Phylogenetic and molecular characterization of equine H3N8 influenza viruses from Greece (2003 and 2007): Evidence for reassortment between evolutionary lineages

<p>Abstract</p> <p>Background</p> <p>For first time in Greece equine influenza virus infection was confirmed, by isolation and molecular analysis, as the cause of clinical respiratory disease among unvaccinated horses during 2003 and 2007 outbreaks.</p> <p>Methods</p> <p>Equine influenza virus (EIV) H3N8 was isolated in MDCK cells from 30 nasal swabs from horses with acute respiratory disease, which were tested positive by Directigen Flu A. Isolation was confirmed by haemagglutination assay and RT-PCR assay of the M, HA and NA gene.</p> <p>Results</p> <p>HA sequences of the Greek isolates appeared to be more closely related to viruses isolated in early 1990s in Europe. These results suggested that viruses with fewer changes than those on the main evolutionary lineage may continue to circulate. On the other hand, analysis of deduced NA amino acid sequences were more closely related to viruses isolated in outbreaks in Europe and Asia during 2003-2007. Phylogenetic analysis characterized the Greek isolates as a member of the Eurasian lineage by the haemagglutinin (HA) protein alignment, but appeared to be a member of the Florida sublineage clade 2 by the neuraminidase (NA) protein sequence suggesting that reassortment might be a possible explanation.</p> <p>Conclusion</p> <p>Our findings suggest that the Greek strains represent an example of "frozen evolution" and probably reassortment between genetically distinct co-circulated strains. Therefore expanding current equine influenza surveillance efforts is a necessity.</p

Report of the FELASA-EFAT Working Group

Publisher Copyright: © The Author(s) 2023.Competent, confident and caring laboratory animal caretakers, technicians and technologists (LAS staff) are vital for good animal welfare, high-quality science and a secure Culture of Care. This requires high-quality education, training, supervision and continuing professional development (CPD) of LAS staff. However, there is a lack of harmonisation regarding how this education and training is conducted among European countries, and nor are there recommendations adapted to Directive 2010/63/EU. Therefore, FELASA and EFAT established a working group with the task of establishing recommendations for education, training and CPD for LAS staff. The working group established five different levels (LAS staff levels 0–4), defining the required level of competence and attitude, as well as suggesting educational requirements for reaching each level. Defining these levels should help to ensure that appropriate educational and CPD activities are in place, and to enable employers and LAS staff to determine the level and career stage attained. Furthermore, proper assessment of competencies and effective CPD schemes for all relevant staff should be established. Regulators should support this by setting standards for competence assessment and ensuring that they are consistently applied. In addition, establishments should involve the LAS staff in defining and developing the Culture of Care. The Animal Welfare Body should be involved and have oversight of education, training and CPD. These recommendations will contribute to harmonisation and increased quality of education, training and CPD, as well as provide clearer career pathways for LAS staff, helping to ensure high standards of animal welfare and science.publishersversionepub_ahead_of_prin

The RNA binding protein HuR is a gatekeeper of liver homeostasis

BACKGROUND AND AIMS: Non-alcoholic fatty liver disease (NAFLD) is initiated by steatosis and can progress via fibrosis and cirrhosis to hepatocellular carcinoma (HCC). The RNA binding protein HuR controls RNAs at the posttranscriptional level; hepatocyte HuR has been implicated in the regulation of diet-induced hepatic steatosis. The present study aimed to understand the role of hepatocyte-HuR in NAFLD development and progression to fibrosis and HCC. APPROACH AND RESULTS: Hepatocyte-specific HuR-deficient mice and control HuR-sufficient mice were fed either a normal diet or a NAFLD-inducing diet. Hepatic lipid accumulation, inflammation, fibrosis and HCC development were studied by histology, flow cytometry, quantitative PCR and RNA sequencing. The liver lipidome was characterized by lipidomics analysis and the HuR-RNA interactions in the liver were mapped by RNA immunoprecipitation-sequencing. Hepatocyte-specific HuR-deficient mice displayed spontaneous hepatic steatosis and fibrosis predisposition, compared to control HuR-sufficient mice. On a NAFLD-inducing diet, hepatocyte-specific HuR-deficiency resulted in exacerbated inflammation, fibrosis and HCC-like tumor development. A multi-omic approach, including lipidomics, transcriptomics and RNA-immunoprecipitation sequencing revealed that HuR orchestrates a protective network of hepatic-metabolic and lipid homeostasis-maintaining pathways. Consistently, HuR-deficient livers accumulated, already at steady-state, a triglyceride signature resembling that of NAFLD livers. Moreover, upregulation of Spp1 and its product osteopontin mediated, at least partially, the fibrosis development in hepatocyte-specific HuR deficiency on a NAFLD-inducing diet, as shown by experiments utilizing antibody blockade of osteopontin. CONCLUSIONS: HuR is a gatekeeper of liver homeostasis preventing NAFLD-related fibrosis and HCC, suggesting that the HuR-dependent network could be exploited therapeutically

