ABSTRACT
Rotavirus(RV) is currently the leading cause of severe diarrhea in infants and young children worldwide, and RV has been found to recognize and bind to histo-blood group antigens(HBGAs) through its structural protein VP8, and the FUT2 gene determines the expression of HBGAs in epithelial tissues and secretion in body fluids.Individuals with loss of functional enzyme activity due to mutations in the FUT2 gene, called non-secretors, are unable to express and secrete HBGAs in the mucosa and body fluids, and non-secretors have been found to be resistant to diarrhea caused by RV.Studies have shown that microbial composition is genetically regulated by the host, and hundreds of genetic loci are involved in regulating the composition of human gut microbes, including FUT2.Sterile animal models reduce the rate of RV infection, suggesting that intestinal bacteria are associated with the process of RV infection.These studies reveal that secretory status directly influences individual susceptibility to RV, and its effect on gut microbial composition indirectly modulates human susceptibility to RV.This article reviews the correlation between FUT2 and gut microbial composition with RV susceptibility, with the aim of opening new avenues for personalized prevention and treatment of RV infection.
ABSTRACT
【Objective】 To establish a stable human megakaryocytic cell line with low expression of Ley antigen to further study the role of Ley on activation of platelets. 【Methods】 The expression level of the Ley antigen in a human megakaryocytic cell line, DAMI, was determined using Western Blot and flow cytometry. The expression level of genes that encode fucosyltransferase (FUTs), which was involved in the biosynthesis of Ley antigen, was also determined to identify the candidate genes to be knocked out. The candidate FUT gene was knocked out via a CRISPR/Cas 9 gene knockout system and cells with low Ley antigen expression were sorted by flow cytometry. The sorted cell line was cultured and characterized. 【Results】 The Ley was expressed intensively on DAMI cell. FUT1 and FUT4 mRNA was expressed relatively higher, both may be key enzymes for the biosynthesis of the Ley antigen. In the DAMI cell line with the knockout of FUT1 gene, the expression of the Ley adntigen was remarkedly reduced, while cell proliferation was not affected compared to the wildtype control cells. 【Conclusions】 Since various FUTs contributes to the biosynthesis of the Ley antigen, the knockout of the primary one of them cannot totally block its biosynthesis, but only reduce its expression. In this study, a stable FUT gene knockout human megakaryocyticcell line is established using CRISPR/Cas 9 technology, which provides basis for the study of the impact of the Ley antigen on platelet functions.
ABSTRACT
Norovirus, as one of the main pathogens causing non-bacterial gastroenteritis, can cause serious public health problems and economic losses around the world. In recent years, the outbreaks caused by the virus in China are on the rise. Human Norovirus (HuNoV) can hardly be cultivated in-vitro. The nucleic acid detection method (such as RT-qPCR) has the highest sensitivity and specificity, but it was not established that the correlation between the detected viral genome and viral infectivity, which leads to inaccurate judgment of safety risks. Here, the in-vitro and in-vivo culture models, viral genome integrity and capsid protein integrity were cut into three aspects. The research progress and characteristics of infectious Norovirus identification technology in recent years were reviewed and discussed, and the future development trend of this technology was prospected. The aim is to further improve the accuracy of Norovirus quantitative detection and provide a theoretical basis for its application in the field of food safety testing.
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Objective@#To explore the receptor binding specificity of VP8* protein of porcine P[19] rotaviruses (RVs) with oligosaccharides.@*Methods@#The porcine P[19] VP8* protein was expressed and purified. The receptor binding specificity of P[19] VP8* was analyzed by oligosaccharide binding and saliva binding assay.@*Results@#The P[19] VP8* protein showed significant binding to mucin cores, especially mucin core 2.@*Conclusions@#Mucin core 2 may be a potential receptor for the porcine P[19] RV, which provides certain basis for the study of virus infection mechanism and RV surveillance.
