Richard B. 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.

Studies on schistosomiasis. XX. Further studies on the immunogenic significance of Schistosoma mansoni eggs in albino mice when subjected to homologous challenge.

A group of experimental mice was given ten subcutaneous injections of living S. mansoni eggs at 4to 7-day intervals. Each mouse received an approximate total of 5,500 eggs. One group of control mice received a like number of injections of the supernate from the egg suspensions; another control group remained untreated. All groups were challenged by percutaneous exposure to 100 S. mansoni cercariae at intervals ranging from 2 to 42 days after the last egg injection. Antibody response due to the egg injections was demonstrated by the miracidial immobilization test, but no increased resistance to S. mansoni challenge infection attributable to the egg injections could be demonstrated. The immunogenic significance of schistosome eggs deposited in the tissues of the mammalian host remains uncertain. Antibody reactive with eggs has been demonstrated by Oliver-Gonzalez (1954) and Oliver-Gonzalez, Bauman, and Benenson (1955) who showed that the anti-egg precipitins were species specific. Circumstantial evidence that eggs in tissues are necessary for the development of acquired resistance to Schistosomatium douthitti and Schistosoma mansoni in mice has been presented (Kagan and Lee, 1953; Crandall and Hunter, 1961; Hunter et al., 1962a). Hunter and Crandall (1962b) in preliminary experiments produced some resistance to homologous challenge infection in mice by the injection of living schistosome eggs, but the results were not entirely consistent. Recently Hsui and Hsii (1961) and Sadun, Yamaki, and Burke (1961) using the zoophilic Formosan strain of S. japonicum as an immunizing infection and the human Japanese strain as a challenge infection have suggested that eggs in the tissues are not necessary to demonstrate some degree of resistance in experimental animals. Hsui, Hsii, and Osborne (1962) using X-irradiated cercariae of S. japonicum for immunization of monkeys produced evidence of partial resistance to homologous challenge as manifested by reduced egg production in immunized animals. Villella, Gomberg, and Gould (1961, 1962) working with S. mansoni in mice have shown that irradiation of cercariae with cobalt 60 results in sterile worms, and that mice infected with irradiated cercariae exhibit some degree of resistance to challenge infections. Smithers (1962) using X-irradiated S. mansoni cercariae and monkeys as the test animal generally confirmed the work of Villella et al. and Hsui et al. in that a partial resistance to reinfection was produced by exposure to irradiated cercariae. Smithers states that even large numbers of irradiated cercariae produce only an incomplete protection against S. mansoni in rhesus monkeys. In some of the studies on irradiation complete sterilization of the worms was not always attained. Ritchie, Garson, and Erickson (1962) failed to demonstrate resistance to S. mansoni in mice after the injection of homogenates of lyophilized cercariae, adult worms, and eggs. These recent findings do not exclude the possibility that eggs, when present, contribute significantly to immunity in schistosome infections. Because previous experiments on inoculation Received for publication 27 August 19Q2. * With the aid of Grants E-1893 and E-2500 from the Division of Allergy and Infectious Diseases of the National Institutes of Health, U. S. Public Health Service. t Department of Microbiology, The University of Texas Southwestern Medical School, Dallas. + Department of Microbiology, College of Medicine, University of Florida, Gainesville. ? Resident Coordinator, L.S.U.-International Center for Medical Research and Training, San Jose, Costa Rica on leave, Department of Microbiology, College of Medicine, University of Florida, Gainesville.

Macroglobulin antibody response to Ascaris suum infection. Detection of a second macroglobulin component in the serum of infected mice.

Two closely associated components were detected by immunoelectrophoresis in the serum macroglobulin response of mice to Ascaris suum. One of these components was demonstrated to be IgM. The second component was shown to be antigenically distinct from IgM by rabbit antisera made to the macroglobulin fraction of infected mouse sera or to antigen-antibody precipitates of ascaris extract and mouse serum. The second component was not detected in normal mouse serum or in sera from mice with a variety of parasitic infections. This component bound to ascaris extract and its presence coincided with precipitin formation in agar gel between ascaris extract and sera from infected mice. Preliminary studies failed to establish that this component was an immunoglobulin or an acute phase reactant. The possible nature of the component is discussed. Helminth infections commonly induce raised serum immunoglobulin levels and a complex antibody response (Smithers, 1967). Larval ascarid infections in man have been associated with hypergammaglobulinemia with an increased level of IgM (Huntley et al., 1965). In a previous study, a precipitating antibody of the IgM immunoglobulin class was demonstrated in the sera of mice following experimental infection with Ascaris suum (Crandall and Crandall, 1967). Gel elution profiles, as well as immunoelectrophoretic studies of serum from mice, suggested that a marked increase in the concentration of macroglobulins occurred after an infection with A. suum. The present investigation was initiated to characterize further this macroglobulin response to larval A. suum infection in mice. MATERIALS AND METHODS

Brugia malayi: Intravenous injection of microfilariae in ferrets as an experimental method for occult filariasis

Microfilaremia, immune responses, and pathology were compared in ferrets infected with 100 third-stage larvae of Brugia malayi (subperiodic strain) or injected intravenously with 10(6) microfilariae. Ferrets (Mustela putorius furo) inoculated with third-stage larvae typically became patent during the third month after infection, with a mean patency of 123 +/- 25 (SE) days. Ferrets injected intravenously with microfilariae exhibited a relatively constant microfilaremia for 3-4 weeks and usually cleared microfilariae before the fourth month. Ferrets that cleared microfilariae after intravenous injection of microfilariae or after infection with third-stage larvae failed to become patent or became amicrofilaremic within 3 weeks after a challenge intravenous injection of 10(6) microfilariae. Clearance of circulating microfilariae was associated with eosinophilia and serum antibody specific for the microfilarial sheath in ferrets injected with microfilariae and in most ferrets infected with third-stage larvae. Ferrets infected with third-stage larvae and necropsied after clearance of microfilariae had tissue inflammatory reactions to microfilariae characteristic of occult filariasis (tropical eosinophilia) in man; these ferrets exhibited immediate cutaneous hypersensitivity and circulating reaginic antibody to antigens of microfilariae. In ferrets necropsied following two intravenous injections of microfilariae, the majority of ferrets examined within 10 days after clearance of microfilariae had visible liver lesions to microfilariae identical to those of the ferrets infected with third-stage larvae; immediate cutaneous hypersensitivity and reaginic antibody were not consistently detected in ferrets injected with microfilariae. Sera from ferrets that had cleared circulating microfilariae were transferred passively into ferrets made microfilaremic by intravenous injection of microfilariae. Sera with microfilarial sheath-reactive IgG antibody titers (greater than or equal to 1:200) and microfilarial agglutination titers (greater than or equal to 1:40) rapidly cleared injected microfilariae (less than 24 hr); this serum also cleared or greatly reduced circulating microfilariae established by an infection with third-stage larvae; only the IgG-containing fraction of the sera was active in immune clearance. Sera that cleared microfilariae of B. malayi did not clear circulating microfilariae of Dirofilaria immitis or prevent recurrence of circulating microfilariae of B. malayi in ferrets infected with adult filariae.(ABSTRACT TRUNCATED AT 400 WORDS)