Our research is investigating the evolutionary factors shaping not only the pain-inducing defensive toxins in cobra venom, but also the parallel evolution ornate markings on their hoods, and the extremely bright warning colourings present in some species. 

We have found that the pain-inducing defensive toxins first evolved alongside the broad hoods that make cobras so distinctive. Further increases in the potency of the toxins subsequently occurred parallel to their warning strategies such as hood markings, body banding, red colouring and spitting. Their spectacular hoods and eye-catching patterns evolved to warn off potential predators because unlike other snakes, which use their venom purely for predation, cobras also use it in defence.

Typically it has been thought that only spitting cobras had these defensive toxins in high amounts in their venoms, however we’ve shown that this toxin type evolved with the ability to hood in the common ancestor of Hemachatus and Naja. Further increases were linked to the evolution of ornate patterns as well as the three separate evolutions of spitting (Hemachatus, once in African Naja and again in Asian Naja). The pain inducting toxin type in Hemachatus and Naja are modified three finger toxins that have switched from the ancestral toxic effect of antagonistic binding of post-synaptic nicotinic acetylcholine receptors to the pain-inducing cytotoxicity. This toxin class is small enough to be absorbable in the eyes and cause immediate pain there. Thus there was a selection pressure for the evolution of spitting.

King cobras (Ophiophagus genus) are genetically distinct from other 'cobras' and convergently evolved their hooding ability. Parallel to this they convergently evolved pain-inducing defensive toxins. While the function is the same (pain) the convergence reflected by differences in the underlying mechanics, with king cobras using L-amino acid oxidase enzymes to accomplish the same outcome. These toxins are far too large to be readily evolved in the eyes, and there there was not a convergent selection pressure for the evolution of spitting. However, like in Hemachatus and Naja, there is a pattern in that the most conspicuously marked populations have the highest levels of these defensive toxins. In the case of the kind cobras, it is the Malaysian population with its bright orange hood markings that that contains the highest amounts of the pain inducing toxins. 

Secondary losses are as evolutionarily interesting as secondary increases. Within Naja, the water cobra has secondarily dramatically reduced the hood as the elongate ribs would impair swim performance, and thus there was a selection pressure for their reduction. As these species do not do the hood displays like other cobras, there has also been a secondary loss of the defensive toxins. In other cobras, some species have secondarily lost the conspicuous hood markings (such as Naja oxiana and Naja philippinensis) and the venoms are similarly much reduced in the relative content of the defensive toxins.

 

Ancestral state reconstructions of hooding, spitting, and cytotoxicity; based on ambiguous Aspidelaps species coded as non-hooding as well as the ambiguous N. atra and N. kaouthia also coded as non-spitting. Reconstruction over branches represents the AUC for the non-transformed NFF cell line (left) and the melanoma (MM96L) cancer cell line (right), where warmer colours represent higher cytotoxicity against cell lines (raw data in Supplementary Tables). Pie charts are the same on both trees and represent estimates of ancestral states for hooding (above branch) and spitting (below branch) where black and white represent the trait being present or absent respectively. States at tips represent the data collected.  CLICk here to download the associated paper

Ancestral state reconstructions of hooding, spitting, and cytotoxicity; based on ambiguous Aspidelaps species coded as non-hooding as well as the ambiguous N. atra and N. kaouthia also coded as non-spitting. Reconstruction over branches represents the AUC for the non-transformed NFF cell line (left) and the melanoma (MM96L) cancer cell line (right), where warmer colours represent higher cytotoxicity against cell lines (raw data in Supplementary Tables). Pie charts are the same on both trees and represent estimates of ancestral states for hooding (above branch) and spitting (below branch) where black and white represent the trait being present or absent respectively. States at tips represent the data collected.  CLICk here to download the associated paper