ABSTRACT
Objective To evaluate the immune effects of virus-like particles ( VLPs) assembled from the capsid protein VP1 of a recombinant norovirus ( NoV) GⅡ. 17 genotype. Methods The recombi-nant NoV GⅡ. 17 VP1 VLPs were purified, and then tested by SDS-PAGE and Western blot to analyze the purity. The size, morphology and diameter distribution of the recombinant VLPs were detected by transmis-sion electron microscopy ( TEM) and dynamic light scattering ( DLS) analyzer. The recombinant VP1 VLPs adsorbed by aluminium adjuvant were used to immunize BALB/c mice. Serum samples were collected after immunization. Specific antibody level and neutralizing antibody activity were evaluated with enzyme linked immunosorbent assay ( ELISA) and histo-blood group antigen ( HBGA)-VLP blocking test. Cross-reactivity of serum samples with GⅠ. 1 and GⅡ. 4 VP1 VLPs were detected. Moreover, cross-protection against GⅠ. 1 and GⅡ. 4 VP1 VLPs was analyzed. Results The purity of the recombinant NoV GⅡ. 17 VP1 VLP was greater than 90％ and specific bands were detected by Western blot. TEM images and DLS experiments showed that VLPs were 30-50 nm in size with good morphology and uniformity, indicating that the recombi-nant VLPs were similar to the wildtype virus. High titers of specific antibodies were detected in serum sam-ples of the immunized mice. A certain degree of cross-reactions between serum samples and VP1 VLPs of NoV GⅠ. 1 and GⅡ. 4 were observed, but no cross-protection was detected. Conclusion The recombinant GⅡ. 17 VP1 VLPs in combination with aluminum adjuvant can induce higher titers of HBGA blocking anti-bodies in mice, suggesting that it could be used as a candidate target antigen for norovirus vaccine.
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Objective To investigate the physicochemical properties and immunogenicity of virus like particles(VLPs) in two different conformations assembled from the essential capsid protein VP1 of GⅡ.4 norovirus(NoV) in Hansenula polymorpha. Methods NoV GⅡ.4 VLPs in two different conforma-tions were prepared from high-density fermentation of recombinant engineered strains and VLPs purification. Physicochemical properties of the two forms of VLPs were identified by Western blot,size-exclusion high per-formance liquid chromatography (SEC-HPLC), dynamic light scattering(DLS) and transmission electron microscopy. Serum VLPs binding activities and blocking activities against VLPs binding to histo-blood group antigen(HBGA-VLPs) were evaluated after immunization of BALB/c mice with the two forms of VLPs. Re-sults VLPs of two different diameters with high homogeneity were obtained after purification. DLS results showed that particle sizes of two VLPs were 53.98 nm and 45.18 nm,respectively. The two VLPs were sim-ilar in binding abilities to HBGA receptors. Serum VLPs binding activities and blocking activities against HBGA-VLPs were found higher in NoV-VLP-L than NoV-VLP-S,but the difference was not statistically sig-nificant (P>0.05). Conclusion VLPs in two different conformations were obtained by expressing NoV GⅡ.4 VP1 proteins in Hansenula polymorpha. Though they were similar in physicochemical properties and immunogenicity,the NoV-VLP-L might be potential antigen candidates for the development of recombinant human norovirus vaccine.
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Objective To evaluate the immune effects of virus-like particles ( VLPs) of VP1 pro-teins derived from norovirus GⅠ. 1 and GⅡ. 4 genotypes expressed in Hansenula polymorpha expression sys-tem. Methods SDS-PAGE and Western blot assay were performed to detect the purity of GⅠ. 1 and GⅡ. 4 VP1 proteins after purification. Morphologies of the recombinant VLPs were observed under transmission electron microscopy ( TEM) . Sizes and distributions of the VLPs were analyzed by dynamic light scattering analyzer. BT50(50% of blocking titer) was detected by HBGA (histo-blood group antigen) blocking assay in BALB/c mice immunized with different regimens. Results SDS-PAGE analysis of the purified recombinant GⅠ. 1 and GⅡ. 4 VP1 proteins showed that their purity were greater than 90%. Western blot assay con-firmed the specific bands of VLPs. TEM images showed that the sizes of purified GⅠ. 1 and GⅡ. 4 VP1 VLPs were at a mean diameter of 30-50 nm with clear border and high homogeneity, which was similar to that of wild virus. BT50 significantly increased in the groups, in which Al( OH) 3 was used as adjuvant. Con-clusion Animal studies have shown that administration of GⅠ. 1 and GⅡ. 4 VP1 VLPs in the presence of Al( OH) 3 induces detectable HBGA-blocking antibody, indicating that GⅠ. 1 and GⅡ. 4 VP1 VLPs are promising candidates for norovirus vaccine.
ABSTRACT
Rotavirus (RV) is one of the major pathogens responsible for acute viral gastro-enteritis in children. The infec-tion of RV is dependent upon the recognition of the host cell speciifc receptors and attachment. Thus, receptors are the important factors of infection. Recent studies have suggested that a genetic factor might play a role in the susceptibility of hosts to RV infec-tion. Histo-blood group antigens have recently been discovered as receptors binding to RV, which are important for the study of evolution. Thus it will be also crucial for the pathogenesis and epidemiology and prevention and treatment for RV. In this article, we will review the correlation of the RV infection and histo-blood group antigens and further discuss the development of optimal vaccine.