Effect of dietary magnesium on Ascaris suum infections of mice

Low amounts of dietary magnesium affected the inflammatory tissue response in nonimmunized mice differently than in immunized mice. Eosinophil numbers and LPL activity in lung tissue following infection with A. suum larvae were altered by the level of magnesium in the diets of mice. Average or higher dietary levels of magnesium resulted in decreased numbers of lung larvae indicating an overall protective effect. Increases in eosinophil numbers or LPL activity were not directly related to the numbers of larvae/lungs. Larvae/livers, eosinophil numbers, and LPL activity were affected by the types of magnesium diets that mice received. Nonimmunized mice had differences in larvae/liver (at 2 days and 7 days pi) and LPL activity (at 2 days pi). Immunized mice had varying findings at 2 days pi but a direct relationship between dietary magnesium and numbers of larvae, numbers of eosinophils, and liver LPL activity at 7 days pi.     

A phosphorylcholine idiotype related to TEPC 15 in mice infected with Ascaris suum.

A serum component, binding antigens having phosphorylcholine (PC) determinants were induced in several strains of mice by infection with Ascaris suum. This component was isolated and demonstrated to be an IgM (K) anti-PC antibody having idiotypic determinants in common with the IgA PC-binding myeloma protein TEPC 15. Rabbit anti-idiotypic antisera prepared with this component had idiotypic specificity for TEPC 15 and cross-idiotypic recognition of MOPC 167 and McPC 603, all IgA PC-binding myeloma proteins. The antisera also recognized determinants not present on TEPC 15. IgM and idiotype levels were quantitated by radial immunodiffusion and PC-specific antibody measured by hemagglutination (HA) with sheep erythrocytes coated with pneumococcal-C-polysaccharide. Mean IgM levels ranged from 2.5 to 8.7 mg/ml, idiotype from 0.54 to 5.3 mg/ml; and HA titers from 1:512 to 1:130,000 in different mouse strains. The high PC-specific antibody response was not duplicated by immunization with dead ascaris larvae or by infection with two other nematode species.     

Nonspecific immunomodulation influences resistance of mice to experimental infection with Mesocestoides corti and Ascaris suum

The influence of nonspecific immunomodulation on the course of experimental infection was examined in larval cestodosis (Mesocestoides corti) and ascaridosis (Ascaris suum) in mice. Immunosuppressive treatment (with azathioprine or hydrocortisone) resulted in a decrease of resistance in both models. The subsequent administration of T-activin to immunosuppressed mice led to the restoration of resistance to a level equal to that of untreated control mice. The administration of different immunomodulators partially protected mice against M. corti (T-activin, thymomodulin) or A. suum (T-activin, thymomodulin, thymosin fr.5, bursa-activin) infection. The protective effect of different treatments did not correlate with the level of specific antibody in the sera of infected mice. These results, which confirmed the decisive role of T-cell immunity in the resistance to the helminth infections, raise the possibility of the use of immunomodulators (thymic preparations) in the immunoprophylaxis of helminthoses.     

Effects of immunoactivity on Ascaris suum infection in mice]

