Nippostrongylus brasiliensis: phospholipase in nonsensitized and sensitized rats after challenge.

Rats given an initial infection with Nippostrongylus brasiliensis showed greatly elevated phospholipase B levels in the small intestines and lungs from 8 through 22 days after challenge. The rise in enzyme concentration occurred earlier (Days 8–11) in the proximal half of the intestine, but at Days 22, 29, and 36 the levels were much higher in the distal segments. This shift in activity correlates with the known elimination of worms and a diminishing inflammatory response in the proximal areas. The increase in enzyme activity in the intestine and lungs was associated with an increased production of eosinophils in the bone marrow 11–22 days after challenge. Rats sensitized with one stimulating infection before challenge showed an anamnestic type of response, as measured by enzyme levels in the small intestines and lungs and by the numbers of eosinophils in the bone marrow. The results are discussed in light of our similar data reported earlier from animals infected with Trichinella spiralis.     

Eosinophi-mediated destruction of Schistosoma mansoni eggs in vitro. II. The role of cytophilic antibody

Peritoneal exudative eosinophils obtained from Schistosoma mansoni-infected CBA/J mice cause morphological damage to isolated S. mansoni eggs in a 24 hr co-cultivation system in vitro. This egg-destructive activity was complement-independent and was abolished by trypsinization of the cells prior to co-cultivation. Trypsinized cells could be passively sensitized to renewed egg-destructive capacity by preincubation or co-gcultivation with immune sera, containing antibodies against a soluble egg antigenic preparation (SEA). Solid phase absorption of immune sera with SEA coupled to Sepharose 4B lowered the anti-egg antibody titers of these sera and eliminated their ability to sensitize trypsinized eosinophils. Sera from uninfected mice or from mice infected with Trichinella spiralis did not sensitize trypsinized cells. Addition of immune sera to eosinophil-rich cell populations obtained from uninfected mice also enhanced the egg-destructive capacity of these otherwise non-reactive cells. Therefore, eosinophil-mediated destruction of S. mansoni eggs may be directed by cytophilic antigen-specific factors in sera from S. mansoni infected hosts.     

Eosinophil-mediated destruction of Schistosoma mansoni eggs III. lymphokine involvement in the induction of eosinophil functional abilities

Granulomas isolated from the livers of CBA/J mice infected for 8 weeks with Schistosoma mansoni produced a chemotactic activity for eosinophils, in a manner which correlated with the production of the lymphokine eosinophil stimulation promoter (ESP). ESP and chemotactic activities were also produced when eosinophilrich peritoneal exudative cells from S. mansoni-infected mice were cultured with S. mansoni eggs. These S. mansoni-related eosinophils destroyed approximately 20% of the eggs whereas eosinophils from normal (uninfected) mice did not have this ability. However, normal cells exposed to ESP-containing fluids in the co-cultivation system actively participated in egg destruction. Eosinophil-rich peritoneal exudative cells obtained from Trichinella spiralis-infected mice were incapable of destroying S. mansoni eggs during the normal 24 hr co-cultivation period, but did achieve destruction if the incubation period was extended to 48 hr. Marginal levels of chemotactic activity for eosinophils were detected in the co-cultivation fluids from T. spiralis-related cells and S. mansoni eggs, although these fluids did not contain demonstrable levels of ESP. Together, these data indicate that ESP/chemotactic factor-containing culture fluids can induce in normal, unreactive eosinophils the functional ability to destroy S. mansoni eggs in vitro. This may account for the ability of T. spiralis-related eosinophils to do so upon extended incubation.     

Immune responses to Trichinella pseudospiralis and Trichinella spiralis in mice

Primary infections with Trichinella pseudospiralis and Trichinella spiralis were followed in rapid- (NIH) and slow- (B10G) responder strains of mice. Expulsion of T. pseudospiralis was slower in both strains, but markedly so only in slower responder B10G mice. Blast cell activity in the mesenteric lymph nodes of the mice correlated with the expulsion patterns. In NIH mice, both parasites stimulated a strong response by day 8 of infection and activity had returned to control levels by day 11. In B10G mice, T. spiralis elicited an earlier peak response (day 12) than T. pseudospiralis (day 18), but in both, activity returned to control levels by day 21. Immunity to T. pseudospiralis and T. spiralis could be stimulated in NIH mice by prior infection with either parasite, by injection of T. spiralis larval antigen and by adoptive transfer of immune mesenteric lymph node cells taken from mice infected with either parasite. This extensive cross reactivity, and the differences seen during primary infections, are discussed in relation to the biology and specific identity of the two worms.     

