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Ring Trials
Ring Trials 2024
Ring Trials 2023
Ring Trials 2022
Ring Trials 2020
Ring Trials 2019
Ring Trials 2018
Ring Trials 2017
Ring Trials 2016
Ring Trials 2015
Ring Trials 2014
Ring Trials 2013
Ring Trials 2012
Ring Trials 2011
Ring Trials 2010

Ring Trials 2024


Histopathological and immunohistochemical diagnosis of tuberculosis

Ring Trial

Feb. 2024 - May 2024
State: In process

Description


According to the article 9 of the Commission Delegated Regulation (EU) 2020/689, an animal can be classified as a suspected case if post-mortem lesions or histological findings are indicative of the infection by members of the Mycobacterium tuberculosis complex (MTBC). To confirm a case, the identification of specific antigens of MTBC by histopathological and immunohistochemical diagnosis can be used.


Objectives


1. To assess the ability of NRLs to provide accurate results for the histopathological and immunohistochemical diagnosis of tuberculosis in animals. 2. To provide an official inter-laboratory evaluation scheme that can be useful to those laboratories which need to provide evidence of their proficiency to perform histopathological and immunohistochemical diagnosis of tuberculosis in animals.


Expected outputs


Retain or improve the NRLs´ competency to perform histopathological and immunohistochemical diagnosis of tuberculosis and study the results of laboratories.
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Ring Trials 2023


Molecular characterization of MTBC

Ring Trial

Oct. 2023 - Feb. 2024
State: Evaluation of results

Description


Molecular characterization of the M. tuberculosis complex strains is an useful tool for epidemiological studies. Therefore, the assignation of profiles should be standardize all over Europe.


Objectives


To assign the proper SB code and VNTR profile for the molecular characterization of the M. tuberculosis complex strains. A set of DNA samples will be sent to NRLs to assign a SB number and the VNTR profile, and to identify the MTBC species.


Expected outputs


Proper assignation of profiles to compare the results around Europe, and identification of MTBC members.
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Ring Trials 2022


Direct extraction and bacteriological culture

Ring Trial

Nov. 2022 - June 2023
State: Final report

Description


Proficiency testing for the recovery of members of the Mycobacterium tuberculosis complex (MTBC) from bovine tissues by culture and/or the detection of DNA belonging to members of the MTBC isolated from bovine tissues.


Objectives


To test the competency of appointed NRLs to recover members of the MTBC from bovine tissues by culture and for detection of MTBC in bovine tissues by PCR.


Expected outputs


Assessment of the NRLs’ competency to perform culture for the recovery of MTBC from bovine tissues and direct PCR on tissue samples to detect MTBC.
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Gamma interferon release assay

Ring Trial

Oct. 2022 - Jan. 2023
State: Evaluation of results

Description


Proficiency testing for the IFNγ release assay as an official method for the detection of interferon gamma in bovine plasma samples for the diagnosis of MTBC infection.


Objectives


To assess the competence of NRLs to carry out the assay on bovine plasma samples under the purposes for granting and maintenance of disease-free status (DR 2020/689), and movements of cattle within the Union (DR 2020/688); and to provide an official inter-laboratory evaluation scheme to the NRLs for accreditation.


Expected outputs


Assessment of the NRLs’ competency to perform the IFNγ release assay as an official diagnostic test and evaluation of the intra and inter laboratory variation of the test.
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Molecular characterization of MTBC

Ring Trial

Feb. 2022 - Aug. 2022
State: Final report

Description


Molecular characterization of the M. tuberculosis complex strains is an useful tool for epidemiological studies. Therefore, the assignation of profiles should be standardize all over Europe.


Objectives


To assign the proper SB code and VNTR profile for the molecular characterization of the M. tuberculosis complex strains. A set of DNA samples will be sent to NRLs to assign a SB number and the VNTR profile.


