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1.
J Enzyme Inhib Med Chem ; 34(1): 171-178, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-30451014

RESUMO

Inosine 5'-monophosphate dehydrogenase (IMPDH) is an essential enzyme for the production of guanine nucleotides. Disruption of IMPDH activity has been explored as a therapeutic strategy for numerous purposes, such as for anticancer, immunosuppression, antiviral, and antimicrobial therapy. In the present study, we established a luciferase-based high-throughput screening system to identify IMPDH inhibitors from our chemical library of known bioactive small molecules. The screening of 1400 compounds resulted in the discovery of three irreversible inhibitors: disulfiram, bronopol, and ebselen. Each compound has a distinct chemical moiety that differs from other reported IMPDH inhibitors. Further evaluation revealed that these compounds are potent inhibitors of IMPDHs with kon values of 0.7 × 104 to 9.3 × 104 M-1·s-1. Both disulfiram and bronopol exerted similar degree of inhibition to protozoan and mammalian IMPDHs. Ebselen showed an intriguing difference in mode of inhibition for different IMPDHs, with reversible and irreversible inhibition to each Cryptosporidium parvum IMPDH and human IMPDH type II, respectively. In the preliminary efficacy experiment against cryptosporidiosis in severe combined immunodeficiency (SCID) mouse, a decrease in the number of oocyst shed was observed upon the oral administration of disulfiram and bronopol, providing an early clinical proof-of-concept for further utilization of these compounds as IMPDH inhibitors.


Assuntos
Descoberta de Drogas , Reposicionamento de Medicamentos , Inibidores Enzimáticos/isolamento & purificação , Inibidores Enzimáticos/farmacologia , Ensaios de Triagem em Larga Escala/métodos , IMP Desidrogenase/antagonistas & inibidores , Animais , Azóis/química , Azóis/isolamento & purificação , Azóis/farmacologia , Cryptosporidium parvum/enzimologia , Dissulfiram/química , Dissulfiram/isolamento & purificação , Dissulfiram/farmacologia , Inibidores Enzimáticos/química , Humanos , IMP Desidrogenase/metabolismo , Isoindóis , Cinética , Camundongos , Camundongos SCID , Compostos Organosselênicos/química , Compostos Organosselênicos/isolamento & purificação , Compostos Organosselênicos/farmacologia , Estudo de Prova de Conceito , Propilenoglicóis/química , Propilenoglicóis/isolamento & purificação , Propilenoglicóis/farmacologia , Bibliotecas de Moléculas Pequenas
2.
Exp Parasitol ; 172: 1-4, 2017 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-27876472

RESUMO

Cats are the only definitive hosts of Toxoplasma gondii and constitute an essential source of infection to all warm blooded animals and humans. Diagnosis of T. gondii infection in cats is fundamental for proper management and control of infection in humans and animals. In the current study, we have evaluated the diagnostic performance of tachyzoite lysate antigen (TLA) and different T. gondii recombinant antigens including surface antigen 2 (SAG2), dense granule proteins 2, 6, 7, 15 (GRA2, GRA6, GRA7, GRA15) and microneme 10 protein (MIC10) in immunoglobulin G enzyme linked-immunosorbent assay (IgG ELISA) using cat serum samples, with reference to latex agglutination test (LAT). Remarkably, TLA showed better performance than other recombinant antigens in IgG ELISAs as compared to LAT, with concordance and Kappa values of 94.27% and 0.93, respectively. Furthermore, to improve the reactivity of the recombinant antigens, we have developed IgG ELISAs using different combinations with these recombinant antigens. Strikingly, a combination of SAG2 and GRAs has relatively similar performance as TLA evidenced by concordance and Kappa values of 94.27% and 0.81, respectively. The developed ELISA with a combination of recombinant antigens can be used as a promising diagnostic tool for routine testing of T. gondii infection and mass screening in cats. The major advantages of this assay are the high sensitivity and specificity, lower cost, safer production and easiness of standardization in various laboratories worldwide.


