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1.
Front Vet Sci ; 8: 778560, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34966810

RESUMO

Apicomplexan parasites possess several unique secretory organelles, including rhoptries, micronemes, and dense granules, which play critical roles in the invasion of host cells. The molecular content of these organelles and their biological roles have been well-studied in Toxoplasma and Plasmodium, but are underappreciated in Cryptosporidium, which contains many parasites of medical and veterinary importance. Only four proteins have previously been identified or proposed to be located in micronemes, one of which, GP900, was confirmed using immunogold electron microscopy (IEM) to be present in the micronemes of intracellular merozoites. Here, we report on the discovery of four new microneme proteins (MICs) in the sporozoites of the zoonotic species C. parvum, identified using immunofluorescence assay (IFA). These proteins are encoded by cgd3_980, cgd1_3550, cgd1_3680, and cgd2_1590. The presence of the protein encoded by cgd3_980 in sporozoite micronemes was further confirmed using IEM. Cgd3_980 encodes one of the three C. parvum rhomboid peptidases (ROMs) and is, thus, designated CpROM1. IEM also confirmed the presence of CpROM1 in the micronemes of intracellular merozoites, parasitophorous vacuole membranes (PVM), and feeder organelles (FO). CpROM1 was enriched in the pellicles and concentrated at the host cell-parasite interface during the invasion of sporozoites and its subsequent transformation into trophozoites. CpROM1 transcript levels were also higher in oocysts and excysted sporozoites than in the intracellular parasite stages. These observations indicate that CpROM1, an intramembrane peptidase with membrane proteolytic activity, is involved in host-parasite interactions, including invasion and proteostasis of PVM and FO.

2.
J Antimicrob Chemother ; 77(1): 124-134, 2021 12 24.
Artigo em Inglês | MEDLINE | ID: mdl-34648615

RESUMO

OBJECTIVES: To rapidly generate host cells with resistance to multiple compounds for differentiating drug action on parasite target or the host cell target (i.e. on-target or off-target effect) against the zoonotic enteric parasite Cryptosporidium parvum. METHODS: Transient overexpression of a multidrug resistance protein 1 (MDR1) gene in host cells (HCT-8 cell line) was explored to increase drug tolerance of the host cells to selected anti-cryptosporidial leads. In vitro cytotoxicity and anti-cryptosporidial efficacy of selected compounds were evaluated on the parasite grown in WT parental and transiently transfected HCT-8 cells. The approach was based on the theory that, for an epicellular parasite receiving consistent exposure to compounds in culture medium, overexpressing MDR1 in HCT-8 cells would increase drug tolerance of host cells to selected compounds but would not affect the anti-cryptosporidial efficacy if the compounds acted solely on the parasite target and the drug action on host cell target played no role on the antiparasitic efficacy. RESULTS: Six known anti-cryptosporidial leads were tested. Transient overexpression of MDR1 increased drug tolerance of HCT-8 cells on paclitaxel, doxorubicin HCl and vincristine sulphate (2.11- to 2.27-fold increase), but not on cyclosporin A, daunorubicin HCl and nitazoxanide. Increased drug tolerance in host cells had no effect on antiparasitic efficacy of paclitaxel, but affected that of doxorubicin HCl. CONCLUSIONS: Data confirmed that, at efficacious concentrations, paclitaxel acted mainly on the parasite target, while doxorubicin might act on both parasite and host cell targets. This model can be employed for studying the action of additional anti-cryptosporidial leads, and adapted to studying drug action in other epicellular pathogens. The limitation of the model is that the anti-cryptosporidial leads/hits need to be MDR1 substrates.


Assuntos
Membro 1 da Subfamília B de Cassetes de Ligação de ATP , Criptosporidiose , Cryptosporidium parvum , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/genética , Linhagem Celular Tumoral , Cryptosporidium parvum/efeitos dos fármacos , Doxorrubicina/efeitos adversos , Doxorrubicina/farmacologia , Humanos , Paclitaxel/efeitos adversos , Paclitaxel/farmacologia
3.
Microorganisms ; 9(9)2021 Sep 10.
Artigo em Inglês | MEDLINE | ID: mdl-34576816

RESUMO

Cryptosporidium parasites are known to be highly divergent from other apicomplexan species at evolutionary and biological levels. Here we provide evidence showing that the zoonotic Cryptosporidium parvum also differs from other apicomplexans, such as Toxoplasma gondii, by possessing only two tubulin-based filamentous structures, rather than an array of subpellicular microtubules. Using an affinity-purified polyclonal antibody against C. parvum ß-tubulin (CpTubB), we observed a long and a short microtubule that are rigid and stable in the sporozoites and restructured during the intracellular parasite development. In asexual development (merogony), the two restructuring microtubules are present in pairs (one pair per nucleus or merozoites). In sexual developmental stages, tubulin-based structures are detectable only in microgametes, but undetectable in macrogametes. These observations indicate that C. parvum parasites use unique microtubule structures that differ from other apicomplexans as part of their cytoskeletal elements.

