Regulation of toxocariasis in mice selectively reared for high and low immune responses to Nematospiroides dubius

Test mice have been selectively reared for high (H) or low (L) immune responses to Nematospiroides dubius. After secondary infection with N. dubius, the L mice voided ten times as many eggs in their faeces as the H mice, and at necropsy, 71% versus 20% of the inoculum of N. dubius were recovered as adult worms from the L and H mice respectively. Furthermore, N. dubius were more fecund in the L than in H mice. High or low immune responsiveness was not restricted to N. dubius infection in these mice but was also observed during Toxocara canis infection. The migration of T. canis larvae from gut via the liver to skeletal muscle and CNS was inhibited in H versus L mice. Many more larvae were recovered from the livers of H compared with L mice which was indicative of greater immunity in the H mice. The protective immune response in H compared with L mice to both N. dubius and T. canis included pronounced eosinophilia and elevated antiparasite antibody titres.     

Nematospiroides dubius: direct and correlated responses to selection for high and low immune responsiveness in mice

High and low immune responder lines of mice were bred selectively from an allogeneic stock over 10 generations, based on their fecal parasite egg count assayed 3 weeks after reinfection with 100 Nematospiroides dubius larvae. By generation 10, (F10), the low immune response mice voided about 10 times as many fecal N. dubius eggs as the high immune response mice. Realized heritability for the selected trait, fecal egg count after secondary infection (= protective immunity), was 0.35 at F7. F7 was considered the selection limit. Selection for change in fecal egg count did not significantly influence the conformational nor reproductive characteristics of these mice. Significant phenotypic and genetic correlations were evident between the selected character and innate immunity to N. dubius, humoral antibody response to N. dubius infection, and establishment, growth, and reproduction of N. dubius in the selected mice.     

Genetic control of immune responses to parasites: immunity to Trichuris muris in inbred and random-bred strains of mice.

A comparison has been made of the responses of random-bred CFLP and inbred NIH mice to infection with Trichuris muris. Random-bred mice showed greater variation in worm burdens and less uniformity in worm expulsion. Irradiation prior to infection reduced variation, but did not increase the mean level of infection above that shown by the most susceptible unirradiated mice. In NIH mice, however, irradiation raised the level of infection in all mice. The factors responsible for variation between CFLP mice and for the level of infection in NIH mice came into play after the fifth day of infection and were inactivated by cortisone acetate. It is suggested that these factors are immunologically mediated and under direct genetic control. Uniformity of infection and expulsion in NIH mice is therefore seen as a consequence of genetic uniformity; variability in CFLP mice as a consequence of genetic variation. The time of worm expulsion was found to differ markedly between inbred strains of mice. Hybrid progeny showed the expulsion time characteristic of the parental strain with the most rapid expulsion; greater resistance was therefore inherited as a dominant characteristic. The genetic control of immunity to T. muris is discussed in the context of the antibody- and cell-mediated components of the expulsion process.     

Immune expulsion of Trichuris muris from resistant mice: suppression by irradiation and restoration by transfer of lymphoid cells.

Lethal irradiation (850 rads) of mice made resistant to Trichuris muris markedly depressed their ability to expel a challenge infection. Expulsion was restored within 7-10 days when MLNC from uninfected mice were transferred on the day of infection, but no significant restoration was evident after transfer of immune serum. Transfer of BM alone had no restorative effect within 10 days and no synergism was seen when both BM and MLNC were transferred. MLNC from uninfected donors did not restore challenge expulsion when transfer was delayed until day 7 and the mice were killed 3 days later, although MLNC from resistant donors were effective within this time. When irradiated mice were given BM and the challenge infection allowed to continue for 15 days expulsion was restored, as it was when challenge was delayed for 7 days after BM transfer in thymectomized mice. The results confirm that expulsion of T. muris involves both antibody-mediated and lymphoid cell-mediated phases and offer no evidence for the involvement of other cell types.     

Suppression of the immune response to Trichuris muris in lactating mice.

