Isolation and characterization of bovine herpesvirus 4 (BoHV-4) from a cow affected by post partum metritis and cloning
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BioMed Central
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Isolation and characterization of bovine herpesvirus 4 (BoHV-4) from a cow affected by post partum metritis and cloning of the genome as a bacterial artificial chromosome Gaetano Donofrio*1, Valentina Franceschi1, Antonio Capocefalo1, Sandro Cavirani1 and Iain Martin Sheldon2 Address: 1Dipartimento di Salute Animale, Facoltà di Medicina Veterinaria, Università di Parma, via del Taglio 10, 43100 Parma, Italy and 2Institute of Life Science, School of Medicine, Swansea University, Singleton Park, Swansea, SA2 8PP, UK Email: Gaetano Donofrio* - [email protected]; Valentina Franceschi - [email protected]; Antonio Capocefalo - [email protected]; Sandro Cavirani - [email protected]; Iain Martin Sheldon - [email protected] * Corresponding author
Published: 19 August 2009 Reproductive Biology and Endocrinology 2009, 7:83
doi:10.1186/1477-7827-7-83
Received: 7 July 2009 Accepted: 19 August 2009
This article is available from: http://www.rbej.com/content/7/1/83 © 2009 Donofrio et al; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
Abstract Background: Bovine herpesvirus 4 (BoHV-4) is a gammaherpesvirus with a Worldwide distribution in cattle and is often isolated from the uterus of animals with postpartum metritis or pelvic inflammatory disease. Virus strain adaptation to an organ, tissue or cell type is an important issue for the pathogenesis of disease. To explore the mechanistic role of viral strain variation for uterine disease, the present study aimed to develop a tool enabling precise genetic discrimination between strains of BoHV-4 and to easily manipulate the viral genome. Methods: A strain of BoHV-4 was isolated from the uterus of a persistently infected cow and designated BoHV-4-U. The authenticity of the isolate was confirmed by RFLP-PCR and sequencing using the TK and IE2 loci as genetic marker regions for the BoHV-4 genome. The isolated genome was cloned as a Bacterial Artificial Chromosome (BAC) and manipulated through recombineering technology Results: The BoHV-4-U genome was successfully cloned as a BAC, and the stability of the pBACBoHV-4-U clone was confirmed over twenty passages, with viral growth similar to the wild type virus. The feasibility of using BoHV-4-U for mutagenesis was demonstrated using the BAC recombineering system. Conclusion: The analysis of genome strain variation is a key method for investigating genes associated with disease. A resource for dissection of the interactions between BoHV-4 and host endometrial cells was generated by cloning the genome of BoHV-4 as a BAC.
Background Uterine infections are important because they disrupt not only the function of the uterus, but also the ovary and the overarching higher control centres in the hypothalamus
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