Meat Adulteration Research Articles

Integrating microneedle DNA extraction to hand-held microfluidic colorimetric LAMP chip system for meat adulteration detection

Most microfluidic-based “sample-in-result-out” systems suffer sophisticated microfluidic production processes, high-cost chips, and expensive instruments. They cannot be used in the meat market as well as farmer's markets in rural areas. Here, we developed a hand-held microfluidic chip system for on-site meat species qualitative authentication detection which integrated a simple microneedle DNA extraction and a visual loop-mediated isothermal amplification (LAMP). The chip can be used by easily pricking meat samples, simply hand-shaking the chip, and readily available isothermal heating instead of a complicated DNA extraction process and microfluidic control device. The system demonstrates high specificity and sensitivity for selected six species of meat samples and low to 1% simulated adulteration could be detected within 60 min. Besides, the whole cost was less than 1 dollar. The integrated hand-held microfluidic detection system offers a simple, fast, low-cost “sample-in-result-out” point-of-care device which could be extended to medical diagnosis and animal/plant disease identification.

A Low-Cost Closed-Tube Method for Detection of Adulteration in Ground Meat

ABSTRACT The increased public concern about adulteration and mislabelling of meat products has fueled a need for the development of fast, reliable, and cost-effective species detection methods. Hence, we developed a high resolution melt analysis (HRMA) for the discrimination of meat species commonly used in the meat industry and their possible adulterants. A universal primer pair spanning a ⁓238 bp fragment of mitochondrial 16S rRNA gene was designed to perform HRMA. The developed workflow was validated on standard meat samples, ternary meat mixtures of eight different species, and 56 commercial meat samples. Distinct melting profiles were generated for each species, several ternary meat mixtures, and commercial food samples. The assay was sufficiently sensitive to detect 0.003 ng/µl of DNA of every targeted species. In short, the developed HRM analysis is a rapid, cost-effective, and efficient species discrimination system for confirmation of meat origin.

Infrared Spectroscopy-Based Chemometric Analysis for Lard Differentiation in Meat Samples

One of the most pressing concerns for the consumer market is the detection of adulteration in meat products due to their preciousness. The rapid and accurate identification mechanism for lard adulteration in meat products is highly necessary, for developing a mechanism trusted by consumers and that can be used to make a definitive diagnosis. Fourier Transform Infrared Spectroscopy (FTIR) is used in this work to identify lard adulteration in cow, lamb, and chicken samples. A simplified extraction method was implied to obtain the lipids from pure and adulterated meat. Adulterated samples were obtained by mixing lard with chicken, lamb, and beef with different concentrations (10%–50% v/v). Principal component analysis (PCA) and partial least square (PLS) were used to develop a calibration model at 800–3500 cm⁻¹. Three-dimension PCA was successfully used by dividing the spectrum in three regions to classify lard meat adulteration in chicken, lamb, and beef samples. The corresponding FTIR peaks for the lard have been observed at 1159.6, 1743.4, 2853.1, and 2922.5 cm⁻¹, which differentiate chicken, lamb, and beef samples. The wavenumbers offer the highest determination coefficient R² value of 0.846 and lowest root mean square error of calibration (RMSEC) and root mean square error prediction (RMSEP) with an accuracy of 84.6%. Even the tiniest fat adulteration up to 10% can be reliably discovered using this methodology.

Modified, innovative method for detection of adulteration of meat pro­ducts with meat of other species

The aim of this study was to detect illegal adulteration of beef meat products with meat from other species. Samples (n=120) of industrial and handmade beef products were randomly collected from retail outlets in Assiut city, Egypt: raw beef burger, oriental beef sausage, beef kofta, and beef luncheon (30 samples each). Samples were analysed using agar gel immunodiffusion (AGID) and modified AGID (MAGID) assays. Results showed that 17.5% of examined products were adulterated with chicken meat. MAGID detected that 14.1% of samples were adulterated with donkey meat, whereas all AGID results were negative. Human tissue was detected in 8.3% (AGID) and 10% (MAGID) of examined samples. Histological examination was then used to detect foreign tissue, and all categories of products were found to be adulterated, and some of them - contaminated with human blood cells. Polymerase chain reaction analysis confirmed that MAGID was more accurate and sensitive than AGID, especially for false negative AGID results. Consumers are advised not to consume too much of the studied meat products to avoid exposure to adulterated or contaminated products that might constitute a health hazard.

