Detection of Bovine Central Nervous System Tissues in Rendered Animal By-Products by One-Step Real-Time Reverse Transcription PCR Assay

Abstract

Contamination of rendered animal byproducts with central nervous system tissues (CNST) from animals with bovine spongiform encephalopathy is considered one of the vehicles of disease transmission. Removal from the animal feed chain of CNST originated from cattle of a specified age category, species-labeling of rendered meat products, and testing of rendered products for bovine CNST are tasks associated with the epidemiological control of bovine spongiform encephalopathy. A single-step TaqMan real-time reverse transcriptase (RRT) PCR assay was developed and evaluated for specific detection of bovine glial fibrillary acidic protein (GFAP) mRNA, a biomarker of bovine CNST, in rendered animal by-products. An internal amplification control, mammalian b -actin mRNA, was coamplified in the duplex RRT-PCR assay to monitor amplification efficiency, normalize amplification signals, and avoid false-negative results. The functionality of the GFAP mRNA RRT-PCR was assessed through analysis of laboratory-generated binary mixtures of bovine central nervous system (CNS) and muscle tissues treated under various thermal settings imitating industrial conditions. The assay was able to detect as low as 0.05 % (wt/wt) bovine brain tissue in binary mixtures heat treated at 110 to 130°C for 20 to 60 min. Further evaluation of the GFAP mRNA RRT-PCR assay involved samples of industrial rendered products of various species origin and composition obtained from commercial sources and rendering plants. Low amounts of bovine GFAP mRNA were detected in several bovine-rendered products, which was in agreement with declared species composition. An accurate estimation of CNS tissue content in industrial-rendered products was complicated due to a wide range of temperature and time settings in rendering protocols. Nevertheless, the GFAP mRNA RRT-PCR assay may be considered for bovine CNS tissue detection in rendered products in combination with other available tools (for example, animal age verification) in inspection programs.

Microwave-accelerated bioassay technique for rapid and quantitative detection of biological and environmental samples

“…These tests conducted in hospital or diagnostic laboratory are based on ELISA (Ahadi et al. 2015; Jiang et al 2014) and RT-PCR (Andrievskaia and Tangorra 2014), that have a turnaround time of 1–3 days, depending upon the availability of manpower and the medical emergency of the patient.…”

The Development of a Risk Assessment Model for Inedible Rendering Plants in Canada: Identifying and Selecting Feed Safety-Related Factors

“…• Presence of non-compliance related to the process control program as assessed by a government inspector (current and past three inspection results)End-product control program[20,22,[24][25][26][27]29,[31][32][33][34]40,41,[57][58][59][60][61][62][63][65][66][67] • Presence of non-compliance related to the end-product control program as assessed by a government inspector (current and past three inspection results)Import control program[36,38,45,46] • Presence of non-compliance related to the import control program as assessed by a government inspector (current and past three inspection results) • Presence of non-compliance related to the water, ice, and steam program as assessed by a government inspector (current and past three inspection results)…”