Antischistosomal activity of hederacochiside C against Schistosoma japonicum harbored in experimentally infected animals.

The present study was undertaken to investigate whether hederacochiside C (HSC) possesses antischistosomal effects and anti-inflammatory response activities in Schistosoma japonicum-infected mice. Different concentrations of HSC were administrated to the mice infected by schistosomula or adult worm by intravenous injection twice a day for five consecutive days. The total worm burden, female worm burden, and the egg burden in liver of mice treated with 400 mg/kg HSC were fewer than those in non-treated ones. Murine immune responses following HSC treatment were investigated using enzyme-linked immunosorbent assays (ELISA). Our results indicated that 200 mg/kg HSC could reduce the expression of IgG, tumor necrosis factor (TNF)-α, interleukin (IL)-4 and IL-17 in comparison to infected group, exhibiting best immunomodulatory effects. In addition, scanning electron microscopical examination revealed that male worms treated with HSC lost their normal surface architecture since its surface showed extensive swelling, erosion, and peeling in tegumental regions. Remarkable amelioration was noticed in histopathological investigations, and 200 mg/kg HSC treatment could reduce the size of granulomatous inflammatory infiltrations in the liver which was reflected in nearly normalization of liver architecture. These results suggested that HSC had potential antischistosomal activity and provided a basis for subsequent experimental.

Effect of photoperiod change on chronobiology of cercarial emergence of Schistosoma japonicum derived from hilly and marshy regions of China.

The chronobiology of cercarial emergence appeared to be a genetically controlled behavior, adapted to definitive host species, for schistosome. However, a few physiological and ecological factors, for example the change of photoperiod, were reported to affect the rhythmic emergence of cercariae. Therefore, the effect of photoperiod change on cercarial emergence of two Schistosoma japonicum isolates, the hilly and the marshland, was investigated. Four shedding experiments each under a different photoperiod were conducted. Under a natural photoperiod, two distinct shedding modes, one from the hilly region and one from the marshland, were observed. Under a reversed photoperiod, the regular pattern (i.e. under a natural photoperiod) of S. japonicum cercarial emergence was reversed for the marshland isolate and disappeared for the hilly isolate. With an input of a 2 h darkness from 7am to 9am, the cercarial emergence peak were delayed for the two isolates; whereas with an input of a 2 h darkness from 5pm to 7pm, neither effect on the cercarial emergence rhythm was observed. The total cercariae emerged for both parasite isolates varied with a different photoperiod. The results indicate that the change of photoperiod could affect the chronobiology of S japonicum cercarial emergence.

A multifactorial analysis of the pregnancy outcomes in cytomegalovirus-infected women.

AIMS: To investigate the impacts of cytomegalovirus (CMV) viral load, TORCH (toxoplasmosis, others, rubella, CMV and herpes) coinfections, CMV glycoprotein B (gB) genotypes and maternal genetic polymorphisms on pregnancy outcomes among CMV-infected women. METHODS: A total of 731 CMV-infected pregnant women (634 and 97 with normal and adverse pregnancy outcomes, respectively) were recruited. CMV load quantification and screening of TORCH coinfections were performed by using real-time polymerase chain reaction (PCR) and immunodetection techniques, respectively. Genotyping of CMV gB and maternal NFKB1 -94 ins/del, NFKBIA -826C/T and -881A/G polymorphisms was performed by using PCR-restriction fragment length polymorphism. RESULTS: We found that the mean CMV viral load in women with adverse pregnancy outcomes was significantly higher than that in women with normal outcomes at all pregnancy stages (p < 0.01). We also found that TORCH coinfections resulted in a 1.65-fold (95% CI = 1.00-2.73) increase in the risk of adverse pregnancy outcomes (p = 0.05). Additionally, we noticed no significant difference in the distribution of CMV gB genotypes between women with normal and adverse pregnancy outcomes (p = 0.42). We also observed that the ins/ins variant genotype of the NFKB1 polymorphism could reduce the risk of adverse pregnancy outcomes (OR = 0.38, 95% CI = 0.15-0.98; p = 0.04). CONCLUSION: CMV viral load, TORCH coinfections and maternal NFKB1 polymorphism could influence pregnancy outcomes among CMV-infected women.

[Cloning, expression and antigenic analysis of merozoite surface protein MSPDBL2-DBL2 domain from Plasmodium falciparum].

