is considered to be lower than several other pathways (e.g. contact with infected live animals and swill feeding), it cannot be ruled out that matrices assessed in this opinion pose a risk. Evidence on survival of ASFV in different matrices from literature and a public consultation was used in an Expert Knowledge Elicitation (EKE) on the possible contamination of products and traded or imported product volumes used on pig farms. The EKE results were used in a model that provided a risk‐rank for each product's contamination likelihood (‘q’), its trade or import volume from affected EU or Eurasian areas (N) and the modelled number of potentially infected pig farms (N × q). The products ranking higher regardless of origin or destination were mash and pelleted compound feed, feed additives and cereals. Bedding/enrichment materials, hydrolysed proteins and blood products ranked lowest regardless of origin or destination. Empty vehicles ranked lower than compound feed but higher than non‐compound feed or bedding/enrichment material. It is very likely (95–99% certainty) that compound feed and cereals rank higher than feed materials, which rank higher than bedding/enrichment material and forage. As this is an assessment based on several parameters including the contamination and delivery to a pig farm, all of which have the same impact on the final ranking, risk managers should consider how the relative rank of each product may change with an effective storage period or a virus inactivation step., This publication is linked to the following EFSA Supporting Publications articles: http://www.efsa.europa.eu/en/supporting/pub/9993e and http://www.efsa.europa.eu/en/supporting/pub/9994e

Background: In patients with Multiple Sclerosis (pwMS) disease-modifying therapies (DMTs) affects immune response to antigens. Therefore, post-vaccination serological assessments are needed to evaluate the effect of the vaccine on SARS-CoV-2 antibody response. Methods: We designed a prospective multicenter cohort study enrolling pwMS who were scheduled for SARS-Cov-2 vaccination with mRNA vaccines (BNT162b2, Pfizer/BioNTech,Inc or mRNA-1273, Moderna Tx,Inc). A blood collection before the first vaccine dose and 4 weeks after the second dose was planned, with a centralized serological assessment (electrochemiluminescence immunoassay, ECLIA, Roche-Diagnostics). The log-transform of the antibody levels was analyzed by multivariable linear regression. Findings: 780 pwMS (76% BNT162b2 and 24% mRNA-1273) had pre- and 4-week post-vaccination blood assessments. 87 (11·2%) were untreated, 154 (19·7%) on ocrelizumab, 25 (3·2%) on rituximab, 85 (10·9%) on fingolimod, 25 (3·2%) on cladribine and 404 (51·7%) on other DMTs. 677 patients (86·8%) had detectable post-vaccination SARS-CoV-2 antibodies. At multivariable analysis, the antibody levels of patients on ocrelizumab (201-fold decrease (95%CI=128–317), p < 0·001), fingolimod (26-fold decrease (95%CI=16–42), p < 0·001) and rituximab (20-fold decrease (95%CI=10–43), p < 0·001) were significantly reduced as compared to untreated patients. Vaccination with mRNA-1273 resulted in a systematically 3·25-fold higher antibody level (95%CI=2·46–4·27) than with the BNT162b2 vaccine (p < 0·001). The antibody levels on anti-CD20 therapies correlated to the time since last infusion, and rituximab had longer intervals (mean=386 days) than ocrelizumab patients (mean=129 days). Interpretation: In pwMS, anti-CD20 treatment and fingolimod led to a reduced humoral response to mRNA-based SARS-CoV-2 vaccines. As mRNA-1273 elicits 3·25-higher antibody levels than BNT162b2, this vaccine may be preferentially considered for patients under anti-CD20 treatment or fingolimod. Combining our data with those on the cellular immune response to vaccines, and including clinical follow-up, will contribute to better define the most appropriate SARS-CoV-2 vaccine strategies in the context of DMTs and MS. Funding: FISM[2021/Special-Multi/001]; Italian Ministry of Health‘Progetto Z844A 5 × 1000′.

