Dysregulated Glucuronidation of Bilirubin Exacerbates Liver Inflammation and Fibrosis in Schistosomiasis Japonica through the NF-κB Signaling Pathway.

Hepatic fibrosis is an important pathological manifestation of chronic schistosome infection. Patients with advanced schistosomiasis show varying degrees of abnormalities in liver fibrosis indicators and bilirubin metabolism. However, the relationship between hepatic fibrosis in schistosomiasis and dysregulated bilirubin metabolism remains unclear. In this study, we observed a positive correlation between total bilirubin levels and the levels of ALT, AST, LN, and CIV in patients with advanced schistosomiasis. Additionally, we established mouse models at different time points following S. japonicum infection. As the infection time increased, liver fibrosis escalated, while liver UGT1A1 consistently exhibited a low expression, indicating impaired glucuronidation of bilirubin metabolism in mice. In vitro experiments suggested that SEA may be a key inhibitor of hepatic UGT1A1 expression after schistosome infection. Furthermore, a high concentration of bilirubin activated the NF-κB signaling pathway in L-O2 cells in vitro. These findings suggested that the dysregulated glucuronidation of bilirubin caused by S. japonicum infection may play a significant role in schistosomiasis liver fibrosis through the NF-κB signaling pathway.

An aquaporin and an aquaglyceroporin have roles in low temperature adaptation of mosquitoes (Anopheles sinensis).

Mosquitoes (Anopheles sinensis), widely geographically distributed in Asia including China, are the primary vector of the malaria parasite Plasmodium vivax and other parasitic diseases such as Malayan filariasis. An. sinensis can survive through low winter temperatures. Aquaporin channels are found in all life forms, where they facilitate environmental adaptation by allowing rapid trans-cellular movement of water (classical aquaporins) or water and solutes such as glycerol (aquaglyceroporins). Here, we identified and characterized 2 aquaporin (AQP) homologs in An. sinensis: AsAQP2 (An. sinensis aquaglyceroporin) and AsAQP4 (An. sinensis aquaporin). When expressed in frog (Xenopus laevis) oocytes, AsAQP2 transported water, glycerol, and urea; AsAQP4 transported only water. Water permeation through AsAQP2 and AsAQP4 was inhibited by mercuric chloride. AsAQP2 expression was slightly higher in adult female mosquitoes than in males, and AsAQP4 expression was significantly higher in adult males. The 2 AsAQPs were highly expressed in Malpighian tubules and midgut. AsAQP2 and AsAQP4 expression was up-regulated by blood feeding compared with sugar feeding. At freezing point (0 °C), the AsAQP4 expression level increased and An. sinensis survival time reduced compared with those at normal temperature (26 °C). At low temperature (8 °C), the AsAQP2 and AsAQP4 expression levels decreased and survival time was significantly longer compared with those at 26 °C. These results suggest that AsAQP2 and AsAQP4 have roles in water homeostasis during blood digestion and in low temperature adaptation of A. sinensis. Together, our results show that the 2 AQPs are important for mosquito diuresis after blood feeding and when exposed to low temperatures.

A retrospective analysis of schistosomiasis related literature from 2011-2020: Focusing on the next decade.

