A new species of Versteria (Cestoda: Taeniidae) parasitizing Neogale vison and Lontra canadensis (Carnivora: Mustelidae) from Western Canada.

Via molecular and morphological analyses, we describe adult specimens of a new species of Versteria (Cestoda: Taeniidae) infecting mink and river otter (Carnivora: Mustelidae) in Western Canada, as well as larval forms from muskrat and mink. These sequences closely matched those reported from adult specimens from Colorado and Oregon, as well as larval infections in humans and a captive orangutan. We describe here a new species from British Columbia and Alberta (Canada), Versteria rafei n. sp., based upon morphological diagnostic characteristics and genetic distance and phylogeny. Versteria rafei n. sp. differs from the three other described species of the genus in the smaller scolex and cirrus sac. It also differs from V. mustelae (Eurasia) and V. cuja (South America) by having an armed cirrus, which is covered in hair-like bristles, and in the shape of its hooks, with a long thorn-like blade, and short or long handle (vs. a short sharply curved blade and no difference in handle size in previously described species). The poorly known V. brachyacantha (Central Africa) also has an armed cirrus and similarly shaped hooks. However, it differs from the new species in the number and size of hooks. Phylogenetic analysis of the cox1 and nad1 mitochondrial regions showed that our specimens clustered with isolates from undescribed adults and larval infections in North America, and separate from V. cuja, confirming them to be a distinct species from the American Clade.

Habitat selection and populations of Corynosoma (Acanthocephala) in the intestines of sea otters (Enhydra lutris) and seals.

Acanthocephalans are common intestinal parasites of marine mammals, the most widespread of which is the genus Corynosoma. In this study, parasite infrapopulations of two closely related species of Corynosoma were examined: Corynosoma enhydri from sea otters (Enhydra lutris) in Alaska (n = 12) and California (n = 19), and Corynosoma strumosum from seals in Germany (n = 22). Prevalence of C. enhydri was 100% in Californian otters, with a mean abundance of 30, and 83% in Alaskan otters, with a mean abundance of 232. In seals, C. strumosum had a prevalence of 65%, with a mean abundance of 33. Female C. enhydri dominated both Californian (82%) and Alaskan (79%) infections, while, in seals, female C. strumosum made up 68% of the parasite population. Reproduction rates for C. enhydri, with 16% (California) and 18% (Alaska) of females mated, were low compared to C. strumosum in seals, of which 40% of females were mated. Habitat selection also differed significantly between the two species. Corynosoma enhydri was found most frequently in the second and third fifths of the small intestine, while C. strumosum was found most frequently in the fourth. The differences in habitat selection and prevalence analysed in this study may be related to a trade-off between growth and reproduction between the two species.

Missing link: California rock crabs serve as intermediate hosts for the parasite Helicometrina nimia.

Parasites can have strong effects on invertebrate host behaviour, fecundity and survival in marine ecosystems. However, parasites are often poorly documented and still rarely integrated into marine ecological modelling; comprehensive surveys of infection in marine invertebrates are sporadic at best. For example, rock crabs are an important part of Californian coastal ecosystems, both as regulators of mussel populations and non-native species, and as prey items for predators like sea otters, but their parasite communities and potential effects on crab population dynamics are seldom studied or understood. Here, we present the first report of infection by the trematode Helicometrina nimia in the economically and ecologically important red rock crab (Cancer productus) and Pacific rock crab (Romaleon antennarium). As intermediate hosts, they are a missing link for infection by H. nimia in Californian fish that was unreported until now. Based on these findings, we advocate for further research into parasite diversity and their potential effects on ecologically and commercially important species.

IFN-gamma is a potent inducer of Ig secretion by sort-purified murine B cells activated through the mIg, but not the CD40, signaling pathway.