Background: The mitochondrial metabolism has been associated with pancreatic ductal adenocarcinoma (PDAC) risk. Recent evidence also suggests the involvement of the genetic variability of the mitochondrial function in several traits involved in PDAC etiology. However, a systematic investigation of the genetic variability of mitochondrial genome (mtSNP) and of all the nuclear genes involved in its functioning (n-mtSNPs) has never been reported. Methods: We conducted a two-phase association study of mtSNPs and n-mtSNPs to assess their effect on PDAC risk. We analyzed 35,297 n-mtSNPs and 101 mtSNPs in up to 55,870 individuals (12,884 PDAC cases and 42,986 controls). In addition, we also conducted a gene-based analysis on 1,588 genes involved in mitochondrial metabolism using Multi-marker Analysis of GenoMic Annotation (MAGMA) software. Results: In the discovery phase, we identified 49 n-mtSNPs and no mtSNPs associated with PDAC risk (P < 0.05). In the second phase, none of the findings were replicated. In the gene-level analysis, we observed that three genes (TERT, SUGCT, and SURF1) involved in the mitochondrial metabolism showed an association below the Bonferroni-corrected threshold of statistical significance (P = 0.05/1588 = 3.1 × 10−5). Conclusions: Even though the mitochondrial metabolism might be involved in PDAC etiology, our results, obtained in a study with one of the largest sample sizes to date, show that neither n-mtSNPs nor mtSNPs are associated with PDAC risk. Impact: This large case–control study does not support a role of the genetic variability of the mitochondrial function in PDAC risk.