BACKGROUND: Schistosomiasis, an ancient and neglected tropical disease, which poses a huge threat to over 200 million people globally. It is necessary to have a general summary of schistosomiasis research after the new roadmap 2021-2030 issued by WHO. This study analyzes the current status of schistosomiasis research from the perspective of the One Health concept by analyzing important research literature published from 2011 to 2020, while further highlighting research priorities, difficulties, and research directions in order to propose suggestions for tropical disease studies research. METHODS: Published literature related to schistosomiasis was searched from the Web of Science Core Collection (WoSCC) database. Focusing on a visual analysis of the main research literature in the field of schistosomiasis, CiteSpace software was used to conduct co-occurrence analysis with keywords, countries, institutions, and authors. Moreover, clustering and burst analyses of keywords and co-citation analysis of authors, publications, and journals were performed. RESULTS: A total of 6638 schistosomiasis-related articles were published from 2011 to 2020, all of which can be sourced from the WoSCC database. The publication of schistosomiasis research has remained stable over the past 10 years, and contains studies in the area of human epidemiology, animal surveillance and the environment. The top five high-frequency keywords included Schistosoma mansoni, schistosomiasis, infection, praziquantel, and Schistosoma japonicum. The keywords formed nine clusters, including praziquantel, epidemiology, Schistosoma japonicum, helminths, protein, diagnosis, schistosomiasis, response, and haematobium. In recent years, most research studies focused on the mechanism of liver fibrosis, eliminating schistosomiasis, controlling risk factors, and the relationship between schistosomiasis infection and host immunity. The most productive countries include the United States, China, and Brazil, and the most productive institutions are the University of Basel, the Swiss Tropical and Public Health Institute, and the University of São Paulo. Highly productive authors include Jürg Utzinger and Donald P. McManus. At the time of writing, the author with the highest co-citation frequency (993 times) was Peter Hotez, and the journal with the highest co-citation frequency (3,720 times) was PLoS Neglected Tropical Diseases. Human schistosomiasis, published by Colley et al. (2014), was the most frequently co-cited publication (494 times). CONCLUSIONS: This study provides a preliminary description of the current status of schistosomiasis research and an initial exploration of future research directions. The One Health concept was applied in the field of schistosomiasis control, as confirmed by this bibliometric analysis. Our study provides guidance for the development of research on schistosomiasis and other neglected tropical diseases.

The Deletion of IL-17A Enhances Helicobacter hepaticus Colonization and Triggers Colitis.

Objective: IL-17 is a key regulator of the inflammatory response, and as such, it is involved in the constraint and clearance of pathogens. The mechanism of IL-17 in the pathogenesis of inflammatory bowel disease (IBD) caused by microbial infection is still unclear. Helicobacter hepaticus infection can induce colitis in many mouse strains, and thus, it has been widely used in the study of IBD pathogenesis. Methods: In this study, male C57BL/6, BALB/c, Il-10-/-, and Il-17a-/- mice were infected with H. hepaticus for several weeks. Histopathology, H. hepaticus colonization and distribution, expression of inflammatory cytokines and lysozyme, and distribution of mucus in proximal colon were examined. Results: The colonic colonization of H. hepaticus was abnormally high in Il-17a-/- mice. H. hepaticus infection caused only mild to moderate colitis symptoms in Il-17a-/- mice, including low levels of lymphocyte infiltration, epithelial cell defects, goblet cell reduction, and crypt atrophy without obvious hyperplasia in the later stage of infection. Furthermore, many inflammatory genes were significantly increased in the proximal colon of H. hepaticus-infected Il-17a-/- mice compared with C57BL/6 mice. In addition, the reduction of colonic mucus and the down-regulation of ZO-1, Claudin-1, and IL-22 were observed in Il-17a-/- mice compared with C57BL/6 mice post H. hepaticus infection. Conclusion: These results demonstrated that the deletion of IL-17A impaired the integrity of the intestinal epithelium, weakened the secretion of mucus, attenuated colonic mucosal regeneration, reduced the ability to resist microbial infection, and finally led to colitis caused by H. hepaticus.

Awareness Status of Schistosomiasis among School-Aged Students in Two Schools on Pemba Island, Zanzibar: A Cross-Sectional Study.

Schistosomiasis elimination has been set as a target in the Neglected Tropical Disease Roadmap of 2021 to 2030. The present study assessed the level of understanding, awareness and behaviors of schistosomiasis among students in Zanzibar and explored the influencing factors as the basis for reliable suggestions for the follow-up policy on schistosomiasis prevention and control. A Knowledge, Attitude and Practices (KAP) survey on students' perceptions of schistosomiasis was conducted on students from grades 4−9 at two selected schools on Pemba, Zanzibar, from May through September in 2021. A total of 217 valid participants responded to the questionnaires. T-test and chi-squared tests were used to examine the association between the dependent and explanatory variables. Multiple linear regressions were used to analyze the influencing factors of KAP. The findings indicated a lack of knowledge about schistosomiasis among the participants. Although respondents were aware of the risks of infection, they continued to engage in high-risk activities. Age, family size and presence of hematuria were found as contributing factors. Elder students performed better on knowledge (p = 0.02) and attitude (p < 0.01) scores, and students with a smaller family received higher attitude scores (p = 0.04). Practice was significantly correlated with gender (p < 0.01) and hematuria (p < 0.01). Several kinds of health education should be adopted to raise students' basic knowledge of schistosomiasis. It is also critical to make the community aware regarding schistosomiasis. Future efforts for the prevention and control of schistosomiasis should employ an integrated strategy combining communities with schools to encourage behavioral change.

