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1.
Vet Parasitol Reg Stud Reports ; 50: 101012, 2024 05.
Artigo em Inglês | MEDLINE | ID: mdl-38644044

RESUMO

A substantial parallel increase in prevalence and geographical spread of the rumen fluke, Calicophoron daubneyi, in livestock in western and central Europe has been recognized in the recent past. In the course of the examination of rectum feces of 471 red deer (Cervus elaphus) and one sika deer (Cervus nippon) from the Fascioloides magna endemic Sumava National Park in the years 2021 and 2022, rumen fluke eggs were detected in four red deer (0.8%) and the sika deer and identified as eggs of C. daubneyi by molecular analysis. Subsequent examination of rectal fecal samples of 247 beef cattle from 22 herds of 14 farms located in or nearby the national park revealed rumen fluke eggs in 53 samples (21.5%) originating from 16 herds of 11 farms, molecularly identified as C. daubneyi eggs as well. One C. daubneyi egg positive red deer and three C. daubneyi egg positive cattle samples also contained fasciolid eggs, respectively, which were detected in 9.5% or 3.6% of the total samples from red deer or cattle, respectively. Results of this investigation reveal the first finding of C. daubneyi in sika deer worldwide and in red deer in mainland Europe and add to the growing number of reports on C. daubneyi in livestock in Europe. Considering that the ratio of cattle excreting rumen fluke eggs exceeded that of deer substantially, it can reasonably be assumed that the C. daubneyi infections in deer are a consequence of the prevalent infection in cattle, illustrating a pathogen spillover event from livestock into wildlife.


Assuntos
Doenças dos Bovinos , Cervos , Fezes , Paramphistomatidae , Rúmen , Infecções por Trematódeos , Animais , Bovinos , Cervos/parasitologia , República Tcheca/epidemiologia , Infecções por Trematódeos/veterinária , Infecções por Trematódeos/epidemiologia , Infecções por Trematódeos/parasitologia , Paramphistomatidae/isolamento & purificação , Doenças dos Bovinos/epidemiologia , Doenças dos Bovinos/parasitologia , Rúmen/parasitologia , Prevalência , Fezes/parasitologia , Parques Recreativos
2.
Vet Parasitol Reg Stud Reports ; 40: 100860, 2023 05.
Artigo em Inglês | MEDLINE | ID: mdl-37068863

RESUMO

Helminth composition and burden data for dairy cows have not been reported for >40 years for Germany and even less information is available for Austria. In the context of two recent studies, helminth parasitism was studied in 32 cows (23 from six farms in Bavaria and Tyrol; 9 from one farm in Saxony) from pasture-based dairy farms necropsied during the housing period. Helminths were enumerated and identified based on morphological characters (all helminths but rumen flukes) or molecular techniques (rumen flukes). Thirteen species of gastrointestinal nematodes and two species each of liver flukes (Fasciola hepatica, Dicrocoelium dendriticum) and rumen flukes (Calicophoron daubneyi, Paramphistomum leydeni) were recorded; no lungworms were recovered from any cow. Early fourth-stage (inhibited) larval Ostertagia species nematodes (210 to 140,600) were recovered from all cows, 31 each had adult Ostertagia ostertagi/Ostertagia lyrata (40 to 2020) and Trichostrongylus axei (10 to 53,400), 23 Oesophagostomum radiatum (1 to 242) and 20 Cooperia punctata (10 to 3330). Other nematodes present in descending order of prevalence were: Cooperia oncophora/Cooperia surnabada, Ostertagia leptospicularis/Ostertagia kolchida, Oesophagostomum venulosum, Bunostomum phlebotomum, Chabertia ovina, Nematodirus helvetianus, Trichostrongylus longispicularis, Haemonchus contortus and Aonchotheca bilobata. The cows from Bavaria and Tyrol harbored more total gastrointestinal nematodes than that from Saxony (geometric mean adult plus inhibited larval nematodes, 6510 vs. 2051, respectively). However, in both cohorts of cows abomasal nematodes accounted for ∼97% of the total nematode burden with inhibited larval Ostertagia species nematodes contributing over 70% of the total gastrointestinal nematode burden and âˆ¼ 96% of the Ostertagia species burden. Approximately 44%, 37% and 19% of the cows harbored <5000, 5000 to 10,000 or > 10,000 total gastrointestinal nematodes, respectively. Fecal nematode egg and coproculture nematode larval counts significantly correlated with the cows' total adult nematode burden (rs = 0.354, p < 0.05, and rs = 0.608, p < 0.001, respectively). Although the magnitude of nematode burden to exert production effects on dairy cows is not well defined and may vary relative to several factors including nutritional supplementation, the level of mixed parasitism found in this investigation supports consideration of grazing dairy cows in helminth control measures, especially at the time of housing in autumn.


