Dir 128 Full Risk Assessment and Risk Management Plan (rarmp)

Table of Contents

Summary of the Risk Assessment and Risk Management Plan 3

Decision 3

The application 3

Risk assessment 3

Risk management plan 4

Risk assessment context 7

Background 7

Regulatory framework 7

The proposed dealings 8

The proposed limits of the dealings (size, locations, duration and people) 9

The proposed controls to restrict the spread and persistence of the GMOs and their genetic material in the environment 9

The parent organisms 10

The GMOs, nature and effect of the genetic modification 10

Introduction to the GMOs 10

The introduced genes, encoded proteins and their associated effects 15

Toxicity/allergenicity associated with the introduced genes, their encoded proteins and associated products 18

Characterisation of the GMOs 19

The receiving environment 19

Relevant abiotic factors 20

Relevant agricultural practices 20

Presence of related plants in the receiving environment 21

Presence of similar genes and encoded proteins in the environment 22

Relevant Australian and international approvals 22

Australian approvals 22

International approvals of GM wheat and barley 22

Risk assessment 23

Introduction 23

Risk Identification 24

Risk source 24

Causal pathway 25

Potential harm 26

Postulated risk scenarios 26

Uncertainty 41

Risk evaluation 42

Risk management 43

Background 43

Risk treatment measures for identified risks 43

General risk management 43

Licence conditions to limit and control the release 43

Other risk management considerations 47

Issues to be addressed for future releases 49

Conclusions of the RARMP 49

Summary of issues raised 61

Summary of issues raised 63

Abbreviations

APVMA	Australian Pesticides and Veterinary Medicines Authority
CaMV	Cauliflower mosaic virus
CCI	Confidential Commercial Information as declared under section 185 of the Gene Technology Act 2000
DAFWA	Department of Agriculture and Food, Western Australia
DIR	Dealings involving Intentional Release
FSANZ	Food Standards Australia New Zealand
GM	Genetically modified
GMO	Genetically modified organism
ha	Hectare
hph	Hygromycin phosphotransferase
LGA	Local government area
m	Metres
NGNE	New Genes for New Environments
NLRD	Notifiable low risk dealings
nptII	neomycin phosphotransferase II
OGTR	Office of the Gene Technology Regulator
PC2	Physical Containment level 2
PR	Pathogenesis-related
RARMP	Risk Assessment and Risk Management Plan
Regulations	Gene Technology Regulations 2001
Regulator	Gene Technology Regulator
TGA	Therapeutic Goods Administration
the Act	The Gene Technology Act 2000

Risk assessment context

Background

1. 
   An application has been made under the Gene Technology Act 2000 (the Act) for Dealings involving the Intentional Release (DIR) of genetically modified organisms (GMOs) into the Australian environment.
   
2. 
   The Act in conjunction with the Gene Technology Regulations 2001 (the Regulations), an inter-governmental agreement and corresponding legislation that is being enacted in each State and Territory, comprise Australia’s national regulatory system for gene technology. Its objective is to protect the health and safety of people, and to protect the environment, by identifying risks posed by or as a result of gene technology, and by managing those risks through regulating certain dealings with genetically modified organisms (GMOs).
   
3. 
   This chapter describes the parameters within which potential risks to the health and safety of people or the environment posed by the proposed release are assessed. The risk assessment context is established within the regulatory framework and considers application-specific parameters (Figure ).
   

PROPOSED DEALINGS

Proposed activities involving the GMO

Proposed limits of the release

Proposed control measures

PARENT ORGANISM

Origin and taxonomy

Cultivation and use

Biological characterisation

Ecology

PREVIOUS RELEASES

GMO

Introduced genes (genotype)

Novel traits (phenotype)

RISK ASSESSMENT CONTEXT
LEGISLATIVE REQUIREMENTS

(including Gene Technology Act and Regulations)

