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ABSTRACT: Neospora caninum is an intracellular protozoan that infects domestic and wild canids as well as many warm-blooded animals as shown by the isolation of viable parasites. The effectiveness of diagnostic tests for detecting specific antibodies against N. caninum is hampered by potential cross-reaction with other Coccidia. So, there is currently an urgent need for a sensitive and specific diagnostic assay for detecting N. caninum in animals. The N. caninum 40-kD surface antigen (p40), similar to NcSAG1 and NcSRS2, was shown to belong to surface antigen super family and thus represents an excellent marker for the diagnosis of neosporosis. In order to test the hypothesis, recombinant Ncp40 (rNcp40) was expressed in Escherichia coli, and an indirect ELISA test was developed using recombinant NCp40 antigen for N. caninum serodiagnosis. The antigen used in this study did not have cross-reactivity with anti-Toxoplasma gondii serum. Anti-p40 antibodies were detected by ELISA in the sera of Yellow cattle and were compared with (IFAT). Optimal sensitivity and specificity (98.2 and 98.6 %) were identified by IFAT. Additionally, 37 positive sera of T. gondii were detected and there was no significant difference with the negative serum of N. caninum. The rNcp40 ELISA developed here provides a specific and sensitive assay for detecting neosporosis in cattle.
Parasitology Research 02/2013; · 2.15 Impact Factor
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ABSTRACT: To obtain novel antigen genes for use as an anti-tumor vaccine, a Trichinella spiralis cDNA expression library was constructed from muscle larvae RNA and screened with sera from Balb/C mice injected with Sp2/0 myeloma cells. One positive clone was obtained after three rounds of immunoscreening of the cDNA expression library and was subsequently excised in vivo using the ExAssist helper phage with SOLR strain. A full-length gene was amplified using 5'-RACE technology and analyzed by BLAST, Protein Analysis System of ELM, and DNAStar Software. The sequencing results showed that the fragment was 569bp in length and contained an open reading frame. It was predicted that the full-length gene encoded 136 amino acids. This gene, TS2, contained four putative N-Arg dibasic convertase (nardilysine) cleavage sites, one peptide C-terminal amidation site, and one glycosaminoglycan attachment site. Six antibody epitopes were predicted by bioinformatic analysis.
Veterinary Parasitology 02/2013; · 2.58 Impact Factor
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ABSTRACT: To study the molecular mechanism of suppressed growth caused by Trichinella spiralis, an SP2/0 myeloma model was established using parasite-infected Balb/c mice. Suppression subtractive hybridization (SSH) was then utilized to identify differentially expressed genes between tumor cells from the infected and non-infected mice. On the 11th day after infection, 2×106 SP2/0 myeloma cells were subcutaneously inoculated into 6-8 week old female Balb/c mice in both the experimental and control groups. Twenty-eight days after tumor cell inoculation, the mice were euthanized and the sizes and weights of the tumors were measured. Messenger RNA was isolated and used to perform SSH. Putative differentially expressed genes were identified, sequenced and analyzed by BLASTn. Among the sequences detected which ranged in size between 180 and 850bp, genes encoding RpL41, NKTR, Rbbp4 and ANXA2 were enriched and considered possible proteins involved in tumor growth inhibition.
