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1.
Cell Rep ; 34(12): 108896, 2021 03 23.
Artigo em Inglês | MEDLINE | ID: mdl-33761358

RESUMO

Severe and often fatal opportunistic fungal infections arise frequently following mucosal damage caused by trauma or cytotoxic chemotherapy. Interaction of fungal pathogens with epithelial cells that comprise mucosae is a key early event associated with invasion, and, therefore, enhancing epithelial defense mechanisms may mitigate infection. Here, we establish a model of mold and yeast infection mediated by inducible epithelial cell loss in larval zebrafish. Epithelial cell loss by extrusion promotes exposure of laminin associated with increased fungal attachment, invasion, and larval lethality, whereas fungi defective in adherence or filamentation have reduced virulence. Transcriptional profiling identifies significant upregulation of the epidermal growth factor receptor ligand epigen (EPGN) upon mucosal damage. Treatment with recombinant human EPGN suppresses epithelial cell extrusion, leading to reduced fungal invasion and significantly enhanced survival. These data support the concept of augmenting epithelial restorative capacity to attenuate pathogenic invasion of fungi associated with human disease.


Assuntos
Fator de Crescimento Epidérmico/farmacologia , Mucosa/microbiologia , Mucosa/patologia , Rhizopus/patogenicidade , Animais , Epigen/farmacologia , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/microbiologia , Células Epiteliais/ultraestrutura , Humanos , Hifas/efeitos dos fármacos , Hifas/crescimento & desenvolvimento , Larva/microbiologia , Modelos Biológicos , Mucosa/efeitos dos fármacos , Neutrófilos/efeitos dos fármacos , Neutrófilos/metabolismo , Proteínas Recombinantes/farmacologia , Rhizopus/ultraestrutura , Esporos Fúngicos/efeitos dos fármacos , Esporos Fúngicos/fisiologia , Fatores de Tempo , Peixe-Zebra/microbiologia
2.
J Vis Exp ; (143)2019 01 07.
Artigo em Inglês | MEDLINE | ID: mdl-30663701

RESUMO

Due to their transparency, genetic tractability, and ease of maintenance, zebrafish (Danio rerio) have become a widely-used vertebrate model for infectious diseases. Larval zebrafish naturally prey on the unicellular protozoan Paramecium caudatum. This protocol describes the use of P. caudatum as a vehicle for food-borne infection in larval zebrafish. P. caudatum internalize a wide range of bacteria and bacterial cells remain viable for several hours. Zebrafish then prey on P. caudatum, the bacterial load is released in the foregut upon digestion of the paramecium vehicle, and the bacteria colonize the intestinal tract. The protocol includes a detailed description of paramecia maintenance, loading with bacteria, determination of bacterial degradation and dose, as well as infection of zebrafish by feeding with paramecia. The advantage of using this method of food-borne infection is that it closely mimics the mode of infection observed in human disease, leads to more robust colonization compared to immersion protocols, and allows the study of a wide range of pathogens. Food-borne infection in the zebrafish model can be used to investigate bacterial gene expression within the host, host-pathogen interactions, and hallmarks of pathogenicity including bacterial burden, localization, dissemination and morbidity.


Assuntos
Doenças Transmitidas por Alimentos/parasitologia , Paramecium caudatum/fisiologia , Peixe-Zebra/parasitologia , Animais , Bactérias/metabolismo , Interações Hospedeiro-Patógeno , Larva/parasitologia , Paramecium caudatum/microbiologia , Comportamento Predatório , Peixe-Zebra/microbiologia
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