Assuntos
Doenças dos Bovinos , Helmintos , Nematoides , Infecções por Nematoides , Trichostrongyloidea , Feminino , Bovinos , Animais , Infecções por Nematoides/veterinária , Áustria/epidemiologia , Habitação , Alemanha/epidemiologia , Trichostrongylus , Ostertagia , Doenças dos Bovinos/epidemiologia , Doenças dos Bovinos/parasitologia
3.
Vet Parasitol ; 301: 109639, 2022 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-34959085

RESUMO

Parasitic respiratory infections in domestic sheep and goats are caused by Dictyocaulus filaria and various species belonging to the Protostrongylidae family of nematodes which frequently occur in mixed infections. Although the parasitism with protostrongylid lungworms is generally considered to be of low pathogenicity, there are reports of clinical disease including cases associated with Protostrongylus rufescens infection. The efficacy against P. rufescens of eprinomectin 5 mg/mL topical solution (EPRINEX® Multi, Boehringer Ingelheim) was thus evaluated in a clinical study compliant with GCP and VICH anthelmintic efficacy testing guidelines in adult sheep with naturally acquired infection. Following ranking on pre-treatment Protostrongylus fecal larval counts and forming into blocks of two animals, the sheep were randomly allocated to either remain untreated (control) or to be administered eprinomectin 5 mg/mL topical solution at 1 mL/5 kg body weight (equivalent to 1 mg eprinomectin per kg body weight) once as a pour-on. Fecal samples of the sheep were examined to monitor the larval excretion weekly for five weeks after treatment; then the animals were necropsied for lungworm recovery and count to determine the efficacy of the treatment. After treatment, Protostrongylus larval excretion decreased to zero within three weeks. Nematode counts demonstrated that the efficacy of the treatment with eprinomectin 5 mg/mL topical solution was 100 % against P. rufescens: no lungworms were recovered from any treated sheep while all controls harbored P. rufescens (range, 17-406) (p < 0.001). The treatment was well accepted; no treatment-related health problems were observed.


Assuntos
Anti-Helmínticos , Doenças das Cabras , Metastrongyloidea , Infecções por Nematoides , Animais , Anti-Helmínticos/uso terapêutico , Doenças das Cabras/tratamento farmacológico , Cabras , Ivermectina/análogos & derivados , Ivermectina/uso terapêutico , Infecções por Nematoides/tratamento farmacológico , Infecções por Nematoides/veterinária , Ovinos
4.
Parasitol Res ; 120(12): 4061-4066, 2021 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-34661730

RESUMO

Rumen flukes have received growing veterinary attention in western and central Europe during the past two decades because of an increase in prevalence of infection in cattle and sheep, including cases of severe clinical disease. Historically, rumen fluke infections in Europe were assumed to be caused mainly by Paramphistomum cervi (or species, which were later considered to be synonymous with P. cervi), but more recently molecular studies demonstrated Calicophoron daubneyi to be the predominating species. For the present investigation, adult rumen flukes isolated from 23 cattle originating from ten farms in Germany (Saxony [1], Baden-Württemberg [4], Bavaria [5]) and one farm in Austria (Tyrol) were analyzed to establish partial sequences of the mitochondrial cytochrome c oxidase subunit I (COI) and the complete sequence of the nuclear internal transcribed spacer 2 (ITS2). Flukes of five animals (dairy cows from three farms in Bavaria) were determined as P. leydeni, and flukes of 18 animals (dairy cows or cattle from cow-calf operations from eight farms in Saxony [1], Baden-Württemberg [4], Bavaria [2], and Tyrol [1]) were identified as C. daubneyi. Based on the molecular analysis of adult rumen flukes collected from cattle, the results of this investigation confirm the common occurrence of C. daubneyi in Germany and reveal the first definitive findings of P. leydeni in Germany and C. daubneyi in Austria.