Assuntos
Anticorpos Antiprotozoários/sangue , Doenças do Gato/diagnóstico , Toxoplasma/imunologia , Toxoplasmose Animal/diagnóstico , Animais , Antígenos de Protozoários/genética , Antígenos de Protozoários/imunologia , Gatos , Ensaio de Imunoadsorção Enzimática , Imunoglobulina G/sangue , Testes de Fixação do Látex , Proteínas de Protozoários/genética , Proteínas de Protozoários/imunologia , Proteínas Recombinantes de Fusão/imunologia , Sensibilidade e Especificidade , Testes Sorológicos
3.
Nat Commun ; 15(1): 4278, 2024 May 22.
Artigo em Inglês | MEDLINE | ID: mdl-38778039

RESUMO

Toxoplasma gondii is a global protozoan pathogen. Clonal lineages predominate in Europe, North America, Africa, and China, whereas highly recombinant parasites are endemic in South/Central America. Far East Asian T. gondii isolates are not included in current global population genetic structure analyses at WGS resolution. Here we report a genome-wide population study that compared eight Japanese and two Chinese isolates against representative worldwide T. gondii genomes using POPSICLE, a novel population structure analyzing software. Also included were 7 genomes resurrected from non-viable isolates by target enrichment sequencing. Visualization of the genome structure by POPSICLE shows a mixture of Chinese haplogroup (HG) 13 haploblocks introgressed within the genomes of Japanese HG2 and North American HG12. Furthermore, two ancestral lineages were identified in the Japanese strains; one lineage shares a common ancestor with HG11 found in both Japanese strains and North American HG12. The other ancestral lineage, found in T. gondii isolates from a small island in Japan, is admixed with genetically diversified South/Central American strains. Taken together, this study suggests multiple ancestral links between Far East Asian and American T. gondii strains and provides insight into the transmission history of this cosmopolitan organism.


Assuntos
Genoma de Protozoário , Filogenia , Toxoplasma , Toxoplasma/genética , Toxoplasma/classificação , Humanos , América do Norte , Genoma de Protozoário/genética , Toxoplasmose/parasitologia , China , América Central , Japão , Haplótipos , Variação Genética , Recombinação Genética
4.
Langmuir ; 29(41): 12737-43, 2013 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-24041254

RESUMO

The interparticle distance between nanoparticles (NPs) dispersed on on SiO2 was shown to be controlled by PEGylation. Ferritins with nanoparticle cores were prepared and PEGylated with poly(ethylene glycol)s (PEGs) of two different molecular weights. It was shown that the thickness of the PEG layer on the ferritin surface determines the interparticle distance under short Debye lengths. Under conditions where the Debye length was greater than the PEG layer thickness, distance between ferritins increased due to the electrostatic repulsive force. Results suggest that the PEG layer accommodated a small amount of counterions insufficient to cancel the ferritin outer surface charges. Simulation showed that ferritins adsorbed randomly and interparticle distance can be predicted theoretically. We demonstrate that PEGylated ferritins, that is, NP cores, can be dispersed on a surface with interval distances between particles determined by the combination of the ionic strength of the solution and the molecular weight of the PEG.


Assuntos
Ferritinas/química , Ácido Poliglicólico/química , Adsorção , Peso Molecular , Tamanho da Partícula , Dióxido de Silício/química , Eletricidade Estática , Propriedades de Superfície
5.
Nanotechnology ; 24(26): 265401, 2013 Jul 05.
Artigo em Inglês | MEDLINE | ID: mdl-23733263

RESUMO

A highly periodical Si nanodisk superlattice has been fabricated by our top-down process. Based on the realistic structure, a 3D simulation program using the finite element method is developed to calculate energy band structure, optical and electrical properties, as well as the intermediate band solar cell operation. Both the experiments and simulations reveal that miniband formation enhances the optical and electrical collections. Consequently, detailed electronic structure and conversion efficiency are examined to guide the optimal design of minibands. A theoretically predicted maximal efficiency of the explored Si nanodisk superlattice is 50.3%, which is promising, compared with well-known complicated Si tandem solar cells.

