The role and implementation of next-generation sequencing technologies in the coordinated action plan against antimicrobial resistance.
Date: 2017-05-29
Institutions: Joint Research Centre
Subjects: action programme, antibiotic, disease prevention, epidemiology, medical research, medicinal product, medicine, public health, research report, technological process, veterinary medicine

The Role and Implementation of 
Next-Generation Sequencing 
Technologies in the Coordinated 
Action Plan against Antimicrobial 
Resistance  

JRC Workshop 

March 21th - 22nd, 2017 
Villa Borghi, Italy 

Angers A., Petrillo M., Patak A., Querci 
M., Van den Eede G. 

2017 

EUR 28619 EN

 
This publication is a Conference and Workshop report by the Joint Research Centre (JRC), the European 
Commission’s science and knowledge service. It aims to provide evidence-based scientific support to the 
European policymaking process. The scientific output expressed does not imply a policy position of the 
European Commission. Neither the European Commission nor any person acting on behalf of the Commission is 
responsible for the use that might be made of this publication. 

JRC Science Hub 
https://ec.europa.eu/jrc 

JRC106743 

EUR 28619 EN 

PDF 

ISBN 978-92-79-69020-4 

ISSN 1831-9424 

doi: 10.2760/745099 

Luxembourg: Publications Office of the European Union, 2017  

© European Union, 2017 

The reuse of the document is authorised, provided the source is acknowledged and the original meaning or 
message of the texts are not distorted. The European Commission shall not be held liable for any consequences 
stemming from the reuse. 

How to cite this report: Angers A., Petrillo M., Patak A., Querci M., Van den Eede G., The Role and 
Implementation of Next-Generation Sequencing Technologies in the Coordinated Action Plan against 
Antimicrobial Resistance, EUR 28619, doi: 10.2760/745099 

All images © European Union 2017, except: Cover page; ©Leigh Prather; source: Fotolia.com 

 
 
 
 
 
 
 
 
 
 
 
 
 
Contents 

Foreword .............................................................................................................. 1 

Acknowledgements ................................................................................................ 2 

1  Introduction ...................................................................................................... 3 

1.1  Monitoring the rise and spread of Antimicrobial Resistance................................ 3 

1.2  A potential role for Next-Generation Sequencing .............................................. 4 

2  Summary of the Workshop discussions ................................................................. 6 

2.1  The added value of NGS technologies in the AMR action plan ............................ 6 

2.2  Technical considerations and challenges in the implementation of NGS-based 

methods for AMR determination .................................................................... 7 

3  Conclusions .................................................................................................... 10 

References ......................................................................................................... 11 

List of abbreviations and definitions ....................................................................... 13 

Annexes ............................................................................................................. 14 

Annex 1. Proposed follow-up: Mapping the evidence on the risk related to AMR 

between the environment, food producing animals, the food chain and human 
health. ..................................................................................................... 14 

Annex 2. Proposed follow-up: Harmonisation efforts in the common aspects regarding 
the use of NGS in the detection of AMR common to environment, food producing 
animals, the food chain and human health. ................................................... 15 

Annex 3: Workshop agenda ............................................................................... 16 

i 

Foreword 

On the 21st and 22nd of March, a two-day workshop was organised by the Knowledge for 
Health  and  Consumer  Safety  Unit  of  the  Joint  Research  Centre,  on  the  Role  and 
Implementation of Next-Generation Sequencing Technologies in the European Action Plan 
against Antimicrobial Resistance. 

The workshop brought together 15 international experts representing the top expertise in 
the  use  of  NGS  to  detect  the  genetic  determinants  of  AMR  in  diverse  fields:  clinical, 
human health, animal health, food and environmental monitoring.  Representatives from 
the JRC, EMBL-EBI and standardisation authorities were also present in the discussions. 

The participating experts were: 

Johan Bengtsson-Palme - University of Gothenburg, Sweden 
Thomas Berendonk - Technische Universität Dresden, Germany 
Burton Blais - Canadian Food Inspection Agency, Canada 
Kok Gan Chan - University of Malaya, Malaysia 
Teresa M. Coque - Hospital Universitario Ramón y Cajal (IRYCIS), Spain 
Derrick Crook - University of Oxford, UK 
Matthew Ellington - Public Health England, UK 
Christoph Endrullat - German Institute for Standardization (DIN), Germany 
Dirk  Höper  -  Friedrich-Loeffler-Institut-  Federal  Research  Institute  for  Animal  Health, 
Germany 
Ole Lund - Technical University of Denmark, Denmark 
Jean  Yves  Madec  -  Agence  nationale  de  sécurité  sanitaire  de  l’alimentation,  de 
l’environnement et du travail, France 
Alan McNally - University of Birmingham  
Thierry Naas - Hôpital de Bicêtre- Service de Bactériologie, France 
Justin O'Grady - University of East Anglia, UK 
Jessica Vamathevan- European Bioinformatics Institute (EMBL-EBI), UK 

Chair: 

Guy Van den Eede - Head of Unit Knowledge for Health and Consumer Safety, JRC. 