RISK ANALYSIS FRAMEWORK
OGTR OPERATIONAL POLICIES AND GUIDELINES
RECEIVING ENVIRONMENT

Environmental conditions

Agronomic practices

Presence of related species

Presence of similar genes

Figure Summary of parameters used to establish the risk assessment context

Regulatory framework

4. 
   Sections 50, 50A and 51 of the Act outline the matters which the Gene Technology Regulator (the Regulator) must take into account, and who must be consulted with, in preparing the Risk Assessment and Risk Management Plans (RARMPs) that inform the decisions on licence applications. In addition, the Regulations outline further matters the Regulator must consider when preparing a RARMP. In accordance with section 50A of the Gene Technology Act 2000 (the Act), this application is considered to be a limited and controlled release application, as its principal purpose is to enable the applicant to conduct experiments and the applicant has proposed limits on the size, location and duration of the release, as well as controls to restrict the spread and persistence of the GMOs and their genetic material in the environment. Therefore, the Gene Technology Regulator (the Regulator) was not required to consult with prescribed experts, agencies and authorities before preparation of the Risk Assessment and Risk Management Plan (RARMP; see section 50 of the Act).
   
5. 
   Section 52 of the Act requires the Regulator to seek comment on the RARMP from the States and Territories, the Gene Technology Technical Advisory Committee, Commonwealth authorities or agencies prescribed in the Regulations, the Minister for the Environment, relevant local council(s), and the public. The advice from the prescribed experts, agencies and authorities and how it was taken into account is summarised in Appendix A. Thirteen public submissions were received and their considerations are summarised in Appendix B.
   
6. 
   The Risk Analysis Framework (OGTR 2013) explains the Regulator’s approach to the preparation of RARMPs in accordance with the legislative requirements. Additionally, there are a number of operational policies and guidelines developed by the Office of the Gene Technology Regulator (OGTR) that are relevant to DIR licences. These documents are available from the OGTR website.
   
7. 
   Any dealings conducted under a licence issued by the Regulator may also be subject to regulation by other Australian government agencies that regulate GMOs or GM products, including Food Standards Australia New Zealand (FSANZ), Australian Pesticides and Veterinary Medicines Authority (APVMA), Therapeutic Goods Administration (TGA), National Industrial Chemicals Notification and Assessment Scheme and Department of Agriculture. These dealings may also be subject to the operation of State legislation declaring areas to be GM, GM free, or both, for marketing purposes.
   

The proposed dealings

8. 
   The University of Adelaide proposes to release up to 1262 lines³ of genetically modified (GM) wheat and barley into the environment under limited and controlled conditions.
   

The purpose of the trial is to evaluate the candidate genes for yield potential under field conditions. In addition, the release will allow the applicant to produce sufficient grain for subsequent replicated trials.

9. 
   The dealings involved in the proposed intentional release include:
   

• 
  conducting experiments with the GMOs
  
• 
  breeding the GMOs
  
• 
  propagating the GMOs
  
• 
  growing or culturing the GMOs
  
• 
  transporting the GMOs
  
• 
  disposing of the GMOs
  

and the possession, supply or use of the GMOs for the purposes of, or in the course of, any of the above. These dealings are detailed further below.

The proposed limits of the dealings (size, locations, duration and people)

10. 
    The applicant proposes to grow GM wheat and barley plants between June 2014 and December 2019.
    
11. 
    The GMOs are proposed to be planted at five sites, two of which are in South Australia and three in Western Australia. The South Australian sites are located in the LGAs of Marion and Wakefield, and the Western Australian sites in the LGAs of Corrigin, Merredin, and Katanning. The latter two sites represent the New Genes for New Environment (NGNE) facilities operated by the Department of Agriculture and Food, Western Australia (DAFWA).
    
12. 
    The collective area of the trial (over 5 sites) would be up to 2.5 hectares (ha) per growing season.
    
13. 
    The applicant proposes that only trained and authorised staff would be permitted to deal with the GM wheat and barley. Any other visitors to the sites would be accompanied by an authorised University of Adelaide representative and would not deal with the GMOs.
    