OBJECTIVE: To clone and express the DBL domain of Plasmodium falciparum merozoite surface protein MSPDBL2(DBL2), and investigate its antigenicity. METHODS: The DBL2 fragment was amplified by PCR and cloned into pET28a vector. The recombinant pET28a-DBL2 plasmid was transformed into E. coli BL21 (DE3) and protein expression was induced by IPTG. The expressed product was purified by Ni-NTA affinity chromatography, and analyzed by SDS-PAGE and Western blotting. RESULTS: DBL2 gene fragment of Plasmodium falciparum merozoite surface protein MSPDBL2 (950 bp) was obtained by PCR. The recombinant pET28a-DBL2 plasmid was identified by PCR, double enzyme digestion, and DNA sequencing. The recombinant DBL2 protein was expressed in an inclusion body form with Mr 340,000 after being induced with IPTG. Moreover, the purified recombinant DBL2 protein was recognized by sera from patients with falciparum malaria. CONCLUSION: The recombinant pET28a-DBL2 plasmid has been constructed. The purified rDBL2 protein shows adequate antigenicity.

Laboratory evaluation of the molluscicidal activity of Pulsatilla chinensis (Bunge) Regel saponins against the snail Oncomelania hupensis.

OBJECTIVE: To observe the toxicity of Pulsatilla chinensis (Bunge) Regel saponins (PRS) against Oncomelania hupensis (O. hupensis). METHODS: O. hupensis snails were exposed to 40% and 80% of 24 h LC50 of PRS for 24 h, and then choline esterase (CHE), alanine aminotransferase (ALT), alkaline phosphatase (ALP), lactate dehydrogenase (LDH) activities in cephalopodium and liver of snails were determined. Niclosamide (NIC) was used as the reference molluscicide. Zebra fish lethality test was evaluated to non-target aquatic species of PRS. RESULTS: The molluscicidal activity of PRS (LC50 at 24 h: 0.48 mg/L) was similar to that of NIC (LC50 at 24 h: 0.16 mg/L). Significant alterations about CHE, ALP, and ALT activities both in the cephalopodium and the liver of snails were observed when O. hupensis was exposed to 40% and 80% LC50 of PRS or NIC for 24 h. PRS and NIC could not affect LDH activity in the cephalopodium and the liver. Lower toxicity to fish of PRS was observed up to the highest concentration tested than NIC. CONCLUSION: PRS, as compared with the reference molluscicide NIC, is thought to be used for the control of harmful vector snails safely.

[Localization of acetylcholinesterase in the nervous system of Cotylophoron indicum].

The nervous system of Cotylophoron indicum was studied by using acetylcholine esterase histochemical staining techniques. Cranial ganglia and transverse commissure situate at dorso-lateral body between oral sucker and genital sucker. From the cranial ganglia four pairs of nerves proceed cephalad and connect with nerve network of the oral sucker. The posterior nerve cords from the cranial ganglia consist of 3 pairs and the ventral ones are the stoutest and longest nerves. A few branches from the 3 pairs of nerve cords connect to ventral sucker. There is a developed nerve network distributed in its genital sucker. The nerve fibers on body surface in pairs and parallel are diagonal and cross to form a nerve network on body surface. Three kinds of neurocytes distribute at the prosomal region. Results show that the nervous system structure of C. indicum is consistent with the essential features of Digenea, but more special and complicated around genital sucker.

[In vitro co-cultivation of Toxoplasma gondii tachyzoites with rat brain astrocytes].

Purified astrocytes were cultured in plates. When astrocytes grew over 80% of the plate, tachyzoites of Toxoplasma gondii RH strain were added for co-culture. In the period of 0-72 h, change of the astrocytes and tachyzoites was observed after Giemsa staining. In 0-48 h, monodansylcadaverine (MDC) was used to study the action of autophagy in the process of tachyzoites invading astrocytes. At 1 h co-culture, tachyzoites had entered in astrocytes and the autophagosomes appeared. At 4 h, the autophagosomes increased pronouncedly. However, after 12 h, number of autophagosomes considerably decreased and damage of the cells occurred. 48 h later, autophagosomes disappeared and more astrocytes were destroyed. At 72 h most cells destroyed and tachyzoites were released. The result showed that autophagy is inhibited when the astrocytes were in vitro infected by tachyzoites.

[Transcription profile of PfRON4 gene in Plasmodium falciparum erythrocytic stage].