4.
Microorganisms ; 9(5)2021 May 08.
Artigo em Inglês | MEDLINE | ID: mdl-34066754

RESUMO

Cryptosporidium parvum is a globally recognized zoonotic parasite of medical and veterinary importance. This parasite mainly infects intestinal epithelial cells and causes mild to severe watery diarrhea that could be deadly in patients with weakened or defect immunity. However, its molecular interactions with hosts and pathogenesis, an important part in adaptation of parasitic lifestyle, remain poorly understood. Here we report the identification and characterization of a C. parvum T-cell immunomodulatory protein homolog (CpTIPH). CpTIPH is a 901-aa single-pass type I membrane protein encoded by cgd5_830 gene that also contains a short Vibrio, Colwellia, Bradyrhizobium and Shewanella (VCBS) repeat and relatively long integrin alpha (ITGA) N-terminus domain. Immunofluorescence assay confirmed the location of CpTIPH on the cell surface of C. parvum sporozoites. In congruence with the presence of VCBS repeat and ITGA domain, CpTIPH displayed high, nanomolar binding affinity to host cell surface (i.e., Kd(App) at 16.2 to 44.7 nM on fixed HCT-8 and CHO-K1 cells, respectively). The involvement of CpTIPH in the parasite invasion is partly supported by experiments showing that an anti-CpTIPH antibody could partially block the invasion of C. parvum sporozoites into host cells. These observations provide a strong basis for further investigation of the roles of CpTIPH in parasite-host cell interactions.

5.
Parasitol Res ; 120(6): 2165-2174, 2021 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-33893549

RESUMO

Neospora caninum is an important pathogen commonly causing spontaneous abortion in livestock. The parasite is known to remain in cysts in an inactive state; or it can undergo expansive development within an intermediate host. However, the mechanisms that trigger the proliferation of N. caninum have not been thoroughly elucidated. For various organisms, it has been demonstrated that microRNAs (miRNAs) can act as important endogenous regulatory factors in gene regulation during cell differentiation and development. However, miRNAs and their function have not been studied in N. caninum. In this study, small RNA libraries from N. caninum tachyzoites (NC-1 strain) were analyzed using a high-throughput RNA sequencing technology combined with systematic bioinformatics analysis. A considerable number of novel miRNAs from N. caninum NC-1 strain tachyzoites were identified. Of the 300 miRNAs found by bioinformatics analysis, 10 were conserved miRNAs belonging to 10 metazoan miRNA families, while 290 were novel miRNAs. The expression of 13 novel miRNAs was verified by real-time quantitative PCR (qRT-PCR). Data from this study provided and identified authentic miRNAs for the first time in N. caninum. The study also introduces a framework for further investigations of RNAi-dependent regulatory mechanisms of the parasite and provides data for further understanding of N. caninum development.


Assuntos
MicroRNAs/metabolismo , Neospora/genética , RNA de Protozoário/metabolismo , Animais , Chlorocebus aethiops , Coccidiose , Regulação da Expressão Gênica , Biblioteca Gênica , Sequenciamento de Nucleotídeos em Larga Escala , Neospora/fisiologia , Reação em Cadeia da Polimerase em Tempo Real , Análise de Sequência de RNA , Células Vero
6.
Pathogens ; 11(1)2021 Dec 24.
Artigo em Inglês | MEDLINE | ID: mdl-35055969

RESUMO

Phosphoglucomutase 1 (PGM1) catalyzes the conversion between glucose-1-phosphate and glucose-6-phosphate in the glycolysis/glucogenesis pathway. PGM1s are typically cytosolic enzymes in organisms lacking chloroplasts. However, the protozoan Cryptosporidium parasites possess two tandemly duplicated PGM1 genes evolved by a gene duplication after their split from other apicomplexans. Moreover, the downstream PGM1 isoform contains an N-terminal signal peptide, predicting a non-cytosolic location. Here we expressed recombinant proteins of the two PGM1 isoforms from the zoonotic Cryptosporidium parvum, namely CpPGM1A and CpPGM1B, and confirmed their enzyme activity. Both isoforms followed Michaelis-Menten kinetics towards glucose-1-phosphate (Km = 0.17 and 0.13 mM, Vmax = 7.30 and 2.76 µmol/min/mg, respectively). CpPGM1A and CpPGM1B genes were expressed in oocysts, sporozoites and intracellular parasites at a similar pattern of expression, however CpPGM1A was expressed at much higher levels than CpPGM1B. Immunofluorescence assay showed that CpPGM1A was present in the cytosol of sporozoites, however this was enriched towards the plasma membranes in the intracellular parasites; whereas CpPGM1B was mainly present under sporozoite pellicle, although relocated to the parasitophorous vacuole membrane in the intracellular development. These observations indicated that CpPGM1A played a house-keeping function, while CpPGM1B played a different biological role that remains to be defined by future investigations.