The immune response to sheep red blood cell (sRBC) was monitored in the mice infected with Ascaris suum or Trichinella spiralis. The effects of the infection with T. spiralis or the injection with cyclophosphamide(CY) as an immunosuppression agent prior to challenge infection with the embryonated eggs of A. suum were monitored in mice by means of the level of infection with A. suum and cellular and humoral immune response to sRBC. Following the oral administration of 1,000 eggs of A. suum to mice, delayed-type hypersensitivity (DTH) and rosette-forming rate were gradually decreased and reached to the lowest levels at the 5th week and 6th week postinfection, respectively, and then returned to normal at the 10th week. The hemagglutinin(HA) and hemolysin(HE) titers were gradually elevated and reached to peak at the 3rd week postinfection, and then returned to normal level. The appearance ratios of the eosinophils and mast cells were in peak at the 4th week and the 2nd week postinfection, respectively. Meanwhile the harvest ratio of A. suum larvae from the liver and lungs was 21.97% at the 1st week postinfection. Following the oral administration of 300 T. spiralis infective larvae, DTH and rosette-forming rate were gradually decreased with the lapse of time and reached the lowest values in the 30th and 21st day of postinfection, and then slightly increased and transiently decreased in the 70th and 80th day of postinfection, respectively. HA and HE titers were the lowest in the 21st and 90th day, whereas the ratios of eosinophils and mast cells were the highest on the 40th and 14th day postinfection, respectively. Following the intraperitoneal injection of CY, the body weight, the spleen weight, DTH, rosette-forming ratio, HA and HE titers, the number of WBC and the ratio of the mast cell were predominantly decreased in the 5th day, and then returned to the same value of the 1st day postinjection. The ratio of eosinophils was gradually decreased following to advance of days. At the 1st, 5th and 10th days after intraperitoneal injection of CY of 400 mg/kg, a dose with 1,000 eggs of A. suum was administered orally to mice, and harvest rate of the larvae at the 7th day postadministration was 7.07% in the 1st day, 14.94% in the 5th day, 10.1% in the 10th day, 8.02% in control group. The effect of prior infection with infective larvae of T. spiralis upon immunological sequelae of a challenge infection of mice with embryonated eggs of A. suum in 30 or 70 days interval was checked.(ABSTRACT TRUNCATED AT 400 WORDS)     

Acquired resistance to migrating larvae of Ascaris suum in young pigs by repeated drug-abbreviated infections

Cross-bred 3- and 8-wk-old pigs were used to test whether drug-abbreviated infections with Ascaris suum can stimulate acquired resistance to challenge. During the immunization period, both age groups of animals were infected with increasing numbers of A. suum eggs (500, 1,000, 2,000, 5,000, 10,000, and 20,000) at 7-day intervals while the pigs were receiving pyrantel tartrate in the feed. Two days after the last infective dose, animals were placed on unmedicated feed for 8 days and then challenged with 10,000 eggs. All pigs were killed 7 days after challenge, and milk spots on the livers and larvae recovered from the lungs were counted. Larval recoveries from lungs of the immunized animals were significantly smaller than those from the unimmunized animals in both age groups, suggesting that the pigs were capable of acquiring strong resistance to parasitic infections. In immunized animals, challenge infection did not contribute significantly to milk spot formation. The number of milk spots was significantly greater in the older animals, indicating that milk spot formation may be age related.     

Cross-reactions with Ascaris suum antigens of sera from mice infected with A. suum, Toxocara canis, and Angiostrongylus cantonensis

Ascaris suum larval excretory-secretory (AsES) antigen and larval (AsLA) as well as adult somatic antigen (AsAA) which were thought to be possibly helpful in the diagnosis of visceral larva migrans (VLM) due to A. suum infection were investigated in the present study. Serum taken from mice orally inoculated with approximately 250 embryonated eggs of A. suum or Toxocara canis, or 40 third-stage larvae of Angiostrongylus cantonensis were assessed by enzyme-linked immunosorbent assay (ELISA) using the AsES antigen, AsLA or AsAA at 1, 2, 3, 4, 6 and 8 weeks post infection (WPI). The titer of serum IgG from mice infected with A. suum increased from 1 WPI and a peak at 4 WPI was observed when it reached approximately three times the level of uninfected control mice. Thereafter, it decreased gradually but remained high as found from 6 to 8 WPI. No cross-reactions of heterologous serum IgG against AsES antigen was observed, whereas heterologous serum IgM exhibited significant cross-reactions to AsES antigen. Cross-reactivities to AsLA and AsAA by heterologous serum IgG as well as IgM antibodies were also observed in the trial. Altogether, the AsES antigen apparently seemed to be superior to the other two somatic antigens when used in the diagnosis of A. suum-induced VLM with serum IgG as tested by ELISA. Moreover, it was the first report to test the possibly antigenic cross-reactivity between A. suum and A. cantonensis.     

Cross-reactions of sera from Toxocara canis-infected mice with Toxascaris leonina and Ascaris suum antigens.