Design and development of a rapid meat detection system based on RPA-CRISPR/Cas12a-LFD

In recent years, meat adulteration safety incidents have occurred frequently, triggering widespread attention and discussion. Although there are a variety of meat quality identification methods, conventional assays require high standards for personnel and experimental conditions and are not suitable for on-site testing. Therefore, there is an urgent need for a rapid, sensitive, high specificity and high sensitivity on-site meat detection method. This study is the first to apply RPA combined with CRISPR/Cas12a technology to the field of multiple meat identification. The system developed by parameter optimization can achieve specific detection of chicken, duck, beef, pork and lamb with a minimum target sequence copy number as low as 1 × 100 copies/μL for 60 min at a constant temperature. LFD test results can be directly observed with the naked eye, with the characteristics of fast, portable and simple operation, which is extremely in line with current needs. In conclusion, the meat identification RPA-CRISPR/Cas12a-LFD system established in this study has shown promising applications in the field of meat detection, with a profound impact on meat quality, and provides a model for other food safety control programs.

Forensic analysis of raw meat adulteration using mtDNA

Meat species identification has become essential with the increasing events of frauds like the illegal slaughter of cows, meat adulteration, and substitution. Food scam directly influences public well-being, trade, and wildlife. In Pakistan, donkey meat is used as adulterants for cow meat and is considered Haram in Islamic concepts. In this study PCR, based detection methods are used for identification purposes. The mitochondrial gene cytochrome b has been used in this study to identify the origin of meat specie. Specie-specific primers of cyt b of cow and donkey were used for identification. DNA from different binary ratios of cow and donkey meat was extracted by the phenol-chloroform method. Ratios were made from 1-10 and extracted DNA was subjected to PCR to amplify the target fragment of the cyt b gene. Primers were sensitive to identifying species origin in all meat ratios. Multiplex PCR was designed to identify both species and the results were analyzed by gel electrophoresis. Fragment size of 309bp for cow and 475bp for donkey was observed. Results of the current study conclude that PCR assays, including multiplex PCR, is efficient and has a high sensitivity for even small amount of meat. It is concluded that multiplex PCR is useful and reliable for adulterated meat detection.

Application of biosensors for detection of meat species: A short review

Biosensors are easy-to-use tools that have made many advances in recent years. In the meat industry, most of these devices are applied in pathogen detection or freshness analysis. However, they are also applicable in the investigation of adulteration in meat by mixtures of different animal species, since the authenticity of food components is a serious problem involving health, economic, legal and religious concerns. This review aimed to summarize the biosensors developed for this purpose and how they manage to detect different meat species in complex mixtures. Only 41 studies were found, the majority involving optical biosensors, from which 63% aimed the detection of pork in meat samples. However, for an appropriate sensitivity, 81% of the proposed devices required extensive purification and/or amplification steps of the samples, unsuitable to point-of-need use. In view of this and the absence of studies on the stability of the platforms over time, it is necessary to develop more robust devices that can be used at any stage of the meat industrial chain and even by the final consumer.

Heat-Treated Meat Origin Tracing and Authenticity through a Practical Multiplex Polymerase Chain Reaction Approach.

Meat adulteration have become a global issue, which has increasingly raised concerns due to not only economic losses and religious issues, but also public safety and its negative effects on human health. Using optimal primers for seven target species, a multiplex PCR method was developed for the molecular authentication of camel, cattle, dog, pig, chicken, sheep and duck in one tube reaction. Species-specific amplification from the premixed total DNA of seven species was corroborated by DNA sequencing. The limit of detection (LOD) is as low as 0.025 ng DNA for the simultaneous identification of seven species in both raw and heat-processed meat or target meat: as little as 0.1% (w/w) of the total meat weight. This method is strongly reproducible even while exposed to intensively heat-processed meat and meat mixtures, which renders it able to trace meat origins in real-world foodstuffs based on the authenticity assessment of commercial meat samples. Therefore, this method is a powerful tool for the inspection of meat adulterants and has broad application prospects.

CRISPR/Cas Precisely Regulated DNA-Templated Silver Nanocluster Fluorescence Sensor for Meat Adulteration Detection.

