Effect of treatment with Lactococcus lactis NZ9000 on intestinal microbiota and mucosal immune responses against Clostridium perfringens in broiler chickens

Alterations in intestinal microbiota can modulate the developing avian intestinal immune system and, subsequently, may impact on resistance to enteric pathogens. The aim was to demonstrate that early life exposure to Lactococcus lactis, could affect either susceptibility or resistance of broilers to necrotic enteritis (NE). L. lactis NZ9000 (rL. lactis) pre-treatment at 1, 7, 14 and 21 days of age (DOA) led to a significant decrease in NE lesion scores in Clostridium perfringens infected chickens. C. perfringens Infection was associated with spatial and temporal decreases in mononuclear phagocytes and CD4+ αβ T cells. However, rL. Lactis pre-treatment and subsequent C. perfringens infection led to a significant increase in mononuclear phagocytes, CD8α + γδ T, αβ T cells (CD4+ and CD8α+) and B cells (IgM+, IgA+ and IgY+), as well as IL-12p40, IFN-γ and CD40. Differential expression of interleukin (IL)-6, IL-8, IL-10, IL-13, IL-18, IL-22, and transforming growth factor (TGF)-β were observed in L. lactis treated chickens when compared to C. perfringens infected chickens. Microbiota analysis in C. perfringens infected chickens demonstrated an increase in abundance of Bacillota, Bacteroidota, Pseudomonadota and Actinomycetota. These findings suggests that modulation of the chicken intestinal immune system by L. lactis confers partial protection 30 against NE

Identification of a Dual-Specific T Cell Epitope of the Hemagglutinin Antigen of an H5 Avian Influenza Virus in Chickens

Avian influenza viruses (AIV) of the H5N1 subtype have caused morbidity and mortality in humans. Although some migratory birds constitute the natural reservoir for this virus, chickens may play a role in transmission of the virus to humans. Despite the importance of avian species in transmission of AIV H5N1 to humans, very little is known about host immune system interactions with this virus in these species. The objective of the present study was to identify putative T cell epitopes of the hemagglutinin (HA) antigen of an H5 AIV in chickens. Using an overlapping peptide library covering the HA protein, we identified a 15-mer peptide, H5246–260, within the HA1 domain which induced activation of T cells in chickens immunized against the HA antigen of an H5 virus. Furthermore, H5246–260 epitope was found to be presented by both major histocompatibility complex (MHC) class I and II molecules, leading to activation of CD4+ and CD8+ T cell subsets, marked by proliferation and expression of interferon (IFN)-γ by both of these cell subsets as well as the expression of granzyme A by CD8+ T cells. This is the first report of a T cell epitope of AIV recognized by chicken T cells. Furthermore, this study extends the previous finding of the existence of dual-specific epitopes in other species to chickens. Taken together, these results elucidate some of the mechanisms of immune response to AIV in chickens and provide a platform for creation of rational vaccines against AIV in this species

Global, regional, and national incidence of six major immune-mediated inflammatory diseases : findings from the global burden of disease study 2019

