Next-Generation sequencing has become an essential tool in the analysis of antibody responses in the settings of health, vaccination, and disease. However, immune receptors comprise two chains encoded by separate mRNA strands, and conventional NextGen sequencing fails to identify the native pairings encoded by individual lymphocytes. To overcome this limitation we have applied recent technical advances in high-throughput sequencing and functional analysis of complete antibodies (i.e., paired heavy and light chain sequencing) to generate a comprehensive understanding of the antibody response to vaccination and natural infection. Here we present a new technology to screen natively-paired human antibody repertoires from millions of B cells. Libraries of natively-paired variable region heavy and light (VH:VL) amplicons were expressed in a yeast display platform that was optimized for human Fab surface expression, and the resulting libraries were interrogated for binding to viral vaccine antigens via FACS paired with next generation sequencing. Using our method we identified HIV-1 broadly neutralizing antibodies (bNAbs) from an HIV-1 slow progressor and high-affinity neutralizing antibodies responding to an Ebola virus glycoprotein vaccination. These next-generation approaches are providing detailed molecular feedback on immune receptor responses and are informing the design and discovery of new vaccines and therapeutics.
Catheter-associated urinary tract infections (CAUTIs) contribute greatly to the burden of healthcare associated infections. Acinetobacter baumannii is a Gram-negative bacterium with high levels of antibiotic resistance that is of increasing concern as a CAUTI pathogen. A. baumannii expresses fibrinogen-binding adhesins (Abp1D and Abp2D) that mediate colonization and biofilm formation on catheters, which become coated with fibrinogen upon insertion. We developed a protein subunit vaccine against Abp1DRBD and Abp2DRBD and showed that vaccination significantly reduced bladder bacterial titers in a mouse model of CAUTI. We then determined that immunity to Abp2DRBD alone was sufficient for protection. Mechanistically, we defined the B cell response to Abp2DRBD vaccination and demonstrated that immunity was transferrable to naïve mice through passive immunization with Abp2DRBD-immune sera. This work represents a novel strategy in the prevention of A. baumannii CAUTI and has an important role to play in the global fight against antimicrobial resistance.
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