Veterinary Parasitology 02/2013; · 2.58 Impact Factor
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Shijie Li,
Wenzhi Ren,
Wei Li,
Na Zhao,
Guangpeng Ma,
Pengtao Gong,
Qianzhong Han,
Weizhi Li,
Ju Yang, Jianhua Li,
Xichen Zhang
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ABSTRACT: The IPSE/alpha-1 gene (IL-4-inducing principle of Schistosoma mansoni eggs) is a major secreted glycoprotein of S. mansoni eggs that has a potent IL-4-inducing effect. To test the hypothesis that the immune evasion mechanism can be used to overcome the xenograft immune response, the IPSE/alpha-1 gene was transferred into pig fibroblasts, and the transgenic cells were transplanted into KM mice by subcutaneously injecting 10(5)cells per mouse. Cytokine levels were measured to examine the immune response polarization by real-time PCR and ELISA. Mice injected with pig fibroblasts containing a pIRES2-EGFP expression vector were used as a control group. In this group, both cellular and humoral immune responses were activated to reject the grafts alongside increases in all measured cytokine levels. In contrast, the experimental group injected with cells constitutively expressing the IPSE/alpha-1 gene demonstrated a significant decrease in Th1 response cytokines and a significant increase in Th2 response cytokines compared with the control group. These results imply that constitutive IPSE/alpha-1 expression can shift the Th1/Th2 balance of xenograft rejections toward the Th2 response while suppressing the Th1 response. In conclusion, IPSE/alpha-1 could influence the polarization of immune responses during xenograft rejection and suppress the Th1 response. Therefore, this parasitic immune evasion mechanism could be helpful in overcoming xenograft rejection.
Veterinary Immunology and Immunopathology 01/2013; · 2.08 Impact Factor
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Pengtao Gong,
Xiangsheng Huang,
Qinlei Yu,
Yunna Li,
Jingui Huang, Jianhua Li,
Ju Yang,
He Li,
Guocai Zhang,
Wenzhi Ren,
Xichen Zhang
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ABSTRACT: Toxoplasmosis is a world-wide zoonosis that causes significant public health and veterinary problems. The study of vaccines remains the most promising method for the future prevention and control of toxoplasmosis. Recombinant Toxoplasma gondii cyclophilin has been shown to have potent PPIase and IL-12-inducing activities, thus promoting the stabilisation of T. gondii's life cycle and maintaining the survival of its host during evolution. In the present study, the T. gondii cyclophilin gene was used to construct a DNA vaccine (pVAX1-TgCyP). The immune response and protective efficacy of the vaccine against T. gondii infection in BALB/c mice were evaluated. All BALB/c mice that were vaccinated with pVAX1-TgCyP developed a high response with TgCyP-specific antibodies, and significant splenocyte proliferation (P<0.05) compared with pVAX1 vector and PBS groups. pVAX1-TgCyP also induced a significant Th1 type immune response, indicated by the higher production of IL-2 and IFN-γ (P<0.05). The survival rate of BALB/c mice increased significantly after vaccination with pVAX1-TgCyP (37.5%) (P<0.05). These results indicate that TgCyP is a highly efficacious vaccine candidate that can generate protective immunity against T. gondii infection in BALB/c mice. © 2012 Blackwell Publishing Ltd.
Parasite Immunology 12/2012; · 2.60 Impact Factor
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Qiang Lv,
Shenyang Xing,
Zhenxiao Li, Jianhua Li,
Pengtao Gong,
Xiaofang Xu,
LE Chang,
Xiaoxia Jin,
Feng Gao,
Wei Li,
Guocai Zhang,
Ju Yang,
Xichen Zhang
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ABSTRACT: IDH2 encodes a mitochondrial metabolic enzyme that converts isocitrate to α-ketoglutarate (α-KG) by reducing nicotinamide adenine dinucleotide phosphate (NADP(+)) to NADPH and participates in the citric acid cycle for energy production. Notably, this gene has been shown to be critical for cell proliferation. The abnormal expression of IDH2 has been reported in several types of cancer, and mutations in IDH2 have been identified in gliomas and acute myelogenous leukemia. The overexpression of IDH2 has been reported in endometrial, prostate and testicular cancer as well as in Kashin-Beck disease. In this study, we observed that IDH2 expression was significantly downregulated in early phase but was upregulated in advanced phase colon carcinoma compared to peritumoral tissues. In addition, we demonstrated that the growth of a colon carcinoma cell line was inhibited by IDH2-siRNA and increased following transfection with an IDH2-overexpressing plasmid. These results indicate that IDH2 may play a unique role in the development of colon carcinoma.
Experimental and therapeutic medicine 11/2012; 4(5):801-806.
