WO2001011961A1 - Control of ectoparasites using spinosyns - Google Patents

Abstract

The invention provides formulations for controlling insects/pests in small ruminants, such as camellids, sheep, and goats, the formulations comprising an effective amount of a spinosyn, or an analogue or derivative thereof, together with an acceptable carrier or diluent, and methods of controlling these insect pests comprising topically applying these formulations to a ruminant.

Description

CONTROL OF ECTOPARASITES USING SPINOSYNS

This invention relates to A83543 compounds including analogues and

derivatives thereof, and in particular the application of such compounds as

insecticides in controlling lice, flies and other ectoparasites and related arthropod

pests which infest sheep, goats, other small ruminant species and camelids (including

alpacas, llamas, vacuna). Historically, the greatest damage to domestic animals has

been caused, and continues to be caused, by pests such as insects. Insects particularly

represent a cause for concern as they are the most numerous of all living organisms

and constitute approximately 72% of all animal species. Approximately 1% of insects

are considered pests in that they attack humans and/or domestic animals, transmit

human, animal and plant diseases, destroy objects and structures and compete for food

and other necessities.

The losses resulting from insect-caused human and animal diseases are

enormous. In fact, insects are considered to be the carriers of more than 250 viruses

which are pathogens of humans and higher animals. The numbers of human deaths

caused by mosquito-transmitted diseases such as malaria and lymphatic filariasis are

huge. Flies also transmit human- and animal-related diseases such as trachoma,

trypanosomiasis and river blindness. Other human and animal diseases are

transmitted by fleas and lice.

To date, the primary method for controlling insects, particularly in respect of

domestic animals and crops, has been by the application of synthetic chemical

insecticide compositions. It is estimated that there are at least 35,000 formulated

insecticide products worldwide with chemicals as the active ingredients. Such insecticide products include antimicrobials, larvicides, insecticides, animal dips,

avicides and disinfectants.

The extensive use of chemical insecticides since the 1940s has resulted

in a large number of problems, including widespread insect resistance, emergence of

secondary pests, hazards to human and animal health, as well as detrimental effects on

fish and birds and environmental pollution.

Many insect species have developed resistance to the action of specific

insecticides so as to necessitate changes in control practices. There is an ever

widening pool of insect pests which are developing multiple resistance. The resistant

genes persist in insect genomes, precluding successful reuse of an insecticide to

control an insect population with resistant genes. Thus, there are increasing reasons to

develop new insecticides due to insect resistance.

Plant-derived insecticides such as nicotine, rotenone, veratrine and

pyrethrum have also been used as natural contact insecticides to kill insect pests.

However, the disadvantage with some plant-derived insecticides is their toxicity to

humans, animals and fish.

Insecticide residues are also seen as a major problems resulting from

chemical usage. With the exception of microbial insecticides, nearly all insecticides

result in residues of various chemicals and their degradation products or metabolites

which may be present in detectable amounts (ppb to ppm) in food despite food

processing. With increasing use of insecticides, the nature and magnitude of such

persisting residues have assumed great significance in public health. Potential

environmental risks include threats to ground water or surface water (lakes, dams,

rivers, streams). The persistence, mobility and potential for accumulation of an insecticide and its primary degradates must be considered in order to assess the

environmental impact of the use of an insecticide. Concern about the fate of bio-

active chemicals introduced into the environment has led to strict regulations on

release of insecticide waste into water, as well as for proper disposal of containers and

waste from use of insecticides. Other factors such as corrosiveness, explosiveness

and flammability of the insecticide must also be considered.

Some of the currently used products, such as pyrethroids and

organophosphates (OP's), are potentially toxic to sheep, goats, other small ruminants

and camelids, to the human operator who applies the treatment, and to the

environment, such as when run-off or the effluent from wool processing plants is

discharged into water ways. Thus, a potentially broad spectrum of toxicities exists

with respect to insecticides used in the treatment or eradication of ectoparasites.

It is acknowledged that the use of insecticides is essential for high

yield production of quality wool and to protect sheep from attack by ectoparasites

(mainly lice and blowfly) through the growing season. Inevitably, however, the use

of insecticides leaves some residue in the wool at shearing time. Accordingly, wool

residues resulting from currently used ^'.. isecticides are of major concern. Particularly,

use of insecticides in late season treatments can lead to the occurrence of residues in

the fleece at shearing. Late season backline and jetting treatments have been studied

to calculate the dissipation of the active agents and to assess the spread of insecticide

through the fleece. Insecticide formulations using representative members of

organophosphate, synthetic pyrethroid or insect growth regulator insecticides

commonly used to control lice and flies on sheep often leave high levels of wool

residues when applied as late season backline treatments or as late season jetting applications. The residues resulting from such standard, currently used products have

been the cause of concern in the wool industry for some time. Further, the use of

these products can also result in environmental discharge limits being exceeded.

The two processing issues with the highest priority relate to 1) the

acceptability of lanolin products that are made from wool wax that is recovered during

scouring, and 2) the residues that associate with the wool wax that is discharged from

the scour with the processing wastes. Insecticides in the wool wax that is recovered

when the wool is scoured may partially survive the lanolin refining process unless

special steps are taken. For example, several years ago, diazinon was found in some

samples of pharmaceutical lanolin and was considered a potential health hazard to

infants.

Aqueous wool scouring produces high volumes of effluent containing

emulsified wool wax, dirt and insecticides. While this effluent is treated before

discharge to the environment, the impact on the environment depends on the extent of

effluent treatment and dilution after scouring and on the location of the scour. If

present in high enough concentrations in the effluent, insecticide residues can kill

beneficial aquatic fauna which help the process of sewage degradation. Once released

from sewage systems into streams or rivers, these chemical residues can interfere with

the waterway food chain.

Currently, Australia and other countries export a substantial amount of

their wool in a greasy form. That wool may be scoured and processed in many

different countries and environmental locations, leaving international environmental

issues as a major concern when considering wool residues. European environmental

authorities have commenced the introduction of tough new guidelines for discharge of chemicals into waterways. These regulations mean that both wool processors and

growers have to change their practices if European customers are to continue to

process wool. The total concentration of organophosphates and synthetic pyrethroids

in the Australian wool clip in 1994-1995 averaged 10 mg/kg (ppm) on greasy wool.

While this amount has been reduced over the last two years, Europe has indicated that

chemical residue levels on greasy wool will need to be reduced to 1-2 mg/kg over the

next 9 years in order to comply with new EC standards. For some pesticides, residue

levels must be even lower.

It is to be noted that the Australian wool industry has set a goal to

reduce the level of insecticide residues in the wool clip over the next few years, and

therefore new safer insecticides are required. Other wool growing countries in the

International wool network are also moving to reduce residues of all insecticides.

Europe, Canada, Japan and New Zealand all have eco-labelling schemes of wool and

textile products which focus on absence of toxic substances, etc. In order to

accomplish these goals, new, safer, low residue insecticides will be required.