Source attribution and microbial risk assessment have proved to be crucial to identify and prioritize food safety interventions as to effectively control the burden of human illnesses (Cassini et al., 2016; Mughini-Gras et al., 2018a, 2019). By comparing human cases and pathogen occurrences in selected animal, food, and environmental sources, microbial subtyping approaches were successfully applied to pinpoint the most important sources of Salmonella, Campylobacter, Shiga toxin-producing Escherichia coli, and Listeria monocytogenes (Hald et al., 2004; Mullner et al., 2009a,b; Barco et al., 2013; Nielsen et al., 2017; Mughini-Gras et al., 2018b; Cody et al., 2019). Microbial risk assessment has been applied to assess known or potential adverse health effects resulting from human exposure to food-borne hazards. Through a scientific structured approach (FAO and WHO, 2021), microbial risk assessment helps to identify and quantify the risk represented by specific foods and the critical points in these foods' production chains for microbial control (Cassini et al., 2016; FAO and WHO, 2021). For both source attribution and risk assessment, one key challenge has been to define the hazard in question: is the whole foodborne pathogen species a hazard, or only some of its subtypes? In this regard the choice of the subtyping method becomes crucial. In recent years, Whole Genome Sequencing (WGS) has represented a major benefit for more targeted approaches, no longer focused on the species/genus level but at the level of subtypes (Franz et al., 2016; Fritsch et al., 2018; EFSA Panel on Biological Hazards, 2019). Besides WGS, metagenomics showed potentialities in source attribution. In particular, this approach was useful in attributing the source of environmental contamination by comparing the abundances of source-specific genetic markers (i.e., resistome) in different reservoirs (Gupta et al., 2019). Therefore, this special issue focuses on traditional and novel source attribution approaches applied on molecular, WGS, and metagenomic data as well as on a fine-tuning genetic characterization of foodborne pathogens useful for hazard identification and characterization. In particular, one study compares the outputs of a modified Hald model, which was applied to different subtyping input data of S. enterica Typhimurium and its monophasic variant (Arnold et al.) whereas two studies proposed a novel network approach and a method based on the core-genome genetic distance to attribute human infections of S. enterica Typhimurium monophasic variant and S. enterica Derby using WGS as input data (Merlotti et al.; Sévellec et al.). Another study by Duarte et al. included the relative abundance of antimicrobial resistance (AMR) associated genes (resistome) as metagenomic input data in an AMR source attribution study. Finally, two studies were focused on the molecular and genomic characterization of human isolates of Campylobacter jejuni and C. coli from China and of Listeria monocytogenes isolates collected from ready-to-eat meat and processing environment from Poland (Zhang et al.; Kurpas et al.). Arnold et al. performed a source attribution study including the genomes of S. enterica Typhimurium and its monophasic variant of 596 human sources and 327 animal sources from England and Wales between 2014 and 2016. Data from Seven Loci Multi Locus Sequence Typing (7-loci MLST), core-genome MLST (cg-MLST), and SNP calling were compared as input data. By applying a modified Hald model, 60% of human genomes were attributed to pork. Comparing different input data, results highlighted MLST as the method with the lowest fit and the lowest discriminatory power. Merlotti et al. applied a network approach to 351 human and animal genomes of S. enterica Typhimurium and its monophasic variant collected from 2013 to 2014. Three datasets of whole-genome MLST (wgMLST), cgMLST, and SNPs were used as input data. Genomes were clustered based on their genetic similarities. Interestingly, a higher percentage of cluster coherence was reported for animal sources in comparison to country and year of isolation, suggesting animal sources as the major driver of cluster formation. The approach showed to be effective in attributing up to 97.2% of human genomes to animal sources represented in the dataset. Among these genomes, the majority (84%) was attributed to pigs/pork. No significant differences were highlighted by comparing the three different input datasets. Core genome analysis was the approach applied by Sévellec et al. to attribute human sporadic cases of S. enterica Derby that occurred in France in 2014–2015 to non-human reservoirs. The authors analyzed 299 S. enterica Derby genomes corresponding to all S. enterica Derby sporadic human cases registered in the time frame, along with 141 non-human genomes. Within the non-human genomes, three main genomic lineages were detected in France: ST39-ST40 and ST682 associated to pork and ST71 associated to poultry. Within human genomes, 94% of S. enterica Derby clustered within the three genetic groups associated with pork, identifying this animal reservoir as the major contributor of S. enterica Derby to sporadic human cases in France. Relative abundance of antimicrobial resistance genes in shotgun metagenomic data was chosen in an antimicrobial resistance source attribution study by Duarte et al.. Starting from the assumption that fecal resistomes are source related, authors compared the resistomes of pooled fecal samples of pigs, broilers, turkeys, and veal calves with the resistomes of individual fecal samples of humans occupationally exposed to livestock production. Five supervised random forest models were applied on a total of 479 observations. Among the four livestock species, the results indicated that pigs have the resistome composition closest to the composition of the human resistome suggesting that occupational exposure to AMR determinants was higher among workers exposed to pigs than workers of broiler farms. Zhang et al. characterized genetic diversity and antimicrobial resistance of 236 Campylobacter jejuni and C. coli isolates collected from 2,945 individual stool samples of hospitalized patients with diarrhea in Beijing from 2017 to 2018. MLST results confirmed the high genetic diversity among isolates as well as CC21 as the most common clonal complex of C. jejuni in diarrhea patients in China. Clonal complex CC828 was the most frequently identified among C. coli isolates. Regarding antimicrobial resistance, rates higher than 88% were identified for the antimicrobials nalidixic acid, ciprofloxacin, and tetracycline. Last but not least, Kurpas et al. genetically characterized 48 L. monocytogenes isolates of PCR-serogroup IIb and IVb collected from ready-to-eat food and food processing environments. Additionally, the authors compared them with public genomes collected from humans in Poland. Among food isolates, 65% belonged to CC1, CC2, and CC6 already described as hypervirulent strains in humans. The clonal complex CC5 was also identified; mostly collected from food processing environments and belonging to PCR-serogroup IIB. Genomes of this clonal complex showed mutations in the inlA gene and a deletion of 144 bp in the inlB gene suggesting them as hypovirulent. Based on these studies, we conclude that the application of NGS data, in particular source attribution models, shows great potential. The results are improved by becoming more specific and to the point, which is considered very valuable for the decision support process. Integrations with phenotypic tests will continue to be essential for confirmation of NGS predicted outcomes.