6.
Nanotechnology ; 24(1): 015301, 2013 Jan 11.
Artigo em Inglês | MEDLINE | ID: mdl-23221349

RESUMO

A sub-10 nm, high-density, periodic silicon nanodisk (Si-ND) array with a SiC interlayer has been fabricated using a new top-down process that involves a 2D array of a bio-template etching mask and damage-free neutral beam etching. Optical and electrical measurements were carried out to clarify the formation of mini-bands due to wavefunction coupling. We found that the SiC interlayer could enhance the optical absorption coefficient in the layer of Si-NDs due to the stronger coupling of wavefunctions. Theoretical simulation also indicated that wavefunction coupling was effectively enhanced in Si-NDs with a SiC interlayer, which precisely matched the experimental results. Furthermore, the I-V properties of a 2D array of Si-NDs with a SiC interlayer were studied through conductive AFM measurements, which indicated conductivity in the structure was enhanced by strong lateral electronic coupling between neighboring Si-NDs. We confirmed carrier generation and less current degradation in the structure due to high photon absorption and conductivity by inserting the Si-NDs into p-i-n solar cells.

7.
Nanotechnology ; 24(28): 285301, 2013 Jul 19.
Artigo em Inglês | MEDLINE | ID: mdl-23787817

RESUMO

We successfully fabricated defect-free, distributed and sub-20-nm GaAs quantum dots (named GaAs nanodisks (NDs)) by using a novel top-down technique that combines a new bio-template (PEGylated ferritin) and defect-free neutral beam etching (NBE). Greater flexibility was achieved when engineering the quantum levels of ND structures resulted in greater flexibility than that for a conventional quantum dot structure because structures enabled independent control of thickness and diameter parameters. The ND height was controlled by adjusting the deposition thickness, while the ND diameter was controlled by adjusting the hydrogen-radical treatment conditions prior to NBE. Photoluminescence emission due to carrier recombination between the ground states of GaAs NDs was observed, which showed that the emission energy shift depended on the ND diameters. Quantum level engineering due to both diameter and thickness was verified from the good agreement between the PL emission energy and the calculated quantum confinement energy.


Assuntos
Arsenicais/química , Gálio/química , Nanoestruturas/química , Nanotecnologia/métodos , Pontos Quânticos , Biotecnologia/métodos , Ferritinas/química , Microscopia Eletrônica de Varredura , Nanoestruturas/ultraestrutura , Tamanho da Partícula , Polietilenoglicóis/química
8.
Exp Parasitol ; 135(1): 42-9, 2013 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-23792005

RESUMO

Host cell invasion by apicomplexan parasites driven by gliding motility and empowered by actin-based movement is essential for parasite survival and pathogenicity. The parasites share a conserved invasion process: actin-based motility led by the coordination of adhesin-cytoskeleton via aldolase. A number of studies of host cell invasion in the Plasmodium species and Toxoplasma gondii have been performed. However, the mechanisms of host cell invasion by Babesia species have not yet been studied. Here, we show that Babesia gibsoni aldolase (BgALD) forms a complex with B. gibsoni thrombospondin-related anonymous protein (BgTRAP) and B. gibsoni actin (BgACT), depending on tryptophan-734 (W-734) in BgTRAP. In addition, actin polymerization is mediated by BgALD. Moreover, cytochalasin D, which disrupts actin polymerization, suppressed B. gibsoni parasite growth and inhibited the host cell invasion by parasites, indicating that actin dynamics are essential for erythrocyte invasion by B. gibsoni. This study is the first molecular approach to determine the invasion mechanisms of Babesia species.


