Protective immunity induced by a DNA vaccine cocktail expressing TgSAG1, TgROP2, and the genetic adjuvant HBsAg against Toxoplasma gondii infection.

Toxoplasma gondii is an intracellular obligate parasitic protozoon that can infect all warm-blooded animals, causing zoonotic toxoplasmosis. So far, there is no commercial toxoplasmosis vaccine for human use. In the present study, we constructed a DNA vaccine cocktail which includes the surface protein (SAG1) and the rhoptry protein ROP2 denoted as pEGFP-N1-SAG1-ROP2.

[Preparation and purification of polyclonal antibody against microneme protein 16 and its application in subcellular localization].

Objective: To prepare and purify the rabbit anti-TgMIC16 polyclonal antibody, so as to apply it in subcellular localization.

Methods: New Zealand white rabbits were immunized with purified recombinant TgMIC16 mixing with the same volume of Freund's adjuvant for three times, respectively. The rabbit serum was collected on the 14th day after the last immunization.

Protective immune response against Toxoplasma gondii elicited by a novel yeast-based vaccine with microneme protein 16.

Toxoplasma gondii is an obligate intracellular protozoan that can invade all eukaryotic cells and infect all warm-blood animals, causing the important zoonosis toxoplasmosis. Invasion of host cells is the key step necessary for T. gondii to complete its life cycle and microneme proteins play an important role in attachment and invasion of host cells.

[Construction and expression of multi-gene recombinant plasmid pEG-FP-N1-HBsAg-ROP2].

Objective: To construct pEGFP-N1-HBsAg-ROP2 recombinant expression plasmid and transfect HEK293T cells for expression, and pay a way for nucleic acid vaccine development.

Methods: According to the HBsAg gene sequence and pcDNA3-p30-ROP2 recombinant plasmid restriction sites, the HBsAg gene was amplified by PCR. The HBsAg gene was cloned into the pcDNA3-p30-ROP2 and instead of p30 gene.

Expression of codon-optimized TgMIC16 in three strains.

In a previous study, we found that rabbit anti- serum was capable of recognizing truncated microneme protein 16 (TgMIC16), indicating that TgMIC16 is an essential antigenic protein. However, the broad application of this recombinant protein is limited by its low expression level. In this study, we performed codon optimization of by changing the codon-adaptation index from 0.

[Allele genetypes and homology analysis of MSP-1 and CSP gene of in Shandong Province].

Objective: To analyze the genotypes and homology of MSP-1 and CSP gene of in Shandong Province, so as to provide the evidence for case traceability.

Methods: A total of 12 blood samples were collected from -infected cases in Shandong Province in 2011. Parasite genomic DNA was extracted.

[Detection of Genetic Mutations Associated with Drug Resistance in Imported Plasmodium falciparum].

Objective: To investigate the mutation of genes associated with drug resistance (Pfcrt, Pfmdr1, Pfdhfr and K13） in imported Plasmodium falciparum in Shandong Province.

Methods: Blood was collected from 94 falciparum malaria cases who returned from Africa in 2014. Genomic DNA for P.

Characteristics of Imported Malaria and Species of Plasmodium Involved in Shandong Province, China (2012-2014).

Malaria remains a serious public health problem in Shandong Province, China; therefore, it is important to explore the characteristics of the current malaria prevalence situation in the province. In this study, data of malaria cases reported in Shandong during 2012-2014 were analyzed, and Plasmodium species were confirmed by smear microscopy and nested-PCR. A total of 374 malaria cases were reported, 80.

[Prokaryotic Expression, Purification and Immunological Characterization of Micronemal Protein 16 of Toxoplama gondii].

Objective: To prokaryotically express three gene fragments of micronemal protein 16 （TgMIC16） of Toxoplasma gondii, and analyze the immunoreactivity of the three recombinant protein products.

Methods: Primers were designed for three fragments of TgMIC16 gene which encode proteins within the functional domain. Reverse-transcription PCR was used to generate cDNA from RNA, and the three fragments were amplified on the cDNA by PCR using the designed primers.

[Extraction Method of Malaria Parasite DNA from Preserved Positive Blood Smears].

Objective: To develop a method for DNA extraction from malaria parasites on preserved blood smears, to provide basis for research on malaria genetic traceability.

Methods: The improved DNA extraction kit (QIAamp DNA Mini Kit) was used to extract plasmodium DNA from 41 giemsa-stained blood smears, and the extraction was compared with that using the Chelex-100 and Na(2)HPO(4) methods. Nested PCR was used to amplify small subunit ribosomal RNA to identify Plasmodium parasite.

[Cloning and Bioinformatics Analysis of Toxoplasma gondii ROP21 Gene].

