Snake Venom Proteomics of Samar Cobra (Naja samarensis) from the Southern Philippines: Short Alpha-Neurotoxins as the Dominant Lethal Component Weakly Cross-Neutralized by the Philippine Cobra Antivenom

Abstract

The Samar Cobra, Naja samarensis, is endemic to the southern Philippines and is a WHO-listed Category 1 venomous snake species of medical importance. Envenomation caused by N. samarensis results in neurotoxicity, while there is no species-specific antivenom available for its treatment. The composition and neutralization of N. samarensis venom remain largely unknown to date. This study thus aimed to investigate the venom proteome of N. samarensis for a comprehensive profiling of the venom composition, and to examine the immunorecognition as well as neutralization of its toxins by a hetero-specific antivenom. Applying C18 reverse-phase high-performance liquid chromatography (RP-HPLC) and tandem mass spectrometry (LC-MS/MS), three-finger toxins (3FTx) were shown to dominate the venom proteome by 90.48% of total venom proteins. Other proteins in the venom comprised snake venom metalloproteinases, phospholipases A2, cysteine-rich secretory proteins, venom nerve growth factors, L-amino acid oxidases and vespryn, which were present at much lower abundances. Among all, short-chain alpha-neurotoxins (SαNTX) were the most highly expressed toxin within 3FTx family, constituting 65.87% of the total venom proteins. The SαNTX is the sole neurotoxic component of the venom and has an intravenous median lethal dose (LD50) of 0.18 μg/g in mice. The high abundance and low LD50 support the potent lethal activity of N. samarensis venom. The hetero-specific antivenom, Philippine Cobra Antivenom (PCAV, raised against Naja philippinensis) were immunoreactive toward the venom and its protein fractions, including the principal SαNTX. In efficacy study, PCAV was able to cross-neutralize the lethality of SαNTX albeit the effect was weak with a low potency of 0.20 mg/ml (defined as the amount of toxin completely neutralized per milliliter of the antivenom). With a volume of 5 ml, each vial of PCAV may cross-neutralize approximately 1 mg of the toxin in vivo. The findings support the potential para-specific use of PCAV in treating envenomation caused by N. samarensis while underscoring the need to improve the potency of its neutralization activity, especially against the highly lethal alpha-neurotoxins.

Inhibition of bacterial biofilms by the snake venom proteome

“…Our data for 3FTx in the N. samarensis venom proteome contrasts sharply with the results obtained by Palasuberniam et al. [65] which revealed a high content of 3FTx (90.5% of total venom) compared to the 15.2% 3FTx detected in this study. There were similar relative percentages of snake venom metalloproteinases (4.2% vs 7.4%), phospholipase A2 (3.8% vs 2.8%), cysteine-rich secretory protein (1.1% vs 2.3%), L -amino acid oxidase (0.3% vs 0.5%), venom nerve growth factor (0.1% vs 0.5%) and vespryn (0.1% vs 0.5%) in the two studies.…”

An immunoinformatic approach to assessing the immunogenic capacity of alpha-neurotoxins in elapid snake venoms

“…This is likely due to most viperid venoms are predominated with high MW toxins (MW > 25 kDa) (e.g., SVMP and snake venom serine protease, SVSP) ( Offor et al, 2022 ), which are capable of eliciting stronger immune response for the production of neutralizing antibodies. However, given that venoms constituting neurotoxins of more than ∼40% failed to raise antivenoms with major improvement in neutralization efficacy ( Tan et al, 2015b ; Tan et al, 2016a ; Tan et al, 2019a ; Palasuberniam et al, 2021 ), it is therefore suggested that the low neutralizing efficacy of elapid antivenom may be also attributed to other factors, such as amino acid composition, that intrinsically modulate the epitope configuration of α-NTXs to be recognized by T-cell receptor (TCR) to elicit a strong humoral response.…”

“…The resultant antisera showed an exceptionally broad para-specificity of neutralization against diverse elapid venoms, while the strategy did not translate into a major improvement of neutralizing potency per individual snake species ( Ratanabanangkoon et al, 2016 ; Ratanabanangkoon et al, 2020 ). Furthermore, even antivenoms raised against elapid venoms that contain abundant α-NTX (>40% of total venom proteins) showed generally low neutralizing potency, questioning if the “dose” of α-NTX is indeed the key determinant of eliciting good immunization response ( Ratanabanangkoon et al, 2016 ; Palasuberniam et al, 2021 ; Tan et al, 2021 ). Hypothetically, there are other factors at play that intrinsically modulate the makeup of epitope(s) in the α-NTX, and this results in variable immunorecognition by the B-cell receptor (BCR) and/or T-cell receptor (TCR) for antibody production.…”

Isolation and Characterization of Two Postsynaptic Neurotoxins From Indian Cobra (Naja Naja) Venom

“…This may explain the successful treatment of bites from Asiatic cobras, which have large quantities of long-chain α-neurotoxins in their venoms ( Lagraulet, 1984 ; Veto et al, 2007 ). Interestingly, a recent study has shown that the venom of the Samar cobra ( N. samarensis ) consists of approximately 66% short-chain α-neurotoxins, no long-chain α-neurotoxins were identified, and envenoming by this species causes paralysis in humans ( Palasuberniam et al, 2021 ). This is an extremely high quantity of short-chain α-neurotoxins, and supports the hypothesis that short-chain toxins only cause envenoming in humans if administered in large quantities ( Silva et al, 2018 ).…”