Postnatal challenge infections of congenitally Schistosoma japonicum-infected piglets

The aim of the present study was to assess the effect of a congenital Schistosoma japonicum infection on the establishment, fecundity, and pathogenicity of a postnatal challenge infection. Five prenatally S. japonicum-infected piglets received a challenge infection (prenatal + challenge group), 5 prenatally infected piglets were followed without challenge (prenatal group), and 10 piglets, born by unexposed sows, served as challenge controls (challenge control group). Challenge infections were given 8 wk after the piglets were born (14 wk after the primary infection of the sows), and the study lasted another 11 wk. Variables included worm burden, tissue egg count, and liver pathology. Worm establishment and tissue egg count were comparable in the prenatal + challenge group and in the challenge control group, both exceeding at a statistically significant level those in the prenatal group. No difference in worm fecundity (eggs/female worms/g tissue) was seen between the 3 groups. Liver pathology (i.e., portal and septal fibrosis) was more severe in the challenge control group compared to the other groups. A congenital S. japonicum infection in piglets thus affected neither establishment nor fecundity of a postnatal challenge infection. In spite of this, the challenge infection gave rise to much less liver pathology than the similarly sized challenge control infection.     

Induction of resistance in Oncomelania hupensis nosophora against Schistosoma japonicum, but not against Paragonimus ohirai, using irradiated miracidia

Oncomelania hupensis nosophora snails sensitized with X-irradiated Schistosoma japonicum miracidia demonstrated resistance against a following challenge infection with non-irradiated homologous miracidia. The resistance in O. h. nosophora against S. japonicum was acquired within 1 day of sensitization, and it was strongest in a group challenged at an interval of 3 days. The resistance persisted for at least 4 weeks. Histological examinations revealed amoebocyte accumulation around the challenged S. japonicum sporocysts. On the other hand, when O. h. nosophora sensitized by exposure to X-irradiated P. ohirai or S. japonicum miracidia were subsequently challenged with normal P. ohirai miracidia, no resistance was observed, although they expressed the resistance against heterologous S. japonicum infection.     

Negative regulation of Schistosoma japonicum egg-induced liver fibrosis by natural killer cells

The role of natural killer (NK) cells in infection-induced liver fibrosis remains obscure. In this study, we elucidated the effect of NK cells on Schistosoma japonicum (S. japonicum) egg-induced liver fibrosis. Liver fibrosis was induced by infecting C57BL/6 mice with 18-20 cercariae of S. japonicum. Anti-ASGM1 antibody was used to deplete NK cells. Toll-like receptor 3 ligand, polyinosinic-polycytidylic acid (poly I:C) was used to enhance the activation of NK cells. Results showed that NK cells were accumulated and activated after S. japonicum infection, as evidenced by the elevation of CD69 expression and IFN-γ production. Depletion of NK cells markedly enhanced S. japonicum egg-induced liver fibrosis. Administration of poly I:C further activated NK cells to produce IFN-γ and attenuated S. japonicum egg-induced liver fibrosis. The observed protective effect of poly I:C on liver fibrosis was diminished through depletion of NK cells. Disruption of IFN-γ gene enhanced liver fibrosis and partially abolished the suppression of liver fibrosis by poly I:C. Moreover, expression of retinoic acid early inducible 1 (RAE 1), the NKG2D ligand, was detectable at high levels on activated hepatic stellate cells derived from S. japonicum-infected mice, which made them more susceptible to hepatic NK cell killing. In conclusion, our findings suggest that the activated NK cells in the liver after S. japonicum infection negatively regulate egg-induced liver fibrosis via producing IFN-γ, and killing activated stellate cells.     

