Plasmodium yoelii: resistance to disease is linked to the mtv-7 locus in BALB/c mice.

We have identified congenic mouse strains that differ dramatically in resistance to infection with the murine malaria parasite Plasmodium yoelii 17X. After infection, BALB/c mice develop severe anemia and a high degree of parasitemia which sometimes results in death. The mtv-7 congenic strain BALB.D2.mlsa, however, develops only a mild degree of anemia and parasitemia. In this paper we describe the course of the disease and discuss the potential role of mtv-7 and linked loci in control of this infection. These mice differ in their response to anemia, which may contribute to their differential susceptibility to disease; however, no influence of mtv-7 reactive T cells was documented.

Host genetics and resistance to acute Trypanosoma cruzi infection in mice: profiles and compartmentalization of IL-2-, -4-, -5-, -10-, and IFN-gamma-producing cells.

Survival of acute Trypanosoma cruzi infection by mice is influenced by genes inside and outside the major histocompatibility complex (MHC) and genes associated with resistance must be expressed in both the MHC and the genetic background or the host will die within a few weeks of infection. Both the levels and the kinetics of cytokine production have also been implicated as important factors for resistance. Antigen-stimulated spleen cells from mice that express the resistant H-2q MHC haplotype produced significantly more interferon (IFN)-gamma than did cells from mice that share the susceptible H-2k haplotype. But, spleen cells from susceptible and resistant mice produce similar levels of IFN-gamma when stimulated with concanavalin A. The kinetics of interleukin (IL)- 10 production by ConA (ConA)-stimulated spleen cell were inversely correlated with IFN-gamma levels throughout the course of acute infection in all mouse strains. Levels of IL-2 produced by ConA-stimulated spleen cells were also initially high (day 0) then decreased as acute infection progressed. Conversely, IL-4 production by ConA-stimulated spleen cells increased during infection, and mice that express the susceptible C3H background produced significantly more IL-4 than those that share the resistant B10 background. IL-2 production by lymph node cells from mice that express the susceptible C3H genetic background also declined during infection, while lymph node cells from B10 background mice showed a moderate increase in IL-2 secretion. These data suggest that both the levels and the kinetics of cytokine production may be genetically regulated and that cytokine responses are compartmentalized in the T. cruzi-infected host.

Novel tyvelose-containing tri- and tetra-antennary N-glycans in the immunodominant antigens of the intracellular parasite Trichinella spiralis.

The larval stage of the intestinal nematode, Trichinella spiralis, secretes and displays on its cuticle a number of antigenically cross-reactive glycoproteins. These so-called TSL-1 antigens induce a powerful antibody response in parasitized animals. In rats, anti-TSL-1 antibodies mediate a protective immunity that expels invading larvae from the intestine. The vast majority of anti-TSL-1 antibodies are specific for glycans. Although the biological functions of TSL-1 antigens are not known, the powerful effect of glycan-specific antibodies on the intestinal survival of T. spiralis suggests that they play an important role in parasite establishment. Little is known about the structures of the glycans present on the TSL-1 glycoproteins. Recent studies have suggested, however, that the antigens contain very unusual glycans (Wisnewski, N., McNeil, M., Grieve, R.B. and Wassom, D.L., Mol. Biochem. Parasitol., 61, 25-36, 1993). Sugar and linkage analysis of the combined secreted products unexpectedly showed that a major terminal sugar is tyvelose (3,6-dideoxy-D-arabino-hexose; Tyv) which has previously been found only in certain gram-negative bacterial lipopolysaccharides. In this paper, we report the first rigorous structural study of oligosaccharides released from TSL-1 antigens by peptide N-glycosidase F digestion. Using strategies based on fast atom bombardment mass spectrometry (FAB-MS), we have discovered a novel family of tri- and tetra-antennary N-glycans whose antennae are comprised of the tyvelose-capped structure: Tyv1,3GalNAc beta 1,4(Fuc alpha 1,3)GlcNAc beta 1-. Thus a major population of TSL-1 glycans contains clusters of hydrophobic terminal structures which are likely to be highly immunogenic.

Characterization of novel fucosyl- and tyvelosyl-containing glycoconjugates from Trichinella spiralis muscle stage larvae.