OBJECTIVE: To reveal the transcription profile of PfRON4 gene in Plasmodium falciparum erythrocytic stage. METHODS: P. falciparum schizonts were purified by differential centrifugation on a Percoll-sorbitol gradient, after which the released merozoites were allowed to invade uninfected erythrocytes for 4 hours before the clearance of all remaining schizonts using 5% D-sorbitol. The cultured synchronous parasites were harvested for RNA assay immediately, 24 hours later, and then at every 6th hour. PfRON4 and related genes (PfAMA1 and PfRhopH2) were amplified by real-time PCR for establishing standard curves to evaluate the copy number of genes. RESULTS: P. falciparum parasites were well synchronized. Those quantative analyses were reliable because the R value of standard curves were more than 0.98 and the melting curve showed a single peak. When parasites were in the schizont stage, PfRON4 gene transcription reached a peak in 36-40 hours after invasion. CONCLUSION: The transcription of PfRON4 peaks at mature schizont stage, suggesting that the PfRON4 gene may involve in erythrocyte-invasion of P. falciparum.

[Analysis of karyotype and preparation of C-band and G-band in Schistosoma japonicum].

Chromosomes of Schistosoma japonicum were prepared by usual air drying method, C-band and G-band were made by modified BSG method and enzyme digestion method respectively. Results showed that the karyotype of S. japonicum was 4m + 6Sm + 4St + 2sex chromosome and the C-band formula was 2n = 5CIt + 4CI(+) + 3CI + 2CT +2CT.

[Studies on haemocytes of Oncomelania hupensis].

OBJECTIVES: To establish method for collecting haemocytes of Oncomelania hupensis and study its morphology and immunological importance. METHODS: Referring to the method of haemocytes collection from peripheral lymphoid organ, suspension technique was used for collection of haemocytes from snails, which were then Giemsa-stained and observed under microscope. Stained by gentian violet, number of haemocytes was counted and compared with that of conventional squashing method and needling method by ANOVA and Dunnett-t test. Supernatant from freeze thawing haemocytes was applied for the tests of immuno-precipitation, bacteriostasis, and phagocytosis. SDS-PAGE was used to analyze relative molecular mass of protein ingredients. RESULTS: Four kinds of haemocytes were found: round cells with filiform filopodia, acidophilic and basophilic round cells both without filiform filopodia, and spindle cells. The average diameter of the 4 type cells was 10.93, 6.13, 6.08, and 11:06 microm, and occupied 50%, 30%, 5%, and 15% respectively. The mean of haemocytes received from suspension, squashing and needling methods was 15 000, 6 600 and 300/ml respectively. ANOVA analysis showed F=281.47, P<0.01, and Dunnett-t test revealed t1=15.67, P<0.01 between suspension and squashing methods, and t2=24.50, P<0.01 between suspension and needling method. The supernatant of haemocytes showed precipitation with SEA, bacteriostasis with Staphylococcus aureus and Escherichia coli, and 86% phagocytosis and 46% bactericide to Candida albicans. SDS-PAGE revealed approximate Mr 112 300, 107 100, 972 00, 73 500, 600 00, 12 000 of the protein ingredients of the haemocytes. CONCLUSIONS: Large amount of haemocytes has been obtained from the snails by suspension method, and the cells show immunological activities.

[The relationship between the efficacy of interferon-alpha treatment and the variation of perforin protein expression in peripheral blood mononuclear cells in chronic hepatitis B patients].

OBJECTIVE: To study the relationship between the efficacy of interferon-alpha and the variation of perforin protein expression in the peripheral blood mononuclear cells in 35 patients with chronic hepatitis B (CHB). METHODS: The perforin protein in the peripheral blood mononuclear cells was detected by immunocytochemistry technique. RESULTS: The level of the perforin protein expression in the peripheral blood mononuclear cells was significantly higher in the post-treatment group with interferon-alpha than in the pre-treatment group. At the end of the treatment with interferon-alpha, there were 12 cases of complete responders, 14 cases of partial responders, and 9 cases of non-responders. After interferon-alpha treatment, the mean level of the perforin protein expression in the peripheral blood mononuclear cells was 12.1%, 6.9% and 3.9% respectively, and there existed significant differences among the three groups. Moreover, before treatment, the level of the perforin protein expression in the complete responder group was significantly higher compared to the partial responder group or the non-responder group. CONCLUSION: Treatment with interferon-alpha can increase the perforin protein expression in the peripheral blood mononuclear cells in patients with CHB. The variation of perforin protein expression in the peripheral blood mononuclear cells may be closely related to the efficacy of interferon-alpha treatment against hepatitis B virus.

Serological Prevalence of Schistosoma japonicum in Mobile Populations in Previously Endemic but Now Non-Endemic Regions of China: A Systematic Review and Meta-Analysis.

