Japonicum Eggs Research Articles

Comprehensive analysis of miRNA profiles reveals the role of Schistosoma japonicum miRNAs at different developmental stages

Schistosomiasis is an important zoonotic disease affecting up to 40 kinds of animals and 250 million people. It has been reported that the miRNAs play a role in the metabolism, differentiation, development and reproduction in many organisms. However, the roles of miRNAs regulating the development, maturation and production in schistosome in both females and males remains unclear. Here we present the dynamic transcriptome analysis of all 79 known Schistosoma japonicum miRNAs from pairing to production, including 14 days post-infection (dpi), 16, 18, 20, 22, 24, 26, 28 dpi female and male, by small RNA sequencing. The miRNA expression profiles showed time-related characteristics in male and female from paring to production, which could be clustered into three patterns, characterized by pairing stage highly expressed (cluster 1), maturating stage highly expressed (cluster 2), and egg producing stage highly expressed (cluster 3). The enrichment of miRNA cluster targeted genes in female and male were distinctly different. Network analysis of miRNAs and their target regulation showed that cluster 1 had 15 miRNAs involved in the regulation of interaction, communication, immune response in female–male and parasite–host. The other 11 miRNAs were involved in gender differentiation and the meiotic cell cycle process. In cluster 2, 11 miRNAs were involved in development and sexual maturation. In cluster 3, 45 miRNAs possibly regulate metabolism and synthesis of the substance for egg production. Analysis of the miRNA regulation network would contribute to understanding the molecular mechanism in S. japonicum development and egg production.

Toll-like receptor-2 regulates macrophage polarization induced by excretory-secretory antigens from Schistosoma japonicum eggs and promotes liver pathology in murine schistosomiasis.

Schistosomiasis is endemic to many regions of the world and affects approximately 200 million people. Conventional adaptive T cell responses are considered to be the primary contributors to the pathogenesis of Schistosoma japonicum infection, leading to liver granuloma and fibrosis. However, the functional polarization of macrophages and the associated underlying molecular mechanisms during the pathogenesis of schistosomiasis remains unknown. In the present study, we found that excretory-secretory (ES) antigens derived from S. japonicum eggs can activate macrophages, which exhibit an M2b polarization. Furthermore, ES antigen-induced M2b polarization was found to be dependent on enhanced NF-κB signaling mediated by the MyD88/MAPK pathway in a TLR2-dependent manner. In addition, the cytokine profile of the liver macrophages from wild-type-infected mice are quite distinct from those found in TLR2 knockout-infected mice by quantitative PCR analysis. More importantly, the size of granuloma and the severity of the fibrosis in the livers of TLR2-/- mice were significantly reduced compared to that in WT mice. Our findings reveal a novel role for M2b polarization in the pathogenesis of schistosome infection.

Praziquantel for a Polyp?

Schistosomiasis is the second leading cause of parasitic morbidity in the world after malaria, with five known species associated with human infection. Of these five, Schistosoma japonicum is endemic to East Asia and the Philippines, and implicated in colorectal carcinoma. We report a case of incidental colon polyps containing calcified S. japonicumeggs in an otherwise healthy man. This case highlights the asymptomatic presentation seen in the majority of patients with chronic schistosomiasis and lends insight to the pathogenesis of S. japonicum associated colorectal cancer. A 75-year-old asymptomatic man presented to his physician for routine care. His colonoscopy in 2008 was normal and he was referred for interval screening. His medical history was non-contributory. He was born in the Philippines, but moved to the United States in his twenties. He returned annually to the Philippines. He reported child-hood exposure to fresh water lakes, but denied subsequent exposure. His physical examination and laboratory studies were normal. Colonoscopy demonstrated two 2-millimeter flat sessile polyps in the cecum which were removed by cold biopsy (Figure 1). Pathology revealed benign colonic mucosa with calcified S. japonicum eggs focally in the lamina propria and predominantly in the submucosa without an associated inflammatory reaction (Figures 2 and 3). Subsequent Schistosoma species IgG antibody was negative. He was treated with praziquantel and remained asymptomatic at routine follow up. The life cycle of schistosoma involves infectious forms of the parasite passing between snails and humans. Acute infection may cause systemic symptoms and abdominal pain. Chronic infection may lead to portal hypertension, chronic diarrhea, or infect the central nervous, urinary or pulmonary systems. However, the majority of patients are asymptomatic, increasing their risk for malignancy due to prolonged untreated infection. S. japonicum is the only species strongly associated with colorectal cancer, likely due to the high volume of egg production of this species. The imbedded eggs cause an inflammatory reaction with associated risk of dysplasia. Despite this risk, there are no guidelines on appropriate interval for repeat colonoscopy in patients with S. japonicum related polyps. However, literature supports treatment with praziquantal in all patients infected with s. japonicum regardless of chronicity or symptoms based on malignancy risk.1562_A Figure 1. Cecal polyps1562_B Figure 2. Calcified eggs of schistosomiasis in the colonic mucosa and submucosa with overlying normal glandular epithelium. No dysplasia is present. (100X)1562_C Figure 3. Higher magnificent of the calcified eggs. There is no associated inflammatory reaction. (400X)

