Investigation of genes coding for potential tetrodotoxin binding proteins in Canthigaster valentini and Taricha granulosa

Tetrodotoxin (TTX) is a powerful neurotoxin that selectively binds to voltage gated sodium channels in excitable tissues causing paralysis, asphyxiation, and death. TTX is found in a variety of unrelated taxa where multiple species have been reported to produce TTX binding proteins. Pufferfish in the Tetraodontidae family (mainly Takifugu spp.) have been demonstrated to accumulate high TTX concentrations however, the presence of TTX is still unknown for many pufferfish species. Canthigaster valentini is pufferfish that is thought to be toxic but has yet to be assayed for the presence of TTX. Other species such as Taricha granulosa are also capable of accumulating high concentrations of TTX however, the origin of TTX in this species is still debated. This study has multiple objectives i) determine if TTX is present in C. valentini liver tissues using CIEIA analysis, ii) identify possible TTX binding protein genes in C. valentini using PCR analysis with TTX binding protein gene specific primers, iii) identify possible TTX binding protein genes in T. granulosa using PCR analysis with TTX binding protein gene specific primers, and iv) to identify bacterial flora present on skin, intestine, liver, and ovary tissues using PCR analysis with 16S rDNA specific primers. In this study, the first instance of TTX identification and quantification in C. valentini is reported. Also, BLAST analysis of amplified products identified gene fragments similar to PSTBPs found in Takifugu spp. and their ancestral TBT-bp2 genes in pufferfish. Here, it is suggested that C. valentini may possess TTX binding proteins similar to those found in Takifugu spp. pufferfish. PCR amplification of T. granulosa DNA using TTX binding protein gene specific primers produced an amplicon of 615 bp however, BLAST analysis did not produce a significant alignment. Although bacterial DNA was amplified from total DNA extracts from T. granulosa tissues, metagenomic analysis did not produce data.