The ELISA method using larval excretory-secretory (E/S) products and homogenized Toxocara canis, Toxascaris leonina and Ascaris suum adult worm extract were used to determine possible cross-reactions in BALB/c and C57BL/10 mice, inoculated with embryonated eggs or adult worm extract of T. canis in single and multiple doses. When we used sera of mice infected with embryonated eggs of T. canis against different heterologous antigens, we observed no cross-reactions in BALB/c mice against A. suum E/S and adult worm extract antigens with a single dose. In multiple doses this was absent too against T. leonina adult worm extract in BALB/c mice, and in both strains against A. suum E/S and adult worm extract. In BALB/c mice inoculated with adult worm extract of T. canis we did not observe cross-reactions with A. suum E/S antigen with both inoculation doses. In the remainder of the experiments, we observed cross-reactions of different intensities.     

Vaccination with recombinant Ascaris suum 24-kilodalton antigen induces a Th1/Th2-mixed type immune response and confers high levels of protection against challenged Ascaris suum lung-stage infection in BALB/c mice

Previous studies have shown that antigens from various life-cycle stages of Ascaris suum can induce host-protective immunity against challenge infections with infective eggs of A. suum. This study evaluated whether Escherichia coli-expressed recombinant 24-kDa antigen from A. suum (rAs24) was a suitable vaccine candidate for the control of Ascaris infections by examining its performance in a mouse model. Immunization of BALB/c mice in three consecutive doses with rAs24 in Freund's Complete Adjuvant (FCA) results in protection against challenge infections as manifested by a 58% reduction (P<0.001) in recovery and stunted development of A. suum lung-stage larvae at day 7 post-challenge. Sera obtained from immune protected mice had a significantly increased level of immunoglobulin G (IgG) (P<0.0001) but had no IgE response. Analysis of IgG-subclass profiles revealed that IgG1 (P<0.0001) showed the greatest increase followed by IgG2b (P<0.005), IgG2a (P<0.006) and IgG3 (P<0.04). Splenic T cells from rAs24-FCA immunized mice secreted significantly high levels of both Th1 cytokine gamma-interferon (P<0.005) and Th2 cytokine interleukin-10 (P<0.001) after stimulation with rAs24 in vitro. Interestingly, affinity purified anti-rAs24 IgG was shown to inhibit moulting of A. suum lung-stage L3 to L4 in vitro by 26%, indicating an in vivo function of the endogenous As24 in the moulting processes. An intense expression of endogenous As24 in the hypodermis and gut epithelium of A. suum lung-stage L3 by immunofluorescence supports a function for endogenous As24. These findings may contribute to the understanding of rAs24-induced Th1/Th2-mediated effector mechanisms required for the protection of A. suum lung-stage larval infection.     

Cryptosporidium infections in inbred strains of mice.

Cryptosporidium, a protozoan parasite of man and animals, is an important etiological agent of diarrhea throughout the world, particularly in children and immunocompromised individuals such as AIDS patients. Unfortunately, because of the lack of both in vivo laboratory models and reliable in vitro parasite culture systems, virtually nothing is known about the immunological events occurring during disease. In order to identify reliable animal models for infection, we studied C. parvum infections in 19 different strains of mice representing 12 H-2 haplotypes: A/J, AKR/J, B10.D2/J, B10.M/J, C3H/HeJ, C57BL/65, C57BL/6J-bgJ, CBA/NJ, DBA/1J, DBA/2J, HRS/J, HTG/J, NZB/B1NJ, NZW/J, P/J, RIII/J, SJL/J, SWR/J, and WB/ReJ, and in one gerbil: Meriones unguiculatus. Fecal samples and histological sections of the intestine taken on day 7 post-Cryptosporidium inoculation indicated that only the beige mouse (C57BL/6J-bgJ) harbored significant numbers of parasites compared to the other strains. The numbers of parasites harbored in these NK cell-deficient beige mice were, however, considerably lower than those seen in neonatal mice. Adult inbred mouse strains susceptible to Cryptosporidium infections are discussed.     

