Abstract

BackgroundBioassessment and biomonitoring of meat products are aimed at identifying and quantifying adulterants and contaminants, such as meat from unexpected sources and microbes. Several methods for determining the biological composition of mixed samples have been used, including metabarcoding, metagenomics and mitochondrial metagenomics. In this study, we aimed to develop a method based on next-generation DNA sequencing to estimate samples that might contain meat from 15 mammalian and avian species that are commonly related to meat bioassessment and biomonitoring.ResultsIn this project, we found the meat composition from 15 species could not be identified with the metabarcoding approach because of the lack of universal primers or insufficient discrimination power. Consequently, we developed and evaluated a meat mitochondrial metagenomics (3MG) method. The 3MG method has four steps: (1) extraction of sequencing reads from mitochondrial genomes (mitogenomes); (2) assembly of mitogenomes; (3) mapping of mitochondrial reads to the assembled mitogenomes; and (4) biomass estimation based on the number of uniquely mapped reads. The method was implemented in a python script called 3MG. The analysis of simulated datasets showed that the method can determine contaminant composition at a proportion of 2% and the relative error was < 5%. To evaluate the performance of 3MG, we constructed and analysed mixed samples derived from 15 animal species in equal mass. Then, we constructed and analysed mixed samples derived from two animal species (pork and chicken) in different ratios. DNAs were extracted and used in constructing 21 libraries for next-generation sequencing. The analysis of the 15 species mix with the method showed the successful identification of 12 of the 15 (80%) animal species tested. The analysis of the mixed samples of the two species revealed correlation coefficients of 0.98 for pork and 0.98 for chicken between the number of uniquely mapped reads and the mass proportion.ConclusionTo the best of our knowledge, this study is the first to demonstrate the potential of the non-targeted 3MG method as a tool for accurately estimating biomass in meat mix samples. The method has potential broad applications in meat product safety.

Highlights

  • Bioassessment and biomonitoring of meat products are aimed at identifying and quantifying adulter‐ ants and contaminants, such as meat from unexpected sources and microbes

  • The results showed that it can accurately determine the biological composition of meat mix samples and accurately estimate biomass

  • Evaluation of the metabarcoding method for the 15 mammalian and avian species To determine whether the mixture containing 15 species can be identified using metabarcoding, we analysed the availability of universal primers and the ability of their amplified products to distinguish the 15 species

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Summary

Introduction

Bioassessment and biomonitoring of meat products are aimed at identifying and quantifying adulter‐ ants and contaminants, such as meat from unexpected sources and microbes. We aimed to develop a method based on next-generation DNA sequencing to estimate samples that might contain meat from 15 mammalian and avian species that are commonly related to meat bioassessment and biomonitoring. Two of the nine beef samples examined by Erol et al contained horse and deer meat [3]. Such adulteration harms consumers’ rights and interests [4] and disrupts market order [5]. Identifying adulterated ingredients in meat and meat products is essential

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