A qualitative binary risk assessment model for regulating the biosecurity and environmental risk of endophytes

Accepted Manuscript A qualitative binary risk assessment model for regulating the biosecurity and environmental risk of endophytes Kate E. Bromfield, Steven Corin, Asela Atapattu PII: DOI: Reference:

S1049-9644(17)30097-X http://dx.doi.org/10.1016/j.biocontrol.2017.05.002 YBCON 3586

To appear in:

Biological Control

Received Date: Revised Date: Accepted Date:

15 December 2016 27 April 2017 9 May 2017

Please cite this article as: Bromfield, K.E., Corin, S., Atapattu, A., A qualitative binary risk assessment model for regulating the biosecurity and environmental risk of endophytes, Biological Control (2017), doi: http://dx.doi.org/ 10.1016/j.biocontrol.2017.05.002

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A qualitative binary risk assessment model for regulating the biosecurity and

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environmental risk of endophytes

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Running title: Endophyte Risk Model

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Kate E Bromfield 1, Steven Corin, Asela Atapattu

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New Zealand Environmental Protection Authority

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Keywords: ERA, regulation, traits, novel, Epichloë, alkaloids

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Corresponding author: [email protected]

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1

Abstract

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We present a model for the regulation of endophytes that focuses on the hazard identification

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and risk assessment components. The goal was to develop a model that can identify and

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characterise risk, and quantify the level of risk. Our model takes a binary approach, either

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categorising risk into ‘acceptable’ or ‘unacceptable’ levels, as regulators are often left with a

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choice of approving or declining a proposal when an organism is unable to be contained after

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it is released. This model provides a framework for practitioners to use in order to identify

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and quantify the risks and benefits to be derived from endophytes and plants with novel traits

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systematically. We present two case studies of the model in practice in New Zealand

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legislation under the Hazardous Substances and New Organisms (HSNO) Act, and indicate

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how the model might be used by researchers and regulators to allow for the safe introduction

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of new traits for biocontrol.

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1. INTRODUCTION

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Biotechnology is a rapidly advancing field both in terms of research and application. The

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pace of research raises questions on how to regulate the commercial products that result from

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this groundswell of innovation.

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At present there are varying approaches to regulating the products of biotechnology research.

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Many countries such as the United States and Australia have instigated methods-based

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regulation, which control the development and use of biotechnology products based on the

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methods used to create them; for example, Genetic Modification (GM) technology is singled

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out for regulation (Ammann 2014). In Canada, the regulatory system is traits based. Any

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newly created organism considered to have a ‘novel trait’ that differs from the original

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organism is regulated whether it was the product of GM, chemical mutagenesis or even

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conventional cross breeding (McLean and Charest 2000). The regulatory framework in New

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Zealand is similar to the Canadian system. It focuses on whether an organism is ‘new’ with

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the definition including new species not present in the environment or any organism created

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through the modification of genetic material using ‘in vitro techniques’. The nature of this

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legislation means that the New Zealand Environmental Protection Authority (EPA), as the

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national regulator, has needed to create a system that is responsive to both assessing the risk

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and benefits of research methods and the resulting organism.

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New Zealand’s primary production is typically cattle based. At June 2015, there were 6.5

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million head of dairy cattle (Statistics NZ 2016) and an additional 3.5 million beef cattle.

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Dairy production is New Zealand’s top export earner, accounting for 7-8% of GDP (~$11

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million per annum; Statistics NZ 2016), and making up over 40% of the world’s trade in

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dairy produce (USDA, 2010). It is therefore not surprising that there is ongoing pressure to

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continue to grow productivity in the sector, and develop high value food products, while at

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the same time improve environmental sustainability (Marshall et al 2012). In order to achieve 3

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these goals, research has begun to focus on fungal endophytes - obligate symbionts that are

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associated with almost all plant species (Saikkonen et al. 1998). Endophytes have the

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potential to be a source of novel products for agriculture, medicine and industry and have

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sometimes been construed as a panacea for all modern problems, with the compounds

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resulting from endophyte/host interactions showing antibiotic, antiviral, anticancer,

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antioxidant, insecticidal and antidiabetic properties (Strobel and Daisy, 2003).

