Snake venom is an ecologically relevant functional trait directly linked with a snake’s fitness and survival, facilitating predation and defence. Snake venom variation occurs at all taxonomic levels, but the study at the intraspecific level is still in its early stages. The common adder (Vipera berus) exhibits considerable variation in colour phenotypes across its distribution range. Melanistic (fully black) individuals are the subject of myths and fairytales, and in German folklore such ‘hell adders’ are considered more toxic than their normally coloured conspecifics despite any formal investigation. Here, we provide the first comparative analysis of venoms from melanistic and normally coloured common adders. Specifically, we compared the venom profiles by sodium dodecylsulfate polyacrylamide gel electrophoresis and reversed-phase high-performance liquid chromatography and tested the venoms’ protease, phospholipase A2 and cytotoxic activities. Phospholipase A2 activity was similar in both phenotypes, whereas general protease activity was higher in the melanistic venom, which was also more cytotoxic at two concentrations (6.25 and 12.5 µg ml−1). These minor differences between the venoms of melanistic and normally coloured adders are unlikely to be of clinical relevance in the context of human envenomation. In light of our results, the claim that melanistic adders produce more toxic venom than their normally coloured conspecifics appears rooted entirely in folklore.

Supplementary Files for the wasp spider Venomics project: S1: Docker Images combined for bioinformatic analysis S2: 2D SDS PAGE Gel used for interpretation and MALDI MS (Zoom in for Spot assignments) S3: Mascot output for MALDI MS S4: Mascot output for nanoLC ESI MS

Supplementary Files for the wasp spider Venomics project: (An image of the species (paper5.jpg for visualisation) has been added to the supplementary files) Figure S1: 2D SDS PAGE Gel used for interpretation and MALDI MS (Zoom in for Spot assignments) Table S1: Docker Images combined for bioinformatic analysis Table S2: Mascot output for MALDI MS Table S3: Mascot output for nanoLC ESI MS Table S4: Annotated Proteome Hits incl Sequences Table S5: Annotated sequences from Araneus ventricosus

Supplementary Files for the wasp spider Venomics project: S1: Docker Images combined for bioinformatic analysis S2: 2D SDS PAGE Gel used for interpretation and MALDI MS (Zoom in for Spot assignments) S3: Mascot output for MALDI MS S4: Mascot output for nanoLC ESI MS S5: Annotated Proteome Hits incl Sequences

Supplementary Files for the wasp spider Venomics project: Figure S1: 2D SDS PAGE Gel used for interpretation and MALDI MS (Zoom in for Spot assignments)Table S1: Docker Images combined for bioinformatic analysis Table S2: Mascot output for MALDI MS Table S3: Mascot output for nanoLC ESI MS Table S4: Annotated Proteome Hits incl Sequences Table S5: Annotated sequences from Araneus ventricosus

Supplementary Files for the wasp spider Venomics project: (An image of the species (paper5.jpg for visualisation) has been added to the supplementary files) Figure S1: 2D SDS PAGE Gel used for interpretation and MALDI MS (Zoom in for Spot assignments) Table S1: Docker Images combined for bioinformatic analysis Table S2: Mascot output for MALDI MS Table S3: Mascot output for nanoLC ESI MS Table S4: Annotated Proteome Hits incl Sequences Table S5: Annotated sequences from Araneus ventricosus