The European Commission asked EFSA to provide study designs for the investigation of four research domains (RDs) according to major gaps in knowledge identified by EFSA in a report published in 2019: (RD 1) African swine fever (ASF) epidemiology in wild boar; (RD 2) ASF transmission by vectors; (RD 3) African swine fever virus (ASFV) survival in the environment, and (RD 4) the patterns of seasonality of ASF in wild boar and domestic pigs in the EU. In this Scientific Opinion, the second RD on ASF epidemiology in wild boar is addressed. Twenty-nine research objectives were proposed by the working group and broader ASF expert networks and 23 of these research objectives met a prespecified inclusion criterion. Fourteen of these 23 research objectives met the predefined threshold for selection and so were prioritised based on the following set of criteria: (1) the impact on ASF management; (2) the feasibility or practicality to carry out the study; (3) the potential implementation of study results in practice; (4) a possible short time-frame study (< 1 year); (5) the novelty of the study; and (6) if it was a priority for risk managers. Finally, after further elimination of three of the proposed research objectives due to overlapping scope of studies published during the development of this opinion, 11 research priorities were elaborated into short research proposals, considering the potential impact on ASF management and the period of one year for the research activities. (C) 2021 European Food Safety Authority.

Pancreatic ductal adenocarcinoma (PDAC) is projected to become the second cancer-related cause of death by 2030. Identifying novel risk factors, including genetic risk loci, could be instrumental in risk stratification and implementation of prevention strategies. Long noncoding RNAs (lncRNAs) are involved in regulation of key biological processes, and the possible role of their genetic variability has been unexplored so far. Combining genome wide association studies and functional data, we investigated the genetic variability in all lncRNAs. We analyzed 9893 PDAC cases and 9969 controls and identified a genome-wide significant association between the rs7046076 SNP and risk of developing PDAC (P = 9.73 × 10-9 ). This SNP is located in the NONHSAG053086.2 (lnc-SMC2-1) gene and the risk allele is predicted to disrupt the binding of the lncRNA with the micro-RNA (miRNA) hsa-mir-1256 that regulates several genes involved in cell cycle, such as CDKN2B. The CDKN2B region is pleiotropic and its genetic variants have been associated with several human diseases, possibly though an imperfect interaction between lncRNA and miRNA. We present a novel PDAC risk locus, supported by a genome-wide statistical significance and a plausible biological mechanism.

Background An isochromosome of the long arm of chromosome 7, i(7)(q10), and an interstitial deletion of the long arm of chromosome 20, del(20)(q), are the most frequent anomalies in the bone marrow of patients with Shwachman-Diamond syndrome, which is caused in most cases by mutations of the SBDS gene. These clonal changes imply milder haematological symptoms and lower risk of myelodysplastic syndromes and acute myeloid leukaemia, thanks to already postulated rescue mechanisms. Results Bone marrow from fourteen patients exhibiting either the i(7)(q10) or the del(20)(q) and coming from two large cohorts of patients, were subjected to chromosome analyses, Fluorescent In Situ Hybridization with informative probes and array-Comparative Genomic Hybridization. One patient with the i(7)(q10) showed a subsequent clonal rearrangement of the normal chromosome 7 across years. Four patients carrying the del(20)(q) evolved further different del(20)(q) independent clones, within a single bone marrow sample, or across sequential samples. One patient with the del(20)(q), developed a parallel different clone with a duplication of chromosome 3 long arm. Eight patients bore the del(20)(q) as the sole chromosomal abnormality. An overall overview of patients with the del(20)(q), also including cases already reported, confirmed that all the deletions were interstitial. The loss of material varied from 1.7 to 26.9 Mb and resulted in the loss of the EIF6 gene in all patients. Conclusions Although the i(7)(q) and the del(20)(q) clones are frequent and clinically benign in Shwachman Diamond-syndrome, in the present work we show that they may rearrange, may be lost and then reconstructed de novo, or may evolve with independent clones across years. These findings unravel a striking selective pressure exerted by SBDS deficiency driving to karyotype instability and to specific clonal abnormalities.

A new adsorbent from Jerivá coconut (a native fruit of the Atlantic Forest) was developed through thermal activation at high temperatures and chemical activation using H3PO4. The characterization of activated carbon revealed the functional groups O-H, NH2 and C=C. The morphology showed an irregular and heterogeneous surface, with a pore volume of 0.442 cm3 g−1 and a surface area of 750 m2 g−1. The main variables of the process were evaluated for statistical significance in the removal of the methylene blue dye showing that only the agitation of the system affected the process. Adsorption kinetics were performed at three different concentrations and fitted to nonlinear models where the Avrami-fractional-order model represented the experimental data. Equilibrium occurred within 80 min and was sufficient to remove more than 99% of the initial concentration. The sorption isotherms were performed at four different temperatures, fitted to four nonlinear models. The Sips model obtained the best adjustment, with a maximum adsorption capacity of 254.40 mg g−1, for the highest temperature investigated. The process occurred spontaneously, favorably and endothermically. The results were promising, pointing out that this adsorbent has the potential to remove methylene blue and the treated effluent can be reused for less noble purposes in the industry.

