Age-related burden and characteristics of embolic stroke of undetermined source in the real world clinical practice.

Few data are available on age-related burden and characteristics of embolic stroke of undetermined source (ESUS) in the real world clinical practice. The aim of our study was to provide information about it. We retrospectively analyzed data of patients consecutively admitted to our Stroke Unit along 1 year (2017, November 1st-2018, October 31st). The etiology of ischemic stroke was defined at hospital discharge; ESUS was considered as a subset of cryptogenic stroke, and defined according to the 2014 international criteria. In the analyzed period, 306 patients, 52.3% females, mean age ± SD 77.9 ± 11.9 years, were discharged with diagnosis of ischemic stroke. Ischemic strokes of cardioembolic and lacunar origin were the most frequent subtypes: 30.1% and 29.4%, respectively. Cardioembolic strokes were particularly frequent in patients ≥ 75 years, and almost always associated with atrial fibrillation. Overall, in 80 patients (26.1%) the etiology of stroke was undetermined; in 25 (8.2%) it remained undefined because of death or severe comorbidity, making further diagnostic work-up not worthy. Cryptogenic stroke occurred in 55 patients (18%), and ESUS criteria were satisfied in 39 of them (12.7%). According to age, cryptogenic stroke was diagnosed in 21.1% (21.1% ESUS) of patients < 65 years, 24.2% (19.4% ESUS) of patients aged 65-74 years, 15.5% (9.2% ESUS) of patients ≥ 75 years. After diagnostic work-up, patent foramen ovale was most commonly associated with ESUS (17.9%), especially in patients < 65 years (62.5%); covert paroxysmal atrial fibrillation was detected in 10.5% of ESUS patients ≥ 75 years. In the real world clinical practice, the frequency of ischemic strokes of undetermined etiology, and of those satisfying ESUS criteria, is not negligible, especially in younger patients. A thorough diagnostic work-up, with an age-specific approach, is therefore necessary and of the utmost importance for the identification of stroke etiology, in order to optimize secondary stroke prevention strategies.

Preparation of Powdered Infant Formula: Could Product's Safety Be Improved?

The recent outbreak of Salmonella Agona linked to the consumption of infant formula (powdered formula) has rekindled the attention about the correct procedures for preparation and use of these products. International guidelines have already been published so far, particularly in association with Cronobacter sakazakii in early 2000s. FAO/WHO suggested to reconstitute formula with water at no less than 70°C. We therefore contaminated powdered formula with low levels of Salmonella spp and C sakazakii to evaluate the pathogens inactivation during the formula preparation using water at 70°C. In these conditions we observed a survival of both pathogens, indicating that the suggested recommendations may be not enough to guarantee the safety of this product. Higher temperatures are needed to reduce the biological risk, even if it may be not easily realized in actual domestic conditions. Moreover, the impact on the nutritional value of reconstituted formulas should be evaluated.

Transformation and Tumorigenicity Testing of Simian Cell Lines and Evaluation of Poliovirus Replication.

The key role of cell cultures in different scientific fields is worldwide recognized, both as in vitro research models alternative to laboratory animals and substrates for biological production. However, many safety concerns rise from the use of animal/human cell lines that may be tumorigenic, leading to potential adverse contaminations in cell-derived biologicals. In order to evaluate the suitability of 13 different cell lines for Poliovirus vaccine production, safety and quality, in vitro/in vivo tumorigenicity and Poliovirus propagation properties were evaluated. Our results revealed that non-human primate cell lines CYNOM-K1, FRhK-4, 4MBr-5 and 4647 are free of tumorigenic features and represent highly susceptible substrates for attenuated Sabin Poliovirus strains. In particular, FRhK-4 and 4647 cell lines are characterized by a higher in vitro replication, resulting indicated for the use in large-scale production field.

Protein mutations following adaptation of avian influenza viruses in different biological systems.

Traditionally, embryonated chicken eggs (ECE) are considered the gold standard for Influenza virus isolation and vaccine production. Nowadays, different biological systems have been improved and performed, in order to evaluate a feasible alternative to ECE. In fact, in a previous study, mammalian and avian cell cultures were successfully used for avian influenza viruses primary isolation from target tissues and virus propagation. This research is focused on the investigation of adaptive mutations that occur after influenza A virus amplification in ECE and cell cultures. The results of the study shows that avian influenza viruses after multiple passages in different biological systems undergo mutations, in particular, the largest number of amino acid substitutions occurred in all biological substrates in the hemagglutinin.

Veterinary biobank facility: development and management for diagnostic and research purposes.