The monosaccharide composition of an affinity-purified family of antigenically-related Trichinella spiralis larval glycoproteins was determined by gas chromatography/mass spectrometry. This group of 6 major glycoproteins, designated TSL-1, originates in the muscle stage (L1) larval stichosome. They are present on the L1 surface and in excretory/secretory products of L1 larvae, are stage-specific, and are highly immunodominant. The glycosyl composition of the TSL-1 antigens was remarkable in 2 respects: (1) fucose accounted for 36 molar percent of the glycosyl residues; and (2) a 3,6-dideoxyhexose was identified, which accounted for at least 24 molar percent of the glycosyl residues. Previously, 3,6-dideoxyhexoses have been found only in certain Gram-negative bacterial lipopolysaccharides and in ascaroside alcohols (ascarylose) of Ascaris eggs. The 3,6-dideoxyhexose found in the TSL-1 antigens also was found in ES. This Trichinella sugar has been chemically identified as a 3,6-dideoxyarabinohexose, the same as found in Ascaris eggs. However, the absolute configuration of the TSL-1 sugar is D-(tyvelose), not L-(ascarylose) as is found in Ascaris eggs. Methylation analysis indicated that the TSL-1 3,6-dideoxy-D-arabinohexose was present entirely as non-reducing terminal residues. Approximately 83% of the fucose was also present as non-reducing terminal residues, with the remaining fucose found as 3,4-linked branched residues.

Immunoecological succession in host-parasite communities.

From the unique perspective of the parasite, each potential host exists as an ecosystem in its own right, comprised of biotic and abiotic components. If the parasite's limits of tolerance are not exceeded by the host environment, parasitism may occur. The organs of the host can be viewed as integrated communities comprised of distinct cell populations. Each population of cells occupies a unique ecological niche and contributes to the community in an essential way. The coordinated interactions among cells within each community and among the communities themselves are essential for the host to maintain itself in a homeostatic state. As with free-living systems in equilibrium, introduction of an exotic population into the community disrupts the balance, and ecological succession resumes until a new equilibrium is established. Such is the case when parasites invade the host. The successful invader must not exceed the carrying capacity of the environment nor modify the habitat so drastically that its own survival is compromised. The specific interactions that occur between invading parasites and the populations of cells that comprise the host's immune system can also be viewed in ecological terms. The successful parasite must "compete" with cells in the host community for available niches and avoid "predation" by cells of the host immune system. As is true for interactions among organisms in a free-living ecosystem, the outcome of the host-parasite interaction is not readily predicted from knowledge of the component parts in isolation.

Plasmodium yoelii: splenectomy alters the antibody responses of infected mice.

The antibody response to Plasmodium yoelii is altered in splenectomized mice. Sera were obtained from sham-operated or splenectomized DBA/2 and C57BL/6 mice on Days 11, 18, and 24 after infection with nonlethal P. yoelii 17x and used to precipitate metabolically radiolabeled parasite antigens. Mice of both strains responded to many antigens. However, only splenectomized DBA/2 mice made strong antibody responses to antigens of approximately 110, 56, 50, 40, 35, and 20 kDa. Metabolically radiolabeled parasite extracts prepared in sham-operated and splenectomized mice appeared identical on SDS-PAGE. Thus it is unlikely that expression of new parasite antigens in splenectomized DBA/2 mice accounts for these results. Parasite-reactive IgM and IgG antibody responses were also modulated by splenectomy. Levels of IgM increased in splenectomized DBA/2 mice and decreased in C57BL/6 mice. Both mouse strains had slight to moderate increases in IgG when infected after splenectomy. The results suggest that when the spleen is present, responses to specific antigens are markedly suppressed. Alternatively, it is possible that in the absence of a spleen, antigen processing and presentation occurs in other tissues such as the lymph nodes or liver, leading to responses that are qualitatively different than those which occur when the spleen is present.

Plasmodium yoelii: cellular immune responses in splenectomized and normal mice.

