Mutation E48K in PB1 Polymerase Subunit Improves Stability of a Candidate Live Attenuated Influenza B Virus Vaccine.

Jongsuk Mo,Daniel R Perez,Daniela S Rajao,Lucas M Ferreri,Ginger Geiger,C Joaquín Cáceres,Stivalis Cardenas-Garcia,Jefferson J S Santos

doi:10.3390/vaccines9070800

Abstract

Influenza B virus (IBV) is a major respiratory pathogen of humans, particularly in the elderly and children, and vaccines are the most effective way to control it. In previous work, incorporation of two mutations (E580G, S660A) along with the addition of an HA epitope tag in the PB1 segment of B/Brisbane/60/2008 (B/Bris) resulted in an attenuated strain that was safe and effective as a live attenuated vaccine. A third attempted mutation (K391E) in PB1 was not always stable. Interestingly, viruses that maintained the K391E mutation were associated with the mutation E48K. To explore the contribution of the E48K mutation to stability of the K391E mutation, a vaccine candidate was generated by inserting both mutations, along with attenuating mutations E580G and S660A, in PB1 of B/Bris (B/Bris PB1att 4M). Serial passages of the B/Bris PB1att 4M vaccine candidate in eggs and MDCK indicated high stability. In silico structural analysis revealed a potential interaction between amino acids at positions 48 and 391. In mice, B/Bris PB1att 4M was safe and provided complete protection against homologous challenge. These results confirm the compensatory effect of mutation E48K to stabilize the K391E mutation, resulting in a safer, yet still protective, IBV LAIV vaccine.

Highlights

Influenza B virus (IBV) is an enveloped virus of the Orthomyxoviridae family with a negative-sense single-stranded RNA genome [1]
The B/Bris PB1 3M and B/Bris PB1att 4M strains were serially passaged at 33 ◦C in Madin–Darby canine kidney (MDCK) cells and specific pathogen-free (SPF) eggs
Sanger sequencing analysis of the PB1 segment confirmed that the B/Bris PB1att 4M virus stably retained all modifications introduced (E48K, K391E, E580G, and S660A) after four serial passages in MDCK cells and in SPF eggs

Summary

Introduction

Influenza B virus (IBV) is an enveloped virus of the Orthomyxoviridae family with a negative-sense single-stranded RNA genome [1]. IBV causes seasonal respiratory disease epidemics in humans. IBV can affect all age groups, studies demonstrate that individuals under the age of 18 are more susceptible to infection [4]. IBV infection can be severe in children and is associated with a high proportion of pediatric deaths related to influenza [5,6]. IBV infections are less common than influenza A virus (IAV) infections. IBV strains are classified into two antigenically distinct lineages, the B/Victoria (B/Vic) and the B/Yamagata (B/Yam) lineages [7,8]. The development of effective vaccines against IBV is important to reduce disease burden, especially in vulnerable age groups

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