The phospholipase B content of the intestines of sensitized rats challenged with varied larvae doses of Nippostrongylus brasiliensis.

The effects of challenging previously infected (sensitized) groups of rats with 25, 100 and 400 Nippostrongylus brasiliensis larvae on the relation between phospholipase B activity, bone marrow eosinophilia and worm burden were studied. The results indicated that the immune expulsion of worms from sensitized rats was temporally related to an increased phospholipase B activity and bone marrow eosinophilia. All three parameters demonstrated an anamnestic-type of response in the previously infected rats when compared to the infected control animals. The results also demonstrated that the size of the worm burden influenced the induction and concentration of phospholipase B and the numbers of bone marrow eosinophilis.     

Immune Correlates of Resistance to Trichinella spiralis Reinfection in Mice

The immune correlate of host resistance induced by reinfection of Trichinella spiralis remains unclear. In this study, we investigated immune correlates between the resistance and serum IgG antibody level, CD23⁺ IgM⁺ B cells, and eosinophil responses induced by T. spiralis reinfection. Mice were primarily infected with 10 or 100 T. spiralis larvae (10 TS, 100 TS), respectively, and after 4 weeks, they were challenge infected with 100 T. spiralis larvae (10–100 TS, 100-100 TS). Upon challenge infections, 10–100 TS mice induced significantly higher levels of T. spiralis-specific total IgG antibody responses in sera and antibody secreting cell responses in spleens compared to 100-100 TS mice, resulting in significantly reduced worm burdens in 10–100 TS mice (60% and 70% reductions for adult and larvae, respectively). Higher levels of eosinophils were found in mice primarily infected with 10 TS compared to those of 100 TS at week 8 upon challenge. CD23⁺ IgM⁺ B cells were found to be increased significantly in mice primarily infected with 10 TS. These results indicate that primary infection of 10 larvae of T. spiralis, rather than 100 larvae, induces significant resistance against reinfection which closely correlated with T. spiralis-specific IgG, eosinophil, and CD23⁺ IgM⁺ B cell responses.     

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

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.     

Interactions between Trichinella spiralis and Hymenolepis nana in the intestine of the mouse

Ferretti G., Gabriele F., Palmas C. and Wakelin D. 1984. Interactions between Trichinella spiralis and Hymenolepis nana in the intestine of the mouse. International Journal for Parasitology14: 29–33. The rejection of the intestinal phase of Trichinella spiralis in outbred CD₁ mice is associated with an inflammatory response. The effects of this inflammation were studied in mice concurrently infected with an unrelated parasite, Hymenolepis nana, which stimulates a slight inflammation but is also immunologically rejected by this strain of mice. Moreover though Cd(In)1 mice seem to be ‘slow responders’, in concurrent infections with T. spiralis and H. nana there was a marked effect upon the cesode survival. At the highest level of T. spiralis infection, H. nana failed to establish at all. On the other hand, if 300 T. spiralis were administered 10 weeks earlier, H. nana become established in all mice and the distribution of parasites suggests that immunodepression is involved. An accelerated loss of the nematode, when H. nana was present, was also demonstrated. There was no evidence of any reciprocal or specific cross-immunity between the two parasites.     

Anti-enteroviral activity of new MDL-860 analogues: Synthesis,in vitro/in vivo studies and QSAR analysis

A series of 60 nitrobenzonitrile analogues of the anti-viral agent MDL-860 were synthesized (50 of which are new) and evaluated for their activity against three types of enteroviruses (coxsackievirus B1, coxsackievirus B3 and poliovirus 1). Among them, six diaryl ethers (20e, 27e, 28e, 29e, 33e and 35e) demonstrated high in vitro activity (SI > 50) towards at least one of the tested viruses and very low cytotoxicity against human cells. Compound 27e possesses the broadest spectrum of activity towards all tested viruses in the same way as MDL-860 does. The most active derivatives (27e, 29e and 35e) against coxsackievirus B1 were tested in vivo in newborn mice experimentally infected with 20 MLD₅₀ of coxsackievirus B1. Compound 29e showed promising in vivo activity (protection index 26% and 4 days lengthening of mean survival time). QSAR analysis of the substituent effects on the in vitro cytotoxicity (CC₅₀) and anti-viral activity of the nitrobenzonitrile derivatives was carried out and adequate QSAR models for the anti-viral activity of the compounds against poliovirus 1 and coxsackievirus B1 were constructed.     

