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Veterinary parasitology2007; 147(1-2); 16-25; doi: 10.1016/j.vetpar.2007.03.031

A quantitative PCR assay for the detection and quantification of Babesia bovis and B. bigemina.

Abstract: The haemoparasites Babesia bovis and Babesia bigemina affect cattle over vast areas of the tropics and temperate parts of the world. Microscopic examination of blood smears allows the detection of clinical cases of babesiosis, but this procedure lacks sensitivity when parasitaemia levels are low. In addition, differentiating between similar haemoparasites can be very difficult. Molecular diagnostic procedures can, however, overcome these problems. This paper reports a quantitative PCR (qPCR) assay involving the use of SYBR Green. Based on the amplification of a small fragment of the cytochrome b gene, this method shows both high sensitivity and specificity, and allows quantification of parasite DNA. In tests, reproducible quantitative results were obtained over the range of 0.1 ng to 0.1 fg of parasite DNA. Melting curve analysis differentiated between B. bovis and B. bigemina. To assess the performance of the new qPCR procedure it was used to screen for babesiosis in 40 cows and 80 horses. B. bigemina was detected in five cows (three of these were also found to be positive by standard PCR techniques targeting the 18S rRNA gene). In addition, B. bovis was detected in one horse and B. bigemina in two horses using the proposed method, while none was found positive by ribosomal standard PCR. The sequences of the B. bigemina cytochrome b and 18S rRNA genes were completely conserved in isolates from Spain and Argentina, while those of B. bovis showed moderate polymorphism.
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Publication Date: 2007-04-26 PubMed ID: 17466458DOI: 10.1016/j.vetpar.2007.03.031Google Scholar: Lookup
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  • Journal Article
  • Research Support
  • Non-U.S. Gov't

Summary

This research summary has been generated with artificial intelligence and may contain errors and omissions. Refer to the original study to confirm details provided. Submit correction.

This study details the development and testing of a highly specific and sensitive molecular diagnostic tool using quantitative PCR for the detection and measurement of Babesia bovis and Babesia bigemina DNA, parasites affecting cattle globally.

Objective of the Research

  • The research aimed to develop a more effective diagnostic method to detect and measure the parasites Babesia bovis and Babesia bigemina. These parasites, which affect cattle and can sometimes infect horses, are prevalent in tropical and temperate regions. The current method of diagnosis, microscopic examination of blood smears, lacks sensitivity, especially when levels of parasites in the blood are low. Additionally, it can be challenging to distinguish between similar types of parasites.

Methodology

  • To overcome the limitations of the current diagnostic method, the researchers developed a quantitative PCR (qPCR) assay that uses SYBR Green, a nucleic acid stain used in molecular biology.
  • This new method amplifies a small fragment of the cytochrome b gene, a part of the parasites’ mitochondrial DNA, which allows for the detection and quantification of parasite DNA.
  • They tested the new procedure’s effectiveness by screening 40 cows and 80 horses for babesiosis, the disease caused by the Babesia parasites.

Findings

  • The results showed that the new method was highly sensitive and specific, allowing for quantifiable results ranging from 0.1 ng to 0.1 fg of parasite DNA.
  • Its accuracy was also demonstrated by the ability to distinguish between B. bovis and B. bigemina through melting curve analysis.
  • The method detected B. bigemina in three cows and B. bovis in one horse, and B. bigemina in two horses, which were not detected by the standard PCR method targeting the 18S rRNA gene.

Significance of the Results

  • The research provided evidence that the developed qPCR assay is a more effective method for diagnosing babesiosis.
  • Early and accurate detection of these parasites can lead to a faster and specific treatment protocol, potentially reducing the risk of outbreaks in the future.

Cite This Article

APA
Buling A, Criado-Fornelio A, Asenzo G, Benitez D, Barba-Carretero JC, Florin-Christensen M. (2007). A quantitative PCR assay for the detection and quantification of Babesia bovis and B. bigemina. Vet Parasitol, 147(1-2), 16-25. https://doi.org/10.1016/j.vetpar.2007.03.031

Publication

Veterinary parasitology
ISSN: 0304-4017
NlmUniqueID: 7602745
Country: Netherlands
Language: English
Volume: 147
Issue: 1-2
Pages: 16-25

Researcher Affiliations

Buling, A
  • Microbiology and Parasitology Department, Faculty of Pharmacy, University of Alcalá, 28871 Alcalá de Henares, Spain.
Criado-Fornelio, A
    Asenzo, G
      Benitez, D
        Barba-Carretero, J C
          Florin-Christensen, M

            MeSH Terms

            • Animals
            • Babesia / isolation & purification
            • Babesia bovis / isolation & purification
            • Babesiosis / diagnosis
            • Babesiosis / veterinary
            • Cattle
            • Cattle Diseases / diagnosis
            • Cytochromes b / genetics
            • Horse Diseases / diagnosis
            • Horses
            • Molecular Sequence Data
            • Plasmids / genetics
            • Polymerase Chain Reaction
            • RNA, Ribosomal, 18S / genetics
            • Reproducibility of Results
            • Sensitivity and Specificity

            Citations

            This article has been cited 18 times.
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