Production of Newcastle Disease Virus by Vero Cells Grown on Cytodex 1 Microcarriers in a 2-Litre Stirred Tank Bioreactor

Mohd Azmir Arifin,Maizirwan Mel,Mohamed Ismail Abdul Karim,Aini Ideris

doi:10.1155/2010/586363

Abstract

The aim of this study is to prepare a model for the production of Newcastle disease virus (NDV) lentogenic F strain using cell culture in bioreactor for live attenuated vaccine preparation. In this study, firstly we investigated the growth of Vero cells in several culture media. The maximum cell number was yielded by culture of Vero cells in Dulbecco's Modified Eagle Medium (DMEM) which was 1.93 × 106 cells/ml. Secondly Vero cells were grown in two-litre stirred tank bioreactor by using several commercial microcarriers. We achieved the maximum cell concentration about 7.95 × 105 cells/ml when using Cytodex 1. Later we produced Newcastle Disease virus in stirred tank bioreactor based on the design developed using Taguchi L4 method. Results reveal that higher multiplicity of infection (MOI) and size of cell inoculums can yield higher virus titer. Finally, virus samples were purified using high-speed centrifugation based on 3∗∗(3-1) Fractional Factorial Design. Statistical analysis showed that the maximum virus titer can be achieved at virus sample concentration of 58.45% (v/v), centrifugation speed of 13729 rpm, and centrifugation time of 4 hours. As a conclusion, high yield of virus titer could be achieved through optimization of cell culture in bioreactor and separation by high-speed centrifugation.

Highlights

Newcastle disease (ND), caused by Newcastle disease virus (NDV), is reported as the most important viral disease of poultry in the world [1]
Later we investigated the effect of different sizes of cell inoculum, serum concentrations during virus growth phase, and multiplicity of infection (MOI) on the virus titer
Dulbecco’s Modified Eagle Medium (DMEM) was selected to be used in further stages of the study because it has the best performance in terms of maximum cell concentration yielded and growth kinetic

Summary

Introduction

Newcastle disease (ND), caused by Newcastle disease virus (NDV), is reported as the most important viral disease of poultry in the world [1]. The disease can vary from clinically inapparent to highly virulent forms, depending on the virus strain and the host species [2]. Nine avian paramyxovirus serotypes (APMV-1 to APMV-9), of which APMV-1 is the most economically important, have been identified among these virus types [3]. NDV isolates display a spectrum of virulence in chickens, from a fatal to an inapparent infection [3]. Strains of NDV are classified into three major pathotypes, depending on the severity of disease produced in chickens. Lentogenic strains do not usually cause disease in adult chickens and are widely used as live vaccines in poultry industries in the United States and other countries. Viruses of intermediate virulence that cause respiratory disease are termed mesogenic, while virulent viruses that cause high mortality are termed as velogenic [5]

Objectives

Methods

Results

Conclusion