RSjP40 suppresses hepatic stellate cell activation by promoting microRNA-155 expression and inhibiting STAT5 and FOXO3a expression.

Activation of hepatic stellate cells (HSCs) is the central event of the evolution of hepatic fibrosis. Schistosomiasis is one of the pathogenic factors which could induce hepatic fibrosis. Previous studies have shown that recombinant Schistosoma japonicum egg antigen P40 (rSjP40) can inhibit the activation and proliferation of HSCs. MicroRNA‐155 is one of the multifunctional noncoding RNA, which is involved in a series of important biological processes including cell development, proliferation, differentiation and apoptosis. Here, we try to observe the role of microRNA‐155 in rSjP40‐inhibited HSC activation and explore its potential mechanisms. We found that microRNA‐155 was raised in rSjP40‐treated HSCs, and further studies have shown that rSjP40 enhanced microRNA‐155 expression by inhibiting STAT5 transcription. Up‐regulated microRNA‐155 can down‐regulate the expression of FOXO3a and then participate in rSjP40‐inhibited expression of α‐smooth muscle actin (α‐SMA) and collagen I. Furthermore, we observed microRNA‐155 inhibitor could partially restore the down‐regulation of FOXO3a, α‐SMA and collagen I expression in LX‐2 cells induced by rSjP40. Therefore, our research provides further insight into the mechanism by which rSjP40 could inhibit HSC activation via miR‐155.

Enhancing the parasitism of insect herbivores through diversification of habitat in Philippine rice fields

The diversification of farming landscapes is proposed as a method to increase farm productivity while at the same time enhancing pest regulation services provided by the natural enemies of insect herbivores. In this study, areas of diversified and conventional rice fields (with 2–14 fields per area = 0.42–1.31 ha) were established at three sites in the Philippines. The diversified areas had a series (> 40 per hectare) of small (< 2 m2) vegetable patches interspersed along rice bunds (levees). The abundance of planthopper and lepidopteran pests in these areas was monitored during two consecutive cropping seasons using sweep nets with rice yield and damage recorded at the end of each season. Mortality of brown planthopper (Nilaparvata lugens) and yellow stemborer (Scirpophaga incertulas) eggs was monitored through field exposures of infested rice plants. There were no differences in rice yields between the conventional and diversified areas; however, a range of vegetables were produced as a supplementary product from the diversified rice areas. Diversification had no effect on planthopper or leaffolder (Cnaphalocrocis medinalis) abundance at the sites. Adult stemborers (Scirpophaga incertulas) were more abundant in the diversified areas; however, stemborer damage was similar between area treatments in both years. Vegetable patches did not increase parasitism of planthopper eggs; however, egg parasitism was highest close to the vegetable patches, suggesting that egg parasitoids interacted with the patches. Mortality of stemborer eggs was higher in the diversified areas than in the conventional areas. Parasitism due to Trichogramma japonicum and other egg parasitoids was also higher in the diversified areas. These results indicate the potential for vegetable patches to enhance the biological control services provided by egg parasitoids in rice fields; however, the study also indicates a need to carefully select vegetable crops that avoid any potential benefits for rice pests. We discuss the need for further research into diversified farms as a means of achieving sustainable rice production in Asia.

Exosomes Derived from Dendritic Cells Treated with Schistosoma japonicum Soluble Egg Antigen Attenuate DSS-Induced Colitis.

