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      Dietary Eugenol Nanoemulsion Potentiated Performance of Broiler Chickens: Orchestration of Digestive Enzymes, Intestinal Barrier Functions and Cytokines Related Gene Expression With a Consequence of Attenuating the Severity of E. coli O78 Infection

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          Abstract

          Recently, the use of essential oils (EOs) or their bioactive compounds encapsulated by nanoparticles as alternative supplements for in-feed antimicrobials is gaining attention, especially in organic poultry production. Focusing on eugenol, its incorporation into the nanoformulation is a novel strategy to improve its stability and bioavailability and thus augment its growth-boosting and antimicrobial activities. Therefore, we explored eugenol nanoemulsion activities in modulating growth, digestive and gut barrier functions, immunity, cecal microbiota, and broilers response to avian pathogenic E. coli challenge (APEC) O78. A total of 1,000 one-day-old broiler chicks were allocated into five groups; negative control (NC, fed basal diet), positive control (PC), and 100, 250, and 400 mg/kg eugenol nanoemulsion supplemented groups. All groups except NC were challenged with APEC O78 at 14 days of age. The results showed that birds fed eugenol nanoemulsion displayed higher BWG, FI, and survivability and most improved FCR over the whole rearing period. Birds fed 400 mg/kg of eugenol nanoemulsion sustained a higher growth rate (24% vs. PC) after infection. Likely, the expression of digestive enzymes' genes ( AMY2A, CCK, CELA1, and PNLIP) was more prominently upregulated and unaffected by APEC O78 challenge in the group fed eugenol nanoemulsion at the level of 400 mg/kg. Enhanced gut barrier integrity was sustained post-challenge in the group supplemented with higher levels of eugenol nanoemulsion as evidenced by the overexpression of cathelicidins-2, β-defensin-1, MUC-2, JAM-2, occludin, CLDN-1, and FABP-2 genes. A distinct modulatory effect of dietary eugenol nanoemulsion was observed on cytokine genes (IL-1β, TNF-α, IL-6, IL-8, and IL-10) expression with a prominent reduction in the excessive inflammatory reactions post-challenge. Supplementing eugenol nanoemulsion increased the relative cecal abundance of Lactobacillus species and reduced Enterobacteriaceae and Bacteriods counts. Notably, a prominent reduction in APEC O78 loads with downregulation of papC, iroN, iutA, and iss virulence genes and detrimental modifications in E. coli morphological features were noticed in the 400 mg/kg eugenol nanoemulsion group at the 3rd-week post-challenge. Collectively, we recommend the use of eugenol nanoemulsion as a prospective targeted delivery approach for achieving maximum broilers growth and protection against APEC O78 infection.

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          Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) Method.

          The two most commonly used methods to analyze data from real-time, quantitative PCR experiments are absolute quantification and relative quantification. Absolute quantification determines the input copy number, usually by relating the PCR signal to a standard curve. Relative quantification relates the PCR signal of the target transcript in a treatment group to that of another sample such as an untreated control. The 2(-Delta Delta C(T)) method is a convenient way to analyze the relative changes in gene expression from real-time quantitative PCR experiments. The purpose of this report is to present the derivation, assumptions, and applications of the 2(-Delta Delta C(T)) method. In addition, we present the derivation and applications of two variations of the 2(-Delta Delta C(T)) method that may be useful in the analysis of real-time, quantitative PCR data. Copyright 2001 Elsevier Science (USA).
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            The antimicrobial activity of nanoparticles: present situation and prospects for the future

            Nanoparticles (NPs) are increasingly used to target bacteria as an alternative to antibiotics. Nanotechnology may be particularly advantageous in treating bacterial infections. Examples include the utilization of NPs in antibacterial coatings for implantable devices and medicinal materials to prevent infection and promote wound healing, in antibiotic delivery systems to treat disease, in bacterial detection systems to generate microbial diagnostics, and in antibacterial vaccines to control bacterial infections. The antibacterial mechanisms of NPs are poorly understood, but the currently accepted mechanisms include oxidative stress induction, metal ion release, and non-oxidative mechanisms. The multiple simultaneous mechanisms of action against microbes would require multiple simultaneous gene mutations in the same bacterial cell for antibacterial resistance to develop; therefore, it is difficult for bacterial cells to become resistant to NPs. In this review, we discuss the antibacterial mechanisms of NPs against bacteria and the factors that are involved. The limitations of current research are also discussed.
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              Antimicrobial properties of chitosan and mode of action: a state of the art review.