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ABSTRACT: Rhomboid protein in Apicomplexa was associated with the process of host cell invasion. To evaluate the potential of the protein in eliciting protective immunity against challenge, a DNA vaccine pVAX1-Rho encoding Eimeria tenella rhomboid was constructed. Recombinant protein was expressed in Hela cells and verified by indirect immunofluorescence and western blotting analysis. In vivo experiments, 1-week-old chickens were randomly divided into three groups. Experimental group of chickens were immunized with DNA vaccines while control group of chickens were injected with pVAX1 plasmid alone or sterile water. Two weeks following the booster dose, all chickens were inoculated orally with 5 × 10(4) sporulated oocysts of E. tenella. The host immunity and protective efficacy of this vaccine against E. tenella challenge in broilers were evaluated. Results showed that specific antibody, the levels of interleukin-2 (IL-2), interferon-γ (IFN-γ), and the percentages of CD4(+) and CD8(+) T lymphocyte cells were significantly increased in the pVAX1-Rho group. Challenge experiments demonstrated that pVAX1-Rho vaccination could reduce oocyst excretion, decrease cecal lesion, increase bodyweight gains and provide chickens with oocysts decrease ratio around 75.8 %. These results suggest that the pVAX1-Rho was able to induce humoral and cellular responses and generate protective immunity against E. tenella infection.
Parasitology Research 10/2012; · 2.15 Impact Factor
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ABSTRACT: Cystoisospora spp. oocysts isolated from dog feces in Changchun, China were morphologically similar to those of Cystoisospora ohioensis and Cystoisospora sp. 1-MM recently isolated from dogs in Japanese. Sequencing results of the 18S subunit RNA gene from isolates in the present study were compared to other Cystoisospora spp. and the results suggested that Cystoisospora spp. from dogs in Changchun was homologous to C. ohioensis and Cystoisospora sp. 1-MM. Phylogenetic analysis of the 18S rRNA sequences showed that the Cystoisospora sp. ChangChun 1 and Cystoisospora sp. ChangChun 2 were nested in a clade with other Cystoisospora spp., including C. ohioensis, Cystoisospora belli, Cystoisospora suis, Isospora sp. Harbin/01/08 and C. orlovi,. Cystoisospora sp. ChangChun 2 was confirmed as C. ohioensis, and the other isolate was in a separate clade but the genetic relationship was relatively close to C. suis after analysis of the ITS-1gene.
Veterinary Parasitology 06/2012; 190(1-2):254-8. · 2.58 Impact Factor
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ABSTRACT: Toxoplasmosis is one of the most world-wide spread zoonosis representing a very serious clinical and veterinary problem. There is still need for vaccines for toxoplasmosis. In the present study, we evaluated the protective efficacy of a recombinant actin depolymerizing factor (ADF) subunit vaccine against Toxoplasma gondii infection in BALB/c mice. The recombinant T. gondii ADF protein (rADF) was expressed in Escherichia coli and used as antigens for BALB/c mice immunization. The results indicated that specific antibody and the increased percentage of CD4(+) T lymphocyte were found in vaccinated BALB/c mice with rADF, when compared with adjuvant or PBS groups. After challenged with T. gondii (RH strain) tachyzoites, the survival time of the mice in rADF group was longer than those in the control group. The numbers of brain cysts of the mice in rADF group reduced significantly when compared with those in control groups (P<0.05), and the rate of reduction could reach to around 30%. These results suggest that rADF can generate protective immunity against T. gondii infection in BALB/c mice.
Experimental Parasitology 03/2012; 130(3):218-22. · 2.12 Impact Factor
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ABSTRACT: Here we have developed methods to transiently and stably transfect the human pathogenic protist Trichomonas vaginalis. The viral RNA-based transfection vector pTVV-EGFP/NEO was constructed by using enhanced green fluorescent protein gene (EGFP) and neomycin resistance gene (NEO) in tandem to replace the whole gene encoding region of T. vaginalis virus (TVV). The in vitro transcripts of linearized pTVV-EGFP/NEO were electroporated into trophozoites and the transfectants transiently expressed EGFP after 16 h postincubation. Stable expression of EGFP was persistently detected by fluorescence microscopy and by RT-PCR in transfected trophozoites under G418 selection. Our study provides a novel and valuable approach for genetic study of T. vaginalis.