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Advances have been made in this field, including the creation of novel plant-endophyte

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associations for the improvement of pasture (Thom et al. 2014). In New Zealand, fungal

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endophytes have been identified as critical to the success of pasture grasses, as they produce

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bioactive compounds that protect the grass host from insect herbivory (Popay & Bonos

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2005). Without these fungal endophytes pastoral agriculture would be limited in large parts of

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New Zealand due to forage losses from insects. Yet the unmanaged use of endophytes poses

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potential risks to the environment. For example, perennial ryegrass (Lolium perenne) can be

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host to the endophyte species Neotyphodium lolii, which in this association produces

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neurotoxins that are harmful to grazing mammals, and can cause the neurological condition

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‘ryegrass staggers’ (Fletcher and Harvey 1981). Importantly, the bioactive compounds

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produced by the fungus are expressed differentially in planta and in culture (Blankenship et

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al. 2001; Tanaka et al. 2005; Young et al. 2006), and that toxin production can be affected by

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both the genotype of the host plant and the environment in which it grows (Easton et al.

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2002). These factors have led some to argue that a more comprehensive understanding of the

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genetic basis and phenotypic plasticity of the plant–microbe interaction in different

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environments is needed (Saikkonen et al. 2013).

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It is clear that there are valid reasons to take a considered approach to the use of novel plant-

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microbial associations. In addition, the implementation of a robust and comprehensive risk

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assessment process to examine new proposals has the potential to lower the costs of 4

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regulation, and speed up the pathway to commercialisation of this potentially valuable

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technology. In this paper we present and describe such a risk methodology and framework. It

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has been developed to enable the New Zealand Environmental Protection Authority (EPA) to

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assess and regulate plant-microbe associations, but there is also the potential for the model to

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inform environmental risk assessments (ERA) in overseas jurisdictions. We present the

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rationale for the model, the specifics of the model itself and then we test the model using two

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case studies; (1) the introduction of the fungal endophyte species Neotyphodium seigellii to

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improve the performance of rye grass pasture, and (2) the introduction of novel

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Neotyphodium endophytes into cereal crops to protect against herbivory.

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2. RISK ASSESSMENT MODEL

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2.1 Risk assessment model

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Our model focuses on the hazard identification and risk assessment components of

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introducing novel traits (Figure 1). The goal was to develop a model that can identify and

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characterise risk, and to some extent quantify the level of risk. For the purposes of decision

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making our model uses a qualitative framework in order to allow for a richer characterisation

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of risk. Our model also takes a binary approach, either categorising risk into ‘acceptable’ or

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‘unacceptable’ levels, as in the case of a novel organism there are few opportunities to impose

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practical controls once an organism has been released in the field.

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The risk assessment model presented here provides a framework for practitioners to use to

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identify and quantify the risks and benefits to be derived from endophytes and plants with

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novel traits. The risks and benefits arising from the use of these organisms are primarily due

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to the changes in their biological characteristics as a result of the plant taking on novel

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‘embedded’ traits. This means that a hazard can be described in terms of new traits arising

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from the modification. The risk assessment process then compares a baseline to the additional

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benefits and risks arising from the hazards identified.

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2.2 Risk assessment process

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As previously mentioned, novel traits can be considered hazards. Therefore the first step of

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the risk assessment model is to determine the taxonomic identity of both the endophyte and

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the host plant (Figure 2.). This categorisation will enable the prediction of potential traits and

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hazards that can arise during the creation of a new association. We note that a taxonomic

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categorisation need not be at the species level. On occasions it may be more applicable to

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focus on sub-populations, or strains as well as higher taxonomic groupings, with the ultimate

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decision being based on the traits of the organisms in question (Baird et al. 2008). Indeed,

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taxonomic changes have already muddied the waters for the specific endophytes considered here

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which are now all considered to be in the same genus, Epichloë (Leuchtmann et al. 2014).

