ENVIRONMENTAL RISK MANAGEMENT
AUTHORITY DECISION
Amended under s67A on 16 August 2007
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3 May 2005
GMD05039
 To develop in containment genetically modified organisms
under section 40(1)(b) of the Hazardous Substances and New
Organisms (HSNO) Act 1996.
 Victoria University of Wellington
Applicant
 To develop in containment genetically modified bacteria to
Purpose
study the mechanisms of growth, adaptation and survival in
mycobacteria
20 18 April 2005
Date received
Consideration date 22 3 May 2005
Chief Executive, ERMA New Zealand
Considered by
Application code
Application type
1 Summary of decision
1.1
The application to develop a new organism in containment is approved, with
controls, having been considered in accordance with the relevant provisions of
the Hazardous Substances and New Organisms (HSNO) Act 1996, the HSNO
(Low-Risk Genetic Modification) Regulations 2003 (the Regulations), and the
HSNO (Methodology) Order 1998.
1.2
The organisms approved for development are the genetically modified
microorganisms listed below in Table 1.
Table 1: Organisms as recorded on the ERMA New Zealand Register
Host Organism Category
of Host
2
1)
Mycobaterium
marinum
laboratory
Strains
(Aronson 1926)
As modified by
Site directed mutagenesis of genes in the
following protein families:
Sensory proteins
Gene expression regulators
Nucleotide cyclases, phosphodiesterases
and cyclic nucleotide binding proteins
Sigma factors
DNA replication and modification
proteins
Transcription and translation proteins
Category of PC
modification level
B
2
2)
Mycobaterium
marinum
laboratory
Strains
(Aronson 1926)
2
3)
Mycobacterium
smegmatis
laboratory
strains
(Trevisan 1889)
(Lehmann &
Neumann 1899)
1
Cell division proteins
Chaperones and stress-related proteins
Protein and peptide secretory proteins
Surface proteins, adhesins and antigens
Cell envelope synthesis proteins
Small molecule and macromolecule
metabolism proteins
Standard non-conjugative plasmid
vectors containing DNA derived from
the genus Mycobacterium (Lehmann and
Neumann 1896).
DNA fragments shall encode regulatory
elements, non-coding regions and open
reading frames of known or predicted
function from the following protein
families:
Sensory proteins
Gene expression regulators
Nucleotide cyclases, phosphodiesterases
and cyclic nucleotide binding proteins
Sigma factors
DNA replication and modification
proteins
Transcription and translation proteins
Cell division proteins
Chaperones and stress-related proteins
Protein and peptide secretory proteins
Surface proteins, adhesins and antigens
Cell envelope synthesis proteins
Small molecule and macromolecule
metabolism proteins
Site directed mutagenesis of genes in the
following protein families:
Sensory proteins
Gene expression regulators
Nucleotide cyclases, phosphodiesterases
and cyclic nucleotide binding proteins
Sigma factors
DNA replication and modification
proteins
Transcription and translation proteins
Cell division proteins
Chaperones and stress-related proteins
Protein and peptide secretory proteins
Environmental Risk Management Authority Decision: GMD05039
B
2
A
1
Page 2 of 12
4)
Mycobacterium
smegmatis
laboratory
strains
(Trevisan 1889)
(Lehmann &
Neumann 1899)
1
5)
Escherichia coli
K12 or B
derivatives
(Migula 1895)
(Castellani and
Chambers
1919)
1
Surface proteins, adhesins and antigens
Cell envelope synthesis proteins
Small molecule and macromolecule
metabolism proteins
Standard non-conjugative plasmid
vectors containing DNA fragments
derived from the genus Mycobacterium
(Lehmann and Neumann 1896).
DNA fragments shall encode regulatory
elements, non-coding regions and open
reading frames of known or predicted
function from the following protein
families:
Sensory proteins
Gene expression regulators
Nucleotide cyclases, phosphodiesterases
and cyclic nucleotide binding proteins
Sigma factors
DNA replication and modification
proteins
Transcription and translation proteins
Cell division proteins
Chaperones and stress-related proteins
Protein and peptide secretory proteins
Surface proteins, adhesins and antigens
Cell envelope synthesis proteins
Small molecule and macromolecule
metabolism proteins
Standard non-conjugative plasmid
vectors containing DNA fragments
derived from the genus Mycobacterium
(Lehmann and Neumann 1896).