BACKGROUND: Schistosomiasis japonica has been resurging in certain areas of China where its transmission was previously well controlled or interrupted. Several factors may be contributing to this, including mobile populations, which if infected, may spread the disease. A wide range of estimates have been published for S. japonicum infections in mobile populations, and a synthesis of these data will elucidate the relative risk presented from these groups. METHODS: A literature search for publications up to Oct 31, 2014 on S. japonicum infection in mobile populations in previously endemic but now non-endemic regions was conducted using four bibliographic databases: China National Knowledge Infrastructure, WanFang, VIP Chinese Journal Databases, and PubMed. A meta-analysis was conducted by pooling one arm binary data with MetaAnalyst Beta 3.13. The protocol is available on PROSPERO (No. CRD42013005967). RESULTS: A total of 41 studies in Chinese met the inclusion criteria, covering seven provinces of China. The time of post-interruption surveillance ranged from the first year to the 31st year. After employing a random-effects model, from 1992 to 2013 the pooled seroprevalence ranged from 0.9% (95% CI: 0.5-1.6%) in 2003 to 2.3% (95% CI: 1.5-3.4) in 1995; from the first year after the disease had been interrupted to the 31st year, the pooled seroprevalence ranged from 0.6% (95% CI: 0.2-2.1%) in the 27th year to 4.0% (95%CI: 1.3-11.3%) in the second year. The pooled seroprevalence in mobile populations each year was significantly lower than among the residents of endemic regions, whilst four papers reported a lower level of infection in the mobile populations than in the local residents out of only 13 papers which included this data. CONCLUSIONS: The re-emergence of S. japonicum in areas which had previously interrupted transmission might be due to other factors, although risk from re-introduction from mobile populations could not be excluded.

Human babesiosis, an emerging tick-borne disease in the People's Republic of China.

Babesiosis is a typical zoonotic, emerging disease caused by a tick-borne intraerythrocytic protozoan of Babesia spp. that also can be transmitted by blood transfusion. Babesiosis imposes an increasing public-health threat. We reviewed and mapped epidemiological studies on Babesia in vectors and/or rodents in the People's Republic of China (P.R. China) and found that B. microti was the predominant species detected in the investigated regions such as Heilongjiang, Zhejiang, Fujian provinces and Taiwan island. We reviewed a series of sporadic human babesiosis cases collected from 1940's to 2013, in Yunnan, Inner Mongolia, Taiwan and Zhejiang and other regions including a main endemic area of malaria on the China-Myanmar border areas in P.R. China. Clinical manifestations of human babesiosis were also reviewed. Human babesiosis may have previously been overlooked in P.R. China due to a lack of medical awareness and the limitation of clinical diagnostic methods.

Control efficacy of annual community-wide treatment against Schistosoma japonicum in China: a meta-analysis.

BACKGROUNDS: Human schistosomiasis is caused by schistosome, with annual loss of over 70 million disability adjusted life years in the world. China is endemic with Schistosoma japonicum and large-scale chemotherapy with praziquantel has become the mainstay of control in China since 1990s. However, the control effects of mass treatment in the field have been uneven. Moreover, mass treatment has come into a wide use in other countries with limited health resources. Therefore, a better understanding of the control effect of mass treatment is in an urgent need. METHODS: We performed a systematic search of the literature to investigate the control efficiency of annual community-wide treatment (ACWT, treatment to an entire community without any preliminary screening) with a single dose of PZQ (40 mg kg(-1) bodyweight) against schistosome in humans in China. Three Chinese literature databases, including China National Knowledge Infrastructure, WanFang and Chinese Scientific Journal Databases, and the PubMed were searched. Pooled prevalence ratios (prevalence after to before treatment) were used to assess effect. Our protocol is available on PROSPERO (No. CRD42013003628). RESULTS: 22 articles were included. Meta-analyses on data from 18 studies on one round of ACWT, 17 studies on two consecutive rounds and 6 studies on three consecutive rounds were performed. The results showed control effects of ACWT plus other measures were statistically significant, with prevalence ratios being 0.38 (0.31, 0.46) for one round, 0.28 (0.22, 0.35) for two rounds and 0.22 (0.10, 0.46) for three rounds. When ACWT was performed alone or with health education only, the values for one and two rounds were 0.389 (0.307, 0.492) and 0.348 (0.300, 0.403), respectively. CONCLUSIONS: The control effect of ACWT alone or with other measures is significant and increases with the number of rounds. Such program is recommended in high endemic areas and the criteria yet merit further assessment.