Parasitology Research 08/2011; 110(3):1305-10. · 2.15 Impact Factor
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ABSTRACT: Infection with the intracellular protozoan parasite Toxoplasma gondii causes serious public health problems and is of great economic importance worldwide. The rhomboid proteins which are responsible for adhesion and invasion of host cells have been suggested as vaccine candidates against toxoplasmosis. A DNA vaccine (pVAX-ROM1) encoding T. gondii rhomboid protein 1 (TgROM1) gene was constructed and the immune response and protective efficacy of this vaccine against lethal challenge in BALB/c mice were evaluated. The results indicated that specific antibody and lymphocyte proliferative responses were elicited in mice receiving pVAX-ROM1. The production levels of IFN-γ, IL-2, IL-4, and IL-10, as well as the percentage of CD4(+) cells in mice vaccinated with pVAX-ROM1 were significantly increased respectively, compared to controls receiving either pVAX1 alone or PBS. After lethal challenge, the mice immunized with pVAX-ROM1 showed an increased survival time compared with the mice in the controls. Our data suggested that a DNA vaccine pVAX-ROM1 encoding T. gondii rhomboid protein 1 triggered strong humoral and cellular responses, and prolonged survival time against T. gondii infection in BALB/c mice.
Veterinary Parasitology 08/2011; 184(2-4):154-60. · 2.58 Impact Factor
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ABSTRACT: The immune responses and protective efficacy against homologous challenge in chickens elicited by recombinant proteins of a rhomboid-like gene (ETRHO1) from Eimeria tenella was investigated in the present study. When chickens were immunized with the recombinant rhomboid antigen, specific antibody was generated by ELISA assay. In comparison with the PBS group, the expression levels of interleukin-2, interferon-γ, as well as the percentages of CD4⁺ and CD8⁺ cells in the group immunized with the recombinant rhomboid proteins were significantly increased (p < 0.01, p < 0.05, and p < 0.05, respectively). These results suggest that rhomboid was capable of eliciting humoral and cell-mediated immunity response in birds. Challenge experiments demonstrated that the recombinant rhomboid protein could provide chickens with a protection rate around 77.3%. Numbers of oocysts and cecal lesion from chickens in the group immunized with recombinant rhomboid proteins decreased significantly, and the body weight increased significantly when compared with chickens in the PBS group (p < 0.05). These results suggested that the recombinant rhomboid antigen was able to impart partial protection against homologous challenge in chicken and could be a potential candidate for an E. tenella vaccine development.
Parasitology Research 08/2011; 110(3):1139-45. · 2.15 Impact Factor
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Yanan Cai,
Yongxing Ai,
Quan Zhao, Jianhua Li,
Guilian Yang,
Pengtao Gong,
Qiuyue Wang,
Honglie Hou,
Guocai Zhang,
Lingdan Li,
Ju Yang,
He Li,
Jun Zheng,
Shuhong Li,
Xichen Zhang
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ABSTRACT: The telomerase reverse transcriptase (TERT) is primarily known for its ability to elongate telomeres for maintaining chromosomal integrity and delaying cellular senescence. It plays an important role in cell proliferation, differentiation, tumorigenesis, and aging. Telomerase includes two core components-an internal RNA moiety acting as a template of DNA extension and a catalytic subunit (TERT) which provides catalytic activity. Here, we described the cloning, sequence, and characterization of the TERT gene from Trichinella spiralis (T. spiralis). The prediction results of amino acid sequence showed that it possessed all the motifs characteristics of the TERT family members. T. spiralis TERT (Ts_TERT) cDNA contains an open reading frame encoding a protein with 1,201 amino acids with moleculer mass of 139 kDa and isoelectric point of 9.673, and the protein contains the conserved reverse transcriptase motifs 1, 2, A, B, C, D, and E, as well as the TERT-specific T motifs and the N-terminal conserved motifs GQ, CP, and QFP. While RT-PCR analysis indicates that TERT mRNA is expressed in T. spiralis adult worm, newborn larvae, and muscle larvae.