DNA fragments shall encode regulatory
elements, non-coding regions and open
reading frames of known or predicted
function from the following protein
families:
Sensory proteins
Gene expression regulators
Nucleotide cyclases, phosphodiesterases
and cyclic nucleotide binding proteins
Sigma factors
DNA replication and modification
proteins
Transcription and translation proteins
Environmental Risk Management Authority Decision: GMD05039
A
1
A
1
Page 3 of 12
Cell division proteins
Chaperones and stress-related proteins
Protein and peptide secretory proteins
Surface proteins, adhesins and antigens
Cell envelope synthesis proteins
Small molecule and macromolecule
metabolism proteins
The applicant has specified the following vector systems to be used:
1. Standard non-conjugative plasmid cloning and expression vectors for
transformation of Mycobacterium marinum, Mycobacterium smegmatis
and Escherichia coli.
These vectors will include some or all of the classes of gene regulatory elements
listed below:
1. Promoters;
2. Enhancers;
3. Transcription termination signals;
4. Operator sequences;
5. Polyadenylation signals;
6. Untranslated regions such as introns/spacers and insulators;
7. Multiple cloning sites;
8. Protein purification tags;
9. Origins of replication;
10. Secretory, targeting and localization signals;
11. Regulatory sequences for induced expression;
12. Other commercially available regulatory elements.
These vectors may contain standard and generally available regulatory elements
sourced from invertebrates, vertebrates, bacteria, viruses or bacteriophages
including the selectable markers Hygromycin resistance, Kanamycin resistance
and Ampicillin resistance.
The genetic material shall not include:
 Genes encoding any known or predicted toxins and pathogenicity islands from
pathogenic mycobacteria.
1.3
Category of low-risk genetic modification:
The genetic modification is a low-risk genetic modification as defined in clause
5 of the Hazardous Substances and New Organisms (Low-Risk Genetic
Modification) Regulations 2003. Mycobacterium smegmatis and Escherichia
coli are category 1 host organisms as defined in clause 7(1) of the HSNO (LowRisk Genetic Modification) Regulations 2003. The transformation of both
microorganisms, as described in Table 1, constitutes a category A genetic
modification as defined in clause 5 of those Regulations and therefore, will be
carried out under a minimum of PC1 containment. Mycobacterium marinum is a
category 2 host organism as defined in clause 7(2) of the HSNO (Low-Risk
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Genetic Modification) Regulations 2003. The transformation of this
microorganism, as described in Table 1, constitutes a category B genetic
modification as defined in clause 5 of those Regulations and therefore, will be
carried out under a minimum of PC2 containment.
2.0 Legislative Criteria for Application
The application was lodged pursuant to section 40(1)(b) of the HSNO Act 1996 and
determined according to the rapid assessment provisions of section 42A of the HSNO
Act 1996.
The application has been approved by Dr Bas Walker, Chief Executive of ERMA
New Zealand, under delegation from the Authority, as provided for in section 19
HSNO Act 1996.
In reaching this decision I have considered matters relevant to the purpose of the
HSNO Act 1996, as specified in Part II, and followed the relevant provisions of the
Hazardous Substances and New Organisms (Methodology) Order 1998.
3.0 Consideration
3.1
Sequence of the consideration
The application was formally received and verified as containing sufficient
information on 18 April 2005.
The decision is based on the information supplied by the applicant in the application
including copies of scientific literature referenced in the application and comments on
the application received from the Department of Conservation (DoC).
In reaching this decision I have used information that is relevant and appropriate to
the scale and significance of the risks, costs, and benefits associated with the genetic
modification.
In accordance with section 42A of the Act (rapid assessment), the approach adopted
was to identify the circumstances of the genetic modification, to evaluate these against
the criteria specified in section 41, and to consider whether there are any residual risks
that require further consideration. This approach covered the following issues:
 Purpose of the application (section 39)
 Assessment against the criteria for low-risk genetic modifications (section
42A)
 Identification and assessment of the risks and other impacts of the organism
 Māori issues and concerns (sections 6(d) and 8)
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Precedents
Proposed controls
In accordance with section 53(2)(b) this application was not publicly notified. The
Department of Conservation (DoC) and the Ministry of Agriculture and Forestry
Biosecurity New Zealand were notified upon receipt of this application.
DoC submitted comments on the application addressing the issues of containment and
potential risks to indigenous flora and fauna. These were summed up in the following
statement:
In general, the research proposed in this application does not pose an increased risk to
indigenous flora and fauna. However, some uncertainty remains regarding the
potential for negative impacts from M. marinum on indigenous fauna including
species of fish and amphibia. Consequently, we believe that particular care should be
taken to ensure this species of bacteria is not released. We consider that the proposed
containment is sufficient to mitigate the risks of release and therefore the Department
would not oppose the approval of this application.