7.
Acta Biochim Biophys Sin (Shanghai) ; 51(1): 104-111, 2019 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-30544221

RESUMO

Cryptosporidium parvum is an important zoonotic parasite that causes significant economic loss in the animal husbandry industry, especially the cattle industry. As there is no specific vaccine or drug against Cryptosporidium, a rapid and accurate method for the detection of C. parvum is of great significance. In this study, colloidal gold strips were developed based on Cryptosporidium parvum virus 1 (CSpV1) for the detection of C. parvum infection in cattle fecal samples. The colloidal gold solution was prepared by reducing trisodium citrate and the CSpV1 #5 monoclonal antibody was labeled with colloidal gold. A polyclonal antibody against the CSpV1 capsid protein and an anti-mouse IgG antibody were coated on the colloidal gold strips for use in the test and control lines, respectively. Our results showed that the detection sensitivity in fecal samples was up to a 1:64 dilution. There was no cross-reaction with Cryptosporidium andersoni or Giardia in the fecal samples. The different preservation conditions (room temperature, 4°C, and 37°C) and preservation time (7, 30, 60, and 90 days) were analyzed. The data showed that the strips could be preserved for 90 days at 4°C and for 60 days at room temperature or 37°C. The colloidal gold strips were used to detect the samples of 120 clinical fecal in Changchun, China. The results indicated that the rate of a positive test was 5% (6/120). This study provides a rapid and accurate method for detecting C. parvum infection in cattle and humans.


Assuntos
Doenças dos Bovinos/parasitologia , Criptosporidiose/parasitologia , Cryptosporidium parvum/fisiologia , Fezes/parasitologia , Animais , Anticorpos Monoclonais/química , Anticorpos Monoclonais/imunologia , Proteínas do Capsídeo/imunologia , Bovinos , Doenças dos Bovinos/diagnóstico , Doenças dos Bovinos/virologia , Criptosporidiose/diagnóstico , Criptosporidiose/virologia , Cryptosporidium parvum/virologia , Fezes/virologia , Coloide de Ouro/química , Humanos , Concentração de Íons de Hidrogênio , Vírus de RNA/imunologia , Vírus de RNA/fisiologia , Sensibilidade e Especificidade , Zoonoses/diagnóstico , Zoonoses/parasitologia , Zoonoses/virologia
8.
Oncol Lett ; 12(1): 405-412, 2016 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-27347159

RESUMO

As a gynecological oncology, ovarian cancer has high incidence and mortality. To study the mechanisms of ovarian cancer, the present study analyzed the GSE37582 microarray. GSE37582 was downloaded from Gene Expression Omnibus and included data from 74 ovarian cancer cases and 47 healthy controls. The differentially-expressed genes (DEGs) were screened using linear models for microarray data package in R and were further screened for functional annotation. Next, Gene Ontology and pathway enrichment analysis of the DEGs was conducted. The interaction associations of the proteins encoded by the DEGs were searched using the Search Tool for the Retrieval of Interacting Genes, and the protein-protein interaction (PPI) network was visualized by Cytoscape. Moreover, module analysis of the PPI network was performed using the BioNet analysis tool in R. A total of 284 DEGs were screened, consisting of 145 upregulated genes and 139 downregulated genes. In particular, downregulated FBJ murine osteosarcoma viral oncogene homolog (FOS) was an oncogene, while downregulated cyclin-dependent kinase inhibitor 1A (CDKN1A) was a tumor suppressor gene and upregulated cluster of differentiation 44 (CD44) was classed as an 'other' gene. The enriched functions included collagen catabolic process, stress-activated mitogen-activated protein kinases cascade and insulin receptor signaling pathway. Meanwhile, FOS (degree, 15), CD44 (degree, 9), B-cell CLL/lymphoma 2 (BCL2; degree, 7), CDKN1A (degree, 7) and matrix metallopeptidase 3 (MMP3; degree, 6) had higher connectivity degrees in the PPI network for the DEGs. These genes may be involved in ovarian cancer by interacting with other genes in the module of the PPI network (e.g., BCL2-FOS, BCL2-CDKN1A, FOS-CDKN1A, FOS-CD44, MMP3-MMP7 and MMP7-CD44). Overall, BCL2, FOS, CDKN1A, CD44, MMP3 and MMP7 may be correlated with ovarian cancer.