              Owing to its high biodegradability, and nontoxicity and antimicrobial properties, chitosan is widely-used as an antimicrobial agent either alone or blended with other natural polymers. To broaden chitosan's antimicrobial applicability, comprehensive knowledge of its activity is necessary. The paper reviews the current trend of investigation on antimicrobial activities of chitosan and its mode of action. Chitosan-mediated inhibition is affected by several factors can be classified into four types as intrinsic, environmental, microorganism and physical state, according to their respective roles. In this review, different physical states are comparatively discussed. Mode of antimicrobial action is discussed in parts of the active compound (chitosan) and the target (microorganisms) collectively and independently in same complex. Finally, the general antimicrobial applications of chitosan and perspectives about future studies in this field are considered. Copyright © 2010 Elsevier B.V. All rights reserved.
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                Author and article information

                Contributors
                Journal
                Front Vet Sci
                Front Vet Sci
                Front. Vet. Sci.
                Frontiers in Veterinary Science
                Frontiers Media S.A.
                2297-1769
                23 June 2022
                2022
                : 9
                : 847580
                Affiliations
                [1] 1Department of Nutrition and Clinical Nutrition, Faculty of Veterinary Medicine, Zagazig University , Zagazig, Egypt
                [2] 2Department of Hygiene and Zoonoses, Faculty of Veterinary Medicine, Mansoura University , Mansoura, Egypt
                [3] 3Department of Pharmacology, Faculty of Veterinary Medicine, Aswan University , Aswan, Egypt
                [4] 4Veterinary Educational Hospital, Faculty of Veterinary Medicine, Zagazig University , Zagazig, Egypt
                [5] 5Department of Pathology and Clinical Pathology, Zagazig Branch, Agriculture Research Center, Animal Health Research Institute , Zagazig, Egypt
                [6] 6Department of Clinical Laboratory Sciences, Turabah University College, Taif University , Taif, Saudi Arabia
                [7] 7Department of Biochemistry, Zagazig Branch, Agriculture Research Center, Animal Health Research Institute , Zagazig, Egypt
                [8] 8Department of Bacteriology, Zagazig Branch, Agriculture Research Center, Animal Health Research Institute , Zagazig, Egypt
                [9] 9Department of Clinical Pathology, Faculty of Veterinary Medicine, Zagazig University , Zagazig, Egypt
                [10] 10Department of Microbiology, Faculty of Veterinary Medicine, Zagazig University , Zagazig, Egypt
                Author notes

                Edited by: Francesco Gai, Italian National Research Council, Italy

                Reviewed by: Hamada A. M Elwan, Minia University, Egypt; Jana Koscova, University of Veterinary Medicine and Pharmacy in Košice, Slovakia

                *Correspondence: Doaa Ibrahim mero_micro2006@ 123456yahoo.com
                Marwa I. Abd El-Hamid doibrahim@ 123456vet.zu.edu.eg

                This article was submitted to Animal Nutrition and Metabolism, a section of the journal Frontiers in Veterinary Science

                Article
                10.3389/fvets.2022.847580
                9260043
                35812892
                1c9fe634-3b9d-49d6-bf1c-b76a61c50fc4
                Copyright © 2022 Ibrahim, Eldemery, Metwally, Abd-Allah, Mohamed, Ismail, Hamed, Al Sadik, Neamat-Allah and Abd El-Hamid.

                This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

                History
                : 02 January 2022
                : 05 April 2022
                Page count
                Figures: 7, Tables: 6, Equations: 0, References: 111, Pages: 17, Words: 13886
                Categories
                Veterinary Science
                Original Research

                broiler chickens,eugenol nanoemulsion,performance,immunity,barrier function,apec o78,virulence gene expression

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