Differential innate immune responses induced by Mycoplasma hyopneumoniae and Mycoplasma hyorhinis in various types of antigen presenting cells.

Mycoplasma (M.) hyopneumoniae is the etiological agent of enzootic pneumonia in pigs and is closely related to M. hyorhinis, which can be isolated from the healthy mucosal surfaces of the upper respiratory tract. In rare cases it can also cause arthritis and polyserositis. Since the innate immune system is an important first line of defense and promotes adaptive immune responses, we characterized the innate immune response of various antigen presenting cells (APCs) to M. hyopneumoniae and M. hyorhinis, which differ in their pathogenicity in vivo. Porcine peripheral blood mononuclear cells were infected with different multiplicities of infection (MOI) of live and inactivated porcine mycoplasmas. Both Mycoplasma species induced strong tumour necrosis factor (TNF) responses in monocytes, with a stronger activation by M. hyorhinis. This higher stimulatory activity was also confirmed for CD40 upregulation. Conventional and plasmacytoid dendritic cells (cDC and pDC, respectively) did not or poorly respond to mycoplasmas in terms of TNF expression but more efficiently in terms of CD40 upregulation. Again, these responses were generally stronger with M. hyorhinis than with M. hyopneumoniae. Both Mycoplasma species also activated B cells in terms of CD25 upregulation, proliferation, and IgM secretion. Interestingly, while the induction of CD25 and in particular proliferation was higher with M. hyorhinis, the IgM secretion did not differ between the two species with the exception of the highest dose of M. hyopneumoniae,which appeared to suppress IgM responses. Taken together, our results provide a comparative analysis of innate immune response with different porcine APCs and demonstrate Mycoplasma species-dependent differences, which could relate to their different pathogenicity in vivo.

Erratum: Author Correction: System immunology-based identification of blood transcriptional modules correlating to antibody responses in sheep.

[This corrects the article DOI: 10.1038/s41541-018-0078-0.].

System immunology-based identification of blood transcriptional modules correlating to antibody responses in sheep.

Inactivated vaccines lack immunogenicity and therefore require potent adjuvants. To understand the in vivo effects of adjuvants, we used a system immunology-based analysis of ovine blood transcriptional modules (BTMs) to dissect innate immune responses relating to either antibody or haptoglobin levels. Using inactivated foot-and-mouth disease virus as an antigen, we compared non-adjuvanted to liposomal-formulated vaccines complemented or not with TLR4 and TLR7 ligands. Early after vaccination, BTM relating to myeloid cells, innate immune responses, dendritic cells, and antigen presentation correlated positively, whereas BTM relating to T and natural killer cells, as well as cell cycle correlated negatively with antibody responses. Interestingly, similar BTM also correlated with haptoglobin, but in a reversed manner, indicating that acute systemic inflammation is not beneficial for early antibody responses. Analysis of vaccine-dependent BTM modulation showed that liposomal formulations induced similar responses to those correlating to antibody levels. Surprisingly, the addition of the TLR ligands appeared to reduce early immunological perturbations and mediated anti-inflammatory effects, despite promoting antibody responses. When pre-vaccination BTM were analyzed, we found that high vaccine responders expressed higher levels of many BTM relating to cell cycle, antigen-presenting cells, and innate responses as compared with low responders. In conclusion, we have transferred human BTM to sheep and identified early vaccine-induced responses associated with antibody levels or unwanted inflammation in a heterogeneous and small group of animals. Such readouts are applicable to other veterinary species and very useful to identify efficient vaccine adjuvants, their mechanism of action, and factors related to low responders.

Porcine B Cell Subset Responses to Toll-like Receptor Ligands.

Toll-like receptors (TLR) triggering of B cells are known to promote B cell expansion, differentiation of B cells into antibody-producing and memory cells, but the TLR responses of porcine B cells is poorly characterized. Therefore, this study investigated the response pattern of porcine B cell subsets to a large collection of TLR ligands and demonstrates that the TLR2 ligand Pam3Cys-SK4 and the TLR7/8 ligands gardiquimod and resiquimod are particularly efficient at inducing proliferation, CD25 and CCR7. This activation was also determined in B-cell subpopulations including a CD21+IgM+ subset, an IgG+ subset and two putative B1-like subsets, defined as CD21-IgMhighCD11R1+CD11c+CD14+ and CD21-IgMhigh CD11R1-CD11c+CD14- B cells. The latter two were larger and expressed higher levels of CD80/86 and spontaneous phospholipase C-γ2 phosphorylation. All porcine B-cell subsets were activated by TLR2, TLR7, and TLR9 ligands. Naïve and memory conventional B cells responded similar to TLR ligands. The CD11R1+ B1-like subset had the highest proliferative responses. While both B1-like subsets did not spontaneously secrete IgM, they were the only subsets to produce high level of TLR-induced IgM. Similar to polyclonal IgM responses, memory B cells were efficiently induced to produce specific antibodies by CpG oligodinucleotide, resiquimod, and to a weaker extend by Pam3Cys-SK4. Depletion of plasmacytoid dendritic cells (pDCs) enhanced TLR-induced antibodies. The same set of TLR ligands also induced CD40 on cDCs, pDCs, and monocytes with the exception of TLR4 ligand being unable to activate pDCs. Gardiquimod and resiquimod were particularly efficient at inducing CCR7 on pDCs. Porcine B cells expressed high levels of TLR7, but relatively little other TLR mRNA. Nevertheless, TLR2 on B cells was rapidly upregulated following stimulation, explaining the strong responses following stimulation. Subset-specific analysis of TLR expression demonstrated a comparable expression of TLR2, TLR7, and TLR9 in all B cell subsets, but TLR3 was restricted to B1-like cells, whereas TLR4 was only expressed on conventional B cells, although both at low levels. Altogether, our data describe porcine innate B1-like cells, and how different B cell subsets are involved in innate sensing.