Parasitology Research 07/2011; 110(1):411-7. · 2.15 Impact Factor
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ABSTRACT: A DNA vaccine (pVAX1-TgADF) encoding Toxoplasma gondii actin depolymerizing factor (ADF) gene was constructed and the immune response and protective efficacy of this vaccine against homologous challenge in BALB/c mice were evaluated. High titers of specific antibody and increases in the percentage of CD4(+) and CD8(+) T lymphocyte cells were observed from BALB/c mice vaccinated with pVAX1-TgADF (P<0.05), when PBS group was used as control. The survival time of BALB/c mice in pVAX1-TgADF group was longer than those in control groups. The numbers of brain cysts in the experimental BALB/c mice immunized with pVAX1-TgADF reduced significantly compared with those in PBS group (P<0.05), and the rate of reduction could reach to around 42.8%. These results suggested that the DNA vaccine pVAX1-TgADF could generate specific humoral and cellular immune responses, prolong survival times, and reduce brain cysts load against T. gondii infection in BALB/c mice.
Veterinary Parasitology 06/2011; 179(1-3):1-6. · 2.58 Impact Factor
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ABSTRACT: The aim of the present study was to estimate the prevalence and risk factors of Dirofilaria immitis infection in dogs from Dandong, China. A total of 886 dogs were examined for D. immitis infection by microscopic examination and PCR, indicating that the prevalence was 16.6% (213/886) and 24.0% (147/886), respectively. The odds of infection were significantly higher in older dogs and dogs sheltered in outdoor, compared to the younger ones and ones sheltered in indoor. No significant difference of infection was observed in different genders, and between pure breed and cross-breed dogs in the same rearing conditions. These results indicated that the risk of exposure to D. immitis in dogs is high in Dandong, China, and prophylaxis against the parasite is advisable to decrease the incidence of canine dirofilariosis.
Veterinary Parasitology 06/2011; 183(1-2):189-93. · 2.58 Impact Factor
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ABSTRACT: Total nucleic acids from sporulated oocysts of Eimeria tenella isolated from Changchun in China were found to contain three extrachromosomal double-stranded RNA segments (dsRNAs) of 1.4, 2.4 and 3.6 kb in sizes. These RNAs were resistant to RNase A digestion under high salt concentration (0.3 M NaCl). RNA-dependent RNA polymerase (RDRP) activity was detected in crude extracts of E. tenella sporulated oocysts containing these nucleic acid species. Virus-like particles (VLPs) were shown to have a diameter of approximately 38 nm under Electron Microscopy (EM) after purification by sucrose density gradient centrifugation. In keeping with the nomenclature generally adopted for protozoan viruses, we have named this isolate as E. tenella virus (ETV) which is the first virus isolated from E. tenella.
Experimental Parasitology 03/2011; 127(3):646-50. · 2.12 Impact Factor
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ABSTRACT: Cryptosporidium andersoni parasited in the abomasum has been demonstrated as a cause of reduction of milk production in dairy cow. In this study, a novel chimeric DNA vaccine pVAX1-AB was constructed and the efficacy against Cryptosporidium parvum was determined. BALB/c mice were divided into 3 groups and immunized with DNA vaccine expressing the oocyst wall protein, AB protein of C. andersoni, the recombinant plasmid containing the AB gene, respectively. After inoculation of 1 × 10(6) oocysts of C. parvum, the humoral and cellular immune responses were detected. Experimental results showed that the recombinant plasmid can induce corresponding specific antibody response, simultaneously influenced cellular immune responses, and provided greater protection rate (48.6%) than the other groups. These results indicated that chimeric DNA vaccine has a potential in Cryptosporidium vaccine development.