Assuntos
Doenças dos Bovinos , Paramphistomatidae , Trematódeos , Infecções por Trematódeos , Animais , Áustria/epidemiologia , Bovinos , Doenças dos Bovinos/epidemiologia , Código de Barras de DNA Taxonômico , Feminino , Alemanha/epidemiologia , Paramphistomatidae/genética , Rúmen , Ovinos , Infecções por Trematódeos/epidemiologia , Infecções por Trematódeos/veterinária
5.
J Vet Pharmacol Ther ; 44(6): 952-960, 2021 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-34542908

RESUMO

Lactation is discussed as a physiological covariate which may influence the exposure characteristics of systemically acting drugs including macrocyclic lactones and potentially alter their pharmacological response. This study characterizes for the first time in the same study, the plasma profile and therapeutic anthelmintic efficacy of eprinomectin 5 mg/ml solution (EPRINEX® Multi, Boehringer Ingelheim) administered as a pour-on at 1 mg per kg body weight to lactating dairy goats. The study was conducted in compliance with VICH GCP and anthelmintic efficacy evaluation guidelines and included 20 goats harboring induced adult gastrointestinal and pulmonary nematode infections. The goats were blocked on pre-treatment body weight and randomly allocated either to remain untreated (control) or to be eprinomectin-treated. Plasma samples to determine the plasma disposition kinetics of eprinomectin (B1a component) were obtained at intervals up to 14 days following treatment when the animals were necropsied for parasite enumeration and identification. Basic pharmacokinetic parameters of eprinomectin determined in the ten eprinomectin-treated goats were as follows: AUClast , 23.8 ± 9.7 day*ng/ml and Cmax , 5.35 ± 2.27 ng/ml; individual maximum plasma concentrations were observed from 8 to 48 h after treatment (median Tmax , 0.5 days). Topical eprinomectin treatment efficacy, based on significant (p < .01) worm burden reductions in eprinomectin-treated animals relative to untreated controls, was ≥97% to 100% against adult Dictyocaulus filaria, Haemonchus contortus, Teladorsagia circumcincta(pinnata/trifurcata), Trichostrongylus axei, T. colubriformis, Cooperia curticei, Nematodirus battus, and Oesophagostomum venulosum. Both pharmacokinetic parameters and anthelmintic activity in lactating dairy goats were similar to those observed in parasitized young growing and adult female non-lactating dairy goats treated with eprinomectin administered as a pour-on.


Assuntos
Anti-Helmínticos , Doenças das Cabras , Infecções por Nematoides , Animais , Anti-Helmínticos/uso terapêutico , Feminino , Doenças das Cabras/tratamento farmacológico , Cabras , Ivermectina/análogos & derivados , Ivermectina/uso terapêutico , Lactação , Infecções por Nematoides/tratamento farmacológico , Infecções por Nematoides/veterinária
6.
Micromachines (Basel) ; 12(3)2021 Mar 06.
Artigo em Inglês | MEDLINE | ID: mdl-33800830

RESUMO

The microelectrode ion flux estimation (MIFE) is a powerful, non-invasive electrophysiological method for cellular membrane transport studies. Usually, the MIFE measurements are performed in a tissue culture dish or directly with tissues (roots, parts of the plants, and cell tissues). Here, we present a transwell system that allows for MIFE measurements on a cell monolayer. We introduce a measurement window in the transwell insert membrane, which provides direct access for the cells to the media in the upper and lower compartment of the transwell system and allows direct cell-to-cell contact coculture. Three-dimensional multiphoton lithography (MPL) was used to construct a 3D grid structure for cell support in the measurement window. The optimal polymer grid constant was found for implementation in transwell MIFE measurements. We showed that human umbilical vein endothelial cells (HUVECs) efficiently grow and maintain their physiological response on top of the polymer structures.