Exosomes are 30–150 nm small membrane vesicles that are released into the extracellular medium via cells that function as a mode of intercellular communication. Dendritic cell (DC)-derived exosomes modulate immune responses and prevent the development of autoimmune diseases. Moreover, Schistosoma japonicum eggs show modulatory effects in a mouse model of colitis. Therefore, we hypothesized that exosomes derived from DCs treated with S. japonicum soluble eggs antigen (SEA; SEA-treated DC exosomes) would be useful for treating inflammatory bowel disease (IBD). Exosomes were purified from the supernatant of DCs treated or untreated with SEA and identified via transmission electron microscopy, western blotting and NanoSight. Acute colitis was induced via the administration of dextran sulfate sodium (DSS) in drinking water (5.0%, wt/vol). Treatment with exosomes was conducted via intraperitoneal injection (i.p.; 50 μg per mouse) from day 0 to day 6. Clinical scores were calculated based on weight loss, stool type, and bleeding. Colon length was measured as an indirect marker of inflammation, and colon macroscopic characteristics were determined. Body weight loss and the disease activity index of DSS-induced colitis mice decreased significantly following treatment with SEA-treated DC exosomes. Moreover, the colon lengths of SEA-treated DC exosomes treated colitis mice improved, and their mean colon macroscopic scores decreased. In addition, histologic examinations and histological scores showed that SEA-treated DC exosomes prevented colon damage in acute DSS-induced colitis mice. These results indicate that SEA-treated DC exosomes attenuate the severity of acute DSS-induced colitis mice more effectively than DC exosomes. The current work suggests that SEA-treated DC exosomes may be useful as a new approach to treat IBD.

MRI findings of cerebral schistosomiasis in acute stage：establishment of experimental model of acute cerebral schistosomiasis with rabbits

To establish an experimental model of acute cerebral schistosomiasis japonica and explore the MRI manifestations of acute cerebral schistosomiasis. Rabbits were divided into 3 groups with 10 rabbits in each group. The rabbits in the experimental group were directly injected with suspension fluid of Schistosoma japonicum eggs (0.9 mg, 1 ml) by the cranial drilling method, those in the negative control group were given saline (1 ml) by the same method above-mentioned, and those in the blank control group were not given any treatment. Antibiotic was given to the first two groups after the operation. The clinical manifestations of the 3 groups were observed, and the magnetic resonance imaging (MRI) was performed in 30 days post-operation, and then the brain tissues were taken for pathological examinations. All the rabbits in the experimental group exhibited inappetence, various neurological symptoms including hemiplegia, and weight loss after the operation; while those in the negative control group showed inappetence in 3 days after the operation, and 1 week later, the symptom disappeared; there were no adverse reactions in the blank control group. MRI of the experimental group showed nodular or patchy enhancement on T1WI enhancement, brain edema, abnormal ventricular dilatation, and needle augmentation. SWI displayed hypointense in the abnormal enhanced nodules and flaky hypointense on the operation brain. In the negative control group, 2 rabbits showed abnormal enhancement of the needle canal, and 1 showed mild dilatation of the ventricle. The blank control group showed normal manifestations. The pathological examinations showed abnormal appearances in 10 rabbits of the experimental group, including 6 with S. japonicum egg granuloma nodules, nonspecific granuloma nodules coexisted with perivascular inflammation; no granuloma nodules were found in the negative control group, but 2 rabbits showed vascular inflammation; the blank control group showed the normal brain tissue. An experimental model of acute cerebral schistosomiasis is successfully established in rabbits by intracranial injection of schistosome eggs. The MRI examination combined with the clinical manifestations can improve the accuracy of early diagnosis of cerebral schistosomiasis.

Clinical diagnostic value of viable Schistosoma japonicum eggs detected in host tissues