Other participants from the Joint Research Centre: 

Alexander Binder - Health in Society Unit, JRC 
Alessia Bogni - Consumer Products Safety Unit, JRC 
Dafni Kagkli - Fraud Detection & Prevention Unit, JRC 
Teresa Lettieri - Water and Marine Resources Unit, JRC 
Valentina Paracchini - Fraud Detection & Prevention Unit, JRC 
Cristian Savini - Food & Feed Compliance Unit, JRC 
Heinz Schimmel - Reference Materials Unit, JRC 

Organising committee: 

Alexandre Angers - Knowledge for Health and Consumer Safety Unit, JRC 
Mauro Petrillo - Knowledge for Health and Consumer Safety Unit, JRC 
Maddalena Querci - Knowledge for Health and Consumer Safety Unit, JRC 
Alex Patak - Knowledge for Health and Consumer Safety Unit, JRC 

1 

 
 
 
Acknowledgements 

We  are  grateful  to  our  colleagues  Dagmar  Tielemans,  Virginie  Tregoat,  Gabriela  Ferrari 
and Rossella Speroni for their invaluable support in organising the workshop. 

2 

 
 
1  Introduction 

1.1  Monitoring the rise and spread of Antimicrobial Resistance 

Since  their  discovery,  antimicrobials  have  played  an  essential  role  in  the  treatment  of 
infections  and  have  significantly  improved  the  population's  health.  However,  the  rise  of 
antimicrobial  resistance  (AMR,  i.e.  the  ability  of  a  microorganism  to  resist  the  action  of 
an antimicrobial agent) now endangers the status quo of our healthcare system. 

Evidence of antibiotic resistance is growing. For example, an alarming development was 
reported last year for one of the last-resort antibiotics, namely colistin, where a form of 
resistance  was  discovered  that  could  be  readily  transferred  to  other  bacteria  (Liu  et  al., 
2016). Researchers first discovered this resistance in China, quickly followed by findings 
–  including  by  the  JRC  (Petrillo  et  al.,  2016)  -  of  similar  resistance  patterns  in  other 
countries,  including  in  Europe.  The  spread  of  multiple-drug  resistant  bacteria  already 
causes  an  estimated  25.000  deaths  annually  in  Europe  alone,  a  toll  that  is  expected  to 
increase1. 

For  many  years,  the  European  Union,  as  well  as  other  countries  and  international 
organisations, have been addressing the issue of the rise and spread of AMR. Their work 
includes the establishment of collaborative programs that raise public awareness and aim 
to  align  international  actions  in  order  to  maximise  their  efficacy.    These  actions  include 
the  promotion  of  research  and  innovation  to  identify  new  antimicrobial  compounds,  the 
prevention of disease to minimise antimicrobial needs, and public awareness about their 
correct use.   

These  efforts  depend,  among  other  things,  on  the  establishment  of  an  efficient 
monitoring  and  surveillance  scheme,  implemented  in  a  coordinated  and  international 
framework.  This  scheme  is  crucial  for  understanding  the  development  and  diffusion  of 
resistance  in  order  to  provide  relevant  risk  assessment  data  and  evaluate  the 
effectiveness of targeted interventions (see Box 1). 

Box 1.  AMR surveillance  

The importance of an efficient framework for AMR surveillance and the efforts needed for 
its improvement have been stressed in the recent years.   

For  example,  the  WHO's  2014  Antimicrobial  Resistance  Global  Report  on  Surveillance 
highlighted  the  existence  of  gaps  in  the  methodology  for  integrated  surveillance  of 
resistance in human and foodborne pathogens and the need for the development of tools 
and standards for harmonized surveillance of AMR (World Health Organization, 2014).  

