Transmission of Plasmodium falciparum malaria parasites requires formation and development of gametocytes, yet all but the most mature of these sexual parasite forms are absent from the blood circulation. We performed a systematic organ survey in pediatric cases of fatal malaria to characterize the spatial dynamics of gametocyte development in the human host. Histological studies revealed a niche in the extravascular space of the human bone marrow where gametocytes formed in erythroid precursor cells and underwent development before reentering the circulation. Accumulation of gametocytes in the hematopoietic system of human bone marrow did not rely on cytoadherence to the vasculature as does sequestration of asexual-stage parasites. This suggests a different mechanism for the sequestration of gametocytes that could potentially be exploited to block malaria transmission.

A collection of Enterococcus faecalis strains from clinical isolates, healthy individuals and the environment was screened for the presence of virulence factor genes, such as those for collagenbinding protein (ace), endocarditis antigen (efaA), haemolysin activator (cylA), gelatinase (gelE), aggregation substances (asa1 and asa373), a surface protein (esp) and two novel putative surface antigens (EF0591 and EF3314). Apart from some genes that were present in all strains (ace, efaA and EF3314), the gelE gene was the most common factor, although its presence did not correlate with its expression. The genes that encode Esp and CylA were never detected in endocarditis isolates, whereas an association was noted between the esp gene and isolates from urinary tract infection (UTI) and bacteraemia. An aggregation substance gene was always present in commensal strains. As for gelatinase, the presence of the cylA and asa genes did not correlate completely with their phenotypic expression. Generally, isolates from endocarditis, biliary stents and the environment were equipped with fewer virulence factors than isolates from other sources. UTI strains possessed the highest number of factors.

SummaryMale gametogenesis occurs directly after uptake of malaria parasites by the mosquito vector and leads to the release of eight nucleated flagellar gametes. Here, we report that one of the two parasite actin isoforms, named actin II, is essential for this process. Disruption of actin II in Plasmodium berghei resulted in viable asexual blood stages, but male gametogenesis was specifically inhibited. Upon activation, male gametocyte DNA was replicated normally and axonemes assembled, but egress from the host cell was inhibited, and axoneme motility abolished. The major actin isoform, actin I, displayed dual localization to the cytoplasm and the nucleus in male gametocytes. After activation actin I was found to be restricted to the cytoplasm. In actII(-) mutant parasites, this re-localization was abolished and actin I remained in both cellular compartments. These findings reveal vital and pleiotropic functions for the actin II isoform in male gametogenesis of the malaria parasite.

SummaryMalaria parasites invade erythrocytes of their host both for asexual multiplication and for differentiation to male and female gametocytes -the precursor cells of Plasmodium gametes. For further development the parasite is dependent on efficient release of the asexual daughter cells and of the gametes from the host erythrocyte. How malarial parasites exit their host cells remains largely unknown. We here report the characterization of a Plasmodium berghei protein that is involved in egress of both male and female gametes from the host erythrocyte. Protein MDV-1/PEG3, like its Plasmodium falciparum orthologue, is present in gametocytes of both sexes, but more abundant in the female, where it is associated with dense granular organelles, the osmiophilic bodies. Dmdv-1/peg3 parasites in which MDV-1/PEG3 production was abolished by gene disruption had a strongly reduced capacity to form zygotes resulting from a reduced capability of both the male and female gametes to disrupt the surrounding parasitophorous vacuole and to egress from the host erythrocyte. These data demonstrate that emergence from the host cell of male and female gametes relies on a common, MDV-1/PEG3-dependent mechanism that is distinct from mechanisms used by asexual parasites.

Natural killer (NK) cells contribute to immunosurveillance and first-line defense in the control of tumor growth and metastasis diffusion. NK-cell-derived extracellular vesicles (NKEVs) are constitutively secreted and biologically active. They reflect the protein and genetic repertoire of originating cells, and exert antitumor activity in vitro and in vivo. Cancer can compromise NK cell functions, a status potentially reflected by their extracellular vesicles. Hence, NKEVs could, on the one hand, contribute to improve cancer therapy by interacting with tumor and/or immune cells and on the other hand, sense the actual NK cell status in cancer patients. Here, we investigated the composition of healthy donors' NKEVs, including NK microvesicles and exosomes, and their interaction with uncompromised cells of the immune system. To sense the systemic NK cell status in cancer patients, we developed an immune enzymatic test (NKExoELISA) that measures plasma NK-cell-derived exosomes, captured as tsg101 + CD56 + nanovesicles. NKEV mass spectrometry and cytokine analysis showed the expression of NK cell markers, i.e., NKG2D and CD94, perforin, granzymes, CD40L, and other molecules involved in cytotoxicity, homing, cell adhesion, and immune activation, together with EV markers tsg101, CD81, CD63, and CD9 in both NK-derived exosomes and microvesicles. Data are available via Proteome Xchange with identifier PXD014894. Immunomodulation studies revealed that NKEVs displayed main stimulatory functions in peripheral blood mononuclear cells (PBMCs), inducing the expression of human leukocyte antigen DR isotype (HLA-DR) and costimulatory molecules on monocytes and CD25 expression on T cells, which was maintained in the presence of lipopolysaccharide (LPS) and interleukin (IL)-10/transforming growth factor beta (TGFβ), respectively. Furthermore, NKEVs increased the CD56 + NK cell fraction, suggesting that effects mediated by NKEVs might be potentially exploited in support of cancer therapy. The measurement of circulating NK exosomes in the plasma of melanoma patients and healthy donors Federici et al.NKEVs: Immune Sensors and Interactors evidenced lower levels of tsg101 + CD56 + exosomes in patients with respect to donors. Likewise, we detected lower frequencies of NK cells in PBMCs of these patients. These data highlight the potential of NKExoELISA to sense alterations of the NK cell immune status.