Meat adulteration can cause consumer fraud, food allergies, and religious issues. Rapid and sensitive detection methods are urgently demanded to supervise meat authenticity. Herein, a clustered regularly interspaced short palindromic repeats (CRISPR)/Cas precisely regulated DNA-templated silver nanocluster (DNA-AgNC) sensor was ingeniously designed to detect meat adulteration. Specific sequence recognition of CRISPR/Cas12a allowed accurate identification of target DNA. The emerging label-free fluorescent probes, DNA-AgNCs, a class of promising fluorophores in biochemical analysis with attractive photostability and remarkably enhanced fluorescence properties, were first introduced as the substrates of CRISPR/Cas12a system, allowing a sensitive output of amplified signals through the precise regulation of the unique target DNA-activated trans-cleavage activity of Cas12a. Based on this specific recognition, efficient signal transduction of CRISPR/Cas12a, and the outstanding fluorescence properties of DNA-AgNCs, the proposed strategy achieved a satisfactory linear range from 10 pM to 1 μM with a limit of detection (LOD) as low as 1.9 pM, which can achieve sensitive detection of meat adulteration.

Rapid solid phase microextraction of DNA using mesoporous metal-organic framework coating for PCR-based identification of meat adulteration.

A rapid, convenient, low-cost, and selective DNA isolation method was developed for identifying meat adulteration. A mesoporous metal organic framework (Meso-UIO-66)-coated solid phase microextraction system was employed as an isolation device to simplify DNA isolation into three steps (lysis, washing, and elution). Meso-UIO-66 was utilized as the adsorbent because of its positively charged surface, high chemical stability, and mesoporous structure. Meso-UIO-66 was characterized by scanning electron microscopy, transmission electron microscopy, powder X-ray diffraction, Fourier transform infrared spectroscopy, ultraviolet‒visible spectroscopy, X-ray photoelectron spectroscopy, and nitrogen adsorption-desorption tests. Parameters that affected DNA isolation were optimized. This method can be used to isolate and purify DNA from meat in 60s, and the DNA concentration and purity are comparable to those of samples isolated with a commercial kit. Multiple DNA detection was achieved by coupling this method with the multiplex polymerase chain reaction technique, and the detection limit was lower than 1% (w/w).

A Unique Oligonucleotide Probe Hybrid on Graphene Decorated Gold Nanoparticles Modified Screen-Printed Carbon Electrode for Pork Meat Adulteration

A Unique Oligonucleotide Probe Hybrid on Graphene Decorated Gold Nanoparticles Modified Screen-Printed Carbon Electrode for Pork Meat Adulteration

Image based beef and lamb slice authentication using convolutional neural networks

Meat adulteration affects customers and the market. Existing meat authentication methods usually rely on special devices, and thus are limited to professional use only. Fake lamb or beef slices made from duck and fat appear in some Chinese hotpot restaurants. This study present a customer–conducted method for detecting such adulteration. The method takes a single image as input, and authenticates it by using the textural features of meat slices. The authentication is rapid and non–destructive. The only device required is a mobile phone with a camera. A lightweight (thus high–efficient) convolutional neural network architecture called MTx–Net was built for this task. Fourteen convolutional layers in four blocks were used for extracting valid visual features. Techniques like residuals, depth–wise convolution, dropout and batch normalization were employed in the neural network. For training and testing the neural network, 77956 meat images were collected using 225Kg of meats. This method achieves 99.38% and 98.20% accuracy on lamb and beef slice authentication, respectively.

Applications of lipidomics in goat meat products: Biomarkers, structure, nutrition interface and future perspectives

Goat meat, as a superior product including low lipids, low cholesterol contents and high-quality proteins, becomes the superior food for the national market. With the increasing demand for goat meat, the production, sensory quality and physicochemical properties of goat meat are also widely observed. Following significant discoveries on the mechanism determining goat meat quality, further research on complex and interactive factors leading to changes of goat meat quality is increasingly based on data-driven “omics” methods, such as lipidomics, which can rapidly identify and quantify >1000 lipid species at same time facilitating comprehensive analyses of lipids in tissues. Molecular mechanism and biomarkers indicating the changes of goat meat quality, authentication, meat analogue, nutrition and health by lipidomics are feasible. According to the analysis results of the classes and of different biomarkers lipids of goat meat quality, the main processes involved the biosynthesis of unsaturated fatty acids, associations with lipids and proteins, lipid oxidation, lipid hydrolysis, lipid degradation, lipid deposition and lipid denaturation, which have been translated into advanced technologies for identifying the goat meat adulteration and faux meat rapidly and accurately. SignificanceIn this review, the research of lipidomics technology, past applications, recent findings and common on the recent advances of lipidomics in the quality assessment of mutton products by lipidomics with MS approaches have been summarized. The information reported in review can serve as a reference to characterize the lipids found in mutton, clarify the application of lipidomics to the field of mutton products and provide new perspectives in producing superior quality mutton products.