Analysis of Intestinal Microflora and Metabolites From Mice With DSS-Induced IBD Treated With Schistosoma Soluble Egg Antigen.

Objective: This study aimed to analyze the changes in intestinal flora and metabolites in the intestinal contents of mice with inflammatory bowel disease (IBD) to preliminarily clarify the mechanism of action of Schistosoma soluble egg antigen (SEA) on IBD, thus, laying a research foundation for the subsequent treatment of IBD. Methods: A total of 40 Institute of Cancer Research (ICR) mice were divided into four groups: control, SEA 50 µg, dextran sulfate sodium salt (DSS), and SEA 50 µg + DSS. The overall state of the animals was observed continuously during modeling. The colonic length was measured after 10 days of modeling. The degree of colonic inflammation was observed by hematoxylin and eosin staining. 16srRNA and liquid chromatography-mass spectrometry sequencing techniques were used to determine the abundance of bacteria and metabolites in the intestinal contents of mice in the DSS and SEA 50 µg + DSS groups, and the differences were further analyzed. Results: After SEA intervention, the disease activity index score of mice with IBD decreased and the colon shortening was reduced. Microscopically, the lymphocyte aggregation, glandular atrophy, goblet cell disappearance, and colonic inflammation were less in the SEA 50 µg + DSS group than in the DSS group (p < 0.0001). After SEA intervention, the abundance of beneficial bacteria prevotellaceae_UCG-001 was upregulated, while the abundance of the harmful bacteria Helicobacter, Lachnoclostridium, and Enterococcus was downregulated in the intestinal tract of mice with IBD. The intestinal metabolite analysis showed that SEA intervention decreased the intestinal contents of glycerophospholipids (lysophosphatidylcholine, lysophosphatidylethanolamine, phatidylcholine, and phatidylethanolamine) and carboxylic acids (L-alloisoleucine and L-glutamate), whereas increased bile acids and their derivatives (3B,7A,12a-trihydroxy-5A-cholanoic acid and 3A,4B, 12a-trihydroxy-5b-cholanoic acid). Combined microbiota-metabolite analysis revealed a correlation between these differential microbiota and differential metabolites. At the same time, the changes in the contents of metabolites and differential metabolites in the two groups also correlated with the abundance of the gut microbiome. Conclusions: The study showed that SEA reduced DSS-induced inflammation in IBD and improved the symptoms of IBD in mice through the combined regulation of intestinal flora and intestinal metabolism. It suggested a potential possibility for the use of SEA in treating and regulating intestinal flora and metabolism in patients with IBD.

UHPLC-MS-Based Metabolomics Analysis Reveals the Process of Schistosomiasis in Mice.