BackgroundSchistosomiasis, one of the neglected tropical diseases, is endemic in more than 70 countries. However, the clinical diagnosis of patients with a low degree of infection is an unsolved technical problem. In areas endemic for schistosomiasis japonica, proctoscopy detection of eggs has been one method used for clinical diagnosis. However, it is often a challenge to find typical live eggs and it is difficult to distinguish live eggs from large numbers of partially degraded and/or completely degraded eggs within colon biopsy tissue. To address this problem, we tested six different morphological and biochemical/molecular markers (ALP; morphological characteristics of egg; CalS (calcified substance); AOS (antioxidase); SDHG (succinic dehydrogenase) and SjR2 mRNA (retrotransposons 2 of S.japonicum genome mRNA)), including four new markers (CalS; AOS; SDHG and SjR2 mRNA.), to determine the viability of S. japonicum eggs deposited in human and mouse colon tissues. Our ultimate aim is to obtain a new method that is more sensitive, practical and accurate to clinically diagnose schistosomiasis.MethodsTissue samples were collected from mice at six different time points during S. japonicum infection with or without treatment with praziquantel (PZQ). Four new biochemical or molecular markers were used for the detection of egg viability from mouse liver and intestinal samples: CalS; AOS; SDHG and SjR2 mRNA. Subsequently, all markers were employed for the detection and analysis of eggs deposited in biopsy materials from patients with suspected schistosomiasis japonica for clinical evaluation. Microscopic examination of the egg morphology, worm burden in vivo and ALP (alkaline phosphatase) levels were used as a reference standard to evaluate the sensitivity and reliability of four new markers detecting egg viability.ResultsThe results of the study showed that the morphology of S. japonicum eggs deposited in tissues of hosts with schistosomiasis, especially cases with chronic schistosomiasis, is complex and egg viability is difficult to judge morphologically, particularly eggs with a fuzzy structure or partially modified eggs. We found that the majority of the viable schistosome eggs determined by four new markers (CalS, AOS, SDHG and SjR2 mRNA) were morphologically difficult to identify.ConclusionsAmong the markers, the most sensitive and specific method was the detection of SjR2 mRNA and the most simple, rapid and practical method was the detection of SDHG. Therefore, the detection of SDHG is the most practical for clinical application and its use could improve the accuracy in diagnosing active schistosome infection.

Egg antigen p40 of Schistosoma japonicum promotes senescence in activated hepatic stellate cells via SKP2/P27 signaling pathway

Schistosomiasis is characterized by egg deposition, granulomatous inflammatory reaction and then subsequent hepatic fibrosis formation. Activated HSCs are regarded as the main effector cells in the progression of liver fibrosis and induction of senescence in hepatic stellate cells (HSCs) is vital to the reversion of hepatic fibrosis. Our previous work has showed that S. japonicum egg antigen p40 (Sjp40) could promote HSCs senescence via a STAT3/p53/p21 mechanism. In this paper, the major aim was to explore whether there are other signaling pathways in the process of Sjp40-induced HSCs aging and the underlying effect of SKP2/P27 signal pathway in this procedure. We observed the Sjp40-induced decrease of α-SMA and the senescence of LX-2 cells, and Sjp40 could upregulate P27 and downregulate the protein level of SKP2. The senescence induced by Sjp40 might be reversed in LX-2 cells that treated with P27-specific siRNA or with SKP2-special over-expression plasmid. In addition, we also demonstrated that the decreased expression of P-Rb and α-SMA induced by Sjp40 were partly restored by SKP2-overexpression. These data suggest that Sjp40 might inhibit HSCs activation by promoting cellular senescence via SKP2/P27 signaling pathway, which put forward novel mechanism in the treatment of liver fibrosis.

Characterization of Schistosoma japonicum CP1412 protein as a novel member of the ribonuclease T2 molecule family with immune regulatory function

