Guideline for Blowfly Specific (Flystrike Preventatives and Treatments) Efficacy Submissions

Guideline 21

1 Introduction

Blowfly strike can be a devastating disease of sheep.  Trial work involving deliberate exposure of sheep to blowfly strike entails significant ethical and animal welfare responsibilities.  While the intention of these Guidelines is to ensure that sufficient blowfly pressure is present to adequately test the candidate treatment, it must always be remembered that the welfare of the trial sheep is to be considered paramount and any deliberate (flyhouse, larval implant) strikes be terminated as soon as the result is unequivocal.

1.1       These guidelines shall apply to all methods of applying chemicals for flystrike by the Australian sheep blowfly, Lucilia cuprina and other blowfly species.  As guidelines the protocols outlined below are not mandatory requirements.  The intention is not to stifle individual initiative or the development of novel technology that may not suit these guidelines. Instead, these guidelines are presented, following requests for direction by several companies with blowfly products under development, to assist companies that may not always be experienced in flystrike prevention/treatment or which are uncertain whether proposed project designs are feasible or appropriate.

For registration, the mandatory requirement is that flystrike preventative products must protect sheep from flystrike, for a specified period, when sheep treated with such products are subjected to reasonable fly pressure.  Ideally, reasonable flystrike pressure would be assessed by strike incidence in an untreated control mob but this is seldom acceptable to co-operative farmers who are not prepared to place sheep at risk.  It may also be undesirable in terms of animal care considerations.  As an alternative to using untreated control sheep, daily rainfall, temperature and fly trap records of catches of L. cuprina, or whatever data demonstrates adequate flystrike pressure, would be acceptable for products seeking registration. In these situations at least one standard, registered product should also used in the trial.  Hearsay by farmers or local advisory people is not acceptable in isolation.

Data from fly exposure shed trials, larval implant trials and laboratory bioassays may support, but not replace, field trial data. Some guidance for shed and larval implant trials is provided to facilitate these types of support trials but it is accepted that available facilities vary considerably and that companies will have their own protocols. The fly exposure house guidelines are provided to alert prospective applicants to the need for statistically significant results if this type of data is to be used to support a registration application. A biometrician should be consulted to determine group size, to ensure this requirement is met. With carefully supervised trials of this sort an untreated control group is recommended but evidence of egg laying by the released flies under flyhouse conditions would suffice.

1.3       These guidelines may be varied to suit different treatments, but major deviations (in draft protocols) should be discussed with the National Registration Authority prior to undertaking the field trial to ensure that the trial will still be valid, statistically or otherwise.

1.4       Trials must be carried out in Australia to demonstrate the value to the product under local conditions. Additional supportive data, collected overseas under similar climatic conditions may be submitted but shall not replace data collected in Australian trials.

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2 Field trials (Guidelines only)

It will be necessary for the applicant to demonstrate to the satisfaction of the reviewer, that the product when used as directed, will be effective in a range of geographic environments as well as in sheep of variable age, breed, sex, wool, length etc. This will normally require a minimum of 5 to 10 well conducted and strategically located Australian field trials. A reduced number of trials attracts the risk that, if one or more trials are deemed, for whatever reason, to be unsuitable further trials may be required or efficacy claims may be diluted. This could unduly delay registration.

Ideally, but not necessarily, field trials should be conducted under a range of climatic conditions. The goal is to conduct trials with large numbers of sheep when flies are active and when weather conditions are conducive to flystrike. Trials should enable useful information to be obtained on the following-

  1. Ability of the chemical and application method to prevent flystrike for a specified period.

  2. Pesticide persistence in the fleece. This would provide information on the effect of climate on the chemical in wool. This may influence (explain?) efficacy (protection period?).

  3. Ability to treat existing strikes (where applicable).

  4. Ability to prevent restrike (where applicable).

Conduct of trials

2.1       Sheep: Merino sheep, mulesed or unmulesed, should be used in the majority of flystrike trials for which information is presented. If breeds other than merinos are used, this should be stated. It is recommended that weaners or unclassed hoggets be used where possible to increase the likelihood of strike. Records of whether sheep have been shorn previously should be included.

2.2       Group size: Treatment groups should each contain at least 100 animals. Sheep in each group should be identified with numbered eartags or similar. Where applicable all treatment groups should be segregated after treatment until dry, and thereafter managed as a single mob. A similarly sized untreated control group (and/or a known standard treatment group) may be included but this is at the applicants discretion.