Expected outputs


Proper assignation of profiles to compare the results around Europe.
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Ring Trials 2020


Histopathological and immunohistochemical diagnosis of tuberculosis

Ring Trial

Nov. 2020 - March 2021
State: Final report

Description


According to the Council Directive 64/432/EEC, the presence of the agents of bovine tuberculosis, in clinical and post-mortem specimens may be demonstrated by examination of stained smears or immunoperoxidase techniques and confirmed by cultivation of the organism on primary isolation medium.


Objectives


1. To assess the ability of NRLs to provide accurate results for the histopathological and immunohistochemical diagnosis of tuberculosis in animals. 2. To provide an official inter-laboratory evaluation scheme that can be useful to those laboratories which need to provide evidence of their proficiency to perform histopathological and immunohistochemical diagnosis of tuberculosis in animals.


Expected outputs


Retain or improve the NRLs´ competency to perform histopathological and immunohistochemical diagnosis of tuberculosis and study the results of laboratories.
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Ring Trials 2019


Direct extraction and bacteriological culture

Ring Trial

Dec. 2019 - July 2020
State: Final report

Description


Proficiency testing for the recovery of members of the Mycobacterium tuberculosis complex (MTBC) from bovine tissues by culture and/or the detection of DNA belonging to members of the MTBC isolated from bovine tissues.


Objectives


To test the competency of appointed NRLs to recover members of the MTBC from bovine tissues by culture and to assess the performance of molecular techniques implemented for the detection of MTBC in bovine tissues.


Expected outputs


Assessment of the NRLs’ competency to perform culture for the recovery of MTBC from bovine tissues and evaluation of the performance of molecular techniques to detect MTBC in animal tissues respect to culture.
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Gamma interferon test

Ring Trial

Nov. 2019 - March 2020
State: Final report

Description


Proficiency testing for the IFNγ release assay as a supplementary official testing to enable detection of the maximum number of infected and diseased animals (64/432/EEC); and for its implementation as a potential official test for the confirmation of OTF herds and/or the intracommunity trade of animals.


Objectives


To assess the competence of appointed NRLs to carry out the enzyme linked immunosorbent assay on plasmas of previously stimulated blood samples and evaluate the intra and inter-laboratory variation of results.


Expected outputs


Assessment of the NRLs’ competency to perform the IFNγ release assay as an official test and evaluation of the intra and inter laboratory variation of the test.
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Ring Trials 2018


Histopathological diagnosis of tuberculosis

Ring Trial

Dec. 2018 - March 2019
State: Final report

Description


The official diagnosis of tuberculosis is based mainly on the detection of the response in the medium of the cells originated in the host after infection. However, other techniques included in Council Directive 64/432 / EEC are still used for the diagnosis of bovine tuberculosis.


Objectives


1. To assess the ability of NRLs to provide accurate results for the histopathological diagnosis of tuberculosis in animals. 2. To provide an official inter-laboratory evaluation scheme that can be useful to those laboratories which need to provide evidence of their proficiency to perform histopathological diagnosis of tuberculosis in animals for accreditation purposes.


Expected outputs


Assessment of the NRLs´ competency to perform histopathological diagnosis of tuberculosis and study the results of laboratories.
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Molecular characterization of MTBC

Ring Trial

Nov. 2018 - Feb. 2019
State: Preliminary report

Description


Molecular characterization of the M. tuberculosis complex strains is an useful tool for epidemiological studies. Therefore, the assignation of profiles should be standardize all over Europe.


Objectives


To assign the proper SB code and VNTR profile for the molecular characterization of the M. tuberculosis complex strains. A set of DNA samples will be sent to NRLs to assign a SB number and the VNTR profile.


Expected outputs


Proper assignation of profiles in order to compare the results around Europe.
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Ring Trials 2017


Gamma interferon test

Ring Trial

Dec. 2017 - March 2018
State: Final report

Description


Proficiency testing for the implementation of the IFNγ release assay as an official test for the confirmation of OTF herds and/or the intracommunity trade of animals (64/432/EU).


Objectives


To assess the competence of appointed NRLs to carry out the enzyme linked immunosorbent assay on plasmas of previously stimulated blood samples and evaluate the intra and inter-laboratory variation of results.