Salmonella enterica subspecies enterica serovar Typhimurium and its monophasic variant are among the most common Salmonella serovars associated with human salmonellosis each year. Related infections are often due to consumption of contaminated meat of pig, cattle, and poultry origin. In order to evaluate novel microbial subtyping methods for source attribution, an approach based on weighted networks was applied on 141 human and 210 food and animal isolates of pigs, broilers, layers, ducks, and cattle collected in Denmark from 2013 to 2014. A whole-genome SNP calling was performed along with cgMLST and wgMLST. Based on these genomic input data, pairwise distance matrices were built and used as input for construction of a weighted network where nodes represent genomes and links to distances. Analyzing food and animal Typhimurium genomes, the coherence of source clustering ranged from 89 to 90% for animal source, from 84 to 85% for country, and from 63 to 65% for year of isolation and was equal to 82% for serotype, suggesting animal source as the first driver of clustering formation. Adding human isolate genomes to the network, a percentage between 93.6 and 97.2% clustered with the existing component and only a percentage between 2.8 and 6.4% appeared as not attributed to any animal sources. The majority of human genomes were attributed to pigs with probabilities ranging from 83.9 to 84.5%, followed by broilers, ducks, cattle, and layers in descending order. In conclusion, a weighted network approach based on pairwise SNPs, cgMLST, and wgMLST matrices showed promising results for source attribution studies.

Salmonella enterica ser. Typhimurium monophasic variant 4,[5],12:i:- has been associated with food-borne epidemics worldwide and swine appeared to be the main reservoir in most of the countries of isolation. However, the monomorphic nature of this serovar has, so far, hindered identification of the source due to expansion of clonal lineages in multiple hosts and food producing systems. Since geographically structured genetic signals can shape bacterial populations, identification of biogeographical markers in S. 1,4,[5],12:i:- genomes can contribute to improving source attribution. In this study, the phylogeographical structure of 148 geographically and temporally related Italian S. 1,4,[5],12:i:- has been investigated. The Italian isolates belong to a large population of clonal S. Typhimurium/1,4,[5],12:i:- isolates collected worldwide in two decades showing up to 2.5% of allele differences. Phylogenetic reconstruction revealed that isolates from the same geographical origin form highly supported monophyletic groups, suggesting discrete geographical segregation. These monophyletic groups are characterized by the gene content of a large sopE-containing prophage. Within this prophage, genome-wide comparison identified several genes overrepresented in strains of Italian origin. This suggests that certain lineages may be characterized by the acquisition of specific accessory genetic markers useful for improving identification of the source in ongoing epidemics.

Background. Pheochromocytomas (PCCs) show the highest degree of heritability in human neoplasms. However, despite the wide number of alterations until now reported in PCCs, it is likely that other susceptibility genes remain still unknown, especially for those PCCs not clearly syndromic. Methods. Whole exome sequencing of tumor DNA was performed on a set of twelve PCCs clinically defined as sporadic. Results. About 50% of PCCs examined had somatic mutations on the known susceptibility VHL, NF1, and RET genes. In addition to these driver events, mutations on SYNE1, ABCC10, and RAD54B genes were also detected. Moreover, extremely rare germline variants were present in half of the sporadic PCC samples analyzed, in particular variants of MAX and SAMD9L were detected in the germline of cases wild-type for mutations in the known susceptibility genes. Conclusions. Additional somatic passenger mutations can be associated with known susceptibility VHL, NF1, and RET genes in PCCs, and a wide number of germline variants with still unknown clinical significance can be detected in these patients. Therefore, many efforts should be aimed to better define the pathogenetic role of all these germline variants for discovering novel potential therapeutic targets for this disease still orphan of effective treatments.