Tissue residues are also a major concern when considering use of

insecticides on domestic animals. App!.^' cation of an insecticide to a domestic animal

may result in toxic chemical residues being collected in the meat/tissue of the animal.

Human consumption of meat or other products from domestic animals can result in

human ingestion of such insecticidal residues.

Accordingly, insect and pest control has been sought to be directed

away from exclusive reliance on insecticides and towards the optimisation of

environmental and economic insect and pest control (integrated pest management).

The application of microbial control in which insects are attacked by pathogens such as viruses, bacteria, fungi and protozoa are favoured as such microbial insecticides are

highly selective for insect pests and do not leave toxic residues. However, microbial

insecticides are not without their problems, such as the difficulty in applying them, as

well as confining the natural enemy/parasite/disease to a large area. Further, they also

have the disadvantages of short residual action and extreme specificity, which limit

general applicability.

Genetic engineering has most recently been applied in the area of

insecticides through the release of sterilised male insects and mass introduction of

deleterious mutations such as chromosomal translocations. However, such procedures

are very expensive, and stringent criteria are required before release of sterile males is

contemplated. Chemosterilants which sterilise large segments of insect pest

populations are also known, but these are strong carcinogens which precludes their

use.

The search for new substances or approaches to pests and insect

control has assumed increasing significance in public health in view of the use of

chemical insecticides (and their environmental and economic viability), the nature and

magnitude of the persisting residues, a; J increased insect and pest resistance, together

with the toxicity levels of many synthetic chemical as well as natural chemical

insecticides.

Of particular concern in the wool industry are the products used by

woolgrowers to control or prevent lice and fly infestations. Currently, such products

include synthetic pyrethroids, insect growth regulators and organophosphate products

which cover nearly all the registered external parasitic treatments on small ruminants

such as sheep and goats. All lice and fly products start to break down from the date of application, but some of the chemicals remain in the fleece and tissue, as discussed

above. This residual amount depends on the chemical used and the timing and rate of

applications prior to shearing.

One specific problem in the wool industry is the late treatment of

flystrike and lice in sheep before shearing. Sheep treated for flystrike and lice in long

wool are a major source of high chemical residues on wool. To date, while the

industry has sought measures such as introducing the lice and flystrike treatment early

in the season and before 6 months before shearing, these measures are not often

effective. Consequently, the search for a lice and flystrike prevention treatment which

can be administered in the last 6 months of the growing season and which will

produce wool with low chemical residues while still controlling lice and flystrike is a

major commercial objective in the wool industry.

There is, therefore, a need for compounds which can be used as one or

more active principles in insecticides, particularly in respect of insects including

ectoparasites which afflict small ruminant economic animals, especially sheep and

goats, and which are effective at low application rates, selective in biologic action and

have low toxicity and a high margin of afety to humans, economic animals, aquatic

organisms and birds. Such compounds should also be ones from which no persisting

wool, tissue or other residues result, as well as being substances for which no insect or

pest resistance exists. The substances must be environmentally friendly in that there

must be demonstrably low impacts on the environment. They must also be

economically viable to use on a large scale, particularly in respect of late season

applications, that is applied within the last six months of the growing season. Fermentation product A83543, also known as spinosyn, includes a

family of related compounds (spinosyns) produced by Saccharopolyspora spinosa.

These are naturally derived fermentation products with a positive safety profile in

contrast to currently used synthetic organically derived compounds (such as synthetic

pyrethroids, organophosphates, organochlorines and carbamates), and have previously

been shown to exhibit insecticidal activity against insects affecting crops and transient

activity against larval blow fly and adult stable fly when administered systemically to

guinea pigs and sheep. By the terms "A83543 compounds" and "spinosyn and

derivatives and analogues thereof is meant components consisting of a 5,6,5-tricyclic

ring system, fused to a 12-membered macrocyclic lactone, a neutral sugar [2N,3N,4N-

(tri-O-methyl)rhamnose] and an amino sugar (forosamine), as described in the patents

discussed infra.

The family of compounds from A83543 fermentation product has been

shown to comprise individual compounds A83543A, A83453B, A83543C, A83453D,

A83543E, A83453F, A83543G, A83453H, A83543 J, A83453L, A83543M,

A83453N, A83543Q, A83453R, A83543S, A83453T, A83453U, A83543V,

A83453W, A83453X. Boeck et al. described spinosyns A-H and J and salts thereof

in US patent Nos 5,362,634, 5,496,932 and 5,571,901. Mynderse et al. described

spinosyns L-N, their N-demethyl derivatives and salts thereof in US patent No,

5,202,242. Turner et al. described spinosyns Q-T, their N-demethyl derivatives and

salts thereof in US patent Nos 5,591,606, 5,631,155 and 5,767,253. Spinosyns

K,O,P,U,V,W, and Y are described in the article by DeAmicis, CN. et al. in

American Chemical Society's Symposium Series: Phytochemicals for Pest Control (1997), Chapter 11 "Physical and Biological Properties of Spinosyns: Novel

Macrolide Pest-Control Agents from Fermentation," pp 146-154.

Spinosyn A (A83543A) was the first spinosyn isolated and identified

from the fermentation broth of Saccharapolyspora spinosa. Subsequent examination

of the fermentation broth revealed that the parent strain oϊ S. spinosa produced a

number of spinosyns (A83543A to H and J). Compared to spinosyn A, spinosyns B

to H and J are characterised by differences in the substitution patterns on the amino

group of the forosamine, at selected sites on the ring system and on the neutral sugar.

The strains of S.spinosa produce a mixture of spinosyns, the primary components of

which are spinosyn A (50-85%) and spinosyn D (15-50%). These are the two

spinosyns that are currently known to be the most active as insecticides. The name

"spinosad" refers to a mixture of spinosyn A and spinosyn D.

The present invention provides an insecticidal formulation for

controlling an insect infestation in a small ruminant animal, said formulation

comprising an effective amount of a spinosyn, or an analogue or derivative thereof,

together with an acceptable carrier or diluent. The formulation may additionally

comprise another insecticidal agent. In me embodiment, the amount of the spinosyn

present in the formulation is such that the spinosyn residue in any wool, milk or tissue

obtained from the animal immediately, or within about a month, following application

of the formulation is at an environmentally acceptable level or safe to humans.

In these formulations, an effective amount of the spinosyn is present

when the amount of spinosyn in the formulation is from 1-500 g of the spinosyn /L of

the formulation. Preferably, the amount of spinosyn in the formulation is selected

from the group consisting of 1-500, 1-400, 1-350 , 1-300, 1-250, 1-200, 1-150, 1-100, 1-90, 1-80, 1-70, 1-60, 1-50, 1-40, 1-30, , 1-25 or 1-20 g of spinosyn per L of the

formulation. A preferred amount is 25 g/L.

In another aspect, this invention relates to an article of manufacture,

comprising packaging material and a formulation for controlling an insect infestation

in a small ruminant animal contained within said packaging material, wherein said

formulation comprises

a topical unit dose of a formulation of this invention, i.e. one comprising an

effective amount of a spinosyn, or an analogue or derivative thereof, together with an

acceptable carrier or diluent; and

wherein said packaging material comprises a label or package insert with instructions

for topically administering the dose to the animal.