Expected outputs


Assessment of the NRLs’ competency to perform the IFNγ release assay as an official test and evaluation of the intra and inter laboratory variation of the test.
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Direct extraction and bacteriological culture

Ring Trial

Nov. 2017 - May 2018
State: Final report

Description


Proficiency testing for the recovery of members of the Mycobacterium tuberculosis complex (MTBC) from bovine tissues by culture and/or the detection of DNA belonging to members of the MTBC isolated from bovine tissues


Objectives


To test the competency of appointed NRLs to recover members of the MTBC from bovine tissues by culture and to assess the performance of molecular techniques implemented for the detection of MTBC in bovine tissues


Expected outputs


Assessment of the NRLs’ competency to perform culture for the recovery of MTBC from bovine tissues and evaluation of the performance of molecular techniques to detect MTBC in animal tissues respect to culture
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Ring Trials 2016


Histopathological diagnosis of tuberculosis

Ring Trial

June 2016 - Sept. 2016
State: Evaluation of results

Description


The official diagnosis of bovine tuberculosis is based mainly on the detection of the cell mediated immune response originated in the host after the infection. However, other techniques included in the Council Directive 64/432/EEC are still used for bovine tuberculosis diagnosis.


Objectives


Evaluation of the methodology employed for the histopathological diagnosis of tuberculosis in different animal species. Samples will be sent to perform the staining (Hematoxylin-eosin, Ziehl-Neelsen) and immunohistochemistry (when possible) protocols together with the pathologist interpretation with the final objective to give a final diagnosis after the examination of the samples.


Expected outputs


Knowledge of the methodology used to perform the histopathological investigation to diagnose bovine tuberculosis and evaluation of the concordance of results between laboratories.
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Ring Trials 2015


Direct extraction and bacteriological culture

Ring Trial

Nov. 2015 - July 2016
State: Final report

Description


Isolation and/or detection of mycobacteria by microbiological culture and direct extraction from bovine tissue samples.


Objectives


Isolation and/or detection of mycobacteria by microbiological culture and direct extraction from bovine tissue samples


Expected outputs


Evaluation of the methodology for the direct extraction in comparison with the bacteriological culture, standardization of techniques, and assurance of the quality of the results offered by the NRLs
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Ring Trials 2014


Biological test of tuberculins

Ring Trial

Sept. 2014 - Dec. 2014
State: Final report

Description


Tuberculins are the reagents currently in use for the diagnosis of bovine tuberculosis. Although guidelines for the determination of the biological potency of the tuberculins in guinea pigs are available, some modifications have been encountered between NRLs (number of inoculations, dilutions of the tuberculins, statistical program, infection or sensitization, period of time).


Objectives


Evaluation of the methodology for determination of biological potency of the PPD in guinea pigs. PPDs will be sent to determine the biological potency in each laboratory


Expected outputs


Knowledge of the methodology for determination of the biological potency of tuberculins in guinea pigs following international guidelines
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DVR-Spoligotyping

Ring Trial

June 2014 - Sept. 2014
State: Final report

Description


Molecular characterization of the M. tuberculosis complex strains is an useful tool for epidemiological studies. Therefore the assignation of profiles should be standardize all over Europe


Objectives


To evaluate the DVR-spoligotyping technique performance for the M. tuberculosis complex strains. A set of DNA will be sent to NRLs to assign a SB number by spoligotyping


Expected outputs


Proper assignation of the spoligotyping profiles in order to compare the results around Europe
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Ring Trials 2013


Direct extraction and microbiological culture

Ring Trial

Sept. 2013 - Feb. 2014
State: Final report

Description


Ring trial for the standardization and setting up of the bacteriological culture and direct extraction from tissue samples


Objectives


Isolation and/or detection of mycobacteria by microbiological culture and direct extraction from bovine tissue samples


Expected outputs


Evaluation of the methodology for the direct extraction in comparison with the bacteriological culture, standardization of techniques, and assurance of the quality of the results offered by the NRLs
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Analytical sensitivity of the PCR

