HSE - Scientific Advisory Committee on Genetically Modified Organisms (SACGM)

Minutes of the first meeting of the Scientific Advisory Committee on Genetic Modification (Contained Use), held on the 21st January 2004 at Rose Court, Southwark Bridge, London

Minutes of the first meeting of the Scientific Advisory Committee on Genetic Modification (Contained Use), held on the 21st January 2004 at Rose Court, Southwark Bridge, London

Chair

Prof. Janet Bainbridge

Members

Prof. Martin Gore - Vice Chair
Dr David Lewis
Dr Brian Robertson
Dr Michael Skinner
Dr John Carr
Dr Penny Hirsch
Dr Phillip Minor
Prof. David Baulcombe
Prof. Peter Williams
Dr Peter Searle
Mr Bob Osbourne
Dr Gary Burns
Dr Martin Carrier
Prof. Ernest Gould
Dr Peter Coyle
Dr Sue Mayer
Dr Keith Howard

Secretariat

Dr Paul Logan
Mr Lee Wilson

Departmental Assessors

Dr Claire Pitcher - DEFRA
Dr Androulla Gilliland - DEFRA
Mr Paul Manser - DEFRA
Mrs Susan Shearman - DEFRA
Dr Leslie Wilkie - Scottish Executive
Dr Andrew Voas - Scottish Executive
Dr Andrew Cottam - HSE
Dr Michael Mackett - HSE
Dr Liz Pollitt - HSE
Ms Liz Sawyer - HSE
Ms Linda West - HSE

Guest Speakers

Dr Graham Belsham
Dr Martin Ryan

Welcome & introductions

1. The meeting opened with the Chair welcoming everyone to the first meeting. There were no apologies.

2. The Chair informed the members that she favoured an informal meeting style, with members addressing one another by first name. She would encourage full member involvement and make every effort to keep discussions within the agreed time frame. She went on to inform the Committee that a Web site would be established for the new Committee, and that it would be useful to include short background descriptions for each member.

Action: Members to provide a short paragraph about themselves and forward to the Secretariat for collation and inclusion on the new Web site.

Induction training for members

3. As this was the inaugural meeting of the new Committee, a short induction session was carried out. The induction covered: role and remit of the Committee; terms of appointment; working methods/decision making; role of the Secretariat; open government issues; declarations of interest and expenses.

4. The Secretariat outlined the Committee's future work plan including updating the Compendium of Guidance, and providing guidance on clinical applications including gene therapy. As part of the updating of the Compendium, the guidance on viral vectors, bacteria, plant and animal viruses would need to be reviewed and where appropriate updated. Furthermore, risk assessment enquiries and a number of individual notification reviews would be put to the Committee. Other areas of work would possibly include scientific aspects of guidance on the Contained Use amending regulations, and research. It is anticipated the Committee will meet three times a year. The need to make minutes and papers publicly available was discussed and agreed. A Committee Internet site would be established, and papers published on line.

Future name of committee

5. The Chair introduced the topic, and invited members' views on an appropriate new name for the Committee. The Chair suggested that the Committee should be called "the Scientific Advisory Committee on Genetic Modification (SACGM)", as this would encompass historical information from the old ACGM Committee, which would be useful, especially when interrogating the Internet. There was a short discussion on the subject, with members suggesting that the name should include a reference to "contained use". This was seen as important in distinguishing between this Committee and ACRE (Advisory Committee on Releases into the Environment). The title "Advisory Committee on Contained Use (ACCU)" was proposed, however it was thought that this might imply that it covered the work of the ACDP (Advisory Committee on Dangerous Pathogens). Furthermore the reference to GM was considered important. The Chair stressed that it was important to maintain links with historical information, especially when updating the Compendium. Members agreed that SACGM should be used, and that reference to "Contained Use" should be made in the logo. The Secretariat stated that HSE's publications section would be asked to help draw up a logo that incorporated both the name and referred to Contained Use.

Action: Secretariat to circulate draft logo.