Recent improvements in our understanding of physiology have altered the way in which bone is perceived: no longer is it considered as simply the repository of divalent ions, but rather as a sophisticated endocrine organ with potential extraskeletal effects. Indeed, a number of pathologic conditions involving bone in different ways can now be reconsidered from a bone-centred perspective. For example, in metabolic bone diseases like osteoporosis (OP) and renal osteodystrophy (ROD), the association with a worse cardiovascular outcome can be tentatively explained by the possible derangements of three recently discovered bone hormones (osteocalcin, fibroblast growth factor 23 and sclerostin) and a bone-specific enzyme (alkaline phosphatase). Further, in recent years the close link between bone and inflammation has been better appreciated and a wide range of chronic inflammatory states (from rheumatoid arthritis to ageing) are being explored to discover the biochemical changes that ultimately lead to bone loss and OP. Also, it has been acknowledged that the concept of the bone-vascular axis may explain, for example, the relationship between bone metabolism and vessel wall diseases like atherosclerosis and arteriosclerosis, with potential involvement of a number of cytokines and metabolic pathways. A very important discovery in bone physiology is the bone marrow (BM) niche, the functional unit where stem cells interact, exchanging signals that impact on their fate as bone-forming cells or immune-competent haematopoietic elements. This new element of bone physiology has been recognized to be dysfunctional in diabetes (so-called diabetic mobilopathy), with possible clinical implications. In our opinion, ROD, the metabolic bone disease of renal patients, will in the future probably be identified as a cause of BM niche dysfunction. An integrated view of bone, which includes the BM niche, now seems necessary in order to understand the complex clinical entity of chronic kidney disease-mineral and bone disorders and its cardiovascular burden. Bone is thus becoming a recurrently considered paradigm for different inter-organ communications that needs to be considered in patients with complex diseases.

Neotropical fishes have highly diversified karyotypic and genomic characteristics and present many diverse sex chromosome systems, with various degrees of sex chromosome differentiation. Knowledge on their sex-specific composition and evolution, however, is still limited. Satellite DNAs (satDNAs) are tandemly repeated sequences with pervasive genomic distribution and distinctive evolutionary pathways, and investigating satDNA content might shed light into how genome architecture is organized in fishes and in their sex chromosomes. The present study investigated the satellitome of Megaleporinus elongatus, a freshwater fish with a proposed Z1Z1Z2Z2/Z1W1Z2W2 multiple sex chromosome system that encompasses a highly heterochromatic and differentiated W1 chromosome. The species satellitome comprises of 140 different satDNA families, including previously isolated sequences and new families found in this study. This diversity is remarkable considering the relatively low proportion that satDNAs generally account for the M. elongatus genome (around only 5%). Differences between the sexes in regards of satDNA content were also evidenced, as these sequences are 14% more abundant in the female genome. The occurrence of sex-biased signatures of satDNA evolution in the species is tightly linked to satellite enrichment associated with W1 in females. Although both sexes share practically all satDNAs, the overall massive amplification of only a few of them accompanied the W1 differentiation. We also investigated the expansion and diversification of the two most abundant satDNAs of M. elongatus, MelSat01-36 and MelSat02-26, both highly amplified sequences in W1 and, in MelSat02-26’s case, also harbored by Z2 and W2 chromosomes. We compared their occurrences in M. elongatus and the sister species M. macrocephalus (with a standard ZW sex chromosome system) and concluded that both satDNAs have led to the formation of highly amplified arrays in both species; however, they formed species-specific organization on female-restricted sex chromosomes. Our results show how satDNA composition is highly diversified in M. elongatus, in which their accumulation is significantly contributing to W1 differentiation and not satDNA diversity per se. Also, the evolutionary behavior of these repeats may be associated with genome plasticity and satDNA variability between the sexes and between closely related species, influencing how seemingly homeologous heteromorphic sex chromosomes undergo independent satDNA evolution.