Additionally, DoC noted that additional import approvals would be required for any
of the Mycobacterium sp. (from which DNA may be sourced for use in the
modifications) that either are not already present in New Zealand or are not covered
by existing import approvals. DoC also noted that exotic strains of Mycobacteria have
been declared as unwanted organisms under the Biosecurity Act 1998 by MAF.
Therefore, the applicant will need to contact the Chief Technical Officer of MAF if
they are intending to communicate or propagate exotic strains of Mycobacteria
species, other than those species approved for import into containment by the
Authority1.
3.2
Purpose of the application
The purpose of this project is to determine how mycobacteria control growth and are
able to adapt to their environment and persist indefinitely. Laboratory models based
on the low pathogenic species Mycobacterium smegmatis and Mycobacterium
marinum will be used for this purpose. The function of regulatory and effector
proteins will be determined on the basis of their expression, biochemical properties
and also their importance to the life cycle of M. smegmatis and M. marinum.
To achieve this, the applicant will develop M. smegmatis and M. marinum organisms
that are specifically mutated in single genes encoding proteins belonging to protein
families involved in sensory functions; gene expression regulators; nucleotide
cyclases, phosphodiesterases and cyclic nucleotide binding proteins; sigma factors;
DNA replication and modification proteins; transcription and translation proteins; cell
division proteins; chaperones and stress-related proteins; protein and peptide secretory
proteins; surface proteins, adhesins and antigens; cell envelope synthesis proteins; and
small molecule and macromolecule metabolism proteins.
1
Sections 52 and 53 of the Biosecurity Act 1998
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In addition, M. smegmatis and M. marinum will be modified to express DNA
fragments encoding regulatory elements, non- coding regions and open reading
frames of known or predicted function from the same mycobacterial protein families
described above. The DNA fragments will be sourced from Mycobacterium avium,
Mycobacterium bovis, Mycobacterium tuberculosis, M. marinum, M. smegmatis and
other species within the genus Mycobacterium (Lehmann and Neumann 1896). The
genetic material shall not include genes encoding any known or predicted toxins and
pathogenicity islands from pathogenic mycobacteria.
I have determined that this application may be approved for the purpose of the
development of a genetically modified organism as provided for in section 39(1)(a) of
the HSNO Act 1996.
3.3
Assessment against the criteria for low-risk genetic
modifications
Category of host organisms:
Laboratory strains of Mycobacterium smegmatis, and Escherichia coli are not capable
of causing disease in humans, animals, plants or fungi nor do they produce
desiccation-resistant structures, such as spores or cysts. As such, M. smegmatis and E.
coli are considered Category 1 host organisms under the HSNO (Low-Risk Genetic
Modification) Regulations 2003.
Mycobacterium marinum is capable of causing disease in humans but the risk posed to
the health of laboratory workers is low and it is classified as a risk group 2 organism.
Mycobacterium marinum is a category 2 host organism as defined in the HSNO (LowRisk Genetic Modification) Regulations 2003.
Category of genetic modification:
The modifications to Mycobacterium smegmatis and Escherichia coli as described in
Table 1 are Category A modifications as defined in the HSNO (Low-Risk Genetic
Modification) Regulations 2003.
The modifications to Mycobacterium marinum as described in Table 1 are Category B
modifications as defined in the HSNO (Low-Risk Genetic Modification) Regulations
2003.
I am satisfied that the development meets the criteria for low-risk genetic
modification specified in the Regulations, made under section 41 of the Act. The
experiments meet the requirements of Category A and Category B modifications as
defined in clause 5 of the Regulations in that the modification involves category 1 and
category 2 host organisms and is to be carried out under a minimum of PC1
containment for developments involving Escherichia coli and Mycobacterium
smegmatis and a minimum of PC2 containment for developments involving
Mycobacterium marinum. The developments, described in Table 1, will not increase
the pathogenicity, virulence, or infectivity of the host organism to laboratory
personnel, the community or the environment. In addition, the developments will not
result in organisms that have a greater ability to escape from containment than the
unmodified organism.
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3.4 Identification and assessment of the risks, costs, and other
impacts of the organism
I consider that the information provided by the applicant is relevant and appropriate to
the scale and significance of the risks, costs, and benefits associated with the
application (as required by clause 8 of the Methodology). In accordance with clauses
9 and 10 of the Methodology the information supplied by the applicant and by DoC
has been evaluated as follows.
I consider that, given the containment controls imposed by this approval, there are no
significant non-negligible adverse effects of the proposed genetically modified
Mycobacterium smegmatis, Mycobacterium marinum and Escherichia coli on
humans, animals, plants, other organisms or the environment.