2.3       Staggered treatment option: This option is offered only to allow greater flexibility to trial design. It may be more appropriate to trials conducted on company's research farms. Trials of this nature could be conducted in autumn as well as spring and so could utilise sheep whose fleece had had the chemical "weathering" in it during summer.

If it is considered helpful in improving the chance of obtaining acceptable fly challenge at critical times after treatment, treatment times may be staggered. For example, if fly problems usually occur in a particular location in (say) late October, it may be advisable to have sheep which were treated 8, 10 and 12 weeks previously, so that if no further fly challenge occurs after October, information for these critical times will still be collected. Shearing times would also have to be manipulated. Treating at least 40 sheep per treatment, at each time, would be appropriate. If further flystrike occurs again later, more data is collected.

2.4       Details of application should be stated. Collection and submission of the following information is recommended:

  1. Technical information on the chemical involved eg. insecticide class (organophosphate, triazine, benzoylphenyl urea etc.), mode of action (if known eg. nerve poison, insect growth regulator etc.), any precautions for safe handling.
  2. Application technique (standard handjetting, Dutjetting, application pressure at the handpiece, backline application, automatic jetting race etc). Regardless of equipment used, any special requirements eg application apparatus, plus specific set up instructions regarding pump, pressure, nozzle setting etc should be given.
  3. Concentration applied (mg/L in solution) plus mean volume applied per sheep (L/sheep) (total volume of fluid applied/total number of sheep treated).
  4. Wool length (cms) at shoulder, back and hip (mean of 10 sheep selected at random) at time of treatment. Time since shearing does not always give a good indication of wool length as nutrition and seasonal conditions can affect wool growth. Weight of sheep at treatment (mean of 10 sheep selected at random).
  5. Dates of treatment applications.
  6. Number of sheep in each group.
  7. Location of trial.

Mention should be made of whether groups were segregated or mixed after treatment.

2.5       Non-standard treatments: Off-shears treatments for blowfly strike control are usually not recommended if they involve thorough wetting of the sheep ie. plunge or shower dipping. This is because of concern about post-dipping infections. However, such a recommendation is not precluded if adequately justified.

Flystrike trials for sheep in short wool (1-42 days after shearing) should compare the candidate treatment with a registered product with a similar claim for flystrike protection in short wool. At present this standard treatment could be an organophosphate treatment (chlorfenvinphos, diazinon or propetamphos) or pyrethroid product at the recommended blowfly rate. Cyromazine (Vetrazin, Jetcon) should not be used for comparison because it is not registered for this purpose. An untreated group may be included to demonstrate adequate fly pressure.

2.6       Details of daily rainfall, maximum temperature and wind velocity should be maintained from the time the sheep are treated until the completion of the trial. As well, it is recommended that fly trap catches of L. cuprina over a 24 h period each week should be recorded if no untreated control mob is included. Standard Western Australian design traps are recommended (see attached reprint).

Most farms will record daily rainfall as a matter of course. Weekly max/maximum temperature and records of days with windspeed exceeding 30 km/h would be adequate. These points are strongly recommended so as to avoid controversy regarding the adequacy of fly pressure during particular trials. Unless a reviewer is convinced that there was adequate flystrike risk at a particular time, that reviewer is obliged to discount that data from his/her appraisal.

2.7       Sheep should be inspected in the paddock daily during flywave conditions and at least every 3 days under less severe conditions. Flystruck sheep should be caught, the struck area clipped and treated and those sheep reinspected the next day to ensure that the treatment has been effective.

2.8       Records should be kept of the position, size, severity and date at which any strikes occur. Pictorial records are recommended. When a trial is conducted on a flystrike preventative/treatment whose efficacy is known, or suspected, of being affected by an existing resistance, it is recommended that the resistance level be determined by laboratory bioassay. This is to ensure that the product is adequately challenged by flies whose resistance level is typical of most areas of Australia.