Assuntos
Actinas/metabolismo , Babesia/enzimologia , Babesia/fisiologia , Eritrócitos/parasitologia , Frutose-Bifosfato Aldolase/metabolismo , Actinas/química , Animais , Babesia/efeitos dos fármacos , Citocalasina D/farmacologia , DNA Complementar/isolamento & purificação , DNA de Protozoário/isolamento & purificação , Feminino , Frutose-Bifosfato Aldolase/química , Frutose-Bifosfato Aldolase/genética , Cinética , Camundongos , Camundongos Endogâmicos ICR , Inibidores da Síntese de Ácido Nucleico/farmacologia , Fases de Leitura Aberta/genética , Polimerização , Proteínas de Protozoários/metabolismo
9.
Nanotechnology ; 23(6): 065302, 2012 Feb 17.
Artigo em Inglês | MEDLINE | ID: mdl-22248504

RESUMO

A sub-10 nm, high-density, periodic silicon-nanodisc (Si-ND) array has been fabricated using a new top-down process, which involves a 2D array bio-template etching mask made of Listeria-Dps with a 4.5 nm diameter iron oxide core and damage-free neutral-beam etching (Si-ND diameter: 6.4 nm). An Si-ND array with an SiO(2) matrix demonstrated more controllable optical bandgap energy due to the fine tunability of the Si-ND thickness and diameter. Unlike the case of shrinking Si-ND thickness, the case of shrinking Si-ND diameter simultaneously increased the optical absorption coefficient and the optical bandgap energy. The optical absorption coefficient became higher due to the decrease in the center-to-center distance of NDs to enhance wavefunction coupling. This means that our 6 nm diameter Si-ND structure can satisfy the strict requirements of optical bandgap energy control and high absorption coefficient for achieving realistic Si quantum dot solar cells.

10.
Therap Adv Gastroenterol ; 15: 17562848221118302, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36082177

RESUMO

Image-enhanced endoscopy (IEE) allows real-time high-contrast imaging of a targeted lesion without any special equipment. Among various IEE technologies, narrow-band imaging, in which a light of shorter wavelength is used, emphasizes the surface and blood vessel patterns on the mucosal surface. This technology has been widely used in endoscopic diagnosis in the gastrointestinal tract. Red dichromatic imaging (RDI) was recently developed; it utilizes lights of longer wavelengths (520-550, 595-610, and 620-640 nm), which have weak light scattering characteristics in contrast to narrow-band imaging. RDI was designed to enhance the visibility of deep-lying blood vessels and areas of bleeding, and it has been installed in the latest Olympus endoscopy system, EVIS X1, as an advanced version of the optical-digital method that was originally developed. Improving the visibility of deep blood vessels allows more accurate evaluation of esophageal varices and the degree of inflammation in ulcerative colitis. Easier identification of a bleeding source makes hemostasis quicker and easier to accomplish during endoscopic resection procedures such as endoscopic submucosal dissection and peroral endoscopic myotomy as well as during treatment of gastrointestinal bleeding from a peptide ulcer or colon diverticulum. The authors herein review the technological development and principles, review the existing literature on RDI, and discuss the utility and effectiveness of this unique IEE technology in gastrointestinal endoscopy.

11.
Vet Parasitol Reg Stud Reports ; 27: 100673, 2022 01.
Artigo em Inglês | MEDLINE | ID: mdl-35012729

RESUMO

Gastrointestinal nematodes cause massive economic losses as an important impediment to the development of animals around the world. This study aimed to investigate the microscopical diagnosis of nematode parasites of buffalo from Minoufiya, Egypt and molecular characterization of Bunostomum phlebotomum. We examined 390 fecal samples with floatation and fecal culture techniques to recognize different genera of nematodes. The results revealed B. phlebotomum (2.56%), Strongyloides papillosus (3.85%), Toxocara vitulorum (7.69%), Haemonchus sp. (1.28%), and Dictyocaulus viviparus (1.28%). The recovered eggs and larvae of nematodes were identified as well as the adults of B. phlebotomum. Age-wise, sex-wise, and seasonal prevalences of the recovered nematodes were recorded. Sequence analysis of the ITS-2 gene of B. phlebotomum was highly identical (99-100%) to sequences from Australia and China and occurred in the same clade with B. trigoncephalum. In conclusion, the study presented the coprological survey of gastrointestinal nematodes, and the genetic characterization of B. phlebotomum from Minoufiya Governorate, Egypt for the first time.