In October 2015, a Declaration of the G7 Health Ministers stated that "We consider that a 
lack of comparable data on the quantity and kind of use of antibiotics and the prevalence 
of  AMR  in  the  population  results  in  an  incomplete  understanding  of  the  antibiotic 
resistance  situation.  The  availability  of  comparable  international  and  national  data  is  a 
pre-condition for targeted action within countries"  
(http://www.g8.utoronto.ca/healthG8/2015-berlin.html) 

Within the EU, data on surveillance of AMR in human health are compiled and released by 
the European Antimicrobial Resistance Surveillance Network (EARS-Net)2 as part of ECDC 
surveillance.  A  “One  Health”  perspective  is  also  encouraged,  evaluating  the  impact  of 

1 http://ec.europa.eu/dgs/health_food-safety/docs/amr_factsheet.pdf 
2 http://ecdc.europa.eu/en/healthtopics/antimicrobial-resistance-and-

consumption/antimicrobial_resistance/EARS-Net/Pages/EARS-Net.aspx 

3 

 
 
 
 
                                           
 
antimicrobial resistance in humans, food-producing animals and food; in this optic, every 
year,  EFSA  and  ECDC  publish  the  European  Union  summary  report  on  antimicrobial 
resistance  in  zoonotic  and  indicator  bacteria  from  humans,  animals  and  food  (see,  for 
example,  European  Food  Safety  Authority  and  European  Centre  for  Disease  Prevention 
and Control, 2017). 

In these efforts, the definition of AMR refers to clinical or epidemiological cut-off (ECOFF) 
breakpoints  defined  by  the  European  Committee  on  Antimicrobial  Susceptibility  Testing 
(EUCAST)3. Their significant implementation across European laboratories  is an essential 
component of the consistency required for clinical reporting of antimicrobial susceptibility 
results (Brown et al., 2015).  These methods are "phenotypic", meaning that they rely on 
observing the extent to which live bacteria are affected by the antimicrobials.  

1.2  A potential role for Next-Generation Sequencing 

At  the  end  of  2014,  a  report  from  the  "The  Review  on  Antimicrobial  Resistance"  stated 
that  "advances  in  genetics,  genomics  and  computer  science  will  likely  change  the  way 
that  infections  and  new  types  of  resistance  are  diagnosed,  detected  and  reported 
worldwide, so that we can fight back faster when bacteria evolve to resist drugs".4  

Indeed,  the  potential  to  predict  antibiotics  resistance  of  bacteria  by  determining  the 
sequences  of  their  genomes  and  the  plasmids  they  host  has  long  been  discussed  (see 
Box 2). With the advent of Next-Generation Sequencing (NGS) technologies, our modern 
capability  to  generate  a  wealth  of  nucleic  acid  sequence  information  -  when  coupled  to 
the  appropriate  bioinformatics  information  systems  -  allows  both  a  profiling  of 
microorganisms  (as  single  clones  or  as  a  community)  and  the  detection  of  potential 
antimicrobial activities in a single experiment. 

Box 2. Detection of AMR using DNA sequencing 

An  important  aspect  for  the  monitoring  of  antimicrobial  resistance  using  sequencing-
based  methods  (compared  to  phenotypic  approaches)  is  the  importance  to  understand 
and  take  into  account  the  molecular  mechanisms  of  these  resistances.  Several 
mechanisms have been described in the literature, and include: 

1. The production of an enzyme that digests/metabolizes the antimicrobial. 

2. The production of efflux pumps that remove the drug(s) from within the cell. 

3. The modification, through mutations, of the intracellular target of the antimicrobial so 
that their interaction is lost. 

4.  The  activation/up-regulation  of  alternate  pathways  that  allow  survival  through  the 
bypass of the pathway disrupted by the antimicrobial. 

5. The down-regulation of the expression of the pores through which the drug enters the 
bacteria. 

Mechanisms 1-3 generally involve modifications in the pathogen’s DNA sequence and/or 
content  through  horizontal  gene  transfers  or  specific  mutations  in  the  endogenous 
genome sequence. As such, these modifications can most probably be efficiently detected 
by  sequencing-based  methods.  Mechanisms  4  and  5,  on  the  other  hand,  represent 
environmental  adaptation  through  signal  transduction  pathways  and  their  detection 
through non-phenotypic approaches will likely present a case-by-case challenge.  

In  the  recent  years,  a  wealth  of  scientific  articles  have  been  published,  describing 
development  and  trials  of  NGS-based  methods  in  the  determination  of  AMR  in  various 
contexts, both for the clinic (see, for example, Bradley et al., 2015; Sherry et al., 2013; 
Votintseva  et  al.,  2017),  food  (see,  for  example,  Hasman  et  al.,  2015),  and  the 

3 http://www.eucast.org/fileadmin/src/media/PDFs/EUCAST_files/Resistance_mechanisms/ 

EUCAST_detection_of_resistance_mechanisms_v1.0_20131211.pdf 

4 "Antimicrobial Resistance: Tackling a crisis for the health and wealth of nations", December 2014 

4 

 
                                           
environment  (see,  for  example,  (Bengtsson-Palme  et  al.,  2014,  2016;  Noyes  et  al., 
2016). 