Risk assessment enquiry: Invasive E.coli K12 cloning host - containment and classification - GMO (CU)/SC/2004/01/P2

6. The Chair introduced the paper and the Secretariat summarised the key scientific issues. HSE had been asked for advice on the classification of a modified E. coli K12 strain that had been engineered to express the inv gene from Yersinia pseudotuberculosis and the hly gene from Listeria monocytogenes from a low copy non-mobilisable plasmid. The modification, which allowed the bacterium to enter mammalian cells and to break out of the endosome, was being developed as a novel method of delivering genetic material to mammalian cells. The strain being used was a DAP auxotroph, and was degraded once inside the cell. However, the genetic material could be expressed and would elicit an immune response.

7. The original proposal that had led to the enquiry was to use the system to deliver clones of mutant vaccine strains of Marek's disease virus (MDV) into cultured avian cells. If successful it would be used in animal (poultry) studies. Concerns had been raised over the chances of these and similarly modified bacteria getting into the gut of an animal, and transferring the plasmid carrying the inv and hly genes into a wild-type E. coli. The Committee was informed that there were few data available on work with this particular E. coli strain.

8. The question of whether or not the strain could colonise the gut was raised. Could it survive long enough to pass on the inv and hly genes to wild type E.coli? Members considered that the disabled strain could survive for some time, but would not colonise the gut.

9. Members agreed that the main concern was that the inv and hly genes could be transferred into a wild-type background, and produce an invasive E. coli. It was recognised that the plasmid was a low copy number non-mobilisable plasmid. However, concerns were raised that this was not a 100% barrier to gene transfer.

10. As the E.coli strain is both a K-12 derivative, and a DAP mutant it was unlikely to survive inside aanimal cells, and would be rapidly degraded. Therefore it was acceptable to give the cloning host the lowest level of classification. The final classification would be dependent on the genes being delivered.

11. The issue of using the system to deliver herpes viruses was discussed, as it could be used to deliver viruses to cells to which they would not normally have access. It was agreed that it would be useful to update the Compendium regarding the development of this type of vector system.

12. The Committee went on to discuss the use of this system in the development of vaccines. Members noted that if this was developed as a vaccine system for use in chickens there may be the potential for gene transfer - as small probability events might be realised if the system was widely used. Members were reminded that the likelihood of recombination was extremely low. It was noted that current practice is that all chickens are inoculated against MDV and there is, therefore, potential for this construct/system to be used on a very large scale, creating greater potential for the small risk of recombination becoming a reality.

13. The Secretariat reminded the Committee that the risk assessment provided to HSE covered only the early stages of the work, which may eventually be used in vivo in chickens. If the project moved on to a much larger scale and progessed to trial stages, it would have to go through the veterinary products licensing process. This would probably alter the risk assessment, as it would require consideration of the numbers of animals and the potential for environmental spread. It was suggested that in the absence of available data, this would be a good area for research to be undertaken to look at the safety of the system.

14. The Chair summarised the discussion. Clearly this is a useful transfer system that may become widely used in the future; therefore there is a need for guidance, which should be produced by this Committee. Level 1 containment was considered appropriate for the host. However, the assessments must be made on a case-by-case basis, and consider the effects of the gene sequences being transferred and expressed. HSE would consider the need for research after consulting with colleagues in Europe and the USA.

Action: Secretariat to obtain more information on how the system is currently being assessed by the user. HSE to consider the need for future research on the likelihood of gene transfer, to and from the modified E. coli (including to the host animal), in the gut of infected animals and guidance on the use of the strain.

Safety of viral replicons: use of a foot-and-mouth disease virus replicon system - GMO(CU)/SC/2004/01/P3

15. The Chair introduced the paper and the Secretariat reminded members that the Committee was being asked to consider the scientific issues relating to the use of viral replicon systems, and in particular the use of an FMDV replicon. HSE had received a risk assessment covering the proposed use of the FMDV replicon, and was seeking advice, which would be provided to the other parts of the Competent Authority, who had a licensing role for high hazard animal pathogens under the Specified Animal Pathogens Order. A three-part review had been produced dealing with:

• Scientific policy and regulatory issues (Part A);
• A review of the use of RNA viral replicon systems (Part B);
• Specific safety issues raised with HSE (Part C).