Spleen and lymph node cells from Plasmodium yoelii 17X-infected, C57BL/6 (B6), and DBA/2 (D2) mice were cultured in vitro with parasite antigens. The ability of these cells to proliferate was quantified by uptake of [3H]thymidine and ELISA was used to measure secretion of IFN-gamma and IL-5. B6 mice are relatively susceptible to P. yoelii 17X infection compared to D2 mice. Susceptible mouse strains develop higher levels of parasitemia, become more anemic, and take longer to resolve their infections than do resistant strains. Following splenectomy, D2 mice resisted P. yoelii 17X infections as well as did sham-operated controls, but splenectomized B6 mice failed to resolve their infections and all died. Spleen cells from infected mice of either strain were activated in vitro as evidenced by their proliferation in the absence of exogenous antigen. When malaria antigen was added to these cultures, cells from resistant D2 mice responded strongly with increased proliferation, whereas cells from susceptible B6 mice responded weakly, and on Day 14 postinfection, responses were actually suppressed. Mesenteric lymph node cells from infected B6 and D2 mice did not proliferate in the presence or absence of P. yoelii 17X antigen unless the spleen was removed. Following splenectomy, mesenteric lymph node cells from D2 mice, but not B6 mice, proliferated strongly compared to cells from sham-operated controls. IFN-gamma and IL-5 production from spleen and lymph node cells was measured following in vitro stimulation with P. yoelii 17X antigen. Spleen cells from D2 mice produced levels of IFN-gamma increased over those of cells from B6 mice.(ABSTRACT TRUNCATED AT 250 WORDS)

Host genetics and resistance to acute Trypanosoma cruzi infection in mice. I. Antibody isotype profiles.

Some strains of inbred mice survive acute infection with Trypanosoma cruzi while others die within a few weeks after infection. Mice which express B10 background genes and either the H-2q or H-2d haplotypes are resistant and survive. However, mice which share the B10 genetic background but express H-2k alleles die, usually within 4 weeks following infection. These data confirm that at least 1 gene in the major histocompatibility complex can determine whether an animal lives or dies during the acute phase. Expression of the H-2q haplotype on the B10 genetic background or in DBA/1 mice is associated with resistance, but H-2q mice expressing the C3H background are susceptible. Therefore, at least 1 gene in the genetic background also influences resistance. Our data suggest that genes associated with resistance must be present in both the MHC and the genetic background or the animal will die. The isotypes and specificities of parasite reactive antibodies found in the serum of different inbred mouse strains were assessed during acute infection. Levels of IgM were higher in sera from mice which express the resistant B10 background than in sera from mice expressing the susceptible C3H background. Conversely, mice which share the C3H background genes produced high levels of anti-parasite IgG2a when compared to B10 congenic strains. Antigen specificity, however, may be influenced by both background and MHC genes, as congenic strains expressing different MHC haplotypes recognized different constellations of T. cruzi antigens.(ABSTRACT TRUNCATED AT 250 WORDS)

Are antibodies important in mice infected with Plasmodium yoelii?

It is unwise to extrapolate results, even from one mouse strain to another, when attempting to define the mechanisms that control and effect anti-malaria immunity. It is important to better characterize the broad range o f possible responses that are likely to occur when individuals in an outbred population are infected. Here, Peter Sayles and Donald Wossom discuss briefly their views on the role of antibody in murine and human malaria infections, based on their work on mice infected with Plasmodium yoelii.

Influence of resistant and susceptible genotype, IL-1, and lymphoid organ on Trichinella spiralis-induced cytokine secretion.