Outbreak of Canine Norovirus Infection in Young Dogs ▿

An outbreak of norovirus (NoV) infection was identified in a kennel. Sequence analysis of a short fragment in the polymerase complex indicated the clonal origin of the strains, which were similar to the prototype canine NoV strain GIV.2/Bari/170/07-4/ITA (94.7% nucleotide identity). The findings demonstrate that canine NoV circulates in dogs in Greece and that it can spread easily across a group of animals

PIK3Cd expression by fibroblasts promotes triple-negative breast cancer progression

As there is growing evidence for the tumor microenvironment’s (TME) role in tumorigenesis, we investigated the role of fibroblast-expressed kinases in triple negative breast cancer (TNBC). Using a high-throughput kinome screen combined with 3D invasion assays, we identified fibroblast-expressed PIK3Cd (f-PIK3Cd) as a key regulator of progression. Although PIK3Cd was expressed in primary fibroblasts derived from TNBC patients, it was undetectable in breast cancer cell lines. Genetic and pharmacologic gain- and loss-of functions experiments verified the contribution of f-PIK3Cd in TNBC cell invasion. Integrated secretomics and transcriptomics analyses revealed a paracrine mechanism via which f-PIK3Cd confers its pro-tumorigenic effects. Inhibition of f-PIK3Cd promoted the secretion of factors, including PLGF and BDNF, which led to upregulation of NR4A1 in TNBC cells where it acts as a tumor suppressor. Inhibition of PIK3Cd in an orthotopic BC mouse model reduced tumor growth only after inoculation with fibroblasts, indicating a role of f-PIK3Cd in cancer progression. Similar results were observed in the MMTV-PyMT transgenic BC mouse model, along with a decrease on tumor metastasis emphasizing the potential immune-independent effects of PIK3Cd inhibition. Finally, analysis of BC patient cohorts and TCGA datasets identified f-PIK3Cd (protein and mRNA levels) as an independent prognostic factor for overall and disease free survival, highlighting it as a therapeutic target for TNBC

INFRAFRONTIER quality principles in systemic phenotyping

International audienceAbstract Improving reproducibility and replicability in preclinical research is a widely discussed and pertinent topic, especially regarding ethical responsibility in animal research. INFRAFRONTIER, the European Research Infrastructure for the generation, phenotyping, archiving, and distribution of model mammalian genomes, is addressing this issue by developing internal quality principles for its different service areas, that provides a quality framework for its operational activities. This article introduces the INFRAFRONTIER Quality Principles in Systemic Phenotyping of genetically altered mouse models. A total of 11 key principles are included, ranging from general requirements for compliance with guidelines on animal testing, to the need for well-trained personnel and more specific standards such as the exchange of reference lines. Recently established requirements such as the provision of FAIR (Findable, Accessible, Interoperable, Reusable) data are also addressed. For each quality principle, we have outlined the specific context, requirements, further recommendations, and key references

PIK3Cδ expression by fibroblasts promotes triple-negative breast cancer progression

As there is growing evidence for the tumor microenvironment’s (TME) role in tumorigenesis, we investigated the role of fibroblast-expressed kinases in triple negative breast cancer (TNBC). Using a high-throughput kinome screen combined with 3D invasion assays, we identified fibroblast-expressed PIK3Cδ (f-PIK3Cδ) as a key regulator of progression. Although PIK3Cδ was expressed in primary fibroblasts derived from TNBC patients, it was undetectable in breast cancer cell lines. Genetic and pharmacologic gain- and loss-of functions experiments verified the contribution of f-PIK3Cδ in TNBC cell invasion. Integrated secretomics and transcriptomics analyses revealed a paracrine mechanism via which f-PIK3Cδ confers its pro-tumorigenic effects. Inhibition of f-PIK3Cδ promoted the secretion of factors, including PLGF and BDNF, which led to upregulation of NR4A1 in TNBC cells where it acts as a tumor suppressor. Inhibition of PIK3Cδ in an orthotopic BC mouse model reduced tumor growth only after inoculation with fibroblasts, indicating a role of f-PIK3Cδ in cancer progression. Similar results were observed in the MMTV-PyMT transgenic BC mouse model, along with a decrease on tumor metastasis emphasizing the potential immune-independent effects of PIK3Cδ inhibition. Finally, analysis of BC patient cohorts and TCGA datasets identified f-PIK3Cδ (protein and mRNA levels) as an independent prognostic factor for overall and disease free survival, highlighting it as a therapeutic target for TNBC