Role of the carbohydrate moiety of phospholipase B from Penicillium notatum in enzyme activity

Two types of phospholipase B from Penicillium notatum—the native enzyme and enzyme modified by endogenous protease (T. Okumura, S. Kimura, and K. Saito (1980) Biochim. Biophys. Acta, 617, 264–273)—were treated with endoglycosidase H (endo-β-N-acetylglucosaminidase H, Streptomyces griseus) to investigate the orientational change of the sugar chains associated with the lower activity of the modified enzyme. On measurement of release of sugar chains, by periodic acid-Schiff staining of endoglycosidase H-treated phospholipase B on sodium dodecyl sulfate-polyacrylamide gel electrophoresis and by direct sugar analysis of the isolated endoglycosidase H-treated phospholipase B, distinct curves were obtained for release of sugar chains from the native and modified enzymes with ultimately loss of about 30 and 55%, respectively, of the carbohydrate. Removal of sugar chains from the two enzymes resulted in similar increases in phospholipase B activity (phosphatidylcholine hydrolysis) and their phospholipase A₁ and A₂ activities in the presence of Triton X-100, but no change of lysophospholipase activity (lysophosphatidylcholine hydrolysis). The three former activities of the native and modified enzymes increased to almost 170 and 350%, respectively, of their initial values. However, little increase in phospholipase B activity was observed when the activity was assayed in the absence of Triton X-100, and none when it was assayed in the presence of sodium taurocholate. These findings suggest that the carbohydrate moiety of phospholipase B greatly influence the phospholipase B activity, especially in the presence of Triton X-100, and that the low phospholipase B activity of the modified enzyme is due to excess exposure of sugar chains on the surface of the molecule as a result of protease attack.     

Synthesis and anti-enterovirus activity of new analogues of MDL-860

A series of twelve novel compounds, analogues of antiviral agent MDL-860 were synthesized and their antiviral activity was evaluated in vitro against enteroviruses poliovirus 1 (PV1), Coxsackieviruses B1 (CVB1) and Coxsackieviruses B3 (CVB3). Compounds 14, 24 and 25 manifested strong antiviral effects against CVB1 and PV1 (SI values of 405 and 118 for CVB1 and PV1 respectively). In contrast to the wide anti-enteroviral activity of MDL-860, these three compounds were inactive against CVB3. Compounds 14, 24 and 25 along with MDL-860 were tested in vivo in mice infected with CVB1. Marked protective effects of compounds 14 and 24 were established, PI values of 50% and 33.3%, respectively. In addition, almost all of the tested compounds manifested very low toxicity.     

Counterregulation of Th2 immunity by interleukin 12 reduces host defenses against Strongyloides venezuelensis infection

The aim of this study was to investigate the role of interleukin 12 (IL-12) during Strongyloides venezuelensis infection. IL-12^−/− and wild-type C57BL/6 mice were subcutaneously infected with 1500 larvae of S. venezuelensis. On days 7, 14, and 21 post-infection, we determined eosinophil and mononuclear cell numbers in the blood and broncoalveolar lavage fluid (BALF), Th2 cytokine secretion in the lung parenchyma, and serum antibody levels. The numbers of eggs in the feces and worm parasites in the duodena were also quantified. The eosinophil and mononuclear cell counts and the concentrations of IL-3, IL-5, IL-10, IL-13, and IgG1 and IgE antibodies increased significantly in infected IL-12^−/− and wild-type mice as compared with uninfected controls. However, the number of eosinophils and mononuclear cells in the blood and BALF and the Th2 cytokine levels in the lungs of infected IL-12^−/− mice were greater than in infected wild-type C57BL/6 mice. In addition, serum IgE and IgG1 levels were also significantly enhanced in the infected mice lacking IL-12. Meanwhile, parasite burden and fecal egg counts were significantly decreased in infected IL-12^−/− mice. Together, our results showed that the absence of IL-12 upregulates the Th2 immune response, which is important for control of S. venezuelensis infection.     

Evidence for two distinct mechanisms of resistance in the rat to reinfection with Fasciola hepatica

Doy T. G. and Hughes D. L. 1982. Evidence for two distinct mechanisms of resistance in the rat to reinfection with Fasciola hepatica. International Journal for Parasitology12: 357–361. Congenitally athymic nude (rnu/rnu) and heterozygous hairy (rnu/ + ) rats were found to be equally highly resistant to oral reinfection with Fasciola hepatica. Accompanying the development of this resistance was a marked increase in intestinal eosinophil numbers. The sensitised rnu/ + rats showed a similar marked resistance to intraperitoneal challenge with newly excysted juvenile (NEJ) flukes. This was much less effective in the rnu/rnu rats, although there was some evidence of reduced numbers and stunting of parasites. Serum from infected rnu/rnu rats, unlike that from the infected rnu/+ rats, failed to induce the adherence of eosinophils to NEJ flukes in vitro. Flukes recovered from rnu/rnu rats were in general considerably larger than comparable flukes from their rnu/ + counterparts.There thus appears to be two distinct mechanisms of resistance to reinfection with F. hepatica operating in the rat. The first a T-independent system effective at the gut wall and the second, effective after penetration of the gut and requiring a T-dependent response for full expression. If eosinophils are involved in protection they can apparently function in the gut wall in the absence of an adherence promoting antibody in the serum.     