Imaging diagnosis of schistosomiasis japonica--the use in Japan and application for field study in the present endemic area

For detecting lesions-related schistosomiasis japonica, X-rays, scintillation scanning, ultrasonography (US), computed tomography (CT), magnetic resonance (MR) and endoscopic examinations with biopsies have been used in Japan. Liver fibrosis and calcified changes are detected by US and CT. Most of the lesions that are detected by endoscopic examinations are due to deposited ova of Schistosoma japonicum. Portal hypertension is detected by US, CT and gastroscopic examination. Because schistosome infection decreased rapidly in Japan, most of the studies on imaging diagnosis were performed on chronic lesions or sequelae of schistosomiasis. Most of the techniques were used on admitted patients in well-equipped hospitals. US was introduced in the 1970s as a safe, rapid, non-invasive and inexpensive technique and has been used for diagnosis in hospitals and screening in the fields. As a typical US image of the liver, septal formation by high echogenic bands like mosaic was described, and this network pattern was reported in the other endemic countries; China and Philippines. As an appropriate technique, US has been broadly used in developing countries. Not only for diagnosis in a hospital, but also for monitoring changes of morbidity, US is used in the community level. Network pattern related to the severity of S. japonicum infection, has not been described in S. mansoni or S. haematobium infection. Appearance of network pattern depends on pathological changes such as periportal fibrosis, postnecrotic fibrosis and calcified ova. For advanced studies on morbidity of schistosomiasis japonica, further research on pathological basis of network pattern and standardization of US diagnosis are necessary.     

Prenatal Schistosoma japonicum infection in piglets: effect of repeated exposure of the dams on treatment efficacy and susceptibility to challenge infections

This study elucidated the fate of prenatal infections in piglets born by dams repeatedly infected before and during pregnancy with Schistosoma japonicum. Independent variables included repeated infections of the dams and treatment or challenge infection (or both) of the prenatally exposed piglets. Dependant variables were worm counts, fecal and tissue egg counts, weight gain, and gross pathological observations. Fifteen female piglets (the dams) were included, of which 6 received repeated infections with S. japonicum during 6 mo. All dams were inseminated and 10 wk pregnant; 12 of the dams were infected with S. japonicum, of which 6 had been repeatedly infected. Three dams remained uninfected. Eight weeks after delivery, the prenatally exposed piglets (the offspring) were grouped, and 6 of the 12 groups were treated with praziquantel. Four weeks after treatment, 5 groups of piglets were infected with S. japonicum. Groups of piglets were killed either 12 or 22 wk after delivery. Repeated infections of the dam did not prevent establishment of a congenital infection in the pig fetuses. Piglets born with a congenital infection were not resistant to a S. japonicum challenge infection given 12 wk after birth. Neither did praziquantel effectively cure the piglets nor did treatment of the prenatally infected piglets prevent establishment of a challenge infection given 4 wk after treatment. Results of the present study indicate that prenatal exposure, independently of the dam's infection status, may change the host response to challenge infections and treatment after birth.     

B cell response is required for granuloma formation in the early infection of Schistosoma japonicum

Schistosoma egg-induced liver granuloma is a dynamic inflammatory reaction that results from complex immune responses to the infection. However, the role of B cells in inflammatory granuloma development is not yet fully understood. We report here that B cell function is required for S. japonicum egg-induced granuloma pathology in early infection. Both OBF-1 knockout mice and microMT mice develop severely reduced hepatic granulomas at five weeks post-infection compared to their wild-type counterparts. In contrast, they display no significant difference in granuloma pathology at eight weeks post-infection. Moreover, we find that B cells and antibodies accumulate in the granulomas of wild-type mice early in the infection, indicating a contribution of the B cell response to the granulomatous inflammation. Furthermore, defects in B cell function markedly reduce liver egg burden. These results suggest an important role for B cells in early granuloma pathology. Surprisingly, we found that the S. japonicum infection destroys the structure of the lymphoid follicles. This disruptive effect is correlated with a severely impaired T cell-dependent antibody response upon challenge with ovalbumin. Thus, these findings reveal a novel aspect of the interaction between Schistosoma and the host immune system.     

