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      The use of cells from ANPEP knockout pigs to evaluate the role of aminopeptidase N (APN) as a receptor for porcine deltacoronavirus (PDCoV)

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          Abstract

          The coronaviruses, porcine epidemic diarrhea virus (PEDV), transmissible gastroenteritis virus (TGEV), and porcine deltacoronavirus (PDCoV) represent important sources of neonatal diarrhea on pig farms. The requirement for aminopeptidase N (APN) as a receptor for TGEV, but not for PEDV, is well established. In this study, the biological relevance of APN as a receptor for PDCoV was tested by using CRISPR/Cas9 to knockout the APN gene, ANPEP, in pigs. Porcine alveolar macrophages (PAMs) from ANPEP knockout (KO) pigs showed resistance to PDCoV infection. However, lung fibroblast-like cells, derived from the ANPEP KO PAM cultures, supported PDCoV infection to high levels. The results suggest that APN is a receptor for PDCoV in PAMs but is not necessary for infection of lung-derived fibroblast cells. The infection of the ANPEP KO pigs with PDCoV further confirmed that APN is dispensable as a receptor for PDCoV.

          Highlights

          • •

            APN is a receptor for PDCoV in PAMS, but not fibroblast cells.

          • •

            ANPEP KO pigs are not resistant to PDCoV.

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          Emergence of Porcine epidemic diarrhea virus in the United States: clinical signs, lesions, and viral genomic sequences.

          Gregory Stevenson, Hai Hoang, Kent Schwartz … (2013)
          During the 10 days commencing April 29, 2013, the Iowa State University Veterinary Diagnostic Laboratory received the first 4 of many submissions from swine farms experiencing explosive epidemics of diarrhea and vomiting affecting all ages, with 90-95% mortality in suckling pigs. Histology revealed severe atrophy of villi in all segments of the small intestines with occasional villus-epithelial syncytial cells, but testing for rotaviruses and Transmissible gastroenteritis virus (Alphacoronavirus 1) were negative. Negative-staining electron microscopy of feces revealed coronavirus-like particles and a pan-coronavirus polymerase chain reaction (PCR) designed to amplify a conserved region of the polymerase gene for all members in the family Coronaviridae produced expected 251-bp amplicons. Subsequent sequencing and analysis revealed 99.6-100% identity among the PCR amplicons from the 4 farms and 97-99% identity to the corresponding portion of the polymerase gene of Porcine epidemic diarrhea virus (PEDV) strains, with the highest identity (99%) to strains from China in 2012. Findings were corroborated at National Veterinary Services Laboratories using 2 nested S-gene and 1 nested N-gene PCR tests where the sequenced amplicons also had the highest identity with 2012 China strains. Whole genome sequence for the virus from 2 farms in 2 different states using next-generation sequencing technique was compared to PEDV sequences available in GenBank. The 2013 U.S. PEDV had 96.6-99.5% identity with all known PEDV strains and the highest identity (>99.0%) to some of the 2011-2012 Chinese strains. The nearly simultaneous outbreaks of disease, and high degree of homology (99.6-100%) between the PEDV strains from the 4 unrelated farms, suggests a common source of virus.
          Article 0 comments Cited 372 times – based on 0 reviews     Review now
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            Detection and Genetic Characterization of Deltacoronavirus in Pigs, Ohio, USA, 2014

            Leyi Wang, Beverly Byrum, Yan-Yan Zhang (2014)
            In Ohio, United States, in early 2014, a deltacoronavirus was detected in feces and intestine samples from pigs with diarrheal disease. The complete genome sequence and phylogenetic analysis of the virus confirmed that the virus is closely related to a porcine deltacoronavirus (porcine coronavirus HKU15) reported in Hong Kong in 2012.
            Article 0 comments Cited 209 times – based on 0 reviews     Review now
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              Cloning of the mouse hepatitis virus (MHV) receptor: expression in human and hamster cell lines confers susceptibility to MHV.

              G Dveksler, M Pensiero, C B Cardellichio … (1991)
              The cellular receptor for murine coronavirus mouse hepatitis virus (MHV)-A59 is a member of the carcinoembryonic antigen (CEA) family of glycoproteins in the immunoglobulin superfamily. We isolated a cDNA clone (MHVR1) encoding the MHV receptor. The sequence of this clone predicts a 424-amino-acid glycoprotein with four immunoglobulinlike domains, a transmembrane domain, and a short intracytoplasmic tail, MHVR1 is closely related to the murine CEA-related clone mmCGM1 (Mus musculus carcinoembryonic antigen gene family member). Western blot (immunoblot) analysis performed with antireceptor antibodies detected a glycoprotein of 120 kDa in BHK cells stably transfected with MHVR1. This corresponds to the size of the MHV receptor expressed in mouse intestine and liver. Human and hamster fibroblasts transfected with MHVR1 became susceptible to infection with MHV-A59. Like MHV-susceptible mouse fibroblasts, the MHVR1-transfected human and hamster cells were protected from MHV infection by pretreatment with monoclonal antireceptor antibody CC1. Thus, the 110- to 120-kDa CEA-related glycoprotein encoded by MHVR1 is a functional receptor for murine coronavirus MHV-A59.
              Article 0 comments Cited 115 times – based on 0 reviews     Review now
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                Author and article information

                Contributors
                Randall S. Prather
                Journal
                Journal ID (nlm-ta): Virology
                Journal ID (iso-abbrev): Virology
                Title: Virology
                Publisher: The Authors. Published by Elsevier Inc.
                ISSN (Print): 0042-6822
                ISSN (Electronic): 1096-0341
                Publication date PMC-release: 24 December 2019
                Publication date (Print): February 2020
                Publication date (Electronic): 24 December 2019
                Volume: 541
                Pages: 136-140
                Affiliations
                [a ]Division of Animal Science, University of Missouri, Columbia, MO, 65211, USA
                [b ]Department of Diagnostic Medicine and Pathobiology, Kansas State University, Manhattan, KS, USA
                [c ]Genus plc, DeForest, WI, USA
                [d ]Veterinary Diagnostics and Production Animal Medicine, Iowa State University, Ames, IA, USA
                Author notes
                [∗ ]Corresponding author. PratherR@ 123456missouri.edu
                Article
                Publisher ID: S0042-6822(19)30349-6
                DOI: 10.1016/j.virol.2019.12.007
                PMC ID: 7112016
                PubMed ID: 32056711
                SO-VID: 4818355a-8857-4275-89b8-537e7fb5b4e2
                Copyright © © 2019 The Authors
                License:

                Since January 2020 Elsevier has created a COVID-19 resource centre with free information in English and Mandarin on the novel coronavirus COVID-19. The COVID-19 resource centre is hosted on Elsevier Connect, the company's public news and information website. Elsevier hereby grants permission to make all its COVID-19-related research that is available on the COVID-19 resource centre - including this research content - immediately available in PubMed Central and other publicly funded repositories, such as the WHO COVID database with rights for unrestricted research re-use and analyses in any form or by any means with acknowledgement of the original source. These permissions are granted for free by Elsevier for as long as the COVID-19 resource centre remains active.

                History
                Date received : 10 October 2019
                Date revision received : 16 December 2019
                Date accepted : 17 December 2019
                Categories
                Subject: Article

                ScienceOpen disciplines: Microbiology & Virology
                Keywords: aminopeptidase n,porcine deltacoronavirus,apn,crispr,anpep,cd13
                Data availability:
                ScienceOpen disciplines: Microbiology & Virology
                Keywords: aminopeptidase n, porcine deltacoronavirus, apn, crispr, anpep, cd13

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