Dados do Trabalho

Título

Proteomic profiling of the in vitro excretory/secretory products of male and female Angiostrongylus costaricensis adult worms

Introdução

Angiostrongylus costaricensis is a nematode that causes a poorly understood intestinal human disease called abdominal angiostrongyliasis. No pharmacological treatment has thus far proven to be effective against the parasite. Excreted/secreted proteins (ESPs) released by parasites are vital players at the host-parasite interface. 

Objetivo (s)

This study aimed to characterize the proteomic profile of ESPs from A. costaricensis.  

Material e Métodos

Adult worms were kept in serum-free RPMI 1640 medium for 24 hours (SisGen A26AB0E, CEUA/IOC-037/2017), and culture supernatants were concentrated by ultrafiltration (3 kDa cutoff), followed by ethanol/acetone precipitation and in-solution trypsin digestion. Peptides were analyzed by nLC-MS/MS on a QExactive Plus Orbitrap (two biological replicates per class, four technical replicates each). 

Resultados e Conclusão

A total of 2026 unique nematode ESPs were identified in the secretome (FDR ≤ 1%) by the PatternLab for Proteomics V software suite. It comprises 15% of the UniProt reference proteome (UP000267027), including 1732 proteins from females and 1643 from males, several of which were identified for the first time. Using a label-free approach (i.e., the mean value of 2-3 most intense unique peptides per protein, normalized by the total signal), approximately 84% of the identified proteins could be quantified in at least one biological replicate. Their absolute abundances were distributed over a wide dynamic range, and a few abundant proteins could explain half the total ion intensity. The highest-ranked proteins included fatty acid and retinol-binding protein (FAR), transthyretin-like family protein, major sperm protein (MSP), peptidases/peptidase inhibitors, and proteins related to protein folding/stress response, energy metabolism, and cytoskeleton. Using an MS1-based relative quantification (i.e., considering the sum of the ionic current of the top 2-n unique peptides per protein, normalized by the total signal), a subset of 1052 proteins was quantified in both biological replicates of each class. It was used to perform a gender-related comparison of A. costaricensis secretome. Most of these proteins (789) were common to both genders, while 123 and 140 were exclusive to females and males, respectively. Only 12% of the common proteins were differentially released/secreted. Our results provide new biological knowledge on A. costaricensis and may contribute to developing new diagnostic, therapeutic, and vaccine tools for abdominal angiostrongyliasis.

Palavras Chave

Helminth; Nematode; Proteomics; secretome; Mass spectrometry.