Metabolomics, as an emerging technology, has been demonstrated to be a very powerful tool in the study of the host metabolic responses to infections by parasites. Schistosomiasis is a parasitic infection caused by schistosoma worm via the direct contact with the water containing cercaria, among which Schistosoma japonicum (S. japonicum) is endemic in Asia. In order to characterize the schistosome-induced changes in the host metabolism and further to develop the strategy for early diagnosis of schistosomiasis, we performed comprehensive LC-MS-based metabolomics analysis of serum from mice infected by S. japonicum for 5 weeks. With the developed diagnosis strategy based on our metabolomics data, we were able to successfully detect schistosomiasis at the first week post-infection, which was 3 weeks earlier than "gold standard" methods and 2 weeks earlier than the methods based on 1H NMR spectroscopy. Our metabolomics study revealed that S. japonicum infection induced the metabolic changes involved in a variety of metabolic pathways including amino acid metabolism, DNA and RNA biosynthesis, phospholipid metabolism, depression of energy metabolism, glucose uptake and metabolism, and disruption of gut microbiota metabolism. In addition, we identified seventeen specific metabolites whose down-regulated profiles were closely correlated with the time-course of schistosomiasis progression and can also be used as an indicator for the worm-burdens. Interestingly, the decrease of these seventeen metabolites was particularly remarkable at the first week post-infection. Thus, our findings on mechanisms of host-parasite interaction during the disease process pave the way for the development of an early diagnosis tool and provide more insightful understandings of the potential metabolic process associated with schistosomiasis in mice. Furthermore, the diagnosis strategy developed in this work is cost-effective and is superior to other currently used diagnosis methods.

Xiaochaihu decorction relieves liver fibrosis caused by Schistosoma japonicum infection via the HSP47/TGF-ß pathway.

BACKGROUND: Hepatic fibrosis caused by chronic infection with Schistosoma japonica remains a serious public health problem in the world. Symptoms include inflammation, liver granuloma and fibrosis, whilst treatment options are still limited. This study aims to investigate whether and how traditional Chinese medicine Xiaochaihu decoction (XCH) could mitigate liver fibrosis caused by S. japonicum infection. METHODS: BALB/c mice were infected with S. japonicum cercariae and treated with XCH for 16 weeks. Liver pathological changes were assessed by H&E and Masson staining. NIH3T3 and Raw264.7 cells were treated with S. japonicum egg antigens with or without XCH treatment. Quantitative real-time PCR, western blot, immunfluorescence and ELISA were performed to determine the changes of levels of fibrogenic markers. RESULTS: XCH protected mouse liver from injuries and fibrosis caused by S. japonicum infection and considerably reduced egg burden in a dose-dependent manner. Infection with S. japonicum caused elevation of serum ALT, AST, ALP, HA and PIIINP levels and reduction of ALB and GLOB levels, which was markedly suppressed by XCH. The upregulation of TGF-ß1, Hsp47, α-SMA, Col1A1 and Col3A1 in S. japonicum-infected mouse liver was also significantly inhibited by XCH. Schistosoma japonicum egg antigens promoted the expression of Hsp47, TGF-ß1, Timp-1, α-SMA, Col1A1 and Col3A1 in NIH3T3 cells, and TGF-ß1, CTGF, IL-13, IL-17 and IL-6 in Raw264.7 cells, which was inhibited by XCH, LY2157299 and shRNA-Hsp47. CONCLUSIONS: These results demonstrated that the hepatic protective effects of Xiaochaihu decoction were mediated by HSP47/TGF-ß axis.

Helicobacter hepaticus infection-induced IL-33 promotes hepatic inflammation and fibrosis through ST2 signaling pathways in BALB/c mice.

It has been documented that Helicobacter hepaticus (H. hepaticus) infection is linked to hepatic inflammation and fibrosis. Interleukin 33 (IL-33) is a cytokine involved in inflammatory and fibrotic diseases, but its relevance to H. hepaticus infection-induced liver inflammation and fibrosis is unknown. In this study, we found that the expression of IL-33 in mice liver was significantly induced by H. hepaticus infection at 24 weeks post infection (WPI). Immunohistochemistry analysis revealed that IL-33 was transferred from the nucleus to the cytoplasm due to infection. The quantitation of inflammatory cytokine and histopathology evaluation showed that IL-33 knockdown attenuated the H. hepaticus-induced hepatic inflammation and fibrosis. More importantly, H. hepaticus promoted the expression of the IL-33 receptor ST2 on cell surfaces, and the expression of ST2 then activated the expression nuclear factor-κB (p65), α-SMA, and Erk1/2. These observations provide novel insights into the pathogenic mechanism of hepatic inflammation and fibrosis during H. hepaticus infection.

Helicobacter hepaticus infection induces chronic hepatitis and fibrosis in male BALB/c mice via the activation of NF-κB, Stat3, and MAPK signaling pathways.