IFN-gamma has been shown to either stimulate or inhibit Ig secretion. No studies have yet addressed the basis for these seemingly conflicting properties nor whether IFN-gamma acted directly at the level of the B cell to mediate its effects. Thus, we studied the ability of IFN-gamma to regulate Ig secretion in sort-purified, resting murine B cells that were >99% Ig+, activated either through membrane Ig using unconjugated or dextran-conjugated anti-IgD antibodies (alphadelta-dex) or through CD40 using soluble or membrane CD40 ligand (CD40L). B cells activated with alphadelta-dex proliferated but do not secrete Ig, even in the presence of IL-1 + IL-2. We demonstrate that IFN-gamma only when added subsequent to B cell stimulation with alphadelta-dex, but not unconjugated anti-IfD antibody, plus IL-1 + IL-2 induces up to 100-fold enhancements in Ig secretion and in the numbers of Ig-secreting cells. The predominant Ig isotype secreted is IgM, with IgG3 and IgG2a comprising the majority of non-IgM antibody. IFN-gamma must act in concert with IL-2 for stimulation of Ig secretion. Further, IFN-gamma synergizes with IL-3 + granulocyte-macrophage colony stimulating factor for induction of Ig synthesis. IFN-gamma also enhances IgA syntheses by transforming growth factor-beta-induced membrane IgA+ cells. By contrast, 125IIFN-gamma fails to stimulate Ig secretion in B cells activated with CD40L in the presence or absence of IL-1 + IL-2 or IL-4. However, the combination of CD40L and alphabeta-dex is strongly synergistic for IFN-gamma-induced Ig secretion. Thus, these data establish that IFN-gamma can act directly on the B cell to induce Ig synthesis without the participation of any other cell and demonstrates that the mode of activation of the B cell plays an important role in directing the action of IFN-gamma.

Regulation of murine B cell growth and differentiation by CD30 ligand.

A ligand for CD30 has been recently cloned, and has been shown to have sequence homology with the tumor necrosis factor family of cytokines. CD30 ligand (CD30L) was found to be induced on helper T cell clones, and its receptor was expressed on freshly isolated and activated murine B cells. Recombinant murine CD30L was found to share many functional properties with CD40 ligand (CD40L) in the regulation of murine B cell growth and differentiation in vitro. CD30L stimulated B cell proliferation, antigen-specific antibody production, and polyclonal immunoglobulin secretion in a cytokine-dependent manner. In particular, the stimulation of B cell proliferation by CD30L required interleukin (IL)-4 and IL-5, induction of anti-sheep red blood cell antibody-secreting B cells by CD30L required IL-2 and IL-5, and optimal induction of polyclonal immunoglobulin secretion required IL-4 and IL-5. Under these conditions, the polyclonal secretion of IgG1, IgA, IgG3 and IgE was induced. The induction of immunoglobulin secretion by CD30L was independent of CD40L, as B cells from CD40L deficient-mice responded normally to CD30L treatment. We conclude that CD30L is a potent mediator of B cell growth and differentiation in vitro and may play a role in cognate T cell-B cell interactions.

Identification and cloning of a novel IL-15 binding protein that is structurally related to the alpha chain of the IL-2 receptor.

Interleukin-15 (IL-15) is a novel cytokine of the four-helix bundle family which shares many biological activities with IL-2, probably due to its interaction with the IL-2 receptor beta and gamma (IL-2R beta and gamma c) chains. We report here the characterization and molecular cloning of a distinct murine IL-15R alpha chain. IL-15R alpha alone displays an affinity of binding for IL-15 equivalent to that of the heterotrimeric IL-2R for IL-2. A biologically functional heteromeric IL-15 receptor complex capable of mediating IL-15 responses was generated through reconstruction experiments in a murine myeloid cell line. IL-15R alpha is structurally similar to IL-2R alpha; together they define a new cytokine receptor family. The distribution of IL-15 and IL-15R alpha mRNA suggests that IL-15 may have biological activities distinct from IL-2.

Elevated levels of shed type II IL-1 receptor in sepsis. Potential role for type II receptor in regulation of IL-1 responses.

Two types of cellular IL-1Rs have been characterized and cloned from both human and murine sources. The type II IL-1R has a very short cytoplasmic domain and does not seem to participate in IL-1 signaling. We demonstrate that type II IL-1Rs are released from the surface of neutrophils in response to treatment with TNF or endotoxin. In addition, serum from patients with sepsis syndrome contains elevated levels of soluble type II IL-1Rs. Neutrophils isolated from patients with sepsis have greatly enhanced expression of type II IL-1R mRNA and cell surface receptors and are therefore a likely source for the shed receptors in serum. Of the three forms of IL-1, soluble type II IL-1R binds IL-1 beta with highest affinity and also selectively inhibits IL-1 beta activity. We propose that increased cell surface expression and rapid release of preformed type II IL-1R from neutrophils, as a soluble IL-1 beta binding protein, represents a mechanism that has evolved for regulating IL-1 activity in sepsis.

Interleukin-12 regulates the proliferation of Th1, but not Th2 or Th0, clones.