This invention further relates to a use of a formulation of this

invention, i.e. one comprising an effective amount of a spinosyn, or an analogue or

derivative thereof, together with an acceptable carrier or diluent, in the preparation of

a medicament for controlling an insect infestation in a small ruminant animal.

In another aspect, this invention provides the use of a formulation of

this invention, i.e. one comprising an elective amount of a spinosyn, or an analogue

or derivative thereof, together with an acceptable carrier or diluent, for controlling an

ectoparasite infestation in a small ruminant animal selected from a sheep, goat, or

camellid, comprising topically administering the formulation to the animal.

This invention further provides a method of controlling an ectoparasite

infestation in a small ruminant selected from a sheep, goat or camellid, comprising

topically administering a formulation of the invention, i.e., one comprising an

effective amount of a spinosyn or an analogue or derivative thereof, to the ruminant. When carrying out this method, it is preferable to administer a formulation wherein

the amount of spinosyn in the formulation is such that the spinosyn residue in any

wool, milk or tissue obtained from the animal immediately, or within about a month,

following administration of the formulation is at an environmentally acceptable level.

Thus, the present invention provides formulations and methods of

controlling, i.e. preventing, ameliorating or eliminating, ectoparasites, including

insects and arachnids, in small ruminant animals such as sheep, goats and camellids

by applying one or more A83543 compounds in an acceptable carrier or diluent.

Another aspect of this invention is a method for preventing, controlling

and/or eliminating insects including ectoparasites in small ruminant animals by

administering an insecticide to the animal, said insecticide including one or more

spinosyn factors or analogues or derivatives thereof present

The invention also provides a method for controlling an ectoparasitic

infestation, wherein the ectoparasite includes lice, fleas, ked, mites, itch mites, ticks

and blowfly strike, in a small ruminant animal by administering to the animal, a

spinosyn, or an analogue or derivative thereof, at a concentration of about 50 ppm or

less, such that any wool, milk or tissue ■ .^jsidue from the spinosyn in the animal is

present in an environmentally acceptable amount immediately or within about a

month following administration. In sheep, for example, an equivalent effect to a

concentration of about 50 ppm of spinosyn is achieved by administering an amount of

approximately 250 mg/head.

The invention further provides a method for controlling an

ectoparasitic infestation, wherein the ectoparasite includes lice, ked and blowfly

strike, in short wool sheep by administering to the sheep, a spinosyn, or an analogue or derivative thereof at a concentration of from 1 to about 25 ppm, such that any wool,

milk or tissue residue from the spinosyn in the sheep is present in an environmentally

acceptable amount immediately or within about a month following administration.

The effect of a concentration of about 25 ppm of spinosyn can be

achieved by administering about 125 mg/head to the sheep.

Other useful spinosyn concentrations for administering to short wool

sheep are about 10, 3, and 1 ppm of spinosyn in the formulation.

In another aspect the invention provides a method for controlling an

ectoparasitic infestation, including lice, ked and blowfly strike, in long wool sheep by

administering to the sheep, a spinosyn, or an analogue or derivative thereof at a

concentration of about 50 ppm or less, such that any wool, milk or tissue residue from

the spinosyn in the sheep is present in an environmentally acceptable amount

immediately or within about a month following administration.

Other useful spinosyn concentrations for administering to long wool

sheep are about 25, 20, or 10 ppm.

Still another aspect of this invention provides a method for controlling

an ectoparasitic infestation, including li e, ked and blowfly strike, in long wool sheep

by administering to the sheep, a formulation wherein the spinosyn, or analogue or

derivative thereof, is present at a concentration of about 50 ppm or less, such that any

wool, milk or tissue residue from the formulation in the sheep is present in an

environmentally acceptable amount immediately or within about a month following

administration of the formulation.

Particularly useful methods for administering the formulations of this

invention in short and long wool sheep are using a dip wash or jetting fluid. Another aspect of this invention provides a method for controlling an

ectoparasitic infestation, including lice, ked and blowfly strike, in long wool sheep by

administering to the sheep in the last 6 months of the growing season, a spinosyn or

an analogue or derivative thereof, at a concentration of about 50 ppm or less , such

that any wool, milk or tissue residue from the spinosyn in the sheep is present in an

environmentally acceptable amount.

For the purposes of the present application, the term 'insecticide' is

defined to include acaracide, ectoparasiticide and miticides. Similarly, for the

purposes of the present application, the term 'insect' is defined to include, but is not

limited to mosquitoes, lice, fleas, flies, ticks and mites. The term 'ectoparasite' is

similarly defined to include, but is not limited to, members of the insect order Diptera,

Phthiraptera, and Acarina, and parasites and other insects which are parasitic during

all of their life cycle or only part of their life cycle, such as only the larval or adult

stage.

The term, 'small ruminant' or 'small ruminant animal' refers to sheep,

goats or camellids (including alpacas, llamas and vacuna, etc.).

The term 'spinosyn or an. logue or derivative thereof is defined to

include an individual spinosyn factor (A83543A-H, J-W or Y) an N-demethyl

derivative of an individual spinosyn factor, or salt thereof, or a combination thereof.

As stated above, the term "A83543 compound" is also used herein to mean an

individual spinosyn factor, or a derivative or salt thereof, or a combination thereof.

The term "controlling an insect infestation" refers to preventing onset

of an infestation of insects in a susceptible animal or to decreasing or eliminating the number of living insects or viable insect eggs on the animal. The extent of reduction

somewhat depends on the application rate of the compound and the compound used.

The term 'effective amount' means the amount which is sufficient to

prevent or cause a measurable reduction in the insect population after treatment .

The term 'short wool sheep' means sheep with 6 weeks or less growth

of wool since the last date of shearing.

The term 'long wool sheep' means sheep with more than 6 weeks

growth of wool since the last date of shearing.

The term 'withholding period' means the period after treatment during

which the wool/meat/milk of animals treated with a insecticide cannot be harvested.

The withholding period of course differs in respect of the chemical nature of the

active in the insecticide.

The term 'environmentally acceptable amount/level' refers to the

amount of residue of an insecticide that is permissible in the wool, tissue and milk of

treated small ruminant animals such as sheep, goats, and camellids. According to the

Australian MRLs (Maximum Residue Levels) for spinosad, published in the

Commonwealth of Australia Gazette (N tional Registration Authority for Agricultural

and Veterinary Chemicals) or "NRA" January 1999 edn, edible offal (mammalian)

has a MRL of 0.05 ppm (= 50 ng residue/g offal). Meat (mammalian in the fat) has an

MRL of 0.2 ppm (=200 ng residue/g meat). In the U.S., the equivalent limits are:

mammalian in the fat: 600 ng residue/g meat; edible offal: 200 ng residue/g offal and

whole muscle: and 40 ng residue/g muscle.

