engineering to human health and the environment are covered by Directives 2001/18/EC and 2009/41/EC and the
guidance notes published in Commission Decision 2000/608/EC [16-18].
Event background information
On 24 March 2017, the Bayerisches Landesamt für Gesundheit und Lebensmittelsicherheit (LGL, Bavarian Health
and Food Safety Authority) issued a press release warning about the risk of exposure to several pathogenic
bacteria associated with the use of a specific DIY bacterial gene engineering CRISPR kit manufactured by a US
company [19]. According to the manufacturer, the kit allows users to perform a molecular biology experiment to
make a
‘genome mutation (K43T) to the
rps
L gene, changing the 43rd amino acid, a lysine (K) to a threonine (T),
thereby allowing the bacteria to survive on a kanamycin and streptomycin culture media which would normally
prevent its growth’ [20].
The kit is based on CRISPR–Cas9 system. To perform the experiment, the kit should contain the Cas enzyme (i.e.
endonuclease protein), exogenous genetic material (i.e. RNA and DNA sequences), and a recipient bacterium able
to acquire the exogenous genetic material [20]. The recipient bacterium included in the kit is described as a
harmless laboratory strain of
Escherichia coli (E. coli)
HME63
.
However, analysis performed by LGL on two kits
ordered from the US in November 2016 and in March 2017 revealed the presence of the facultative pathogenic
bacteria of risk group 2.
According to the Genetic Engineering Act in Germany, DIY genetic engineering kits should only be used in genetic
engineering installations [21]. Moreover, the risk-group-2 bacteria that were detected in two kits are pathogens
according to the German Infection Protection Act. Their import, export, storage or handling require permission by
the competent authority [22]. Consequently, LGL recommends not opening the kit, not to perform the
experiments, and to contact the public health authority [19,23,24]. After detecting risk-group-2 bacteria in the kits,
the LGL stated that the risk of infection cannot be excluded during the manipulation of these kits, even if the
spread of an infection in the general population remains unlikely [19].
ECDC threat assessment for the EU
Risk of infection for users of the contaminated kit
The isolates of
Klebsiella pneumoniae,
Enterobacter
spp. and
Enterococcus faecalis
that have been detected as
contaminants in the kits are bacterial species that are part of the normal human gastrointestinal as well as
environmental microbiome. They are also facultative pathogens with the ability to cause severe human infections
in certain conditions, especially in healthcare settings or when natural host defences are impaired. They belong to
biological risk group 2, a group which requires handling in containment facilities with appropriate safety measures
(e.g. worker protection) (Directive 2000/54/EC) [25].
In the ECDC point prevalence survey of healthcare-associated infections 2011–2012,
Enterococcus
spp.,
K. pneumoniae
and
Enterobacter
spp. were the third, fifth and eighth most frequently encountered microorganisms
associated with healthcare-associated infections in European acute care hospitals [26]. The most frequently
associated infections were surgical site and urinary tract infections for
Enterococcus
spp., respiratory tract and
urinary tract infections for
K. pneumoniae
, and surgical site and lower respiratory tract infections for
Enterobacter
spp. [26]. In addition, some of the
K. pneumoniae
and
Enterobacter
spp. isolates detected in the kits were
multidrug-resistant and ESBL producing bacteria that lead to resistance to penicillins and cephalosporins. The
detection of imipenem-resistant
K. pneumoniae
likely implies the carriage of carbapenemase genes encoding
resistance to the last-line agents that are used in hospitals for treatment of severe infections.
The risk of infection for users of the kits who are unaware of the contamination with pathogenic agents is low
because the manipulation of the kit does not involve percutaneous injury-prone manipulations. However, infection
resulting from the contamination of broken skin or mucous membranes may occur, even though the kit
recommends and provides disposable gloves. Furthermore, the kit includes lyophilised materials that need to be
reconstituted, which may lead to contamination of the mucosae of the eyes, mouth and nose. Finally, the risk of
infection may be increased for immunocompromised or immunosuppressed persons.
Risk of releasing and spreading antimicrobial-resistant
strains into the human population and the environment
The ESBL and carbapenemase resistance genes carried by the pathogenic bacteria present in the contaminated kits
are usually located on plasmids which can be transmitted easily between bacteria of the same species and to other
bacteria of the same family of
Enterobacteriaceae
that inhabit the human gastrointestinal tract. Bacteria with
resistance can persist for several months in the intestinal tract of asymptomatic carriers. If a carrier develops