The full-length gene sequence of Toxoplasma gondii ROP21 (TgROP21) gene was amplified with PCR. The signaling peptide and transmembrane domain of TgROP21 protein were predicted by SignaIP and TMHMM online predictive sites, and the hydrophilicity and antigenic index of this protein were ananlyzed with DNAStar software. Meanwhile, the functional domains and tertiary structure were modeled by combined use of ExPASY and PRODATA online sites.

Establishment of lymphatic filarial specific IgG4 indirect ELISA detection method.

Objective: To establish the lymphatic filarial specific IgG4 indirect ELISA detection method and develop the kits.

Methods: ELISA and the developed specific IgG4 reagent was used to explore the best way for detecting filarial specific IgG4. Combined with the production process of commercialized enzyme immunoassay kit to develop economical lymphatic filarial specific IgG4 test kit, and to explore the value of the kit in the laboratory.

Construction and identification of Complex DNA vaccine of hepatitis B and Toxoplasma gondii.

Objective: To construct and identify multi-gene recombinant expression vector pcDNA3-HBsAg-p30-ROP2.

Method: Primers were designed according to the gene sequences of restriction enzyme cutting site of recombinant pcDNA3-p30-ROP2 and hepatitis B surface antigen (HBsAg). The target fragment of HBsAg was amplified and cloned to expression vector pcDNA3-p30-ROP2 by restriction enzyme digestion and ligation.

[Cloning, expression, purification and identification of Toxoplasma gondii SAG2 gene in Escherichia coli].

Objective: To construct a recombinant plasmid containing surface antigen 2(SAG2) gene of Toxoplasma gondii and express it in Escherichia coli.

Methods: The truncated SAG2 gene was amplified from the genomic DNA of T. gondii RH strain and cloned into plasmid pGEX-4T.

[Application of Nested PCR in the Diagnosis of Imported Plasmodium Ovale Infection].

Objective: To identity Plasmodium ovale infection by 18S rRNA gene nested PCR.

Methods: Whole blood and filter paper blood samples of malaria patients in Shandong Province were collected during 2012-2013. The parasites were observed under a microscope with Giemsa staining.

[Survey of intestinal parasitic infections and related knowledge and behavior of residents in Jiaodong area of Shandong Province].

Objective: To understand the status of intestinal parasitic infections and the related knowledge and behavior in residents of Jiaodong area of Shandong Province, so as to provide the evidence for making an appropriate preventive and control strategy.

Methods: A total of 18 villages from 6 counties in Jiaodong area were selected as investigation sites according to the stratified sampling method. The feces samples of the permanent residents aged above 3 years were collected and examined by Kato-Katz technique to find the intestinal parasite eggs, and the children under 12 years old were examined by the method of cellophane anal swab to detect the Enterobius vennrmicularis eggs.

[Research progress on polymorphism in the merozoite surface protein-3alpha gene of Plasmodium vivax].

The merozoite surface protein-3alpha of Plasmodium vivax (PvMSP-3alpha) is a surface protein that is expressed during the asexual blood stages. With a high polymorphism, PvMSP-3alpha gene has been used as a suitable epidemiology and genotype marker. Moreover, it might be an important malaria vaccine candidate.

[Epidemiological analysis of malaria prevalence in Shandong Province in 2012].

Objective: To understand the characteristics of malaria prevalence and epidemic in Shandong Province in 2012 so as to provide the evidence for improving the work of the elimination of malaria.

Methods: The epidemiological data of malaria cases collected from the Disease Surveillance Information Reporting System of Chinese Center for Disease Control and Prevention were analyzed with the descriptive epidemiological method for epidemiological characteristics of malaria.

Results: A total of 93 malaria cases were reported in Shandong Province in 2012 with the incidence of 0.

XPD codon 312 and 751 polymorphisms, and AFB1 exposure, and hepatocellular carcinoma risk.

Background: Genetic polymorphisms in DNA repair genes may influence individual variation in DNA repair capacity, which may be associated with risk of hepatocellular carcinoma (HCC) related to the exposure of aflatoxin B1 (AFB1). In this study, we have focused on the polymorphisms of xeroderma pigmentosum complementation group D (XPD) codon 312 and 751 (namely Asp312Asn and Lys751Gln), involved in nucleotide excision repair.

Methods: We conducted a case-control study including 618 HCC cases and 712 controls to evaluate the associations between these two polymorphisms and HCC risk for Guangxi population by means of TaqMan-PCR and PCR-RFLP analysis.

[Cloning and identification of ROP2 gene of Toxoplasma gondii].

Objective: To amplify ROP2 from the genomic DNA of Toxoplasma gondii RH strain and construct eukaryotic expression plasmid pc-DNA3-ROP2.

Methods: Tachyzoites of T. gondii RH strain were collected and depurated to obtain genome.