In vitro gene silencing of independent phosphoglycerate mutase (iPGM) in the filarial parasite Brugia malayi

Abstract

Background: The phosphoglycerate mutase (PGM) enzyme catalyzes the interconversion of 2- and 3-phosphoglycerate in the glycolytic /gluconeogenic pathways that are present in the majority of cellular organisms. They can be classified as cofactor-dependent PGM (dPGM) or cofactor-independent PGM (iPGM). Vertebrates, yeasts, and many bacteria have only dPGM, while higher plants, nematodes, archaea, and many other bacteria have only iPGM. A small number of bacteria, including Escherichia coli and certain archaea and protozoa, contain both forms. The silencing of ipgm in Caenorhabditis elegans (C. elegans) has demonstrated the importance of this enzyme in parasite viability and, therefore, its potential as an anthelmintic drug target. In this study, the role of the Brugia malayi (B. malayi) ipgm in parasite viability, microfilaria release, embryogenesis, and in vivo development of infective larvae post-gene silencing was explored by applying ribonucleic acid (RNA) interference studies.

Results: The in vitro ipgm gene silencing by small interfering RNA (siRNA) leads to severe phenotypic deformities in the intrauterine developmental stages of female worms with a drastic reduction (~90%) in the motility of adult parasites and a significantly reduced (80%) release of microfilariae (mf) by female worms in vitro. Almost half of the in vitro-treated infective L3 displayed sluggish movement. The in vivo survival and development of siRNA-treated infective larvae (L3) was investigated in the peritoneal cavity of jirds where a ~45% reduction in adult worm establishment was observed.

Conclusion: The findings clearly suggest that iPGM is essential for both larval and adult stages of B. malayi parasite and that it plays a pivotal role in female worm embryogenesis. The results thus validate the Bm-iPGM as a putative anti-filarial drug target.