The health problem of antimicrobial resistance (AMR) involves several species. AMR surveillance is essential to identify its development and design control strategies, however, available data are still limited in some contexts. The AMR profiles of 2612 E. coli strains isolated over a period of 15 years (2002–2016) from calf enteric cases were analyzed to determine the presence of resistance and their temporal dynamics. Furthermore, the AMR profiles and the presence of the major virulence genes of 505 E. coli strains isolated from 1-week- and 2-week-old calves, 406 treated with antimicrobials and 99 untreated, were analyzed and compared to investigate the potential effects of treatment on AMR and strain pathogenicity. Resistance to tetracycline (90.70%) was the most common, followed by resistance to sulfamethoxazole/trimethoprim (77.70%) and flumequine (72.10%). The significantly higher percentage of AMR and virulence gene expression recorded in treated calves, combined with the statistically higher resistance to sulfamethoxazole/trimethoprim in E. coli with K99, corroborates the notion of resistance being induced by the frequent use of antimicrobials, leading to treatments potentially becoming ineffective. The significantly higher resistance to amoxicillin/clavulanic acid, enrofloxacin, and florfenicol in isolates from 1-week-old calves suggests the role of the environment as a source of contamination that should be investigated further.

Purpose Aim of the study was to investigate whether aPL positivity correlated with thrombosis in COVID-19 patients and whether it was transient or persistent. Methods We enrolled COVID-19 patients who underwent aPL tests: Lupus Anticoagulant (LA); IgM, IgG, IgA anticardiolipin antibodies (aCL); and IgM, IgG anti-β2-Glycoprotein-I antibodies (aβ2GPI). Results Twenty-eight out of 73 (38.4%) patients resulted positive for at least one aPL assay: 32.8% for IgA aCL, 6.8% for IgM aCL and 4.1% for IgM aβ2GPI. No patients tested positive for IgG aPL or LA at the first determination. Seven (9.6%) patients developed thrombotic events during hospitalization, with 4 of them resulting positive for aPL. In patients with thrombotic events during hospitalization the risk of death was increased 9-fold (LR+8.9, p=0.003). Patients with double positivity for aCL and aβ2GPI IgM had a LR+ of 6.3 to have thrombotic events (p=0.012) and a LR+ of 4.9 to have elevated D-dimer levels (p=0.027). In 10 out of 28 positive patients, aPL was detected in a second occasion at least 12-weeks apart and two patients confirmed the positivity. Conclusion Results suggest that double positivity for aCL and aβ2GPI IgM increases the risk of thrombosis in COVID-19, unlike IgA aCL positivity. APL positivity may be persistent, and it is advisable to monitor it over time.

It has been hypothesized that the occurrence of central serous chorioretinopathy (CSC) might be associated with stress. Therefore, our purpose was to investigate the diurnal trajectories of salivary cortisol and α-amylase (α-Amy) - markers of hypothalamic-pituitary-adrenal (HPA) axis and sympathetic-adrenal-medullary (SAM) system activity, respectively - and psychological profiles in idiopathic acute CSC. This cross-sectional observational case-control study, which included self-reported psychometric questionnaires, was formally approved by the Ethics Committee. Written informed consent was obtained from all participants. Home diurnal saliva collection was scheduled at several timepoints: at awakening, 30 and 60 min later, and at approximately 13:00 h and 20:00 h. Twenty consecutive male subjects with first-episode CSC attending the outpatient clinic of the Retina Medical Service at the Bietti Foundation were enrolled in the study. Twenty age-matched subjects were recruited as controls. After their initial enrollment, 3 subjects per group were excluded. The production of cortisol and α-Amy and the scores on the negative subscale of the Positive/Negative Affect Schedule, the Daily Hassles and Stress Scale and the Beck Depression Inventory were higher in the CSC group than in the control group. To estimate the diurnal trends in the production of salivary cortisol and α-Amy, an equation was derived for each group of the study population. The equations describing the interpolated regression lines gave salivary cortisol and salivary α-Amy slopes that were determined to be significantly different by Student's t-test (cortisol: t = 3.533

