Catherine A. Crandall

Trichinella spiralis: immunologic response to infection in mice.

Abstract Serum IgG 1 , IgG 2 , IgM, and IgA levels were measured by radial immunodiffusion in C57B1 mice after infection with 200 Trichinella larvae . IgG 1 concentrations increased 2- to 5-fold during the second week of infection; IgA and IgG 2 levels showed little consistent change; IgM concentrations increased from the second week through the fourth week of infection. Similar changes in serum immunoglobulin levels, with the exception of IgM, were found in ICR mice infected with Trichinella . An increase in IgG 1 -containing cells relative to cells containing immunoglobulins of other classes was observed in the immunocyte populations of the spleens, mesenteric lymph nodes, and Peyers patches of mice hyperinfected with Trichinella . The immunocyte population of the intestinal mucosa showed a relative increase in IgM- and IgG 1 -containing cells during the second week after primary infection. Antibody in the four immunoglobulin classes was measured by the indirect fluorescent antibody technique using frozen sections of muscle larvae as antigens. By day 15 of infection antibody was detected in all immunoglobulin classes. Antibodies in IgG 1 and IgG 2 exhibited specificity for the stichosome cells and cuticle; antibodies in IgA and IgM showed specificity principally for membranes of the larvae. Antibody in the content of the small intestine was detected during the second week of infection. These antibodies were predominantly in IgA. Antigen ( Trichinella extract)-sensitive cells in spleen, mesenteric lymph node and Peyers patches of Trichinella -infected mice were detected by incorporation of tritiated thymidine in cell cultures. The cells were detected in the spleen during the first week of infection and by the second week in the other lymphoid tissues. Transfer of spleen cells or lymph node cells from Trichinella -infected C57B1 mice to uninfected C57B1 mice induced an accelerated, humoral antibody response to Trichinella antigens in the recipient mice after inoculation of the mice with infective Trichinella larvae.

Heligmosomoides polygyrus (=Nematospiroides dubius): Suppression of antibody response to orally administered sheep erythrocytes in infected mice

Abstract Mice infected with 200 to 300 Heligmosomoides polygyrus had reduced serum hemagglutinin titers following a series of oral inoculations of sheep erythrocytes (SRBC) when compared to similarly inoculated uninfected mice. Study of antibody-producing cells by the indirect hemolytic plaque technique demonstrated a low splenic response to oral immunization in which IgA predominated. No alteration was evident in the proportions of antibody-containing cells in the different Ig classes with infection. Comparison of the immune response to oral and intraperitoneal routes of SRBC inoculation in infected and uninfected mice demonstrated a similar reduction in antibody titer with both routes of inoculation, although immunosuppression following intraperitoneal inoculations was not consistantly observed. The data are discussed in relation to the influence of the helminth infection on intestinal immune response and systemic immune response.

Ascaris suum: Immunoglobulin responses in mice

Abstract The immune response in mice to infection with Ascaris suum was characterized by determining (1) changes in serum immunoglobulin levels and (2) changes in the relative proportions of immunoglobulin-containing cells in major lymphoid tissues and sites of possible local immunoglobulin production. Significant increases in IgG 1 and IgA occurred during the third week of infection. IgM was the only immunoglobulin exhibiting a major change in relative serum concentrations; levels increased approximately 15-fold during the second week of infection. Absorption experiments with ascaris extract indicated that up to 50% of the serum IgM may be antibody. Inhibition studies showed that the IgM anti-body precipitating with ascaris extract has a phosphoryl choline specificity. A relative increase in IgM-containing cells was observed in spleens and mesenteric lymph nodes early after infection and preceded the rise in serum IgM concentration. The relative distribution of immunocytes by immunoglobulin class in the lung and liver was similar to that of the spleen.