Escherichia coli K12 derivatives contain genetic modifications that make them
dependent upon supplemental growth factors to survive. For this reason it is very
unlikely that the organism could establish a self-sustaining population should it escape
containment.
I have also considered the potential Māori cultural effects in accordance with sections
6(d) and 8 of the HSNO Act and clauses 9(b)(i), 9(c)(iv) of the Methodology, in
consultation with the Manager, Māori. As this application does not involve the use of
genetic material from native or valued flora and fauna or humans, and as this
application is for a development in containment, there is no requirement for the
applicant to consult with Māori.
Although I recognise that Māori maintain an ongoing interest and concern in the
potential long term cultural implications of genetic modification, I consider that this
application poses negligible risk of adverse effects to the relationship of Māori culture
and traditions with their ancestral lands, water, sites, waahi tapu, valued flora and
fauna, and other taonga.
3.5
Precedents
I must consider each application on its merits, and am therefore not bound by the
stance taken in previous decisions. However, in reflecting on previous decisions that
involved similar issues to those raised by this application, I note that low-risk genetic
developments of Mycobacterium smegmatis have been considered and approved
subject to PC1 containment (most recently for application GMD04055).
Whereas the modification in the current application is similar to that approved in
GMD04055, a different host organism is to be used. Under the HSNO (Low-Risk
Genetic Modification) Regulations 2003 M. marinum is considered a Category 2 host
and must therefore, be contained at PC2.
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3.6
Proposed Controls
The experiments proposed in this application, to develop a genetically modified
Mycobacterium smegmatis and Escherichia coli, meet the requirements of Category A
genetic modification as defined in clause 5 of the Hazardous Substances and New
Organisms (Low-Risk Genetic Modification) Regulations 2003. Category A
experiments are required to be contained within a Physical Containment level 1
facility (PC1) registered under MAF/ERMA New Zealand Standard 154.03.02
‘Containment Facilities for Microorganisms’. This containment regime contains clear
guidelines for the safe handling and disposal of bacterial cultures.
The experiments proposed in this application, to develop a genetically modified
Mycobacterium marinum, meet the requirements of Category B genetic modification
as defined in clause 5 of the Hazardous Substances and New Organisms (Low-Risk
Genetic Modification) Regulations 2003. Category B experiments are required to be
contained within a Physical Containment level 2 facility (PC2) registered under
MAF/ERMA New Zealand Standard 154.03.02 ‘Containment Facilities for
Microorganisms’.
The facility in which the organisms will be maintained shall comply with the
requirements of the Australian New Zealand Standard AS/NZS 2243.3:2002 Safety in
Laboratories: Part 3: Microbiological aspects of containment and facilities, except for
the deviations specified in the MAF/ERMA New Zealand Standard 154.03.02. The
laboratory proposed to be used by the Applicant is currently approved and registered
as a containment facility under section 39 of the Biosecurity Act, in accordance with
the MAF/ERMA New Zealand Standard 154.03.02.
4
Decision
4.1
I am satisfied that this application is for one of the purposes specified in section
39(1) of the HSNO Act, being section 39(1)(a): the development of any
genetically modified organism.
4.2
Based on consideration and analysis of the information provided, and having
considered the characteristics of the organisms and the modification and the
criteria for low-risk genetic modification detailed in the HSNO (Low-Risk
Genetic Modification) Regulations 2003, I am of the view that the organisms
meet the criteria for rapid assessment under section 42A(2) of the HSNO Act.
4.3
I am satisfied that the proposed containment regime together with the additional
controls imposed in accordance with section 42A(3)(b) of the HSNO Act 1996
will adequately contain the organisms.
In accordance with section 42(A)(3)(c) of the HSNO Act 1996 I have considered
the provision of progress reports on this development to the Authority and do
not consider that it is necessary for this approval.
4.4
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4.5
Pursuant to section 42A(3)(a) of the HSNO Act 1996, and acting under
delegation from the Authority provided for in section 19, I have approved this
application subject to the controls specified herein.
4.6
In reaching this decision I have relied upon the following criteria in the HSNO
Act and the Methodology:
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5
Criteria for assessing the purpose of the application (section39).
Criteria for rapid assessment of adverse effects for the development of a
genetically modified organism in containment (section 42A).
Criteria for a low-risk genetic modification specified in the HSNO (LowRisk Genetic Modification) Regulations 2003, made under section 41 of
the Act.
The information provided by the applicant was assessed against the criteria
in clauses 9, 10 and 12 of the HSNO (Methodology) Order 1998.