16. A set of questions had been included in Part A, and HSE was seeking the Committee's scientific advice to help answer these. Members were reminded that replicon technology was being widely used in research, and this use was certain to develop further. Furthermore, as a result of the WHO eradication programme, consideration would at some stage need to be given to the safety aspects of the poliovirus replicon system, and the following discussions may inform that debate.

17. There then followed two short presentations to the Committee. First, Dr Graham Belsham, from the Institute for Animal Health (IAH) at Pirbright, gave the background to the development of the FMDV replicon system. After giving an overview of the molecular biology of FMDV and the picornavirus replication cycle, he went on to describe the replicon system.

18. Dr Belsham informed members that recombination between replicon RNA and capsid sequences to generate live virus could possibly occur if both were expressed within the same cell. However, he explained that there appeared to be some evidence that both the replicon RNA and the capsid RNA have to be in replication complexes, as is the case with pestiviruses. Finally Dr Belsham explained that the complexity of the capsid, which comprises 60 copies of the 4 structural proteins VP1 - VP4, means it is very unlikely that any sequences other than FMDV capsid sequences could encapsidate the viral RNA.

19. There followed a discussion on trans-encapsidation and recombination. The summary of these discussions has been incorporated into the answers to the questions and is given in Annex 1 to these minutes.

20. Dr Martin Ryan from the University of St. Andrews Department of Biological Sciences then gave a presentation on his work with FMDV proteins, in particular on the mechanism of the 'primary' 2A/2B polyprotein cleavage, and wider studies on the 2A and 2C proteins. He informed the Committee as to why he would like to work with the replicon system. He outlined the studies he proposed to carry out, with a view to increasing our understanding of FMDV molecular biology, which may assist the development of a vaccine system. Dr Ryan reminded members that the Royal Society report, which was published in the aftermath of the last FMDV outbreak, had recommended that the UK FMDV research base should be expanded.

21. Dr Ryan explained that the risk assessment he had submitted to HSE was not intended to be the "finished article". It did not contain He then gave members some background to the risk assessment he had submitted to HSE. He asked members to note that there was an experimental programme of work, and the assessment had been used to test the 'opinion' of co-workers (academic, technical, secretarial, manual) to the proposal that work with the FMDV replicon could be carried out safely at St. Andrews University.  Dr Ryan said that he had agreed that if he were to obtain the replicon, he would destroy his existing clones encoding the capsid protein sequences. On the subject of biosecurity, Dr Ryan stated that he considered this a matter for HSE / SE / DEFRA to decide on the appropriate level of containment. He said that the local committees had raisedquestions as to whether containment level 1 was appropriate. Dr Ryan then recommended a range of additional security measures that in his opinion would be appropriate including, vetting of staff, restricted access to replicons, non-release to third parties, use of secure storage facilities and standard working practices (SOPs), controlled waste disposal, strict record keeping / periodic local safety committee audit of records, and unannounced regulatory inspections.

22. Dr Ryan finished his presentation by referring to the Wellcome Trust's statement on Bioterrorism and Biomedical Research, and suggested that contracts could be drawn up detailing conditions under which the replicons could be released and worked with at sites other than within the containment level 4 facility at Pirbright.

23. Dr Ryan then answered members' questions, which focussed on his proposed working practices and procedures. Once again the detail of the discussions has been incorporated into the answers to the questions in Annex 1.

Any Other Business

Members were informed that the date of the next meeting would be sent out after the meeting. The date has now been set for the 19^th May.

There was no other business and the meeting closed at 16.10

Annex 1

Questions from Part A of paper GMO(CU)/SC/2004/01/P3

1) How far can we read across information from other viral systems, for example, poliovirus, to inform our assessment of work involving FMDV?

The Secretariat explained that the purpose of the question was to see whether members considered that the approach taken in the review was valid, that is, can what is known about other viral systems, and in particular picornaviruses, inform our understanding of the molecular biology of FMDV. It was noted that the review had covered the existing literature on FMDV, but had also drawn extensively on data relating to poliovirus.

Members agreed that it was valid to consider the molecular biology of picornaviruses, particularly poliovirus, when considering the safety aspects of the FMDV replicon system.

2) Are there any areas not covered in the review that could impinge on the safety of work with the FMDV replicons?