Heligmosomoides polygyrus (= Nematospiroides dubius): Humoral and intestinal immunologic responses to infection in mice

Measurements of serum IgG1, IgG2a, IgM, and IgA levels and antibody titers in these immunoglobulin classes were made at intervals after initial infection and challenge infection of mice immunized by two or three previous infections. Identical measurements were made on the content of the small intestine in mice which had been exposed to the same infection schedule. Sections of small intestine taken after initial infection and challenge infection were examined by the fluorescent antibody technique for changes in populations of immunoglobulin-containing cells and by routine histologic procedures for histopathologic changes. In serum, only IgG1 was consistently increased after initial infection, and antibody in IgG1 was detected within the first 2 wk of infection. In immunized animals, only IgG1 and antibody of this class always responded to challenge infection, although antibody in other immunoglobulin classes was detected. IgA concentration of the intestinal content did not differ significantly after initial infection or challenge infection of immunized mice. Immunized mice had about twice the IgG1 concentration in intestinal content as singly infected animals. No intestinal antibody was detected after initial infection; only IgG1 antibody was detected in the intestinal content of immunized and challenged mice. Cell infiltrates in the intestinal mucosa and submucosa of immunized animals contained numerous IgG1-containing cells. Mast cells and globular leukocytes were observed in the intestine of immunized animals.

T-dependent suppression of the primary antibody response to sheep erythrocytes in mice infected with Trichinella spiralis

Abstract Mice infected for 20 days with the parasitic mematode Trichinella spiralis had significantly reduced numbers of splenic antibody-forming cells (AFC) and decreased serum hemagglutinin titers following intraperitoneal immunization with sheep erythrocytes (SE). Similarly, when immunized in vitro to SE, cultures of splenocytes from infected mice developed fewer AFC than cultures of normal cells. Splenocytes from infected mice actively suppressed the in vitro response of normal cells to SE, and this in vitro suppression was abolished by lysis with anti-thy 1 antiserum and enhanced by lysis with anti-immunoglobulin antiserum. The addition of supernatant fluids from cultures of splenocytes from infected mice to cultures of normal cells on Day 0 of culture reduced by 70% the number of AFC produced by these cultures. These results indicate the presence of T-suppressor cells and suggest that antigen-induced suppression (antigenic competition) is one mechanism of Trichinella -induced suppression.

Microfilaremia and antibody responses in CBA/H and CBA/N mice following injection of microfilariae of Brugia malayi.

leave the surface at an angle that was slightly larger than their arrival angle. The reverse was true for arrival angles larger than 14.7?. In either case, the variance in departure angles was quite large. No evidence was seen of attraction toward solid surfaces so that it seems likely that observations of affinity for margins (Chernin and Dunavan, 1962, loc. cit.) may have been due to surface configuration as these authors had suggested. As a result, miracidia in natural surroundings may not be expected to be concentrated around all solid surfaces.

Ascaris suum: Immunosuppression in mice during acute infection

Abstract Mice inoculated by stomach intubation with 10,000 embryonated Ascaris suum eggs, 4, 11, or 21 days before an intraperitoneal (ip) immunization with 2 × 108 sheep erythrocytes (SRBC) had reduced numbers of direct (IgM) splenic hemolytic plaques measured at 4 days after immunization and only a marginal reduction in indirect plaques (IgG) measured at 9 days after immunization. Lower dosages of Ascaris eggs or simultaneous inoculation of Ascaris eggs and SRBC did not suppress antibody responses to SRBC. No reduction in a secondary antibody response to SRBC injected 4 days after Ascaris inoculation was observed. IgM and IgG hemagglutinin titers, as distinguished by 2-mercaptoethanol sensitivity, were suppressed in mice injected ip with 108 SRBC 10 days following inoculation of 10,000 Ascaris eggs, but titers in both Ig classes were similar in infected and control mice injected with 2 × 109 SRBC. At Day 20, antibody titers following ip injection of 1.0 or 100 μg of ovalbumin in alum were reduced in mice infected with 10,000 Ascaris eggs 4 days before antigen injection. Contact hypersensitivity to oxazalone was not altered in mice sensitized at 5 or 14 days after inoculation of 10,000 Ascaris eggs. The delayed hypersensitivity response, measured by footpad swelling, to an optimum intravenous sensitizing dosage of SRBC was inhibited in mice sensitized 10 days after Ascaris infection, but not inhibited in mice sensitized at 21 or 32 days after infection. In contrast, the delayed hypersensitivity response to subcutaneous sensitization with SRBC 10 days after Ascaris infection was not altered.