Current Analytical Methods and Research Trends Are Used to Identify Domestic Pig and Wild Boar DNA in Meat and Meat Products.

The pig, one of the most important livestock species, is a meaningful source of global meat production. It is necessary, however, to prove whether a food product that a discerning customer selects in a store is actually made from pork or venison, or does not contain it at all. The problem of food authenticity is widespread worldwide, and cases of meat adulteration have accelerated the development of food and the identification methods of feed species. It is worth noting that several different molecular biology techniques can identify a porcine component. However, the precise differentiation between wild boar and a domestic pig in meat products is still challenging. This paper presents the current state of knowledge concerning the species identification of the domestic pig and wild boar DNA in meat and its products.

Volatile compounds, texture, and color characterization of meatballs made from beef, rat, wild boar, and their mixtures

The purpose of this research was to characterize the volatile compounds, texture, and color profile of meatballs made from beef, rat, wild boar, and their combinations. Volatile compounds were analyzed using SPME/GC-MS and multivariate data analysis (PCA, PLS-DA). Additionally, several textural features such as hardness, gumminess, chewiness, cohesiveness, and colour (L, a∗, b∗, C, and h) were also analyzed. The findings revealed that texture and color characteristics can only be used to differentiate meatballs based on their raw meat materials when meat adulterants are used in high concentrations (≥50%). PLS-DA analysis of volatile data revealed distinct groupings among various types of meatballs, including meatballs adulterated with rat or wild boar meat at the lowest percentage used in this study (20%). By using VIP and correlation coefficient, the strongest markers in beef, rat, and wild boar meatballs were identified as (Z)-2-amino-5-methyl-benzoic acid, 2-heptenal, and cyclobutanol, respectively. Nonanal was consistently found as a significant marker in the meatballs made from a mixture of beef-rat and beef-wild boar at different ratios. This study demonstrated that the volatile profile of meat is more reliable than physicochemical profiles for developing an analytical tool for quickly identifying undesired meat in meat-derived products.

Profiling of Volatile Compounds in Beef, Rat and Wild Boar Meat using SPME-GC/MS

The high beef price triggers adulteration of beef and other non-halal animal meat, such as wild boar and rats. An appropriate and effective analytical method is needed to differentiate halal and non-halal animal meat. The SPME/GC-MS method could authenticate meat based on specific volatile compounds in each meat. The objective of this study was to characterize volatile compounds and determine the volatile marker in raw beef, rat, wild boar meat, and their mixtures using SPME/GC-MS. The chemometrics of principal component analysis (PCA) and orthogonal projection to latent structure-discriminant analysis (OPLS-DA) classified raw beef, rat, wild boar, and their mixture. Correlation coefficients and VIP values were used to determine the volatile marker compounds for each meat in the OPLS-DA classes. The OPLS-DA results that the most robust marker in the beef class was dimethylfulvene, benzyl alcohol in the rat class, and 1,3,5-cycloheptatriene in the wild boar class. Furthermore, the most robust marker in the mixture of beef and rat class was benzaldehyde, 3-ethyl-, while 2,6-dimethyldecane was dominant in the mixture of beef and wild boar class. However, further study using larger number of samples which include commercial meat is required to confirm these results.

Cleavable molecular beacon-based loop-mediated isothermal amplification assay for the detection of adulterated chicken in meat.