In  June  2016,  in  its  conclusions  on  the  next  steps  under  a  “One  Health”  approach  to 
combat  antimicrobial  resistance,  the  Council  of  the  European  Union  called  upon  the 
Commission  and  the  Member  States  to  "align  surveillance  on  AMR  in  humans,  food, 
animals and environment at EU level"5. 

This  mention  of  environmental  surveillance  on  AMR  is  a  challenge  with  the  currently 
available  methods.  Phenotypic  interpretation  of  AMR  for  environmental  bacteria  is 
difficult, largely because there are no guidelines for resistance due to their lack of clinical 
relevance  and  difficulty  (if  not  impossibility)  in  culturing  them.  Still,  AMR  in  non-
pathogenic  environmental  bacteria  could  be  a  crucial  factor  in  the  development  of 
resistance pathogens due to the potential of horizontal gene transfer of resistance genes 
from non-pathogenic to pathogenic hosts. 

In summary, in the framework of Antimicrobial Resistance detection and monitoring, NGS 
technologies have the potential to: 

  Provide  an  harmonised  link  between  the  surveillance  in  the  environment  and  in 
the other important aspects of the  “One  Health” approach (clinic, food and  food-
producing animals) 

  Provide added value to the monitoring currently established in each of these fields 

individually. 

For this reason, a Workshop was organised in order to discuss the potential impacts NGS 
technologies  could  have,  specifically,  on  the  current  international  action  plans  against 
AMR, as well as to understand the next steps for their development and implementation 
in this context. 

5 http://www.consilium.europa.eu/en/press/press-releases/2016/06/17-epsco-conclusions-antimicrobial-

resistance/ 

5 

 
 
 
 
                                           
2  Summary of the Workshop discussions 

Structured in three distinct sessions, the workshop addressed: 

1.  The added value of NGS technologies in the AMR action plan 

2.  Technical considerations in the determination of AMR using NGS technologies 

3.  Challenges  in  the  implementation  of  eventual  NGS-based  methods  for  AMR 

determination 

Because  of  a  large  overlap in  the  discussions  of  sessions  2  and  3,  they  are  reported  as 
one in this document. 

2.1  The added value of NGS technologies in the AMR action plan 

Next-Generation Sequencing, also known as high-throughput sequencing, is the catch-all 
term used to describe a number of different modern technologies that allow nucleic acids 
sequences to be generated much more quickly and cheaply than previously. Coupled with 
Bioinformatics, NGS has revolutionised the study of genomics and microbiology. Applied 
to  the  prediction  of  antimicrobial  resistance  of  an  unknown  isolate  or  within  an 
environmental sample, it could provide many significant advantages to the currently used 
methods, including: 

It can contribute to clinical decision making.  

In  a  clinical  setting,  sequencing-based  approaches  the  potential  to  provide  different 
levels of information that can guide treatment with the appropriate antimicrobials.  These 
include a) whether the infection is bacterial or viral, b) the type of bacteria, allowing the 
development  and  use  of  narrow  spectrum  antimicrobials,  c)  the  eventual  presence  of 
genetic  determinant  of  resistance  and  d)  the  actual  predicted  susceptibility  to 
antimicrobials  of  the  isolate.    Efforts  are  still  necessary  and  ongoing,  in  particular  for 
points c) and d). 

The  time  to  complete  the  analyses  remains  an  important  factor  with  most  sequencing 
technologies.    This  is  not  a  concern  for  infections  with  slow-growing  bacteria,  such  as 
tuberculosis,  for  which  the  advantages  of  sequencing  over  culture-based  assays  makes 
no  doubt  (see  Votintseva  et  al.,  2017;  Walker  et  al.,  2015).    For  faster-growing 
pathogens,  the  possibility  to  fully  replace  phenotypic  testing  remains  to  be  evaluated, 
despite high accuracy in reported prediction rates. 

Advances  in  sequencing  technologies  also  shorten  of  time  between  sample  acquisition 
and AMR prediction.  For example, studies showed that, in the treatment of patients with 
urinary  tract  infections,  metagenomics  analyses  on  nanopore  sequencing  data  provided 
information to adapt antimicrobial treatment in time for the second dose, within 8 hours 
of the first (Schmidt et al., 2017).   

