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We genetically characterized highly pathogenic avian influenza virus A(H5N6) clade 2.3.4.4b isolates found in Germany in 2017–2018 and assessed pathogenicity of representative H5N5 and H5N6 viruses in ferrets. These viruses had low pathogenicity; however, continued characterization of related isolates is warranted because of their high potential for reassortment.

The main findings of the post-mortem examination of poultry infected with highly pathogenic avian influenza viruses (HPAIV) include necrotizing inflammation and viral antigen in multiple organs. The lesion profile displays marked variability, depending on viral subtype, strain, and host species. Therefore, in this study, a semiquantitative scoring system was developed to compare histopathological findings across a wide range of study conditions. Briefly, the severity of necrotizing lesions in brain, heart, lung, liver, kidney, pancreas, and/or lymphocytic depletion in the spleen is scored on an ordinal four-step scale (0 = unchanged, 1 = mild, 2 = moderate, 3 = severe), and the distribution of the viral antigen in parenchymal and endothelial cells is evaluated on a four-step scale (0 = none, 1 = focal, 2 = multifocal, 3 = diffuse). These scores are used for a meta-analysis of experimental infections with H7N7 and H5N8 (clade 2.3.4.4b) HPAIV in chickens, turkeys, and ducks. The meta-analysis highlights the rather unique endotheliotropism of these HPAIV in chickens and a more severe necrotizing encephalitis in H7N7-HPAIV-infected turkeys. In conclusion, the proposed scoring system can be used to condensate HPAIV-typical pathohistological findings into semiquantitative data, thus enabling systematic phenotyping of virus strains and their tissue tropism.

The immune system is dysfunctional for years after hematopoietic stem cell transplantation (HSCT). A potential cause is an intrinsic B cell deficiency. In a cohort of pediatric HSCT patients few CD27(+) B cells formed after transplantation with the number of CD27(+)IgM(high) cells more affected than class-switched ones. A previously unacknowledged population of CD27(-)IgM(high) cells made up the majority of B cells and this population was also enlarged in healthy children compared to adults. Only a minority of these CD27(-)IgM(high) B cells expressed markers typical for transitional B cells, and the non-transitional CD27(-)IgM(high) cells could be further divided into subpopulations based on their ability to extrude the dye Rhodamine 123 and their expression of CD45RB(MEM55), a glycosylation-dependent epitope. Thus, we define several novel human CD27(-)IgM(high) B cell subpopulations in blood, all of which are present in higher frequencies and numbers in young children and after HSCT than in adults.

Viral entry, compartmentalization and transmission depend on the formation of membrane lipid/protein microdomains concentrating receptors and signalosomes. Dendritic cells (DCs) are prime targets for measles virus (MV) infection, and this interaction promotes immune activation and generalized immunosuppression, yet also MV transport to secondary lymphatics where transmission to T cells occurs. In addition to MV trapping, DC-SIGN interaction can enhance MV uptake by activating cellular sphingomyelinases and, thereby, vertical surface transport of its entry receptor CD150. To exploit DCs as Trojan horses for transport, MV promotes DC maturation accompanied by mobilization, and restrictions of viral replication in these cells may support this process. MV-infected DCs are unable to support formation of functional immune synapses with conjugating T cells and signalling via viral glycoproteins or repulsive ligands (such as semaphorins) plays a key role in the induction of T-cell paralysis. In the absence of antigen recognition, MV transmission from infected DCs to T cells most likely involves formation of polyconjugates which concentrate viral structural proteins, viral receptors and with components enhancing either viral uptake or conjugate stability. Because DCs barely support production of infectious MV particles, these organized interfaces are likely to represent virological synapses essential for MV transmission.

Assembly and budding of enveloped RNA viruses rely on viral matrix (M) proteins and host proteins involved in sorting and vesiculation of cellular cargoes, such as the endosomal sorting complex required for transport (ESCRT). The measles virus (MV) M protein promotes virus-like particle (VLP) production, and we now show that it shares association with detergent-resistant or tetraspanin-enriched membrane microdomains with ebolavirus VP40 protein, yet accumulates less efficiently at the plasma membrane. Unlike VP40, which recruits ESCRT components via its N-terminal late (L) domain and exploits them for particle production, the M protein does this independently of this pathway, as (i) ablation of motifs bearing similarity to canonical L domains did not affect VLP production, (ii) it did not redistribute Tsg101, AIP-1 or Vps4A to the plasma membrane, and (iii) neither VLP nor infectious virus production was sensitive to inhibition by dominant-negative Vps4A. Importantly, transfer of the VP40 L domain into the MV M protein did not cause recruitment of ESCRT proteins or confer sensitivity of VLP release to Vps4A, indicating that MV particle production occurs independently of and cannot be routed into an ESCRT-dependent pathway.