The relative importance of cell-mediated inflammatory responses and antibody-mediated responses in controlling parasitic helminth infection is debated. To study the relationship between these responses and resistance or susceptibility to primary Trichinella spiralis infection, we infected resistant AKR mice and susceptible B10.BR mice and analyzed the lymphokines IL-2, IFN-gamma, and IL-5 produced by their T cells as a function of time and lymphoid organ. IL-2-secretors occurred maximally between days 3 and 6 postinfection, whereas IL-5-secretors peaked between days 6 and 9. Previously, we found that IFN-gamma producers peaked before day 6, whereas IL-4 producers peaked between days 6 and 9. Most cytokine secretors were CD4+. The simultaneous development of IL-2- and IFN-gamma-secreting cells, and IL-4- and IL-5-secreting cells, suggests that the infection may be stimulating T cells to differentiate into cells capable of secreting specific cytokine sets, analogous to the postulated Th1 and Th2 subsets. In the spleen and mesenteric lymph nodes, cells from B10.BR mice secreted more IL-5 than cells from AKR mice, as we found previously for IL-4. For both strains, mesenteric lymph node cells produced more IL-5 than splenocytes. The AKR mesenteric lymph node cells produced more IL-2 than the B10.BR cells, but the reverse occurred in splenocytes. The AKR peripheral lymph node cells also secreted more IFN-gamma than the B10.BR cells, but the strains were equivalent for peritoneal exudate cell IFN-gamma production. Thus, the lymphoid organ microenvironment plays an important role in regulating cytokine-secreting cell development in this system. We also tested the possible regulatory role of IL-1. Exogenous rIL-1 alpha increased IFN-gamma secretion early but not late in mesenteric lymph node cells from both strains; this reflected an increased IFN-gamma-secreting cell frequency, not a change in IFN-gamma mRNA transcript level. Exogenous rIL-1 alpha did not consistently affect IL-2, IL-4, or IL-5 secretion. These data suggest that IL-1 alpha availability in vivo may regulate IFN-gamma-secreting cell development. In sum, early activation of IFN-gamma-secreting T cells in lymph nodes, with little subsequent activation of IL-4- and IL-5-secreting cells, distinguished the resistant from susceptible strain responses to T. spiralis infection, and IL-1 alpha and lymphoid organ environment influence IFN-gamma-secreting cell activation.

Immunity to Trichinella spiralis infection in vitamin A-deficient mice.

Vitamin A-deficient (A-) mice make strikingly poor IgG responses when they are immunized with purified protein antigens. Previously, we showed that A- T cells overproduce interferon gamma (IFN-gamma), which then could inhibit interleukin 4 (IL-4)-stimulated B cell IgG responses. To determine if the altered IFN-gamma regulation pattern and its immunological consequences would extend to a natural infection, we studied mice infected with the parasitic helminth Trichinella spiralis. The course of the infection was similar in A- and A-sufficient (A+) mice. These mice did not differ with respect to newborn larvae/female/hour produced in the intestine, or muscle larvae burden 5 wk postinfection. They also did not differ in the intestinal worm expulsion rate until day 15, when A- mice still harbored parasites, whereas A+ mice had cleared intestinal worms. Vitamin A deficiency reduced both the frequency of B lymphocytes secreting IgG1 antibodies to parasite antigens, and the bone marrow eosinophilia associated with helminth infection. The cytokine secretion patterns in infected mice were consistent with these observations and with previous studies. Mesenteric lymph node cells from infected A- mice secreted significantly more IFN-gamma, and significantly less IL-2, IL-4, and IL-5 than infected A+ controls. A- splenocytes secreted significantly more IFN-gamma, and equivalent amounts of IL-2, IL-4, and IL-5 compared with A+ controls. Interestingly, CD4-CD8- cells secreted the majority of the IL-4 produced in the spleen. The IL-2, IL-4, and IL-5 steady-state transcript levels correlated with secreted protein levels, but IFN-gamma transcripts did not. Although they secreted more protein, A- cells contained fewer IFN-gamma transcripts than A+ cells. These results suggest two vitamin A-mediated regulation steps in IFN-gamma gene expression: positive regulation of IFN-gamma transcript levels, and negative regulation posttranscriptionally. The essentially unaltered outcome of T. spiralis infection in vitamin A-deficient mice probably reflects a balance between cellular and humoral responses. The IFN-gamma overproduction might have a positive effect on the gut inflammatory response, but the decrease eosinophilia, cytokine production in mesenteric lymph node, and IgG1-secreting cell frequency might have a negative effect on T. spiralis immunity.

Plasmodium yoelii: antibody response in resistant and susceptible mouse strains.

The numbers of antigen-reactive antibody-secreting cells, levels of parasite antigen-specific serum antibodies and numbers of red blood cells staining positive for surface immunoglobulin were determined for susceptible and resistant mouse strains following infection with Plasmodium yoelii 17x. As a control, these parameters also were measured using antigen prepared from normal red blood cells. The relatively susceptible C57BL/6 mice produced more antigen-specific antibody-secreting cells and had higher levels of immunoglobulin positive red blood cells than did DBA/2 mice, but the DBA/2 mice had more antigen-specific IgG in their sera. Both mouse strains possessed cells secreting antibody reactive with soluble normal red blood cell antigen; however, C57BL/6 mice had more IgG positive unparasitized RBC than did DBA/2 mice. Despite possessing fewer antibody positive normal RBC, DBA/2 mice had significantly higher levels of serum antibodies that reacted with soluble red blood cell antigen. These data indicate that levels of serum antibody may not reflect the amounts of antibody produced and that use of any single assay to assess the magnitude of the antibody response may give rise to misleading results.