It  provides  information  (in  addition  to  AMR  predictions)  that  can  help  in 
understanding outbreaks and guide intervention.  

NGS is, by its technical nature, a technology able to provide a "complete" set of data on 
the  genetic  material  in  the  analysed  sample.    The  same  data  can  then  be  analysed, 
through separate bioinformatics processes, to answer different questions.  

If  established  around  NGS,  the  same  diagnostics/monitoring  framework  can  provide 
information on many crucial aspects in addition to  the resistance determinants for AMR, 
such as epidemiological typing for outbreak investigation, organism species and virulence 
factors of clinical relevance (Eyre et al., 2012; Quick et al., 2015).  

By storing WGS sequence data, it is possible to retroactively analyse when new 
information appear. 

6 

 
 
This "completeness" of information includes the possibility for future analyses which were 
not  planned  or  known  to  be  relevant  at  the  time  the  samples  were  sequenced,  such  as 
the  rise  and  spread  of  new  AMR.  This  was  seen  with  the  recent  discovery  of  the  mcr-1 
gene  in  colistin  resistance  and  its  retrospective  monitoring  in  established  genome 
sequence databases (Falgenhauer et al., 2016; Hasman et al., 2015) 

It provides mechanistic information about the resistance. 

Unlike  phenotypic  tests  that  provide  information  only  regarding  resistance/susceptibility 
to antimicrobials, NGS can reveal the molecular basis for this resistance. This information 
can feed in monitoring schemes, helping to understand the events leading to acquisition 
of resistance. In addition, NGS can characterise novel resistance mechanisms when they 
arise, through sequencing of isolates that are phenotypically proven to be resistant.  This 
is a remarkable added value if compared to  other nucleic-acid based techniques such as 
the polymerase chain reaction (PCR).  

It  is  a  technology  that  has  potential  to  link  the  different  fields:  clinics, 
environment, food and animals. 

Standardised  methods  for  AMR  monitoring  that  are  specifically  applicable  to  the 
environment  have  never  been  developed;  culture-dependent  methods  established  for 
clinical  samples  can't  readily  be  applied  to  environmental  samples  (Berendonk  et  al., 
2015)  since  the  numbers  of  isolates  necessary  for  the  tests  endpoints  are  different  (set 
of  species/prevailing  species),  most  environment  bacteria  are  not  recovered  in  culture 
conditions  and  established  criteria  are  not  applicable  (the  main  objective  of  these  tests 
being  to  identify  likelihood  of  therapeutic  failure).  In  addition,  AMR  in  non-pathogenic 
environmental  bacteria  (for  which  there  are  no  guidelines)  is  relevant  due  to  the 
possibility of horizontal gene transfer.  

In  order  to  achieve,  as  described  by  the  Council  of  the  European  Union,  a  coordinated 
surveillance  of  AMR  in  humans,  food,  animals  and  environment  at  the  EU  level,  new 
technologies  are  needed  on  which  to  base  the  framework.    For  this,  NGS  is  a  strong 
candidate, as extensive work is currently being done with this technology in all the fields. 

Data accumulation allows better understanding and improvement of the system. 

The information available from a set of whole genome sequences grows as the amount of 
available information increases.  Building a monitoring framework on NGS will thus allow 
continuous self-improvement of the whole framework. 

2.2  Technical considerations and challenges in the 

implementation of NGS-based methods for AMR determination 

Recently,  numerous  articles,  reports  and  guidelines,  both  in  the  scientific  literature  and 
regulatory frameworks, have been published assessing and promoting the use of NGS for 
pathogen detection in the clinic, environment and food chain. Although these documents 
often include AMR determination in their discussion, it is becoming clear that the use of 
NGS  for  AMR  has  its  own  -  and  not  always  overlapping  -  set  of  technical  and 
implementation challenges. 

The workshop brought together experts having approached the specific problem of using 
NGS  for  detecting  the  genetic  determinants  of  AMR  in  the  different  areas.    From  these 
discussions, it became apparent that: 

Each field has its own set of technical challenges and realities. 

Despite  many  studies  reporting  the  use  of  a  common  technology,  NGS,  for  AMR 
determination, it is important to note that there exist very specific sets of requirements, 
difficulties and gaps that will need to be addressed on a field-by-field basis.  Outside of a 
simple  "monitoring"  framework,  additional  needs  can  include,  for  example,  proper  risk 

7 

 
 
assessment  (food,  environment)  and  correct  prediction  of  therapeutic  success  (human 
and veterinary medicine). 