DATA SHARING STATEMENT : Data used for the analyses are publicly available from the Institute of Health Metrics and Evaluation (http://www.healthdata.org/; http:// ghdx.healthdata.org/gbd-results-tool).BACKGROUND : The causes for immune-mediated inflammatory diseases (IMIDs) are diverse and the incidence trends of IMIDs from specific causes are rarely studied. The study aims to investigate the pattern and trend of IMIDs from 1990 to 2019. METHODS : We collected detailed information on six major causes of IMIDs, including asthma, inflammatory bowel disease, multiple sclerosis, rheumatoid arthritis, psoriasis, and atopic dermatitis, between 1990 and 2019, derived from the Global Burden of Disease study in 2019. The average annual percent change (AAPC) in number of incidents and age standardized incidence rate (ASR) on IMIDs, by sex, age, region, and causes, were calculated to quantify the temporal trends. FINDINGS : In 2019, rheumatoid arthritis, atopic dermatitis, asthma, multiple sclerosis, psoriasis, inflammatory bowel disease accounted 1.59%, 36.17%, 54.71%, 0.09%, 6.84%, 0.60% of overall new IMIDs cases, respectively. The ASR of IMIDs showed substantial regional and global variation with the highest in High SDI region, High-income North America, and United States of America. Throughout human lifespan, the age distribution of incident cases from six IMIDs was quite different. Globally, incident cases of IMIDs increased with an AAPC of 0.68 and the ASR decreased with an AAPC of −0.34 from 1990 to 2019. The incident cases increased across six IMIDs, the ASR of rheumatoid arthritis increased (0.21, 95% CI 0.18, 0.25), while the ASR of asthma (AAPC = −0.41), inflammatory bowel disease (AAPC = −0.72), multiple sclerosis (AAPC = −0.26), psoriasis (AAPC = −0.77), and atopic dermatitis (AAPC = −0.15) decreased. The ASR of overall and six individual IMID increased with SDI at regional and global level. Countries with higher ASR in 1990 experienced a more rapid decrease in ASR. INTERPRETATION : The incidence patterns of IMIDs varied considerably across the world. Innovative prevention and integrative management strategy are urgently needed to mitigate the increasing ASR of rheumatoid arthritis and upsurging new cases of other five IMIDs, respectively.The Global Burden of Disease Study is funded by the Bill and Melinda Gates Foundation. Support from Scientific Research Fund of Sichuan Academy of Medical Sciences & Sichuan Provincial People’s Hospital; Shaqra University; the School of Pharmacy, University of Botswana; the Indian Council of Medical Research (ICMR); an Australian National Health and Medical Research Council (NHMRC) Investigator Fellowship; the Italian Center of Precision Medicine and Chronic Inflammation in Milan; the Department of Environmental Health Engineering of Isfahan University of Medical Sciences, Isfahan, Iran; National Health and Medical Research Council (NHMRC), Australia; Jazan University, Saudi Arabia; the Clinician Scientist Program of the Clinician Scientist Academy (UMEA) of the University Hospital Essen; AIMST University, Malaysia; the Department of Community Medicine, Kasturba Medical College, Mangalore, Manipal Academy of Higher Education, Manipal, India; a Kornhauser Research Fellowship at The University of Sydney; the National Research, Development and Innovation Office Hungary; Taipei Medical University; CREATE Hope Scientific Fellowship from Lung Foundation Australia; the National Institute for Health and Care Research Manchester Biomedical Research Centre and an NIHR Clinical Lectureship in Respiratory Medicine; Kasturba Medical College, Mangalore and Manipal Academy of Higher Education, Manipal; Author Gate Publications; the Cleveland Clinic Foundation and Nassau University Medical center; the Italian Ministry of Health (RRC); King Abdulaziz University (DSR), Jeddah, and King Abdulaziz City for Science & Technology (KACSAT), Saudi Arabia, Science & Technology Development Fund (STDF), and US-Egypt Science & Technology joint Fund: The Academy of Scientific Research and Technology (ASRT), Egypt; partially supported by the Centre of Studies in Geography and Spatial Planning; the International Center of Medical Sciences Research (ICMSR), Islamabad Pakistan; Ain Shams University and the Egyptian Fulbright Mission Program; the Belgian American Educational Foundation; Health Data Research UK; the Spanish Ministry of Science and Innovation, Institute of Health Carlos III, CIBERSAM, and INCLIVA; the Clinical Research Development Unit, Imam Reza Hospital, Mashhad University of Medical Sciences; Shaqra University; Saveetha Institute of Medical and Technical Sciences and SRM Institute of Science and Technology; University of Agriculture, Faisalabad-Pakistan; the Chinese University of Hong Kong Research Committee Postdoctoral Fellowship Scheme; the institutional support of the Department of Microbiology and Immunology, Faculty of Pharmacy, Zagazig University, Egypt; the European (EU) and Developing Countries Clinical Trials Partnership, the EU Horizon 2020 Framework Programme, UK-National Institute for Health and Care Research, the Mahathir Science Award Foundation and EU-EDCTP.http://www.thelancet.comam2024School of Health Systems and Public Health (SHSPH)SDG-03:Good heatlh and well-bein

1α,25(OH)2 Vitamin D3 Modulates Avian T Lymphocyte Functions without Inducing CTL Unresponsiveness.

1,25-Dihydroxyvitamin D3 (Vitamin D) is a naturally synthesized fat soluble vitamin shown to have immunomodulatory, anti-inflammatory and cancer prevention properties in human and murine models. Here, we studied the effects of Vitamin D on the functional abilities of avian T lymphocytes using chicken Interferon (IFN)-γ ELISPOT assay, BrdU proliferation assay, Annexin V apoptosis assay and PhosFlow for detecting phosphorylated signalling molecules. The results demonstrate that Vitamin D significantly inhibited the abilities of T lymphocytes to produce IFN-γ and proliferate in vitro (P≤0.05), but retained their ability to undergo degranulation, which is a maker for cytotoxicity of these cells. Similarly, Vitamin D did not inhibit Extracellular signal-Regulated Kinase (ERK) 1/2 phosphorylation, a key mediator in T cell signalling, in the stimulated T lymphocytes population, while reduced ERK1/2 phosphorylation levels in the unstimulated cells. Our data provide evidence that Vitamin D has immuno-modulatory properties on chicken T lymphocytes without inducing unresponsiveness and by limiting immuno-pathology can promote protective immunity against infectious diseases of poultry

Vitamin D treatment does not induce cell death/apoptosis of chicken splenocytes.