2.9       Group size is critical to ensuring that statistically significant results are obtained. With group sizes of 100 sheep, there would need to be 0 % strike in a treated group, if only 6 % control strike occurred, for the result to be significant. Similarly, to be 95 % sure of proving that 5 % strike incidence in a treated mob was significantly different to 25 % strike in an untreated group, treatment groups would have to contain at least 90 sheep each. Information on critical group sizes for significant differences between treatments is available in Fleiss, J.L. ( 1981 ) Statistical methods for rates and proportions. 2nd Edition. Wiley, New York. (see attached table). In trials with untreated controls. 15% of control sheep (excluding restrikes) should sustain strikes throughout the observation period of the trial ie cumulative strike for the product to be considered adequately challenged. Depending on group size, the number of strikes in the treated group may have to be 0 for the result to be significant. Alternatively, in trials utilising positive controls where proof of adequate fly challenge is provided, the limit of the protection period of the product under review is reached where > 1% of treated sheep sustain strikes. This would include crutch strike if a claim is made for protection of the crutch, and/or if the crutch was treated.

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3 Semi-field/exposure house trials

Conduct of trials

3.1       Semi-field/exposure house trials may support, but not replace, field data.

3.2       Sheep: Merino sheep, mulesed or unmulesed, should be used in the majority of flystrike trials for which information is presented. If breeds other than merinos are used, this should be stated. It is recommended that weaners be used where possible to increase the likelihood of strike. Records of whether the sheep have been shorn previously should be included. Unless information on protection from crutch strike is required, sheep should be crutched prior to entry into the fly exposure shed.

3.3       Group size: Although it is acknowledged that the available facilities will place the ultimate constraints, in deciding treatment group size, consideration should be given to the need for statistically significant results. An untreated (control) group, and a standard treatment group are recommended. Extra control sheep may be used to offset any sheep which are dropped during the trial because of flystrike. All sheep, however, should be exposed to the same wetting regimen. Sheep in each group should be identified with numbered eartags or similar.

3.4       At the applicants discretion, treatment times may be staggered to reduce the number of fly releases (see 2.3 above).

3.5       Details of application should be stated. Information is required for:

  1. Technical information on the chemical involved eg. insecticide class (organophosphate, triazine, benzoylphenyl urea etc.), mode of act~on (if known eg. nerve poison, insect growth regulator etc.), any precautions for safe handling. v
  2. Application technique (standard handjetting, Dutjetting, application pressure at the handpiece, backline application, automatic jetting race etc.). Regardless of equipment used, any special requirements eg. application apparatus, plus specific set up instructions regarding pump, pressure, nozzle setting etc. should be given.
  3. Concentration applied (mg/L in solution) plus mean volume applied per sheep (L/sheep) (total volume of fluid applied/total number of sheep treated).
  4. Wool length (cms) at shoulder, back and hip (mean of 10 sheep) at time of treatment. Weight of sheep at treatment (mean of 10 sheep).
  5. Dates of treatment applications.
  6. Number of sheep in each group.
  7. Location of trial.

Mention should be made of whether groups were segregated or mixed after treatment.

3.6       Non-standard treatments: Off-shears treatments for blowfly strike control are usually not recommended if they involve thorough wetting of the sheep ie plunge or shower dipping. This is because of concern about post-dipping infections. However, such a recommendation is not precluded if adequate justification is provided.

Flystrike trials for sheep in short wool (1-42 days after shearing) should compare the candidate treatment with a registered product with a similar claim for flystrike protection in short wool. At present this standard treatment could be an organophosphate treatment (chlorfenvinphos, diazinon, or propetamphos) or a pyrethroid product at the recommended blowfly rate. Cyromazine (VetrazinR, JetconR) should not be used for comparison if treating sheep with less than 6 weeks wool growth as cyromazine is not registered for this purpose.

3.7       Details of temperature, rainfall, both real and simulated, should be maintained from the time the sheep are treated until the completion of the trial. Mention should be made of any attractant/oviposition stimulants applied to target sheep.

3.8       Between fly exposures sheep should be inspected at least every 2-3 days. Daily inspections of sheep are required while sheep are housed.

3.9       Records should be kept of the position, size, severity and date at which any strike occur. Pictorial records are recommended.

3.10     The trial may cease when 20% of treated animals have sustained strikes during the trial period (cumulative strike excluding restrikes). At least 20% control strike should occur after each fly release. Very high control strike at earlier fly releases, however, may preclude the inclusion of sufficient control sheep to reach a further 20% strike incidence unless extra sheep are kept. Cumulative strike incidence should be presented.