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2.3 Hybridisation

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There is a risk that introducing a new endophyte into a previously unutilised host could lead

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to hybridisation, introgression or recombination events between the novel endophyte and any

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naturally occurring endophytes (for example Shoji et al. 2015). This could have potential

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impacts: novel genetic mixing causing new traits or interactions with the plant host to arise

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(Faeth et al. 2016), or theoretically threaten the diversity of native endophytes. Therefore the

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next question in the risk assessment model is whether or not hybridisation can occur. If the

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answer is affirmative, the potential impacts need to be considered, and if further information

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is unavailable, we suggest that such risks should be considered unacceptable.

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2.4 Horizontal transmission

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Fungal endophytes have three distinct dispersal mechanism types: type I where stroma are

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obligatory on infected plants (termed horizontal transmission); type II where stroma are 6

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optional; and type III where no stroma are formed (termed vertical transmission)

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(Leuchtmann and Clay 1997). If an endophyte can transmit horizontally, a new and different

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spectrum of traits could be expressed when the endophyte is associated with a different host.

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Without knowing more detail about this potential, any horizontal transmission indicates that

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the activity may not be predictable, lowering the certainty of any ERA. For this reason, if

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there is evidence that an endophyte can spread between hosts without assistance, via

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horizontal transmission, we consider this an unacceptable risk, unless information is available

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to identify the range of possible hosts. Conversely, if information is available that confirms

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that the endophyte in question only transmits vertically, we consider this would constitute an

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acceptable risk in terms of the model.

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2.5 Alkaloid profile

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One of the main hazards of endophytes is the production of bioactive alkaloids (Hume et al.

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2016). Four classes of bioactive alkaloid compounds have been identified; ergot alkaloids,

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lolines, indole-diterpenes and peramine (Panaccione et al. 2014). These can adversely affect

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organisms which feed on the host plant and production of these alkaloids is therefore

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considered a hazard. Both benefits and risks can arise from the hazard depending on the

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species impacted, whether it has been accorded pest status or is considered economically

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significant, and the level of the effect. For example, these alkaloids have been implicated in

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resistance to invertebrates (Shymanovich et al. 2015; Hennessy et al. 2016), but also have the

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potential to harm vertebrates, with effects including reduced weight gain, reduced milk

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production (Bush et al. 1997), and the neurological condition ‘ryegrass staggers’ (Fletcher

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and Harvey 1981).

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While hazards in the model are considered in a binary fashion, an assessment of bioactive

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compounds needs to be conducted much more intensively. There needs to be a

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comprehensive assessment on how stable alkaloid production is across a range of hosts in a 7

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range of environments, enabling a focus on the assessment of the specific alkaloids produced.

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If there is wide variation in the profile expressed, it is very difficult to predict the impacts of

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the novel plant-endophyte association. In this event, more information is required, or the risks

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should be considered unacceptable.

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If an accurate description of the alkaloid profile can be created, then an assessment can be

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made as to whether or not the effects of the bioactive compounds are beneficial or adverse.

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Specific information is provided in the case studies below. However, in a generalised

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approach, the regulator should consider: the environmental baseline given that fungal and

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bacterial endophytes are globally widespread and even considered ubiquitous (Stone et al.

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2004), which species could be further affected by the introduction of a novel association, and

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any potential mitigation measures available.

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2.6 Competitive advantage

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Fungal endophytes have been credited with improving host fitness through the production of

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alkaloids, but additional improvements in a plant’s ability to withstand stressors such as

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fungal pathogens and drought have also been observed (Rodriguez et al. 2009). If this fitness

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advantage is significant, there is the potential for the host plant to begin to outcompete other

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desirable plants including native and valuable species. The risk is that the plant inoculated

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with the novel endophyte could become weedier or invasive in ways it wasn’t previously, and

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if there is any evidence to suggest this, the risk should be considered unacceptable.

