Reconstitution and Mutagenesis of Avian Infectious Laryngotracheitis Virus from Cosmid and Yeast Centromeric Plasmid Clones.

التفاصيل البيبلوغرافية
العنوان:	Reconstitution and Mutagenesis of Avian Infectious Laryngotracheitis Virus from Cosmid and Yeast Centromeric Plasmid Clones.
المؤلفون:	Spatz S; US National Poultry Research Center, Athens, Georgia, USA., García M; Poultry Diagnostic and Research Center, College of Veterinary Medicine, University of Georgia, Athens, Georgia, USA., Fuchs W; Institute of Molecular Virology and Cell Biology, Friedrich Loeffler-Institut, Federal Research Institute for Animal Health, Greifswald-Insel Riems, Germany., Loncoman C; Asia-Pacific Centre for Animal Health, Melbourne Veterinary School, Faculty of Veterinary and Agricultural Sciences, University of Melbourne, Parkville, Victoria, Australia., Volkening J; BASE2BIO, Oshkosh, Wisconsin, USA., Ross T; US National Poultry Research Center, Athens, Georgia, USA., Riblet S; Poultry Diagnostic and Research Center, College of Veterinary Medicine, University of Georgia, Athens, Georgia, USA., Kim T; US National Poultry Research Center, Athens, Georgia, USA., Likens N; US National Poultry Research Center, Athens, Georgia, USA., Mettenleiter T; Institute of Molecular Virology and Cell Biology, Friedrich Loeffler-Institut, Federal Research Institute for Animal Health, Greifswald-Insel Riems, Germany.
المصدر:	Journal of virology [J Virol] 2023 Apr 27; Vol. 97 (4), pp. e0140622. Date of Electronic Publication: 2023 Apr 06.
نوع المنشور:	Journal Article; Research Support, U.S. Gov't, Non-P.H.S.
اللغة:	English
بيانات الدورية:	Publisher: American Society For Microbiology Country of Publication: United States NLM ID: 0113724 Publication Model: Print-Electronic Cited Medium: Internet ISSN: 1098-5514 (Electronic) Linking ISSN: 0022538X NLM ISO Abbreviation: J Virol Subsets: MEDLINE
أسماء مطبوعة:	Publication: Washington Dc : American Society For Microbiology Original Publication: Baltimore, American Society for Microbiology.
مواضيع طبية MeSH:	Cosmids/genetics , Herpesvirus 1, Gallid/genetics , Herpesvirus 1, Gallid/pathogenicity , Mutagenesis , Plasmids*/genetics, Animals ; Male ; Chickens ; Herpesviridae Infections/virology ; Poultry Diseases/virology ; Saccharomyces cerevisiae/genetics ; Cell Line ; Genome, Viral/genetics ; Viral Proteins/genetics ; Viral Proteins/metabolism
مستخلص:	The genomes of numerous herpesviruses have been cloned as infectious bacterial artificial chromosomes. However, attempts to clone the complete genome of infectious laryngotracheitis virus (ILTV), formally known as Gallid alphaherpesvirus-1, have been met with limited success. In this study, we report the development of a cosmid/yeast centromeric plasmid (YCp) genetic system to reconstitute ILTV. Overlapping cosmid clones were generated that encompassed 90% of the 151-Kb ILTV genome. Viable virus was produced by cotransfecting leghorn male hepatoma (LMH) cells with these cosmids and a YCp recombinant containing the missing genomic sequences - spanning the TR S /UL junction. An expression cassette for green fluorescent protein (GFP) was inserted within the redundant inverted packaging site (ipac2), and the cosmid/YCp-based system was used to generate recombinant replication-competent ILTV. Viable virus was also reconstituted with a YCp clone containing a BamH I linker within the deleted ipac2 site, further demonstrating the nonessential nature of this site. Recombinants deleted in the ipac2 site formed plaques undistinguished from those viruses containing intact ipac2. The 3 reconstituted viruses replicated in chicken kidney cells with growth kinetics and titers similar to the USDA ILTV reference strain. Specific pathogen-free chickens inoculated with the reconstituted ILTV recombinants succumbed to levels of clinical disease similar to that observed in birds inoculated with wildtype viruses, demonstrating the reconstituted viruses were virulent. IMPORTANCE Infectious laryngotracheitis virus (ILTV) is an important pathogen of chicken with morbidity of 100% and mortality rates as high as 70%. Factoring in decreased production, mortality, vaccination, and medication, a single outbreak can cost producers over a million dollars. Current attenuated and vectored vaccines lack safety and efficacy, leaving a need for better vaccines. In addition, the lack of an infectious clone has also impeded understanding viral gene function. Since infectious bacterial artificial chromosome (BAC) clones of ILTV with intact replication origins are not feasible, we reconstituted ILTV from a collection of yeast centromeric plasmids and bacterial cosmids, and identified a nonessential insertion site within a redundant packaging site. These constructs and the methodology necessary to manipulate them will facilitate the development of improved live virus vaccines by modifying genes encoding virulence factors and establishing ILTV-based viral vectors for expressing immunogens of other avian pathogens.
فهرسة مساهمة:	Keywords: Gallid alphaherpesvirus-1; alphaherpesvirus; cosmid libraries; infectious laryngotracheitis virus; origin of DNA replication; packaging sites; yeast centromeric plasmids
المشرفين على المادة:	0 (Viral Proteins)
تواريخ الأحداث:	Date Created: 20230406 Date Completed: 20230517 Latest Revision: 20231007
رمز التحديث:	20231007
مُعرف محوري في PubMed:	PMC10134816
DOI:	10.1128/jvi.01406-22
PMID:	37022163
قاعدة البيانات:	MEDLINE

الوصف
تدمد:	1098-5514
DOI:	10.1128/jvi.01406-22