<p>Chicken splenocytes were treated with Vitamin D (100nM) or vehicle (DMSO) for 4 hrs. The cells were stained with 7AAD (dead cell marker) and Annexin V (early apoptotic marker) and the data were analysed using flow cytometry. (<b>A</b>) Shows dot plots of the stained cells and the numbers in each quadrant represents percentage of cells. (<b>B</b>) The percentages (as mean ± SD) of live cells (Annexin V-7AAD- cells), apoptotic cells (Annexin V+7AAD- cells) and dead cells (Annexin V+7AAD+ cells) are shown from four independent experiments with three biological replicates in each experiment (12 biological replicates in total). (<b>C</b>) The data represent the percentage of live, apoptotic and dead cells 24hrs post Vitamin D (100 nM) or vehicle (DMSO) treatment. <b>(D)</b> The data represents the percentage of live, apoptotic, dead cells 3 days after Con A (10 μg/ml) stimulation.</p

Vitamin D limits ERK1/2 phosphorylation in the resting chicken CD3+ T cells.

<p>Following incubation of splenocytes with Vitamin D or vehicle only for 4hrs, phosphorylation of ERK1/2 (T202/Y204) was evaluated in CD3+ T cells in the resting cells (unstimulated) or cells stimulated with PMA for 5 minutes using flow cytometry. The data represents flow cytometry analysis of ERK 1/2 phosphorylation in cells treated with vehicle (thick line), Vitamin D (thin line) or Isotype control (grey area) in the resting cells and PMA-stimulated T cells.</p

Degranulation of chicken T cells is not influenced by Vitamin D.

<p>(<b>A</b>) Representative flow cytometry dot plots of spleen mononuclear cells that were cultured in medium containing Vitamin D (100nM or 10 nM) or vehicle only (DMSO) for 4 hours and were stimulated with a T-cell stimulation cocktail of PMA and Ion for an additional 4 hrs. The expression of CD107a in CD3+ and CD3- cells was analysed using flow cytometry. Upper right quadrant and lower right quadrant shows the percentages of CD107a+ cells within CD3+ and CD3- T cells, respectively. The bar graphs represents the percentages of CD3-CD107a+. (<b>B</b>) and CD3+CD107a+ cells (<b>C</b>) in the treated cells. The results are shown as mean ± SD of the study population of CD107a expression (<i>P</i> = NS indicates no statistical significance). † (symbol) represents the cells treated with the Vehicle only (DMSO). Similar data were obtained in four independent experiments.</p

Vitamin D inhibits chicken T-cell proliferation and IFN-γ production <i>ex vivo</i>.

<p>Splenocytes were pre-treated for 4hrs with Vitamin D (100 and 10 nM) or vehicle only (DMSO). (<b>A</b>) BrdU incorporation measured by ELISA assay to quantify T cell proliferation after stimulation with different concentrations of Concanavalin A. The results are presented as absorbance at OD₄₅₀ from three independent experiments with 3–4 biological replicates in each experiment. (B, C) The combined results from proliferation from 6 independent experiments for absorbance at OD₄₅₀ (B) and the percentages of inhibition by Vitamin D are shown. (<b>D</b>) The frequency of IFN-γ producing mononuclear cells stimulated with a T-cell stimulation cocktail of PMA and Ion was detected using a chicken IFN-γ ELISPOT assay. The results are presented as spots forming unit (SFU) per 1.0 x 10⁶ cells from five independent experiments with 3–5 biological replicates in each experiment (19 replicates in total). (<b>E</b>) Represents the percentage of inhibition for IFN-γ production by mononuclear cells after Vitamin D₃ pre-treatment from five independent experiments. Each dot represent a biological replicate. Non-parametric Wilcoxon tests (Mann-Whitney) was used to assess normal distribution and test significance. The results are shown as mean ± SD. † (symbol) represents cells treated with Vehicle only (DMSO). * indicates a statistically significant difference (<i>P</i> < 0.05).</p