9.
Nat Commun ; 7: 11537, 2016 05 06.
Artigo em Inglês | MEDLINE | ID: mdl-27151551

RESUMO

Neutrophil extracellular traps (NETs), composed primarily of DNA and proteases, are released from activated neutrophils and contribute to the innate immune response by capturing pathogens. Plasmodium falciparum, the causative agent of severe malaria, thrives in its host by counteracting immune elimination. Here, we report the discovery of a novel virulence factor of P. falciparum, a TatD-like DNase (PfTatD) that is expressed primarily in the asexual blood stage and is likely utilized by the parasite to counteract NETs. PfTatD exhibits typical deoxyribonuclease activity, and its expression is higher in virulent parasites than in avirulent parasites. A P. berghei TatD-knockout parasite displays reduced pathogenicity in mice. Mice immunized with recombinant TatD exhibit increased immunity against lethal challenge. Our results suggest that the TatD-like DNase is an essential factor for the survival of malarial parasites in the host and is a potential malaria vaccine candidate.


Assuntos
Desoxirribonucleases/imunologia , Armadilhas Extracelulares/imunologia , Produtos do Gene tat/imunologia , Fatores de Virulência/imunologia , Animais , Desoxirribonucleases/genética , Desoxirribonucleases/metabolismo , Técnicas de Inativação de Genes , Produtos do Gene tat/genética , Produtos do Gene tat/metabolismo , Vacinas Antimaláricas , Malária Falciparum/prevenção & controle , Camundongos , Plasmodium berghei/genética , Plasmodium berghei/imunologia , Plasmodium berghei/metabolismo , Plasmodium falciparum/genética , Plasmodium falciparum/imunologia , Plasmodium falciparum/metabolismo , Vacinação , Fatores de Virulência/genética , Fatores de Virulência/metabolismo
10.
Exp Parasitol ; 154: 20-4, 2015 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-25845754

RESUMO

Giardia duodenalis is an important zoonotic intestinal parasite responsible for diarrhea in humans and other animals worldwide. The present study was conducted to assess the prevalence of bovine giardiosis and to perform molecular characterization of Giardia duodenalis in the northeast of China. A total of 655 fecal specimens were collected from dairy cattle in 15 farms located in three different provinces. G. duodenalis assemblages and subtypes were determined by sequence analysis of the triosephosphate isomerase (TPI) gene. As a whole, the G. duodenalis infection rate in dairy cattle was 7.9% (52/655), as determined by Lugol's iodine staining. Two assemblages were identified, namely, the potentially zoonotic assemblage A (n = 1), the livestock-specific assemblage E (n = 50), and a mixed infection case of assemblages A and E. Seven distinct subtypes of E assemblages were identified and E-XI, E-I and E-III are the major subtypes. Only subtype A-I was identified in assemblage A. Findings relevant to assemblage A are of public health importance. The results indicated the livestock-specific assemblage E is the major genotype and zoonotic assemblage A or B occurs very seldomly which is significantly different with previous report in the same area. So that determination of genotypes in individual epidemiological setting can make important contributions to public health.


Assuntos
Doenças dos Bovinos/epidemiologia , Giardia lamblia/genética , Giardíase/veterinária , Animais , Bovinos , Doenças dos Bovinos/parasitologia , China/epidemiologia , DNA de Protozoário/química , DNA de Protozoário/isolamento & purificação , Fezes/parasitologia , Feminino , Genótipo , Giardia lamblia/classificação , Giardia lamblia/isolamento & purificação , Giardíase/epidemiologia , Giardíase/parasitologia , Reação em Cadeia da Polimerase/veterinária , Polimorfismo Genético , Prevalência , Alinhamento de Sequência/veterinária
11.
J Appl Genet ; 56(3): 393-401, 2015 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-25604266

RESUMO

The digestive systems of mammals harbor a complex gut microbiome, comprising bacteria and other microorganisms that confer metabolic and immunological benefits to the host. Ruminants that digest plant-based foods have a four-compartment stomach consisting of the rumen, reticulum, omasum, and abomasum. The microorganisms in the stomach are essential for providing the host with critical nutrients. However, the majority of these microorganisms are unknown species. The microbiome of the stomach is diverse, and the majority of these organisms cannot be cultured. Next-generation sequencing (NGS) combined with bioinformatic analysis tools have allowed the dissection of the composition of the microbiome in samples collected from a specific environment. In this study, for the first time, the bacterial composition in two compartments, the reticulum and the omasum, of bovine were analyzed using a metagenomic approach and compared to the bacterial composition of the rumen. These data will assist in understanding the biology of ruminants and benefit the agricultural industry. The diversity and composition of the bacterial community in samples collected from the rumen, reticulum, and omasum of bovines in the Changchun Region of Northeast China were analyzed by sequencing the V3 region of the 16S rRNA gene using a barcoded Illumina paired-end sequencing technique, and the primary composition of the microbiome in the rumen, reticulum, and omasum of the bovines was determined. These microbiomes contained 17 phyla and 107 genera in all three samples. Five phyla, Bacteroidetes, Firmicutes, Proteobacteria, Spirochaetes, and Lentisphaerae, were the most abundant taxonomic groups. Additionally, the different stomach compartments harbored different compositions of the microorganisms.