The proposed controls to restrict the spread and persistence of the GMOs and their genetic material in the environment

The applicant has proposed a number of controls to restrict the spread and persistence of the GM wheat and barley lines and the introduced genetic material in the environment, including:

• 
  locating the trial sites at least 200 m from other wheat and barley plants, unless such plants represent those (GM or non-GM) planted at another location as part of this trial or those approved for trial at the site under a separate DIR licence
  
• 
  surrounding each trial site with a fence, and controlling plant growth in a 10 m wide zone outside the fence by mowing, herbicide treatment and/or weeding
  
• 
  surrounding each location (in the fenced trial site) by a 2 m wide buffer where plant growth will be controlled by mowing, herbicide treatment and/or weeding
  
• 
  monitoring each location for volunteers, beginning two weeks prior to the expected start of flowering and continuing on a fortnightly basis until flowering has finished
  
• 
  during flowering, inspecting a 190 m wide zone surrounding the 10 m wide zone for wheat and barley (but see next point)
  
• 
  if there has been no cultivation or detection of wheat and barley in the 190 m wide zone in the previous two years, reducing inspection of that area to the 50 m wide area directly surrounding the 10 m wide zone
  
• 
  after harvest, monitoring locations for volunteers and related species at least once every 35 days for a period of two years; during this time period at least three irrigations will be performed to encourage germination of volunteers
  
• 
  destroying volunteers and related species
  
• 
  cleaning equipment used in harvesting, and threshing on site or transporting the heads to approved facilities for analysis or processing
  
• 
  storing seed remaining from analysis in an approved facility or destroying the seed by autoclaving or any other method approved by the Regulator
  
• 
  destroying waste material derived from harvest, and any stubble, by ploughing back into the soil, burning or burial
  
• 
  transporting and storing GM material in accordance with the Regulator’s Guidelines for the Transport, Storage and Disposal of GMOs (2011)
  
• 
  restricting access to trial sites to authorised staff
  
• 
  not allowing GM plant material or products to be used for human food or animal feed.
  

14. 
    Figure shows the proposed site layout including some of these controls. These controls, and the limits outlined above, have been taken into account in establishing the risk assessment context (this Chapter), and their suitability for containing the proposed release is evaluated in , Section .
    

 

 

 

 

 

190 m wide

Isolation zone inspected for wheat and barley during flowering of GMOs

10 m wide

Monitoring zone where the growth of plants is controlled

Fence

2 m wide buffer

Zone where the growth of plants is controlled

Location

(planting area)

Figure Proposed trial layout, including some of the controls (not drawn to scale)

The parent organisms

15. 
    The parent organisms are bread wheat (Triticum aestivum L.) and barley (Hordeum vulgare L.), both exotic to Australia. Commercial wheat cultivation occurs in the wheat belt from south eastern Queensland through New South Wales, Victoria, Tasmania, southern South Australia and southern Western Australia. Barley is cultivated in these same areas, although a small amount is also grown in Tasmania.
    
16. 
    The wheat cultivars used to generate the GM wheat lines are Bobwhite and Gladius, but some genes have been backcrossed into the commercial cultivars EGA Bonnie Rock, EGA-Burke, IGW-2971, Magenta, Frame and Drysdale. GM barley lines were generated in Golden Promise and WI4330, but some genes were backcrossed into the commercial cultivars Flagship, Gairdner, Commander.
    
17. 
    Detailed information about the parent organisms are contained in the reference documents The Biology of Triticum aestivum L. em Thell (bread wheat) (OGTR 2008b) and The Biology of Hordeum vulgare L. (barley) (OGTR 2008a), which were produced to inform the risk assessment process for licence applications involving GM wheat and barley plants. These documents are available from the OGTR website.
    

The GMOs, nature and effect of the genetic modification

Introduction to the GMOs

The applicant proposes to release up to 1262 lines of GM wheat and barley. The lines were produced using either Agrobacterium tumefaciens or biolistics mediated plant transformation. Information about these transformation methods can be found in the risk assessment reference document Methods of plant genetic modification available from the Risk Assessment References page on the OGTR website.

Each GM wheat and barley line has been transformed with a single gene from 33 genes of interest. On the basis of the trait that they are expected to induce, the genes can be divided into five groups: (i) drought tolerance; (ii) salt tolerance; (iii) aluminium tolerance; (iv) nitrogen use efficiency; and (v) micronutrient uptake. The GMOs also contain selectable marker genes, these being the neomycin phosphotransferase II (nptII) gene and/or the hygromycin phosphotransferase (hptII) gene, both originating from E.coli.

