Development of nucleotide signatures for common poisonous organisms provides a new strategy for food poisoning diagnosis.

DNA barcoding is widely used in toxic species authentication, but due to serious DNA degradation of forensic materials, the application of full-length barcode sequences in food poisoning diagnosis is greatly limited. Nucleotide signature, a shorter specific molecular marker, derived from traditional DNA barcoding has been proposed as an emerging tool of toxic species detection in deeply processed materials. In this study, to resolve the frequent food poisoning accidents with unknown origin, we envisioned developing a nucleotide signature data set of common poisonous organisms and combining high-throughput sequencing (HTS) to reveal the poisoning cause. Ninety-three individuals and 1093 DNA barcode sequences of twelve common poisonous plants, fish, mushrooms and their related species were collected. Through sequence alignment and screening, the nucleotide signatures were respectively developed and validated as their specific molecular markers. The sequence length varied from 19 bp to 38 bp. These fragments were conserved within the same species or genera, and the specificity between related species has been also demonstrated. To further evaluate the application potential of nucleotide signature in forensic diagnosis, simulated forensic specimens (SFS) containing different poisonous ingredients were sequenced by HTS with PCR-free libraries. As a result, the nucleotide signature was successfully captured from original HTS data without assembly and annotation, accompanied by a high detection sensitivity of 0.1 ng/µl in mixture system. Therefore, this method was suitable for the assay of forensic materials with serious DNA degradation. The present study undoubtedly provides a new perspective and strong support for the detection of toxic ingredients and the diagnosis of food poisoning.