7.
ACS Nano ; 15(2): 2984-2993, 2021 02 23.
Artigo em Inglês | MEDLINE | ID: mdl-33480670

RESUMO

High-resolution imaging is essential for analysis of the steps and way stations of cargo transport in in vitro models of the endothelium. In this study, we demonstrate a microfluidic system consisting of two channels horizontally separated by a cell-growth-promoting membrane. Its design allows for high-resolution (down to single-molecule level) imaging using a high numerical aperture objective with a short working distance. To reduce optical aberrations and enable single-molecule-sensitive imaging, an observation window was constructed in the membrane via laser cutting with subsequent structuring using 3D multiphoton lithography for improved cell growth. The upper channel was loaded with endothelial cells under flow conditions, which showed polarization and junction formation. A coculture of human vascular endothelial cells with pericytes was developed that mimics the blood-brain barrier. Finally, this dual channel microfluidics system enabled 3D localization microscopy of the cytoskeleton and 3D single-molecule-sensitive tracing of lipoprotein particles.


Assuntos
Barreira Hematoencefálica , Microfluídica , Técnicas de Cocultura , Células Endoteliais , Humanos , Pericitos
8.
Vet Parasitol X ; 4: 100032, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-33083781

RESUMO

The efficacy of the eprinomectin, praziquantel, fipronil and (S)-methoprene combination parasiticide Broadline® (Boehringer Ingelheim Animal Health) was evaluated against developing larval and adult stages of Troglostrongylus brevior, a metastrongyloid pulmonary nematode which is reported to parasitize domestic cats in southern Europe with increasing frequency. Twenty four purpose-bred cats were experimentally infected with 100 third-stage T. brevior larvae each and randomly allocated to either remain untreated (control) or to be treated with the combination product when T. brevior were developing larval (6 days post inoculation, dpi) or adult nematodes (28 dpi) (eight cats per group). Treatments were administered topically at the minimum label dose of 0.12 mL/kg. Fecal samples of the cats were examined to confirm the presence of patent (adult) nematode infections prior to treatment at 28 dpi and to monitor the larval excretion. At necropsy (49 dpi), the weight of the pulmonary lymph nodes and lungs were determined, and T. brevior lungworms were recovered and counted. All control animals and cats to be treated at 28 dpi excreted T. brevior larvae 24 dpi and 26 dpi while no larvae were excreted by the cats treated at 6 dpi. Following treatment at 28 dpi, T. brevior larval excretion decreased immediatetly and ceased prior to necropsy. Nematode counts demonstrated that treatment with the combination product was 100 % efficacious against both developing larval and adult T. brevior: no lungworms were recovered from any treated cat while all control animals harbored T. brevior (range, 6-52) (p < 0.001). No treatment-related health problems or any other clinical signs were observed in the cats. However, significantly higher absolute and relative (organ weight to body weight ratio) pulmonary lymph node weights of the control animals compared with the treated cats at 6 dpi (p < 0.001 and p < 0.001, respectively) and at 28 dpi (p = 0.003 and p = 0.019, respectively) indicated the pathology of the T. brevior infection. In conclusion, the combination product was demonstrated to be 100 % efficacious against developing larval and adult T. brevior. Furthermore, it was demonstrated that indicators of impaired respiratory and immune systems resultant from T. brevior infection can be prevented with an efficacious treatment when administered during the pre-patent period of infection or are improving substantially within three weeks of treatment of cats harboring adult lungworms.