BackgroundSchistosome infection typically induces a polarized Th2 type host immune response. As egg antigen molecules play key roles in this immunoregulatory process, clarifying their functions in schistosomiasis would facilitate the development of vaccine and immunotherapeutic methods. Schistosoma japonicum (Sj) CP1412 (GenBank: AY57074.1) has been identified as a new member of the RNase T2 family with immune regulatory functions.MethodsThe expression plasmid Sj CP1412-pET28a was constructed and transformed into bacteria for production of recombinant Sj CP1412 protein (rSj CP1412) via IPTG induction. The RNase activity of Sj CP1412 was predicted by bioinformatic analysis and confirmed by digesting the yeast tRNA with rSj CP1412.C57BL/6j mice were immunized with rSj CP1412, and its immune regulatory effects in vivo and in vitro were investigated. Meanwhile, the relationship between the RNase activity of Sj CP1412 and its immune regulation was observed.ResultsSj CP1412 was confirmed as a novel RNase T2 family protein with RNase activity. Immunoblotting and RT-PCR analyses demonstrated Sj CP1412 as a protein exclusively secreted/excreted from eggs, but not cercariae and adult worms. Stimulating RAW264.7 macrophages with rSj CP1412 raised the expression of CD206, Arg-1 and IL-10, which are related to M2 type macrophage differentiation. Stimulating dendritic cells (DCs) with rSjCP1412 failed to induce their maturation, and the recombinant protein also inhibited LPS-stimulated DC maturation. Depletion of Sj CP1412 from soluble egg antigen (SEA) impaired the ability of SEA to induce M2 type polarization of RAW264.7 macrophages. Immunizing mice with rSj CP1412 induced high antibody titers, increased serum IL-4 and TGF-β levels and splenic CD4 + CD25 + Foxp3 + T cells, downregulated serum IFN-γ levels and alleviated the egg granuloma pathology of schistosome infection. In vitro stimulation by rSj CP1412 significantly increased CD4 + CD25 + Foxp3 + T cell numbers in splenocytes of healthy mice. The rSj CP1412 protein with RNase activity inactivated by DEPC failed to induce M2 surface marker CD206 expression in RAW264.7 macrophages.ConclusionsThe Sj CP1412 protein expressed specifically in S. japonicum eggs is a novel member of the RNase T2 family. Similar to Omega-1 of Schistosoma mansoni, the Sj CP1412 protein drives polarization of the host Th2 immune response, which is dependent on its RNase activity. These data provide new evidence towards understanding the immune regulatory role of RNase T2 family proteins during schistosome infection.

Release of extracellular vesicles containing small RNAs from the eggs of Schistosoma japonicum.

Background Schistosoma japonicum is a parasitic flatworm that causes human schistosomiasis. Secreted extracellular vesicles (EVs) play a key role in pathogen-host interfaces. Previous studies have shown that S. japonicum adult worms can release microRNA (miRNA)-containing EVs, which can transfer their cargo to mammalian cells and regulate gene expression in recipient cells. Tissue-trapped eggs are generally considered the major contributor to the severe pathology of schistosomiasis; however, the interactions between the host and parasite in this critical stage remain largely unknown.MethodsThe culture medium for S. japonicum eggs in vitro was used to isolate EVs. Transmission electron microscopy (TEM) analysis was used to confirm that vesicles produced by the eggs were EVs based on size and morphology. Total RNA extracted from EVs was analyzed by Solexa technology to determine the miRNA profile. The in vitro internalization of the EVs by mammalian cells was analyzed by confocal microscopy. The presence of EVs associated miRNAs in the primary hepatocytes of infected mice was determined by quantitative real-time PCR (qRT-PCR).ResultsEVs were isolated from the culture medium of in vitro cultivated S. japonicum eggs. TEM analysis confirmed that nanosized vesicles were present in the culture medium. RNA-seq analysis showed that the egg-derived EVs contained small non-coding RNA (sncRNA) populations including miRNAs, suggesting a potential role in host manipulation. This study further showed that Hepa1-6, a murine liver cell line, internalized the purified EVs and their cargo miRNAs that were detectable in the primary hepatocytes of mice infected with S. japonicum.Conclusions Schistosoma japonicum eggs can release miRNA-containing EVs, and the EVs can transfer their cargo to recipient cells in vitro. These results demonstrate the regulatory potential of S. japonicum egg EVs at the parasite-host interface.Electronic supplementary materialThe online version of this article (doi:10.1186/s13071-016-1845-2) contains supplementary material, which is available to authorized users.

Role of goat in transmission of schistosomiasis japonica III Effect of temperature and humidity on survival of eggs in goat feces and egg survival in natural environments