Schistosoma mekongi: a prominent neutrophil chemotactic activity of egg antigen with reference to that of Schistosoma japonicum

Schistosoma mekongi causes granulomatous lesions around eggs deposited in the liver with neutrophil-rich inflammatory reactions in the early stage of the egg laying. To define the aspects of the typical pathogenesis of S. mekongi infection, we determined the difference between soluble egg antigen (SEA) from S. mekongi and S. japonicum with a focus on chemotactic factors for neutrophils or eosinophils. Mean volume and protein amount of S. mekongi eggs was 71 and 58% of those of Schistosoma japonicum eggs, respectively. Neutrophil chemotactic activity of S. mekongi SEA was about two times higher than that of S. japonicum. In contrast, eosinophil chemotactic activity of S. mekongi SEA was about half of that of S. japonicum SEA. Molecular analysis revealed that S. mekongi SEA contains higher molecular-weight components with a lower level of glycosylation, and this is likely to be related to the intense neutrophil chemotactic activity in comparison with S. japonicum SEA. The prominent chemotactic reactivity for neutrophils is likely to be involved in the typical pathogenesis of mekongi schistosomiasis.     

Schistosoma mansoni,S. haematobium,and S. japonicum: early events associated with penetration and migration of schistosomula through human skin

Migratory pattern of schistosomula of Schistosoma mansoni, S. haematobium, and S. japonicum through human skin were analyzed in skin organ cultures. These studies showed that the schistosomula of S. mansoni and S. haematobium has similar migratory patterns through human skin. During the first 24h after infection nearly 90% of S. mansoni and S. haematobium schistosomula were present only in the epidermis. Majority of the schistosomula were found in the dermis only after 48h and they appear to reach the dermal vessels around 72h after infection. Migratory pattern of S. japonicum on the other hand was significantly different from the other two species in that over 90% of the parasites had already reached the dermis within the first 24h and schistosomula were present in the dermal vessels within 2h after infection. Analysis of the cytokine pattern at 8h after infection by a macro gene array and RT-PCR analysis showed that out of 24 different cytokines analyzed only IL-1ra, IL-10, and TNF-alpha were increased in the human skin following infections with S. mansoni and S. haematobium, whereas, after infection with S. japonicum there was significant increases in IL-1beta, IL-1ra, IL-2, IL-6, IL-8, IL-10, IL-15, IL-18, and TNF-alpha. Immunohistochemical analysis of epidermal sheets showed focal accumulation of HLA-DR(+) cells in areas where schistosomula of S. mansoni had entered the human skin.     

Study on differences in the pathology, T cell subsets and gene expression in susceptible and non-susceptible hosts infected with Schistosoma japonicum

More than 40 kinds of mammals in China are known to be naturally infected with Schistosoma japonicum (S. japonicum); Microtus fortis (M. fortis), a species of vole, is the only mammal in which the schistosomes cannot mature or cause significant pathogenic changes. In the current study, we compared the differences in pathology by Hematoxylin-eosin staining and in changes in the T cell subsets with flow cytometry as well as gene expression using genome oligonucleotide microarrays in the lung and liver, before challenge and 10 days post-infection with schistosomes in a S. japonicum-susceptible mouse model of infection, a non-susceptible rat model and the non-permissive host, M. fortis. The results demonstrated that S. japonicum promoted a more intensive immune response and more pathological lesions in M. fortis and rats than in mice. Hematoxylin-eosin staining revealed that the immune effector cells involved were mainly eosinophilic granulocytes supplemented with heterophilic granulocytes and macrophages. The analysis of splenic T cell subsets showed that CD4(+) T cell subsets and the CD4(+)/CD8(+) ratio were increased, while the CD8(+) T cell subsets decreased remarkably in rats; whereas the CD8(+) T cell subsets were increased, but the CD4(+)/CD8(+) ratio was decreased significantly in mice. The analysis of the pattern of gene expression suggested that some immune-associated genes and apoptosis-inducing genes up-regulated, while some development-associated genes were down-regulated in the infected M. fortis compared to the uninfected controls; the three different hosts have different response mechanisms to schistosome infection. The results of this study will be helpful for identifying the key molecules in the immune response to S. japonicum in M. fortis and for understanding more about the underlying mechanism of the response, as well as for elucidating the interaction between S. japonicum and its hosts.     

