Excretory-secretory products from the brown stomach worm, Teladorsagia circumcincta, exert antimicrobial activity in in vitro growth assays

Abstract

Background Over the past decade, evidence has emerged of the ability of gastrointestinal (GI) helminth parasites to alter the composition of the host gut microbiome; however, the mechanism(s) underpinning such interactions remain unclear. In the current study, we (i) undertake proteomic analyses of the excretory-secretory products (ESPs), including secreted extracellular vesicles (EVs), of the ‘brown stomach worm’ Teladorsagia circumcincta, one of the major agents causing parasite gastroenteritis in temperate areas worldwide; (ii) conduct bioinformatic analyses to identify and characterise antimicrobial peptides (AMPs) with putative antimicrobial activity; and (iii) assess the bactericidal and/or bacteriostatic properties of T. circumcincta EVs, and whole and EV-depleted ESPs, using bacterial growth inhibition assays. Methods Size-exclusion chromatography was applied to the isolation of EVs from whole T. circumcincta ESPs, followed by EV characterisation via nanoparticle tracking analysis and transmission electron microscopy. Proteomic analysis of EVs and EV-depleted ESPs was conducted using liquid chromatography-tandem mass spectrometry, and prediction of putative AMPs was performed using available online tools. The antimicrobial activities of T. circumcincta EVs and of whole and EV-depleted ESPs against Escherichia coli were evaluated using bacterial growth inhibition assays. Results Several molecules with putative antimicrobial activity were identified in both EVs and EV-depleted ESPs from adult T. circumcincta. Whilst exposure of E. coli to whole ESPs resulted in a significant reduction of colony-forming units over 3 h, bacterial growth was not reduced following exposure to worm EVs or EV-depleted ESPs. Conclusions Our data points towards a bactericidal and/or bacteriostatic function of T. circumcincta ESPs, likely mediated by molecules with antimicrobial activity. Graphical Abstract

Helminthic host defense peptides: using the parasite to defend the host

“…Another interesting study carried out in the nematode Teladorsagia circumcincta combined proteomics and bioinformatics screening to detect antimicrobial ESP in secreted EVs. Although these peptides were not individually tested, the authors were able to demonstrate the antimicrobial activity carried by EVs and ESP, validating the reliability of their approach [66]. Despite these encouraging outputs, the computer-assisted identification of novel HDP in helminths will remain a challenging task for the years to come, due to the variable quality of available helminth genome assemblies, the different technical approaches used for sequencing and assembly, the inherent complexity of these genomes and the risk of exogenous contamination [67].…”

Structural and functional analyses of nematode-derived antimicrobial peptides support the occurrence of direct mechanisms of worm-microbiota interactions

“…Indeed, A. suum ESPs contain several antimicrobial molecules, including cecropin-P2 [25] , A. suum antibacterial factor (ASABF), lysozymes, as well as C-type lectin domain-containing proteins [5] , that can disrupt bacterial membranes to cause cell lysis, or promote lethal agglutination of bacterial cells. More recently, we also demonstrated that in vitro exposure of E. coli to ESPs from the ‘brown stomach worm’, Teladorsagia circumcincta , an abomasal parasite of small ruminants, results in significant reductions of colony forming units over a 3-h period [24] . Proteomics analyses of T. circumcincta ESPs, coupled with bioinformatics screening of amino acid sequence data, identified several putative AMPs, including a histone H4-like protein, an Ancylostoma -secreted protein (ASP)-like protein, and an invertebrate-specific lysozyme-like enzyme known as destabilase [24] .…”

“…More recently, we also demonstrated that in vitro exposure of E. coli to ESPs from the ‘brown stomach worm’, Teladorsagia circumcincta , an abomasal parasite of small ruminants, results in significant reductions of colony forming units over a 3-h period [24] . Proteomics analyses of T. circumcincta ESPs, coupled with bioinformatics screening of amino acid sequence data, identified several putative AMPs, including a histone H4-like protein, an Ancylostoma -secreted protein (ASP)-like protein, and an invertebrate-specific lysozyme-like enzyme known as destabilase [24] . Previously, we hypothesised that the antimicrobial properties of ESPs from T. circumcincta , as well as of ESPs from other GI helminths of humans and animals, may be attributed to the activity of secreted AMPs, free and/or encapsulated in secreted extracellular vesicles (EVs) [24] .…”

“…Until recently, the vast majority of host-parasite interaction studies have focussed on the ability of helminths to communicate with and modulate the immune system of the host, particularly through the activity of their excretory/secretory products (ESPs) [21] , [22] . Intriguingly, recent evidence points towards a role of helminth ESPs in worm-microbiota interactions, namely by exerting antimicrobial activity [5] , [23] , [24] . For instance, ESPs of the swine roundworm, Ascaris suum, have been shown to exert significant bactericidal activity against Escherichia coli , Staphylococcus aureus, and Salmonella typhimurium , and to disrupt biofilm formations of E. coli in vitro [5] .…”

A dominance of Mu class glutathione transferases within the equine tapeworm Anoplocephala perfoliata

“…For 2D SDS PAGE protein spots, the method of Duncan et al (2018) was utilized for LC-MSMS. For whole A. perfoliata Orbitrap Fusion™ Tribrid™ mass spectrometer (Thermo Scientific™), with EASY-Spray™ Source, coupled to an UltiMate™ 3000 RSLCnano System (Thermo Scientific™) as per methods of Rooney et al (2022).…”