A summary of these groups is presented in Table 1, with details provided in the following sections.

Table 1 Summary of GM wheat and barley lines proposed for release.

GMO class	Gene of interest^#	Plant transformed	Approx no. of lines
Drought tolerance
1	SIGNALLING PROTEIN 2	T. aestivum	18
2	TRANSCRIPTION FACTOR 1	T. aestivum	72
3	TRANSCRIPTION FACTOR 3	T. aestivum	54
4	TRANSCRIPTION FACTOR 4	T. aestivum	54
5	TRANSCRIPTION FACTOR 5	T. aestivum	54
6	TRANSCRIPTION FACTOR 6	T. aestivum	54
		H. vulgare	36
7	TRANSCRIPTION FACTOR 7	T. aestivum	90
8	PHOTOSYNTHESIS AND METABOLISM GENE 3	T. aestivum	18
9	PHOTOSYNTHESIS AND METABOLISM GENE 4	T. aestivum	18
10	PHOTOSYNTHESIS AND METABOLISM GENE 5	T. aestivum	18
11	CELL SPECIFICATION, PROLIFERATION AND DIVISION GENE 2	T. aestivum	36
12	CELL SPECIFICATION, PROLIFERATION AND DIVISION GENE 3	T. aestivum	36
13	RNA METABOLISM PROCESSING GENE 1	H. vulgare	6
14	RNA METABOLISM PROCESSING GENE 2	H. vulgare	6
15	RNA METABOLISM PROCESSING GENE 3	T. aestivum	72
		H. vulgare	72
16	RNA METABOLISM PROCESSING GENE 4	T. aestivum	72
		H. vulgare	72
Salt tolerance
17	ScNHA1 (Saccharomyces cerevisiae)	H. vulgare	40
		T. aestivum	20
18	PpENA1 (Physcomitrella patens)	H. vulgare	40
		T. aestivum	20
19	AtAVP (A. thaliana)	H. vulgare	10
		T. aestivum	5
20	AtCIPK16 (A. thaliana)	T. aestivum	20
		H. vulgare	20
Aluminium tolerance
21	TaALMT1 (T. aestivum)	T. aestivum	14
		H. vulgare	12
22	TaALMT1_minus_insert (T. aestivum)	H. vulgare	9
23	ION TRANSPORTER 5	T. aestivum	9
		H. vulgare	9
24	ION TRANSPORTER 6	T. aestivum	9
		H. vulgare	9
25	ION TRANSPORTER 7C	T. aestivum	9
		H. vulgare	9
26	ScALMT1.M39.1_wt (Secale cereale)	T. aestivum	8
		H. vulgare	6
27	ScALMT1.M39.1_plus insert (Secale cereale)	H. vulgare	6
28	HvAACT1 (H. vulgare)	T. aestivum	2
		H. vulgare	4
Nitrogen use efficiency
29	TRANSCRIPTION FACTOR 2	T. aestivum	20
30	TRANSCRIPTION FACTOR 8	T. aestivum	12
31	Aminotransferase	T. aestivum	32
		H. vulgare	32
32	CELL SPECIFICATION, PROLIFERATION AND DIVISION GENE 1	T. aestivum	12
Micronutrient uptake
33	OsNAS2 (Oryza sativa)	T. aestivum	6

# The identities of some of these genes have been declared as CCI. The applicant has assigned an identifier to these genes.

The vectors used for Agrobacterium mediated transformation are variants of a pMDC backbone, while the vectors used for biolistic transformation include variants of a pMDC backbone and those based on PHP. Promoters, genomic sequences, and terminators used in these vectors come from Agrobacterium tumefaciens, Cauliflower mosaic virus, barley, maize, potato, rice, sorghum, tobacco, durum wheat and bread wheat. They are listed in Tables 2, 3 and 4.

Table 2 Promoters used in constructs.