Immunoglobulins inside Schistosoma japonicum eggs from the livers of mice

Using fluorescent antibody techniques, immunoglobulins (Ig's), mainly IgG class, were detected inside Schistosoma japonicum eggs lodged in mouse liver. Ig's were observed as a focal pattern between the miracidium and the eggshell during early infection (5-7 wk), particularly in lightly infected mice (20 cercariae). With advancement of time of infection (8-18 wk), a diffuse pattern of staining over the miracidial body developed and became predominant. The diffuse pattern of staining could be observed in the eggs taken from heavily infected mice (50 cercariae), during early stage. Eggs showing the focal pattern in a restricted area appeared to be morphologically intact, whereas eggs showing the diffuse pattern exhibited some types of eggshell destruction. We conclude that the focal pattern reflects disintegration of eggs in the initial stage and the diffuse pattern in the advanced stage. This spatial relationship between Ig's and eggs is discussed in relation to destruction of eggs.     

Prevention of Schistosoma japonicum infection in mice with long-acting praziquantel implants

This work reports the prevention outcomes of a praziquantel (PZQ) implant against the infection of Schistosoma japonicum in mice. The PZQ implant produced stable plasma PZQ concentrations in a range of 100-1300 ng/mL for a period of 70 days, by releasing PZQ in subcutaneous tissues in a sustained manner. To assess the prevention effects, the mice were infected at varying times after implantation. All the mice were sacrificed at 6 weeks after infection for worm and egg recovery and counting, worm morphological examination, determination of egg-hatching rates, and analysis of hepatic histology. The infection was successfully prevented for mice with early infection times (within 2-3 weeks), as nearly no worms, paired worms, eggs, or miracidia were recovered. However, in mice with late infection times (after 3 weeks), the prevention effects were diminished due to the decreased plasma PZQ concentrations at late times. Interestingly, the implants showed robust prevention effects on repeated infection at 1 and 3 weeks. In the infection-prevented mouse livers, no granuloma formation or granulomatous inflammation was observed. The results demonstrated that by blocking the development of infecting miracidia and by deactivating the eggs, the PZQ implants encouragingly prevented the S. japonicum infection and avoided liver damage.     

Paeoniflorin: a monomer from traditional Chinese medical herb ameliorates Schistosoma japonicum egg-induced hepatic fibrosis in mice

Treatment of liver fibrosis associated with Schistosoma japonicum ova-induced granulomas remains a challenging proposition. There is a close relationship between high levels of interleukin-13 (IL-13) and the development of severe schistosome fibrosis. In contrast, IL-13 receptor (R) α2 has an effective role in attenuation of profibrosis. Several Chinese herbs have significant beneficial effects in liver disease. Accordingly, the purpose of the present study was to investigate the therapeutic effect of Paeoniflorin (PAE) on liver fibrosis. A mouse model for liver fibrosis was established, using infection with S. japonicum cercariae via the skin. Liver tissue was used to examine the effect of PAE on hydroxyproline, collagen I and III, and IL-13 and IL-13Rα2. The results showed that PAE has significant suppressive effect on the increase of both hepatic hydroxyproline and collagen I and III, which are the main components of extracellular matrix (ECM). Meanwhile, PAE not only inhibits IL-13 production, it also elevates IL-13Rα2 in PAE-pretreated groups compared with controls. These results suggested that PAE can improve liver fibrosis due to S. japonicum infection. The effect of PAE appears to depend on a decrease of IL-13 and an increase of IL-13Rα2.     

