Identification and characterization of the Onchocerca volvulus Excretory Secretory Product Ov28CRP, a putative GM2 activator protein

Abstract

Onchocerca volvulus is the nematode pathogen responsible for human onchocerciasis also known as “River blindness”, a neglected tropical disease that affects up to 18 million people worldwide. Helminths Excretory Secretory Products (ESPs) constitute a rich repertoire of molecules that can be exploited for host-parasite relationship, diagnosis and vaccine studies. Here, we report, using a range of molecular techniques including PCR, western blot, recombinant DNA technology, ELISA, high performance thin-layer chromatography and mass spectrometry that the 28 KDa cysteine-rich protein (Ov28CRP) is a reliable component of the O . volvulus ESPs to address the biology of this parasite. We showed that (1) Ov28CRP is a putative ganglioside GM2 Activator Protein (GM2AP) conserved in nematode; (2) OvGM2AP gene is transcriptionally activated in all investigated stages of the parasitic life cycle, including larval and adult stages; (3) The full-length OvGM2AP was detected in in-vitro O . volvulus ESPs of adult and larval stages; (4) the mass expressed and purified recombinant OvGM2AP purified from insect cell culture medium was found to be glycosylated at asparagine 173 and lacked N-terminal signal peptide sequence; (5) the recombinant OvGM2AP discriminated serum samples of infected and uninfected individuals; (6) OvGM2AP competitively inhibits MUG degradation by recombinant β-hexosaminidase A but not MUGS, and could not hydrolyze the GM2 to GM3; (7) humoral immune responses to the recombinant OvGM2AP revealed a negative correlation with ivermectin treatment. Altogether, our findings suggest for the first time that OvGM2AP is an antigenic molecule whose biochemical and immunological features are important to gain more insight into our understanding of host-parasite relationship, as well as its function in parasite development at large.

In Silico Design and Validation of OvMANE1, a Chimeric Antigen for Human Onchocerciasis Diagnosis

“…Possibility of cross-reaction with OvMANE1 chimeric antigen in individuals infected with other related nematodes was investigated using serum samples from patients infected with Brugia malayi , Mansonella perstans , Ascaris lumbricoides and Wuchereria bancrofti by ELISA. In contrast to our previous observed cross-reaction of this set of serum samples with Ov28CRP/OvGM2AP, an excretory secretory product of O. volvulus [ 31 ] OvMANE1 chimeric antigen significantly discriminated onchocerciasis sera from that of related nematodes (see Figure 7 A).…”

Computational Design and Preliminary Serological Analysis of a Novel Multi-Epitope Vaccine Candidate Against Onchocerciasis and Related Filarial Diseases

“…Preventative and/or therapeutic NTD vaccines that will complement the other tools already in use are needed as long-term solutions [ 27 , 71 ]. Lustigman et al (2017) have reiterated the urgent need for a vaccine [ 2 ] to ensure achievement of the onchocerciasis elimination set by Expanded Special Project for Elimination of Neglected Tropical Diseases (ESPEN) [ 72 ]. Though many onchocerciasis vaccine candidates (including subunit and whole-parasite vaccines) have already been assessed in animal model studies) [ 10 , 51 , 73 , 74 , 75 ], none has, at the moment, been successfully translated for human use.…”

“…The infective larval stages of the parasite are transmitted through the repeated bites of infective black flies of the genus Simulium . These later give rise to adult worms that dwell in subcutaneous tissues in the human hosts where they can survive for up to 15 years (even under drug pressure), with adult female worms hatching approximately 1600 microfilariae daily [ 2 ]. Though infections were previously associated mainly with skin and eye lesions, recent records show a trend towards increased mortality.…”

The Functional Parasitic Worm Secretome: Mapping the Place of Onchocerca volvulus Excretory Secretory Products

“…However, ESPs could also play essential roles in ensuring cuticle integrity. As an example, GM2AP is a nematode ESP that was recently characterized in Onchocerca volvulus [ 56 ]. The GM2-activator protein was recently shown, for the first time in a nematode species, to be essential for cuticular integrity in Caenorhabditis elegans ( N ) (Njume et al, in preparation).…”