Assuntos
Bovinos/microbiologia , Metagenoma , Omaso/microbiologia , Retículo/microbiologia , Animais , Bactérias/classificação , DNA Bacteriano/genética , Sequenciamento de Nucleotídeos em Larga Escala , Metagenômica , RNA Ribossômico 16S/genética , Rúmen/microbiologia , Análise de Sequência de DNA
12.
Parasit Vectors ; 7: 433, 2014 Sep 08.
Artigo em Inglês | MEDLINE | ID: mdl-25199527

RESUMO

BACKGROUND: MicroRNAs (miRNAs) have been shown to be present in plasma, which are remarkably stable, and have been suggested as disease biomarkers. Toxoplasma gondii (T. gondii) is a protozoan parasite that is infective to a wide range of animals and human beings. Previous studies have found that the parasite generated a large number of miRNAs during proliferation and it is known that the spectrum of miRNA expression in the infected hosts is pathogen-specific. To date, there are no reports regarding the application of microRNAs as biomarkers for the early detection of T. gondii infection. METHODS: In this study, we investigated the expression patterns of 414 murine miRNAs and tested their expression levels in the plasma after T. gondii infection by real-time PCR, with an ultimate purpose of identifying infection-related miRNAs. Three miRNAs in particular, exhibiting prominently elevated expressions, were further validated in a large number of infected mice. The Toxoplasma infection-specific miRNAs were confirmed by comparing their expression levels with those of mice infected with Plasmodium berghei, P. yoelii, P. chabaudi, Cryptosporidium parvum, Mouse hepatitis virus, and Staphylococcus aureus. RESULTS: Among the 414 miRNA candidates identified by a real-time PCR array, 71 were found to be up-regulated in the plasma of T. gondii infected mice. Three of those miRNAs (mmu-miR-712-3p, mmu-miR-511-5p and mmu-miR-217-5p) were prominently expressed in mice infected by both the RH and ME49 strains of T. gondii. Additionally, the elevated expression of these miRNAs was Toxoplasma-specific. CONCLUSIONS: The levels of the three miRNAs, mmu-miR-712-3p, mmu-miR-511-5p and mmu-miR-217-5p miRNAs, were found specifically up-regulated in plasma of mice after T. gondii infection.


Assuntos
MicroRNAs/sangue , Toxoplasma/isolamento & purificação , Toxoplasmose/diagnóstico , Animais , Biomarcadores/sangue , Modelos Animais de Doenças , Diagnóstico Precoce , Feminino , Regulação da Expressão Gênica , Humanos , Camundongos , Camundongos Endogâmicos BALB C , MicroRNAs/genética , Toxoplasma/genética , Regulação para Cima
13.
BMC Infect Dis ; 14: 429, 2014 Aug 04.
Artigo em Inglês | MEDLINE | ID: mdl-25091724

RESUMO

BACKGROUND: Vaccines are the most effective agents to control infections. However, recombinant vaccines often do not elicit strong immune responses. Protein antigens combined with proper adjuvants have been widely used to induce immune responses, especially the humoral immune responses, against various pathogens, including parasites. The extracellular matrix protein mindin has been recognised as an immune facilitator for initiating innate immune responses. It has therefore been expected to be a potentially potent adjuvant in the development of novel vaccines. The aim of this study was to investigate whether mindin could facilitate the induction of antigen-specific immune responses to recombinant antigens (rBAG1, rSRS4 and rSRS9) of Toxoplasma gondii in BALB/c mice. METHODS: In this study, we explored the adjuvant effect of the recombinant mindin in the generation of specific Th1 and Th2 responses to each of three T. gondii antigens, BAG1, SRS4 and SRS9. All mice in the experimental groups received either antigen alone or in combination with Freund's adjuvant or with the recombinant mindin. The immune responses after immunisation were measured by ELISA and lymphoproliferative assays. The immunised mice were challenged with live T. gondii tachyzoites, and the protection efficiency was compared between the groups. RESULTS: Our results revealed that mindin as an adjuvant could facilitate the recombinant proteins to efficiently stimulate humoral and cellular responses, including antigen-specific IgG1 and IgG2a, as well as lymphocyte proliferation. Furthermore, significantly improved protection against T. gondii infection was observed in the mindin group compared with that of Freund's adjuvant and no-adjuvant groups. CONCLUSIONS: The extracellular matrix protein mindin can effectively induce antigen-specific humoral and cell-mediated immune responses. Our study provides a valuable basis for the development of an efficient, safe, non-toxic vaccine adjuvant for future use in humans and animals.