Biobanking is an essential tool for ensuring easy availability of high-quality biomaterial collections that combine essential samples and epidemiological, clinical, and research data for the scientific community. Specimen collection is an integral part of clinical research. Indeed, every year throughout the world, millions of biological samples are stored for diagnostics and research, but in many fields the lack of biological material and models is a major hindrance for ongoing research. A biobank facility provides suitable samples for large-scale screening studies and database repositories. Software dedicated to biological banks simplify sample registration and identification, the cataloging of sample properties (type of sample/specimen, associated diseases and/or therapeutic protocols, environmental information, etc.), sample tracking, quality assurance, and specimen availability characterized by well-defined features. Biobank facilities must adopt good laboratory practices (GLPs) and a stringent quality control system and also comply with ethical issues, when required. The creation of a veterinary network can be useful under different aspects: the first one is related to the importance of animal sciences itself to improve research and strategies in the different branches of the veterinary area, and the second aspect is related to the possibility of data management harmonization to improve scientific cooperation.

Exploring the larval transcriptome of the common sole (Solea solea L.).

BACKGROUND: The common sole (Solea solea) is a promising candidate for European aquaculture; however, the limited knowledge of the physiological mechanisms underlying larval development in this species has hampered the establishment of successful flatfish aquaculture. Although the fact that genomic tools and resources are available for some flatfish species, common sole genomics remains a mostly unexplored field. Here, we report, for the first time, the sequencing and characterisation of the transcriptome of S. solea and its application for the study of molecular mechanisms underlying physiological and morphological changes during larval-to-juvenile transition. RESULTS: The S. solea transcriptome was generated from whole larvae and adult tissues using the Roche 454 platform. The assembly process produced a set of 22,223 Isotigs with an average size of 726 nt, 29 contigs and a total of 203,692 singletons. Of the assembled sequences, 75.2% were annotated with at least one known transcript/protein; these transcripts were then used to develop a custom oligo-DNA microarray. A total of 14,674 oligonucleotide probes (60 nt), representing 12,836 transcripts, were in situ synthesised onto the array using Agilent non-contact ink-jet technology. The microarray platform was used to investigate the gene expression profiles of sole larvae from hatching to the juvenile form. Genes involved in the ontogenesis of the visual system are up-regulated during the early stages of larval development, while muscle development and anaerobic energy pathways increase in expression over time. The gene expression profiles of key transcripts of the thyroid hormones (TH) cascade and the temporal regulation of the GH/IGF1 (growth hormone/insulin-like growth factor I) system suggest a pivotal role of these pathways in fish growth and initiation of metamorphosis. Pre-metamorphic larvae display a distinctive transcriptomic landscape compared to previous and later stages. Our findings highlighted the up-regulation of gene pathways involved in the development of the gastrointestinal system as well as biological processes related to folic acid and retinol metabolism. Additional evidence led to the formation of the hypothesis that molecular mechanisms of cell motility and ECM adhesion may play a role in tissue rearrangement during common sole metamorphosis. CONCLUSIONS: Next-generation sequencing provided a good representation of the sole transcriptome, and the combination of different approaches led to the annotation of a high number of transcripts. The construction of a microarray platform for the characterisation of the larval sole transcriptome permitted the definition of the main processes involved in organogenesis and larval growth.

Expression of interleukin-1ß, interleukin-8, and interferon-γ in blood samples obtained from healthy and sick neonatal foals.

OBJECTIVE: To evaluate and compare the gene expression of interleukin(IL)-1ß, IL-8, and interferon-γ during the first 72 hours after birth in healthy foals and during the first 72 hours after hospitalization in sick neonatal foals and investigate correlations of clinicopathologic variables with cytokine expressions in healthy and sick neonatal foals. ANIMALS: 33 foals < 7 days old (10 healthy foals, 7 foals with sepsis, 6 foals with peripartum asphyxia syndrome, and 12 foals with other diseases [2 with failure of passive transfer of immunity only were not further evaluated]). PROCEDURES: A blood sample (15 mL) was collected from each foal immediately after birth or hospital admission (0 hours) and at 24 and 72 hours later. Clinicopathologic variables were evaluated, and cytokine gene expression in WBCs was measured with an absolute quantitative real-time reverse transcriptase PCR assay. RESULTS: At all time points, gene expression of interferon-γ was low in all groups. No time-dependent changes in cytokine expressions were detected in healthy or sick foals. Foals with sepsis had significantly higher IL-1ß gene expression than did healthy foals, foals with peripartum asphyxia syndrome, or foals with other diseases. At 0 hours, IL-1ß expression was correlated with plasma fibrinogen concentration in healthy foals and with the neutrophil-to-lymphocyte ratio in foals with sepsis; IL-8 expression was correlated with monocyte count in foals with sepsis and with arterial pH, plasma fibrinogen concentration, and plasma lactate concentration in foals with peripartum asphyxia syndrome. CONCLUSIONS AND CLINICAL RELEVANCE: Data have suggested that evaluation of IL-1ß expression in sick neonatal foals could help identify those with sepsis.

Managing behavioural problems in human-dog interactions.

