Detection of bovine mitochondrial DNA in ruminant feeds: A molecular approach to test for the presence of bovine-derived materials

Marco Tartaglia, Ernestina Saulle, Simonetta Pestalozza, Luisella Morelli, Giovanni Antonucci, Piero A. Battaglia

Research output: Contribution to journalArticle

Abstract

A ban on ruminant-derived proteins in ruminant feeds has been introduced as a preventive measure to avoid the spread of bovine spongiform encephalopathy (BSE), as well as to minimize any potential risk of BSE transmission from bovines to humans. In the absence of commercially available efficient methods for identification of bovine-derived proteins in animal feeds, we developed a rapid and sensitive polymerase chain reaction (PCR)-based assay which allows detection and identification of a bovine-specific mitochondrial DNA sequence from feedstuffs. The amplified product encodes for the whole ATPase subunit 8 and the amino-terminal portion of the ATPase subunit 6 proteins, which are known to exhibit a relatively low degree of conservation among vertebrates. The specific amplification of such a bovine mitochondrial sequence from reference feedstuff samples was demonstrated by means of both direct sequencing and single-strand conformational analysis of the PCR product. Specificity was also confirmed by the absence of detectable homologous PCR product when using reference feedstuff samples lacking bovine-derived meat and bonemeals, or genomic DNA samples from vertebrates whose offals are commonly included in animal feeds. This method allows detection of the presence of bovine mitochondrial DNA in feedstuffs containing less than 0.125% of bovine-derived meat and bonemeals. Furthermore, it does not appear to be considerably affected by prolonged heat treatment. DpnII and SspI restriction endonuclease digestions of the unpurified PCR product may be used routinely to confirm the bovine origin of the amplified sequence. Since this method is specific, rapid, and sensitive, it could be successfully utilized as a routine control assay to evaluate the presence of bovine-derived meat and bonemeals in ruminant feeds.

Original languageEnglish
Pages (from-to)513-518
Number of pages6
JournalJournal of Food Protection
Volume61
Issue number5
Publication statusPublished - May 1998

Fingerprint

Ruminants
Mitochondrial DNA
ruminants
mitochondrial DNA
cattle
testing
polymerase chain reaction
Meat
Bovine Spongiform Encephalopathy
Polymerase Chain Reaction
bovine spongiform encephalopathy
meat
adenosinetriphosphatase
Adenosine Triphosphatases
Vertebrates
vertebrates
proteins
restriction endonucleases
Protein Subunits
assays

ASJC Scopus subject areas

  • Food Science
  • Biotechnology
  • Applied Microbiology and Biotechnology

Cite this

Detection of bovine mitochondrial DNA in ruminant feeds : A molecular approach to test for the presence of bovine-derived materials. / Tartaglia, Marco; Saulle, Ernestina; Pestalozza, Simonetta; Morelli, Luisella; Antonucci, Giovanni; Battaglia, Piero A.

In: Journal of Food Protection, Vol. 61, No. 5, 05.1998, p. 513-518.

Research output: Contribution to journalArticle

Tartaglia, Marco ; Saulle, Ernestina ; Pestalozza, Simonetta ; Morelli, Luisella ; Antonucci, Giovanni ; Battaglia, Piero A. / Detection of bovine mitochondrial DNA in ruminant feeds : A molecular approach to test for the presence of bovine-derived materials. In: Journal of Food Protection. 1998 ; Vol. 61, No. 5. pp. 513-518.
@article{bcd25a3c905c4ac5b828f222d276bbf6,
title = "Detection of bovine mitochondrial DNA in ruminant feeds: A molecular approach to test for the presence of bovine-derived materials",
abstract = "A ban on ruminant-derived proteins in ruminant feeds has been introduced as a preventive measure to avoid the spread of bovine spongiform encephalopathy (BSE), as well as to minimize any potential risk of BSE transmission from bovines to humans. In the absence of commercially available efficient methods for identification of bovine-derived proteins in animal feeds, we developed a rapid and sensitive polymerase chain reaction (PCR)-based assay which allows detection and identification of a bovine-specific mitochondrial DNA sequence from feedstuffs. The amplified product encodes for the whole ATPase subunit 8 and the amino-terminal portion of the ATPase subunit 6 proteins, which are known to exhibit a relatively low degree of conservation among vertebrates. The specific amplification of such a bovine mitochondrial sequence from reference feedstuff samples was demonstrated by means of both direct sequencing and single-strand conformational analysis of the PCR product. Specificity was also confirmed by the absence of detectable homologous PCR product when using reference feedstuff samples lacking bovine-derived meat and bonemeals, or genomic DNA samples from vertebrates whose offals are commonly included in animal feeds. This method allows detection of the presence of bovine mitochondrial DNA in feedstuffs containing less than 0.125{\%} of bovine-derived meat and bonemeals. Furthermore, it does not appear to be considerably affected by prolonged heat treatment. DpnII and SspI restriction endonuclease digestions of the unpurified PCR product may be used routinely to confirm the bovine origin of the amplified sequence. Since this method is specific, rapid, and sensitive, it could be successfully utilized as a routine control assay to evaluate the presence of bovine-derived meat and bonemeals in ruminant feeds.",
author = "Marco Tartaglia and Ernestina Saulle and Simonetta Pestalozza and Luisella Morelli and Giovanni Antonucci and Battaglia, {Piero A.}",
year = "1998",
month = "5",
language = "English",
volume = "61",
pages = "513--518",
journal = "Journal of Food Protection",
issn = "0362-028X",
publisher = "International Association for Food Protection",
number = "5",

