Wensen Liu

Paragonimiasis: an important food-borne zoonosis in China

The lung fluke, Paragonimus westermani, is of major socioeconomic importance in Asia. The parasite is transmitted via snails to freshwater crabs or crayfish, then to humans and other mammals, such as cats and dogs, and causes paragonimiasis. This review provides a background on the parasite and its life cycle; summarizes key aspects regarding the pathogenesis, diagnosis and treatment of paragonimiasis; describes the geographic distribution and prevalence of paragonimiasis; and makes some recommendations for future research and the control of this important disease in China.

Isolation and characterisation of a new antimicrobial peptide from the skin of Xenopus laevis

A new antimicrobial peptide (AMP) named PGLa-H has been isolated from the skin of the African clawed frog (Xenopus laevis) using gel filtration and reverse-phase high-performance liquid chromatography (RP-HPLC). Its amino acid sequence was determined as KIAKVALKAL by Edman degradation, with a molecular weight of 1053.727 Da as analysed by matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI-TOF/TOF-MS). No similar AMP was found by BLAST search. Purified PGLa-H demonstrated antimicrobial ability against the reference bacteria Escherichia coli ATCC 25922 [minimum inhibitory concentration (MIC)=23.6 μg/mL], Staphylococcus aureus ATCC 25923 (MIC=8.7 μg/mL) and Bacillus subtilis (MIC=14.4μg/mL) and was active against multidrug-resistant meticillin-resistant S. aureus (MRSA) (MIC=67.8 μg/mL). The antimicrobial mechanism for this new peptide was further investigated by transmission electron microscopy. PGLa-H killed cells by destroying the cell membrane.

Protection in mice immunized with a heterologous prime-boost regime using DNA and recombinant pseudorabies expressing TgSAG1 against Toxoplasma gondii challenge

An effective vaccine of animals can block transmission of Toxoplasma gondii to humans. In this study, mice have been protected against lethal T. gondii challenge by a prime-boost vaccination strategy using DNA vaccine pVAX/TgSAG1 and recombinant pseudorabies virus rPRV/TgSAG1, both expressing the major immunodominant surface antigen of T. gondii (TgSAG1). High levels of splenocyte proliferative responses and significant levels of IFN-gamma resulted, with strong cytotoxic T lymphocyte (CTL) responses in vitro. After lethal challenge, prime-boost vaccinated mice showed an increased survival time (15.4+/-5.0 days) and a 40% survival rate compared with controls who all died within 11 days of challenge. Results of the present study indicated that this novel immunization strategy is useful in enhancing immune protection in mice against lethal T. gondii infection, which would provide foundation for the development of effective vaccines against T. gondii.

Genomic analysis of Newcastle disease virus strain NA-1 isolated from geese in China

Newcastle disease virus (NDV) has been thought to infect only domestic avian species, with waterfowl such as geese either not being infected, even by virulent strains, or developing only in apparent infection. In 1997, a new infectious disease producing high morbidity and mortality among geese broke out in many provinces of China, which was caused by a serotype I avian paramyxovirus (APMV-1)—NDV. To investigate how NDV spreads between chickens and geese, the complete genome of one NDV strain isolated from a goose was cloned and analyzed. The results indicate that there is conservation in NDV structural genetic evolution but that there are also considerable differences between goose and chicken NDV strains. Separate patterns of NDV evolution exist among wild bird species. Meanwhile, there is evidence indicating that the goose NDV may have evolved from chicken NDV strain Herts/33. In addition, the possibility was investigated that this new strain of NDV may bind to different sialic acid receptor binding sites than the normal NDV strains that have been investigated so far. This might provide clues to the evolution of the goose NDV.

IL-18 enhances protective effect in mice immunized with a Schistosoma japonicum FABP DNA vaccine.

Two recombinant plasmids, pVAX/SjFABP and pVAX/mIL-18 containing Schistosoma japonicum 14 kDa fatty acid binding protein (SjFABP) and murine IL-18, were constructed and evaluated for their ability to induce immune responses and to protect against S. japonicum challenge in mice. Mice were intramuscularly immunized twice at three-weekly intervals, and challenged with S. japonicum cercariae at 4 weeks after the last vaccination. All animals vaccinated with pVAX/SjFABP alone or plus pVAX/mIL-18 developed specific anti-SWAP ELISA antibody and T lymphocyte proliferation. Co-injection of pVAX/mIL-18 significantly increased the production of IFN-gamma and IL-2 compared with pVAX/SjFABP alone, indicating that IL-18 enhances the Th1-dominant immune response. The challenge experiment showed that co-injection of plasmid encoding IL-18 significantly enhances protective effect against S. japonicum infection, as demonstrated by worm reduction rates and the hepatic egg reduction rates 45 days post-challenge. These results indicated that IL-18 may become a novel vaccine adjuvant for development of vaccines against schistosomiasis.