BACKGROUND: It has been documented that Helicobacter hepaticus (H hepaticus) infection is linked to chronic hepatitis and liver cancer. However, our understanding of the molecular mechanisms underlying progression of the H hepaticus-induced hepatic inflammation to cellular hepatocarcinoma is still limited. MATERIALS AND METHODS: In our study, male BALB/c mice were infected by H hepaticus for 8, 12, 16, 20, and 24 weeks. Histopathology, H hepaticus colonization dynamics, select signaling pathways, and expression of key inflammatory cytokines in the liver were examined. RESULTS: We found that H hepaticus was detectible in feces of mice at 7 days postinfection (DPI) by PCR, but it was not detected in the livers by PCR until 8 weeks postinfection (WPI). In addition, abundance of colonic and hepatic H hepaticus was progressively increased over the infection duration. H hepaticus-induced hepatic inflammation and fibrosis were aggravated over the infection duration, and necrosis or cirrhosis developed in the infected liver at 24 WPI H hepaticus infection increased levels of alanine aminotransferase and aspartate aminotransferase. Moreover, mRNA levels of Il-6 and Tnf-α were significantly elevated in the livers of H hepaticus-infected mice compared to uninfected control from 8 WPI to 24 WPI. Furthermore, Stat3, nuclear factor-κB (p65), and MAPK (Erk1/2 and p38) were activated by H hepaticus infection. CONCLUSIONS: These data demonstrated that male BALB/c mice can be used as a new mouse model of H hepaticus-induced liver diseases and that the H hepaticus-induced liver injury is triggered by NF-κB, Jak-Stat, and MAPK signaling pathways.

Identification of newly developed advanced schistosomiasis with MALDI-TOF mass spectrometry and ClinProTools analysis.

Cases of newly developed advanced schistosomiasis (NDAS) have occurred in areas where schistosomiasis transmission has been blocked for more than 25 years. The causes and pathogenesis of NDAS are still unknown. Diagnosis of NDAS relies on historical investigation and clinical symptoms, such as liver fibrosis, hepatic ascites and abnormal biochemical indexes in serum. It is important but difficult at this stage to develop a new tool for early screening and rapid diagnosis. In this study, serum peptides from thirty patients with NDAS and thirty healthy controls were captured with weak cation exchange magnetic beads, and subjected to MALDI-TOF mass spectrometry and ClinProTools analysis. Eleven peaks with m/z 924, 2661, 2953, 2991, 3241, 3884, 5337, 5905, 5943, 7766 and 9289 were decreased and three peaks with m/z 1945, 2082 and 4282 were increased in the NDAS group. The proteomic detection pattern (PDP) was established with 14 different peptide peaks, and its sensitivity and specificity were investigated with a blind test. The peptide mass fingerprints of sera from 50 NDAS patients and 100 healthy controls were double-blind subjected to the PDP method, and 50 patients and 92 healthy controls were classified as NDAS and healthy separately, which showed 100% sensitivity and 92% specificity. Our results showed that the PDP could be a new and useful method to detect NDAS.

Membrane Vesicles Are the Dominant Structural Components of Ceftazidime-Induced Biofilm Formation in an Oxacillin-Sensitive MRSA.

