Baicalin Protects Broilers against Avian Coronavirus Infection via Regulating Respiratory Tract Microbiota and Amino Acid Metabolism
Abstract
:1. Introduction
2. Results
2.1. Assessment of Anti-IBV Effect of Bai in CEK Cell
2.2. Bai Inhibited TLR7/MyD88/NF-κB Signalling Pathway and its Downstream Inflammatory Factors in CEK Cell
2.3. Observation of Clinical Signs and Follow up of the Changes in Body Weight
2.4. Histopathology and Scanning Electron Microscopy Analysis
2.5. Anti-IBV Antibody Level in Serum
2.6. Analysis of IBV and G3BP1 in Trachea
2.7. Effect of Bai on the TLR7/MyD88/NF-κB Signalling Pathway and Its Downstream Signal Protein in Trachea
2.8. Bai Corrected the Dysbiosis of Respiratory Flora Caused by IBV Infection
2.8.1. Alpha and Beta Diversity Analysis
2.8.2. The Regulatory Effect of Bai on the Abundance of Respiratory Microbiota
2.9. Bai Changed Metabolomics Changes in Trachea
2.10. Correlation Analysis of Differential Metabolites and Microbiota Composition
3. Discussion
4. Materials and Methods
4.1. Virus, Cell and Drugs
4.2. Animals
4.3. Cytotoxicity Assay
4.4. Antiviral Activity
4.5. Animal Experimentation
4.6. ELISA for Anti-IBV Antibodies
4.7. Histopathological Detection and Immunohistochemistry Analysis (IHC)
4.8. Scanning Electron Microscopy Analysis (SEM)
4.9. Real-Time RT-qPCR
4.10. Western Blot (WB) Analysis
4.11. Respiratory Microbial Structure Changes by 16S rDNA Analysis
4.12. Analysis and Identification of Metabolites
4.13. Statistical Analysis
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Conflicts of Interest
References
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Groups | Control | Bai | IBV | IBV + Bai | |
---|---|---|---|---|---|
Days | |||||
1 dpi | 108.72 ± 13.37 | 114.69 ± 12.24 | 106.00 ± 11.29 | 115.46 ± 12.17 | |
3 dpi | 140.65 ± 8.63 a | 123.49 ± 8.41 a | 137.15 ± 9.61 ab | 131.93 ± 9.50 ab | |
5 dpi | 151.56 ± 12.19 a | 152.47 ± 6.64 a | 128.52 ± 7.28 b | 143.69 ± 7.88 a |
Name | Dilution Ratio | kDa | Resource |
---|---|---|---|
IBV N | 1:1000 | 50 | Dayao Biotechnology (Hangzhou, China) |
IL-6 | 1:1000 | 24 | Cell Signaling Technology (Boston, MA, USA) |
IL-1β | 1:300 | 17 | WanLei Biotechnology (Shenyang, China) |
TNF-α | 1:1000 | 17–26 | SouthernBiotech (Birmingham, MD, USA) |
TLR7 | 1:1000 | 140 | Cell Signaling Technology (Boston, MA, USA) |
MyD88 | 1:1000 | 33 | Cell Signaling Technology (Boston, MA, USA) |
NF-κB p65 | 1:250 | 65 | Invitrogen (Carlsbad, CA, USA) |
NF-κB p-p65 | 1:1000 | 65 | Invitrogen (Carlsbad, CA, USA) |
IκB-α | 1:2000 | 36 | Proteintech (Chicago, IL, USA) |
p-IκB-α | 1:500 | 35 | Bioss (Beijing, China) |
G3BP1 (WB) | 1:1000 | 50 | Abmart (Shanghai, China) |
β-actin | 1:1000 | 40 | Abmart (Shanghai, China) |
G3BP1 (IHC) | 1:200 | — | Abmart (Shanghai, China) |
Goat Anti-Rabbit and Mouse IgG-HRP | 1:5000 | — | Abmart (Shanghai, China) |
Goat anti-Mouse IgG AF488 | 1:300 | — | Abmart (Shanghai, China) |
Gene | Primers (5′-3′) | Accession Number | Product Size (bp) |
---|---|---|---|
IBV N | F:GACGGAGGACCTGATGGTAA R: CCCTTCTTCTGCTGATCCTG | MK937830.1 | 206 |
IL-6 | F:GTTCGCCTTTCAGACCTACCTG R:ATCGGGATTTATCACCATCTGC | NM_20468.1 | 130 |
IL-1β | F: CCTTCGACATCTTCGACATCAA R: AATGTTGAGCCTCACTTTCTGG | NM_204524.1 | 113 |
TNF-α | F: TGCTGTTCTATGACCGCC R:CTTTCAGAGCATCAACGCA | AY765397 | 219 |
TLR7 | F:GCACACATTCAACTGGGGCAAAC R:TTCGGGGAACGGTAGTCAGAAGG | NM_001011688.3 | 115 |
MyD88 | F: CGGAACTTTTCGATGCCTTTAT R: CACACACAACTTAAGCCGATAG | NM_001030962.5 | 107 |
NF-κB p65 | F:ACCACCACCACCACAACACAATG R: GCGGCGTCGATGGTATCAAAGG | NM_001396038.1 | 116 |
IκB-α | F: GGATACCTGGCTGTTGTCGAATACC R: AAGTGTAGTGCTGTTCTCCCATTGC | NM_020529.3 | 89 |
G3BP1 | F: AGGGTGAACAAGGTGATGTGGAAAC R: GCCATAGCCTGCAAGAGAAGAGC | NM_001006150.2 | 149 |
β-actin | F:CCCAAAGCCAACAGAGAGAA R: CCATCACCAGAGTCCATCAC | NM_205518 | 140 |
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Feng, H.; Zhang, J.; Wang, X.; Guo, Z.; Wang, L.; Zhang, K.; Li, J. Baicalin Protects Broilers against Avian Coronavirus Infection via Regulating Respiratory Tract Microbiota and Amino Acid Metabolism. Int. J. Mol. Sci. 2024, 25, 2109. https://doi.org/10.3390/ijms25042109
Feng H, Zhang J, Wang X, Guo Z, Wang L, Zhang K, Li J. Baicalin Protects Broilers against Avian Coronavirus Infection via Regulating Respiratory Tract Microbiota and Amino Acid Metabolism. International Journal of Molecular Sciences. 2024; 25(4):2109. https://doi.org/10.3390/ijms25042109
Chicago/Turabian StyleFeng, Haipeng, Jingyan Zhang, Xuezhi Wang, Zhiting Guo, Lei Wang, Kang Zhang, and Jianxi Li. 2024. "Baicalin Protects Broilers against Avian Coronavirus Infection via Regulating Respiratory Tract Microbiota and Amino Acid Metabolism" International Journal of Molecular Sciences 25, no. 4: 2109. https://doi.org/10.3390/ijms25042109