Members agreed that the review was comprehensive, however, it was noted that the issue of bacterial recombination hadn't been considered, that is, recombination between the bacterial plasmid carrying the cDNA coding for the replicon, and other plasmids carrying the capsid genes. It was noted that when the replicon RNA is expressed from the cDNA clone, it is not generally purified prior to transfection. Consequently bacterial plasmids carrying the replicon sequences are co-transfected with the replicon, meaning that DNA recombination could also be an issue for consideration if any DNA in cells contained capsid-coding sequences.

There was some discussion on this, however, members concluded that it would only be an issue if the laboratory had clones carrying the capsid genes. Recombination with other sequences was not considered to be an issue.

3) Do members consider that all the replicons described by McInerney et al. should be treated as requiring the same containment, or is the replicon with the largest deletion "safer" than the other functional replicon?

The Chair noted that in her opinion this question was answered in the presentation given by Dr Belsham. Essentially all the replicons with deletions in the capsid genes could be treated as requiring the same levels of containment, and were equally safe. Dr Belsham confirmed to members that all the capsid deleted replicons behaved in the same way. He was asked whether or not the residual sequences from P1 (1D, and part of 1C) could act as a "hotspot" or focus for recombination, and stated that this was unlikely, and that the capsid genes would need to be present for recombination to occur.

4) Does trans-encapsidation pose a risk? Are there any molecular mechanisms that could lead to a risk to the environment or human health following trans-encapsidation?

Members were in general agreement that trans-encapsidation did not pose a risk. It was noted that if trans-encapsidation of FMDV replicon RNA did occur, for example with poliovirus, the resulting virus would behave more like poliovirus than FMDV in terms of species specificity, transmissibility and detection, and would need to be followed by recombination to produce a novel virus. The absence of the capsid genes means that recombination that leading to the production of an FMDV like virus would not occur. Members considered that packaging constraints together with the need for a recombination event meant that the likelihood of producing a viable infectious hybrid was extremely low, and that the resulting virus would adopt the properties of the packaging virus, rather than FMDV. Dr Belsham confirmed that his group had tried to get encapsidation of FMDV replicons by FMDV capsids with little success.

It was noted that trans-encapsidation could lead to a single cycle infection and this may induce an antibody response. The question as to whether or not this could lead to a "false positive" antibody test, and cause significant economic impact was raised. Dr Belsham explained that the primary test for FMDV was based on an antibody response to the FMDV capsid proteins. He agreed that antibodies to other proteins expressed from non-capsid genes were assayed as secondary confirmation, but only following an initial capsid based test.

The overall view of members was that potential trans-encapsidation of replicons does not pose a risk either to human health or the environment.

5) When considering the issues relating to recombination:

5a) What factors will influence successful recombination in a laboratory environment? Could these arise accidentally, for example, in an "uncontrolled" laboratory environment?

Members considered that this would depend on a number of factors; however, recombination under non-ideal conditions was highly unlikely. Once again it was noted that the recombination event would need to be between the FMDV replicon and RNA from plasmids carrying the capsid genes. Members observed that in the risk assessment submitted to HSE the work had been classified as requiring containment level 1. This might mean that the segregation of waste material was not rigorous, and the opportunity for recombination could arise between plasmids in waste. Nevertheless recombination was considered highly unlikely, particularly as the bacteria or cells would require optimal growth temperature of 37^oC, which meant that recombination was even less likely, and the recombined nucleic acid would have to survive to be taken up by permissive cells or be [accidentally] inoculated into a susceptible animal. However, it was noted that if the laboratory had both clones carrying capsid genes and the FMDV replicon then recombination was a potential problem. The issue of plasmid recombination rather than RNA recombination was again raised.

Dr Ryan repeated that should he obtain access to the replicon system he would destroy all his stocks of clones carrying the capsid genes. He stated that current practice in his laboratory was to autoclave all waste containing any FMDV sequences.

Overall, members agreed that recombination was only an issue if the laboratory had both the replicon and the capsid sequences, but unintentional recombination was still an unlikely scenario.