Methicillin-resistant Staphylococcus aureus (MRSA) has received increasing attention in recent years. However, the characteristics and relevant mechanisms of biofilm formation in oxacillin-sensitive MRSA (OS-MRSA) are poorly understood. This study was designed to characterize biofilm formation in OS-MRSA BWSA15 in response to ceftazidime (TZ) by comparing the methicillin-sensitive S. aureus (MSSA) strain BWSA23 and the oxacillin-resistant MRSA (OR-MRSA) strain BWSA11. The biofilms and biofilm-forming cells were observed by electron microscopy. Biofilms grown on microtiter plates were chemically decomposed and analyzed by Fourier transform infrared spectroscopy. The transcriptional regulation of genes associated with methicillin resistance, surface adhesion, fatty acid biosynthesis, and global regulation (sigma B) was investigated. A significant increase in biofilm formation ability (10.21-fold) and aggregation ability (2.56-fold) was observed in BWSA15 upon the treatment with TZ (16 µg/ml). The TZ-induced biofilm formation in BWSA15 was characterized by a disappearance of polysaccharide-like extracellular substances and an appearance of a large number of intercellular MVs from extracellular matrix. Few MVs were identified in the biofilms formed by BWSA11 and BWSA23. There was a significant upregulation of mecA, sigB, and fatty acid biosynthesis-associated genes and downregulation of icaA, icaD, clfA, clfB, and fnaA in BWSA15 upon the treatment with TZ. The formation of intracellular junctions of MVs in the biofilms of BWSA15 was mediated by a significant increase in the proportion of proteins as well as by an increase in the proportion of non-ionized carboxyl groups in fatty acids. This study demonstrated that beta-lactam antibiotics can induce biofilm formation in OS-MRSA, and the biofilm induction in OS-MRSA can mainly be attributed to exposed MVs with increased hydrophobicity rather than polysaccharide intercellular adhesins, cell wall-anchored surface proteins, and extracellular DNA.

Cloning and in vitro characterization of a Schistosoma japonicum aquaglyceroporin that functions in osmoregulation.

As one of the three major human pathogens that cause schistosomiasis, Schistosoma japonicum is the only one that is endemic in China. Despite great progress on schistosomiasis control over the past 50 years in China, S. japonicum transmission still occurs in certain endemic regions, which causes significant public health problems and enormous economic losses. During different life stages, parasites are able to survive dramatic osmolality changes between its vector, fresh water, and mammal host. However, the molecular mechanism of parasite osmoregulation remains unknown. To address this challenging question, we report the first cloning of an S. japonicum aquaglyceroporin (SjAQP) from an isolate from Jiangsu province, China. Expressing SjAQP in Xenopus oocytes facilitated the permeation of water, glycerol, and urea. The water permeability of SjAQP was inhibited by 1 mM HgCl2, 3 mM tetraethylammonium, 1 mM ZnCl2, and 1 mM CuSO4. SjAQP was constitutively expressed throughout the S. japonicum life cycle, including in the egg, miracidia, cercaria, and adult stages. The highest expression was detected during the infective cercaria stage. Our results suggest that SjAQP plays a role in osmoregulation throughout the S. japonicum life cycle, especially during cercariae transformation, which enables parasites to survive osmotic challenges.

New detection method in experimental mice for schistosomiasis: ClinProTool and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.

Oncomelania hupensis snails along the Yangtze River and the low positive rate and infectiosity of human and livestock schistosomiasis still pose a threat to public health in China. Adult blood flukes were recognized as Schistosoma japonicum, which are found in the portal system of the sentinel mice bred in the laboratory for 35 days after contact with the water. However, 35 days was too long from the field test to dissection, and the dissection in the laboratory was also time-consuming and labor-intensive. Serum peptides in mice at different times after infection were measured by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. ClinProTool was used to establish the proteomic detection pattern (PDP), based on the differentially expressed peptide between the infection and healthy control groups. Under experimental conditions, characteristic PDP were detected in 5 % (3/60), 35 % (21/60), 75 % (45/60), 87.93 % (51/58), and 98.15 % (53/54) of infected mice from weeks 1 to 5 post-infection, whereas ELISA and dissection examination for adult blood flukes missed the first 2 weeks. At 35 days post-infection, the infectiosity assay showed 40 % (4/10), 50 % (5/10), and 80 % (8/10) positivity with the PDP test in mice infected with 4, 6, and 10 cercariae, respectively, as well as 100 % (10/10) positivity in mice infected with 14, 18, and 22 cercariae. Five stored sera of positive sentinel mice with parasite detection were verified correctly in the PDP test. The results confirm that PDP can be used as a rapid and early detection method for S. japonicum infection in experimental mice, which are expected to apply in early surveillance for schistosomiasis.

Aquaglyceroporin function in the malaria mosquito Anopheles gambiae.