Assuntos
Doenças dos Bovinos , Nematoides , Ancylostomatoidea , Animais , Búfalos , Bovinos , Doenças dos Bovinos/parasitologia , Egito/epidemiologia
12.
Parasitol Int ; 87: 102533, 2022 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-34968753

RESUMO

Toxoplasmosis is a cosmopolitan protozoan zoonosis caused by Toxoplasma gondii infamous for inducing severe clinical manifestations in humans. Although the disease affects at least one billion people worldwide, it is neglected in many countries including developed ones. In literature, the epidemiological data documenting the actual incidence of the disease in humans and domestic animals from Japan are limited and importantly many earlier papers on T. gondii infections were published in Japanese and a considerable part is not available online. Herein, we review the current summary about the epidemiological situation of T. gondii infection in Japan and the potential associated risk factors in humans and animals as well as the different T. gondii genotypes isolated in Japan. Several T. gondii isolates have been identified among cats (TgCatJpTy1/k-3, TgCatJpGi1/TaJ, TgCatJpObi1 and TgCatJpOk1-4) and goats (TgGoatJpOk1-13). This literature review underscores the need for a nationwide investigation of T. gondii infection in Japanese people and assessment of the socioeconomic impact of the disease burden. Furthermore, epidemiological studies in domestic and wild animals and estimation of degree of contamination of soil or water with T. gondii oocysts are needed, for a better understanding of the scope of this public health concern.


Assuntos
Toxoplasma , Toxoplasmose Animal/epidemiologia , Toxoplasmose/epidemiologia , Animais , Animais Selvagens , Gatos , Humanos , Japão/epidemiologia , Oocistos , Toxoplasma/classificação , Toxoplasma/genética
13.
Parasitol Int ; 90: 102618, 2022 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-35777654

RESUMO

The one-humped camel (Camelus dromedarius) or dromedary is an economically important domestic animal. However, infectious diseases, including those caused by vector-borne hemopathogens, frequently compromise the health and production of camels. In this study, we examined infections caused by Babesia, Theileria, Trypanosoma, and Anaplasma species in camels in Egypt. We analyzed blood DNA samples from 148 camels reared in six Egyptian governorates (Giza, Asyut, Sohag, Qena, Luxor, and the Red Sea) using pathogen-specific Polymerase Chain Reaction (PCR) assays. Our results indicated that 29 (19.6%), 22 (14.9%), 1 (0.7%), 2 (1.4%), 1 (0.7%), 2 (1.4%), and 28 (18.9%) of the surveyed animals were infected with Babesia bovis, B. bigemina, Babesia sp. Mymensingh, Theileria sp. Yokoyama, Theileria equi, Trypanosoma evansi, and Anaplasma marginale, respectively. We found that a total of 68 (45.9%) animals were infected with at least one of the detected hemopathogens. Sequencing analyses of PCR amplicons confirmed our diagnostic results. This study is the first to report Theileria sp. Yokoyama and Babesia sp. Mymensingh in Egypt. This is also the first report of infection with these two species in one-humped camel. In conclusion, this study found that camels in Egypt are infected with several vector-borne hemopathogens, including novel parasite species.


Assuntos
Anaplasmose , Babesia , Babesiose , Doenças dos Bovinos , Theileria , Trypanosoma , Anaplasma/genética , Anaplasmose/epidemiologia , Animais , Babesia/genética , Babesiose/epidemiologia , Babesiose/parasitologia , Camelus , Bovinos , Doenças dos Bovinos/parasitologia , Egito/epidemiologia , Theileria/genética , Trypanosoma/genética
14.
Nanotechnology ; 22(10): 105301, 2011 Mar 11.
Artigo em Inglês | MEDLINE | ID: mdl-21289397