Our results indicate that interleukin (IL)-12 is an important costimulator of antigen-dependent proliferation of murine Th1 clones. In addition, we demonstrate that IL-10 inhibits splenic antigen-presenting cell (APC)-dependent proliferation of Th1 clones, at least in part, via down-regulation of APC-derived IL-12. Moreover, the failure of activated B cells to provide costimulation via IL-12 accounts for their inability to support optimal proliferative responses of Th1 clones. We also show that IL-12 regulates the ability of Th1 clones to respond to IL-4 and enhances their proliferation in response to IL-2, IL-7, or IL-15. In contrast, Th2 and Th0 clones appear refractory to the effects of IL-12, on antigen-dependent or growth factor-induced proliferation.

Utilization of the beta and gamma chains of the IL-2 receptor by the novel cytokine IL-15.

We have recently cloned a novel cytokine, IL-15, with shared bioactivities but no sequence homology with IL-2. We found high affinity IL-15 binding to many cell types, including cells of non-lymphoid origin. Analysis of IL-15 interaction with subunits of the IL-2 receptor (IL-2R) revealed that the alpha subunit was not involved in IL-15 binding. We demonstrated directly in cells transfected with IL-2R subunits that both the beta and gamma chains are required for IL-15 binding and signaling. Hence, IL-15, like IL-2, IL-4 and IL-7, utilizes the common IL-2R gamma subunit found to be defective in X-linked severe combined immunodeficiency in humans. IL-15 is the only cytokine other than IL-2 that has also been shown to share the beta signaling subunit of IL-2R. The differential ability of some cells to bind and respond to IL-2 and IL-15 implies the existence of an additional IL-15-specific component.

Cloning of a T cell growth factor that interacts with the beta chain of the interleukin-2 receptor.

A cytokine was identified that stimulated the proliferation of T lymphocytes, and a complementary DNA clone encoding this new T cell growth factor was isolated. The cytokine, designated interleukin-15 (IL-15), is produced by a wide variety of cells and tissues and shares many biological properties with IL-2. Monoclonal antibodies to the beta chain of the IL-2 receptor inhibited the biological activity of IL-15, and IL-15 competed for binding with IL-2, indicating that IL-15 uses components of the IL-2 receptor.

Induction of B cell costimulatory function by recombinant murine CD40 ligand.

T cell-dependent regulation of B cell growth and differentiation involves an interaction between CD40, a B cell surface molecule, and the CD40 ligand (CD40L) which is expressed on activated CD4+ T cells. In the current study, we show that recombinant membrane-bound murine CD40L induces B cells to express costimulatory function for the proliferation of CD4+ T cells. CD40L- or lipopolysaccharide (LPS)-activated, but not control-cultured B cells were strong costimulators of anti-CD3 or alloantigen-dependent T cell responses. The molecular interactions responsible for the increased costimulatory functions were examined by analyzing the activated B cells for changes in the expression of two costimulatory molecules, B7 and heat-stable antigen (HSA), as well as by the use of antagonists of B7 and HSA (CTLA4.Fc and 20C9, respectively). The expression of both B7 and HSA was enhanced on B cells activated with LPS. As observed in previous studies, the costimulatory activity of the LPS-activated B cells was dependent on both B7 and HSA and was completely inhibited in the presence of a combination of CTLA4.Fc and 20C9. In contrast, activation of B cells with CD40L induced the expression of B7 but did not enhance the expression of HSA. In addition the costimulatory activity of the CD40L-activated B cells was partially, but not completely, inhibited by the combination of CTLA4.Fc and 20C9. These results demonstrate that CD40L regulates costimulatory function of B cells in part by inducing the expression of B7 and suggest that CD40L-activated B cells express an additional costimulatory activity that is not associated with LPS-activated B cells.

Interleukin 1 signaling occurs exclusively via the type I receptor.

Two receptors for the proinflammatory cytokine interleukin 1 (IL-1) have been cloned and characterized biochemically. While it has been well established that the type I (80-kDa) IL-1 receptor can mediate responses to IL-1, the function of the type II (60-kDa) IL-1 receptor has been unknown. In this manuscript we describe experiments designed to ask whether the type II receptor is capable of delivering a biological signal. We have examined two types of experimental situation: responses to IL-1 in cells which express predominantly the type II receptor, and responses to IL-1 which have been suggested previously in the literature to be mediated by type II receptors. In both situations we find that the responses instead are mediated via type I receptors. A blocking antibody against the type II receptor never inhibits, and in fact sometimes enhances, the responses. We conclude that a very small number of type I receptors is sufficient to mediate all of the actions of IL-1 which we have examined here and that the function of the type II receptor may not be to transduce signals.

The regulation of T cell-dependent antibody formation in vitro by CD40 ligand and IL-2.

