Comprehensive characterization of extracellular vesicles produced by environmental (Neff) and clinical (T4) strains of Acanthamoeba castellanii
Elisa Gonçalves Medeiros,
Michele Ramos Valente,
Leandro Honorato,
Marina da Silva Ferreira,
Susana Ruiz Mendoza,
Diego de Souza Gonçalves,
Lucas Martins Alcântara,
Kamilla Xavier Gomes,
Marcia Ribeiro Pinto,
Ernesto S. Nakayasu,
Geremy Clair,
Isadora Filipaki Munhoz da Rocha,
Flavia C. G. dos Reis,
Marcio L. Rodrigues,
Lysangela R. Alves,
Leonardo Nimrichter,
Arturo Casadevall,
Allan Jefferson Guimarães
Affiliations
Elisa Gonçalves Medeiros
Departamento de Microbiologia e Parasitologia, Laboratório de Bioquímica e Imunologia das Micoses, Universidade Federal Fluminense, Niterói, Rio de Janeiro, Brazil
Michele Ramos Valente
Departamento de Microbiologia e Parasitologia, Laboratório de Bioquímica e Imunologia das Micoses, Universidade Federal Fluminense, Niterói, Rio de Janeiro, Brazil
Leandro Honorato
Departamento de Microbiologia Geral, Laboratório de Glicobiologia de Eucariotos, Universidade Federal do Rio de Janeiro (UFRJ), Rio de Janeiro, Brazil
Marina da Silva Ferreira
Departamento de Microbiologia e Parasitologia, Laboratório de Bioquímica e Imunologia das Micoses, Universidade Federal Fluminense, Niterói, Rio de Janeiro, Brazil
Susana Ruiz Mendoza
Departamento de Microbiologia e Parasitologia, Laboratório de Bioquímica e Imunologia das Micoses, Universidade Federal Fluminense, Niterói, Rio de Janeiro, Brazil
Diego de Souza Gonçalves
Programa de Pós-Graduação em Doenças Infecciosas e Parasitárias, Faculdade de Medicina, Universidade Federal do Rio de Janeiro (UFRJ), Rio de Janeiro, Brazil
Lucas Martins Alcântara
Departamento de Microbiologia e Parasitologia, Laboratório de Bioquímica e Imunologia das Micoses, Universidade Federal Fluminense, Niterói, Rio de Janeiro, Brazil
Kamilla Xavier Gomes
Departamento de Microbiologia e Parasitologia, Laboratório de Bioquímica e Imunologia das Micoses, Universidade Federal Fluminense, Niterói, Rio de Janeiro, Brazil
Marcia Ribeiro Pinto
Departamento de Microbiologia e Parasitologia, Laboratório de Bioquímica e Imunologia das Micoses, Universidade Federal Fluminense, Niterói, Rio de Janeiro, Brazil
Ernesto S. Nakayasu
Biological Science Division, Pacific Northwest National Laboratory, Richland, Washington, USA
Geremy Clair
Biological Science Division, Pacific Northwest National Laboratory, Richland, Washington, USA
Isadora Filipaki Munhoz da Rocha
Instituto Carlos Chagas, Fundação Oswaldo Cruz, Fiocruz, Curitiba, Paraná, Brazil
Flavia C. G. dos Reis
Instituto Carlos Chagas, Fundação Oswaldo Cruz, Fiocruz, Curitiba, Paraná, Brazil
Marcio L. Rodrigues
Instituto Carlos Chagas, Fundação Oswaldo Cruz, Fiocruz, Curitiba, Paraná, Brazil
Lysangela R. Alves
Instituto Carlos Chagas, Fundação Oswaldo Cruz, Fiocruz, Curitiba, Paraná, Brazil
Leonardo Nimrichter
Departamento de Microbiologia Geral, Laboratório de Glicobiologia de Eucariotos, Universidade Federal do Rio de Janeiro (UFRJ), Rio de Janeiro, Brazil
Arturo Casadevall
Department of Molecular Microbiology and Immunology, Johns Hopkins Bloomberg School of Public Health, Baltimore, Maryland, USA
Allan Jefferson Guimarães
Departamento de Microbiologia e Parasitologia, Laboratório de Bioquímica e Imunologia das Micoses, Universidade Federal Fluminense, Niterói, Rio de Janeiro, Brazil
ABSTRACT We conducted a comprehensive comparative analysis of extracellular vesicles (EVs) from two Acanthamoeba castellanii strains, Neff (environmental) and T4 (clinical). Morphological analysis via transmission electron microscopy revealed slightly larger Neff EVs (average = 194.5 nm) compared to more polydisperse T4 EVs (average = 168.4 nm). Nanoparticle tracking analysis (NTA) and dynamic light scattering validated these differences. Proteomic analysis of the EVs identified 1,352 proteins, with 1,107 common, 161 exclusive in Neff, and 84 exclusively in T4 EVs. Gene ontology and Kyoto Encyclopedia of Genes and Genomes (KEGG) mapping revealed distinct molecular functions and biological processes and notably, the T4 EVs enrichment in serine proteases, aligned with its pathogenicity. Lipidomic analysis revealed a prevalence of unsaturated lipid species in Neff EVs, particularly triacylglycerols, phosphatidylethanolamines (PEs), and phosphatidylserine, while T4 EVs were enriched in diacylglycerols and diacylglyceryl trimethylhomoserine, phosphatidylcholine and less unsaturated PEs, suggesting differences in lipid metabolism and membrane permeability. Metabolomic analysis indicated Neff EVs enrichment in glycerolipid metabolism, glycolysis, and nucleotide synthesis, while T4 EVs, methionine metabolism. Furthermore, RNA-seq of EVs revealed differential transcript between the strains, with Neff EVs enriched in transcripts related to gluconeogenesis and translation, suggesting gene regulation and metabolic shift, while in the T4 EVs transcripts were associated with signal transduction and protein kinase activity, indicating rapid responses to environmental changes. In this novel study, data integration highlighted the differences in enzyme profiles, metabolic processes, and potential origins of EVs in the two strains shedding light on the diversity and complexity of A. castellanii EVs and having implications for understanding host-pathogen interactions and developing targeted interventions for Acanthamoeba-related diseases.IMPORTANCEA comprehensive and fully comparative analysis of extracellular vesicles (EVs) from two Acanthamoeba castellanii strains of distinct virulence, a Neff (environmental) and T4 (clinical), revealed striking differences in their morphology and protein, lipid, metabolites, and transcripts levels. Data integration highlighted the differences in enzyme profiles, metabolic processes, and potential distinct origin of EVs from both strains, shedding light on the diversity and complexity of A. castellanii EVs, with direct implications for understanding host-pathogen interactions, disease mechanisms, and developing new therapies for the clinical intervention of Acanthamoeba-related diseases.