Ring Trial

June 2013 - Sept. 2013
State: Final report

Description


A wide variety of PCR protocols for the identification of mycobacteria are currently used by the National Reference Laboratories. The EU-RL for bovine tuberculosis looks for the standardization and setting up of techniques, and to assure the quality of the result


Objectives


Determination of the analytical sensitivity of the PCR for the identification of members of the Mycobacterium tuberculosis complex


Expected outputs


Selection of the most sensitive PCR for the detection of M. tuberculosis complex isolates. Recommendation of the use of a protocol based on the results
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Mycobacteria recovery from different culture media

Ring Trial

April 2013 - July 2013
State: Final report

Description


The EU-RL has collected information regarding the different media (solid/liquid) used in the NRLs for primary isolation or subculture of members of the Mycobacterium tuberculosis complex and there are more than 10 manufacturers and media available


Objectives


To evaluate the sensitivity of the culture media used in the NRLs


Expected outputs


To define the sensitivity of the culture media to be able to recommend the media that should be used for isolation of Mycobacteria in the Member States
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Ring Trials 2012


Gamma interferon test

Ring Trial

Oct. 2012 - April 2013
State: Final report

Description


The interferon gamma assay, an in vivo celular immune response technique, is an ancillary test that allows the detection of the maximum number of infected animals in a herd or region of high prevalence


Objectives


Compare the qualitative results using different kits (specificity and sensitivity) and the repeatability intralaboratory (precision)


Expected outputs


Evaluation of the concordance of results between laboratories
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Comparative test to evaluate the performance of the MIRU-VNTR typing

Ring Trial

June 2012 – Oct. 2012
State: Final report

Description


Variable Number Tandem Repeat (VNTR) sequences are valuable markers for genotyping of the Mycobacterium tuberculosis complex species. VNTR genotyping relies on the PCR amplification of a VNTR locus using specific primers for its flanking region and the determination of the size of amplicons.


Objectives


To test the reproducibility of the VNTR typing across different settings and to evaluate the proficiency of the NRLs to correctly assign the VNTR profile to the isolates for epidemiology purposes.


Expected outputs


To provide NRLs with a set of standardized DNA samples to ensure the reliability of their own results.
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Ring Trials 2011


Comparative test to evaluate the Spoligotyping membranes

Ring Trial

Dec. 2011 – April 2012
State: Final report

Description


The direct repeat (DR) region is one of the most important genomic targets used for molecular typing of the Mycobacterium tuberculosis complex isolates and spoligotyping is the most extensively applied technique for the study of this locus.


Objectives


Comparing the results obtained using different spoligotyping membranes (commercial or home-made membranes).


Expected outputs


Define the quality of the comercial and home-made membranes for molecular characterization.
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Comparative test for the diagnosis of Bovine tuberculosis by direct Extraction of DNA

Ring Trial

Oct. 2011 – April 2012
State: Final report

Description


The diagnosis of bovine tuberculosis by direct extraction of DNA from tissue samples is one of the priorities for the European Reference Laboratory (EU-RL) in order to achieve a rapid diagnosis of bovine tuberculosis without the need to perform microbiological culture.


Objectives


Evaluation of the methodology for the direct detection and identification of mycobacteria by PCR from tissue samples and evaluation of the ability of the laboratories for the isolation and identification of mycobacteria from the same tissue samples by microbiological culture.


Expected outputs


Selection of the most sensitive DNA extraction protocol from tissue samples.
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Ring Trials 2010


Identification and molecular characterization

Ring Trial

Nov. 2010 – Feb. 2011
State: Final report

Description


In 2009, a series of questionnaires on logistics of the laboratories, plus techniques used for diagnosis and epidemiology were submitted to all NRLs. From the results collected in these questionnaires, it was evident that there is variety of expertise and techniques amongst NRLs. With this in mind, the ring trial was organised to assess the potential impact of this diversity.


Objectives


Evaluation of the proficiency of the NRLs to correctly identify and assign spoligotyping profiles to the main members of the Mycobacterium tuberculosis complex.


Expected outputs


Evaluation of the methodology for the identification of members of the Mycobacterium tuberculosis complex.
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