Probiotic as Adjuvant Significantly Improves Protection of the Lanzhou Trivalent Rotavirus Vaccine against Heterologous Challenge in a Gnotobiotic Pig Model of Human Rotavirus Infection and Disease
Abstract
:1. Introduction
2. Materials and Methods
2.1. Vaccine
2.2. Probiotic Bacteria Used as Vaccine Adjuvant
2.3. Challenge Virus
2.4. Experimental Design: Gn Pig Inoculation, Challenge, and Sample Collection
2.5. Clinical Signs and Rotavirus and Probiotic Bacteria Shedding
2.6. Shedding of Vaccine Virus by RT-PCR Typing Primer for G2, G3, and G4
2.7. Extraction of Mononuclear Cells (MNCs)
2.8. Preparation of Sf9 Cell Plates Expressing Rotavirus D VP7 (G1), DS1 VP7 (G2), PV P7 (G3), and ST3 VP7 (G4)
2.9. Immunocytochemical Staining Assay for Detection of Rotavirus G1, G2, G3, and G4 VP7 G-Type-Specific IgA and IgG Antibody Responses in Serum
2.10. ELISPOT Assay for Rotavirus G1, G2, G3, G4 VP7 G-Type-Specific ASC
2.11. Intracellular Cytokine Staining and Flow Cytometry Analysis of IFN-γ-Producing CD4+ and CD8+ T Cells
2.12. Statistical Analysis
3. Results
3.1. Vaccine Replication in Gn Pigs: Shedding of G3 and G4 Reassortant Strains was Detected after Oral Inoculations with TLV+LGG
3.2. LGG Enhanced TLV Vaccine’s Protection against Heterotypic Virulent Wa HRV Challenge
3.3. The TLV Induced Intestinal VP7-Specific IgA and IgG ASC Responses to Homotypic and Heterotypic VP7 G-Types; The Presence of LGG Promoted Stronger Intestinal Vaccine Antigen-Specific ASC Responses
3.4. LGG Adjuvant Enhanced VP7 G-Type Specific IgA and IgG Antibody Responses in Serum Pre- and Postchallenge, including the Cross-Reactive Response to the Heterotypic G1 HRV Challenge Strain
3.5. The TLV Primed for Strong Anamnestic Serum VN Antibody Responses to Heterotypic G1P[8] HRV Post-Challenge with or without LGG
3.6. IFN-γ-Producing CD4+ and CD8+ T Cell Responses in Intestinal and Systemic Lymphoid Tissues
3.6.1. Both LGG and TLV Alone Induced IFN-γ-Producing CD4+ and CD8+ T-Cell Responses in Intestinal and Systemic Lymphoid Tissues
3.6.2. The TLV Induced Heterotypic G1- and Homotypic G2-, G3- and G4-Specific IFN-γ-Producing CD4+ and CD8+ T-Cell Responses in Intestinal and Systemic Lymphoid Tissues Pre- and Postchallenge
3.6.3. The LGG Modulates IFN-γ-Producing CD4+ and CD8+ T-Cell Responses Differentially in Intestinal and Systemic Lymphoid Tissues
4. Discussion
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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Primers | Name | Sequence (5′→3′) |
---|---|---|
Primers for VP7 gene reverse transcription | VP7-F | tct ggc taa cgg tta gct cc |
VP7-R | aat act tgc cac cat ttt ttc c | |
Specific primers for G2 | G2-F | caa tga tat taa cac att ttc tgt |
G2-R | aat act tgc cac cat ttt ttc c | |
Specific primers G3 | G3-F | ctt ttg aag aag ttg cga cag |
G3-R | aat act tgc cac cat ttt ttc c | |
Specific primers for G4 | G4-F | cgc ttc tgg tga gga gtt g |
G4-R | aat act tgc cac cat ttt ttc c |
Treatments | n | Clinical Signs (Diarrhea) | Fecal Virus Shedding | |||||||
---|---|---|---|---|---|---|---|---|---|---|
% with Diarrhea *,a | Mean Days to Onset **, b | Mean Duration Days ** | Mean AUC of Diarrhea Score c | Mean Cumulative Score ** | % Virus Shedding * | Mean Days to Onset **,b | Mean Duration Days ** | Mean AUC of Virus Shedding d | ||
TLV+LGG | 3 | 100 A | 4.7 (1.7) fA | 1.3 (0.7) B | 7.7 (0.8) C | 9.3 (0.8) B | 100 A | 2.0 (0.3) A | 1.0 (0) B | 2.2 (0.2) B |
TLV | 3 | 67 A | 1.3 (0.3) B | 5.0(1.2) A | 11.3 (1.1) AB | 12.8 (1.6) A | 100 A | 1.7 (0.3) A | 3.3 (0.7) AB | 1.9 (0.9) B |
LGG | 4 | 100 A | 1.8 (0.8) B | 3.8 (0.9) A | 9.9 (0.7) B | 11.9 (0.8) A | 100 A | 2.0 (0) A | 5.3 (0.5) A | 4.2 (0.1) A |
Mock control | 4 | 100 A | 1.3 (0.3) B | 5.5 (0.3) A | 13.2 (0.9) A | 16.9 (1.3) A | 100 A | 2.0 (0.3) A | 4.5 (0.9) A | 3.9 (0.2) A |
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Parreno, V.; Bai, M.; Liu, F.; Jing, J.; Olney, E.; Li, G.; Wen, K.; Yang, X.; Castellucc, T.B.; Kocher, J.F.; et al. Probiotic as Adjuvant Significantly Improves Protection of the Lanzhou Trivalent Rotavirus Vaccine against Heterologous Challenge in a Gnotobiotic Pig Model of Human Rotavirus Infection and Disease. Vaccines 2022, 10, 1529. https://doi.org/10.3390/vaccines10091529
Parreno V, Bai M, Liu F, Jing J, Olney E, Li G, Wen K, Yang X, Castellucc TB, Kocher JF, et al. Probiotic as Adjuvant Significantly Improves Protection of the Lanzhou Trivalent Rotavirus Vaccine against Heterologous Challenge in a Gnotobiotic Pig Model of Human Rotavirus Infection and Disease. Vaccines. 2022; 10(9):1529. https://doi.org/10.3390/vaccines10091529
Chicago/Turabian StyleParreno, Viviana, Muqun Bai, Fangning Liu, Jiqiang Jing, Erika Olney, Guohua Li, Ke Wen, Xingdong Yang, Tammy Bui Castellucc, Jacob F. Kocher, and et al. 2022. "Probiotic as Adjuvant Significantly Improves Protection of the Lanzhou Trivalent Rotavirus Vaccine against Heterologous Challenge in a Gnotobiotic Pig Model of Human Rotavirus Infection and Disease" Vaccines 10, no. 9: 1529. https://doi.org/10.3390/vaccines10091529