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2.7 Endpoint assessment

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It is important to note that most of this model is based on a binary concept of acceptable or

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unacceptable risks. This means that ultimately there is a level of subjectivity involved in

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determining what can be considered significant and therefore unacceptable. As such, these

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decisions will ultimately be value based (Corin et al. 2012). How this is dealt with depends

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on the regulatory environment and the way it considers subjectivity. It is important to focus

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attention on the biophysical risks to human health, environment and economy. This allows for

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quantification against an established baseline, and if the effects are considered acceptable,

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there is good reason to approve the proposal (Figure 2).

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3. CASE STUDY 1 - Neotyphodium siegelii AND PASTURE GRASSES

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In 2013 an application was made to the EPA to import and release Neotyphodium siegelii into

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the New Zealand environment. Neotyphodium fungal species are natural endophytic

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associates of many grass genera and tribes of the subfamily Pooideae (Schardl et al. 2004),

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including ryegrasses, fescues, and other pasture grasses ((Easton and Fletcher 2007; Easton

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2007). The specific characteristics of these associations vary (Faeth and Saikkonen 2007;

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Rudgers and Clay 2007), but the presence of an endophyte typically enhances host fitness by

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increasing tolerance to nutrient stress, increasing tolerance to insect and herbivore grazing,

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and also potentially conferring a degree of drought tolerance (Elmi and West 1995;

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Malinowski and Belesky 2006; Kuldau and Bacon 2008). Major risks associated with the

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release of a “new to the environment” fungal endophyte result from the potential for toxins

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produced as a result of the new interactions between hosts, endophytes and the growing

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conditions, to cause unintended harm to animals, and the potential for plant fitness to increase

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to the point of weediness.

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3.1 Hybridisation

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In the case of N. siegelii phylogenetic analysis shows that it is likely to be an asexual hybrid

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of the sexually reproducing endophytes Epichloë festucae and E. bromicola. Epichloë

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festucae is present in New Zealand but at the time of writing we found no evidence in the

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available literature that E. bromicola is present. There are six closely-related species of

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Neotyphodium endophytes known to be present in New Zealand, including the native

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N. aotearoae which lives within the endemic forest hedgehog grass (Echinopogon ovatus)

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(Moon et al. 2002). The other, introduced species: N. coenophialum, N. lolii, N. occultans, N.

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typhinum, and N. uncinatum, are found respectively in the pasture grasses tall fescue;

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perennial ryegrass; Persian, Italian, and Wimmera ryegrasses; Canary Islands ryegrass; and

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meadow fescue (Moon et al. 2000; Clay and Schardl 2002; Moon et al. 2004). In addition,

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Neotyphodium has genetic affinities with native endophytes found in grasses (Moon et al.

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2002).

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For hybridisation to occur, N. siegelii and another endophyte would need to co-infect a host

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plant, the two endophytes would need to hybridise through somatic fusion and parasexual

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processes, and the resulting host strain would need to survive and persist in the environment.

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Such an event is considered extremely unlikely, and there is no evidence to suggest that it

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occurs except over evolutionary timescales (A. Leuchtmann, personal communication, 2013).

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Introduced Neotyphodium endophytes have been associated with endemic and native flora for

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some time, and there is no evidence to suggest that they regularly hybridise.

