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Trueperella pyogenes is an opportunistic pathogen that causes suppurative infections in animals including humans. Data on phenotypic and genotypic properties of T. pyogenes isolated from ruminants, particularly goats and sheep, are lacking. We characterized, by phenotypic and genotypic means, T. pyogenes of caprine and ovine origin, and established their phylogenetic relationship with isolates from other ruminants. T. pyogenes isolates ( n = 50) from diagnostic specimens of bovine ( n = 25), caprine ( n = 19), and ovine ( n = 6) origin were analyzed. Overall, variable biochemical activities were observed among the T. pyogenes isolates. The fimbriae-encoding gene, fimE, and neuraminidase-encoding gene, nanH, were, respectively, more frequently detected in the large ( p = 0.0006) and small ( p = 0.0001) ruminant isolates. Moreover, genotype V ( plo/ nanH/ nanP/ fimA/ fimC) was only detected in the caprine and ovine isolates, whereas genotype IX ( plo/ nanP/ fimA/ fimC/ fimE) was solely present in the isolates of bovine origin ( p = 0.0223). The 16S rRNA gene sequences of all T. pyogenes isolates were clustered with the reference T. pyogenes strain ATCC 19411 and displayed a high degree of identity to each other. Our results highlight phenotypic and genotypic diversity among ruminant isolates of T. pyogenes and reinforce the importance of characterization of more clinical isolates to better understand the pathogenesis of this bacterium in different animal species.

Author ContributionsThe manuscript was written through contributions of all authors. NDT, LCP, MRK1 (Karver), and MDS designed TF-targeted peptide sequences. MKK performed all conventional TEM. MKK conceived and designed the rat experiments and performed the surgeries. HAK assisted with the rat hemorrhage model. MKK and RHL conceived and designed the mouse experiments, with laser injury performed by RHL. DCG and BRD provided assistance with tissue processing and handling. MKK conceived, designed, and performed the TEG experiments. DCG provided additional assistance with TEGs. MDS and TDC performed CD spectroscopy. TDC performed cryo-TEM, SAXS, and WAXS. MRK1 synthesized all PAs. WB, EBP, LCP, NDT, TAP, SIS, MRK2 (Kibbe) helped guide the research. MKK, NDT, MRK2 interpreted all results. EBP assisted with statistical analysis. EBP and JRR performed CAC measurements. MKK prepared the initial draft of the manuscript. NDT and MRK2 critically revised the manuscript. MRK2, SIS, and BG provided oversight and funding of the entire project. All authors have given approval to the final version of the manuscript. Supporting InformationA table of the three letter codes, amino acid sequences, and corresponding Factor VII residues for the peptides incorporated into PA molecules; crystal structure models of the putative interaction sites of the targeting peptides on TF; HPLC-MS traces showing purity of synthesized PAs; cryogenic TEM of 25% SFE and 75% SBC-2 PA nanofibers; WAXS analysis of backbone, 25% SFE, and 75% SBC-2 PA nanofibers; CD spectroscopy of FKD and TQD PA nanofibers; fluorescent quantification of tested ratios of SFE and SBC-2 PA nanofibers; the critical aggregation concentration determinations for the 75% SBC-2, 25% SFE, and backbone PAs; and real-time localization of 75% SBC-2 in a mouse laser injury model. This material is available free of charge via the internet at http:// pubs.acs.org.

Lyme disease (LD), the most prevalent tick-borne illness in North America, is caused by Borrelia burgdorferi. The long-term survival of B. burgdorferi spirochetes in the mammalian host is achieved though VlsE-mediated antigenic variation. It is mathematically predicted that a highly variable surface antigen prolongs bacterial infection sufficiently to exhaust the immune response directed toward invariant surface antigens. If the prediction is correct, it is expected that the antibody response to B. burgdorferi invariant antigens will become nonprotective as B. burgdorferi infection progresses. To test this assumption, changes in the protective efficacy of the immune response to B. burgdorferi surface antigens were monitored via a superinfection model over the course of 70 days. B. burgdorferi-infected mice were subjected to secondary challenge by heterologous B. burgdorferi at different time points postinfection (p.i.). When the infected mice were superinfected with a VlsEdeficient clone (ΔVlsE) at day 28 p.i., the active anti-B. burgdorferi immune response did not prevent ΔVlsE-induced spirochetemia. In contrast, most mice blocked culture-detectable spirochetemia induced by wild-type B. burgdorferi (WT), indicating that VlsE was likely the primary target of the antibody response. As the B. burgdorferi infection further progressed, however, reversed outcomes were observed. At day 70 p.i. the host immune response to non-VlsE antigens became sufficiently potent to clear spirochetemia induced by ΔVlsE and yet failed to prevent WT-induced spirochetemia. To test if any significant changes in the anti-B. burgdorferi antibody repertoire accounted for the observed outcomes, global profiles of antibody specificities were determined. However, comparison of mimotopes revealed no major difference between day 28 and day 70 antibody repertoires.