9.
PLoS Comput Biol ; 16(6): e1007902, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32603371

RESUMO

We present the software platform 2CALM that allows for a comparative analysis of 3D localisation microscopy data representing protein distributions in two biological samples. The in-depth statistical analysis reveals differences between samples at the nanoscopic level using parameters such as cluster-density and -curvature. An automatic classification system combines multiplex and multi-level statistical approaches into one comprehensive parameter for similarity testing of the compared samples. We demonstrated the biological importance of 2CALM, comparing the protein distributions of CD41 and CD62p on activated platelets in a 3D artificial clot. Additionally, using 2CALM, we quantified the impact of the inflammatory cytokine interleukin-1ß on platelet activation in clots. The platform is applicable to any other cell type and biological system and can provide new insights into biological and medical applications.


Assuntos
Plaquetas/metabolismo , Proteínas de Membrana/metabolismo , Microscopia/métodos , Modelos Biológicos , Trombose/metabolismo , Humanos , Aprendizado de Máquina , Selectina-P/metabolismo , Estudo de Prova de Conceito
10.
Vet Parasitol ; 277S: 100032, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-34392943

RESUMO

The efficacy of the eprinomectin, praziquantel, fipronil and (S)-methoprene combination parasiticide Broadline® (Boehringer Ingelheim Animal Health) was evaluated against developing larval and adult stages of Troglostrongylus brevior, a metastrongyloid pulmonary nematode which is reported to parasitize domestic cats in southern Europe with increasing frequency. Twenty four purpose-bred cats were experimentally infected with 100 third-stage T. brevior larvae each and randomly allocated to either remain untreated (control) or to be treated with the combination product when T. brevior were developing larval (6 days post inoculation, dpi) or adult nematodes (28 dpi) (eight cats per group). Treatments were administered topically at the minimum label dose of 0.12mL/kg. Fecal samples of the cats were examined to confirm the presence of patent (adult) nematode infections prior to treatment at 28 dpi and to monitor the larval excretion. At necropsy (49 dpi), the weight of the pulmonary lymph nodes and lungs were determined, and T. brevior lungworms were recovered and counted. All control animals and cats to be treated at 28 dpi excreted T. brevior larvae 24 dpi and 26 dpi while no larvae were excreted by the cats treated at 6 dpi. Following treatment at 28 dpi, T. brevior larval excretion decreased immediatetly and ceased prior to necropsy. Nematode counts demonstrated that treatment with the combination product was 100 % efficacious against both developing larval and adult T. brevior: no lungworms were recovered from any treated cat while all control animals harbored T. brevior (range, 6-52) (p<0.001). No treatment-related health problems or any other clinical signs were observed in the cats. However, significantly higher absolute and relative (organ weight to body weight ratio) pulmonary lymph node weights of the control animals compared with the treated cats at 6 dpi (p<0.001 and p<0.001, respectively) and at 28 dpi (p=0.003 and p=0.019, respectively) indicated the pathology of the T. brevior infection. In conclusion, the combination product was demonstrated to be 100 % efficacious against developing larval and adult T. brevior. Furthermore, it was demonstrated that indicators of impaired respiratory and immune systems resultant from T. brevior infection can be prevented with an efficacious treatment when administered during the pre-patent period of infection or are improving substantially within three weeks of treatment of cats harboring adult lungworms.

11.
Nanoscale Adv ; 2(6): 2422-2428, 2020 Jun 17.
Artigo em Inglês | MEDLINE | ID: mdl-36133392

RESUMO

The fabrication of two- and three-dimensional scaffolds mimicking the extracellular matrix and providing cell stimulation is of high importance in biology and material science. We show two new, biocompatible polymers, which can be 3D structured via multiphoton lithography, and determine their mechanical properties. Atomic force microscopy analysis of structures with sub-micron feature sizes reveals Young's modulus values in the 100 MPa range. Assessment of biocompatibility of the new resins was done by cultivating human umbilical vein endothelial cells on two-dimensionally structured substrates for four days. The cell density and presence of apoptotic cells has been quantified.