Allergic inflammation as a hypothesis for the expulsion of worms from tissues: A review

Introduction; comparison of studies of mice infected with Trichinella spiralis; discussion; summary; references.     

Basal metabolic rate and lipid and liver glycogen in mice infected by the nematode Nematospiroides dubius

The basal metabolic rate (BMR) of mice losing weight about two weeks after infection by Nematospiroides dubius was lower than that of uninfected mice gaining weight when fed ad libitum or losing weight on quantitatively reduced rations. There was no difference in BMR between the latter two groups.Following the injection of ¹⁴C-glucose, the high specific activity of expired CO₂ from infected and reduced ration mice was considered to be due to the utilization of energy reserves. The levels of lipid and liver glycogen were low in these two groups of mice and their specific activities, particularly in the severely affected animals, were high.It was concluded that the depressed BMR of infected mice is unrelated to anorexia, which did, however, explain the low levels of lipid and liver glycogen.     

Eosinophils and IL-4 Support Nematode Growth Coincident with an Innate Response to Tissue Injury

It has become increasingly clear that the functions of eosinophils extend beyond host defense and allergy to metabolism and tissue regeneration. These influences have strong potential to be relevant in worm infections in which eosinophils are prominent and parasites rely on the host for nutrients to support growth or reproduction. The aim of this study was to investigate the mechanism underlying the observation that eosinophils promote growth of Trichinella spiralis larvae in skeletal muscle. Our results indicate that IL-4 and eosinophils are necessary for normal larval growth and that eosinophils from IL-4 competent mice are sufficient to support growth. The eosinophil-mediated effect operates in the absence of adaptive immunity. Following invasion by newborn larvae, host gene expression in skeletal muscle was compatible with a regenerative response and a shift in the source of energy in infected tissue. The presence of eosinophils suppressed local inflammation while also influencing nutrient homeostasis in muscle. Redistribution of glucose transporter 4 (GLUT4) and phosphorylation of Akt were observed in nurse cells, consistent with enhancement of glucose uptake and glycogen storage by larvae that is known to occur. The data are consistent with a mechanism in which eosinophils promote larval growth by an IL-4 dependent mechanism that limits local interferon-driven responses that otherwise alter nutrient metabolism in infected muscle. Our findings document a novel interaction between parasite and host in which worms have evolved a strategy to co-opt an innate host cell response in a way that facilitates their growth.     

Depletion of eosinophils by anti-IL-5 monoclonal antibody treatment of mice infected with Trichinella spiralis does not alter parasite burden or immunologic resistance to reinfection.

Mechanisms of parasite killing by eosinophils are widely studied and are often implicated in mediating resistance to parasitic infection, especially in conjunction with specific antibodies. Evidence for the eosinophil as an anti-parasite killer cell in vivo is limited and may not justify the belief that eosinophils engage and/or kill infective helminths. We reexamined this question in a mouse model of trichinosis in which antisera to eosinophils were previously used to show the requirement for eosinophils in resistance to this nematode. The current studies used mAb to IL-5 to suppress eosinophil levels in CF1 mice infected with Trichinella spiralis. In mice given a primary infection and injected with an isotype control mAb or left untreated, the medullary and peripheral blood eosinophil numbers peaked at 3 wk postinfection (PI) and returned to baseline levels by 4 wk PI. Peripheral blood eosinophil numbers in infected mice injected with anti-IL-5 were maintained at levels below those of uninfected normal mice through 4 wk of infection. Histologically, there was a prominent eosinophil accumulation in infected, untreated, or control-mAb-treated mice associated with nurse cell complexes containing infective juveniles in skeletal muscle at 3 and 4 wk PI. This was largely eliminated in mice treated with anti-IL-5 mAb. However, the number of muscle stage juvenile worms recovered 3 and 4 wk PI after acid pepsin digestion was unaffected by eosinophil depletion. Challenge infections, in which mice were infected at day 0 with 125 muscle stage worms and challenged at day 28 PI with 350 muscle stage worms, developed peak eosinophil numbers in bone marrow and peripheral blood 3 wk after primary infection and 2 wk after challenge infection in mice receiving either no treatment or control mAb. In challenged mice receiving anti-IL-5 mAb, medullary and peripheral blood eosinophil numbers remained at or below those of uninfected animals. Although all groups exhibited significant resistance measured as muscle stage worm burdens 56 days PI, eosinophil depletion did not affect resistance of muscle worm recovery. These results suggest that eosinophils are not essential in the control of T. spiralis in either primary or challenge infections of CF1 mice. This in vivo study illustrates the questionable value of in vitro killing assays to assign effector function to any single inflammatory cell type.     