RESUMO

We created a two-dimensional array of sub-10 nm Si-nanodiscs (Si-NDs), i.e. a 2D array of Si-NDs, with a highly ordered arrangement and dense NDs by using a new top-down technique comprising advanced damage-free neutral-beam (NB) etching and a bio-template (iron oxide core) as a uniform sub-10 nm etching mask. The bandgap energy (E(g)) of the fabricated 2D array of Si-NDs can be simply controlled from 2.2 to 1.3 eV by changing the ND thickness from 2 to 12 nm. Due to weak quantum confinement existing in the diameter direction resulting from the sub-10 nm Si-ND diameter, even though the thickness of the Si-ND is much larger than the Bohr radius of Si, E(g) is still larger than the 1.1 eV E(g) of bulk Si. Si-ND not only has wide controllable E(g) but also a high absorption coefficient due to quantum confinement in three dimensions. This new technique is a promising candidate for developing new nanostructures and could be integrated into the fabrication of nanoelectronic devices.

15.
Exp Parasitol ; 127(4): 795-803, 2011 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-21281637

RESUMO

The DnaK-tetratricopeptide repeat (DnaK-TPR) gene (ToxoDB ID, TGME49_002020) is expressed predominantly at the bradyzoite stage. DnaK-TPR protein has a heat shock protein (DnaK) and tetratricopeptide repeat (TPR) domains with amino acid sequence similarity to the counterparts of other organisms (40.2-43.7% to DnaK domain and 41.1-66.0% to TPR domain). These findings allowed us to infer that DnaK-TPR protein is important in the tachyzoite-to-bradyzoite development or maintenance of cyst structure although the function of this gene is still unknown. An immunofluorescence assay (IFA) revealed that DnaK-TPR protein was expressed in Toxoplasma gondii-encysted and in vitro-induced bradyzoites and distributed in the whole part of parasite cells. We conducted yeast two-hybrid screening to identify proteins interacting with DnaK-TPR protein, and demonstrated that DnaK-TPR protein interacts with p23 co-chaperone protein (Tgp23). It was expected that DnaK-TPR protein would have a function as a molecular chaperon in bradyzoite cells associated with Tgp23. Possible mechanisms for this gene are discussed.


Assuntos
Chaperonas Moleculares/metabolismo , Domínios e Motivos de Interação entre Proteínas , Proteínas de Protozoários/metabolismo , Toxoplasma/metabolismo , Sequência de Aminoácidos , Animais , Sequência de Bases , Primers do DNA/química , Feminino , Células HEK293 , Humanos , Imunoprecipitação , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos ICR , Mitocôndrias/química , Chaperonas Moleculares/química , Chaperonas Moleculares/genética , Dados de Sequência Molecular , Sondas de Oligonucleotídeos/química , Proteínas de Protozoários/química , Proteínas de Protozoários/genética , Coelhos , Toxoplasma/genética , Toxoplasmose Animal/parasitologia
16.
Artigo em Inglês | MEDLINE | ID: mdl-21130891

RESUMO

The effect of intra-gastric infusion of skipjack tuna (Katsuwonus pelamis) muscle soluble extract and of carnosine, anserine and histidine, on pepsin-like protease activity was studied in the isolated, externally batch-cultured stomach of the rockfish (Sebastes schlegelii). Stomach was isolated from the fish, then intragastrically infused with the above solutions or with a balanced saline solution (control solution) as the stimulant at 0.1 mL/min for 6h. Intra-gastric efflux was collected for measurement of pepsin-like protease activity. Skipjack tuna soluble extract, but not carnosine, anserine or histidine alone caused significant enhancement of pepsin-like protease activity during the infusion. Pepsin-like protease activity from skipjack tuna soluble extract responded immediately after infusion and was kept for 150 min after infusion. Stomach motility at the end of infusion was also observed. These results suggest that isolated stomach can receive mucosal side stimulants in vitro. It is likely that some effective components for stomach digestion in fish exist in skipjack tuna soluble extract.