On  a  technical  basis,  considerations  of  varying  relevance  include  whether  the  detected 
AMR gene is expressed or not, the presence of the resistance in the chromosome or in a 
plasmid,  understanding  interaction  of  bacteria  harbouring  the  resistance  with  other 
bacteria,  the  limit  of  detection  of  the  whole  methodology  (including  the  sample 
preparation), etc.   

There  are  points of  contact  between  the  different  fields,  where  the  “problems” 
to solve are similar.  

For all these differences, a "core" problem emerged that is common to all fields, which is 
the need to correctly and reliably identify the known genomic determinants of AMR from 
a set of NGS reads produced from the whole genome sequencing of a  sample/isolate. 

Different approaches for this have been and are being developed and used.  There would 
be a lot to gain in involving, in future activities, scientists and experts active in this type 
of work from the different fields in order to share experience and identify best practices. 
Common challenges in this aspect include: 

  Identify  a  "best  practice"  bioinformatics  strategy  and  implementation. 
Define  what  approach 
to  use  (shotgun  metagenomics,  short/long  read 
technologies,  assembly  or  single  reads  analysis,  …);  minimal  metadata 
requirements;  how  to  determine  "true"  outcome  to  which  the  results  should  be 
compared when evaluating a strategy; minimum recommended sequencing depth; 
etc. 

  What reference database to be used in the analysis. A lot of resources exist, 
such  as  ResFinder,  CARD  and  SRST2  (see  Xavier  et  al.,  2016),  which  are  often 
complemented  with  in  house  databases.    The  scope  and  quality  of  the  reference 
database  affects  the  outcome  as  the  AMR  screen  will  only  return  genetic 
determinants  you  are  looking  for.  A  recent  review  from  a  EUCAST  subcommittee 
identified  the  establishment  of  a  sustainable  reference  database  as  one  of  the 
main  recommendations  towards  the  use  of  NGS  for  bacterial  antimicrobial 
susceptibility testing (Ellington et al., 2017). 

  A  harmonised  way  to  record  and  share  information.  Sharing  schemes  exist 
and  produce  benefits,  in  particular  collaborations  between  national  reference 
centres.    However,  the  situation  varies  between  countries,  and  the  existing 
collaborations rely on ad hoc networks.  In this aspect, the COMPARE project, an 
EU  project  (funded  by  Horizon  2020)  whose  aim  is  to  speed  up  the  detection  of 
and  response  to  disease  outbreaks  among  humans  and  animals  through  the  use 
of NGS, is expected to play an important role.  

requirements 

  Appropriate  quality  controls/reference  materials/harmonisation.  There  is 
currently no comprehensive quality management approach in NGS present which 
includes  necessary 
for  proper  documentation,  containing 
standardised  information  about  identified  AMR  resistance  genes,  for  example.  
Several  NGS  standardization  efforts  have  been  done  by  American  work  groups, 
authorities  and  societies,  thus  the  majority  of  standardisation  efforts  has  been 
taken  place  in  US.  In  addition,  Illumina  Inc.,  an  US-based  sequencing  company, 
strives  towards  the  establishment  of  internal  developed  standards  due  to  the 
current  held  position  as  the  market  leader  in  NSG.  However,  due  to  the  strong 
presence  and  importance  of  European  bodies  like  CEN  (European  Committee  for 
Standardization), the standardisation of NGS will become inspired and lifted up on 
the EU level in the future. There exists a high demand for NGS standardisation in 
clinical  diagnostics,  including  AMR  detection  in  a  clinical  setting.  However,  the 
same standards which will be established there, will not be necessarily applicable 
or  reasonable  in  other  application  like  food  chain  or  environment.  Hence,  there 
are  two  options  possible.  First,  the  development  of  one  common  standard  for  all 

8 

 
possible  applications  or  second,  the  primary  development  of  NGS  standards  in  a 
clinical  setting,  which  could  become  translated  into  other  applications  like 
environment or food chain later on. The decision for one of these options will rely 
on  standardisation  bodies  and  the  involved  experts.    Efforts  in  this  direction 
should  take  into  account  specificities  of  the  field  such  as  fast  obsolescence  of 
systems  and  technologies  (e.g.  454  and  SOLiD),  possibly  by  focusing  on  setting 
up  quality  metrics  (mainly  addressing,  in  a  first  instance,  NGS  performance 
characteristics)  and  best  practices  rather  than  restrictive  prescriptions  (Endrullat 
et al., 2016; Lambert et al., 2017). 

