| Schistosomiasis is a major infectious disease and a public health concern in many areas in China and other countries. Traditionally, detection of Schistosoma japonicum relied on microscopic detection of Schistosoma eggs in stool specimens which is also considered the golden standard for diagnosing Schistomiasis. However, this method lacks the sensitivity in low prevalence and in post-treatment situations as S. japonicum eggs are round and have fewer conspicuous characteristics than those of S. mansoni. Even when the disease is active, the analyzed samples might not contain eggs due to the random distribution and highly variable shedding of the eggs. In testing Schistosoma an important delay must be considered from the time of parasite exposure because the process of adult worm maturation after cercarial penetration is slow. Furthermore, detection of circulating parasite antigens lacks the sensitivity in the low prevalence areas and in post-treatment cases. Also, detection of anti-Schistosoma antibodies lacks specificity. Recently, several groups have employed more specific and sensitive diagnostic methods, mainly using the polymerase chain reaction (PCR) techniques. However, those PCR protocols require further processing of the amplification products, which is time consuming and prone to false-positive results because of possible cross-contamination. As the currently available clinical diagnostic methods are far from ideal, developing and evaluation of new strategies and tools for the control of Schistosomiasis are recommended by the WHO.In our study, on one hand, we established a highly sensitive assay for early detection of S. japonicum, and provided a new viable way for the detection of Schistosomiasis. On the other hand, we generated a kind of inducible transgenetic mice with specific expression of IL-22on which will help us to develop new vaccine, find an adjuvant to enhance and augment the protection of vaccine against S. japonicum or make a preparation for the further research about the effect and mechanism on the immune evasion of S. japonicumIn this study, we developed a highly sensitive TaqMan real-time PCR assay for the detection of Schistosoma japonicum DNA in mouse feces and serum samples. Compared to conventional PCR, TaqMan real-time PCR assay offers some advantages in minimizing the risk of carrying over contamination in which DNA amplification and detection is in a sealed tube or well. In addition, further down-stream analysis is not required, which shortens the time needed to obtain results by conventional electrophoresis on agarose gel. A major breakthrough was achieved by detection of cell-free S. mansoni DNA in mouse and human plasma using real-time PCR. Subsequently, the data of a clinical trial showed that this detection method had reliable sensitivity and specificity for diagnosing S. mansoni and S. haematobium infection. However, it was mentioned in the Wichmann report that the sensitivity of this assay may vary among Schistosoma species and the target gene has not been formally evaluated for S. japonicum. Our assay was based on the DNA sequence of the S. japonicum18S rRNA gene and was able to detect10fg of S. japonicum genomic DNA, which is100times more sensitive than conventional PCR. We were able to detect the S. japonicum DNA one week post-infection in mouse sera and4 weeks post-infection in feces, which was one week earlier than egg detection by microscopy in feces. This assay was also highly specific for Asian Schistosomes which are causative species of human Schistosomiasis. In single sex male cercariae infected mice, parasite DNA was only detected in the first4weeks post-infection, suggesting that the DNA was derived from decaying worms’corpse whereas the DNA detected in other infected groups was from both decaying worms’corpse and parasite eggs. Therefore we conclude that the established TaqMan real-time PCR assay is a sensitive, specific and convenient method that could be used for the early diagnostic evaluation of S. japonicum infection in humans and for monitoring outbreaks in endemic areas with low prevalence.Developing an effective immune response to an invading pathogen is critical for host defense and survival. IL-22is derived from T cells and preferentially made by Th17T cells. And IL-22participates in host defense at environmental interfaces, including mucosal surfaces of airways (mainly in the trachea and bronchia) and gastrointestinal tract as in the skin where it maintains barrier integrity and regulates wound repair. We hypothesis that IL-22has a role in the process of S. japonicum invading which could help us to find a way to protect from this helminth or enhance our current vaccines. So in our part2study, we have initiated to generate a series of transgenic mice based on Tet-off/on system, and we have got the CC10-rtTA-hGH/TRE-tight-IL-22mice successfully which expressed pretty high concentration of IL-22in the lung, and we also demonstrate here over-expression of IL-22doesn’t resulted in mortality in adult mice as we could inhibit the transgenic expression of IL-22without feeding of doxcycline before they are mature. |
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