Assuntos
Peixes/metabolismo , Pepsina A/metabolismo , Estômago/enzimologia , Atum , Aminoácidos/análise , Animais , Feminino , Peixes/anatomia & histologia , Técnicas In Vitro , Masculino , Perfusão/métodos , Estômago/anatomia & histologia , Extratos de Tecidos/química , Extratos de Tecidos/farmacologia
17.
Parasitol Int ; 80: 102199, 2021 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-32961305

RESUMO

Apicomplexan parasite Toxoplasma gondii has three distinct clonal lineages: high, medium and low virulent strains, type I, II and III, respectively. T. gondii avoids the immune response by transforming from fast multiplying tachyzoite to slow multiplying bradyzoite, and establishing a chronic infection. In the present study, we isolated a new strain of T. gondii from cat feces in the Tokachi subprefecture, Hokkaido, Japan and named it as TgCatJpObi1 (Obi1) strain. Genotyping analysis of 12 loci revealed atypical characters close to type II, genotype 4 according to ToxoDB classification. Phenotypically, Obi1 strain shows slow growth rate and the ability of spontaneous cyst formation in both human foreskin fibroblast (HFFs) and mouse peritoneal macrophages in vitro without bradyzoite induction. Parasite virulence was assessed by means of mouse survival upon infection with either Obi1 or ME49 strains. Obi1 strain displayed no mortalities in comparison to type II clonal lineage, ME49 at LD50 to LD100 range (1 × 103-106 tachyzoites). Although virulence of Obi1 strain is significantly lower than that of ME49, nucleotide sequences analyses revealed that genes of virulence factors such as Gra15, Rop5, 16, 17, and 18 in Obi1 strain were 100% identical to those in the type II strain. Thus, characterization of a newly isolated strain, Obi1, is crucial to clarify the development of toxoplasmosis in both humans and animals.


Assuntos
Doenças do Gato/parasitologia , Toxoplasma/genética , Toxoplasma/patogenicidade , Fatores de Virulência/genética , Animais , Gatos , Fezes/parasitologia , Feminino , Japão , Camundongos , Camundongos Endogâmicos BALB C , Organismos Livres de Patógenos Específicos , Toxoplasma/isolamento & purificação , Virulência/genética
18.
J Vet Med Sci ; 82(5): 571-575, 2020 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-32224553

RESUMO

Cryptosporidium, a waterborne protozoan parasite, has a substantial veterinary and medical impact worldwide. This parasite is more often recognized during waterborne outbreaks because of its resistance to chlorine disinfection, small size making it difficult to inactivate/eliminate through filtration, and presence in many animal species including humans. Migratory waterfowl, in addition to acting as mechanical carriers of Cryptosporidium oocysts, can also serve as natural reservoirs of infection by host-specific Cryptosporidium species. For better understanding of the extent of genetic diversity and inter-relationships among avian isolates of Cryptosporidium, 200 fecal samples of migratory ducks from the Tokachi subprefecture, Hokkaido, Japan were collected and analyzed by nested PCR (N-PCR) at the 18S rRNA gene. N-PCR revealed that 11.5% (23/200) were positive for Cryptosporidium spp. Among all samples, sequence analysis identified that 10% (20/200) were 98-100% identical to Cryptosporidium avian genotype III. On the other hand, 1.5% (3/200) were 99-100% identical to C. baileyi. This is the first molecular study reporting the prevalence of Cryptosporidium in migratory ducks in Japan. Genetic diversity among Cryptosporidium isolates from humans and birds has been reported worldwide. Nevertheless, further studies are important to assess genetic variety and to elucidate the transmission dynamics of Cryptosporidium parasites.