[Effect of Pulsatilla chinensis (Bunge) Regel saponins on eggs, miracidia and cercariae of Schistosoma japonicum].

OBJECTIVE: To investigate the effect of Pulsatilla chinensis (Bunge) Regel saponins (PRS) against the eggs, miracidia, cercariae of Schistosoma japonicum in vitro and compare its efficacy with praziquantel. METHODS: ICR mice were infected with the cercariae of S. japonicum by the patching abdominal method. The livers of the mice were grinded, screened, and then the eggs of S. japonicum were obtained 42 days post-infection. The miracidia were hatched by using the eggs, and the cercariae were obtained by using the infected Oncomelania snails on the light. The eggs, miracidia and cercariae of S. japonicum were incubated with 0, 1, 2, 3, 4, 5, 6, 7, 8 microg/ml PRS for different time, and praziquantel (PZQ) was used as the control. RESULTS: PRS suppressed the hatching rates of eggs for 24 h slightly superior to that of the control drug PZQ at different concentrations, especially in the 4 microg/ml concentration. After the miracidia were incubated with 1, 2, 3, 4, 5, 6, 7, 8 microg/ml PRS for 30 min, the dead rates of miracidia were 13.47, 26.05, 60.99, 90.84, 100, 100, 100, 100%, respectively. After the cercariae were incubated with 1, 2, 3, 4, 5, 6, 7, 8 microg/ml PRS for 30 min, the dead rates of cercariae were 5.32, 18.81, 44.7, 76.87, 98.28, 100, 100, 100%, respectively. PRS showed time- and dose-dependent mortality effects on the miracidia and cercariae of S. japonicum. CONCLUSION: PRS has the effects against eggs, miracidia, cercariae of S. japonicum in vitro, and it may become a new anti-schistosome agent.

[Effect of Pulsatilla chinensis (Bunge) Regel saponins against juvenile and adult Schistosoma japonicum in vitro].

OBJECTIVE: To explore the effect of Pulsatilla chinensis (Bunge) Regel saponins (PRS) against juvenile and adult Schistosoma japonicum and to compare its efficacy with praziquantel (PZQ) in vitro. METHODS: 3 h, 7 d, 14 d schistosomula and 42 d adult schistosomes were incubated with 0, 1, 5, 10, 20 and 30 microg/ml PRS for 4, 24, 48 and 72 hours, then the states of them were observed. The changes of the surface of S. japonicum incubated with 30 microg/ml PRS and PZQ within 4 hours were observed by a scanning electron microscope. RESULTS: The sensitivity of 3 h, 7 d, 14 d schistosomula and adults of S. japonicum to 1, 5, 10, 20, 30 microg/ml PRS displayed a time and dose dependence. All the worms died in 30 microg/ml PRS after 4 hours. The dead worm body appeared a gray-white color accompanied with their altered morphogenesis and opaque body. The tegumental surface of adults with different degrees of damages was observed by the electron microscope within 4 hours affected by PRS in vitro. CONCLUSIONS: The effects of PRS against S. japonicum in different developmental stages in vitro show that PRS may eventually have a therapeutic potential in the treatment or prevention of S. japonicum infection and is expected to become a new anti-schistosome drug.

[CD4+CD25+high regulatory T cells in patients with autoimmune hepatitis].

OBJECTIVE: To investigate the role of CD4+CD25+high regulatory T cells in the pathogenesis of autoimmune hepatitis. METHODS: CD4+CD25+ high regulatory T cells and CD4+ T cells were measured by using flow cytometry in 16 patients with autoimmune hepatitis, 22 patients with chronic hepatitis B and 20 healthy blood donors. Foxp3 protein was detected by immunohistochemical assay in liver tissues from the patients with autoimmune hepatitis or chronic hepatitis B. RESULTS: The percentage of CD4+CD25+high/CD4+ in patients with autoimmune hepatitis was significantly lower than that in healthy controls and patients with chronic hepatitis B. Meanwhile, the percentage of CD4+CD25+high/CD4+ highly increased in patients with chronic hepatitis B, compared with healthy controls; Foxp3 positive cells were mostly located in the hepatic lobular perisinusoidal spaces and the portal tract, and there was a significant difference in the quantity of Foxp3 positive cells between patients with autoimmune hepatitis and chronic hepatitis B. CONCLUSION: Patients with autoimmune hepatitis harbor a decreased percentage of CD4+CD25+ high regulatory T cells, which may be associated with development of autoimmunity.