12.
Vet Parasitol ; 264: 42-46, 2018 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-30503090

RESUMO

The present study was conducted to further characterize the anthelmintic activity of the 0.5% w/v topical formulation of eprinomectin (EPRINEX® Pour-on, Merial) when administered at 1 mg/kg body weight to sheep in preventing the establishment of induced gastrointestinal and pulmonary nematode infections. Thirty-six female Merino sheep (∼4 months of age, weighing 27.0-36.0 kg) were blocked by pre-treatment body weight to form blocks of four animals. Within blocks, the animals were randomly allocated to either remain untreated (control) or be treated once with EPRINEX® either on Day 0, Day 7 or Day 14. Starting on Day 15, the sheep were given trickle infections with infective larvae of seven species of gastrointestinal nematodes and Dictyocaulus filaria lungworms daily for seven consecutive days. Five weeks after completion of the daily challenge (Day 56), the animals were necropsied for parasite recovery and count. Treatment with EPRINEX® prevented the establishment (>90%, p ≤ 0.027) of D. filaria, Teladorsagia circumcincta (pinnata/trifurcata), Cooperia curticei, Nematodirus battus, Trichostrongylus colubriformis and Oesophagostomum venulosum for at least 21 days, and of Haemonchus contortus and Trichostrongylus axei for at least 14 days. Sheep in the groups treated with EPRINEX® at Days 7 and 14 had significantly (p ≤ 0.018) higher Day -1 to Day 56 wt gains than the untreated controls. No treatment-related health problems or any other health problems were observed throughout the study.


Assuntos
Administração Tópica , Antinematódeos/administração & dosagem , Ivermectina/análogos & derivados , Infecções por Nematoides/veterinária , Doenças dos Ovinos/prevenção & controle , Animais , Feminino , Trato Gastrointestinal/parasitologia , Ivermectina/administração & dosagem , Pulmão/parasitologia , Infecções por Nematoides/tratamento farmacológico , Infecções por Nematoides/prevenção & controle , Distribuição Aleatória , Ovinos , Doenças dos Ovinos/tratamento farmacológico , Resultado do Tratamento
13.
Anal Chem ; 90(21): 12372-12376, 2018 11 06.
Artigo em Inglês | MEDLINE | ID: mdl-30350628

RESUMO

Mobility of proteins and lipids plays a major role in physiological processes. Platforms which were developed to study protein interaction between immobilized and mobile proteins suffer from shortcomings such as fluorescence quenching or complicated fabrication methods. Here we report a versatile platform comprising immobilized histidine-tagged proteins and biotinylated proteins in a mobile phase. Importantly, multiphoton photolithography was used for easy and fast fabrication of the platform and allows, in principle, extension of its application to three dimensions. The platform, which is made up of functionalized polymer structures embedded in a mobile lipid bilayer, shows low background fluorescence and allows for mobility of arbitrary proteins.


Assuntos
Acrilatos/química , Bicamadas Lipídicas/química , Polímeros/química , Proteínas/química , Difusão , Fluorescência , Processos Fotoquímicos
14.
Atherosclerosis ; 277: 53-59, 2018 10.
Artigo em Inglês | MEDLINE | ID: mdl-30173079

RESUMO

BACKGROUND AND AIMS: Exchange of cholesterol between high-density lipoprotein (HDL) particles and cells is a key process for maintaining cellular cholesterol homeostasis. Recently, we have shown that amphiphilic cargo derived from HDL can be transferred directly to lipid bilayers. Here we pursued this work using a fluorescence-based method to directly follow cargo transfer from HDL particles to the cell membrane. METHODS: HDL was either immobilized on surfaces or added directly to cells, while transfer of fluorescent cargo was visualized via fluorescence imaging. RESULTS: In Chinese hamster ovary (CHO) cells expressing the scavenger receptor class B type 1 (SR-B1), transfer of amphiphilic cargo from HDL particles to the plasma membrane was observed immediately after contact, whereas hydrophobic cargo remained associated with the particles; about 60% of the amphiphilic cargo of surface-bound HDL was transferred to the plasma membrane. Essentially no cargo transfer was observed in cells with low endogenous SR-B1 expression. Interestingly, transfer of fluorescently-labeled cholesterol was also facilitated by using an artificial linker to bind HDL to the cell surface. CONCLUSIONS: Our data hence indicate that the tethering function of SR-B1 is sufficient for efficient transfer of free cholesterol to the plasma membrane.