To investigate the survival of Schistosoma japonicum eggs in goat feces in natural marshlands and the factors affecting its survival, so as to provide evidences for understanding of the role of eggs in goat feces in the transmission of schistosomiasis and the development of the interventions pertaining to disease control and elimination. The goat animals of schistosomiasis japonica were modeled in laboratory, and the feces of infected goat were collected. In laboratory, the effects of environmental temperature and water content in goat feces on egg hatching were evaluated, and in the field, the effect of duration of goat feces on marshland on egg hatching and the effect of direct sunshine on egg survival were evaluated. At 25°C in laboratory, the hatching rate of eggs in goat feces washigh-positively correlated with the water content in goat feces (r = 0.87). If the water content reduced to 7.6% in goat feces, the eggs in goat feces lost the ability to hatch. Under the same water content in goat feces, the hatching rate of eggs gradually decreased with the extension of the duration of exposure of goat feces to -5 °C, which reduced to 0 following 5 h exposure. At 5, 15 and 25 °C, the hatching rates of eggs gradually decreased with the extension of the duration of exposure of goat feces, and themiracidium hatching ratesof eggs were 2.3%, 5% and 0.9% respectively following the exposure for 52 d. At 35°C, the hatching rate of eggs gradually decreased with the extension of the duration of exposure, which reduced to 0 following 13 d exposure. In winter (-2-10 °C), the hatching rate of eggs gradually decreased with the extension of the duration of exposure of goat feces on marshlands, which reduced to 0 after 21 d of exposure, and in spring (16-19 °C), the hatching rate of eggs gradually decreased with the extension of the duration of exposure of goat feces on marshlands, which reduced to 0.9% after 5 d of exposure. At the same time point on the same marshland, the hatching rate of eggs in goat feces exposed to marshlands with direct sunshine was lower than that without direct sunshine. The survival of S. japonicum eggs in goat feces is associated with environmental temperature and water content (humidity) in goat feces, and the temperature and humidity are major natural factors affecting egg hatching.

Strengthening the control of goat schistosomiasis to facilitate the progress towards elimination of schistosomiasis in China

Although great success has been achieved in schistosomiasis control, schistosomiasis japonica remains a publichealth concern in China. Schistosoma japonicum is found to naturally infect over 40 mammalian animals. The implementation of the integrated schistosomiasis control strategy emphasizing infectious source control since 2004, which integrates replacement of bovines with machines, breeding domestic animals in fences, building safe pastures and chemotherapy of infected bovines, results in a clear-cut reduction in the prevalence of S. japonicum infection in both humans and bovines, as well as the areas of infected snail habitats, and the national schistosomiasis control program is moving from transmission control towards transmission interruption and elimination. It has been found that goat is highly susceptible to S. japonicum infection, and previous epidemiological data have shown a high prevalence of infection in goat. However, the role of goat in the transmission of schistosomiasis japonica has not been paid much attention, and there are few systematic surveys to evaluate the role of goat in schistosomiasis transmission in China to date. Professor Liang Yousheng's group investigated S. japonicum development and reproduction (egglaying) in goat body, environmental contamination by goat feces, and the effect of temperature and humidity on the survival of S. japonicum eggs in goat feces. Their findings further demonstrate the role of goat in the transmission of schistosomiasis japonica. In addition, they proposed, based on their findings and previous reports, that the management of goat should be integrated into the national schistosomiasis control program in China, since goat is virtually one of the major infectious sources of schistosomiasis japonica in China. Moreover, this group improved the fecal hatching test and optimized the parasitological technique for diagnosis of S. japonicum infection in goats. These innovative studies fill in the gaps of goat schitsosomiasis japonica research in China, and the research outcomes will enrich the currently implemented integrated schistosomiasis control strategy emphasizing infectious source control, and are believed to play a critical role in schistosomiasis elimination in China. Since schistosomiasis seriously affects goat husbandry development and local famer income, and goat has become a major infectious source of schistosomiasis japonica in China, the control of goat schistosomiaiss will facilitate the progress towards the elimination of schistosomiasis in China, and the following research priorities are suggested: (1) to emphasize the control of goat schistosomiasis, and to integrate the comprehensive management of goat into the national schistosomiasis control program in China; and (2) to develop new techniques, products and interventions for the control of goat schistosomiasis.

The effect of Bassiatin as a novel antiseptic agent on methicillin-resistant Staphylococcus aureus (MRSA)

Pyriproxyfen is a juvenile hormone analog that has been widely used to control whiteflies and various other insect pests. Serangium japonicum is a dominant coccinellid predator of Bemisia tabaci (biotype B or MEAM1) in many agricultural systems in China. We assessed the toxicity of pyriproxyfen to S. japonicum to explore the compatibility of using pyriproxyfen and S. japonicum together to control B. tabaci. Eggs and pupae of S. japonicum were dipped into the pyriproxyfen solutions [100 mg (AI)/L and 200 mg (AI)/L] or solvent and water controls. Larvae were fed pyriproxyfen-treated B. tabaci eggs and the adults were fed pyriproxyfen-treated fourth instar nymphs. Significant effects of pyriproxyfen on all immature stages of S. japonicum were found, and the degree of effects depends on the stage treated and the concentration tested. Pyriproxyfen caused significant mortality of S. japonicum eggs, with only 3.3% survival rates at the higher concentration and 17.3% at the lower concentration. Development time of S. japonicum of treated larval instars to adult emergence was longer than that of the untreated larvae. The fourth instar was the most susceptible larval stage, while the egg was the most sensitive stage of S. japonicum. Moreover, pyriproxyfen significantly decreased oviposition by S. japonicum adults and subsequent offspring hatching rate. In conclusion, the S. japonicum egg was the most sensitive stage to pyriproxyfen. Thus for a combined pyriproxyfen–S. japonicum whitefly control program, applying pyriproxyfen after S. japonicum egg hatch would minimize the effects of pyriproxyfen on the S. japonicum population.