The management of dog behavioural problems requires the expertise of professionals such as the veterinary behaviourist. Clinical assessment of behavioural disorders allows the veterinary behaviourist to formulate a diagnosis and prescribe a behavioural and/or pharmacological therapy. The objective of such therapy is to produce a stable change in the perception of a stimulus and the resulting emotion, leading to the correction of the behavioural problem. It may be crucial to evaluate the subject's pathological state in response to the observed symptoms in order to identify the functional impairment of the pivotal neurotransmitter systems involved in the disorder. This allows selecting a suitable pharmacological treatment. In order to implement behavioural therapy, the veterinary behaviourist collaborates, where necessary, with a team of qualified canine trainers.

Real-time quantitative PCR using hairpin-shaped clone-specific primers for minimal residual disease assessment in an animal model of human non-Hodgkin lymphoma.

A multitude of molecular techniques for monitoring minimal residual disease in lymphoproliferative disorders have been described to date. Real-Time Quantitative PCR targeting Immunoglobulin Heavy chain patient-specific sequences is increasingly being used for molecular detection of residual neoplastic B-cells using allele-specific oligos. The establishment of individually tailored PCR assays with the extensive use of patient-specific fluorescent-labeled oligos may be cumbersome and expensive. The present study was aimed at evaluating the usefulness of recently described hairpin-shaped allele-specific primers, originally intended for typing single-nucleotide polymorphisms, for the assessment of minimal residual disease using SYBR Green intercalating dye. Three cloned and 2 sequenced clonogenic Ig heavy chain rearranged gene loci, obtained from 5 cases of canine spontaneous B-cell lymphoma, were used as an experimental model. Both standard linear and hairpin-shaped forward and reverse clone-specific primers were evaluated in terms of specificity, sensitivity and PCR efficiency. Hairpin-shaped primers were demonstrated to have achieved accurate results more consistently than the respective linear primers allowing the specific and sensitive quantification of minimal residual disease of lymphoproliferative disorders with fewer validation procedures and more flexibility on the assay design.

Complete sequencing of full-length canine ataxia telangiectasia mutated mRNA and characterization of its putative promoter.

Ataxia telangiectasia mutated (ATM) protein is considered a "caretaker" of the genome integrity and a defective ATM has been correlated with increased cancer risk in human beings. In an effort to explore the reliability of dog as a spontaneous animal model of genetic susceptibility to lymphoid malignancies, we have carried out the complete sequencing of the canine ATM mRNA. 5' RACE analysis and sequencing were used to obtain the full-length canine cDNA. The transcription start site was found at CFA5: 27307661 (Dog Genome assembly 2.0, release 49). Two exons were found in the 5'UTR. A putative TATA-less bi-directional promoter region was found in the region 5' upstream of the cap site. The core promoter harbours different conserved regulatory motifs: CREB, CCAAT boxes (NF-binding sites), Sp1, AP-2, GCF, XRE, Ets, Cre and c-Myb. The major ORF, corresponding to the ortholog human and pig ATM isoform 1, has 64 exons and codes a protein of 3056 aa. The homology between dog and human ATM at the aa level was 89% identities-93% positives, even higher than the homology between pig and human. When compared with the canine genomic sequences, 3 sequence variants yielding to aa substitution were found. Canine ATM is highly conserved and may represent a candidate gene to evaluate lymphoid malignancies predisposition in dogs.

Effect of passive transfer status on preweaning growth performance in dairy lambs.

OBJECTIVE: To evaluate the effect of passive transfer status, determined by measuring serum IgG concentration 24 hours after parturition, on preweaning growth performance in dairy lambs. DESIGN: Prospective observational study. ANIMALS: 20 healthy Sardinian dairy lambs. PROCEDURES: Serum IgG concentration was measured 24 hours after birth. Body weight was measured at birth and at the time of weaning 28 days (ie, 27 to 29 days) after birth. Mean daily gain from birth to day 28 and day 28 weight were used as measures of preweaning growth performance. Regression analysis was used to evaluate associations between serum IgG concentration 24 hours after birth and measures of preweaning growth performance. RESULTS: Mean +/- SD serum IgG concentration 24 hours after birth was 24.6 +/- 17.5 mg/mL. Mean body weights at birth and weaning were 2,696 +/- 937 g and 9,253 +/- 2,116 g, respectively, and mean daily gain was 234 +/- 63 g/d. No significant association was detected between serum IgG concentration 24 hours after birth and birth weight. However, serum IgG concentration 24 hours after birth was significantly associated with mean daily gain (R(2) = 0.25). Each 1 mg/mL increase in serum IgG concentration 24 hours after birth was associated with a 1.8 g/d increase in mean daily gain and a 60.8-g increase in day 28 weight. CONCLUSIONS AND CLINICAL RELEVANCE: Results indicated that passive transfer status, determined as serum IgG concentration 24 hours after birth, was a significant source of variation in preweaning growth performance in dairy lambs.