Assuntos
Doenças das Aves/parasitologia , Criptosporidiose/epidemiologia , Cryptosporidium/genética , Animais , Cryptosporidium/classificação , Cryptosporidium/isolamento & purificação , DNA de Protozoário/isolamento & purificação , Patos , Fezes/parasitologia , Japão/epidemiologia , Reação em Cadeia da Polimerase/veterinária , RNA Ribossômico 18S/genética
19.
Exp Parasitol ; 121(1): 55-63, 2009 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-18950626

RESUMO

Toxoplasma gondii undergoes stage conversion from tachyzoites to bradyzoites in intermediate hosts. There have been many reports on bradyzoite-specific genes which are thought to be involved in stage conversion. Here, we described a novel T. gondii deoxyribose phosphate aldolase-like gene (TgDPA) expressing predominantly in bradyzoites. The TgDPA gene encodes 286 amino acids having a predicted molecular weight of 31kDa. Sequence analysis revealed that TgDPA had a deoxyribose phosphate aldolase (DeoC) domain with about 30% homology with its Escherichia coli counterpart. RT- and quantitative PCR analyses showed that the TgDPA gene was more expressed in bradyzoites and that its expression gradually increased during in vitro tachyzoite-to-bradyzoite stage conversion. A polyclonal antibody against recombinant TgDPA protein was raised in rabbits, and immunofluorescent analysis demonstrated that TgDPA was expressed in bradyzoites in vivo and in vitro. These findings indicate that the TgDPA gene is a new bradyzoite-specific marker and might play a role in bradyzoites.


Assuntos
Aldeído Liases/genética , Regulação Enzimológica da Expressão Gênica , Toxoplasma/genética , Aldeído Liases/biossíntese , Aldeído Liases/química , Sequência de Aminoácidos , Animais , Anticorpos Antiprotozoários , Western Blotting , Clonagem Molecular , Eletroforese em Gel de Poliacrilamida , Feminino , Técnica Indireta de Fluorescência para Anticorpo , Cinética , Masculino , Camundongos , Camundongos Endogâmicos ICR , Microscopia Confocal , Dados de Sequência Molecular , Reação em Cadeia da Polimerase/métodos , RNA Mensageiro/análise , Coelhos , Análise de Sequência , Toxoplasma/enzimologia , Toxoplasma/imunologia
20.
Mol Cell Biol ; 25(2): 671-84, 2005 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-15632068

RESUMO

Two competing models for fibroblast growth factor (FGF) receptor (FGFR) dimerization have recently emerged based on ternary FGF-FGFR-heparin crystal structures. In the symmetric two-end model, heparin promotes dimerization of two FGF-FGFR complexes by stabilizing bivalent interactions of the ligand and receptor through primary and secondary sites and by stabilizing direct receptor-receptor contacts. In the asymmetric model, there are no protein-protein contacts between the two FGF-FGFR complexes, which are bridged solely by heparin. To identify the correct mode of FGFR dimerization, we abolished interactions at the secondary ligand-receptor interaction site, which are observed only in the symmetric two-end model, using site-directed mutagenesis. Cellular studies and real-time binding assays, as well as matrix-assisted laser desorption ionization-time of flight analysis, demonstrate that loss of secondary ligand-receptor interactions results in diminished FGFR activation due to decreased dimerization without affecting FGF-FGFR binding. Additionally, structural and biochemical analysis of an activating FGFR2 mutation resulting in Pfeiffer syndrome confirms the physiological significance of receptor-receptor contacts in the symmetric two-end model and provides a novel mechanism for FGFR gain of function in human skeletal disorders. Taken together, the data validate the symmetric two-end model of FGFR dimerization and argue against the asymmetric model of FGFR dimerization.


Assuntos
Fatores de Crescimento de Fibroblastos/genética , Fatores de Crescimento de Fibroblastos/metabolismo , Mutação , Estrutura Quaternária de Proteína , Receptores de Fatores de Crescimento de Fibroblastos , Animais , Células Cultivadas , Dimerização , Fator 10 de Crescimento de Fibroblastos , Heparina/química , Heparina/genética , Heparina/metabolismo , Humanos , Modelos Moleculares , Estrutura Secundária de Proteína , Receptor Tipo 2 de Fator de Crescimento de Fibroblastos , Receptores de Fatores de Crescimento de Fibroblastos/química , Receptores de Fatores de Crescimento de Fibroblastos/genética , Receptores de Fatores de Crescimento de Fibroblastos/metabolismo , Transdução de Sinais/fisiologia , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Ressonância de Plasmônio de Superfície
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