GHTM

Global Health and Tropical Medicine

  • GHTM
    • About GHTM
    • Governance
    • Impact
    • Members
      • Population health, policies and services
        • PPS PhD members
        • PPS non PhD members
      • TB, HIV and opportunistic diseases and pathogens
        • THOP PhD members
        • THOP non PhD members
      • Vector-borne diseases
        • VBD PhD members
        • VBD non PhD members
      • Individual Health Care
        • IHC PhD members
        • IHC non PhD members
      • Tech & Admin support
    • Scientific Advisory Board
  • Research
    • Cross-cutting issues
      • Global Pathogen Dispersion and Population Mobility
      • Drug Discovery and Drug Resistance
      • Diagnostics
      • Public Health Information
      • Fair Research Partnerships
    • Research Groups
      • PPS – Population health, policies and services
      • THOP – TB, HIV and opportunistic diseases and pathogens
      • VBD – Vector borne diseases
      • IHC – Individual health care
    • Research in numbers
      • 2023
      • 2022
      • 2021
      • 2020
      • 2019
      • 2018
      • 2017
    • Projects
      • Ongoing Projects
      • Completed Projects
  • Outreach
    • Events
    • News
    • Policy Support & Community Outreach
  • Publications
    • 2024
    • 2023
    • 2022
    • 2021
    • 2020
    • 2019
    • 2018
    • 2017
    • 2016
    • 2015
  • Capacity Building
    • Education
      • Master Theses
      • PhD Theses
    • International
  • Infrastructures
  • Networks & Partnerships
  • Reports
    • GHTM
    • Scientific Advisory Board
    • FCT
Home / Publications / Enhanced Detection of Tuberculous Mycobacteria in Animal Tissues Using a Semi-Nested Probe-Based Real-Time PCR

Enhanced Detection of Tuberculous Mycobacteria in Animal Tissues Using a Semi-Nested Probe-Based Real-Time PCR

  • Authors: Albuquerque T, Amaro A, Botelho A, Costa P, Couto I, Cunha MV, Ferreira AS, Inácio J, Viveiros M
  • Journal: PLoS One
  • Link: http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0081337

Bovine tuberculosis has been tackled for decades by costly eradication programs in most developed countries, involving the laboratory testing of tissue samples from allegedly infected animals for detection of Mycobacterium tuberculosis complex (MTC) members, namely Mycobacterium bovis. Definitive diagnosis is usually achieved by bacteriological culture, which may take up to 6–12 weeks, during which the suspect animal carcass and herd are under sanitary arrest. In this work, a user-friendly DNA extraction protocol adapted for tissues was coupled with an IS6110-targeted semi-nested duplex real-time PCR assay to enhance the direct detection of MTC bacteria in animal specimens, reducing the time to achieve a diagnosis and, thus, potentially limiting the herd restriction period. The duplex use of a novel β-actin gene targeted probe, with complementary targets in most mammals, allowed the assessment of amplification inhibitors in the tissue samples. The assay was evaluated with a group of 128 fresh tissue specimens collected from bovines, wild boars, deer and foxes. Mycobacterium bovis was cultured from 57 of these samples. Overall, the full test performance corresponds to a diagnostic sensitivity and specificity of 98.2% (CIP95% 89.4–99.9%) and 88.7% (CIP95% 78.5–94.7%), respectively. An observed kappa coefficient was estimated in 0.859 (CIP95% 0.771–0.948) for the overall agreement between the semi-nested PCR assay and the bacteriological culture. Considering only bovine samples (n = 69), the diagnostic sensitivity and specificity were estimated in 100% (CIP95% 84.0–100%) and 97.7% (CIP95% 86.2–99.9%), respectively. Eight negative culture samples exhibiting TB-like lesions were detected by the semi-nested real-time PCR, thus emphasizing the increased potential of this molecular approach to detect MTC-infected animal tissues. This novel IS6110-targeted assay allows the fast detection of tuberculous mycobacteria in animal specimens with very high sensitivity and specificity, being amenable and cost effective for use in the routine veterinary diagnostic laboratory with further automation possibilities.

Share this:

  • Click to share on Facebook (Opens in new window) Facebook
  • Click to share on X (Opens in new window) X
  • Click to share on LinkedIn (Opens in new window) LinkedIn
  • Click to share on Pinterest (Opens in new window) Pinterest
  • Click to share on WhatsApp (Opens in new window) WhatsApp
  • Click to print (Opens in new window) Print

About GHTM

GHTM is a R&D Unit that brings together researchers with a track record in Tropical Medicine and International & Global Health. It aims at strengthening Portugal's role as a leading partner in the development and implementation of a global health research agenda. Our evidence-based interventions contribute to the promotion of equity in health and to improve the health of populations.

Contacts

Rua da Junqueira, 100
1349-008 Lisboa
Portugal

+351 213 652 600

  • E-mail
  • Facebook
  • LinkedIn
  • Twitter
  • YouTube

Map

  • Events
  • Research Groups
  • Cross-cutting issues
© Copyright 2025 IHMT-UNL All Rights Reserved.
  • Universidade Nova de Lisboa
  • Fundação para a Ciência e a Tecnologia

    UIDB/04413/2020
    UIDP/04413/2020

We use cookies to ensure that we give you the best experience on our website. If you continue to use this site we will assume that you are happy with it.Ok