Pathogenesis of congenital infection with Schistosoma japonicum in pigs

To elucidate aspects of pathogenesis of congenital infections with Schistosoma japonicum, 5 Danish crossbred sows were infected during late pregnancy with a Chinese isolate of S. japonicum, and 17 of their offspring (fetuses and piglets) were examined 7, 20. 34, 54, and 69 days postinfection (PI). Organ samples were collected for histopathological examination with emphasis on liver and lung. Samples of the corresponding placenta were also collected from fetuses at postmortem examinations. Perfusions were performed on some of the fetuses to recover schistosomes, and in addition, amniotic fluid was examined for schistosomes. A schistosomulum was found in a 99-day-old fetus 3 wk PI. Eggs were found in meconium from 109-day-old fetuses 34 days after infection of the dam, showing that the prepatent time was the same as in postnatal infections. Piglets examined 54 and 69 days PI had inflammatory reactions in their livers, and progression toward healing and repair of the inflammatory reaction occurred from 54 to 69 days PI. This pilot study is one of the bases for the model of congenital schistosomiasis used currently at the Danish Centre for Experimental Parasitology.     

Congenital infection with Schistosoma japonicum but not with Schistosoma bovis in sheep

The present study investigated whether Schistosoma japonicum or Schistosoma bois could establish prenatally in lambs. Three ewes were exposed to S. japonicum by intramuscular injection of cercariae, and 3 ewes were exposed to S. bovis cercariae using the leg-emerging technique approximately 2 mo before delivery, and 1 age-matched pregnant ewe served as an uninfected control. The study lasted 18-20 wk after infection, which was 8-9 wk after delivery. All 6 exposed ewes became infected with either S. bovis or S. japonicum. Eight lambs were borne by the 7 ewes, of which 1 (S. bovis exposed) was dead and 1 (S. japonicum exposed) died at delivery. Of the 3 S. japonicum-exposed lambs, 2 were found infected. Four lambs born of S. bovis-exposed ewes were negative. Despite having no worms, these 4 S. bovis-exposed lambs as well as the 1 negative S. japonicum-exposed lamb had, in contrast to the nonexposed control lamb, few, but distinct, liver granulomas dominated by eosinophils and giant cells with large central necrotic areas but with no remnants of eggs or worms. Hence, congenital infection was demonstrated in S. japonicum-infected lambs, but not in S. bovis-infected ones.     

Reduced levels of mutagen processing potential in the Schistosoma japonicum-infected mouse liver

Epidemiological studies have shown the presence of a positive correlation between the infection of Schistosoma japonicum and colorectal and/or liver cancers in the humans. To explore the mechanism underlying this correlation, we have investigated the mutagen-activating potentials of the liver homogenate fraction (S9) from Schistosoma japonicum infected mice and those from control mice, by use of the Ames test with 2-acetylaminofluorene, aflatoxin B1 and 3-amino-1-methyl-5H-pyrido[4,3-b]indole (Trp-P-2) as test mutagens. Liver S9 prepared from the infected group at the 15th week after the infection showed a potential significantly lower than that from the control group. The hepatic cytochrome P-450 concentration in the infected mice was persistently low, about a half of that in the uninfected mice, during the period of 6-18 weeks after the infection. Thus, in mice bearing chronic schistosomiasis, mutagen-processing potentials are decreased.     

Tuberculosis of the lymphatic nodes and coexisting invasion with Toxoplasma gondii]

Six cases of the peripheral lymphatic nodes tuberculosis with positive serologic reactions to Toxoplasma gondii antigen are presented. It was shown, that independently of a complex of clinical examinations histologic examination is decisive for the diagnosis of lymphatic nodes tuberculosis with coexisting toxoplasmosis. A positive serologic reaction with T. gondii antigen in patients with lymphatic nodes tuberculosis may reflect inactive infection with T. gondii. Use of anti-toxoplasmosis drugs may be not necessary in such cases.     