Localization of antibody binding sites in the larvae of Ascaris lumbricoides var. suum by means of fluorescent technics

Abstract Antibody binding sites in Ascaris lumbricoides var. suum were determined by direct and indirect fluorescent antibody technics. Binding sites on embryonated eggs, freshly hatched second stage larvae, and larvae isolated from and within the lungs and livers of experimentally infected mice, as well as sections of adult worms were investigated. Specific fluorescent staining was observed in oral, anal, cuticular, and excretory pore precipitates of larvae isolated from tissues and in the cuticle of whole and sectioned larvae. No precipitates were observed on second stage larvae; however, the cuticle stained specifically in both whole and sectioned second stage larvae when incubated in immune fluorescent globulin. Similarly, specific staining of the larval cuticle was noted in sections of embryonated eggs. The “fertilization” membrane of the latter stained nonspecifically. No staining of the cuticle in sections of adult worms was observed; only the periphery of the muscle cells showed specific staining.

In vivo Studies of Ascaris suum Larvae planted in Diffusion Chambers in Immune and Nonimmune Mice.

Second-stage Ascaris suum larvae were placed in Millipore diffusion chambers which excluded host cells. The chambers were planted intraperitoneally in immune and nonimmune mice and removed for study at various time intervals. The larvae survived equally well in the chambers planted in immune and normal mice, but growth of the larvae occurred only in chambers placed in normal mice. Larvae recovered from immune mice resumed growth when transferred to nonimmune mice. Acquired immunity to nematode infection is usually associated with a complex antibody response and an increased inflammatory reaction to tissue invasion and migration by the helminth. The relative immunologic significance of this antibody response and the increased cellular reactivity to helminth infection are difficult to evaluate individually in vivo. The diffusion chamber developed by Algire et al. (1954) has been used in many experimental studies to isolate tissues and cells from the cellular reaction of the host. Huff (1956) proposed that the diffusion chamber offers considerable promise for the study of problems in parasitology, and in 1960 Huff et al. grew exoerythrocytic stages of malaria in diffusion chambers. Recently, Coleman and Fotorny (1962) used a Sellotape diffusion chamber in a study of antibody binding sites on Hymenolepis nana. This investigation was undertaken to study the survival of Ascaris suum larvae in diffusion chambers in both immune and nonimmune mice and to evaluate the influence of humoral factors in immunity to Ascaris infection when cells are not allowed to contact the larvae. MATERIALS AND METHODS Adult female Ascaris suum were obtained from a local abattoir. Eggs were removed and embryonated according to a previously described method (Arean and Crandall, 1962).

Electron microscope observations on the cuticle and submicroscopic binding of antibody in Ascaris suum larvae.

An electron microscope study of the cuticles of secondand third-stage Ascaris suum larvae did not reveal the presence of submicroscopic pores. Ferritin-conjugated immune rabbit globulin was bound to the surface of the cuticle of these larvae. Ferritin-labeled normal rabbit globulin was observed on the cuticles of some secondbut not third-stage larvae. Preliminary electron microscope studies of second-stage larvae isolated from the peritoneal cavity of immune mice after an intraperitoneal injection showed that the peritoneal cells adhered to the cuticles in 4 hr. The cells were mainly macrophages and appeared to be in direct contact with the cuticles of the larvae but no apparent morphologic alteration in the larvae was observed. Fluorescent antibody studies have shown that the cuticles of several nematode species are antigenic (Crandall et al., 1963; Taffs and Voller, 1963; Baratawidjaja et al., 1963). It has been suggested that at least some of the fluorescent antibody binding might be due to antigens within the helminth which pass through submicroscopic pores and adhere to the cuticle (Taffs and Voller, 1963; Jackson, 1963). This investigation was undertaken to determine by electron microscopy whether pores are present in the cuticle of Ascaris suum larvae and to localize the submicroscopic antibody-binding sites on the larvae with ferritinlabeled immune globulin. A preliminary electron microscope study showing the cellular reaction to larvae isolated from the peritoneal cavity of immune mice is