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Human infections by type B influenza virus constitute about 25% of all influenza cases. The viral hemagglutinin is comprised of two subunits, HA1 and HA2. While HA1 is constantly evolving in an unpredictable fashion, the HA2 subunit is highly conserved, making it a potential candidate for a universal vaccine. However, immunodominant epitopes in the HA2 subunit remain largely unknown. To delineate MHC Class I epitopes, we first identified 9-mer H-2Kd-restricted CD8 T cell epitopes in the HA2 domain by in silico analyses, followed by evaluating the immunodominance of these peptides in mice challenged with the virus. Of three peptides selected through in silico analysis, the universally conserved peptide, YYSTAASSL (B/HA2-190), possessed the highest predicted binding affinity to MHC Class I and was most effective in inducing IL-2 and TNF-α in mouse splenocytes. Importantly, the peptide demonstrated best capability of stimulating peptide-specific ex-vivo cytotoxicity against target cells. Taken together, this finding would be of value for assessment of cell-mediated immune responses elicited by vaccines based on the highly conserved HA2 stalk domain.

Chlamydia trachomatis is an important human pathogen causing both ocular and sexually transmitted disease. Recently, we identified CT135 as an important virulence determinant in a mouse infection model. Results from CEL 1 digestion assays and sequencing analyses indicated that CT135 was much more polymorphic in high in vitro passage reference serovars than it was in clinical strains that had undergone limited passaging. Herein, we used targeted next-generation sequencing of the CT134-135 locus, from reference strains and clinical isolates, enabling accurate discovery of single nucleotide polymorphisms and other population genetic variations. Our results indicate that CT134 is stable in all C. trachomatis serovars examined. In contrast, CT135 is highly polymorphic in high-passaged reference ocular and non-LGV genital serovars, with the majority of the mutations resulting in gene disruption. In low-passaged ocular clinical isolates, CT135 was frequently disrupted, whereas in genital clinical isolates CT135 was intact in almost all instances. When a serovar K isolate, with an intact CT134 and CT135, was subjected to serial passage in vitro CT134 remained invariable, while numerous gene interrupting mutations rapidly accumulated in CT135. Collectively, our data indicate that, for genital serovars, CT135 is under strong positive selection in vivo, and negative selection in vitro.