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3.2 Horizontal transmission

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Neotyphodium endophytes form asymptomatic and, for the most part, mutualistic symbioses

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with their hosts, and are transmitted vertically via host seed colonisation (Schardl and Clay

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1997). The fungus invades the developing seed and ultimately the leaf tissues of the

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developing plant (Philipson and Christey 1986). In this clonal and efficient means of

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propagation, nearly 100% of seeds from infected mother plants transmit the endophyte

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(Siegel et al. 1984). Endophyte growth is synchronised with the host plant tissues, and when

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the leaf tissues hosting the endophyte senesce and die, the endophyte dies with them. The

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fungus does not survive independently of its host. While Neotyphodium endophytes are

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obligate symbionts with no capacity to exist on their own, it is possible to isolate them in the

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laboratory and culture them on agar preparations (Latch and Christensen 1985). When 10

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artificially cultured, N. siegelii is known to produce abundant conidia. Asexual conidia

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produced by N. siegelii in vitro and in planta are functional in the sense that they are able to

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germinate and establish new colonies in vitro, but it has not been experimentally

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demonstrated that N. siegelii can spread to uninfected plants via these conidia (A.

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Leuchtmann, personal communication, 2013). Neotyphodium siegelii should therefore be

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considered to reproduce wholly by vertical transmission.

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3.3 Alkaloid profile

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Neotyphodium endophytes a range of produce bioactive alkaloids (Schardl et al. 2012;

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Schardl et al. 2013). These alkaloiods accumulate in host grass tissues to varying degrees

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(Faeth et al. 2006; Matsukura et al. 2014; Adhikari et al. 2016). Many species of

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Neotyphodium produce one or more of these alkaloids, and the spectrum of alkaloids

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produced by the endophyte can vary depending on which host grass it is associated with. For

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example, ryegrass and tall fescue, when hosting other Neotyphodium species produce

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ergovaline and lolitrem B, which are toxic to grazing vertebrates. In contrast, when in

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association with N. siegelii, they produce only the alkaloid loline. Lolines increase the

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resistance of host grasses to invertebrate herbivory, and improve resistance to drought

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conditions, but do not have any effect on vertebrate herbivores, such as cattle, sheep and birds

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(Bush et al. 1997; Kuldau and Bacon 2008; Zhang et al. 2009). Because N. siegelii produces

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only lolines, grass species hosting the endophyte are considered harmless to grazing animals

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and birds, but continue to show resistance to insect pests and improved drought tolerance.

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Although some variation in the expression of lolines has been observed, this appears to cause

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variation in the efficacy against specific invertebrate pests and environmental stresses

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(Malinowski and Belesky 2006).

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The agronomic potential presented by endophytes involves the transfer of favourable traits

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conferred by the endophyte, such as beneficial bioactive production, from the original host to 11

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new plant species. Benefits include the ability for farmers to replace existing endophyte

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infected grasses with new grass-endophyte associations that do not produce animal toxins, for

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example, by replacing perennial ryegrass that carries a toxin-producing endophyte with

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ryegrass hosting N. siegelii, which produces only alkaloids toxic to insects, and not

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vertebrates. Such a change would improve animal health while retaining pasture protection

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from invertebrate pests. Conversely, potential risks arise for non-target species to be harmed

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by the presence of these alkaloids. For example, some species of moth endemic to New

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Zealand are found on pasture (Barlow et al. 1986) and may be adversely affected by the

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alkaloids produced when N. siegelii is sown in association with ryegrass. However, any

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regulatory assessment of a new proposal must be mindful of existing environmental

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baselines. In New Zealand, perennial grasses already hosting endophytes naturally, produce a

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wide range of alkaloids. For example, in Neotyphodium lolii/Lolium perenne perennial

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ryegrass associations, the alkaloids lolitrems, paxilline, peramine and ergovaline are

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produced (Schardl et al. 2012). These confer significant insect pest protection to the host,

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with ergovaline, an ergot alkaloid, being associated with resistance to African black beetle

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(Heteronychus arator) (Ball et al. 1997), while peramine increases resistance to Argentine

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stem weevil (Listronotus bonariensis) (Prestidge et al. 1991). Therefore, any introduction of

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N. siegelii would be made against an alkaloid rich environment, and it is unlikely that the

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levels of alkaloids to which native invertebrates are exposed would be significantly altered.