Assuntos
Antígenos de Protozoários/imunologia , Proteínas da Matriz Extracelular/imunologia , Proteínas de Protozoários/imunologia , Toxoplasma/imunologia , Toxoplasmose/imunologia , Adjuvantes Imunológicos/administração & dosagem , Adjuvantes Imunológicos/genética , Animais , Anticorpos Antiprotozoários/imunologia , Antígenos de Protozoários/administração & dosagem , Antígenos de Protozoários/genética , Proteínas da Matriz Extracelular/administração & dosagem , Proteínas da Matriz Extracelular/genética , Feminino , Humanos , Imunidade Celular , Imunidade Humoral , Imunização , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Proteínas de Protozoários/administração & dosagem , Proteínas de Protozoários/genética , Vacinas Protozoárias/administração & dosagem , Vacinas Protozoárias/genética , Vacinas Protozoárias/imunologia , Células Th1/imunologia , Células Th2/imunologia , Toxoplasma/genética , Toxoplasmose/parasitologia , Toxoplasmose/prevenção & controle , Vacinas Sintéticas/administração & dosagem , Vacinas Sintéticas/genética , Vacinas Sintéticas/imunologia
14.
Opt Lett ; 39(13): 3942-5, 2014 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-24978777

RESUMO

A Ho³âº-doped PbF2 mid-IR laser crystal was successfully grown using the vertical Bridgman method. An intense 2.8 µm emission in Ho:PbF2 crystal was observed for the first time. By analyzing the absorption and emission measurements of the Ho:PbF2 crystal with the Judd-Ofelt theory, the intensity parameters Ω(2,4,6), exited state lifetimes, branching ratios, and emission cross-sections were calculated. It is found that the Ho:PbF2 crystal has high fluorescence branching ratio (20.99%), large emission cross section (1.44×10⁻²° cm²), long fluorescence lifetime (5.4 ms), and high quantum efficiency (88.4%) corresponding to the stimulated emission of Ho³âº: 5I6→5I7 transition. The structure of Ho:PbF2 crystal was also analyzed by the Raman spectrum, and it was found that the Ho:PbF2 crystal possesses low phonon energy of 257 cm⁻¹. We propose that the Ho:PbF2 crystal may be a promising material for 2.8 µm laser applications.


Assuntos
Fluoretos , Hólmio , Lasers de Estado Sólido , Chumbo , Cristalização , Fenômenos Ópticos , Espectrometria de Fluorescência , Espectrofotometria Infravermelho
15.
Proteomics ; 14(15): 1737-45, 2014 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-24888565

RESUMO

Toxoplasma gondii is an obligatory intracellular apicomplexan parasite which exploits host cell surface components in cell invasion and intracellular parasitization. Sulfated glycans such as heparin and heparan sulfate have been reported to inhibit cell invasion by T. gondii and other apicomplexan parasites such as Plasmodium falciparum. The aim of this study was to investigate the heparin-binding proteome of T. gondii. The parasite-derived components were affinity-purified on the heparin moiety followed by MS fingerprinting of the proteins. The heparin-binding proteins of T. gondii and P. falciparum were compared based on functionality and affinity to heparin. Among the proteins identified, the invasion-related parasite ligands derived from tachyzoite/merozoite surface and the secretory organelles were prominent. However, the profiles of the proteins were different in terms of affinity to heparin. In T. gondii, the proteins with highest affinity to heparin were the intracellular components with functions of parasite development contrasted to that of P. falciparum, of which the rhoptry-derived proteins were prominently identified. The profiling of the heparin-binding proteins of the two apicomplexan parasites not only explained the mechanism of heparin-mediated host cell invasion inhibition, but also, to a certain extent, revealed that the action of heparin on the parasite extended after endocytosis.


Assuntos
Heparina/metabolismo , Plasmodium falciparum/química , Proteoma/análise , Proteoma/metabolismo , Proteínas de Protozoários/análise , Proteínas de Protozoários/metabolismo , Toxoplasma/química , Sequência de Aminoácidos , Heparina/química , Dados de Sequência Molecular , Plasmodium falciparum/metabolismo , Ligação Proteica , Proteoma/química , Proteínas de Protozoários/química , Toxoplasma/metabolismo
16.
Opt Lett ; 39(9): 2747-50, 2014 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-24784093

RESUMO

In order to meet the demands for applications of optical refrigerators in the fields of spaceflight, aviation, space science, and detection, a 2 wt. % Yb3+-doped LuLiF4 crystal, as a new laser cooling material, was prepared and demonstrated by using a 178 mW diode laser centered at 1015 nm and a resonant extra-cavity scheme with an enhancement factor of 12.8. Cooling efficiency of 1.27% and a temperature drop of 14.3 K/W are obtained with 79% of the incident laser power being absorbed. Based on our results, a sample with background absorption of α=4.2×10(-4) cm(-1) can be potentially cooled down to ∼145 K. Our investigation shows that Yb3+-doped LuLiF4 crystal is potentially a promising candidate for solid-state refrigeration.