Efforts will also be needed in the optimisation and, possibly, harmonisation of the early 
steps of the workflow - such as a sample preparation, DNA extraction and, if needed, 
target enrichment - as these are expected to have a strong impact on the results.  

9 

 
 
 
3  Conclusions 

Apart  from  the  specific  conclusions  linked  to  the  different  topics,  one  of  the  major 
recognised impacts of the workshop was that the experts involved in the different fields 
appreciated  the  opportunity  to  meet  and  interact  with  experts  of  other  fields  of 
applications.  

In  the  monitoring  of  AMR  in  general  and  in  the  implementation  of  NGS  technologies  in 
particular,  each  field  has  its  own  set  of  technical  challenges,  requirements  and 
realities.   However,  it  emerged  that  there  are  points  of  contacts  between  them,  where 
the “problems” to solve are similar.  

In this optic, one of the major requests from the participants was for the JRC to follow up 
this  type  of  discussion  in  a  cross-discipline  setup,  in  order  to  identify,  discuss  and 
possibly harmonise the elements which are common to the different fields. 

For  some  fields,  such  as  human  and  animal  pathogen  monitoring  (including  antibiotics 
resistance), some countries, such as France and the UK, reported ongoing collaborations 
with  evident  benefits  for  public  health  and  for  the  management  of  outbreaks.   It  was 
suggested  that  understanding  (mapping)  the  situation  in  the  different  countries,  with  a 
possible expanded sharing and coordination effort, would be greatly beneficial. 

It  was  highlighted  that  the  legislation  in  the  different  areas  do  not  always  take  into 
accounts  the  importance  of  AMR  monitoring  and  its  translation  into  intervention,  in 
particular in the environment. A need was identified to better understand the abundance 
and  processes  of  AMR  spread  and  evolution  in  the  environment  (and  its  relation  to 
environmental and human health). 

For all the effort involved, the experts emphasised the potential benefits of using NGS for 
AMR determination. 

A list of potential follow-up activities has been suggested, as summarised in the Annexes. 

10 

 
 
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(2013).  Outbreak  Investigation  Using  High-Throughput  Genome  Sequencing  within  a 
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List of abbreviations and definitions 

AMR 

Antimicrobial Resistance 

ECDC  European Centre for Disease Prevention and Control 

EFSA 

European Food Safety Authority 

EUCAST European Committee on Antimicrobial Susceptibility Testing 

NGS 

Next-Generation Sequencing 

13 

 
Annexes 

Annex 1. Proposed follow-up: Mapping the evidence  on the risk related to AMR 
between  the  environment,  food  producing  animals,  the  food  chain  and  human 
health. 

Rationale: An important element in correctly designing and implementing a surveillance 
scheme for AMR  spread that would span humans, food, animals and  the environment is 
to  understand  the  risk  associated  with  the  spread  of  the  genetic  determinants  of  AMR 
between the different frameworks. 

Proposal:  An  expert  group  should  be  set  up  to  review  the  available  evidence  (see  for 
example Ohidul and Tianlin; Williams-Nguyen et al., 2016), assess the demonstrated and 
potential  risks  and  identify  existing  gaps  in  the  current  understanding  of  the  impact  of 
antimicrobial resistance spreading between humans, food, animals and the environment.  
A report would be produced and presented to the appropriate policy makers. 

Possible  impacts:  Mapping  this  evidence  and  understanding  when  there  is  sufficient 
demonstrated risk to justify intervention (and understanding, if not, whether to apply the 
precautionary  principle)  will  help  the  development  of  proportional  AMR  monitoring 
schemes,  in  particular  for  the  environment  which  is  currently  not  well  developed  nor 
encouraged.    It  could  also  lead  to  the  establishment/strengthening  of  official  networks 
between  the  existing  clinic,  food  and  veterinary  frameworks  in  the  different  Member 
States.  It  may  also  affect  existing  risk  assessment  regulatory  frameworks  for 
environmental  policies  (e.g.  the  Water  Framework  Directive  2000/60/EC  and  the 
Committee  for  Medicinal  Products  for  Human  Use),  that  currently  evaluate  safe  level  of 
antimicrobials in the environment based on their toxicity and do not take into account the 
emergence  of  AMR,  which  is  expected  to  occur  at lower  concentrations  (see  Bengtsson-
Palme and Larsson, 2016). 