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4 Larval implant trials

Conduct of trials

4.1       Larval implant trials (Hughes and Shanahan 1978) may support, but not replace field/semi-field trial data.

4.2       It is recommended that only merino sheep should be used.  Age and weight of sheep should be recorded.  There should be a minimum of 3 sheep per treatment, but allowance should be made so that no sheep is implanted on more than 6 occasions. Group size should be recorded.  Up to 2 implant sites per sheep may be used on each.

4.3       Ideally, implants would be made using field-derived and insecticide-susceptible (LS strain) larvae but use of field-derived flies is sufficient. When a trial is conducted on a flystrike preventative/treatment whose efficacy is known, or suspected, of being affected by organophosphate resistance it is recommended that the organophosphate resistance status of field-derived larvae should be determined according to the method of Hughes and Levot (1987). It is suggested that only flies with a Resistance Factor of >20x (median field response) to diazinon should be used.

4.4       If larval implant trial results are submitted to support an application for registration, at least one treatment reported should be the proposed use pattern ie concentration and method of application.

4.5       Implants with Insect Growth Regulators/Insect Development Inhibitors should be checked 48 or 72 hours later. Larval implants onto control sheep, or sheep treated with fast-acting chemicals should be checked after 24 hours.

4.6       Implants are positive if any larvae are alive at time of checking. Larvae may be removed at this time and reared in the laboratory to determine whether affected larvae are capable of completing development, or killed insitu to avoid unnecessary stress to the sheep.

4.7       Details of application should be stated. Information is required for:

  1. Technical information on the chemical involved eg. insecticide class (organophosphate, triazine, benzoylphenyl urea etc.), mode of action (if known eg. nerve poison, insect growth regulator etc.), any precautions for safe handling.
  2. Application technique (standard handjetting, Dutjetting, application pressure at the handpiece, backline application, automatic jetting race etc.).  Regardless of equipment used, any special requirements eg. application apparatus, plus specific set up instructions regarding pump, pressure, nozzle settings etc. should be given.
  3. Concentration applied (mg/L in solution) plus mean volume applied per sheep (L/sheep) (total volume of fluid applied/total number of sheep treated).
  4. Wool length (cms) at shoulder, back and hip (mean of 3 sheep) at time of
  5. Dates of treatment applications.
  6. Number of sheep in each group.

Mention should be made of whether the treatment groups were mixed or kept separate after treatment.

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5 Treatments for existing strikes

Conduct of Trials

Flystrike dressings

5.1       Product claiming activity against existing strikes should demonstrate efficacy in killing active, advanced third instar field-derived larvae on sheep as this is the stage most likely to be detected by farmers. The number of sheep used should be the minimum required to demonstrate effectiveness. Strikes on sheep may be induced naturally or by exposure to flies in a fly house and/or by implant. This information should be supported by data from laboratory bioassays. Laboratory tests should mimic field use as far as possible eg larvae immersed in dressing solution. Treatment times should reflect realistic exposure times on sheep ie maximum 3 minutes exposure. Information on efficacy at shorter treatment times is recommended. When a trial is conducted on a flystrike preventative/treatment whose efficacy is known, or suspected, of being affected by organophosphate resistance it is recommended that the OP resistance level be determined by laboratory bioassay (see 2.8 above). An untreated control group is recommended, especially where the strikes are shorn back prior to product application.

Protection from restrike

5.2       Products claiming a period of protection from restrike should provide supportive data from trials with sheep.  Sheep which have had strikes dressed in accordance with label directions should be exposed to blowflies in the paddock and/or in a fly exposure house at intervals (eg. 3, 7, 14 and 21 days etc.) after treatment and inspected daily for egg masses and restrikes.  A standard treatment should be included for comparison.  Group size should take account of the need for statistically significant results. An untreated control group is recommended.  Records of egg masses laid onto dressed strikes and incidence of restrike will provide information on blowfly repellency and protection from restrike.

5.3       Details of application should be stated. Information is required for:

  1. Technical information of the chemical involved eg. insecticide class (organophosphate, triazine, benzoylphenyl urea etc.), mode of action (if known eg. nerve poison, insect growth regulator etc.), any precautions for safe handling.
  2. Application technique.
  3. Concentration applied (mg/L in solution); mean amount (volume) applied per strike.
  4. Any special requirements eg. application apparatus etc.
  5. Number of sheep in each group.
  6. Pictorial records of strikes and restrikes.
  7. Weather (rainfall, temperature).

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