RESUMEN
Acanthamoeba keratitis (AK) is an infection that is mostly observed in contact lens wearers. It is often misdiagnosed causing delays in the administration of the correct treatment. The aim of this study was to report the outcome of clinical and molecular diagnosis of AK cases during the summer of 2019 in the southern region of Brazil. Three suspected cases of AK were discovered after an ophthalmic examination at a public hospital in the city of Porto Alegre. These cases were then confirmed through laboratory diagnosis (cell culture and molecular analysis by PCR and sequencing). In each of the three clinical sample cell cultures of corneal scraping and molecular analysis confirmed the presence of Acanthamoeba spp., all belonging to the morphological group II and to the genotype T4, which is the most common genotype associated with AK. In addition, Acanthamoeba spp. isolated from one of the clinical samples was found to harbor the Candidatus Paracaedibacter acanthamoeba, a bacterial endosymbiont. The presence of Ca. Paracaedibacter acanthamoeba in clinical isolates requires further research to reveal its possible role in the pathogenicity of Acanthamoeba infections.
Asunto(s)
Queratitis por Acanthamoeba , Acanthamoeba , Amebiasis , Lentes de Contacto , Acanthamoeba/genética , Queratitis por Acanthamoeba/diagnóstico , Queratitis por Acanthamoeba/etiología , Amebiasis/complicaciones , Brasil , Lentes de Contacto/efectos adversos , Genotipo , HumanosRESUMEN
Glycolytic enzymes, such as enolase, have been described as multifunctional complex proteins that also display non-glycolytic activities, termed moonlighting functions. Although enolase multifunctionality has been described for several organisms, the conservation of enolase alternative functions through different phyla has not been explored with more details. A useful strategy to investigate moonlighting functions is the use of systems biology tools, which allow the prediction of protein functions/interactions by graph design and analysis. In this work, available information from protein-protein interaction (PPI) databases were used to design enolase PPI networks for four eukaryotic organisms, namely Homo sapiens, Drosophila melanogaster, Caenorhabditis elegans, and Saccharomyces cerevisiae, covering a wide spectrum of this domain of life. PPI networks with number of nodes ranging from 140 to 411 and up to 15,855 connections were generated, and modularity and centrality analyses, and functional enrichment were performed for all of them. The performed analyses showed that enolase is a central node within the networks, and that, in addition to its canonical interactions with proteins related to glycolysis and energetic metabolism, it is also part of protein clusters related to different biological processes, like transcription, development, and apoptosis, among others. Some of these non-glycolytic clusters, are partially conserved between networks, in terms of overall sharing of orthologs, overall cluster structure, and/or at the levels of key regulatory proteins within clusters. Overall, our results provided evidences of enolase multifunctionality and evolutionary conservation of enolase PPIs at all these levels.
Asunto(s)
Proteínas de Caenorhabditis elegans/metabolismo , Proteínas de Drosophila/metabolismo , Fosfopiruvato Hidratasa/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Secuencia de Aminoácidos , Animales , Caenorhabditis elegans , Proteínas de Caenorhabditis elegans/genética , Secuencia Conservada , Proteínas de Drosophila/genética , Drosophila melanogaster , Evolución Molecular , Humanos , Fosfopiruvato Hidratasa/genética , Mapeo de Interacción de Proteínas , Saccharomyces cerevisiae , Proteínas de Saccharomyces cerevisiae/genética , Alineación de Secuencia , Biología de SistemasRESUMEN
Listeria monocytogenes infections have been frequently reported in many food poisoning outbreaks around the world. In this work, the protein repertoires of L. monocytogenes ATCC 7644 cells treated or not with a 10(-3)mg/mL nisin sublethal concentration, established by antimicrobial susceptibility tests, were analyzed by LC-MS/MS. Overall, 179 proteins were identified, 9 of them more abundant in nisin-treated samples, and 4 more abundant in non-treated control samples. In nisin treated cells, proteins associated to oxidative stress response showed higher abundance. Also, the higher abundance of an enzyme related to the production of cell membrane lipids upon nisin exposure is suggestive of both a failure in conventional cell division mechanism and the activation of an alternative L-form mediated division mechanism. Finally, flagellar and motility proteins' overexpression upon nisin exposure is indicative of increased bacterial motility in response to the bacteriocin. Taken together, these results provide new insights on nisin effects on L. monocytogenes cells and on how this bacterium may overcome a bacteriocin-containing environment. BIOLOGICAL SIGNIFICANCE: The antimicrobial mechanism of nisin on target bacterial cells has been extensively studied since discovery of this bacteriocin. The nisin pore-forming mechanism is mediated by its binding to the pyrophosphate portion of membrane lipid II [1], but some evidences point out to alternative mechanisms. Results from assays with mutacin 1140 hybrids [2] showed that the portion of nisin that is not involved with lipid II binding could damage the bacterial cell, independently of pore formation [3,4]. Moreover, there are insufficient data to explain how nisin affects the bacterial survival. In this scenario, proteomics is an interesting approach, as a comparison between treated and untreated cells may provide insights of both antimicrobial mechanisms of action and bacterial response mechanisms [5].