A simple, sensitive, specific and fast method based on the loop-mediated isothermal amplification (LAMP) technique and cleavable molecular beacon (CMB) was developed for chicken authentication detection. LAMP and CMB were used for DNA amplification and amplicon analysis, respectively. Targeting the mitochondrial cytochrome b gene of chickens, five primers and one CMB probe were designed, and their specificity was validated against nine other animal species. The structure of CMB and concentrations of dNTPs, MgSO4, betaine, RNase H2, primers and CMB were optimized. The CMB-LAMP assay was completed within 17 min, and its limit of detection for chicken DNA was 1.5 pg μL-1. Chicken adulteration as low as 0.5% was detected in beef, and no cross-reactivity was observed. Finally, this assay was successfully applied to 20 commercial meat products. When combined with our developed DNA extraction method (the extraction time was 1 min: lysis for 10 s, washing for 20 s and elution for 30 s), the entire process (from DNA extraction to results analysis) was able to be completed within 20 min, which is at least 10 min shorter than other LAMP-based methods. Our method showed great potential for the on-site detection of chicken adulteration in meat.

Spectroscopic techniques for authentication of animal origin foods.

Milk and milk products, meat, fish and poultry as well as other animal derived foods occupy a pronounced position in human nutrition. Unfortunately, fraud in the food industry is common, resulting in negative economic consequences for customers as well as significant threats to human health and the external environment. As a result, it is critical to develop analytical tools that can quickly detect fraud and validate the authenticity of such products. Authentication of a food product is the process of ensuring that the product matches the assertions on the label and complies with rules. Conventionally, various comprehensive and targeted approaches like molecular, chemical, protein based, and chromatographic techniques are being utilized for identifying the species, origin, peculiar ingredients and the kind of processing method used to produce the particular product. Despite being very accurate and unimpeachable, these techniques ruin the structure of food, are labor intensive, complicated, and can be employed on laboratory scale. Hence the need of hour is to identify alternative, modern instrumentation techniques which can help in overcoming the majority of the limitations offered by traditional methods. Spectroscopy is a quick, low cost, rapid, non-destructive, and emerging approach for verifying authenticity of animal origin foods. In this review authors will envisage the latest spectroscopic techniques being used for detection of fraud or adulteration in meat, fish, poultry, egg, and dairy products. Latest literature pertaining to emerging techniques including their advantages and limitations in comparison to different other commonly used analytical tools will be comprehensively reviewed. Challenges and future prospects of evolving advanced spectroscopic techniques will also be descanted.

Multiple crRNAs-assisted CRISPR/Cas12a assay targeting cytochrome b gene for amplification-free detection of meat adulteration

Clustered regularly interspaced short palindromic repeats (CRISPR)/Cas systems have been widely applied in nucleic acid analysis for the high specificity. Coupled with pre-amplification steps, the sensitivity of CRISPR-based detection is greatly improved. However, an extra pre-amplification step not only complicates the detection procedures but may also cause aerosol contaminations in the process of transferring amplified solution into CRISPR system. In this study, we demonstrate that combination of multiple crRNAs in CRISPR/Cas12a system can enhance the detection sensitivity. Based on it, we establish a multiple crRNAs enhanced CRISPR (meCRISPR) method and apply it to meat adulteration identification. Take cytochrome b (Cyt b) gene as a target, meCRISPR method can directly detect as low as 1.13 ng/μL extracted pork DNA and 5% (w/w) pork contamination in pork and beef meat mixtures. There is no cross-reaction with extracted chicken, beef, duck and fish DNA. meCRISPR reaction is incubated at an isothermal temperature, and the detection process can be completed in a designed portable apparatus with a heat block, a light emitting diode and filters. For the simplicity, specificity and sufficient sensitivity of meCRISPR method, it will have great prospects in species identification, food adulteration, and genetically modified food detection.

Detection and characterization of meat adulteration in various types of meat products by using a high-efficiency multiplex polymerase chain reaction technique.

Identification of meat authenticity is a matter of increasing concerns due to religious, economical, legal, and public health reasons. However, little is known about the inspection of eight meat species in one tube reaction due to technological challenge of multiplex polymerase chain reaction (PCR) techniques. Here, a developed multiplex PCR method can simultaneously authenticate eight meat species including ostrich (753 bp), cat (564 bp), goose (391 bp), duck (347 bp), chicken (268 bp), horse (227 bp), dog (190 bp), and sheep (131 bp). The detectable deoxyribonucleic acid (DNA) contents for each target species was as low as 0.01 ng in both raw and heat-treated meat or target meat down to 0.1% (w/w) of total meat weight reflecting high stability of the assay in heat processing condition, indicating that this method is adequate for tracing meat origin in real-world meat products, which has been further validated by authenticity assays of commercial meat products. Overall, this method is a powerful tool for accurate evaluation of meat origin with a good application foreground.