14 

 
 
 
 
 
 
 
 
Annex  2.  Proposed  follow-up:  Harmonisation  efforts  in  the  common  aspects 
regarding the use of NGS in the detection of AMR common to environment, food 
producing animals, the food chain and human health. 

Rationale:  In  view  of  the  identified  points  of  contacts  between  the  “problems”  faced 
when  using  NGS  to  identify  genetic  determinants  of  AMR  whatever  the  context  (clinic, 
food  control,  environmental  monitoring…),  there  would  be  added  value  in  continuing 
discussions across the different frameworks. 

Proposal: Follow-up discussions should be organised, with the appropriate format to be 
determined (workshops, working groups …) in view of harmonising, as much as possible 
and useful, the common steps and resources.  This could involve: 

 

  Mapping  existing  networks  in  different  areas  (AMR/metagenomics  for  human 
health,  animal  health,  food  monitoring  and  environmental  monitoring,  AMR 
sequence databases, …).  When missing, such a network could be  initiated. (E.g. 
application of metagenomics for human health) 
Invite  representatives  from  these  networks  (in  particular,  bioinformaticians)  to 
identify  and  discuss  the  elements  which  are  truly  common  between  the  different 
frameworks  (to  confirm/complement 
the  present 
workshop).  
Identify  the  best  forum  to  proceed  with  efforts  to  identify  best  practices,  quality 
aspects, reference databases, etc. 

identified  during 

those 

 

Possible  impacts:  With  the  current  impetus  for  the  eventual  establishment  of  an 
efficient and useful AMR monitoring framework that combines all the different aspects of 
the  “One  Health”  initiative  and  the  environment,  it  will  be  invaluable  that  the  elements 
that can be harmonised are harmonised. 

15 

 
 
 
 
 
 
Annex 3: Workshop agenda 

16 

 
 
 
JRC WORKSHOP:  THE ROLE AND IMPLEMENTATION OF NEXT-GENERATION SEQUENCING 
TECHNOLOGIES IN THE COORDINATED ACTION PLAN AGAINST ANTIMICROBIAL 
RESISTANCE. 

JRC Ispra, 21-22 March, 2017 

 1st day: 21 March 2017 

9:30   - 10:00  Arrival and coffee 

 2nd  day: 22 March 2017 

Session 3: Implementation 

10:00 - 10:30  Welcome and opening remarks  

9:30  -  10:45 

Invited presentations (15 min each), followed by round table  
discussions. 
Introductory presentations by: 
Dr Burton Blais, Canadian Food Inspection Agency, Canada 
Dr Christoph Endrullat, German Institute for Standardization (DIN),  
Germany 
Dr Jessica Vamathevan, European Bioinformatics Institute (EMBL- 
EBI), UK 

10:45-11:15 

coffee break   

11:15  - 12:30  Continuation of discussions 

12:30-13:30 

LUNCH BREAK  

Session 4: Conclusions and recommendations 

13:30  - 15:00  Summary of the conclusions from the different sessions. Identification 

of the next steps and follow-up activities  

15:00 

End of Workshop 

10:30 - 12:30 

Setting the scene: background, format & aim of the workshop 
(Guy Van den Eede, JRC) 

Session 1: Added value of NGS technologies in the AMR action plan 

Invited presentations (15 min each), followed by round table  
discussions  
Introductory presentations by: 
Dr Kok Gan Chan, University of Malaya, Malaysia 
Dr Thierry Naas, Hôpital de Bicêtre- Service de Bactériologie, France 
Dr Justin O'Grady, University of East Anglia, UK 

12:30 - 14:00  LUNCH BREAK  

Session 2: Technical considerations 

14:00  - 15:00 

Invited presentations (15 min each), followed by round table  
discussions  
Introductory presentations by: 
Dr Thomas Berendonk, Technische Universität Dresden, Germany  
Dr Derrick Crook, University of Oxford, UK 
Dr Dirk Höper, Friedrich-Loeffler-Institut- Federal Research Institute 
for Animal Health, Germany 
Dr Ole Lund, Technical University of Denmark, Denmark 

15:00-15:30 

coffee break   

15:30  - 17:30  Continuation of discussions 

17:30 

20:00 

End of day 1 

WORKSHOP DINNER (Villa Borghi)  

 
 
  
 
 
 
 
 
 
 
 
  
 
 
 
 
 
 
 
 
 
 
  
 
 
 
  
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
  
  
 
  
 
 
 
 
 
 
 
  
 
 
 
 
 
 
 
 
 
 
 
 
 
 
  
  
  
 
  
 
  
 
 
  
 
  
 
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