In respect of wool residues, a recent report issued in November 1998

by the NRA and The Woolmark Company entitled 'The residue implications of sheep ectoparasiticides' addresses the 'environmentally acceptable amounts' of residues in

sheep meat and wool in great detail. The current Australian guidelines state that the

residue levels of insecticide on wool must not exceed 13 mg/kg raw wool and the

residue levels in scouring effluent must not exceed 1 mg/L scouring effluent. The

allowable amounts depend on the aquatic toxicity profile of the chemical. For the

UK, synthetic pyrethroids (SPs), must be < 0.06 mg/kg.

From the environmental toxicology the average allowable scouring lot

concentration of spinosad is estimated to be approximately 15 mg/kg or higher.

An advantage of this invention is that the critical environmental

residue limit for harvested wool and the relevant MRL in respect of meat and milk is

met with a zero (0) withholding period in respect of meat/milk/tissue from sheep,

goats and camellids treated with the formulations and methods of the present

invention.

In other words, there is a zero withholding period in respect of

meat/milk from small ruminant animals such as sheep, goats and camellids treated

,vith the formulations and methods of tr ^~- present invention, as well as a zero

withholding period in respect of wool harvested from these animals treated with the

formulations and methods of the present invention.

This invention is based on the surprising discovery in sheep that the

spinosyns, i.e. a spinosyn factor or an analogue and derivative thereof, exhibit

insecticidal activity at very low concentrations, much lower than those used or

previously contemplated in sheep. It has been discovered that at even at 1 ppm,

spinosyn compounds have 100% efficacy as insecticides, particularly ectoparasiticides. Spinosyn compounds are usually in the form of either an emulsion

or a suspension, but at such low concentrations they are advantageously in a solution

form. As the spinosyn formulations of the invention are highly effective, they can be

manufactured and supplied as low volume products which are easier to supply, take

up less storage space and are easier to deliver to the farmer. The smaller quantities

needed also means that the formulations can be supplied as dilute ready-to-use

products in smaller packages which are easily disposed of.

Most importantly, these formulations and methods can be applied even

late in the growing season (i.e., in the last 6 months). Due to the low concentrations

at which the formulations are being used, they are practically non-toxic to humans and

animals. Further, residues in the wool and tissue of animals treated with the

formulations are reduced to environmentally acceptable levels. Minimal chemical

residues are found in the milk of such animals treated with the formulations and

methods of the present invention, which is of economic advantage to the farmer as

processing costs are reduced. No skin irritation or dermal toxicity to shearers results

from the methods and formulations of this invention. Environmental contamination is

also minimised, with waste dip wash, w ste jetting fluid and other run off from treated

animals having minimal levels of chemicals and therefore representing only low

environmental contamination.

Another advantage is that the spinosyns are very efficacious at low

levels and have no apparent cross resistance to existing compounds. Thus, the present

invention is of utility against parasite populations on sheep or small ruminants that

have existing levels of resistance to currently used products. As discussed supra, the formulations of the present invention can be

used in controlling pests on small ruminant animals such as sheep, goats and

camellids (including alpacas, llamas and vacuna etc.). Such pests include

ectoparasites such as blowfly strike, lice, ked, mites, itch mite, sheep scab, screw

worm, bot flies, ticks, fleas, and related arthropod pests. The spinosyns can be

formulated into a variety of end use formulations and products which are easy to use.

In summary, food safety, operator safety and minimisation of adverse

environmental effects are advantages resulting from the present invention. Thus, the

present formulations provide advantages over commercially available ectoparasitic

formulations for use in sheep and small ruminants.

Formulations containing the spinosyns are generally administered

topically to the relevant animal. Such topical application can take the form of

dipping, showering, jetting, spraying, manually applying such as dusting, or otherwise

placing or laying the formulation containing the active substance/s on the animal.

Accordingly, usually the spinosyn factor/s and analogues and derivatives thereof are

formulated into a number of topically applied insecticidal formulations.

Topical insecticidal formulations include spot-ons, pour ons, sprays,

dips, dusts, lotions, gels, ointments, salves, dressings, towels, cremes, sticks, soaps,

shampoos, collars, medallions, eartags and tail bands. Pour-on formulations,

including both aqueous and organic-solvent-based ones, as well as emulsions and

suspensions, are preferred. More preferred are dip wash formulations, jetting fluid

formulations and jetting/spray race formulations. The active component-the spinosyn factor/compound- may be present

as a single compound, a mixture of two or more compounds, a mixture including at

least one of A83543A and A83543D, or a mixture of at least one A83543 compound

together with the dried portion of the fermentation medium in which it is produced.

One preferred active component used in the method of the present invention is

spinosad which is a product comprised of 50-85% spinosyn A and 50-15% spinosyn

D.

The active component-the spinosyn factor/compound- may also be

present as a salt in the formulations and methods of this invention. The salts would be

prepared using standard procedures for salt preparation. For example, spinosyn A can

be neutralised with an appropriate acid to form an acid additional salt. The acid

addition salts of spinosyns which can be used in the present invention are useful and

include salts formed by reaction with either an organic or inorganic acid such as, for

example, sulfuric, hydrochloric, phosphoric, acetic, succinic, citric, lactic, maleic,

fumaric, cholic, pamoic, mucic, glutamic, camphoric, glutaric, glycolic, phthalic,

tartaric, formic, lauric, stearic, salicyclic, methanesulfonic, benzenesulfonic, sorbic,

picric, benzoic, cinnamic and other like xids.

The spinosyn factor/s or analogues or derivatives thereof can be mixed

with one or more physiologically acceptable carriers or excipients to formulate the

insecticide formulations of the present invention. The spinosyn factor/s or analogues

or derivatives thereof can also be mixed with other commonly used ectoparasiticidal

compounds in the insecticide formulations used in the present invention.

The formulations of the present invention can take the form of

concentrated formulations which are dispersed in water for application, such as pour on formulations, or dust or granular formulations which are applied without further

treatment.

Preferably, the formulations of the active spinosyn compound are in

the form of aqueous suspensions or emulsions prepared from concentrated

formulations of the compounds. Such water soluble, water-suspendible or

emulsifiable formulations are generally liquids, generally called emulsifiable

concentrates or aqueous suspensions.

It is particularly preferable that the active formulations are in the form

of a suspension concentrate. Emulsifiable concentrates and solution concentrates, as

well as wettable powders, are also preferred.

Generally, emulsifiable concentrates of the A83543 compounds

comprise a convenient concentration of an A83543 compound dissolved in an inert

carrier which is either a water-miscible solvent or a mixture of a water immiscible

organic solvent and emulsifiers. A preferred concentration range is 1-500 g/L of

spinosyn compound. More preferably the concentration range is selected from the

group consisting of 1-400 g/L, 1-350 g/L, 1-300 g/L, 1-250 g/L, 1-200 g/L, 1-150 g/L,

1-100 g/L, 1-90 g/L, 1-80 g/L, 1-70 g/L. 1 -60 g/L, 1-50 g/L, 1-40 g/L, 1-30 g/L, 1-20

g/L, even more preferably 25 g/L. Useful organic solvents include aromatics

including xylenes and petroleum fractions. Other organic solvents may also be used,

such as the terpenic solvents, including rosin derivatives, aliphatic ketones such as

cyclohexanone and complex alcohols such as 2-ethoxyethanol.