Morphological changes of ricin toxin-induced apoptosis in human cervical cancer cells

The morphological changes of ricin-induced apoptosis in a human cervical cancer cell line were studied. To shed light on the mechanism of action of ricin toxin (RT) at the cellular level, we examined cell growth, apoptosis, changes of mitochondrial membrane potential (MMP) and cytochrome C translocation in HeLa cells by exposing these cells to RT for indicated times. The effect of RT on cell proliferation was measured by 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS), inner salt; MTS assay and apoptosis were measured using flow cytometry, fluorescence microscopy and electron microscopy. Changes in MMP were monitored using flow cytometry. Western blot analysis was used to evaluate the release of mitochondrial cytochrome C. RT noticeably inhibited the proliferation of HeLa cells, and the half maximal inhibitory concentration dose was about 100 ng/ml. HeLa cells treated with RT showed typical characteristics of apoptosis rather than necrosis, including phosphatidylserine exposed from the inner to the outer leaflet of the plasma membrane, abnormal cell morphology, chromatin condensation and nuclear fragmentation. In contrast, during the process of cellular apoptosis, the messenger RNA (mRNA) and protein expression of cytochrome C in treated and untreated Hela cells were not significantly changed (data not shown). However, when cells were treated with RT, the massive translocation of cytochrome C to the nucleus was evident. Our results indicate that RT-induced HeLa cell apoptosis, especially for cytochrome C translocation, may play an important role in apoptosis induced by RT.

A laboratory-attenuated vesicular stomatitis virus induces apoptosis and alters the cellular microRNA expression profile in BHK cells.

In the present study, we characterized the pathways by which a laboratory-attenuated vesicular stomatitis virus (La-VSV) induces apoptosis in BHK cells. It was found that La-VSV induced a loss of mitochondrial membrane potential (ΔΨm) and activated caspase-9 and -3, but not caspase-8, indicating that the induction of apoptosis by La-VSV may involve an intrinsic apoptotic pathway. Although aberrant expression of microRNAs (miRNAs) has been linked to viral infection, little is known about changes in the cellular miRNA expression profile following VSV infection. Here, we attempted to identify miRNA expression profiles in VSV-infected BHK cells using miRNA microarray. Data analysis revealed that 28 miRNAs consistently responded to VSV-infection, 12 of which were down-regulated and 16 of which were up-regulated. miR-146a of these miRNAs has been found to be up-regulated in LPS-stimulated monocytes and VSV-infected macrophages, suggesting that VSV-induced miR-146a expression occurs not only in immune cells but also in other host cells. We further found that miR-706 inhibited VSV-induced apoptosis by decreasing caspase-3 and -9 activation, suggesting that induction of miR-706 expression may be a novel strategy for survival of VSV, allowing it to escape the apoptosis response of the host. In summary, our results indicate that miRNAs might play important roles in VSV infection and that their aberrant expression could be involved in VSV pathogenesis.

Molecular Detection and Genetic Diversity of Leishmania donovani in Naturally Infected Phlebotomus chinensi from Southwestern China

Zoonotic visceral leishmaniasis is an important vector-borne infectious disease in western China. In this study, an epidemiological study was carried out on the vector of zoonotic visceral leishmaniasis in rural areas from Sichuan Province, southwestern China. In the 1263 phlebotomine sandflies captured, 859 (68.01%) were females and 404 (31.99%) males, belonging to Phlebotomus chinensis (83.37%), Sergentomyia koloshanensis (6.57%), Sergentomyia squamirostris (4.04%), and Sergentomyia barraudi (6.02%), respectively. The average prevalence of Leishmania parasites in P. chinensis females was 1.98%, which was detected by real-time quantitative PCR. Phylogenetic analysis based on ITS2-rDNA revealed that Leishmania parasites detected in sandflies belonged to the L. donovani group and formed a novel haplotype. This was the first report on molecular detection of L. donovani in naturally infected P. chinensi from China.

Detection of ricin intoxication in mice using serum peptide profiling by MALDI-TOF/MS.

Ricin toxin has been regarded as one of the most potent poisons in the plant kingdom, and there is no effective therapeutic countermeasure or licensed vaccine against it. Consequently, early detection of ricin intoxication is necessary. In this study, we took mice as test subjects, and used the technique of Matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF/MS) and ClinProt™ microparticle beads to set up an effective detection model with an accuracy of almost 100%. Eighty-two peaks in the mass range 1000–10,000 m/z were detected by ClinProTools software, and five different peaks with m/z of 4982.49, 1333.25, 1537.86, 4285.05 and 2738.88 had the greatest contribution to the accuracy and sensitivity of this model. They may therefore provide biomarkers for ricin intoxication.

Analysis of intestinal injuries induced by ricin in vitro using SPR technology and MS identification.

The present study found that ricin toxicity did not only manifest itself as inhibition of protein synthesis, but also induced apoptosis of immune cells and played an extremely significant role in intestinal injury. In this report, we describe a novel method to estimate binding events occurring on intestinal brush border membranes (BBM) based on SPR technology in an attempt to mimic the real intestinal surface capable of interacting physically and/or actively with certain biological molecules. Combined with HPCE-ESI-MS indentification, we obtained 28 kinds of proteins in BBM that interacted with ricin.