Mice infected with Trichuris muris during lactation were unable to expel the infection at the normal time, but expulsion occurred when lactation was terminated. Suppression of expulsion was uniform in mice suckling more than five young but variable with smaller litters. Mice exposed to a primary infection while lactating were shown to have serum antibodies capable of passively transferring immunity to recipient mice and showed near normal immunity to a secondary infection given after lactation had ceased. Acquired immunity to T. muris was also suppressed by lactation, but the worms which became established in lactating resistant mice were fewer and smaller than those in non-lactating, non-resistant controls. It is suggested that the suppressive effect of lactation in this host-parasite relationship is exerted on the second, lymphoid cell-mediated phase of worm expulsion.     

The characterization of intraepithelial lymphocytes, lamina propria leukocytes, and isolated lymphoid follicles in the large intestine of mice infected with the intestinal nematode parasite Trichuris muris

Despite a growing understanding of the role of cytokines in immunity to the parasitic helminth Trichuris muris, the local effector mechanism culminating in the expulsion of worms from the large intestine is not known. We used flow cytometry and immunohistochemistry to characterize the phenotype of large intestinal intraepithelial lymphocytes (IEL) and lamina propria leukocytes (LPL) from resistant and susceptible strains of mouse infected with T. muris. Leukocytes accumulated in the epithelium and lamina propria after infection, revealing marked differences between the different strains of mouse. In resistant mice, which mount a Th2 response, the number of infiltrating CD4+, CD8+, B220+, and F4/80+ IEL and LPL was generally highest around the time of worm expulsion from the gut, at which point the inflammation was dominated by CD4+ IEL and F4/80+ LPL. In contrast, in susceptible mice, which mount a Th1 response, the number of IEL and LPL increased more gradually and was highest after a chronic infection had developed. At this point, CD8+ IEL and F4/80+ LPL were predominant. Therefore, this study reveals the local immune responses underlying the expulsion of worms or the persistence of a chronic infection in resistant and susceptible strains of mouse, respectively. In addition, for the first time, we illustrate isolated lymphoid follicles in the large intestine, consisting of B cells interspersed with CD4+ T cells and having a central zone of rapidly proliferating cells. Furthermore, we demonstrate the organogenesis of these structures in response to T. muris infection.     

Host specificity in mice selected for innate immunity to Nematospiroides dubius: infections with Nippostrongylus brasiliensis, Mesocestoides corti and Salmonella typhimurium

The selection of mice for innate immunity to Nematospiroides dubius was not specific. Mice bred as refractory (R) to N. dubius infection were more refractory to primary infections with Nippostrongylus brasiliensis than other mice bred as liable (L) to infection with N. dubius. R and L, as well as randomly-bred (Rd) and inbred C3H mice were all immune to challenge infection with N. brasiliensis. Previous infections with N. brasiliensis failed to influence the course of N. dubius infections or the status of the selected mice as R, Rd and L to infection with N. dubius. R and L mice were equally susceptible to Mesocestoides corti infection but more resistant than Rd mice. R and L mice died sooner after infection with Salmonella typhimurium than Rd, although R survived longer than L mice.     

Experimental infection of Nematospiroides dubius in Swiss albino mice. VII. Worm burdens and serum protein fractions

The paper deals with the alterations in serum protein profiles and its possible correlation with the worm burden in Swiss white mice infected with single and repeated doses of N. dubius infective larvae. Marked changes observed in serum protein fractions in all the groups were found to be further raised with increase in dose levels. However, maximum (optimal) changes of different fractions depend on the total number of larvae inoculated irrespective of the number of inoculations given. Percentage of worm retention was found to be inversely proportional to the number of larvae inoculated. Worm expulsion during repeated infections was attributed to the therefore, participation of increased amount of immunoglobulins and, humoral immunity (immediate type) has been suggested to play a major role in the immune mechanism of this model.     

Studies in vitro on the reaction of peritoneal exudate cells from mice immune to infection with Nematospiroides dubius with the infective third stage larvae of this parasite.

