Large-Scale Microcarrier Culture of Chinese Perch Brain Cell for Viral Vaccine Production in a Stirred Bioreactor.

Xia Luo,Lihui Liu,Xiaozhe Fu,Yinjie Niu,Hongru Liang,Ningqiu Li,Qiang Lin

doi:10.3390/vaccines9091003

Abstract

Mandarin fish (Siniperca chuatsi) is one of the important cultured fish species in China. Infectious spleen and kidney necrosis virus (ISKNV) and Siniperca Chuatsi rhabdovirus (SCRV) have hindered the development of mandarin fish farming industry. Vaccination is the most effective method for control of viral diseases, however viral vaccine production requires the large-scale culture of cells. Herein, a suspension culture system of Chinese perch brain cell (CPB) was developed on Cytodex 1 microcarrier in a stirred bioreactor. Firstly, CPB cells were cultured using Cytodex 1 microcarrier in 125 mL stirring flasks. With the optimum operational parameters, CPB cells grew well, distributed uniformly, and could fully cover the microcarriers. Then, CPB cells were digested with trypsin and expanded step-by-step with different expansion ratios from the 125 mL stirring bottle to a 500 mL stirring bottle, and finally to a 3-L bioreactor. Results showed that with an expansion ratio of 1:3, we achieved a high cell density level (2.25 × 106 cells/mL) with an efficient use of the microcarriers, which also confirmed the data obtained from the 125 mL stirring flask. Moreover, obvious cytopathic effects (CPE) were observed in the suspended CPB cells post-infection with ISKNV and SCRV. This study provided a large-scale culture system of CPB cells for virus vaccine production.

Highlights

This study provided technical parameters for scale production of Chinese perch brain (CPB) cells for infectious spleen and kidney necrosis virus (ISKNV) and Siniperca chuatsi rhabdovirus (SCRV) vaccine production
CPB cells were distributed homogeneously on Cytodex 1 (Figure 3B), and with a highh) was carried out, CPB cells were distributed homogeneously on Cytodex 1 (Figure 3B), and with a attachment efficiency (i.e., 88.2%) and low empty-loading ratio
Data in our current study has proved that CPB cells could grow well on this kind of microcarrier under a suitable stirring regime

Summary

Introduction

Mandarin fish (Siniperca chuatsi) is one of the important cultured fish species in China. In recent years epidemic diseases, mainly caused by infectious spleen and kidney necrosis virus (ISKNV) and Siniperca chuatsi rhabdovirus (SCRV), have seriously hindered the development of the mandarin fish farming industry [1,2,3]. In China, Dong [4] and Fu [5] have developed the sensitive cell lines mandarin fish fry (MFF-1) and Chinese perch brain (CPB) for ISKNV and SCRV multiplication. Several effective inactivated or live vaccines against the mentioned viruses have been developed. Zhang et al [6] developed an inactivated ISKNV vaccine with chitosan as the adjuvant, which can play a significantly protective role against ISKNV infections. Zhang et al [7] reported a clonal Micropterus salmoides rhabdovirus Sanshui-7

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