BACKGROUND INFORMATION: Anopheles gambiae is the major mosquito vector for Plasmodium falciparum malaria in sub-Saharan Africa, where it survives in stressful climates. Aquaporin water channels are expressed in all life forms, where they provide environmental adaptation by conferring rapid trans-cellular movement of water (classical aquaporins) or water plus glycerol (aquaglyceroporins). Here, we report an aquaglyceroporin homolog in A. gambiae, AgAQP3 (A. gambiae aquaglyceroporin 3). RESULTS: Despite atypical pore-lining amino acids, AgAQP3 is permeated by water, glycerol and urea, and is not significantly inhibited by 1 mM HgCl2 . AgAQP3 is expressed more heavily in male mosquitoes, yet adult female A. gambiae abundantly express AgAQP3 in Malpighian tubules and gut where large amounts of fluid exchange occur during blood meal digestion, water and nutrient absorption and waste secretion. Reducing expression of AgAQP3 by RNA interference reduces median mosquito survival at 39°C. After an infectious blood meal, mosquitoes with depleted AgAQP3 expression exhibit fewer P. falciparum oocysts in the midgut compared to control mosquitoes. CONCLUSIONS: Our studies reveal critical contributions of AgAQP3 to A. gambiae heat tolerance and P. falciparum development in vivo. SIGNIFICANCE: This study indicates that AgAQP3 may be a major factor explaining why A. gambiae is an important malaria vector mosquito in sub-Saharan Africa, and may be a potential target for novel malaria control strategies.

Impact of trehalose transporter knockdown on Anopheles gambiae stress adaptation and susceptibility to Plasmodium falciparum infection.

Anopheles gambiae is a major vector mosquito for Plasmodium falciparum, the deadly pathogen causing most human malaria in sub-Saharan Africa. Synthesized in the fat body, trehalose is the predominant sugar in mosquito hemolymph. It not only provides energy but also protects the mosquito against desiccation and heat stresses. Trehalose enters the mosquito hemolymph by the trehalose transporter AgTreT1. In adult female A. gambiae, AgTreT1 is predominantly expressed in the fat body. We found that AgTreT1 expression is induced by environmental stresses such as low humidity or elevated temperature. AgTreT1 RNA silencing reduces the hemolymph trehalose concentration by 40%, and the mosquitoes succumb sooner after exposure to desiccation or heat. After an infectious blood meal, AgTreT1 RNA silencing reduces the number of P. falciparum oocysts in the mosquito midgut by over 70% compared with mock-injected mosquitoes. These data reveal important roles for AgTreT1 in stress adaptation and malaria pathogen development in a major vector mosquito. Thus, AgTreT1 may be a potential target for malaria vector control.

[Proteomic study of the hippocampus tissue from rats with chronic Toxoplasma gondii infection].

OBJECTIVE: To observe the proteome changes in the hippocampus tissue of rats with chronic Toxoplasma gondii infection. METHODS: Six male SD rats were randomly divided into control group and infection group. Each rat in infection group was intraperitoneally injected with 4 x 10(7) purified T. gondii tachyzoites. Rats in the control group received equivalent volumes of sterile normal saline. At the fifth day post-infection, blood samples were taken from the lateral tail vein and Ciemsa staining of blood cells was performed to find Toxoplasma gondii. Rats were dissected at the 10th week post-infection, total protein in the hippocampus was separated by using two-dimensional gel electrophoresis (2-DE). After Coomassie blue staining, the Image Analysis software was used to select and separate proteins on the gel. Matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF-MS) was used for peptide mass fingerprint PMF). Proteins were identified by using Mascot software to search the MSDB and SwissProt databases. RESULTS: Microscopy examination of blood smears confirmed that the rats in infection group were all infected by 11 gondii. The number of protein spots of rats from infection group and control group was 311 +/- 19 and 327 +/- 13 respectively. Compared with the control group, 5 protein spots disappeared, 4 protein spots were up-regulated and 7 were down-regulated in the infection group. The 9 differentially expressed protein spots were identified by MALDL-TOF-MS: phosphoglycerate kinase 1, similar to alpha-enolase, glutamine synthetase, creatine kinase, creatine kinase B-type, ATP synthase, aconitase 2, mitochondrial precursor, actin and an unnamed protein. The first three proteins were up-regulated and the other five proteins were down-regulated in infection group. CONCLUSION: Nine differential expression proteins are found from the hippocampus tissue in rats chronically infected with T. gondii and normal SD rats.