Viral entry, compartmentalization and transmission depend on the formation of membrane lipid/protein microdomains concentrating receptors and signalosomes. Dendritic cells (DCs) are prime targets for measles virus (MV) infection, and this interaction promotes immune activation and generalized immunosuppression, yet also MV transport to secondary lymphatics where transmission to T cells occurs. In addition to MV trapping, DC-SIGN interaction can enhance MV uptake by activating cellular sphingomyelinases and, thereby, vertical surface transport of its entry receptor CD150. To exploit DCs as Trojan horses for transport, MV promotes DC maturation accompanied by mobilization, and restrictions of viral replication in these cells may support this process. MV-infected DCs are unable to support formation of functional immune synapses with conjugating T cells and signalling via viral glycoproteins or repulsive ligands (such as semaphorins) plays a key role in the induction of T-cell paralysis. In the absence of antigen recognition, MV transmission from infected DCs to T cells most likely involves formation of polyconjugates which concentrate viral structural proteins, viral receptors and with components enhancing either viral uptake or conjugate stability. Because DCs barely support production of infectious MV particles, these organized interfaces are likely to represent virological synapses essential for MV transmission.

A screen of cell surface markers differentially expressed during peripheral B cell differentiation identified that the CD45RB epitope detected by the mAb MEM-55 was highly expressed on CD27+ memory B cells and absent on CD27– naïve B cells. IgG+CD27– memory and a previously unacknowledged CD27– population in blood also expressed high levels of CD45RBMEM55. Naïve and memory B cells from tonsils followed the pattern observed in blood, and CD38high B cells had a bimodal expression pattern when analyzed using flow cytometry. No CD38high GC B cells, however, expressed the CD45RBMEM55 epitope when assayed using immunohistochemistry. Rather, CD38highCD45RBMEM55high B cells had a distinct cellular phenotype and were localized outside of GCs. CD45RB epitopes, detected by other antibody clones, were expressed at high levels through B cell differentiation, and no changes in splicing of the CD45RB exon were observed during B cell differentiation. Instead, B cells regulated their expression of the CD45RBMEM55 epitope through site-specific modifications of an O-linked glycochain. CD4+ T cells differentially spliced CD45 but did not vary the glycosylation of the CD45RBMEM55 epitope, and CD8+ cells modified CD45RBMEM55 expression in a similar manner as B cells. Monocytes expressed the CD45RB exon but not the CD45RBMEM55 epitope. As CD45 is a highly expressed tyrosine phosphatase that regulates antigen receptor signaling strength in lymphocytes, we conclude that regulated O-linked glycosylation of CD45RB can be used to follow B cell differentiation and that this regulation may be involved in fine-tuning antigen signaling in the cell.

Transmission of measles virus (MV) to T cells by its early CD150 + target cells is considered to be crucial for viral dissemination within the hematopoietic compartment. Using cocultures involving monocyte-derived dendritic cells (DCs) and T cells, we now show that T cells acquire MV most efficiently from cis -infected DCs rather than DCs having trapped MV ( trans -infection). Transmission involves interactions of the viral glycoprotein H with its receptor CD150 and is therefore more efficient to preactivated T cells. In addition to rare association with actin-rich filopodial structures, the formation of contact interfaces consistent with that of virological synapses (VS) was observed where viral proteins accumulated and CD150 was redistributed in an actin-dependent manner. In addition to these molecules, activated LFA-1, DC-SIGN, CD81, and phosphorylated ezrin-radixin-moesin proteins, which also mark the HIV VS, redistributed toward the MV VS. Most interestingly, moesin and substance P receptor, both implicated earlier in assisting MV entry or cell-to-cell transmission, also partitioned to the transmission structure. Altogether, the MV VS shares important similarities to the HIV VS in concentrating cellular components potentially regulating actin dynamics, conjugate stability, and membrane fusion as required for efficient entry of MV into target T cells.