Asunto(s)
Antibacterianos/farmacología , Proteínas Bacterianas/biosíntesis , Listeria monocytogenes/metabolismo , Nisina/farmacología , Relación Dosis-Respuesta a Droga , ProteómicaRESUMEN
Glycolytic enzymes, such as fructose-bisphosphate aldolase (FBA) and enolase, have been described as complex multifunctional proteins that may perform non-glycolytic moonlighting functions, but little is known about such functions, especially in parasites. We have carried out in silico genomic searches in order to identify FBA and enolase coding sequences in Echinococcus granulosus, the causative agent of cystic hydatid disease. Four FBA genes and 3 enolase genes were found, and their sequences and exon-intron structures were characterized and compared to those of their orthologs in Echinococcus multilocularis, the causative agent of alveolar hydatid disease. To gather evidence of possible non-glycolytic functions, the expression profile of FBA and enolase isoforms detected in the E. granulosus pathogenic larval form (hydatid cyst) (EgFBA1 and EgEno1) was assessed. Using specific antibodies, EgFBA1 and EgEno1 were detected in protoscolex and germinal layer cells, as expected, but they were also found in the hydatid fluid, which contains parasite's excretory-secretory (ES) products. Besides, both proteins were found in protoscolex tegument and in vitro ES products, further suggesting possible non-glycolytic functions in the host-parasite interface. EgFBA1 modeled 3D structure predicted a F-actin binding site, and the ability of EgFBA1 to bind actin was confirmed experimentally, which was taken as an additional evidence of FBA multifunctionality in E. granulosus. Overall, our results represent the first experimental evidences of alternative functions performed by glycolytic enzymes in E. granulosus and provide relevant information for the understanding of their roles in host-parasite interplay.
Asunto(s)
Echinococcus granulosus/enzimología , Echinococcus granulosus/genética , Fructosa-Bifosfato Aldolasa/genética , Fructosa-Bifosfato Aldolasa/metabolismo , Proteínas del Helminto/genética , Proteínas del Helminto/metabolismo , Fosfopiruvato Hidratasa/genética , Fosfopiruvato Hidratasa/metabolismo , Secuencia de Aminoácidos , Animales , Secuencia de Bases , ADN de Helmintos/genética , Equinococosis/parasitología , Echinococcus granulosus/patogenicidad , Echinococcus multilocularis/enzimología , Echinococcus multilocularis/genética , Fructosa-Bifosfato Aldolasa/química , Perfilación de la Expresión Génica , Genes de Helminto , Proteínas del Helminto/química , Interacciones Huésped-Parásitos , Isoenzimas/química , Isoenzimas/genética , Isoenzimas/metabolismo , Modelos Moleculares , Datos de Secuencia Molecular , Fosfopiruvato Hidratasa/química , Mapeo de Interacción de Proteínas , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Homología de Secuencia de Aminoácido , Especificidad de la EspecieRESUMEN
Transposable elements (TEs) comprise a significant fraction of the genome, and some models of the TE "life cycle" suggest that, in the last phases of the cycle, TEs should be represented, in the genomes, by inactive and degenerated copies. In this study, we analyzed, using a bioinformatics approach, the autonomous hAT elements and their derivatives (active non-autonomous, MITE relatives and degenerated copies) in 12 Drosophila genomes. We found 28 hAT elements that had derivatives. Most copies had features that suggested that they were active, while only a few degenerated copies were found. Because hAT elements comprise an evolutionarily old superfamily, one should expect to find many degenerated copies within the genome, although this was not observed in our study. These results suggest that primarily active copies of hAT elements are maintained in the euchromatic regions of the genome and that degenerated copies are removed from the genome by natural selection.