Relative degree of aposematic hood marking between (A,B) African (Naja haje) and (C,D) Asian (Naja naja) cobras with their higher levels of cytotoxicity;. Similar patterning to N. haje are seen in the the basally coloured African spitting cobras lacking aposematic marking like N. haje (E) N. nubiae and (F) N. ashei. Convergent reversal from aposematic markings to the basal drab coloured state accompanied by a lowering of cytotoxicity has occurred in (G) the Asian non‐spitting cobra N. oxiana and (H) the spitting cobra N. phillipinensis. The Pakistan Sindh desert population of N. naja displays the aposematic hood marking as juveniles and subadults but not (I) as adults, without any loss of cytotoxicity. Photos: (A,B) Arno Naude; (C,D) Gowri Mallapur; (E) HG Hjim; (F) Anothony Childs; (G,H) Randy Ciuros; (I) Bryan Fry.  CLICk here to download the associated paper

Relative degree of aposematic hood marking between (A,B) African (Naja haje) and (C,D) Asian (Naja naja) cobras with their higher levels of cytotoxicity;. Similar patterning to N. haje are seen in the the basally coloured African spitting cobras lacking aposematic marking like N. haje (E) N. nubiae and (F) N. ashei. Convergent reversal from aposematic markings to the basal drab coloured state accompanied by a lowering of cytotoxicity has occurred in (G) the Asian non‐spitting cobra N. oxiana and (H) the spitting cobra N. phillipinensis. The Pakistan Sindh desert population of N. naja displays the aposematic hood marking as juveniles and subadults but not (I) as adults, without any loss of cytotoxicity. Photos: (A,B) Arno Naude; (C,D) Gowri Mallapur; (E) HG Hjim; (F) Anothony Childs; (G,H) Randy Ciuros; (I) Bryan Fry.  CLICk here to download the associated paper

Convergent aposematic banding in the strongly cytotoxic species (A) Naja annulifera; (B) Hemachatus haemachatus; (C) Naja nigricincta; (D) Naja siamensis; and (E) Ophiophagus hannah; Photos by (A,D) Randy Ciuros; (B) Giuseppe Mazza; (C,E) Tom Charlton. CLICk here to download the associated paper

Convergent aposematic banding in the strongly cytotoxic species (A) Naja annulifera; (B) Hemachatus haemachatus; (C) Naja nigricincta; (D) Naja siamensis; and (E) Ophiophagus hannah; Photos by (A,D) Randy Ciuros; (B) Giuseppe Mazza; (C,E) Tom Charlton. CLICk here to download the associated paper

Convergent aposematic hood colouring in the African spitting cobras such as (A) Naja katiensis; (B) Naja pallida; (C) Naja mossambica; (D) Naja nigricollis; and (E) convergently in the adult colouring in the Malaysian population of Ophiophagus hannah (the most cytotoxic O. hannah population). Photos (A) Stephen Sprawls, (B) Wikimedia Commons; (C,D) Randy Ciuros; (E) Kevin Messenger. CLICk here to download the associated paper

Convergent aposematic hood colouring in the African spitting cobras such as (A) Naja katiensis; (B) Naja pallida; (C) Naja mossambica; (D) Naja nigricollis; and (E) convergently in the adult colouring in the Malaysian population of Ophiophagus hannah (the most cytotoxic O. hannah population). Photos (A) Stephen Sprawls, (B) Wikimedia Commons; (C,D) Randy Ciuros; (E) Kevin Messenger. CLICk here to download the associated paper

Disruptive camouflage patterning in aquatic snakes (A,B) Naja annulata which also has a secondarily extremely reduced hood and also secondarily lost its cytotoxicity; (C) Laticauda colubrina; (D) Emydocephalus annulatus; (E) Eunectes murinus and fish (F) Cichla orinocensis; (G) Salmo trutta. Photos (A) Markus Oulehla; (B,D) Wikimedia Commons; (C) Jan Messersmith; (E) Rhett A. Butler; (F) Ivan Mikolji; (G) Phil Skinner.  CLICk here to download the associated paper

Disruptive camouflage patterning in aquatic snakes (A,B) Naja annulata which also has a secondarily extremely reduced hood and also secondarily lost its cytotoxicity; (C) Laticauda colubrina; (D) Emydocephalus annulatus; (E) Eunectes murinus and fish (F) Cichla orinocensis; (G) Salmo trutta. Photos (A) Markus Oulehla; (B,D) Wikimedia Commons; (C) Jan Messersmith; (E) Rhett A. Butler; (F) Ivan Mikolji; (G) Phil Skinner.  CLICk here to download the associated paper