Proteome analysis of the cardiac and pericardial tissue of Biomphalaria tenagophila populations susceptible and resistant to Schistosoma mansoni infection

Abstract

For a better comprehension of the parasite–host interaction, proteins expressed by the cardiac and pericardial tissues were compared between susceptible (Cabo Frio) and resistant (Taim) Biomphalaria tenagophila populations, challenged (c) and non-challenged (nc) with Schistosoma mansoni. Proteins were separated by two-dimensional gel electrophoresis (2DE) and stained with Coomassie blue. A total of 146 and 135 spots were observed in Cabo Frio (CFnc) and in Taim (Tnc) non-challenged populations, respectively, whereas 153 spots were detected in both Cabo Frio (CFc) and Taim (Tc) challenged populations. Regarding comparisons between CFnc and CFc, the numbers of exclusive spots obtained were one and nine, respectively, whereas Tnc yielded 17 and Tc eight exclusive spots. By comparing the total of spots in CF (nc + c) with T (nc + c) populations, we obtained: four exclusive spots for CFc; zero for CFnc; four for Tc and; one for Tnc. A quantitative comparison (reason >2.5) of the total spots of CF (nc + c) with T (nc + c) populations allowed us to distinguish five more intense spots for Tc, 14 for Tnc, 15 for CFnc and 11 for CFc. In the CFnc population, two proteins were identified: actin and ATP synthase alpha chain; in the CFc population, four proteins: actin, calmodulin, HSP70, and dehydrogenase; in the Tnc population, five proteins: matrilin, HSP70, actin, ATP synthase alpha chain and intermediate filament of the protein; and in the Tc population, three proteins: actin, alpha-S1 casein and ATP synthase alpha chain. Out of a total of 79 spots, only nine proteins were identified due to the low number of available nucleotide sequences in the GenBank. Nevertheless, knowing proteins regarded as differentially expressed is indispensable for hitherto unidentified genes implicated in B. tenagophila resistance and or susceptibility to S. mansoni infection.

Introduction

Biomphalaria tenagophila is the second major intermediate host of schistosomiasis in Brazil, responsible for the disease transmission in the state of São Paulo and several disease foci in the states of Santa Catarina, Minas Gerais, Rio de Janeiro and Rio Grande do Sul (Paraense, 1986). Schistosoma mansoni-susceptible and -resistant populations have already been described for B. tenagophila (Santos et al., 1979; Rosa et al., 2006, Coelho and Bezerra, 2006).

The defense system in mollusks is believed to be responsible for their resistance to S. mansoni infection (Van Der Knaap and Loker, 1990). The defense system of these organisms comprises hemocytes and proteins (lectins). Hemocytes are produced by the hematopoietic organ or the amebocyte-producing organ (APO) and other tissues. The APO is formed by a small quantity of primary ameboblasts resting on epithelial cells lining the pericardium. Ameboblasts are held in loose reticulum formed by extensions from smooth muscle extensions and few fibroblast cells (Jeong et al., 1983). Exposure to miracidia results in significant morphological changes in the organ in the presence of a large quantity of primary and secondary ameboblasts, undergoing mitosis. Fully activated APOs consist of masses of cells loosely arranged in zones of progressive maturation. Hemocytes act jointly with soluble factors present in the hemolymph in the process of encapsulation and destruction of pathogenic agents. Sporocysts can excrete and secrete substances that stimulate hemocytes mobility of resistant strains, inhibiting susceptible ones (Lodes and Yoshino, 1990).

Considering S. mansoni infection, variations in the intensity of defense effector mechanisms amongst different species and geographical lineages of mollusks are observed, which might be due to different degrees of susceptibility or resistance to infection (Martins-Souza et al., 2003). The relationship between susceptibility and the internal defense system in Biomphalaria lineages against S. mansoni infections remains unclear and little studied. Such investigation may provide a valuable knowledge on effector defense mechanisms and genetic features of Biomphalaria snails.

Herein, a proteomic approach may broaden knowledge on such defense mechanisms. Differently from the genomic approach, a proteome may be altered under different conditions and phenotypes, once proteins are known to determine phenotypes. The expressed proteins in a given moment must be known in order to obtain information on the genes function investigated. The search for differences in snails gene expression between resistance and susceptibility to infection can lead to the identification of markers responsible for these features. For protein expression analysis, the chosen method for the proteins separation was the bidimensional electrophoresis (2DE) and mass spectrometry (MS-MALDI-TOF/TOF) for their identification.

In this study, a comparative proteomic analysis of the cardiac and pericardial tissues of S. mansoni-resistant and -susceptible B. tenagophila mollusks is presented. Both phenotypes undergoing S. mansoni challenge or not have been analyzed by using two-dimensional gel electrophoresis coupled to mass spectrometry.