The results of the present study show that peritoneal exudate cells from mice immune to infection with Nematospiroides dubius are able in vitro to damage the third stage infective larvae as measured by a loss in infectivity. The ability of these cells to function in the absence of specific antibody seems to be related to the presence of trypsin labile factors on their surfaces. Lymphocytes from immune mice are also able to damage the larvae. The suggestion is made that 'activated' macrophages may play an important role in immunity to this infection.     

Effects of Toxoplasma gondii (Gleadle strain) on the host-parasite relationship in trichinosis.

The effects of concurrent infection with Toxoplasma gondii on the host-parasite relationship in trichinosis were studied. Infected mice showed a delay in expulsion of Trichinella spiralis adults from the gut. Persisting adult female worms were fecund but the numbers of larvae recovered from the muscles were not increased. Increased resistance to the systemic phase of trichinosis was shown by reduced numbers of muscle larvae after intravenous injection of newborn larvae in animals with toxoplasmosis as compared with control mice. There were no differences in small bowel pathology of trichinous mice with and without toxoplasmosis but inflammation around muscle cysts of T. spiralis was reduced in mice with toxoplasmosis. The eosinophilia which normally develops in mice with trichinosis was suppressed by concurrent toxoplasmosis. Trichinella infection did not alter the numbers of T. gondii cysts recovered from the brain 4 weeks after infection. It is suggested that the delay in expulsion of adult worms, decrease in muscle inflammation around T. spiralis cysts, and inhibition of eosinophilia result from immune suppression, while the reduction in numbers of muscle larvae after intravenous injection of newborn larvae reflects enhanced nonspecific resistance to infection in toxoplasmosis.     

Nematospiroides dubius: susceptibility to infection and the development of resistance in hypothymic (nude) BALB/c mice.

BALB/c-nu/nu mice and their intact nu/+ littermates are equally susceptible to infection with third-stage larvae of Nematospiroides dubius. Unlike their heterozygous littermates, however, the nu/nu mice are unable to form ganulomata in the intestinal wall and become only partially resistant to rechallenge. Following two or more infections, nu/nu mice maintain a high burden of adult intestinal worms, whereas worms are lost from immune nu/+ mice. Studies in T cell-injected nu/nu mice suggest that a full complement of T cells is needed to develop maximum resistance against the infective third-stage larvae and to expel adult worms. Measurement of serum immunoglobulin levels indicate that infected nu/+ mice have very high levels of IgG1 whereas the levels of IgG2a are reduced. In infected T cell-injected nu/nu mice, IgG1 levels increase with the number of T cells injected, whereas IgG2a levels are variable but always higher than in infected nu/+ mice.     

Intestinal parasites of conventionally maintained BALB/c mice and Mastomys coucha and the effects of a concomitant schistosome infection

A longitudinal study was carried out to identify the spectrum of intestinal parasites present in conventionally maintained BALB/c mice and Mastomys coucha and to determine the effects of concomitant schistosome infections on their parasite status. Six parasites were observed during the course of the study, namely the nematodes Aspiculuris tetraptera and Syphacia obvelata, Entamoeba muris and the flagellates Trichomonas muris, Spironucleus muris and Chilomastix spp. Although the 2 rodents shared common facilities, the overall prevalences of S. obvelata, T. muris and S. muris were significantly higher in M. coucha than BALB/c mice. BALB/c mice with concomitant schistosome infection had increased prevalences of E. muris, T. muris and S. muris. In M. coucha, in contrast, there were no significant increases in parasite prevalences. Infection intensities of T. muris and S. muris were significantly greater in M. coucha than BALB/c mice. Concomitant schistosome infection resulted in increased intensities of T. muris infection in BALB/c mice only. The influence of immune status in determining the susceptibilities of rodents to environmentally transmitted parasites is discussed.     

Genetic control of immune responses to parasites: selection for responsiveness and non-responsiveness to Trichuris muris in random-bred mice.

Populations of Schofield strain, random-bred mice were shown to have a bimodal variation in ability to bring about immune expulsion of the nematode Trichuris muris. This variation was genetically determined and independent of the size of infection experienced. The proportion of mice unable to achieve worm expulsion (non-responders) was relatively constant in various populations of the strain but was increased by selective breeding from mice of known status. Crosses made between non-responder and responder mice produced progeny that were almost all (92%) of responder phenotype, showing that the ability to achieve worm expulsion was inherited as a dominant characteristic. It is suggested that the genetic control involves a small number of genes; the possible immunological mechanisms by which control is mediated are briefly discussed.     

