Periodic Reporting for period 2 - SpecSpiderVenom (Venomics of Prey-Specialised Spiders in the Evolutionary and Ecological Context)
Reporting period: 2024-04-01 to 2025-03-31
Venom is a trait that has evolved more than 100 times across various organisms. Venoms represent huge natural libraries of bioactive compounds that can be potentially utilized as drug leads or bio-pesticides. Yet, many venomous organisms are still understudied. Moreover, the venom toxins are usually investigated for their pharmacological activities, but the ecological context is often neglected. Spiders represent the most numerous terrestrial predators that utilize venom to paralyse their prey. Most spiders are generalists that feed on a broad range of prey. However, a small proportion of spiders are specialists that feed only on a single prey type, such as ants, termites, or other spiders. They utilise a plethora of morphological, behavioural and venomic adaptations to overcome their prey.
The aim of this project was to investigate venomic adaptations of prey-specialised spiders more thoroughly. Specifically, our aim was to elucidate the venom composition in selected specialists and the prey-specificity of selected toxins.
The aim of this project was to investigate venomic adaptations of prey-specialised spiders more thoroughly. Specifically, our aim was to elucidate the venom composition in selected specialists and the prey-specificity of selected toxins.
The outgoing phase of the project was carried out in the lab of Prof. Glenn King at the University of Queensland. Prof. King is a world expert on spider venoms, and members of his lab provided comprehensive expertise in the field of venomics. The lab was well-equipped to analyse the venom composition of selected prey-specialised spiders and to produce selected toxins for further activity and potency testing.
During the return phase, conducted in the lab of Prof. Stano Pekár at Masaryk University, Czechia, I continued the bioinformatic analysis of the obtained proteo-transcriptomic data using approaches I learned during the outgoing phase. Moreover, we began testing the efficacy of toxins produced during the outgoing phase. The knowledge gained in venomics allowed us start new project and develop ideas for future research projects.
Since the prey specificity of predatory venoms had not been comprehensively reviewed, we summarised the current knowledge on this topic in a review article. We evaluated existing hypotheses on the evolution of prey-specific venoms, with emphasis on the effects of restricted diets, prey defences, and prey resistance. We provided an overview of the prey specificity of crude venoms and prey-specific toxins across various venomous predators, with a focus on snakes, cone snails, and spiders—the most extensively studied groups. As evidence for venom prey specificity remains limited, we also outlined the best approaches and methods for its investigation, highlighted promising model groups, and discussed potential applications of prey-specific toxins. The review thus provides a comprehensive overview and serves as a guideline for future studies on venom prey specificity (Michálek et al., 2024).
We also highlighted the potential application of prey-specific toxin in another review paper focusing on the ecosystem services provided by spiders (Cardoso et al., 2024).
The main objective of the project was to analyse the venom composition of selected prey-specialised spiders. We focused on the proteomes of four spider species: three araneophagous species (Lampona sp., Palpimanus gibbulus, and P. cypreus) and one termitophagous species (Ammoxenus amphalodes). To analyse venom composition, we employed a combined proteo-transcriptomic approach.
White-tailed spiders of the genus Lampona have been wrongly associated with necrotic lesions after bites, although several studies have disproven this. Their primary prey are other spiders. Our previous study revealed that Lampona utilises several morphological and behavioural adaptations to capture spider prey. During this project, we focused on its venomic adaptations. We showed that its venom is more potent against spider prey than against alternative prey such as crickets. Venom composition analysis revealed 208 putative toxins, with smaller peptides (<10 kDa) being more abundant. Most lampotoxins belonged to two families characterised by unique scaffolds containing eight or ten cysteine residues. Several selected lampotoxins were produced, and we plan to further test their efficacy on various prey types (Michálek et al., 2022).
We also analysed the venom proteomes of two araneophagous Palpimanus species that are unrelated to Lampona. Data analysis is nearly complete, and the results will be published soon (Michálek, in prep.).