Application of automatic photography in Schistosoma japonicum miracidium hatching experiments

To explore the value of automatic photography in the observation of results of Schistosoma japonicum miracidium hatching experiments. Some fresh S. japonicum eggs were added into cow feces, and the samples of feces were divided into a low infested experimental group and a high infested group (40 samples each group). In addition, there was a negative control group with 40 samples of cow feces without S. japonicum eggs. The conventional nylon bag S. japonicum miracidium hatching experiments were performed. The process was observed with the method of flashlight and magnifying glass combined with automatic video (automatic photography method), and, at the same time, with the naked eye observation method. The results were compared. In the low infested group, the miracidium positive detection rates were 57.5% and 85.0% by the naked eye observation method and automatic photography method, respectively (χ2 = 11.723, P < 0.05). In the high infested group, the positive detection rates were 97.5% and 100% by the naked eye observation method and automatic photography method, respectively (χ2 = 1.253, P > 0.05). In the two infested groups, the average positive detection rates were 77.5% and 92.5% by the naked eye observation method and automatic photography method, respectively (χ2 = 6.894, P < 0.05). The automatic photography can effectively improve the positive detection rate in the S. japonicum miracidium hatching experiments.

Screening and identification of DNA aptamers toward Schistosoma japonicum eggs via SELEX.

Schistosomiasis is a major parasitic disease caused by blood flukes of the genus Schistosoma. Several million people all over the world are estimated to suffer from severe morbidity as a consequence of schistosomiasis. The worm’s eggs, which cause the symptoms of schistosomiasis, are generally used to diagnose the disease. In this study, we employed egg-based systematic evolution of ligands by exponential enrichment (egg-SELEX) and identified a panel of ssDNA aptamers specifically binding to eggs derived from S. japonicum. Among these, two aptamers LC6 and LC15 exhibited strong binding to and specific recognition of S. japonicum eggs, but not eggs from Fasciolopsis buski, Enterobius, Ascaris or Clonorchis sinensis. Furthermore, tissue imaging results revealed that LC15 could recognize S. japonicum eggs laid in liver tissues with a detection ratio of 80.5%. Collectively, therefore, we obtained useful aptamers specifically recognizing S. japonicum eggs, which will facilitate the development of an effective tool for both schistosomiasis diagnosis and drug delivery.

Evaluation of Lethal Giant Larvae as a Schistosomiasis Vaccine Candidate

Schistosomiasis is a neglected tropical disease of humans, and it is considered to be the second most devastating parasitic disease after malaria. Eggs produced by normally developed female worms are important in the transmission of the parasite, and they responsible for the pathogenesis of schistosomiasis. The tumor suppressor gene lethal giant larvae (lgl) has an essential function in establishing apical-basal cell polarity, cell proliferation, differentiation, and tissue organization. In our earlier study, downregulation of the lgl gene induced a significant reduction in the egg hatching rate of Schistosoma japonicum (Sj) eggs. In this study, the Sjlgl gene was used as a vaccine candidate against schistosomiasis, and vaccination achieved and maintained a stable reduction of the egg hatching rate, which is consistent with previous studies, in addition to reducing the worm burden and liver egg burden in some trials.

Heat Shock Protein 60 in Eggs Specifically Induces Tregs and Reduces Liver Immunopathology in Mice with Schistosomiasis Japonica.