Name of promoter #	Name of gene that promoter derived from^#	Source organism
CaMV35S	Viral promoter	Cauliflower mosaic virus
pAct	OsAct	O. sativa
pB4L	TdHDZipl-4	T. durum
pB7L	TdHDZipl-3	T. durum
pCor39	TdCor39	T. durum
pCor410b	TdCor410b	T. durum
pCor410H1(truncated)	TdCor410H1	T. durum
pCor410H2 (truncated)	TdCor410H2	T. durum
PROMOTER 1	GENE A	T. durum
pDHN8	HvDHN8	H. vulgare
PROMOTER 2	GENE B	T. durum
PROMOTER 3	GENE C	T. durum
PROMOTER 4	GENE D	T. durum
PROMOTER 5	GENE E	O. sativa
PROMOTER 6	GENE F	O. sativa
PROMOTER 7	GENE G	O. sativa
pOsAnt1	Aldehyde dehydrogenase se1	O. sativa
PROMOTER 8	GENE H	Z. mays
PROMOTER 9	GENE I	T. durum
pUbi	Polyubiquitin	Z. mays
PROMOTER 10	GENE J	T. durum
pWRKY71 (pJRCO189)	OsWRKY71	O. sativa
PROMOTER 11	GENE K	T. durum
PROMOTER 12	GENE L	Z. mays

# The identities of some of the promoters and the genes from which they originate have been declared as CCI. The applicant has assigned an identifier to these promoters and genes.

Table 3 Genomic elements used in constructs.

Name of sequence^#	Description^#	Source organism^#
Act intron (MOD1)	intron	O. sativa
APS	amplification promoting sequences	N. tabacum
Mini ALLSTOPS	Stop codon	-
STLS1 intron 2	intron	S. tuberosum
Ubi intron	intron	Z. mays
Ubi1 5’UTR	5’ untranslated region	Z. mays
UNTRANSLATED SEQUENCE 1	GENE M	CCI
Ubi1 intron	intron	Z. mays
UNTRANSLATED SEQUENCE 2	GENE N	CCI

# The identities of some of the genomic elements, the genes they come from and their source organisms have been declared as CCI. The applicant has assigned an identifier to these genomic elements and genes.

Table 4 Terminators used in constructs.

Name of terminator^#	Description^#	Source organism^#
35S ter	Viral terminator	Cauliflower mosaic virus
TERMINATOR SEQUENCE 1	Terminator of the GENE A gene	T. durum
TERMINATOR SEQUENCE 2	Terminator of the GENE B gene	T. durum
TERMINATOR SEQUENCE 3	Terminator of the GENE O gene	CCI
TERMINATOR SEQUENCE 4	Terminator of the GENE P gene	T. durum
Nos ter	Nopaline synthase gene terminator	A. tumefaciens
OsUBI ter	Polyubiquitin gene terminator	O. sativa
TERMINATOR SEQUENCE 5	Terminator of the GENE Q gene	CCI
TERMINATOR SEQUENCE 6	Terminator of the GENE R gene	CCI
TERMINATOR SEQUENCE 7	Terminator of the GENE S gene	CCI
TERMINATOR SEQUENCE 8	Terminator of the GENE T gene	S. bicolor
TERMINATOR SEQUENCE 9	Terminator of the GENE U gene	A. tumefaciens

# The identities of some of the terminators, the genes they come from and their source organisms have been declared as CCI. The applicant has assigned an identifier to these terminators and genes.

The introduced genes, encoded proteins and their associated effects

Four of the traits, drought tolerance, salt tolerance, aluminium tolerance and nitrogen use efficiency, can be grouped as abiotic stress tolerances. The natural mechanisms by which plants deal with abiotic stresses can be classified as either avoidance or tolerance, the former referring to those that obviously affect the morphology and/or physiology of a plant, and the latter to molecular changes that may have no visual effects (Howles & Smith 2013). The availability of water is the most important factor that limits crop yield globally (Castiglioni et al. 2008), but salinity and the presence of aluminium in acid soils likewise both have large negative impacts upon agriculture (Brunner & Sperisen 2013; Munns et al. 2012; Rengasamy 2010). Abiotic stress tolerances are multigenic traits, involving the interaction of genes where the protein products constitute different biochemical pathways. Further information on plant responses to abiotic stresses can be found in the RARMP for DIR 102 (available at