Suspension concentrates (SC) comprise suspensions of the active

water-insoluble compound (including one or more spinosyn compounds) dispersed in

an aqueous vehicle at a concentration in the range of from about 1-500 g/L. Preferably, the concentration range is selected from the group consisting of about 1-

400 g/L, about 1-300 g/L, about 1-250 g/L, about 1-200 g/L about 1-150 g/L, about 1-

100 g/L, about 1-50 g/L, about 1-45 g/L, about 1-40 g/L, about 1-30 g/L, more

preferably about 25 g/L. Generally the suspensions are prepared by finely grinding

the spinosyn compound and mixing it into a vehicle comprised of water and

surfactants chosen from such types as nonionic, sulfonated lignins and alkylsulfates

and suspending agents such as xanthan and guar gums. Other inert ingredients may

also be added.

The suspension concentrates and emulsions are preferably diluted with

water to obtain the desired spinosad concentration in the final formulation which is

applied in the methods of the invention. The active A83543 compounds can also be

applied as insecticides to small ruminant animal species in the form of an aerosol

composition. In such compositions, the active spinosyn compound is dissolved in an

inert carrier, which is a propellant mixture.

Preferably, the carriers or excipients used in the /insecticidal

formulations of the invention include dust carriers, solvents, emulsifϊers, wetting and

dispersing agents and water. Selection < "the carrier is of course made on the basis of

compatibility with the desired insecticide, including such considerations as pH,

moisture content and stability.

In one preferred formulation, the insecticide takes the form of an

emulsion comprising a solution of the active spinosyn compound/s in an organic

solvent, with the optional addition of a surfactant. Water emulsion sprays from such

an emulsion formulation are commonly used. Other preferred formulations of the present invention can take the form

of solids, e.g., wettable powders, dusts or granular compositions. These can then be

compacted to form water dispersible granules. Wettable powders can be used as spray

applications. Other possible formulations include granules or pellets where the

carrier, such as an absorbtive clay, is impregnated with the active.

Advantageously, the method of the invention can be applied as an early

season treatment, or as a late season treatment within the last 6 months of the growing

season when the wool is long. The active spinosyn compound or compounds are

generally present in a concentration of about 50 ppm or less upon application to sheep

or other small ruminants. Other useful concentrations of the active spinosyn

compound or compounds for application to sheep and other small ruminants are 35

ppm, 25 ppm, 20 ppm, 10 ppm, 5 ppm, 4 ppm, 3 ppm, 2 ppm, or 1 ppm.

Generally, the rate, timing and manner of effective application will

vary with the identity of the parasite and other factors. In general, ectoparasite control

is obtained with topical application of liquid formulations containing from about 1 to

10 ppm of spinosyn compound or compounds in respect of short wool animals and 10

_Lo 50 ppm in respect of long wool anim. !s. Conventional veterinary practices are

used in application of the active compound to the animal. Such practices include

spray, back rubbers, and dip tanks.

A preferred composition for use in the present methods of treatment is

a suspension concentrate containing spinosad at a concentration of 25 g/L. If desired,

a dispersant may be added to improve suspendability.

A preferred formulation contains the following ingredients: Ingredient Percent (w/w) Sample batch (2)

Spinosad 2.76 41.4

Propylene glycol 10 150

Surfactant 2 30

Mineral Thickener 2 30

Xanthan Gum 0.2 3

Antimicrobial Agent 0.2 3

Antifoam Agent 0.1 1.5

Water, deionised 82.74 1241.1

Total 100 1500

In one embodiment of an aqueous suspension, the spinosyn is wet-

milled as a "grind batch," i.e., spinosad, surfactant, water, and antifoam as needed. A

typical "grind batch" contains the following ingredients:

(g)

Spinosad 41.4

Surfactant 30

Water, deionised 100

Antifoam Agent 1

Total 172.9

A preferred nonionic surfactant to incorporate into the aqueous

suspension of spinosad is PLURONIC P-123™.

A "hydrated suspension batch" is also formed by blending a hydrated

suspension of the mineral thickener wi '- xanthan gum hydrated in propylene glycol,

along with an antimicrobial agent A typical hydrated suspension batch contains the

following ingredients:

(g)

Propylene glycol 150 Mineral Thickener 30

Xanthan Gum 3 Antimicrobial Agent 3 Water, deionised 1041.1 Total 1227.1 The hydrated suspension batch and additional water as needed are

blended with the grind batch to prevent syneresis, or separation of clear watery fluid

from suspended milled solids. The appropriate amount of the hydrated suspension

batch to be blended with the grind batch to complete the formulation is determined

based upon the percent recovery of the grind batch after particle size reduction.

A particularly useful mineral thickener is colloidal magnesium

aluminum silicate, such as VEEGUM, that disperses and wells in water. A suitable

xanthan gum in one that is heat stable with a good tolerance for strongly acidic and

basic solutions such as RHODOPOL 23™.

Other formulations can also be used in the present methods of

application of ectoparasiticidal formulations to sheep and small ruminants. For

example, emulsifiable concentrates containing spinosad can be made and used for

dilution and subsequent application to sheep. For example, a 25 g/L spinosad

emulsifiable concentrate can be prepared in an aromatic hydrocadrbon such as

Aromatic 150 containing the following ingredients: spinosad (90% active ingredient,

3.05 weight percent), sulfonate/nonionic surfactant blends (a total of 10 weight

percent and aromatic hydrocarbon (86.9 ^" weight percent). As an alternative, spinosad

can be formulated as an emulsifiable concentrate in methyl oleate, potentially safer

solvent, than the aromatic hydrocarbon.

The methods of the present invention, particularly the methods of

treating ectoparasites in sheep can be performed in the following manner:

1) To eradicate lice, sheep should be throughly wetted to the skin, 2 to

6 weeks after shearing, with a solution containing 5 to 20 ppm spinosad. A plunge or shower dip charged at about 5 to 20 ppm and reinforced at about 20 ppm is

recommended.

2) To control lice in sheep with long wool (> 6 weeks after shearing),

sheep should be hand jetted with about 25 ppm spinosad; 0.5 L should be used per

month of wool per sheep. Plunge and shower dips and spray races may also be used.

3) To treat fly stike, the wound should be thoroughly wetted with

about 25 ppm spinosad - applied as a wound dressing or using a jetting hand piece.

4) To prevent fly stike for around 4 (to 6) weeks, 25-50 ppm spinosad

should be applied along the back, around the breech, around the pizzle of wethers and

on the heads of rams; 0.5 L should be used per month of wool per sheep.

The methods as described above can also be used to control sucking

lice, sheep ked and mites.

