8th EQAsia External Quality Assessment trial: E. coli, K. pneumoniae, Acinetobacter spp., and S. aureus – 2024

The EQAsia project was launched in 2020 aiming to strengthen the provision of External Quality Assessment (EQA) services across the One Health sector among National Reference Laboratories / Centres of Excellence in South and Southeast Asia. EQAsia is supported by the Fleming Fund and strives to increase the quality of laboratory-based surveillance of World Health Organization (WHO) Global Antimicrobial Resistance and Use Surveillance System (GLASS) priority pathogens [1] and Food and Agricultural Organization (FAO) priority pathogens [2]. EQAsia has transitioned to a second phase and continues to deliver the established EQA programme for both the Human Health (HH) sector and Food and Animal Health (AH) sector in the region until the end of 2025.

The EQAsia Consortium includes the Technical University of Denmark, National Food Institute (DTU Food) as the Lead Grantee, the International Vaccine Institute (IVI) in South Korea, and the Faculty of Veterinary Science, Chulalongkorn University (CUVET) in Thailand.

EQAsia provides a state-of-the-art EQA program at no cost for the South and Southeast Asian region distributed (?) through CUVET Thailand, a leading regional provider. The EQAsia program is designed to enable the laboratories to select and participate in relevant proficiency tests of both pathogen identification and antimicrobial susceptibility testing (AST), in accordance with the requirements of the WHO GLASS [1]. The EQA program is supported by an informatics module where laboratories can report their results and methods used.

A total of eight EQA trials have taken place since 2021, all of which focused on the WHO GLASS [1] and FAO priority pathogens [2]: Salmonella spp., Escherichia coli, Klebsiella pneumoniae, Shigella spp., Acinetobacter spp., Pseudomonas aeruginosa, Staphylococcus aureus, Campylobacter (C. coli and C. jejuni), Enterococcus (E. faecium and E. faecalis), Streptococcus pneumoniae and Neisseria gonorrhoeae. In addition, a Matrix EQA trial was offered three times, consisting of a complex food sample spiked with AmpC beta-lactamases (AmpC), extended-spectrum beta-lactamases (ESBLs) or carbapenemase-producing E. coli for surveillance purposes.

The aim was to align with the scope of WHO Tricycle project and, as recommended by FAO, to evaluate the capacity of veterinary laboratories to detect multidrug-resistant bacteria from food matrices.

For a given organism, candidate strains are assessed and validated by DTU Food and an external partner (The Peter Doherty Institute for Infection and Immunity, Australia). The validation includes both phenotypic determination of minimum inhibitory concentration (MIC) by broth microdilution, and whole-genome sequencing (WGS) to detect antimicrobial resistance (AMR) genes and chromosomal point mutations. The test strains are then selected based on the phenotypic AMR profile to include a heterogeneous panel, allowing for strain variation from almost pan-resistant to fully susceptible isolates.

This report contains results from the eight EQA trial of the EQAsia project (EQA8) carried out in April – June 2024. The trial included four EQA panels, each containing seven test strains. Of these, five were the organism in question (target organism, i.e., K. pneumoniae), whereas the other two test strains were different from the targeted species (reported as non-[organism], i.e., non-K. pneumoniae). For each of the seven test strains, participants were requested to report which five strains belong to the expected target organism. For the two organisms different from the expected, no additional testing was needed. For the remaining five test strains of the target organism, AST results were requested.

This eight EQA trial includes identification and AST of E. coli, K. pneumoniae, Acinetobacter spp. and S. aureus. The aim of this EQA trial was to monitor the quality of AST results produced by the participating laboratories and identify underperforming laboratories requiring additional support and assistance to improve their performance in bacterial identification and AST.

The evaluation of the participants’ results is based on international guidelines, specifically the Clinical and Laboratory Standards Institute (CLSI) and the European Committee on Antimicrobial Susceptibility Testing (EUCAST). Interpretative criteria referring to both disk diffusion and MIC determination are listed in the EQA8 protocol (Appendix 1) and allow for the obtained results to be interpreted into categories as resistant, intermediate, or susceptible depending on the method used. Results in agreement with the expected interpretation are scored ‘4’ (correct), while results deviating from the expected interpretation are scored as either ‘0’ (incorrect: very major error), ‘1’ (incorrect: major error) or ‘3’ (incorrect: minor error), as described in the EQA8 protocol (Appendix 1). This standardized interpretation of results is necessary to allow comparison of performance between laboratories. Laboratory performance is considered acceptable if there is < 5 % deviation from the expected results.

Evaluation of a result as “deviating from the expected interpretation” should be carefully analysed in a root cause analysis procedure performed by individual participants (self-evaluation) when the EQA results are disclosed to the respective participating laboratory. The methods applied have limitations in reproducibility, thus, on repeated testing, the same strain/antimicrobial combination can result in different MIC or inhibition zone diameter values differing by one-fold dilution or ± 3 mm, respectively. If the expected MIC / zone diameter is close to the threshold for categorising the strain as susceptible, intermediate, or resistant, a one-fold dilution / ± 3 mm difference may result in different interpretations. Since this report assesses the interpretation of MIC/zone diameter rather than the actual values, some participants may find their results classified as incorrect (score of 0, 1 or 3) even though the actual MIC / zone diameter measured is only one-fold dilution / ± 3 mm apart from the expected MIC / zone diameter. In these cases, the participants should be confident in the high quality of their AST performance.

In this report, results from laboratories affiliated with the HH or AH sectors are presented separately. The laboratories are identified by codes and each code is known only by the corresponding laboratory and the organizers. The full list of laboratory codes is confidential and known only by the EQAsia consortium.

This report, in its final version, is approved by a Technical Advisory Group consisting of members from the EQAsia consortium, and by the EQAsia Advisory Board members Ben Howden (The Peter Doherty Institute for Infection and Immunity, Australia), Monica Lahra (WHO Collaborating Centre for STI and AMR, NSW Health Pathology Microbiology, New South Wales, Australia) and Russel Cole (Pacific Pathology Training Centre, New Zealand).