BackgroundParasitic helminths need to suppress the host immune system to establish chronic infections. Paradoxically, immunosuppression induced by the worm also benefits the host by limiting excessive inflammation and tissue damage, which remains the major cause leading to serious morbidity and mortality. Regulatory T cells (Tregs) are key immune regulators of this mutualism. The successive rise in Tregs during schistosome infection plays a critical role in immunoregulation. We and others previously showed that Schistosoma japonicum (S. japonicum) egg antigens (SEA) induce Tregs both in vitro and in vivo. In addition, we identified that SjHSP60 derived from SEA significantly induces Tregs in vivo and in vitro. However, the contribution of SjHSP60 in SEA to Treg induction and the related mechanisms of the Treg induction have not yet been identified.Methodology/Principal FindingsIn this study, we showed that S. japonicum stress protein HSP60 (SjHSP60) was constitutively and extensively expressed in eggs of S. japonicum. SjHSP60 specially induced Tregs in vivo and in vitro without inducing other CD4+ T sub-populations including Th1, Th2 and Th17 cells. Furthermore, we showed that the SjHSP60-depleted SEA almost lost the ability in vitro and displayed a significant impaired ability to induce Tregs in vivo. Finally, our study illustrated that the mechanisms of SjHSP60-mediated induction of Tregs are through both conversion of CD4+CD25- T cells into CD4+CD25+Foxp3+ Tregs and expansion of preexisting CD4+CD25+Foxp3+ Tregs in a TLR4-dependent manner.Conclusions/SignificanceCollectively, our findings identify SjHSP60 as a major parasitic contributor of Treg induction in S. japonicum egg antigens, which not only contributes to the better understanding of the mechanism of immunoregulation during helminth infection, but also suggests its potential as a therapeutic target for control of immunopathology, allergic and autoimmune diseases.

Antibody Signatures Reflect Different Disease Pathologies in Patients With Schistosomiasis Due to Schistosoma japonicum.

Infection with Schistosoma japonicum causes high levels of pathology that is predominantly determined by the cellular and humoral response of the host. However, the specific antibody response that arises during the development of disease is largely undescribed in Asian schistosomiasis-endemic populations. A schistosome protein microarray was used to compare the antibody profiles of subjects with acute infection, with early or advanced disease associated with severe pathology, with chronic infection, and subjects exposed but stool negative for S. japonicum eggs to the antibody profiles of nonexposed controls. Twenty-five immunodominant antigens were identified, including vaccine candidates, tetraspanin-related proteins, transporter molecules, and unannotated proteins. Additionally, individuals with severe pathology had a limited specific antibody response, suggesting that individuals with mild disease may use a broad and strong antibody response, particularly against surface-exposed proteins, to control pathology and/or infection. Our study has identified specific antigens that can discriminate between S. japonicum-exposed groups with different pathologies and may also allow the host to control disease pathology and provide resistance to parasite infection.

An ex vivo model for studying hepatic schistosomiasis and the effect of released protein from dying eggs.

BackgroundWe report the use of an ex vivo precision cut liver slice (PCLS) mouse model for studying hepatic schistosomiasis. In this system, liver tissue is unfixed, unfrozen, and alive for maintenance in culture and subsequent molecular analysis.Methods and FindingsUsing thick naive mouse liver tissue and sterile culture conditions, the addition of soluble egg antigen (SEA) derived from Schistosoma japonicum eggs, followed 4, 24 and 48hrs time points. Tissue was collected for transcriptional analysis and supernatants collected to quantitate liver enzymes, cytokines and chemokines. No significant hepatotoxicity was demonstrated by supernatant liver enzymes due to the presence of SEA. A proinflammatory response was observed both at the transcriptional level and at the protein level by cytokine and chemokine bead assay. Key genes observed elevated transcription in response to the addition of SEA included: IL1-α and IL1-β, IL6, all associated with inflammation. The recruitment of antigen presenting cells was reflected in increases in transcription of CD40, CCL4 and CSF1. Indications of tissue remodeling were seen in elevated gene expression of various Matrix MetalloProteinases (MMP3, 9, 10, 13) and delayed increases in TIMP1. Collagen deposition was significantly reduced in the presence of SEA as shown in COL1A1 expression by qPCR after 24hrs culture. Cytokine and chemokine analysis of the culture supernatants confirmed the elevation of proteins including IL6, CCL3, CCL4 and CXCL5.ConclusionsThis ex vivo model system for the synchronised delivery of parasite antigen to liver tissue provides an insight into the early phase of hepatic schistosomiasis, corresponding with the release of soluble proteins from dying schistosome eggs.