Introduction
The Australian dingo is endangered due to hybridisation in the wild with domestic dogs. The extent of this problem needs thorough investigation but it is thought to be so severe that there is even a worry that many of the dingoes in captivity may not be purebred. At the University of New South Wales we have developed genetic markers that can be used to differentiate DNA from dingoes and domestic dogs and these can be used to assess the purity of any individual dingo or dingo population. These tests have generated many questions on how they work and what they can do. The following may help answer some of those questions.

What are the tests based on and are they routine?
At this stage the tests are still in development and could be improved. They are not yet routine. The tests are based on DNA extracted from blood, tissue, hides or mouth swabs. The extracted DNA is amplified in the lab using the polymerase chain reaction (PCR) which amplifies the small amounts of DNA. Twenty different sites (or loci) in the dog have been chosen for the test. These were chosen because they show a difference between a set of reference dingoes from captivity and domestic dogs of mixed breed from the pound. The sites tested are called microsatellites, which are the type of loci commonly used in paternity testing in dogs and humans. We have chosen loci that have many different forms found in the dog but which are rarely found or not found at all in the dingoes. So if one or more of these "dog types" are found in dingo, it is most likely to have been inherited from a dog ancestor.

Why test 20 different sites? The more sites that are tested the larger the chance of detecting presence of any dog ancestry. For example, in an animal that is three quarters dingo it will have inherited "dog types" at only about half of the sites. If only one or two were tested they dog ancestry could be missed. By testing many sites we are very unlikely to miss it.

Will the test detect every animal that has dog ancestry no matter how far back?
No, it is impossible to detect every case but the most hybrids should be detected. The more generations since the outcrossing to dog the less dog content in the genes and the harder it is to detect. Dog types will be found at fewer sites. One way to improve the chance of finding dog types when they are rare is to type more sites. However, this makes testing more expensive and requires more research so a compromise has to be reached and 20 sites gives us a good detection power at a reasonable cost.

Can a pure dingo ever show "dog types" in the tests?
This is unlikely but still possible for several reasons. Firstly, it could be a laboratory error and any questionable result should be resampled and checked. Secondly, it is not impossible that types not identified in the initial reference dingoes do rarely occur naturally in dingoes. As we gather more results from dingoes from across the country such anomalies should become evident. For these reasons a confirmed "dog type" in a single test should not be taken as conclusive but it is suspicious. However, more than one measure is used to assess purity and all do not rely on results at a single site but combine the results from all sites.

How are results combined from all sites to give a single number? If a type occurs in both dogs and dingoes is it still useful?
Say a type is common in dogs but also found occasionally in dingoes. When we find a unknown sample that has that type we would say it is more likely to come from a dog than a dingo but could possibly be from either. We can assign a probability to the chance it will have come from one group or the other. We can combine this type of probability information from all types at all sites to give our best estimate of which group the animal comes from, dog, dingo or hybrid. We compare the probability it is likely to be from a pure dingo to the probability it is from a hybrid which is three quarters dingo and one quarter dog. The higher the value the more likely the animal is pure dingo. This score from a group of dingoes in captivity is used as a guide to the value expected for pure dingoes.

Can dingo samples be sent for testing?
We are happy to receive samples from dingoes from around the country but the lack of funds means we have little resources to do testing and the samples will be stored until funds are available. A donation of $50 towards the research for each animal tested could mean we are able to process that samples sooner. Even then, to make the process efficient, the testing has to been done in batches and the analysis must be carefully checked before results can be sent out. This means delays of several months. Why have results taken longer in the past? The reference material has had to be double-checked and the computer software to analyse the results developed. Will the situation improve? Not unless further funding can be found to employ experienced personnel. The current funding will not last to the end of 2001. The funding to develop these tests has been from collaborative SPIRT grant from the Australian Research Council to Dr Wilton at University of New South Wales in partnership with the Native Dog Conservation Society and the Australian Dingo Conservation Association. The dingo conservation groups give time, resources and some funds to the project.

What types of samples are used for testing?
The easiest samples to process are blood samples collected in EDTA tubes and mailed by Express post to the lab. The tests can also be done on tissue samples, which is usually an ear clipping from deceased animals. Mouth scrapings, a sample of cells from the inside of the mouth, make convenient samples for owners of friendly captive animals. These are taken with small nylon brushes (the typed used for pap smear tests) which are available on request. For application to monitoring wild populations methods are being developed to do testing on faecal pellets (droppings). Plans have been drawn up with National Parks and Wildlife Service of NSW to monitor hybridisation in several populations but the work is dependent on acquiring funding.

If the captive dingoes that were used as a reference population are not all pure dingoes how would it affect the results?
If there are some "dog types" contaminating the reference samples it will slightly reduce the ability to detect hybrids because that type will be recognised as dingo as well as dog. It will not affect all of the sites or all types at a single site. Since we are testing 20 sites with many dog types at each site the test would still be effective. The consistency between results from different sites within individual reference animals and between different groups of captive dingoes suggest contamination of the reference material, if present at all, is at a very low level.

Would it be better to use samples from dingoes before contact with Europeans and domestic dogs as a reference group?
Yes it would because then we would be sure that reference dingoes are pure dingoes. However, we have not located a source of such material that we can use. Fossil bones are very difficult to work with and do not give reliable results for this type of testing. Hides from early explorers or even scalps from dingo bounties would be useful if they could be located.

Can DNA be extracted from bones for testing?
It is possible to do testing on bone or teeth but it can be difficult to do. The age and condition of the material both affect whether it will work or not. We hope to improve the tests so that it works more reliably from this type of material.

Can the tests be used to identify dogs that have some dingo ancestry?
We do not do this type of testing. The tests have not been designed to do this. Often types found in dingo are also found in dogs so it would not be as effective. We have not investigated the effectiveness of this type of testing.

What is the future for testing purity of dingoes?
Effective testing with a short turn around time will be available for testing of individual animals in the not too distant future. Methods will be developed for sampling and testing wild populations using non-invasive techniques such as collection of droppings. Collaboration with state government orgaisations, wildlife protection organisations and pest control organisation will allow assessment of the wild populations for conservation value and the information incorporated into sensible wild dog management programs.