Examples

Example 1 : Spinosad Dip Formulations for the Control of Bovicola Ovis on Sheep

Spinosad can be formulated as a suspension concentrate. Three

formulations of spinsoad were made into two emulsifiable concentrates, one utilising

an aromatic hydrocarbon solvent and one utilising oleate, and an aqueous

suspension/suspension concentrate. Each formulation was prepared at two

concentrations, namely containing 0.2 and 1 g/L spinosad. These were each diluted

1 :5000 in water to give 0.04 and 0.2 ppm dip washes.

Six fine-wool Merino sheep infested with a strain of lice highly

resistant to synthetic pyrethroids were plunge dipped in each dip two weeks after

shearing. Lice counts were conducted on the day of treatment before dipping and

7,14, 28, 42 and 56 days after treatment. The numbers of lice on sheep treated with

water only (i.e. 0 ppm) increased slightly during the study.

The results with regards to lice numbers on sheep treated with the

various formulations of spinosad in this example, are set out below in Table 1.

Subsections (a) - (g) provide the lice counts observed with the control animals and

with each concentration of the three listed spinosad-containing formulations.

Table 1. Lice Counts

a) Control

b) Emulsifiable Concentrate in Aromatic Hydrocarbon at 0.04 ppm

c) Emulsifiable Concentrate in Aromatic Hydrocarbon at 0.2 ppm

d) Emulsifiable Concentrate in Methyl Oleate at 0.04 ppm

e) Emulsifiable Concentrate in Methyl Oleate at 0.2 ppm

f) Aqueous Suspension at 0.04 ppm

g) Aqueous Suspension at 0.2 ppm

Example 2a: Dose determination for the treatment of sheep body lice

(Bovicola ovis)

i) Short wool sheep, application < six (6) weeks after shearing in the form of a dip

Two dose titration studies were conducted in respect of spinosad

applied as a dip for the control of Bovicola ovis on sheep. Spinosad, prepared as an

emulsifiable concentrate, was diluted 1 :1000 in water to obtain formulations of 0, 1.0,

5.0, 20 and 100 ppm concentration and formulations of 0, 0.04, 0.2 and 1.0 ppm

concentration. Fine, short-wool Merino sheep infested with a strain of lice highly

resistant to synthetic pyrethroids were plunge dipped in the dilute solution two (2)

weeks after shearing. During the first titration study, 5 dips were made, with spinosad

being applied at 0, 1.0, 5.0, 20 and 100 ppm to 6 sheep per each concentration by way

of plunge dipping each group of six sheep. During the second titration study,

spinosad was applied at four different concentrations: 0, 0.04, 0.2 and 1.0 ppm, again

each concentration was applied to six (6) sheep by plunge dipping. Lice counts were conducted on the day of treatment before dipping and

7, 14, 28, 42, 56 and 84 days after treatment. The number of lice on sheep treated

with vehicle and water only (0 ppm) declined following dipping although remained at

a satisfactory level for the duration of the trials. In the first titration study, a dose

response was not observed as all spinosad treatments were 100% effective and no live

lice were found after treatment.

In the second titration study, a dose response was observed. An

average of 1.5 lice per sheep was seen 1 week after dipping in 1.0 ppm but on days

14, 28, 42, 59 and 84, 0-0.3 lice/sheep were seen. The 0.2 ppm formulation resulted

in 73-94%o efficacy and the 0.04 ppm dip gave 38 to 60% efficacy, depending on the

day of counting. Based on the second study, the estimated concentration giving 100%

efficacy was 2.0 ppm.

ii) Long wool sheep, application in the form of a jetting spray. 6 to 52 weeks after shearing

Thirty fine-wool Merino ewes with 6 months wool (5 cm long) were

ranked and block allocated according to lice count into 5 groups of 6 sheep. Spinosad

suspension concentrate (25 g/L) was diluted in water to give 0, 2, 5, 15 and 45 ppm

dilutions. 5 L of each diluted concentrate was jetted onto six (6) sheep according to

standard agricultural practice. The numbers of adult and nymphal lice at each of 40

fleece partings on each sheep was determined the day before jetting and 14, 28, 56

and 84 days after jetting. The control sheep (i.e., those treated with 0 ppm) were

jetted with 350 ppm temephos 70 days into the study to relieve rubbing and skin

irritation.

Spinosad applied at all rates provided >95% efficacy against sheep

body lice within 14 days of treatment. At 56 days post treatment, spinosad at the same rate of 45 ppm provided 100% efficacy and the 2.0 ppm rate gave 98.4%

efficacy. The level of efficacy given by 2.0 ppm spinosad 14 days post treatment was

greater than that provided by 350 ppm temephos 14 days post treatment.

Example 2b: Evaluation of spinosad for the treatment of blowfly strike on sheep caused by Lucilia cuprina

i) Fine wool sheep, application in the form of a wound dressing

Fifteen fine wool Merino sheep with approximately 6 months wool

growth were implanted at six (6) sites with approximately 100 first instar of Lucilia

cuprina. Forty eight hours after implantation, the sites were assessed and the viable

sites treated. Emulsifiable concentrates containing 0, 2, 5, 15, 45 and 135 g/L

spinosad were diluted 1 : 1000 in water to give emulsions of 2, 5, 15, 45 and 135 ppm.

Each treatment was applied to 3 sheep (18 sites) in 20, 40 or 60-mL volumes-2

volumes per sheep per concentration. Each site was examined for larval survival and

development 24 hours after treatment.

As concentration increased, the treatments became progressively more

lethal. For quick 100% efficacy, 80 ppm or more was required. Nonetheless, 15 and

20 ppm gave 100% efficacy at 83% of sites after 24 hours. All concentrations,

however, had an adverse effect on larval survival and no strike in any treated site

developed normally. Indeed, the surviving larvae seen in all the treated strikes in the

first study would probably have died in the next 24 hours as was seen in the second

study where all treatments gave 100% mortality at all sites 48 hours after application.

Varying the volume of treatment applied from 20 to 60 mL had no effect on efficacy.

iO Long wool sheep, application in the form of a jetting fluid Ten cross bred Merino/Suffolk wethers with approximately 12 months

wool growth and 2 Merino wethers with 6 months wool growth were implanted at 6

sites with approximately 100 first instar oϊ Lucilia cuprina. Seventy two (72) hours

after implantation, the sites were assessed and the sheep were treated. Suspension

concentrate containing 25 g/L spinosad was diluted 1 : 1000 and 1 :500 in water to give

suspensions of 25 and 50 ppm. A 25 ppm suspension was also prepared with wetting

agent added at the rate of 1 :1000. Each treatment was applied to 4 sheep (24 sites)

using a commercial jetting wand. Each site was examined for larval survival and

development 1, 2.5-3 and 5-6 hours after treatment.

