significance and power to detect a difference. The European Food Safety Authority has compiled tables showing required sample sizes for different antimicrobial resistance monitoring programme
objectives 11.
1.6. Laboratory testing methodology
Laboratories providing isolates or antimicrobial susceptibility results for a programme of integrated surveillance of antimicrobial resistance in foodborne bacteria should be able to
isolate target bacteria from different specimen types, and identify bacteria to the genus and species levels using internationally accepted microbiological methods. It will also be helpful for
participating laboratories to be able to determine pathotypes of E. coli using validated methods and perform antimicrobial susceptibility testing using validated methods according to established
standards such as those of the Clinical and Laboratories Standards Institute CLSI or the European Committee on Antimicrobial Susceptibility Testing EUCAST. It is also helpful for
participating laboratories to be able to or have access to a reference laboratory that is able to determine serotypes of Salmonella and characterize isolates using molecular methods such as
PCR and sequencing, PFGE, multilocus sequence typing MLST and whole genome sequencing WGS. WHO capacity building activities, such as the WHO Global Foodborne Infections Network
GFN or AGISAR projects, help provide technical support and training in food microbiology to participants. In addition, participation in an external quality assurance programme EQAS, such
as WHO GFN’s EQAS 12,13 or others, is recommended.
1.6.1 Bacterial culture and isolate identification
Different recovery methods can be used based on the type of samples e.g. food samples which can differentially enrich bacterial subpopulations within a sample. However, it is important
to be aware that media selecting for resistance e.g. broth with cefotaxime to select for ESBL- producing bacteria leads to different recovery rates than non-selective media. Culture methods
and media should meet recognized international laboratory standards. As with other design considerations, culture methods should be defined beforehand and be described in surveillance
reports. Differences in culture methodology should be taken into account when data from different surveillance programmes are compared. Monitoring laboratories are encouraged to
collaborate with established monitoring systems, national reference laboratories, WHO Collaborating Centres and other partners to provide long-term storage for a representative
number of isolates that can be used for future testing and analysis. Bacteria should be identified to the species level using standard methods conventional or automated biochemical tests,
matrix-assisted laser desorptionionization time-of-flight mass spectrometry [MALDI], or any other validated methods. For some species, it will be necessary to identify the bacteria to the
type level e.g. serotype, MLST, etc..
1.6.2 Standardized antimicrobial susceptibility testing
Only in vitro antimicrobial susceptibility testing methods that have been standardized and validated under the auspices of an internationally recognized consensus standards organization,
such as CLSI or EUCAST, should be used. This is a critical feature of a sound antimicrobial resistance surveillance programme, and is the only way to ensure reliable data. The steps in
these official standards should be strictly followed and should not be modified for local use. Standard breakpoints for other bacteria of interest not described in this guidance can be found
in CLSI documents; M100S, M45, or VET01-S2 or via the web site of the EUCAST
2
. See Appendix 1 for a description of the differences between the two systems.
1 2