IFN-gamma- and IL-5-producing cells compartmentalize to different lymphoid organs in Trichinella spiralis-infected mice.

The differential induction of cytokines associated with Th1 and Th2 subsets has recently been described during Trichinella spiralis infection. Increased levels of resistance appear to correlate with elevated levels of the Th1-associated cytokines, IFN-gamma and IL-2. In the present report, a filter immunoplaque assay is used to quantify the actual numbers of cells that secrete IFN-gamma and IL-5. It is demonstrated that, in T. spiralis-infected B10.Q mice, Th1- and Th2-associated responses are compartmentalized to different lymphoid organs. Thus, Ag-induced IFN-gamma-producing cells predominate in the spleen, whereas IL-5-producing cells prevail in the mesenteric lymph nodes (MLN). A corresponding compartmentalization of Ag-specific IgA and IgG1 antibody-secreting cells to the MLN is also noted. The virtual absence of Th1-associated responses in the MLN appears to be an Ag-associated phenomenon. MLN from either naive or T. spiralis-infected mice do have the capacity to secrete IFN-gamma if stimulated with Con A. The striking compartmentalization of Ag-driven cytokine responses seen in this parasite system may facilitate study of the mechanisms that regulate the induction of Th1 and Th2 subsets.

H-2 (I-A) control of the antibody repertoire to secreted antigens of Trichinella spiralis in infection and its relevance to resistance and susceptibility.

Humans infected with the parasitic nematode Trichinella spiralis vary in the specificity of their antibody responses to the antigens of the parasite. The possibility that such host variation in antigen recognition has a genetic basis was examined in infected inbred mice whose antigen recognition profiles were characterized by immunoprecipitation of biosynthetically labelled secreted materials of adult parasites and SDS-PAGE. The strains varied considerably in repertoire and none produced detectable antibody to all the potential antigens. Using a panel of H-2 congenic and recombinant strains it was established that the repertoire was determined by the major histocompatibility complex (MHC), the I-A region in particular. Other factors, such as level of infection and variation between individuals, affected antigen recognition profiles, but this was always within limits imposed by the MHC. Lastly, an attempt to correlate antibody repertoire with relative susceptibility or resistance to T. spiralis failed to reveal any clear association. This also applied to the AKR/J and AKR-Fv-1b strains, which are H-2-identical but differ in a non-MHC susceptibility locus. These findings would argue, therefore, that the I-A region controls the antibody repertoire in this nematode infection but that the repertoire overall has little influence on the efficiency with which the infections are controlled by the immune system. Should this also apply for other nematode infections, then antigen recognition profiles of infected individual humans and domestic animals might not, therefore, be useful indicators of relative resistance or susceptibility to infection.

Trichinella spiralis: influence of an immunodominant, carbohydrate-associated determinant on the host antibody response repertoire.

Host antibody responses to the G2.1 epitope, a carbohydrate-associated determinant shared by several Trichinella spiralis glycoproteins, were examined by competitive inhibition enzyme-linked immunosorbent assay (ELISA). The G2.1 epitope dominated the AKR/J mouse antibody response whether the antigens were injected or introduced through infection, as determined by the serum blocking ability of a G2.1 epitope-specific monoclonal antibody (mAb). Serum T. spiralis-binding activity from several other infected mouse strains was blocked 22 to 86% by the G2.1 epitope-specific mAb. In addition to mice, the G2.1 epitope evoked powerful antibody responses in four other species. The binding activity of Trichinella-reactive antibodies from infected rats and pigs was inhibited 56 and 34%, respectively, by the mAb. Greater than 48% of the T. spiralis serum-binding activity from 4/5 infected humans was G2.1-specific. Most of the rabbit antibody response induced by injection of a previously characterized 43-kDa antigen was also directed to the G2.1 determinant. The specificities of 10 T. spiralis-reactive mAb were examined, and 7 reacted with the immunodominant epitope. Finally, of nine helminth species examined, only T. spiralis and T. pseudospiralis extracts efficiently blocked G2.1-specific antibody binding to solid-phase antigens. These results suggest that the responses to G2.1 epitope may play an important role during infection.