Matters to be addressed by containment controls for development of
genetically modified organisms specified in Part 1 of the Third Schedule to
the HSNO Act.
Notification under section 53(4) was given to DoC and MAF.
Controls
In order to provide for the matters detailed in Part 1 of the Third Schedule of the
HSNO Act, Containment Controls for Importation, Development and Field
Testing of Genetically Modified Organisms, the approved organisms are subject to
the following controls:2
1
To limit the likelihood of any accidental release of any organism
or any viable genetic material.3
1.1
The approved organisms shall be developed and maintained within a
containment facility which complies with these controls.
1.2
The person responsible for a particular research area and/or the person
responsible for the operation of the containment facility shall inform all
personnel involved in the handling of the organisms of the Authority’s
controls.
2
Bold headings in the following text refer to Matters to be Addressed by Containment Controls for
Development and Field Testing of Genetically Modified Organisms, specified in the Third Schedule of
the HSNO Act 1996.
3
Viable Genetic Material is biological material that can be resuscitated to grow into tissues or
organisms. It can be defined to mean biological material capable of growth even though resuscitation
procedures may be required, e.g. when organisms or parts thereof are sub lethally damaged by being
frozen, dried, heated, or affected by chemical.
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1.3
The construction and operation of the containment facility in which the
organisms are maintained, shall be in accordance with the:
a) MAF/ERMA New Zealand Standard 154.03.024: Containment Facilities for
Micro-organisms, at laboratory Physical Containment Levels 1 (PC1) for
experiments involving Mycobacterium smegmatis and Escherichia coli, and
PC2 for experiments involving Mycobacterium marinum.
b) Australian New Zealand Standard AS/NZS 2243.3:20024 Safety in
Laboratories: Part 3: Microbiological aspects of containment and facilities,
except for the deviations specified in the Standard referred to in (a).
1.4
2
2.1
3
3.1
4
The facility shall be approved and registered by MAF as a containment facility
under section 39 of the Biosecurity Act, in accordance with the MAF/ERMA
New Zealand Standard 154.03.024, and controls imposed by the Authority.
To exclude unauthorised people from the facility.
Construction and operation of the containment facility shall comply with the
requirements of the standards listed in control 1.3 relating to the identification
of entrances, numbers of and access to entrances and security requirements for
the entrances and the facility.
To exclude other organisms from the facility and to control
undesirable and unwanted organisms within the facility.
Construction and operation of the containment facility shall comply with the
requirements of the standards listed in control 1.3 relating to the exclusion of
other organisms from the facility and the control of undesirable and unwanted
organisms within the facility.
To prevent unintended release of the organism by experimenters
working with the organism.
4.1
Construction and operation of the containment facility shall comply with the
requirements of the standards listed in control 1.3 relating to the prevention of
unintended release of the organism by experimenters working with the
organism.
5
To control the effects of any accidental release or escape of an
organism.
5.1
Construction and operation of the containment facility shall comply with the
requirements of the standards listed in control 1.3 relating to controlling the
effects of any accidental release or escape of an organism.
4
Any reference to this standard in these controls refers to any subsequent version approved or endorsed by ERMA
New Zealand
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5.2
If a breach of containment occurs, the facility operator must ensure that the
MAF Inspector responsible for supervision of the facility has received
notification of the breach within 24 hours.
5.3
In the event of any breach of containment of the organism, the contingency
plan for the attempted retrieval or destruction of any viable material of the
organisms that have escaped shall be implemented immediately. The
contingency plan shall be included in the containment manual in accordance
with the requirements of standards listed in control 1.3.
6
Inspection and monitoring requirements for containment facilities.
6.1
The operation of the containment facilities shall comply with the requirements
contained in the standards listed in control 1.3 relating to the inspection and
monitoring requirements for containment facilities.
6.2
The containment manual shall be updated, as necessary, to address the
implementation of the controls imposed by this approval, in accordance with
the standards listed in control 1.3.
7
Qualifications required of
implementing those controls.
7.1
The training of personnel working in the facility shall be in compliance with
the standards listed in control 1.3.
the
persons
responsible
for
_____________________
_______________
Dr Bas Walker,
3 May 2005
Chief Executive ERMA New Zealand
Approval codes: GMD003724, GMD003725, GMD003726
Amendment: November 2006
Changes to controls:
 Addition of footnotes to the containment facility references and the
Australian/New Zealand containment facility references to “future proof” the
decision
 Standardise the wording of the breach of containment control
 Removal of the control regarding inspection of facilities by the Authority, its
agent or enforcement officers
16 August 2007
____________________________
Date:
Mr Rob Forlong
Chief Executive, ERMA New Zealand
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