The sand-diving spider Ammoxenus amphalodes is the only known monophagous predator, feeding exclusively on a single termite species, Hodotermes mossambicus. Previous research revealed its unique predatory behaviour and rapid prey paralysis, which led us to investigate its venom in greater detail. We identified 116 putative toxins, with six ammoxotoxins (family 1) comprising over one-third of the venom proteome transcripts. These findings support the hypothesis that venoms of predators with highly specialised diets are dominated by a few structurally similar toxins. However, bioassays with selected recombinant peptides showed limited prey specificity, challenging our assumptions of venom specialization (Michálek, in prep.).
During the return phase, conducted in the lab of Prof. Stano Pekár at Masaryk University, Czechia, I continued the bioinformatic analysis of the obtained proteo-transcriptomic data using approaches I learned during the outgoing phase. Moreover, we began testing the efficacy of toxins produced during the outgoing phase. The knowledge gained in venomics allowed us start new project and develop ideas for future research projects.
Since the prey specificity of predatory venoms had not been comprehensively reviewed, we summarised the current knowledge on this topic in a review article. We evaluated existing hypotheses on the evolution of prey-specific venoms, with emphasis on the effects of restricted diets, prey defences, and prey resistance. We provided an overview of the prey specificity of crude venoms and prey-specific toxins across various venomous predators, with a focus on snakes, cone snails, and spiders—the most extensively studied groups. As evidence for venom prey specificity remains limited, we also outlined the best approaches and methods for its investigation, highlighted promising model groups, and discussed potential applications of prey-specific toxins. The review thus provides a comprehensive overview and serves as a guideline for future studies on venom prey specificity (Michálek et al., 2024).
We also highlighted the potential application of prey-specific toxin in another review paper focusing on the ecosystem services provided by spiders (Cardoso et al., 2024).
The main objective of the project was to analyse the venom composition of selected prey-specialised spiders. We focused on the proteomes of four spider species: three araneophagous species (Lampona sp., Palpimanus gibbulus, and P. cypreus) and one termitophagous species (Ammoxenus amphalodes). To analyse venom composition, we employed a combined proteo-transcriptomic approach.
White-tailed spiders of the genus Lampona have been wrongly associated with necrotic lesions after bites, although several studies have disproven this. Their primary prey are other spiders. Our previous study revealed that Lampona utilises several morphological and behavioural adaptations to capture spider prey. During this project, we focused on its venomic adaptations. We showed that its venom is more potent against spider prey than against alternative prey such as crickets. Venom composition analysis revealed 208 putative toxins, with smaller peptides (<10 kDa) being more abundant. Most lampotoxins belonged to two families characterised by unique scaffolds containing eight or ten cysteine residues. Several selected lampotoxins were produced, and we plan to further test their efficacy on various prey types (Michálek et al., 2022).
We also analysed the venom proteomes of two araneophagous Palpimanus species that are unrelated to Lampona. Data analysis is nearly complete, and the results will be published soon (Michálek, in prep.).
The sand-diving spider Ammoxenus amphalodes is the only known monophagous predator, feeding exclusively on a single termite species, Hodotermes mossambicus. Previous research revealed its unique predatory behaviour and rapid prey paralysis, which led us to investigate its venom in greater detail. We identified 116 putative toxins, with six ammoxotoxins (family 1) comprising over one-third of the venom proteome transcripts. These findings support the hypothesis that venoms of predators with highly specialised diets are dominated by a few structurally similar toxins. However, bioassays with selected recombinant peptides showed limited prey specificity, challenging our assumptions of venom specialization (Michálek, in prep.).
Our research elucidated the venom composition of several prey-specialised spider species. As most spider venoms remain uncharacterised, our data help to fill a significant gap in current knowledge. We found that, while these venoms contain tens to hundreds of components, they are typically dominated by a few families of putative toxins with intriguing structural scaffolds, characterised by numerous disulphide bridges typical of spider toxins. We hypothesise that these dominant, yet largely uncharacterised, toxins are responsible for the prey-specific potency observed in these venoms. However, sequence similarity to known toxins was generally low, and bioassays with selected recombinant peptides revealed only limited prey specificity. The activity, physiological roles, and ecological functions of these compounds therefore remain to be further investigated.