Our study demonstrates the central role for the murine CD40 ligand and IL-2 in contact-dependent T cell help for Ag-specific primary antibody responses in vitro. Helper T cell clones were found to express CD40 ligand after activation with CD3 mAb. Membrane-bound recombinant CD40 ligand expressed on fixed CV1/EBNA cells had similar B cell-activating properties as T cell clones that had been activated and then fixed. These activities include the induction of B cell proliferation, induction of polyclonal secretion of multiple Ig isotypes in a cytokine-dependent manner, and induction of Ag-specific antibody responses by purified B cells. The induction of polyclonal Ig secretion by the recombinant CD40 ligand required IL-4 and IL-5 although optimal Ag-specific antibody formation required IL-2. Finally, soluble CD40.Fc inhibited the induction of Ag-specific antibody responses by fixed, activated Th cell clones. The requirement for both CD40 ligand and IL-2 for induction of Ag-specific antibody responses was mediated, in part, by the induction of B cell IL-2R expression by CD40 ligand. We conclude that the interaction of CD40 on B cells with its ligand on activated T cells is an integral event in the early activation of B cells to grow and differentiate to antibody formation.

Regulation of T cell proliferation by IL-7.

The regulation of murine T cell proliferation by IL-7 was investigated. Highly purified resting splenic T cells were induced to proliferate in a short term assay by IL-7 in the presence of the comitogen, Con A. The proliferation of these resting T cells showed both IL-2-dependent and -independent components as determined by the susceptibility of the response to the blocking effects of anti-IL-2 mAb. Furthermore, IL-7 was found to augment the Con A-induced production of IL-2 and expression of IL-2R by resting splenic T cells. In contrast, Con A blasts and long term, Ag-dependent cloned T cells proliferated in response to IL-7 independently of any involvement of IL-2. Finally, differences were observed between IL-7 and IL-6 with regard to the regulation of T cell growth and activation. As with IL-7, IL-6 stimulated resting splenic T cells to proliferate in the presence of comitogen. However, in contrast to IL-7, IL-6 failed to stimulate the proliferation of Con A blasts or T cell clones and did not augment the Con A-induced expression of IL-2R on resting T cells.

Recombinant granulocyte/macrophage colony-stimulating factor activates macrophages to inhibit Trypanosoma cruzi and release hydrogen peroxide. Comparison with interferon gamma.

Recombinant granulocyte/macrophage colony-stimulating factors (rGM-CSF) of mouse and human origins activated macrophages of the homologous species to inhibit the replication of the protozoan parasite T. cruzi. Activation could be induced with 10-100 ng/ml of rMu-GM-CSF, whether it was added before or after uptake of the parasite, in either adherent or suspension cultures. However, the degree of inhibition of parasite replication after exposure to rMu-GM-CSF was not as great as after treatment with rMu-IFN-gamma, and much more rMu-GM-CSF than rMu-IFN-gamma was required to achieve an equivalent antimicrobial effect. These results were mirrored by effects of the cytokines on enhancement of H2O2-releasing capacity in resident mouse peritoneal macrophages. In the latter tests, rMu-IFN-gamma and rHu-TNF-alpha afforded a 44-51-fold enhancement over the untreated control, with a 50% effective concentration (EC50) for rMu-IFN-gamma of approximately 0.05 ng/ml. Using rMu-GM-CSF or rM-CSF, enhancement of H2O2-releasing capacity was 14-15-fold over control, with EC50s of 1 and 14 ng/ml, respectively. However, peak enhancement of macrophage H2O2-releasing capacity was seen at least 24 h earlier with rMu-GM-CSF or rHu-M-CSF than with r-Mu-IFN-gamma or rHu-TNF-alpha. Thus, rMu-GM-CSF and rHu-GM-CSF displayed clear-cut macrophage-activating activity in vitro, but rMu-GM-CSF was less potent and less effective than rMu-IFN-gamma in the tests used.

Purification to homogeneity of B cell stimulating factor. A molecule that stimulates proliferation of multiple lymphokine-dependent cell lines.

Murine B cell stimulating factor 1 (BSF-1) was purified to homogeneity from supernatants of a stimulated thymoma cell line. A protein of 18.4 kD with a unique N-terminal amino acid sequence was identified. BSF-1 had a sp act of at least 3.28 X 10(8) U/mg. In addition to its B cell-stimulatory activity, BSF-1 also stimulated the proliferation of several IL-2- and IL-3-dependent cell lines. We conclude that BSF-1 is both a growth factor and a differentiation factor. Finally, these results also suggest additional biologic properties of BSF-1 on lineages besides B lymphocytes.