Detection of inducible nitric oxide synthase in Schistosoma japonicum and S. mansoni

Schistosoma japonicum and S. mansoni were tested for reactivity with an anti-inducible nitric oxide (iNOS) antibody and the distribution of iNOS was studied by immunofluorescent tests in different stages of the parasites. Reactivity was associated with the tegument in both larval schistosomes (sporocysts and cercariae) and eggs. With adult worms, the majority of the immunofluorescence was predominantly subtegumental in S. japonicum and parenchymal in S. mansoni. Fluorescence was also observed in host tissues (snails and mouse liver). In Western blots, the enzyme of S. japonicum had an apparent molecular weight of about 210 kDa. The possible role of worm and host iNOS in the parasite-host interrelation remains to be clarified.     

Evaluation of the anthelmintic effects of artesunate against experimental Schistosoma mansoni infection in mice using different treatment protocols

The therapeutic effects of artesunate against experimental Schistosoma mansoni infection in mice were analyzed. Previous studies showed that artesunate is highly effective against S. japonicum infection, but the action of this drug against S. mansoni remained uncovered. The present study examines the optical conditions for artesunate against S. mansoni and evaluates the effects of inhibiting the sexual maturation of adult worms. Mice infected with S. mansoni were orally administered with artesunate according to different schedules. Four consecutive administrations of 300 mg/kg of artesunate at 2-week intervals conferred almost total protection without the development of pathological lesions in the liver. The significant reduction in the number of eggs produced by surviving worms and the status of egg maturation suggested that artesunate inhibits sexual maturation. Electron microscopy revealed that artesunate caused morphological damage, especially on the worm tegument. Artesunate was also very effective in iron-deficient mice. Furthermore, the efficacy of artesunate was equal to or better than that of artemether against S. japonicum infection. Considering that artemether is more toxic, artesunate is currently one of the most efficient drugs against immature S. mansoni.     

Efficacy of administration of praziquantel on 2 days 2 weeks apart against Schistosoma japonicum eggs in mice

We compared a group of mice given multiple oral doses of praziquantel (PZQ) on 1 day 7 weeks after infection with Schistosoma japonicum and another group given multiple doses of PZQ over 2 days (7 and 9 weeks after infection) by examining the number of schistosome eggs and oograms in their tissues and feces. In the 1-day-protocol group (4 x 100 mg/kg x 1 day), calcified dead eggs or shells accounted for 77.4 and 70.1% of the total EPG (the total number of immature eggs, mature eggs, and calcified eggs and shells per 1 g tissue) in the liver and small intestine, respectively, 11 weeks after infection. Shells accounted for nearly half of the total EPG (54.3% liver and 46.6% small intestine). Mature eggs accounted for 8.4% (liver) and 5.1% (small intestine) of the total EPG. Only shells were found in the feces. In the 2-day-protocol group (4 x 100 mg/kg x 2 days, 2 weeks apart), dead eggs accounted for 87.2 and 89.5% of the total EPG in the liver and small intestine, respectively, 11 weeks after infection. Shells accounted for 62.5% (liver) and 75.8% (small intestine) of the total EPG. In the 2-day group, mature eggs accounted for 0.9% (liver) and 0.6% (small intestine) of the total EPG. In liver and small intestine, the EPG of immature and mature eggs in the 2-day group was significantly smaller than in the 1-day group. Especially, the tendency was clear in the case of the EPG of mature eggs. There were no schistosome eggs in the feces of the 2-day group. We found that administration of PZQ over 2 days separated by a 2-week interval was effective against S. japonicum.     

Host--parasite relationships of Schistosoma japonicum in mammalian hosts.

Control of schistosomiasis caused by Schistosoma japonicum has been severely hindered by the fact that several non-human mammalian species, including domesticated as well as wild animals, serve as zoonotic carriers of this infection. For effective control, it is imperative that the full host spectrum of this infection is understood. Although about 46 species of mammals are known to carry natural infection with S. japonicum, only a few might be of potential threat to human infection. Generally, in an endemic area, transmission of schistosomiasis to human depends largely on the availability and abundance of permissive hosts. Another important factor that needs to be taken into consideration in developing control measures against S. japonicum is potential strain differences. This review collates pertinent host-parasite relationship of S. japonicum in mammals in an endemic area and assesses the epidemiological significance of these findings for human infection.