Assuntos
Antígenos CD36/metabolismo , Membrana Celular/metabolismo , HDL-Colesterol/sangue , Microscopia de Fluorescência , Imagem Individual de Molécula/métodos , Animais , Células CHO , Cricetulus , Células Hep G2 , Células Endoteliais da Veia Umbilical Humana , Humanos , Interações Hidrofóbicas e Hidrofílicas , Transporte Proteico , Propriedades de Superfície , Fatores de Tempo
15.
Int J Mol Sci ; 19(4)2018 Apr 11.
Artigo em Inglês | MEDLINE | ID: mdl-29641438

RESUMO

Here, we measure the actin cytoskeleton arrangement of different morphological states of human platelets using a new protocol for photo-switching of rhodamine class fluorophores. A new medium composition was established for imaging the cytoskeleton using Alexa Fluor 488 conjugated to phalloidin. Morphological states of platelets bound to a glass substrate are visualized and quantified by two-dimensional localization microscopy at nanoscopic resolution. Marker-less drift correction yields localization of individual Alexa 488 conjugated to phalloidin with a positional accuracy of 12 nm.


Assuntos
Actinas/metabolismo , Plaquetas/ultraestrutura , Actinas/ultraestrutura , Plaquetas/metabolismo , Células Cultivadas , Corantes Fluorescentes/química , Humanos , Microscopia de Fluorescência/métodos , Sensibilidade e Especificidade
16.
BMC Vet Res ; 13(1): 148, 2017 May 30.
Artigo em Inglês | MEDLINE | ID: mdl-28558828

RESUMO

BACKGROUND: The anthelmintic efficacy of the 0.5% w/v topical formulation of eprinomectin (EPN), EPRINEX® Pour-on (Merial) when administered at 1 mg/kg body weight was evaluated in sheep in two dose confirmation laboratory studies and one multicenter field study. In addition, the pharmacokinetics of EPN when administered at that dosage to adult sheep was determined. RESULTS: In the two dose confirmation studies, which included 10 sheep each, sheep treated with topical EPN had significantly (p < 0.05) fewer of the following nematodes than the untreated sheep with overall reduction of nematode counts by >99%: adult Dictyocaulus filaria, Haemonchus contortus, Teladorsagia circumcincta(pinnata/trifurcata), Trichostrongylus axei, T. colubriformis, T. vitrinus, Cooperia curticei, Nematodirus battus, Strongyloides papillosus, Chabertia ovina and Oesophagostomum venulosum, and inhibited fourth-stage Teladorsagia larvae. A total of 196 sheep harboring naturally acquired gastrointestinal nematode infections were included in the field efficacy study at two sites each in Germany (48 Merino x Ile de France lambs, 52 adult Merino females) and in Italy (adult male and female Bagnolese, Lacaune, Lacaune x Bagnolese, Bagnolese x Sarda sheep; 48 animals per site). Animals were blocked on pre-treatment body weight and within each block, one animal was randomly assigned to the control (untreated) group and three animals were randomly assigned to be treated with topical EPN. Examination of feces 14 days after treatment demonstrated that, relative to the controls, topical EPN-treated sheep had significantly (p < 0.0001) lower strongylid egg counts. Reduction was ≥97% at each site and 98.6% across all sites. Pharmacokinetics of EPN following single treatment with topical EPN were determined in eight ~4.5 year old female Merino cross sheep based on the analysis of plasma samples which were collected from two hours to 21 days following treatment. The main pharmacokinetic parameters were: Cmax 6.20 ± 1.71 ng/mL, AUClast 48.8 ± 19.2 day*ng/mL, Tmax 3.13 ± 2.99 days and T1/2 6.40 ± 2.95 days. No treatment-related health problems or adverse drug events were observed in any study. CONCLUSION: These studies demonstrated 0.5% w/v EPN administered topically at 1 mg/kg body weight to be highly efficacious against a broad range of ovine gastrointestinal nematodes and D. filaria lungworms and well tolerated by sheep of different ages, breeds, gender and physiological status.