Nematospiroides dubius: response of the late fourth-stage larva to anthelmintics in vitro

Approximately 60% of fourth-stage larvae of Nematospiroides dubius recovered from mice 6 days after infection, developed to the young adult stage when cultured over a 7-day period in a complex medium in vitro. Larvae at the late fourth stage of development were found to be highly susceptible to most broad spectrum anthelmintics under in vitro conditions, the benzimidazole, imidazothiazole, pyrimidine, isothiocyanate and macrocyclic lactone compounds all being active at very low concentrations. Narrow spectrum anthelmintics active only against ascarids, pinworms, filariae, cestodes or trematodes had little or no effect on these larvae. Ineffective also were those chlorinated hydrocarbon, substituted phenol and salicylanilide compounds known to affect Haemonchus but not trichostrongylid worms in general. It is concluded that in vitro assays employing larvae of N. dubius are useful for the stringent screening of compounds for broad spectrum antitrichostrongyle activity. Used in conjunction with in vivo screens employing N. dubius in mice, they also afford means for detecting intrinsic activity against the parasite in a system free from any complicating host pharmacokinetics.     

Response of Peyer's patch lymphocyte subsets to Giardia muris infection in BALB/c mice. I. T-cell subsets

Initial cellular events in the intestinal immune response occur within Peyer's patches and are subject to complex regulation by T cells. The aim of this study was to analyze the response of murine Peyer's patch T, T-helper (Th), and T-suppressor (Ts) cells to Giardia muris infection. Immunocompetent BALB/c mice were infected with G. muris cysts and, at serial times during the infection, Peyer's patch leukocyte suspensions were incubated with fluorescent monoclonal antibodies that identified murine leukocytes, T, Th, or Ts lymphocytes. These suspensions were examined by flow cytometry to quantify each T-cell subset as a percentage of total leukocytes. Total Peyer's patch leukocytes more than doubled in number during the course of G. muris infection and returned to control levels as the infection was cleared. The percentages of Peyer's patch Th and Ts lymphocytes were 34.1 +/- 0.8% (mean +/- SEM) and 6.2 +/- 0.3%, respectively, in the absence of infection, and did not change significantly during Giardia infection. The Th/Ts ratio in Peyer's patches was 5.6 +/- 0.2 in uninfected BALB/c mice and also did not change significantly during clearance of G. muris. We conclude that Peyer's patch leukocytes double in number in response to G. muris infection in immunocompetent mice, G. muris infection does not lead to altered percentages of Peyer's patch T, Th, or Ts lymphocytes, and clearance of G. muris infection is associated with a Peyer's patch Th/Ts ratio of greater than 5.     

Trichinella spiralis: Delayed rejection in mice concurrently infected with Nematospiroides dubius

The immune response of mice to the nematode Trichinella spiral's was markedly altered when the infection was superimposed upon an existing infection with Nematospiroides dubius. The expulsion of a primary infection of T. spiralis was delayed in such mice, and the worms persisted for at least 4 weeks longer than they did in control mice. The degree to which expulsion was suppressed was related to the number of N. dubius present. It would appear that both adult and larval stages of N. dubius can exert a suppressive effect, since the expulsion of T. spiralis was affected within days of a super-imposed (i.e., larval) N. dubius infection. When adult N. dubius were removed from mice 4 days before infection with T. spiralis, the mice expelled the latter parasite within the normal time, indicating that recovery from the suppressive effects of concurrent infection occurred rapidly. Concurrent infection with N. dubius appeared to affect both the afferent and efferent arms of the immune response to T. spiralis, since sensitization by, and memory of, prior infection were impaired and the expression of acquired immunity was inferior to that of controls.     

