Recombinant outer membrane protein A induces a protective immune response against Escherichia coli infection in mice

Abstract

Pathogenic Escherichia coli (E. coli) is an important infectious Gram-negative bacterium causing millions of death every year. Outer membrane protein A (OmpA) has been suggested as a potential vaccine candidate for conferring protection against bacterial infection. In this study, a universal vaccine candidate for E. coli infection was developed and evaluated. Bioinformatics analysis revealed the OmpA protein from E. coli shares 96~100%, 90~94%, and 45% identity with Shigella, Salmonella, and Pseudomonas strains, respectively. The ompA gene was cloned from the genomic DNA of E. coli, and then the OmpA protein was expressed in BL21 (DE3) using the auto-induction method. The recombinant OmpA (rOmpA) protein had an average molecular weight of 36 kDa with the purity of 93.5%. Immunological analysis indicated that the titers of anti-rOmpA sera against rOmpA and whole cells were 1:642,000 and 1:140,000, respectively. Moreover, rOmpA not only conferred a high level of immunogenicity to protect mice against the challenge of E. coli, but also generated cross-protection against Shigella and Salmonella. The anti-rOmpA sera could enhance the phagocytic activity of neutrophils against E. coli. The survive ratios of mice immunized with rOmpA and PBS were 50% and 20% after 48 h post-challenge, indicating mice were protected from E. coli infection after immunization with rOmpA. All these results clearly indicate that rOmpA may be a promising candidate for the development of a subunit vaccine to prevent E. coli infection.

Escherichia coli outer membrane protein F (OmpF): an immunogenic protein induces cross-reactive antibodies against Escherichia coli and Shigella

“…The positive transformants were cultured for 24 h at 30 °C in ZYM-5052 auto-inducing media (300 mL in the 1 L shaking flask, 100 μg/mL kanamycin) (Studier 2005; Guan et al 2015). The positive transformant was cultured in LB medium on a platform shaker (37 °C, 250 rpm) to an optical density at 600 nm (OD 600 nm ) of 0.4–0.6.…”

“…The rOmpF protein was purified by Ni 2+ -NTA affinity chromatography and refolded according to the previous methods (Guan et al 2015; Saleem et al 2012). Briefly, the cell lysis solution was loaded onto a Ni 2+ -nitriloacetate (NTA) resin column (QIAGEN, Germany), which was pre-equilibrated with 20 mM Tris–HCl buffer [pH 7.9, containing 0.5 M NaCl, 0.1% (v/v) LDAO and 40 mM imidazole].…”

“…The titers or capacities of antisera against rOmpF and bacteria ( E. coli CVCC 1515, E. coli CICC 21530, S. dysenteriae CMCC 51252, S. flexneri CMCC 51571, S. enteritidis CVCC 3377, S. pullorum CVCC 503, P. aeruginosa CICC 21630, and P. aeruginosa CICC 10419) were measured by iELISA (Guan et al 2015; Hu et al 2010). rOmpF was dissolved in coating buffer (pH 9.6, 0.015 M sodium carbonate, 0.035 M sodium bicarbonate).…”

“…Murine peritoneal macrophages cells were isolated as previously described and adjusted to 4 × 10 6 CFU/mL (Guan et al 2015; Zhang et al 2008). Briefly, after incubation with 100 μL of anti-rOmpF sera or anti-PBS sera at 37 °C for 30 min, 400 μL of E. coli CVCC 1515 cells (4 × 10 6 CFU/mL) were incubated with 500 μL of macrophage suspension and 100 μL of baby rabbit complement (Cedarlane, Hornby, ON, Canada) at 30 °C for 1 h. Macrophages were then lysed by adding sterile water into the mixture (Rennermalm et al 2001; Xu et al 2011; Gressler et al 2016).…”

A Universal Vaccine against Leptospirosis: Are We Going in the Right Direction?

“…and serovars would appear to be farfetched. However, some progress has been made with other pathogens such as influenza (13, 14), dengue (15), and others (16, 17). This perspective focuses on current advances, limitations, possible solutions, and looks forward to the possibility of a universal leptospirosis vaccine.…”