Veterinary Parasitology 02/2011; 179(1-3):7-13. · 2.58 Impact Factor
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Pengtao Gong,
Jingzhi Zhang,
Lili Cao,
Zhang Nan, Jianhua Li,
Ju Yang,
Hengtong Fang,
Huping Jiao,
Tao Jiang,
Libo Su,
Xichen Zhang
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ABSTRACT: To investigate the presence of myeloma-associated antigens in Trichinella spiralis and their anti-tumor effect, cross-immune responses between antigens of the myeloma cell SP2/0 versus positive sera to T. spiralis, and antigens of T. spiralis versus positive sera to myeloma cell SP2/0 were determined using T. spiralis and myeloma specific enzyme-linked immunosorbent assays (ELISA). The myeloma-associated antigens in T. spiralis were separated by ultrafiltration and 2-D electrophoresis, and the amino acid sequences and molecular weights were determined by spectrometry. An obvious reaction was found between a 33 kDa antigen and positive sera, and the major component of the antigen was tropomyosin (TM), which is an surface acidic protein with 284 amino acids. Mice were immunized with TM to determine the anti-tumor effect in vivo. The results showed that CD4(+), CD8(+) T lymphocyte, and CD19(+) B lymphocyte were significantly increased (P<0.05). The anti-tumor effects were significantly different between mice immunized with the antigens or adjuvant alone (P<0.05), while the difference between mice immunized with antigens and whole T. spiralis was not significant (P>0.05). The results indicated that TM identified in this study may play a role in eliciting cross-protective immunity.
Experimental Parasitology 01/2011; 127(4):784-8. · 2.12 Impact Factor
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ABSTRACT: Asthma is an inflammatory disease of the lungs that is characterised by increased inflammatory cell infiltration into the airways and poor respiratory function. Ivermectin is a semi-synthetic derivative of a family of macrocyclic lactones that shows broad-spectrum anti-parasitic activity. This drug has been shown to possess anti-inflammatory activity, but whether it can be used in asthma treatment has not yet been investigated. In this study, we aimed to investigate the inhibitory effects of ivermectin on allergic asthma symptoms in mice.
We used a mouse asthma model, in which allergic airway inflammation and airway remodelling were induced by ovalbumin (OVA) sensitisation and challenge. Ivermectin or PBS treatment was administered 1 h before OVA challenge. Ivermectin at 2 mg/kg significantly diminished recruitment of immune cells, production of cytokines in the bronchoalveolar lavage fluids and secretion of OVA-specific IgE and IgG1 in the serum. Histological studies indicated that ivermectin suppressed mucus hypersecretion by goblet cells in the airway.
This is the first study to demonstrate that ivermectin is an effective suppressor of inflammation and may be efficacious in the treatment of non-infectious airway inflammatory diseases such as allergic asthma.
Agents and Actions 01/2011; 60(6):589-96. · 1.59 Impact Factor
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ABSTRACT: The development of Neospora caninum tachyzoites, an apicomplexan protozoan parasite, was studied in vitro using the human breast carcinoma cell 7 (MCF-7) as the host cell line. The extracellular NC-1 tachyzoites in MCF-7 cells were observed and counted daily for 6 consecutive days post-infection to establish the growth curve. The intracellular parasites were observed by acridine orange staining using Laser scanning confocal microscope. The results indicated that NC-1 tachyzoites invaded MCF-7 cells and multiplied intracellularly. The number of extracellular NC-1 tachyzoites started to increase rapidly around day 3 and reached the maximum number around day 4. Results from the present study suggested that MCF-7 cells were susceptible to NC-1 tachyzoites and could be used as an alternative cell line for in vitro studies.
Experimental Parasitology 12/2010; 126(4):536-9. · 2.12 Impact Factor