5b)Could capsid genes from other picornaviruses, such as poliovirus, recombine with the FMDV replicon? Do shed vaccine strains pose a risk?

Members considered that it was highly unlikely that a viable virus would be produced by such a recombination event. It was further noted that such a virus would be more like the virus donating the capsid genes rather than FMDV. Furthermore, there were very few viruses related to FMDV. Shed vaccines, such as poliovirus vaccine, were not considered a likely source of capsid. Recombination under non-ideal conditions was not a realistic concern, and even if it occurred it would produce a polio-like virus, which would be highly unlikely to be viable. It was noted that in plants there was little evidence of new viruses emerging as a result of a recombination between two unrelated positive strand viruses. Members noted that recombination does occur within the enteroviruses, and was an important feature of viral evolution. However, although it occurs widely between viruses in the species human enterovirus B (cocksackie viruses B1 to B6, echoviruses and enteroviruses), it is not seen between viruses outside the group. It was generally considered that recombination would only occur with another FMDV virus, but was a theoretical possibility with another apthovirus. The only other known apthovirus was equine rhinitis virus A, which has a different host range making recombination in cell culture highly unlikely. Dr Belsham informed members that another bovine apthovirus had been discovered but was as yet undescribed.

Members noted that FMDV was endemic in a number of countries, and although recombination was seen between FMDV serotypes, there was no evidence of recombination with other types of virus.

In summary members considered that recombination was a highly unlikely scenario. The most likely candidate would be equine rhinitis virus A, although this was highly unlikely.

5c) Could recombination events take place between distantly related viruses and the FMDV virus from which the replicon was produced?

Members were all in agreement that this was a highly unlikely scenario. The more distantly related, the lower the likelihood of recombination. Given that recombination between viruses within the Picornaviridae was limited to viruses within the same group, such as between human enterovirus B's, members did not consider that recombination could occur between a FMDV replicon and a distantly related virus.

5d) Does the completeness of the genetic sequence in the replicon affect the risks of recombination, e.g. if the replicon contains 80% of the FMDV genome, does it pose a higher risk than a replicon with 65% of the genome?

Members noted that the potential for recombination depended on what sequences were present. Consequently in an 80% replicon there were more likely to be "recombination hotspots" than in a 60% replicon. However, recombination to produce infectious FMDV would require the presence of capsid genes, and there was some evidence that recombination only occurs between actively replicating RNAs. The best practice would be to use the shortest sequence possible to do the experiment. The question was raised as to whether or not researchers could delete the leader protein to make the replicon safer. This was not considered a viable option as it was considered to be crucial to its research use.

5e) Are there particular sequences that affect the risk, e.g. would a replicon with 50% of the genome, but with sequences that contribute to overall pathogenicity, pose a higher risk than a replicon with 75% of the genome, but without such sequences? In other words, should risk management be influenced by the nature of the sequences rather than the overall quantity of genetic material?

It was noted that it was difficult to define which sequences were responsible for virulence. The question of whether or not polio/FMDV hybrids could have altered pathogenicity was raised. However, members considered that such a hybrid would be predicted to have a much lower level of virulence due to incompatibility between structural and non-structural proteins and the RNA. Dr Belsham considered that such a hybrid would be non-viable because the 3C proteins were so different.

6) Do the cell lines being used pose a significant risk?

6a) Should work be limited to established cell lines, or could primary bovine cell lines be used?

Members were informed that primary bovine cell lines are used in research. Some members suggested that only established, characterised cell lines should be used if the replicon system was going to be used outside of containment level 4. This was considered to be a prudent measure to increase the safety of the work. Other members noted that the earlier discussions had led to the conclusion that the replicon would only recombine with FMDV capsid sequences, which are not likely to be present, even in primary cell lines. Nevertheless it was considered that even though this was an unlikely event, use of established cell lines would be good practice.

6b) could cell lines used in work on replicons be contaminated with the missing genetic material (capsid proteins)?

This was not considered to be likely, but would depend on the origins of the cell lines. If the cell lines were derived from material obtained from an infected animal it was a possibility. However, given that the UK and Europe is currently free of FMDV this was not a realistic scenario. Despite this members considered that the use of established cell lines would further militate against this remote possibility.