Production and characterization of hirudin variant-1 by SUMO fusion technology in E. coli.

Hirudin is the most potent non-covalent inhibitor of thrombin. Several expression systems have been used to produce recombinant hirudin for pharmaceutical purposes. However, high expression of active hirudin in Escherichia coli cytoplasm has not been successful owing to the fact that heterogenetic small peptide is easily degraded in the cell. To solve this problem, we constructed a recombinant form of the hirudin variant-1 (HV1) as a fusion protein with the small ubiquitin-related modifier gene (SUMO) by use of over-lap PCR. The fusion gene His(6)-SUMO-HV1 was highly expressed in E. coli BL21 (DE3) in which the SUMO-HV1 accounts for over 30% of the soluble fraction. The fusion protein was purified by Ni-NTA affinity chromatography and cleaved by a SUMO-specific protease Ulp1 to release the HV1 with natural N-terminal. The recombinant HV1 (rHV1) was further purified by Ni-NTA affinity chromatography and then by Q anion-exchange chromatography. N-terminal sequencing result demonstrated the purified rHV1 had the same N-terminal sequence as the native hirudin. MALDI-TOF/MS analysis indicated that the molecular weight of the purified rHV1 protein was 6939.161 Da, which was similar to the theoretical molecular weight of rHV1 6,944 Da. The Chromozym TH assay result showed that the anti-thrombin activity of purified rHV1 was 8,800 ATU/mg and comparable to the specific activity of native hirudin.

[Serological investigation on Toxoplasma gondii infection in patients with unknown central nervous system diseases].

Totally 207 patients with unknown central nervous system diseases and 203 healthy persons were investigated for serum IgG of anti-Toxoplasma antibody assessed by ELISA. The serum IgG positive rate in 207 patients with unknown central nervous system diseases was 19.81%, and that in 203 health people was 5.42%, and there was a significant difference between them (P < 0.01). The IgG positive rates in different types of central nervous system diseases were different, which were 22.81%, 24.32%, 16.05%, and 18.75%, respectively in encephalopathy, epilepsy, mental disorder and neurasthenia. The IgG positive rate in different types of central nervous system diseases were significantly higher than that in healthy population (P < 0.01). The IgG positive rates in patients who contacted or did not contact cats or dogs were 32.97% and 9.48% respectively (P < 0.01). In conclusion, the infection rate in patients with unknown central nervous system diseases is higher than that in healthy persons; therefore, it is necessary to assay the serum IgG in them.

[Preliminary study on serum proteomics in mice with acute toxoplasma gondii infection by using functionalized magnetic beads and MALDI-TOF-MS technique].

OBJECTIVE: To detect and analyze the serum protein biomarkers in mice with acute Toxoplasma gondii infection. METHODS: The serum samples from 8 C57BL/6J mice with acute Toxoplasma gondii infection and 8 normal healthy paired mice were prepared with WCX magnetic beads, and then analyzed on PBS II -C mass spectrometer reader. The protein spectra of the serum samples were normalized by the Ciphergen Protein Chip software. The peak labeling was performed by the Biomarker Wizard software. The specific protein biomarkers were screened by the Biomarker Pattern software to construct a diagnostic model for acute Toxoplasma gondii infection. RESULTS: A total of 13 distinguished proteomic peaks were detected. Nine peaks were of up-regulated expressions including m/z values of 1 932.76, 1 976.85, 2 090.53, 5 004.5, 5 776.01, 5 803.05, 5 847.99, 5 877.51 and 7 501.58, respectively; and four peaks were of down-regulated expressions including m/z values of 1 866.40,4 063.71, 8 120.31 and 8 203.83, respectively. CONCLUSION: The potential protein biomarkers for acute Toxoplasma gondii infection are discovered in mouse serum by MALDI-TOF-MS combined with WCX magnetic beads.