The absence of resistance in congenitally athymic nude mice toward infection with the intestinal nematode, Trichuris muris: resistance restored by lymphoid cell transfer

Congenitally athymic (nude) mice maintained infections of Trichuris muris for at least 40 days post-infection, whereas phenotypically normal mice expelled the worms by 18 days post-infection. Complete worm expulsion occurred in nude mice which had been given spleen cells. On the other hand, a partial resistance to the infection was observed in nude mice which received thymus or mesenteric lymph node cells. No significant worm reduction was seen by injection of immune serum.     

Nematospiroides dubius and Nippostrongylus brasiliensis: delayed type hypersensitivity responses to ovalbumin in the infected mouse

Mice (C57BL) infected with the intestinal nematode Nematospiroides dubius showed depressed delayed type hypersensitivity responses to ovalbumin administered subcutaneously in Freund's complete adjuvant. IgG and IgM responses to this inoculum were unaffected. It is unlikely that the depression arose from impairment of the ear test response because responses to an extract of the adult parasite were measurable and ear testing with lipopolysaccharide yielded normal responses in infected mice. Furthermore, mice immunized on the day of infection responded normally, whilst long term infected mice ear challenged with antigen pulsed macrophages gave depressed responses. The in vitro proliferative responses of cells from the spleens and from the lymph nodes draining the site of immunization were enhanced marginally by N. dubius infection. Furthermore, these cells induced normal or elevated adoptive delayed-type hypersensitivity and IgG responses in irradiated recipients. These findings suggest that N. dubius does not compromise the development of ovalbumin specific T cells involved in a delayed type hypersensitivity response. Evidence for the induction of suppressor cells by N. dubius is discussed, and the findings are compared with results obtained with Nippostrongylus brasiliensis, a parasite which is rejected rapidly from the mouse.     

Effects of spleen cells and serum on transfer of immunity to Strongyloides venezuelensis infection in hypothymic (nude) mice

The course of Strongyloides venezuelensis infection in congenitally hypothymic (nu/nu) mice and their heterozygous thymus-bearing littermates (nu/+) was followed. Unlike the infected nu/+ mice, the nu/nu mice were unable to expel the worms until the end of the observation period (98 days post-infection). In addition, about three times as many eggs were counted at the peak level of infection in faeces of the infected nu/nu mice in comparison with the nu/+ mice. No acquired resistance to rechallenge was observed among the nu/nu mice. Auto-reinfection within the infected nu/nu mice could not be supposed in the present study. The worm expulsion mechanism was generated by nu/nu mice which had been given syngeneic spleen cells from intact +/+ mice. The expulsion of adult worms, as well as the protection against migrating larvae, occurred anamnestically when spleen cells from immune +/+ mice were transferred. The serum transfer, however, only caused a retardation of larval migration. The results support the hypothesis that direct worm immunity and worm expulsion are a T cell-dependent phenomenon.     

Intestinal IgA responses to Giardia muris in mice depleted of helper T lymphocytes and in immunocompetent mice

Immunocompetent mice infected with Giardia muris generate an intestinal antibody response to this parasite and clear G. muris infection. Previous work has shown that G. muris infection is prolonged in mice that have been depleted of helper (CD4+) T lymphocytes by treatment with a monoclonal antibody (mAb) directed against the murine CD4 antigen. The aim of the present study was to compare the intestinal anti-Giardia antibody response in immunocompetent mice and in mice depleted of helper T (Th) lymphocytes by treatment with anti-CD4 mAb. Immunocompetent mice generated an IgA response to G. muris, as judged by the presence of IgA on Giardia trophozoites harvested from the intestine of these animals more than 10 days after the start of the infection. The anti-Giardia IgA response was impaired in mice depleted of Th lymphocytes, as judged by virtual absence of immunofluorescent staining of trophozoites from these animals for surface-bound IgA. Clearance of G. muris infection was impaired by treatment of mice with anti-CD4 mAb. The results suggest that Th (CD4+) lymphocytes are important for the generation of a local IgA response against G. muris trophozoites in the mouse intestine and that IgA anti-trophozoite antibody may contribute to the clearance of G. muris from the intestine of immunocompetent mice.