Within one hour of treatment at most sites, there was movement of

larvae away from the site of implantation, up the wool staple. After 2.5 - 3 hours

there were no normal larvae (except 1 site that jetting fluid had not reached) and many

abnormal larvae. By 5-6 hours after treatment, most larvae were dead or missing

from the site of implantation. Increasing concentration from 25 to 50 ppm did not

improve efficacy and neither did adding surfactant.

iii) Prevention, eval. ation of spinosad for the prevention of blowfly strike on sheep caused by Lucilia cuprina

Spinosad emulsifiable concentrates were diluted 1 :1000 in water to

give 0, 2, 5, 15 and 45 ppm emulsions and applied as jetting fluids to the backs of

each of 3 sheep. Fifteen fine wool merino sheep with approximately 6 months wool

growth were used. 3, 8, 15, 22, 29 and 36 days after treatment each sheep was

implanted at two sites on the back with approximately 100 first instar Lucilia cuprina

maggots. Each site was examined for maggot survival and development 24, 48 and 72

hours after implantation. The degree of maggot killing or inhibition of maggot development was related to concentration and time since treatment. 2 ppm gave

100%> mortality of applied maggots at 3 or more of 6 sites for less than 2 weeks.

Increasing the concentration to 5 ppm extended maggot death at 3 of the 6 sites to 4

weeks. The 15 ppm results were variable, but only lasted 4 weeks. 45 ppm gave

complete mortality of maggots at 4 of 6 or 5 of 6 sites for 4 weeks and at 3 of 6 sites

at 5 weeks.

Example 2c: Evaluation of spinosad for the treatment of sheep ked (M. ovinus)

i) Three (3) shorn sheep and 3 unshorn sheep sprayed with 1 L

each of 100 ppm spinosad.

Shorn sheep - 100% keds dead by 1 day after treatment

Unshorn sheep- some live ked seen 1, 3 and 7 days after

treatment. All were dead by 14 days after treatment.

ii) Six (6) recently shorn sheep infested with ked were divided in 3

groups of 2 sheep. The sheep in group 1 were sprayed with 1 L

containing 25 ppm spinosad; those in group 2 with 50 ppm

spinosad, and the group 3 sheep were treated with vehicle only

(controls). Sprays containing 25 and 50 ppm spinosad killed all

keds within 1 day. Counts on control sheep declined after

shearing but much less rapidly than treated sheep.

Example 3a: Dose confirmation and protection period for the treatment of sheep body lice (Bovicola ovis) i) Short wool sheep, application < six (6) weeks after shearing in the form of a dip

Thirty-five lice-free Merino sheep were plunge dipped in an aqueous

suspension containing 5 ppm spinosad. 35 similar lice-free sheep were used as

controls. 1, 2, 3, 4 and 6 weeks after treatment, five (5) treated and 5 untreated

(control) sheep were implanted with 50 viable lice on the shoulder and hip. The

implantation sites were examined 4 weeks later and again another 4 weeks later if no

live lice were found at the first examination.

Lice placed on treated sheep 1 week after treatment failed to survive.

The lice placed on the untreated sheep survived. Lice implanted 2 weeks after

treatment survived in low numbers on 2 of 5 treated sheep and on all of the 5 control

sheep. Numbers of surviving lice increased with time between dipping and times of

subsequent examination. By five (5) weeks after treatment, lice numbers on treated

sheep were approaching those seen on untreated control sheep.

Example 4: Decay profile of spinosad in sheep wool following dipping

Spinosad suspension at 5 g/L and 20 g/L was diluted 1:1000 with

water. The concentration of the two dip washes were therefore 5 ppm and 20 ppm,

respectively. Nine Merino ewes with 2 weeks wool and having had no ectoparasite

treatment for 12 months were used and divided into 3 groups of 3 sheep. Each group

was housed in an outdoor pen, which was raised with a mesh floor. Each pen had a

roof over 1/3 of the pen and had metal walls between each pen.

Two sheep dip washes were prepared in a calibrated stainless steel

bath. 475 mL of dam water was added to the bath, 475 mL of test article was then

added to the bath and mixed thoroughly and a 40-mL sample of the dip was collected. 6 of the sheep were individually dipped for 30 seconds with the head submerged twice

in the 5 ppm dip wash, while 3 sheep were similarly individually treated in the 20

ppm dip wash.

Following dipping, circumferential wool samples were collected using

electric hair clippers with a cutting width of 4.5 cm from all sheep 3 days prior to

treatment and on a range of days post treatment. Three of the sheep treated with 5

ppm spinosad were sampled on days 1, 2, 4, 7, 9, 11, 14, 17 and 21 post treatment,

while the remaining three sheep were sampled on days 7, 14, 21, 28, 35, 42, 49, 56

and 64. The three sheep treated with 20 ppm spinosad were sampled on days 7,14,

21, 28, 42, 56, 70, 86 and 98. The wool samples were then analysed by

immunoassay.

Following dipping in 5 ppm spinosad, there was a maximum residue of

3.3 ppm which declined with a half life of about 2 weeks. After 64 days, the residue

was 0.27 ppm which was 8% of the initial residue. Following dipping in 20 ppm, a

residue of 16.4 ppm was seen, which took 5 weeks to decline to 8 ppm, but after that

the half life was about 2 weeks. After 98 days, the residue was 1.36 ppm, which was

?>% of the initial residue.

Example 5: Spinosad residues in sheep meat, fat and offal four days following dipping in 50 ppm spinosad dip formulation

Two previously sheared sheep were dipped in an aqueous suspension dip formulation containing 50 ppm spinosad for 30 seconds, with the head being dunked twice, and then placed in a floor pen for four days. The sheep were sacrificed and necropsied at the end of those four days; and samples of kidney, liver, muscle, perirenal and abdominal fat were collected. The tissues were transferred to a -20°C freezer until processing. The tissue samples were chopped, frozen with dry ice and then ground using a hammermill. The samples were then placed in a -20°C freezer until assay. Samples were assayed using Elanco Method B05873 (EPA Method MRID 44058822, Dow Agro Science Method GRM 95.03) "Determination of spinosad and metabolites in Beef tissues, milk and cream by HPLC with UV detection" which is certified for use with sheep tissues. The samples were assayed in duplicate with the results shown below in Table 2. The residue definition of spinosad is the sum of spinosyn A and spinosyn D. However, all residues are spinosyn A, as residues of spinosyn D, N-demethyl spinosyn D and spinosyn B were less than the limit of quantitation of the certified method.

Table 2: Total Spinosad Residue (ng/g) - Day 4 after 50 ppm dipping

Animal A Animal B Average

Abdominal fat 150.0 177.5 163.8 Perirenal fat 99.9 142.7 121.3 Liver 34.2 38.9 36.6 Kidney 23.3 28.3 25.8 Muscle 14.9 13.5 14.2

Example 6: Measurement of the residue decay profile of spinosad in sheep wool following dipping in 10 ppm spinosad and jetting with 25 ppm spinosad.

Table 3. Mean concentration (mg/kg) and total mass (μg) of spinosad on wool band samples collected after sheep were dipped in 10 ppm spinosad suspension in water at days 1 through to 170 after treatment.

Merino sheep with 6 weeks wool growth were dipped in an aqueous suspension containing 10 ppm spinosad. Circumferential strips of wool were collected from each sheep after treatment to allow the estimation of spinosad residues and residue decay profiles. Wool residues decayed with an average degradation rate (half- life) of 26 days. Table 4. Mean concentration (mg/kg) and total mass (μg) of spinosad on wool band samples collected after sheep were jetted with an aqueous suspension containing 25 ppm spinosad at days 1 through to 84 after treatment.