Ascaris suum: biosynthesis and isoinhibitor profile of chymotrypsin/elastase isoinhibitors

Chymotrypsin/elastase isoinhibitors were radiolabeled when live, adult Ascaris suum were incubated in tissue culture medium (NCTC-135) supplemented with L-[35S]cysteine. This is the first demonstration that the synthesis of these proteins occurs in A. suum; the isoinhibitors are not host products utilized by the parasite against its host. Of five chymotrypsin/elastase isoinhibitors demonstrable in A. suum, only isoinhibitors 1, 4, and 5 were found in each worm. The amino acid sequences of these three isoinhibitors indicate that they are gene products and are not obtained by modification after translation. The two inhibitors that are not observed could arise by limited proteolysis. The same chymotrypsin/elastase isoinhibitor profile present in each nematode eliminates any speculation that the multiple forms arise from an adaptation between A. suum and its host. A new chymotrypsin/elastase isoinhibitor nomenclature is proposed, so that isoinhibitor 1 is now Isoinhibitor A, and isoinhibitors 4 and 5 are now Isoinhibitors B and C, respectively.     

Reaction of Ascaris suum phosphofructokinase with diethylpyrocarbonate. Inactivation and desensitization to allosteric modulation

Reaction of the phosphofructokinase from Ascaris suum with the reagent, diethylpyrocarbonate (DEPC), results in the loss of enzymatic activity. Treatment of the inactivated enzyme with hydroxylamine brings about the recovery of almost 80% of the original activity suggesting that the modified residues are histidines. Further evidence for the modification of histidines is that concomitant with the loss of activity, there is a change in A242 nm that corresponds to the derivatization of 5-6 histidines per subunit. There is no change in A278 nm during the derivatization process, thereby ruling out the modification of tyrosines by DEPC. Analyses of the first order inactivation rate constant for DEPC derivatization at different pH values resulted in the determination of a pKa of 6.4 +/- 0.1 for the group on the enzyme that reacts with DEPC. Derivatization of the enzyme with DEPC in the presence of fructose 6-phosphate (Fru-6-P) protected the enzyme against inactivation by 80%. ATP or MgATP gave no protection against DEPC inactivation. When the Fru-6-P-protected enzyme was further reacted with DEPC in the absence of Fru-6-P, a total of 2 histidines were modified per subunit, and the derivatization of one of these could be correlated with activity loss. When the phosphofructokinase that had been derivatized by DEPC in the presence of Fru-6-P was assayed, it was found that it no longer exhibited allosteric properties and appeared to be desensitized to ATP inhibition. This loss of ATP inhibition could be correlated with the modification of 2 histidines per subunit by DEPC. The first order rate constant for desensitization was determined at different pH values and a pKa value of 7.0 +/- 0.2 was obtained for the group(s) responsible for the desensitization. Regulatory studies with the desensitized enzyme revealed that the enzyme was not stimulated by AMP, NH4+, K+, phosphate, sulfate, or hexose bisphosphates. It is concluded that histidine may be involved both in the active site and the ATP inhibitory site of the ascarid phosphofructokinase.     

Eosinophils mediate SIgA production triggered by TLR2 and TLR4 to control Ascaris suum infection in mice

Human ascariasis is the most prevalent but neglected tropical disease in the world, affecting approximately 450 million people. The initial phase of Ascaris infection is marked by larval migration from the host’s organs, causing mechanical injuries followed by an intense local inflammatory response, which is characterized mainly by neutrophil and eosinophil infiltration, especially in the lungs. During the pulmonary phase, the lesions induced by larval migration and excessive immune responses contribute to tissue remodeling marked by fibrosis and lung dysfunction. In this study, we investigated the relationship between SIgA levels and eosinophils. We found that TLR2 and TLR4 signaling induces eosinophils and promotes SIgA production during Ascaris suum infection. Therefore, control of parasite burden during the pulmonary phase of ascariasis involves eosinophil influx and subsequent promotion of SIgA levels. In addition, we also demonstrate that eosinophils also participate in the process of tissue remodeling after lung injury caused by larval migration, contributing to pulmonary fibrosis and dysfunction in re-infected mice. In conclusion, we postulate that eosinophils play a central role in mediating host innate and humoral immune responses by controlling parasite burden, tissue inflammation, and remodeling during Ascaris suum infection. Furthermore, we suggest that the use of probiotics can induce eosinophilia and SIgA production and contribute to controlling parasite burden and morbidity of helminthic diseases with pulmonary cycles.