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3.4 Competitive advantage

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As a result of new, beneficial traits conferred by N. siegelii onto perennial ryegrass, the host

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could become more dominant in its environment, even to the point of weediness. In

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particular, traits that enhance tolerance to environmental stress could allow the plant to spread

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and persist into previously unaffected habitats. Furthermore, other secondary effects, such as

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enhanced tolerance to invertebrate pests could help release the plant from natural enemies 12

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and help it establish more widely. However, it is widely recognised that many of the benefits

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conferred by novel endophytes are moderate or variable and unlikely to provide a high level

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of competitive advantage over existing species (Faeth 2002). In this particular case-study, the

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biological traits of the perennial ryegrass-N. siegelii association were assessed against the

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baseline of perennial ryegrass in association with its natural endophyte, and the modification

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would not be expected to enhance weediness.

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3.6 Endpoint evaluation

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Based on the information available, the EPA considered that the risks of N. siegelii in

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association with perennial ryegrass were acceptable and not significant (EPA 2013a). This

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was in part due to the low likelihood of risks occurring, and the low magnitude of potential

11

effects. The EPA considered the baseline situation where endophyte-infected grasses

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expressing a wide range of alkaloids, some potentially harmful to animals, was already

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present in New Zealand. The EPA also considered that the potential benefits to New

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Zealand’s agricultural industry and the economy were significant and on this basis approved

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the proposal, allowing N. siegelii to be imported and released into New Zealand (EPA 2013b).

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4. CASE STUDY 2 – Neotyphodium AND CEREAL CROPS

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Soon after the first case study was approved, a new proposal was received by the EPA to

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release non-toxic Neotyphodium fungi in order to form novel associations with cereal crops.

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The intention of the application was to introduce the beneficial properties of Neotyphodium

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endophytes, such as drought and insect resistance, to agronomically important cereal crops.

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The applicant pointed out that modern cereals are not naturally infected with epichloae fungi,

22

but that their wild relatives are, and they considered that the infection of cereal crops could

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also improve their resistance to plant pathogens. The application identified a number of

13

1

strains of Neotyphodium that produce the alkaloids loline, peramine, chanoclavine and

2

terpendole E.

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The risk assessment process was similar to that used in case study 1. In fact, as the biology of

4

the organisms proposed for release was so similar to N. siegelii the only traits or hazards

5

identified that differed are the expressed alkaloid profile and any interactions resulting from

6

the novel association with the plant host. However, unlike case study 1, the baseline for

7

assessment was a situation where modern cereal crops lack endophytes, and the risk analysis

8

compared cereal crops with Neotyphodium endophytes to this baseline.

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4.1 Alkaloid profile

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As previously discussed, endophytes induce effects through the production of bioactive

11

alkaloids. This case study focuses on a variety of Neotyphodium strains that produce not only

12

lolines but also chanoclavine, terpendole E and peramines. These alkaloids, individually or in

13

combination, can offer considerable advantages over endophyte-free crops. Neotyphodium in

14

association with rye corn has been shown experimentally to cause a significant reduction in

15

the numbers of nematodes per root system, to inhibit bird cherry oat aphid and Aceria mites

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in wheat, inhibit light brown apple moth in numerous horticultural crops, and inhibit mycelial

17

growth of the plant pathogens Fusarium head blight and bare patch - both devastating

18

diseases of cereal crops, including wheat and barley (AgResearch, unpublished data). This

19

evidence presents a strong case that new endophyte-cereal crop associations will reduce the

20

impact of pest invertebrates on plants, and increase fungal resistance, and these factors will

21

have a positive impact on plant growth. Furthermore, the use of pesticides and fungicides are

22

likely to reduce, lowering the environmental impact of croplands, and improving farm

23

economic margins.