Assuntos
Antinematódeos/uso terapêutico , Gastroenteropatias/veterinária , Ivermectina/análogos & derivados , Pneumopatias Parasitárias/veterinária , Infecções por Nematoides/veterinária , Doenças dos Ovinos/parasitologia , Animais , Antinematódeos/farmacocinética , Feminino , Gastroenteropatias/parasitologia , Helmintíase Animal/tratamento farmacológico , Ivermectina/farmacocinética , Ivermectina/uso terapêutico , Pneumopatias Parasitárias/tratamento farmacológico , Masculino , Infecções por Nematoides/tratamento farmacológico , Ovinos , Doenças dos Ovinos/tratamento farmacológico
17.
Sci Rep ; 6: 32317, 2016 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-27580632

RESUMO

In transfusion medicine, the identification of the Rhesus D type is important to prevent anti-D immunisation in Rhesus D negative recipients. In particular, the detection of the very low expressed DEL phenotype is crucial and hence constitutes the bottleneck of standard immunohaematology. The current method of choice, adsorption-elution, does not provide unambiguous results. We have developed a complementary method of high sensitivity that allows reliable identification of D antigen expression. Here, we present a workflow composed of high-resolution fluorescence microscopy, image processing, and machine learning that - for the first time - enables the identification of even small amounts of D antigen on the cellular level. The high sensitivity of our technique captures the full range of D antigen expression (including D+, weak D, DEL, D-), allows automated population analyses, and results in classification test accuracies of up to 96%, even for very low expressed phenotypes.


Assuntos
Aprendizado de Máquina , Sistema do Grupo Sanguíneo Rh-Hr/classificação , Eritrócitos/metabolismo , Humanos , Microscopia de Fluorescência , Fenótipo , Sistema do Grupo Sanguíneo Rh-Hr/sangue , Imunoglobulina rho(D)/metabolismo , Estatística como Assunto
19.
Acta Crystallogr Sect F Struct Biol Cryst Commun ; 68(Pt 10): 1217-21, 2012 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-23027751

RESUMO

The mechano-enzyme dynamin-related protein 1 plays an important role in mitochondrial fission and is implicated in cell physiology. Dysregulation of Drp1 is associated with abnormal mitochondrial dynamics and neuronal damage. Drp1 shares structural and functional similarities with dynamin 1 with respect to domain organization, ability to self-assemble into spiral-like oligomers and GTP-cycle-dependent membrane scission. Structural studies of human dynamin-1 have greatly improved the understanding of this prototypical member of the dynamin superfamily. However, high-resolution structural information for full-length human Drp1 covering the GTPase domain, the middle domain and the GTPase effector domain (GED) is still lacking. In order to obtain mechanistic insights into the catalytic activity, a nucleotide-free GTPase-GED fusion protein of human Drp1 was expressed, purified and crystallized. Initial X-ray diffraction experiments yielded data to 2.67 Šresolution. The hexagonal-shaped crystals belonged to space group P2(1)2(1)2, with unit-cell parameters a = 53.59, b = 151.65, c = 43.53 Å, one molecule per asymmetric unit and a solvent content of 42%. Expression of selenomethionine-labelled protein is currently in progress. Here, the expression, purification, crystallization and X-ray diffraction analysis of the Drp1 GTPase-GED fusion protein are presented, which form a basis for more detailed structural and biophysical analysis.


Assuntos
GTP Fosfo-Hidrolases/química , Proteínas Associadas aos Microtúbulos/química , Proteínas Mitocondriais/química , Cristalização , Cristalografia por Raios X , Dinaminas , GTP Fosfo-Hidrolases/isolamento & purificação , Humanos , Proteínas Associadas aos Microtúbulos/isolamento & purificação , Proteínas Mitocondriais/isolamento & purificação , Proteínas Recombinantes/química , Proteínas Recombinantes/isolamento & purificação
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