An Influenza HA and M2e Based Vaccine Delivered by a Novel Attenuated Salmonella Mutant Protects Mice against Homologous H1N1 Infection

Abstract

Attenuated Salmonella strains constitute a promising technology for the development of a more efficient multivalent protein based vaccines. In this study, we constructed a novel attenuated strain of Salmonella for the delivery and expression of the H1N1 hemagglutinin (HA) and the conserved extracellular domain of the matrix protein 2 (M2e). We demonstrated that the constructed Salmonella strain exhibited efficient HA and M2e protein expressions and little cytotoxicity and pathogenicity in mice. Using BALB/c mice as the model, we showed that the mice vaccinated with a Salmonella strain expressing HA and M2e protein antigens, respectively, induced significant production of HA and M2e-specific serum IgG1 and IgG2a responses, and of anti-HA interferon-γ producing T cells. Furthermore, immunization with Salmonella-HA-M2e-based vaccine via different routes provided protection in 66.66% orally, 100% intramuscularly, and 100% intraperitoneally immunized mice against the homologous H1N1 virus while none of the animals survived treated with either the PBS or the Salmonella carrying empty expression vector. Ex vivo stimulated dendritic cells (DCs) with heat killed Salmonella expressing HA demonstrated that DCs play an important role in the elicitation of HA-specific humoral immune responses in mice. In summary, Salmonella-HA-M2e-based vaccine elicits efficient antigen-specific humoral and cellular immune responses, and provides significant immune protection against a highly pathogenic H1N1 influenza virus.

Oral immunization with a novel attenuated Salmonella Gallinarum encoding infectious bronchitis virus spike protein induces protective immune responses against fowl typhoid and infectious bronchitis in chickens

“…These types of vaccines are easy to prepare, stable, convenient, economical, and easy to administer in chickens. Moreover, these types of vaccines does not require exogenous adjuvants as Salmonella bacteria provide an appropriate danger signals to the immune system, acting as natural adjuvant, thereby promoting efficient maturation and activation of dendritic cells (DCs) [ 25 , 30 ], which is prerequisite for the induction of potent adaptive immune responses. Also SG-based IB vaccination can differentiate vaccinated birds from infected ones as only structural and immunogenic antigens are exploited for the vaccine development.…”

“…The S1 nucleotide sequence was codon optimized for efficient expression in SG. The optimized gene sequence was chemically synthesized (Bionee, Korea) and then built into the pJHL65 plasmid, an asd + constitutive expression vector, and propagated in an asd mutated Escherichia coli strain as described previously [ 25 ]. The S1 gene sequence was cloned in frame downstream to the beta-lactamase signal sequence ( bla SS ) of pJHL65 vector, so that the intended protein would get secreted out of the bacteria [ 26 ].…”

Oral immunization with a novel attenuated Salmonella Typhimurium encoding influenza HA, M2e and NA antigens protects chickens against H7N9 infection

“…The HA consensus sequence derived from H7N3, H7N7 and H7N9 viruses was constructed and built into pMMP65 expression vector as described elsewhere [ 15 ]. The four tandem repeats of M2e sequence (MSLLTEVETPTRNGWECKCSDSSD) was constructed and built into pMMP65 vector as described elsewhere [ 18 ]. The NA consensus sequence was deduced from 605 sequences of H7N9 virus strains based on the data available in the GenBank (Additional file 1 ) and optimized for efficient gene expression in Salmonella , then synthesized (Bionee, Korea) and built into the pMMP65 vector as described earlier [ 15 ].…”

Intranasally administered protein coated chitosan nanoparticles encapsulating influenza H9N2 HA2 and M2e mRNA molecules elicit protective immunity against avian influenza viruses in chickens

“…The recombinant pET28a-HA2-HA1 plasmid was subsequently transformed into E. coli BL21 (DE3) pLysS host strain (Novagen, San Diego, USA) for expression and purification of HA2-HA1 recombinant protein, as described previously [30]. The four tandem repeats of conserved M2e gene sequence (MSLLTE-VETPTRNGWECKCSDSSD) were cloned in-frame into pET32a (+) prokaryotic expression vector (Novagen, San Diego, USA) and expressed in E. coli BL21 (DE3) pLysS host strain (Novagen, San Diego, USA), as discussed and reported elsewhere [31].…”

“…Western blotting indicated that HA2-HA1 protein reacted specifically with the H9N2 polyclonal anti-HA antibody giving a characteristic band at the expected size, thus confirming the authenticity of the expressed protein (Additional file 2B). The expression of the conserved H9N2 M2e tandem repeat sequence is described and reported elsewhere [31].…”