Wharton's jelly cells from sheep umbilical cord maintained with different culture media are permissive to in vitro infection by Small Ruminant Lentiviruses

R.P Dias,A.L.M Sousa,A.C Farias,M.F.S Teixeira,A Andrioli,J.F Araújo,R.R Pinheiro,D.A.A Azevedo,T.D.F Aguiar

doi:10.1590/1678-4162-9008

Abstract

ABSTRACT This study aimed to isolate cells from the Wharton's jelly of umbilical cord (WJUC) of sheep collected during natural parturition using different culture media, in addition to reporting for the first time the permissiveness of these cells to in vitro infection by small ruminant lentiviruses. Ten umbilical cords were collected from healthy sheep. Each cord explants were grown in different media consisting of MEM, low glucose DMEM, M199, and RPMI-1640. The permissiveness of infection of sheep cells from WJUC was tested with CAEV-Cork and MVV-K1514 strains, inoculating 0.1 MOI of each viral strain. Four supernatants from each strain were obtained from WJUC sheep cell cultures infected in different media. The results demonstrated the presence of cytopathic effect after the in vitro infection by CAEV-Cork and MVV-K1514 with all of the tested culture media. Nested-PCR detected proviral DNA in all supernatants. Supernatants containing CAEV-Cork viruses had TCID50/ml titres of 105.5 in MEM, 104.0 in low glucose DMEM, 105.0 in M199, and 105.7 in RPMI-1640. Supernatants containing the MVV-K1514 virus had TCID50/ml titres of 104.3 in MEM, 103.5 in low-glucose DMEM, 104.7 in M199, and 103.5 in RPMI-1640. Sheep cells from WJUC are permissive to in vitro infection by small ruminant lentivirus.

Highlights

SRLV are a group of highly heterogenic viruses capable of infecting sheep and goats, causing different clinical manifestations depending on the virulence of the strain involved
In order to do so, the in vitro production of viral proteins is essential due to their use in indirect diagnostic tests. This production is performed with viral replication in permissive cells, which are most frequently extracted from goat synovial membrane (GSM) and choroid plexus, and used to replicate caprine arthritis encephalitis virus (CAEV), and Maedi-Visna virus (MVV), respectively (Caprine..., 2008)
Each cord was washed with sterile PBS added 2% penicillin and streptomycin (P/S) (GibcoTM, California) to remove the excess of blood and to disinfect, placed in 50mL falcon tubes immersed in low glucose DMEM added with 4% P/S and 1% amphotericin B (Cristália - Produtos Químicos Farmacêuticos Ltda., Brasil)

Summary

Introduction

SRLV are a group of highly heterogenic viruses capable of infecting sheep and goats, causing different clinical manifestations depending on the virulence of the strain involved. In order to do so, the in vitro production of viral proteins is essential due to their use in indirect diagnostic tests This production is performed with viral replication in permissive cells, which are most frequently extracted from goat synovial membrane (GSM) and choroid plexus, and used to replicate caprine arthritis encephalitis virus (CAEV), and Maedi-Visna virus (MVV), respectively (Caprine..., 2008). These tissues are usually collected from fetuses or neonates, which are euthanized to obtain cells with a higher potential for in vitro growth. In this context, using umbilical cord tissue is an alternative that provides cells without harming ethical principles or causing unnecessary suffering in animals, in addition to being an easy source to obtain these cells with an elevated capacity for in vitro proliferation (Vita et al, 2012)

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