17.
Parasit Vectors ; 6(1): 241, 2013 Aug 20.
Artigo em Inglês | MEDLINE | ID: mdl-23958280

RESUMO

BACKGROUND: Toxoplasmosis is one of the most common parasitic zoonoses. The seroprevalence of Toxoplasma gondii infection in humans varies widely worldwide. Detection of Toxoplasma-specific antibodies has been a gold standard method for both epidemiological investigation and clinical diagnosis. Genetic investigation indicated that there is a wide distribution of different genome types or variants of the parasite prevalent in different areas. Thus the reliability of using antigens from parasites of a single genome type for diagnosis and epidemiology purposes needs to be extensively evaluated. METHODS: In this study, the prevalence of T. gondii infection among 880 clinically healthy individuals in China was systematically tested using crude soluble native antigens and purified recombinant antigens of type I and II T. gondii. The T. gondii-specific IgG and IgM in the sera was further confirmed using commercial Toxoplasmosis Diagnosis Kits and Western blot assays. RESULTS: The sero-prevalence of T. gondii-specific IgG detected with crude native Type I and type II antigens was 12.2% and 11.3% respectively. Whereas the overall prevalence was more than 20% when combined with the results obtained with recombinant tachyzoite and bradyzoite antigens. There was an obvious variation in immune-recognition of parasite antigens among the individuals studied. CONCLUSIONS: The general prevalence of anti-T. gondii IgG in the study population was likely much higher than previously reported. The data also suggested that there is more genetic diversity among the T. gondii isolates in China. Further, combination of recombinant antigens with clear immuno-recognition will be able to generate more sensitive diagnostic results than those obtained with crude antigens of T. gondii tachyzoites.


Assuntos
Antígenos de Protozoários/imunologia , Toxoplasma/imunologia , Toxoplasmose/diagnóstico , Toxoplasmose/epidemiologia , Anticorpos Antiprotozoários/imunologia , Antígenos de Protozoários/genética , China/epidemiologia , Ensaio de Imunoadsorção Enzimática , Feminino , Humanos , Imunoglobulina G , Imunoglobulina M/imunologia , Masculino , Prevalência , Estudos Soroepidemiológicos , Toxoplasma/genética , Toxoplasma/isolamento & purificação , Toxoplasmose/imunologia , Toxoplasmose/parasitologia
18.
Parasit Vectors ; 6: 161, 2013 Jun 04.
Artigo em Inglês | MEDLINE | ID: mdl-23734932

RESUMO

BACKGROUND: Toxoplasma gondii is an intracellular parasite that can modulate host responses and presumably host behavior. Host responses as well as pathogenesis vary depending on the parasite strains that are responsible for infection. In immune competent individuals, T. gondii preferentially infects tissues of the central nervous systems (CNS), which might be an additional factor in certain psychiatric disorders. While in immune-compromised individuals and pregnant women, the parasite can cause life-threatening infections. With the availability of the genome-wide investigation platform, the global responses in gene expression of the host after T. gondii infection can be systematically investigated. METHODS: Total RNA of brain tissues and peripheral lymphocytes of BALB/C mice infected with RH and ME 49 strain T. gondii as well as that of healthy mice were purified and converted to cRNA with incorporated Cy5-CTP (experimental samples), or Cy3-CTP (control samples). The labeled cRNA probes were hybridized to the Whole Mouse Genome Microarray. The impact of parasite infection on gene expression in both brain tissues and peripheral lymphocytes were analyzed. Differentially expressed genes were revalidated with real-time quantitative reverse transcriptase-polymerase chain reaction (Q-PCR). RESULTS: Data indicated that the genes associated with immunity were up-regulated after infection by the two parasite strains, but significant up-regulation was observed in both brain tissues and peripheral lymphocytes of mice infected with ME49 strain compared to that infected by RH strain. The pathways related to pathogenesis of the nervous system were more significantly up-regulated in mice infected with RH strain. CONCLUSIONS: Genetically distinct T. gondii strains showed clear differences in modulation of host pathophysiological and immunological responses in both brain tissue and peripheral lymphocytes. It was likely that some of the host responses to T. gondii infection were universal, but the immune response and CNS reaction were in a strain-specific manner.