6c) could cell lines be effectively screened to guarantee that they did not contain other members of the virus group? If so, how would this need to be done?

Some members considered that cell lines could effectively be screened, by PCR/RT-PCR. However, these methods (RT/PCR) would only detect capsid if it were being expressed. Furthermore it would depend on the nature of the primers used. Dr Belsham added that in practical reality it would not be possible to screen for the presence of other viruses. The numbers of serotypes of particular viruses meant that the results would be very dependent on the primer used, and guaranteeing that the cell line did not contain another picornavirus would be difficult. The question was raised as to whether or not a general screen for the 5^/ non-coding region of a picornavirus could be used. If such a sequence was found, then another cell line could be used. Members generally did not consider that it was feasible or required, given the absence of infection with FMDV in the UK and Europe.

7) Given current experience with replicon systems, how can we minimise the risks of any recombination events within a laboratory:

7a) Should experimental work and storage of replicons derived from serious pathogens be carried out in dedicated/separate facilities away from work with other pathogens/replicons in the same laboratory?

Where the replicon system could be used to generate infectious virus, standards would have to be appropriate for work with that virus. Some members considered that researchers should never work with any more than one virus at a time. This was considered to be standard practice at the higher levels of containment, however, at lower containment, mixed working was common. Separation could be obtained by dedicating safety cabinets and incubators. Freezer storage of samples was likely to be in communal freezers, although clear marking of samples, the use of locked storage boxes, and appropriate training of staff would all mitigate against accidental mis-use of the replicon system. The use of local rules and SOPs, together with good laboratory practices was considered essential. There should be no storage of the clones carrying the capsid sequences and the replicon in the same laboratory or freezers.

The classification of the work as containment level 1 was considered to be a major problem, and would make segregation more difficult. Dr Ryan informed members that the classification did not take into account additional measures that would be required for bio-security reasons, and that if he were to obtain the replicon, strict security would be enforced.

7b) What practical measures can be taken to ensure adequate separation of different replicon systems/viruses within the same laboratory?

Members were in agreement that separation of work with the replicon from work with other viruses was appropriate. Some members considered that to ensure safety you would need to use dedicated areas, with complete separation. The fact that most academics have shared use of facilities was noted. Some members reminded the Committee that the earlier discussions had highlighted that the only realistic scenario for recombination leading to virus production was co-expression of the FMDV replicon and the FMDV capsid sequences. If there was absolute segregation of these sequences then there should be no safety concerns. Good segregation and good laboratory practice were considered crucial.

7c)What precautionary measures or category of containment, would be advised where work was to be undertaken with replicons of viruses that are categorised as DEFRA 4 (i.e. are either exotic or produce notifiable disease and have a high risk of spread from the laboratory, and severe impact on the wider environment and rural industry in the event of an escape)?

There was a lot of discussion as to what the appropriate level of containment should be for different replicon systems. It was agreed that a case-by-case approach was required. The question of whether or not these replicon systems should be treated as genetically modified viruses was raised. It was noted that the FMDV replicon system could only replicate RNA and not package RNA into viruses. Viruses require both replication of the RNA/DNA and subsequent packaging. Furthermore, Dr Belsham informed members that the FMDV replicon system kills the cells within a few hours of transfection, presumably through toxicity of expressed proteins, or the hijacking of the cellular transcription and translation machinery.

Members agreed that if a replicon system were capable of producing virus then the containment level would be at least the minimum required for the parental virus. Furthermore if there was any possibility of producing virus then the containment level should be appropriate to work with the virus.

There was general agreement that containment level 2 should be used as a minimum for the FMDV system, with additional measures covering segregation and access to the facility. The risk assessment submitted to HSE stated that access would be restricted and that gloves would be worn. Members noted that these were measures from containment level 2, so this was the minimum standard necessary.