}

TY - JOUR

T1 - Detection of bovine mitochondrial DNA in ruminant feeds

T2 - A molecular approach to test for the presence of bovine-derived materials

AU - Tartaglia, Marco

AU - Saulle, Ernestina

AU - Pestalozza, Simonetta

AU - Morelli, Luisella

AU - Antonucci, Giovanni

AU - Battaglia, Piero A.

PY - 1998/5

Y1 - 1998/5

N2 - A ban on ruminant-derived proteins in ruminant feeds has been introduced as a preventive measure to avoid the spread of bovine spongiform encephalopathy (BSE), as well as to minimize any potential risk of BSE transmission from bovines to humans. In the absence of commercially available efficient methods for identification of bovine-derived proteins in animal feeds, we developed a rapid and sensitive polymerase chain reaction (PCR)-based assay which allows detection and identification of a bovine-specific mitochondrial DNA sequence from feedstuffs. The amplified product encodes for the whole ATPase subunit 8 and the amino-terminal portion of the ATPase subunit 6 proteins, which are known to exhibit a relatively low degree of conservation among vertebrates. The specific amplification of such a bovine mitochondrial sequence from reference feedstuff samples was demonstrated by means of both direct sequencing and single-strand conformational analysis of the PCR product. Specificity was also confirmed by the absence of detectable homologous PCR product when using reference feedstuff samples lacking bovine-derived meat and bonemeals, or genomic DNA samples from vertebrates whose offals are commonly included in animal feeds. This method allows detection of the presence of bovine mitochondrial DNA in feedstuffs containing less than 0.125% of bovine-derived meat and bonemeals. Furthermore, it does not appear to be considerably affected by prolonged heat treatment. DpnII and SspI restriction endonuclease digestions of the unpurified PCR product may be used routinely to confirm the bovine origin of the amplified sequence. Since this method is specific, rapid, and sensitive, it could be successfully utilized as a routine control assay to evaluate the presence of bovine-derived meat and bonemeals in ruminant feeds.

AB - A ban on ruminant-derived proteins in ruminant feeds has been introduced as a preventive measure to avoid the spread of bovine spongiform encephalopathy (BSE), as well as to minimize any potential risk of BSE transmission from bovines to humans. In the absence of commercially available efficient methods for identification of bovine-derived proteins in animal feeds, we developed a rapid and sensitive polymerase chain reaction (PCR)-based assay which allows detection and identification of a bovine-specific mitochondrial DNA sequence from feedstuffs. The amplified product encodes for the whole ATPase subunit 8 and the amino-terminal portion of the ATPase subunit 6 proteins, which are known to exhibit a relatively low degree of conservation among vertebrates. The specific amplification of such a bovine mitochondrial sequence from reference feedstuff samples was demonstrated by means of both direct sequencing and single-strand conformational analysis of the PCR product. Specificity was also confirmed by the absence of detectable homologous PCR product when using reference feedstuff samples lacking bovine-derived meat and bonemeals, or genomic DNA samples from vertebrates whose offals are commonly included in animal feeds. This method allows detection of the presence of bovine mitochondrial DNA in feedstuffs containing less than 0.125% of bovine-derived meat and bonemeals. Furthermore, it does not appear to be considerably affected by prolonged heat treatment. DpnII and SspI restriction endonuclease digestions of the unpurified PCR product may be used routinely to confirm the bovine origin of the amplified sequence. Since this method is specific, rapid, and sensitive, it could be successfully utilized as a routine control assay to evaluate the presence of bovine-derived meat and bonemeals in ruminant feeds.

UR - http://www.scopus.com/inward/record.url?scp=0032076235&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0032076235&partnerID=8YFLogxK

M3 - Article

C2 - 9709219

AN - SCOPUS:0032076235

VL - 61

SP - 513

EP - 518

JO - Journal of Food Protection

JF - Journal of Food Protection

SN - 0362-028X

IS - 5

ER -