Virome Characterization in Commercial Bovine Serum Batches-A Potentially Needed Testing Strategy for Biological Products.

Willian P Paim,Matheus N Weber,Shollie M Falkenberg,Fernando V Bauermann,Mayara F Maggioli,Cláudio W Canal,Akhilesh Ramachandran

doi:10.3390/v13122425

Abstract

Bovine serum has been widely used as a universal supplement in culture media and other applications, including the manufacture of biological products and the production of synthetic meat. Currently, commercial bovine serum is tested for possible viral contaminants following regional guidelines. Regulatory agencies’ established tests focused on detecting selected animal origin viruses and are based on virus isolation, immunofluorescence, and hemadsorption assays. However, these tests may fail to detect new or emerging viruses in biological products. High-throughput sequencing is a powerful option since no prior knowledge of the viral targets is required. In the present study, we evaluate the virome of seven commercial batches of bovine serum from Mexico (one batch), New Zealand (two batches), and the United States (four batches) using a specific preparation and enrichment method for pooled samples and sequencing using an Illumina platform. A variety of circular replicase-encoding single-stranded (CRESS) DNA families (Genomoviridae, Circoviridae, and Smacoviridae) was identified. Additionally, CrAssphage, a recently discovered group of bacteriophage correlated with fecal contamination, was identified in 85% of the tested batches. Furthermore, sequences representing viral families with single-stranded DNA (Parvoviridae), double-stranded DNA (Polyomaviridae and Adenoviridae), single-stranded RNA (Flaviviridae, Picornaviridae, and Retroviridae), and double-stranded RNA (Reoviridae) were identified. These results support that high-throughput sequencing associated with viral enrichment is a robust tool and should be considered an additional layer of safety when testing pooled biologicals to detect viral contaminants overlooked by the current testing protocols.

Highlights

Bovine serum has been used as a universal supplement in cell culture applications [1]
The United States Department of Agriculture (USDA) regulations require the testing of bovine serum for Bovine Adenovirus (BAV), Bovine Herpesvirus (BoHV), Bovine Parvovirus (BPV), Bovine Respiratory Syncytial Virus (BRSV), Bovine Viral Diarrhea Virus (BVDV), Bluetongue Virus (BTV), Parainfluenza Virus 3 (PI3), Rabies Virus (RABV), and Vesicular Stomatitis Virus (VSV)
The complete coding genomes were obtained for Bovine Parvovirus 2 (BPV-2), Bovine Parvovirus 3 (BPV-3), Bosavirus (BosaV), Bovine Hokovirus 2 (BHoV-2) (Figure 1), and Bovine Polyomavirus 1 (BPyV-1) (Figure 2)

Summary

Introduction

Bovine serum has been used as a universal supplement in cell culture applications [1]. The cell culture uses include in-vitro diagnostics, vaccine production, gene therapy, stem cell therapy, and, more recently, the production of synthetic meat [2,3,4]. Bovine origin products used to manufacture biologicals are currently tested for viral contaminants following regional guidelines. The USDA regulations require the testing of bovine serum for Bovine Adenovirus (BAV), Bovine Herpesvirus (BoHV), Bovine Parvovirus (BPV), Bovine Respiratory Syncytial Virus (BRSV), Bovine Viral Diarrhea Virus (BVDV), Bluetongue Virus (BTV), Parainfluenza Virus 3 (PI3), Rabies Virus (RABV), and Vesicular Stomatitis Virus (VSV). In Europe, the use of bovine serum for the manufacture of human medicinal products with a biological origin is regulated by the European Medicines Agency (EMA/CHMP/BWP/457920/2012 Rev.). The veterinary products in Europe are regulated by the EMA-CVMP-743-00-Rev.

Methods

Results

Conclusion