14

1

The potent effect of these endophyte-cereal associations on invertebrate pests introduces the

2

possibility that native species associated with cereal crops could be harmed. For example,

3

numbers of some native invertebrate species, such as pasture mealybug, are significantly

4

lower on endophyte-infected ryegrass (Pennell et al. 2005) and tall fescue (Pennell and Ball

5

1999). Although this suggests that effects on native species are likely, cereal crops make up

6

an extremely small area of land management in New Zealand – approximately 0.5% of the

7

total country’s size (Millner et al. 2013) and these areas are generally found to be low in

8

native invertebrate biodiversity (for example Fraser et al. 1996). Therefore any impacts are

9

likely to be localised and have little to no effect on the wider population dynamics of native

10

species.

11

Given the wide range of alkaloids that could potentially be produced by endophytes in cereals

12

it is also important to consider risks to vertebrates, including both animal and human health.

13

To manage this risk, the applicant selected particular strains that have alkaloid phenotypes (or

14

chemotypes) with anti-insect alkaloids, but very low, or no impacts on mammalian species.

15

Testing of the bioactive substances produced by these endophytes supported this, with tests

16

including mice bioassays using lolines (Finch 2012), testing of peramine using lambs

17

(Pownall DB et al. 1995) and testing of chanoclavine in livestock (Fleetwood 2007). The

18

applicant also conducted mouse bioassays for loline, peramine and indole-diterpenes and

19

found no indications of sub-chronic or chronic toxicity that could impact livestock health or

20

food safety (AgResearch, unpublished data). While these results indicate that the

21

Neotyphodium-cereal associations are likely to be benign to grazing animals, unexpected

22

toxicity can arise from novel endophyte associations in response to localised conditions. For

23

example, 48 horses grazed on Mediterranean tall fescue infected with Neotyphodium

24

coenophialum in Australia subsequently developed Equine Fescue Oedema (EFO). Four

25

animals in that instance died, and (Bourke et al. 2009) reported “It is significant that the 15

1

period of pasture toxicity…followed a protracted period of dry weather that ended with

2

sufficient rainfall to initiate pasture growth. In all the outbreaks [of EFO] so far there has

3

been an extended period of dry weather, varying in length from 4 to 6 weeks, prior to toxicity

4

developing”. The implication is therefore that the toxins are either produced or concentrated

5

under specific environmental conditions, and with knowledge of these conditions, toxicity

6

resulting from new host/endophyte relationships can be managed. Reid et al. (1978) indicated

7

that the effect is a result of the interaction between pasture growth following drought, and

8

grazing behaviour. Furthermore, trials with this endophyte/Mediterranean-type fescue

9

combination in the US failed to duplicate the Australian experience (Ray Smith, personal

10

communication, 2013), suggesting that events such as these are rare.

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4.2 Competitive advantage

12

While the motivation for using fungal endophytes in cereals is to increase resistance to insect

13

pests and plant pathogens, this could be considered a hazard due to the potential for cereal

14

crops to become weedy. Evidence suggests that endophytes in wild relatives of cereals

15

provide improved drought tolerance of the host plant (Zhang and Nan 2007; Zhang and Nan

16

2010), so any risk assessment must establish whether these traits provide a competitive

17

advantage, creating ‘super-cereals’ that outcompete other valued plants. Modern cereals are

18

typically grown in conditions of tilled and fertile soils, are intensively managed for weeds,

19

and are generally poor competitors. In addition, cereal growers traditionally select for larger

20

seed sizes rather than aggressive growth characteristics, or competitive advantage. Therefore

21

unless endophytes confer an extreme competitive advantage they are unlikely to become

22

weeds, although quantification of the precise changes in competitive ability are difficult. It is

23

increasingly apparent that any effect is dependent on environmental context, including the

24

assemblage of plant competitors and insect herbivores (Richmond et al. 2004) and at least

25

one study has found a competitive decrease when the grass is infected with an endophyte 16

1

(Faeth et al. 2004). Therefore while in isolation the performance benefits can be large, there

2

is no indication that in complex real world environments, where a variety of successful weed

3

species are present, that cereal crops would outcompete these and obtain weed status.

