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Efficiency of NHEJ-CRISPR/Cas9 and Cre-LoxP Engineered Recombinant Turkey Herpesvirus Expressing Pasteurella multocida OmpH Protein for Fowl Cholera Prevention in Ducks

Fowl cholera is caused by the bacterium Pasteurella multocida, a highly transmissible avian ailment with significant global implications, leading to substantial economic repercussions. The control of fowl cholera outbreaks primarily relies on vaccination using traditional vaccines that are still in...

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Autores principales: Apinda, Nisachon, Yao, Yongxiu, Zhang, Yaoyao, Muenthaisong, Anucha, Sangkakam, Kanokwan, Nambooppha, Boondarika, Rittipornlertrak, Amarin, Koonyosying, Pongpisid, Nair, Venugopal, Sthitmatee, Nattawooti
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2023
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10535566/
https://www.ncbi.nlm.nih.gov/pubmed/37766174
http://dx.doi.org/10.3390/vaccines11091498
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author Apinda, Nisachon
Yao, Yongxiu
Zhang, Yaoyao
Muenthaisong, Anucha
Sangkakam, Kanokwan
Nambooppha, Boondarika
Rittipornlertrak, Amarin
Koonyosying, Pongpisid
Nair, Venugopal
Sthitmatee, Nattawooti
author_facet Apinda, Nisachon
Yao, Yongxiu
Zhang, Yaoyao
Muenthaisong, Anucha
Sangkakam, Kanokwan
Nambooppha, Boondarika
Rittipornlertrak, Amarin
Koonyosying, Pongpisid
Nair, Venugopal
Sthitmatee, Nattawooti
author_sort Apinda, Nisachon
collection PubMed
description Fowl cholera is caused by the bacterium Pasteurella multocida, a highly transmissible avian ailment with significant global implications, leading to substantial economic repercussions. The control of fowl cholera outbreaks primarily relies on vaccination using traditional vaccines that are still in use today despite their many limitations. In this research, we describe the development of a genetically engineered herpesvirus of turkeys (HVT) that carries the OmpH gene from P. multocida integrated into UL 45/46 intergenic region using CRISPR/Cas9-NHEJ and Cre-Lox system editing. The integration and expression of the foreign cassettes were confirmed using polymerase chain reaction (PCR), indirect immunofluorescence assays, and Western blot assays. The novel recombinant virus (rHVT-OmpH) demonstrated stable integration of the OmpH gene even after 15 consecutive in vitro passages, along with similar in vitro growth kinetics as the parent HVT virus. The protective efficacy of the rHVT-OmpH vaccine was evaluated in vaccinated ducks by examining the levels of P. multocida OmpH-specific antibodies in serum samples using ELISA. Groups of ducks that received the rHVT-OmpH vaccine or the rOmpH protein with Montanide™ (SEPPIC, Paris, France) adjuvant exhibited high levels of antibodies, in contrast to the negative control groups that received the parental HVT or PBS. The recombinant rHVT-OmpH vaccine also provided complete protection against exposure to virulent P. multocida X-73 seven days post-vaccination. This outcome not only demonstrates that the HVT vector possesses many characteristics of an ideal recombinant viral vaccine vector for protecting non-chicken hosts, such as ducks, but also represents significant research progress in identifying a modern, effective vaccine candidate for combatting ancient infectious diseases.