Assuntos
Interações Hospedeiro-Patógeno , Toxoplasma/crescimento & desenvolvimento , Toxoplasmose Animal/imunologia , Toxoplasmose Animal/patologia , Transcriptoma , Animais , Encéfalo/patologia , DNA Complementar/genética , DNA Complementar/isolamento & purificação , Modelos Animais de Doenças , Feminino , Linfócitos/patologia , Camundongos , Camundongos Endogâmicos BALB C , Análise em Microsséries , RNA/genética , RNA/isolamento & purificação , Reação em Cadeia da Polimerase em Tempo Real , Toxoplasma/imunologia , Toxoplasma/patogenicidade , Toxoplasmose Animal/parasitologia
19.
J Proteome Res ; 12(5): 2185-93, 2013 May 03.
Artigo em Inglês | MEDLINE | ID: mdl-23566259

RESUMO

The malaria parasite Plasmodium falciparum utilizes host glycosaminoglycans (GAGs) as receptors for erythrocyte invasion and intravascular sequestration. Heparin and heparan sulfate (HS) are GAGs which can block erythrocyte invasion of the P. falciparum merozoite, albeit the molecular mechanisms remain poorly understood. Characterization of these heparin-binding merozoite proteins and key ligands in the host-parasite interplay will lead to a better understanding of the mechanism of erythrocyte invasion by malaria parasites. Here, schizont-derived proteins that bind heparin were enriched by affinity chromatography, and 6062 peptides from 811 P. falciparum-derived proteins were identified by two-dimensional liquid chromatography-mass spectrometry (LC/LC-MS/MS). The proteins were categorized into 14 functional groups ranging from pathogenesis, protein catabolic process to signal transduction. Proteins with predominant peptide counts were found to mainly originate from the rhoptry organelle of merozoites and the parasitized erythrocyte membrane. The profile of the heparin/HS-binding proteome of P. falciparum suggests they have important functions in the biology of the parasite.


Assuntos
Antimaláricos/química , Heparina/química , Plasmodium falciparum/metabolismo , Proteoma/metabolismo , Proteínas de Protozoários/metabolismo , Sequência de Aminoácidos , Antimaláricos/farmacologia , Células Cultivadas , Eritrócitos/parasitologia , Heparina/farmacologia , Interações Hospedeiro-Parasita , Humanos , Dados de Sequência Molecular , Plasmodium falciparum/efeitos dos fármacos , Ligação Proteica , Proteoma/química , Proteoma/isolamento & purificação , Proteômica , Proteínas de Protozoários/química , Proteínas de Protozoários/isolamento & purificação , Esquizontes/efeitos dos fármacos , Esquizontes/metabolismo , Espectrometria de Massas em Tandem
20.
Artigo em Chinês | MEDLINE | ID: mdl-24812834

RESUMO

OBJECTIVE: To clone and express Plasmodium falciparum erythrocyte membrane protein 1 DBLalpha (PfEMP1-DBLalpha) and three fragments genes, and screen the strongest affinity sequence with the red blood cell surface receptors-heparin or heparin sulfate in the structure of PfEMP1-DBLalpha. METHODS: The sequence of PfEMP1-DBLalpha1245 was optimized according to the characteristics of E. coli codon, synthesized, and divided into three fragments (DBLaA, DBLalphaB, and DBLalphaC) by PCR. Full-length gene and three gene fragments were subcloned into PGEX-4T-1 vector, and transformed into E. coli BL21 and then induced with IPTG for expression. The recombinant protein was purified from bacterial lysates using glutathione-Sepharose 4B. Heparin affinity test and glycosaminoglycan (GAG) inhibition test were used to analyze the affinity between recombinant protein and heparin. RESULTS: Four recombinant proteins(DBLalpha1245, DBLalphaA, DBLalphaB, and DBLalphaC) were expressed as solubility and the relative molecular weight (M(r) 73 600, M(r) 41 600, M(r) 42 500, and M(r) 41 500) were conformed to the prediction size. Heparin affinity test and GAG inhibition test showed that the four recombinant proteins were binded to the heparin-Sepharose, but not for the GST control. DBLalphaC (Q285-Y415) had the strongest affinity to heparin. CONCLUSION: The strongest affinity sequence with heparin or heparin sulfate in the structure of PfEMP1-DBLalpha is Q285-Y415, which plays a role in binding of Plasmodium falciparum infected red blood cells to the peripheral red blood cells.


Assuntos
Eritrócitos/parasitologia , Heparina/metabolismo , Plasmodium falciparum/genética , Proteínas de Protozoários/metabolismo , Sequência de Aminoácidos , Expressão Gênica , Ligação Proteica , Proteínas de Protozoários/genética , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo
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