The issue of bio security was raised. It was noted that there were major implications for centres working with a range of viruses and bacteria, if containment levels were greatly increased for security reasons. Members were concerned that it should be made clear what measures were required to work safely with the replicon system, and what additional measures were required for "security" reasons. The issue of whether or not the system posed an "inherent risk" should be balanced with any risk to "national security". For example, could someone obtain both the replicon and clones carrying the capsid sequence and artificially re-create the virus? Members noted that this would be an illegal activity and would require intentional recreation of FMDV. Control of the spread of the capsid genes was considered necessary to mitigate against this. One member asked what controls were currently in place, and Defra officials confirmed that movements of genetic material derived from specified animal pathogens is controlled under the Specified Animal Pathogens Order as well as under the Genetically Modified Organisms (Contained Use) Regulations.

One member asked whether or not it is realistic to consider that the FMDV replicon system poses a risk to national security? A more likely scenario was that anyone who wanted FMDV would bring it in from outside the UK, from areas where the virus is endemic. The scientific discussion concluded that the risk from use of the replicon was extremely low.

8) If a replicon were introduced into an animal or human would it produce an antibody response? Would this be the same for all replicons?

Members noted that if directly injected, replicons could induce an immune response to non-structural proteins, whereas if ingested, they would be destroyed through RNAse activity. However, one member commented that if a microgram quantity was ingested then sufficient might pass through the stomach to induce an antibody response. Other members considered that this would require very poor working practices and is an unlikely scenario and was very theoretical. Furthermore the screening procedures are only carried out in farmed animals, and it would be impossible for such a quantity of replicon RNA to get into an animal accidentally.

An antibody response would be induced if the replicon was injected into an animal. Dr Belsham reminded members that the primary diagnosis for FMDV is based on antibody response to the capsid proteins. The antibody response to the non-structural proteins is used as a secondary test. The secretariat asked for confirmation that the antibody response would be based only on the FMDV capsid and consequently even if there were a mechanism by which the replicon could get into an animal, it would not give a positive antibody response to the standard initial diagnostic test. Members agreed this was the case.

9) Given the early stages of the use of this technology, which precautionary approaches does the Committee recommend that will allow work to take place whilst addressing risks, for example:

9a)case-by-case assessment and containment;

Members agreed that containment for all replicon work would require case-by-case assessment. Where the replicon system is combined with a packaging system to produce infectious virus, containment would be based on that required for the virus. For disabled systems such as the FMDV replicon where no virus can be produced a lower level of containment would apply. The general feeling from members was that containment level 2 was appropriate based on the need for restricted access and the need to wear gloves.

9b)restriction of work with other viruses/replicons systems;

Some members considered it would be appropriate to restrict the work with FMDV replicon to a dedicated laboratory, however, the Secretariat noted that in universities it would be unusual to have a laboratory dedicated to an individual project and sharing was likely. Members noted that Dr Ryan had said that he would destroy his capsid sequences and these were realistically the only sequences that could potentially result in production of infectious virus by recombination.

9c) qualifications and experience of staff;

It was noted that in Universities, post-graduate students are usually recruited with little experience and trained on the job. Appropriate training would be required with written training records. Dr Ryan stated that although he had a large research group he had experience of working with infectious FMDV and was fully aware of the expected standards and procedures. He stated that anyone working with the replicon would be fully trained in working practices and procedures.

9d) use of SOPs;

It was generally agreed that standard operating procedures (SOPs) should be used for this work, and that workers should be appropriately trained.

9e) overall laboratory security/containment?

Members again reiterated that it was important to distinguish between bio security and biosafety, and that HSE should not set precedents that could have a major impact on industry as there were a wide range of organisms for which there are bio security concerns and this should not drive the biosafety containment level.

The Secretariat agreed that it is important to separate the two issues. The risk assessment leading to containment should be carried out as described in the Genetically Modified Organisms (Contained Use) Regulations 2000, and any additional security measures should be added afterwards.

A suggestion was made that one solution would be to classify all replicons based on pathogens at a minimum of containment level 2 and decide the final level of containment on a case-by-case basis in consultation with the Competent Authority.

The Secretariat noted that this would be a retrograde step, pointing out that the Genetically Modified Organisms (Contained Use) Regulations 2000 were introduced to be more risk based and that it would be wrong to change the whole system just to deal with this FMDV replicon case. It was noted that replicon technology is extensively used for work with plant viruses, and a blanket approach to classification could hinder a lot of research that is currently carried out at lower levels of containment.