Spinosad decayed with a half-life of 30 days in sheep with 9 months wool growth jetted with 25 ppm spinosad.

On the basis of the toxicity of spinosad to aquatic organisms, a provisional environmental assessment of discharges from wool scouring has established target concentrations for wool processing lots in Australia and in Europe of 66 mg/kg and 15 mg/kg respectively. (I M Russell, N T Campbell, J T Rothwell, Proc Aust Sheep Vet Soc 2000, pp 108-115, ANA Conf Perth 2000). The wool residues seen immediately following treatment are well below the projected maximum permissible residues for scouring lots in Europe which has the most stringent Environmental Quality Standards. Even following late season jetting treatment for blowfly prevention, sheep will be left at least a month before shearing and following other treatments for up to 12 months after treatment allowing wool residues to decay. In addition the wool in a processing lot is made up of wools treated in early season, wools treated in late season, untreated wools, and wools treated with other chemicals. This blending will allow the spinosad residue from any farm to be diluted by these other wools. Therefore treatment of sheep with spinosad at the recommended rates will not cause wool residues to breach any wool scour environmental parameters and will not require the establishment of a wool with-holding period following treatment.

Spinosad has a high margin of safety for farm workers due to the short half-life of spinosad in wool grease, the very low level of dermal absorption and low mammalian toxicity of the active ingredient. Sheep treated with spinosad present no risk to shearers, wool handlers or other farm workers.

Example 7: Tissue residues following dipping in 10 ppm spinosad and jetting with 25 ppm spinosad. Thirty seven fine wool Merino ewes and wethers with 6 weeks wool

growth were dipped in 10 ppm spinosad suspension in water. Thirty seven Merinos

with 9 months wool growth were jetted with 25 ppm spinosad. Animals were

slaughtered 5, 12, 15, 21, 39and 56 days after treatment and samples of back fat,

muscle, liver, peri-renal fat and kidney collected and assayed for spinosad residue.

The limit of quantification (LOQ) was 0.01 mg/kg and the limit of detection (LOD)

was 0.003 mg/kg. Residues of spinosad greater than the limit of quantification were

found in fat samples from animals treated via a plunge dip and traces (values between

the LOQ and LOD) were found in fat from jetted animals. Traces of spinosad were

found in the liver and kidney tissues of animals that were plunge dipped, but except

for one liver sample collected 12 days after treatment, not from jetted animals. Offal

residues were below the LOD 15 days after treatment. Residues of spinosad in the

main edible tissue of sheep, meat were undetectable at all times.

The highest residues recorded were in tissues collected 5 days after

dipping and were 3 to 5 times lower than the lowest relevant residue limit set.

Therefore dipping sheep in 10 ppm spinosad or jetting them with 25 ppm spinosad

does not infringe established minimuni residue limits (MRL) or tolerances and no

with-holding period is required. In countries were a MRL is not established and a nil

residue is required a withholding period of 56 days would ensure that the tissue with

the highest residue - fat - would be below the LOD. Likewise offal would be below

the LOD 15 days after treatment and meat residues would be below the LOD at all

times after treatment.

Tables 5. Mean spinosad residues (mg/kg) in fat after dipping in 10 ppm spinosad.

Claims

CLAIMS:

1. An insecticidal formulation for controlling an insect infestation

in a small ruminant animal, said formulation comprising an effective amount of a

spinosyn, or an analogue or derivative thereof, together with an acceptable carrier or

diluent.

2. A formulation of Claim 1 wherein the spinosyn is a component

of spinosad.

3. A formulation of Claim 1 or 2 wherein the amount of spinosyn

in the formulation is selec^ted from the group consisting of 1-500, 1-400, 1-350 , 1-

300, 1-250, 1-200, 1-150, 1-100, 1-90, 1-80, 1-70, 1-60, 1-50, 1-40, 1-30, , 1-25 or 1-

20 g of spinosyn per L of the formulation.

4. A formulation of Claim 1, 2 or 3 wherein the ruminant animal

is a sheep, a goat or a camellid.

5. A formulation of Claim 1, 2, 3 or 4 wherein the amount of

spinosyn present in the formulation is such that the spinosyn residue in any wool,

milk or tissue obtained from the animal immediately, or within about a month,

following application of the formulation is at an environmentally acceptable level.

6. A formulation of Claim 1, 2, 3, 4, or 5 which additionally

comprises another insecticidal agent.

7. An article of manufacture, comprising packaging material and a

formulation for controlling an insect infestation in a small ruminant animal contained

within said packaging material, wherein said formulation comprises a topical unit dose of a formulation of Claim 1, 2, 3, 4, 5 or 6; and wherein

said packaging material comprises a label or package insert with instructions for

topically administering the dose to the animal.

8. The use of a formulation of Claim 1, 2, 3, 4, 5 or 6 in the

preparation of a medicament for controlling an insect infestation in a small ruminant

animal.

9. A use of a formulation of any one of Claims 1 to 6 for

controlling an ectoparasite infestation in a small ruminant animal selected from a

sheep, goat, or camellid, comprising topically administering the formulation to the

animal.

10. A method of controlling an ectoparasite infestation in a small

ruminant animal selected from a sheep, goat, or camellid, comprising topically

administering a formulation of any one of Claims 1 to 6 to the animal.

11. A method of Claim 10 wherein the animal is a sheep, and the

sheep is a short wool sheep.

12. A method of Claim 11 wherein the effective amount of the

spinosyn in the formulation is 10 ppm or less.

13. A method of Claim 12 wherein the formulation is administered

in a dip wash or jetting fluid.

14. A method of Claim 13 wherein the effective amount of the

spinosyn in the formulation is 5 ppm or less.

15. A method of Claim 14 wherein the amount is 3 ppm or less.

16. A method of Claim 15 wherein the amount is about 1 ppm.

17. A method of Claim 10 wherein the animal is a sheep, and the

sheep is a long wool sheep.

18. A method of Claim 17 wherein the effective amount of the

spinosyn in the formulation is 50 ppm or less.

19. A method of Claim 18 wherein the formulation is administered

in a dip wash or jetting fluid.

20. A method of Claim 19 wherein the effective amount of the

spinosyn in the formulation is 25 ppm or less.

21. A method of Claim 20 wherein the amount is 20 ppm or less.

22. A method of Claim 21 wherein the amount is 10 ppm or less.

23. A method of Claim 17 wherein the formulation is administered

within the last six months of the season.

24. A method of Claim 10 wherein the ruminant is a goat.

25. A method of Claim 10 wherein the ruminant is a camellid.

26. An insecticidal formulation for controlling an insect infestation

in a small ruminant animal, said formulation comprising a spinosyn, or an analogue or

derivative thereof, together with an acceptable carrier or diluent, substantially as

hereinbefore described with reference to any one of the Examples.