Toxoplasma gondii is a protozoan with worldwide distribution, responsible for causing toxoplasmosis, a disease that affects mammals, reptiles, amphibians and birds. The objective of this study was to evaluate T. gondii seroprevalence in dogs and cats domiciled in cities from West and Serrana mesoregions of Santa Catarina. A total of 338 blood samples, 211 from dogs and 127 from cats were collected and submitted to the indirect haemagglutination technique (IHT) for the detection of anti-T. gondii antibodies. Epidemiological questionnaire asking for information related to gender, breed, temperament, behavior, immunity, body index, diet, ectoparasites, access to the streets, hunting habit was applied to the animals owners. The results were 71.89% (243/338) of seropositive animals, being 69.19% (146/211) dogs and 76.38% (97/127) cats. The variable that presented statistical significance was commercial ration for dogs as protection factor. The present study demonstrates a high prevalence of anti-T. gondii antibodies in dogs and cats domiciled in West and Serrana mesoregions of Santa Catarina state, Brazil. Through the obtained results we can reinforce the importance of feeding dogs with commercial food, rather than home-made food, avoiding the consumption of raw or undercooked meat by the animals.

Producción Científica, Fusarium circinatum is a harmful pathogenic fungus mostly attacking Pinus species and also Pseudotsuga menziesii, causing cankers in trees of all ages, damping-of in seedlings, and mortality in cuttings and mother plants for clonal production. This fungus is listed as a quarantine pest in several parts of the world and the trade of potentially contaminated pine material such as cuttings, seedlings or seeds is restricted in order to prevent its spread to disease-free areas. Inspection of plant material often relies on DNA testing and several conventional or real-time PCR based tests targeting F. circinatum are available in the literature. In this work, an international collaborative study joined 23 partners to assess the transferability and the performance of nine molecular protocols, using a wide panel of DNA from 71 representative strains of F. circinatum and related Fusarium species. Diagnostic sensitivity, specifcity and accuracy of the nine protocols all reached values >80%, and the diagnostic specifcity was the only parameter difering signifcantly between protocols.The rates of false positives and of false negatives were computed and only the false positive rates difered signifcantly, ranging from 3.0% to 17.3%.The diference between protocols for some ofthe performance values were mainly due to cross-reactions with DNA from non-target species, which were either not tested or documented in the original articles. Considering that participating laboratories were free to use their own reagents and equipment, this study demonstrated that the diagnostic protocols for F. circinatum were not easily transferable to endusers. More generally, our results suggest that the use of protocols using conventional or real-time PCR outside their initial development and validation conditions should require careful characterization of the performance data priorto use under modifed conditions (i.e. reagents and equipment). Suggestions to improve the transfer are proposed., European Cooperation in Science and Technology (COST Action FP1406 PINESTRENGTH), Estonian Science Foundation (grants PSG136 and IUT21-04), Centro de Estudios do Ambiente e do Mar (CESAM) (projects UID/AMB/50017/2013 and POCI-01- 0145- FEDER-007638), Slovenian Research Agency (project P4-0107)

Over an eight-year period, 686 winter wheat grain samples and information on their cropping history were obtained from Swiss growers. To estimate the risk of Fusarium head blight (FHB), grains were examined for Fusarium species incidence, mycotoxin content as well as the abundance of F. graminearum (FG) and F. poae (FP) DNA and three chemotypes, 15-acetyl-deoxynivalenol (15ADON), 3-acetyl-deoxynivalenol (3ADON) and nivalenol (NIV). Of all Fusarium species, FG and FP were predominant, and the average abundance of the FG DNA was three times higher compared with that of FP. The average detection of the 15ADON chemotype was twice as high as those of 3ADON and NIV, respectively. Deoxynivalenol (DON), zearalenone (ZEA) and nivalenol (NIV) were the most frequently detected toxins. For DON, 11% and for ZEA, 7% of all samples exceeded the European maximum limits for unprocessed cereals intended for human consumption. Furthermore, NIV was most likely produced by four different Fusarium species. A multiple correspondence analysis revealed that high levels of FG and DON were mainly observed in grain samples from fields with the previous crop maize, reduced tillage, cultivars with poor FHB resistance and strobilurin-based fungicides. Other previous crops and/or ploughing decreased the DON content by 78 to 95%. ZEA showed a similar pattern. In contrast, high levels of FP and NIV were associated with samples from ploughed fields and the previous crop canola. These findings and the negative correlations between FP DNA and FG incidence, ZEA and DON suggest a different ecological niche for FP or diverging requirements for growth and infection.