Immunosuppressive activity in serum from mice infected with Nematospiroides dubius following passive serum transfer

Dobson C. and Cayzer C. J. R. 1982. Immunosuppressive activity in serum from mice infected with Nematospiroides dubius following passive serum transfer. International Journal for Parasitology12: 561–566. Anti-N. dubius antibody titres increased with time after primary and secondary infections with 100 larvae in mice and 4 days after anthelmintic termination of a 28 day infection. Mice injected with serum from infected mice harboured fewer, smaller worms with reduced fecundities compared with control mice; the difference was greater with serum from donors given two compared with those given one infection.Mice injected with serum from donors infected for 14 days had fewer N. dubius than recipients of serum from mice infected for 28 days. Serum taken from mice 4 days after termination of a primary infection of 28 days duration was more protective when passively transferred to mice than serum from mice infected for 28 days. Serum from mice infected with 50 N. dubius larvae was more protective than serum, with a higher anti-N. dubius antibody titre, from, mice infected with 400 larvae. These observations are discussed in relation to immunosuppressive activities in the donated serum and associated with N. dubius.     

TLR7 modulating B-cell immune responses in the spleen of C57BL/6 mice infected with Schistosoma japonicum

B cells played an important role in Schistosoma infection-induced diseases. TLR7 is an intracellular member of the innate immune receptor. The role of TLR7 on B cells mediated immune response is still unclear. Here, C57BL/6 mice were percutaneously infected by S. japonicum for 5–6 weeks. The percentages and numbers of B cells increased in the infected mice (p < 0.05), and many activation and function associated molecules were also changed on B cells. More splenic cells of the infected mice expressed TLR7, and B cells were served as the main cell population. Moreover, a lower level of soluble egg antigen (SEA) specific antibody and less activation associated molecules were found on the surface of splenic B cells from S. japonicum infected TLR7 gene knockout (TLR7 KO) mice compared to infected wild type (WT) mice (p < 0.05). Additionally, SEA showed a little higher ability in inducing the activation of B cells from naive WT mice than TLR7 KO mice (p < 0.05). Finally, the effects of TLR7 on B cells are dependent on the activation of NF-κB p65. Altogether, TLR7 was found modulating the splenic B cell responses in S. japonicum infected C57BL/6 mice.     

Susceptibility of Laboratory Rodents to Trichinella papuae

Members of the genus Trichinella are small nematodes that can infect a wide range of animal hosts. However, their infectivity varies depending on the parasite and host species combination. In this study, we examined the susceptibility of 4 species of laboratory rodents, i.e., mice, rats, hamsters, and gerbils to Trichinella papuae, an emerging non-encapsulated Trichinella species. Trichinella spiralis and Trichinella pseudospiralis were also included in this study for comparison. Fifteen animals of each rodent species were infected orally with 100 muscle larvae of each Trichinella species. Intestinal worm burden was determined at day 6 and 10 post-inoculation (PI). The numbers of muscle larvae were examined at day 45 PI. The reproductive capacity index (RCI) of the 3 Trichinella species in different rodent hosts was determined. By day 6 PI, 33.2-69.6% of the inoculated larvae of the 3 Trichinella species became adult worms in the small intestines of the host animals. However, in rats, more than 96% of adult worms of all 3 Trichinella species were expelled from the gut by day 10 PI. In gerbils, only 4.8-18.1% of adult worms were expelled by day 10 PI. In accordance with the intestinal worm burden and the persistence of adults, the RCI was the highest in gerbils with values of 241.5±41.0 for T. papuae, 432.6±48 for T. pseudospiralis, and 528.6±20.6 for T. spiralis. Hamsters ranked second and mice ranked third in susceptibility in terms of the RCI, Rats yielded the lowest parasite RCI for all 3 Trichinella species. Gerbils may be an alternative laboratory animal for isolation and maintenance of Trichinella spp.