A spleen is not necessary to resolve infections with Plasmodium yoelii.

The role of the spleen in resistance to infections with nonlethal Plasmodium yoelii 17x is dependent upon the genotype of the host. Thus, DBA/2 (D2) mice infected with P. yoelii 17x were not adversely affected by removal of the spleen, while splenectomized C57BL/6 (B6) or Balb/c mice failed to resolve their infections and eventually died. The levels of parasitemia were lower in splenectomized mice compared to intact controls; however, splenectomized mice became as anemic as did spleen-intact controls. Splenectomy resulted in the appearance of large aggregates of mononuclear cells in the livers of infected mice and also altered the liver/body weight ratios. These results indicate that D2 mice have a spleen-independent mechanism of clearing parasites which is lacking in B6 and Balb/c mice.

MHC-restricted antibody responses to Trichuris muris excretory/secretory (E/S) antigen.

Two panels of H-2 recombinant mice were used in a detailed serological study to analyse the role of H-2-linked genes in the control of the antibody response to excretory/secretory (E/S) antigens of Trichuris muris. An apparent H-2q (I-Aq) restriction on the early development of high levels of IgG1 antibody to E/S antigen was revealed by ELISA. No such restriction was demonstrated for the specific IgG2a response patterns. Recognition of two high molecular weight antigens (90-95 kDa, 105-110 kDa) by IgG antibodies was also shown to be almost exclusively H-2q restricted and may be related at least in part to the high antibody levels seen for H-2q strains of mice. Immune serum from resistant (B10.BRxB10.G) F1 hybrid mice (H-2q/k) containing high levels of IgG1 antibodies specific for T. muris E/S and IgG antibodies which recognized the 90-95 kDa and 105-110 kDa E/S antigens was effective in transferring protection to the non-responsive B10.BR mouse strain as seen on day 35 post-infection (p.i.). It is suggested that the IgG responses described for the generally very resistant H-2q mouse strains may contribute to, but not be an absolute requirement for, protective immunity, antibody-mediated damage facilitating a subsequent cellular attack in certain strains of mice.

The influence of genes mapping within the major histocompatibility complex on resistance to Trichuris muris infections in mice.

Two panels of H-2 recombinant strains of mice were used in an attempt to map the H-2-linked genes which control resistance to infection with Trichuris muris. Response phenotypes could be related to the presence of 'resistance' (q,b) or 'susceptibility' (k,d) alleles at I-A. The influence of these genes was modulated by other alleles, particularly q or d alleles, at the D end of the H-2. Absence of I-E molecules correlated with resistance to infection in some but not all strains studied. Thus the (B10.BR x B10.G) F1 strain which expressed I-Ek gene products was resistant to infection. A study of the time-course of infection in strains of mice expressing q alleles throughout the H-2 on 4 different genetic backgrounds (NIH, SWR, DBA and B10) revealed that most strains were resistant to infection. However, the DBA/1 strain exhibited differential responsiveness, 4 out of 6 individuals harbouring mature adult parasites on day 35 post-infection.

Characterization of Trichinella spiralis antigens sharing an immunodominant, carbohydrate-associated determinant distinct from phosphorylcholine.

The biochemical and immunochemical characteristics of T. spiralis molecules (group II antigens) sharing an immunodominant epitope were examined. Six major proteins, ranging from 43-68 kDa, and from pI 5.0-6.3, express the determinant. Together, they account for at least 3% by weight of the total protein in L1 larval homogenate. The antigens are glycosylated. Following periodate oxidation, they reacted with biotin aminocaproyl hydrazide, and treatment with trifluoromethanesulfonic acid decreased their Mr. Deglycosylated group II antigens lost immunoreactivity with a monoclonal antibody specific for the determinant, and oligosaccharides released by treatment with mild base blocked binding of the monoclonal antibody to native antigens. The determinant on one of the group II antigens (43 kDa) was removed by N-glycanase. Neither phosphorylcholine nor antibody to phosphorylcholine interfered with monoclonal antibody binding to native group II antigens. Together, these results suggest that the immunodominant group II antigen epitope is associated with N- and O-linked oligosaccharides, and that it is not phosphorylcholine.