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spelling pubmed-105355662023-09-29 Efficiency of NHEJ-CRISPR/Cas9 and Cre-LoxP Engineered Recombinant Turkey Herpesvirus Expressing Pasteurella multocida OmpH Protein for Fowl Cholera Prevention in Ducks Apinda, Nisachon Yao, Yongxiu Zhang, Yaoyao Muenthaisong, Anucha Sangkakam, Kanokwan Nambooppha, Boondarika Rittipornlertrak, Amarin Koonyosying, Pongpisid Nair, Venugopal Sthitmatee, Nattawooti Vaccines (Basel) Article Fowl cholera is caused by the bacterium Pasteurella multocida, a highly transmissible avian ailment with significant global implications, leading to substantial economic repercussions. The control of fowl cholera outbreaks primarily relies on vaccination using traditional vaccines that are still in use today despite their many limitations. In this research, we describe the development of a genetically engineered herpesvirus of turkeys (HVT) that carries the OmpH gene from P. multocida integrated into UL 45/46 intergenic region using CRISPR/Cas9-NHEJ and Cre-Lox system editing. The integration and expression of the foreign cassettes were confirmed using polymerase chain reaction (PCR), indirect immunofluorescence assays, and Western blot assays. The novel recombinant virus (rHVT-OmpH) demonstrated stable integration of the OmpH gene even after 15 consecutive in vitro passages, along with similar in vitro growth kinetics as the parent HVT virus. The protective efficacy of the rHVT-OmpH vaccine was evaluated in vaccinated ducks by examining the levels of P. multocida OmpH-specific antibodies in serum samples using ELISA. Groups of ducks that received the rHVT-OmpH vaccine or the rOmpH protein with Montanide™ (SEPPIC, Paris, France) adjuvant exhibited high levels of antibodies, in contrast to the negative control groups that received the parental HVT or PBS. The recombinant rHVT-OmpH vaccine also provided complete protection against exposure to virulent P. multocida X-73 seven days post-vaccination. This outcome not only demonstrates that the HVT vector possesses many characteristics of an ideal recombinant viral vaccine vector for protecting non-chicken hosts, such as ducks, but also represents significant research progress in identifying a modern, effective vaccine candidate for combatting ancient infectious diseases. MDPI 2023-09-18 /pmc/articles/PMC10535566/ /pubmed/37766174 http://dx.doi.org/10.3390/vaccines11091498 Text en © 2023 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Apinda, Nisachon
Yao, Yongxiu
Zhang, Yaoyao
Muenthaisong, Anucha
Sangkakam, Kanokwan
Nambooppha, Boondarika
Rittipornlertrak, Amarin
Koonyosying, Pongpisid
Nair, Venugopal
Sthitmatee, Nattawooti
Efficiency of NHEJ-CRISPR/Cas9 and Cre-LoxP Engineered Recombinant Turkey Herpesvirus Expressing Pasteurella multocida OmpH Protein for Fowl Cholera Prevention in Ducks
title Efficiency of NHEJ-CRISPR/Cas9 and Cre-LoxP Engineered Recombinant Turkey Herpesvirus Expressing Pasteurella multocida OmpH Protein for Fowl Cholera Prevention in Ducks
title_full Efficiency of NHEJ-CRISPR/Cas9 and Cre-LoxP Engineered Recombinant Turkey Herpesvirus Expressing Pasteurella multocida OmpH Protein for Fowl Cholera Prevention in Ducks
title_fullStr Efficiency of NHEJ-CRISPR/Cas9 and Cre-LoxP Engineered Recombinant Turkey Herpesvirus Expressing Pasteurella multocida OmpH Protein for Fowl Cholera Prevention in Ducks
title_full_unstemmed Efficiency of NHEJ-CRISPR/Cas9 and Cre-LoxP Engineered Recombinant Turkey Herpesvirus Expressing Pasteurella multocida OmpH Protein for Fowl Cholera Prevention in Ducks
title_short Efficiency of NHEJ-CRISPR/Cas9 and Cre-LoxP Engineered Recombinant Turkey Herpesvirus Expressing Pasteurella multocida OmpH Protein for Fowl Cholera Prevention in Ducks
title_sort efficiency of nhej-crispr/cas9 and cre-loxp engineered recombinant turkey herpesvirus expressing pasteurella multocida omph protein for fowl cholera prevention in ducks
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10535566/
https://www.ncbi.nlm.nih.gov/pubmed/37766174
http://dx.doi.org/10.3390/vaccines11091498
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