4

4.3 Endpoint valuation

5

Based on the information available, the EPA did not identify any adverse effects, risks or

6

costs from the release of Neotyphodium (EPA 2014a). The EPA also considered that the

7

potential benefits to New Zealand’s agricultural industry and the economy were significant,

8

and that novel Neotyphodium-host plant associations may increase host plant resistance to

9

insect pests and disease, which has potential flow on beneficial effects, including:

10

•

synthetic chemicals and increased cereal crop yields; and

11 12 13

Improved farm productivity, specifically due to reduced costs of applying

•

Environmental and human health benefits as a result of reduced agrichemical use.

14

On this basis the EPA approved the proposal, allowing 45 non-sporulating strains of

15

Neotyphodium to be imported and released into New Zealand (EPA 2014b).

16

5 CONCLUSION

17

A risk assessment model is useful for assessing any new biotechnology that might pose risks

18

to people, the environment or the economy. While novel associations of microbes and plants

19

could provide wide ranging benefits, they also pose risks. In the case studies presented, we

20

have provided an explanation for how the ERA process developed in New Zealand can be

21

used to make a practical determination. We have found the model can be used to identify the

22

biophysical risks of developing new endophyte-plant associations, which could also be

23

applicable in instances where GM technology is used. At its core, the ERA process is based

17

1

on a comparative analysis of benefits and risks, combined with a binary qualitative

2

framework. It treats new traits as hazards and is agnostic to the method of modification.

3

We recognise that any ERA process needs to be flexible to the context and underlying biology

4

of the organisms. As novel associations between hosts and endophytes are being created, as

5

well as efforts to explore new traits and properties that endophytes can embed into a host, a

6

larger number of hazards and their interactions will be identified. Furthermore, recent

7

advances now include the use of bacterial endophytes with potential applications such as

8

biological control, phytoremediation and the sustainable production of biomass (Ryan et al.

9

2008). We consider that the model presented here begins to fulfil the requirements of

10

establishing an ERA framework. It can be used by researchers when selecting host plants,

11

microbes or GM modifications to understand a regulator’s perspective and undertake

12

hypothetical risk assessments. It can also be used by regulators to assist in their decision

13

making process. The model can set basic standards that pave the way for the adoption of the

14

framework by multiple parties, which will in turn establish standards, familiarity and set a

15

benchmark in risk analysis.

16

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17

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11

26

1

Figure 1. The five components of risk analysis

27

1

Figure 2. An outline of the risk assessment process

2

28

1

A risk assessment model is useful for assessing any new biotechnology that might pose risks

2

to people, the environment or the economy. While novel associations of microbes and plants

3

could provide wide ranging benefits, they also pose risks. In the case studies presented, we

4

have provided an explanation for how the ERA process developed in New Zealand can be

5

used to make a practical determination. We have found the model can be used to identify the

6

biophysical risks of developing new endophyte-plant associations, which could also be

7

applicable in instances where GM technology is used. At its core, the ERA process is based

8

on a comparative analysis of benefits and risks, combined with a binary qualitative

9

framework. It treats new traits as hazards and is agnostic to the method of modification.

10

We recognise that any ERA process needs to be flexible to the context and underlying biology

11

of the organisms. As novel associations between hosts and endophytes are being created, as

12

well as efforts to explore new traits and properties that endophytes can embed into a host, a

13

larger number of hazards and their interactions will be identified. Furthermore, recent

14

advances now include the use of bacterial endophytes with potential applications such as

15

biological control, phytoremediation and the sustainable production of biomass (Ryan et al.

16

2008). We consider that the model presented here begins to fulfil the requirements of

17

establishing an ERA framework. It can be used by researchers when selecting host plants,

18

microbes or GM modifications to understand a regulator’s perspective and undertake

19

hypothetical risk assessments. It can also be used by regulators to assist in their decision

20

making process. The model can set basic standards that pave the way for the adoption of the

21

framework by multiple parties, which will in turn establish standards, familiarity and set a

22

benchmark in risk analysis.

23

29