This protocol is to assist in the collection and treatment of deceased primates to ensure possibilities for future research using their hard tissues, consisting of the skeletal remains and dentition. It is a step by step guide for how to deal with the remains from the death of an individual to the storage and archiving of the hard tissue. This is followed by additional notes and a skeletal inventory form used during step 10.
1. Notice the absence and possible death of an individual
Daily observation of the mandrill colony allows observers to note the absence of individuals at feeding times. If an individual is not seen for several days in a row, the chances are high this individual has died. Record the absence in the individual’s file. After 3 days of absence, search the enclosure for the missing individual. The smell of decomposition also serves as an indication of a death. Search the enclosure as soon as the smell is detected.
2. Confirm and record the death of an individual
When an individual is found dead in the enclosure, dies on the operating table or dies under other circumstances, record this in the individual’s file with the exact date of death. If a decomposed body is retrieved, note the date, the state of decomposition and estimate the date of death based on the state of decomposition and the date on which the animal was last observed at feeding time.
3. Collect the remains of a recently deceased individual
Collect all remains available for controlled decomposition. If the body is fresh, collect the remains as a whole. If the remains of an individual are already in advanced decomposition and skeletal elements have become detached from one another, pay attention to the collection of loose elements. If the body has fallen apart, collect the material according to skeletal regions. These regions are:
- The cranium, consisting of the skull and lower jaw. Check the area around the cranium for loose teeth.
- The thorax (consisting of the vertebral column, the sternum, shoulder blades, pelvis (hip bones) and sacrum) and the extremities (consisting of the arms and legs).
148 - The hands and feet. Collect material from each side separately to avoid difficulties
with determining the side of the material later. Pay extra attention to checking the area around the hands and feet, since there are many small bones here which could get lost.
The most important thing is not to miss any material. In cases where more than one individual died at the same time, make sure the material is kept separated during collection.
4. Place the remains of the individual in a plastic bag for controlled decompositions.
Puncture the bag to facilitate the decomposition process so that liquids can leave the bag and insects and other animals can enter to remove the soft tissue. In the case of large individuals, place the head, feet and hands in bags for decomposition on the cemetery, and place the rest of the body for decomposition in the forest next to the enclosures for decomposition. Check the remains regularly.
5. Add clear ID labels made of non-degradable material (ear-tags are ideal) to the bag and place an individual in the cemetery for decomposition
Do not just write the ID number on the plastic bag in which the remains are kept for
decomposition, because this will degrade, losing the ID information which is vital for further research. Note the date on which the remains are placed in the cemetery in the individual’s file. When more individuals are placed on the cemetery at the same time, place the bags at least 1m from one another to avoid mixing of material.
6. Check the state of decomposition monthly
During decomposition, the first material to degenerate is the soft tissue such as organs and muscles, followed by tougher material such as hair, cartilage and ligaments. Later, mainly hard tissues are left, consisting of dental elements and the skeleton. Although hard tissue is more resistant to degeneration than the other softer tissues, it also degenerates when the remains of an individual are left for too long. The remains of infants and juveniles are less robust than the remains of adults, so need extra attention when monitoring decomposition, shorter decomposition times and more regular checks.
149
7. Collect the remains from the cemetery
Once all the soft tissue is gone, collect the remains.Check the bag in which the remains were placed carefully for all bone and dental elements, as well as the soil under and surrounding of the bag. Ensure that you collect all dental elements (teeth) after decomposition. Molars
are usually securely fixed in the jaw, even when soft tissues are lost, but single rooted teeth, such as the incisors and the canines become detached from the maxilla and mandibular bones easily when soft tissues are no longer present. It is very important to collect these too.
8. Clean the bone material with water and alcohol. Take all the material out of the bag, gently
remove excess soft tissue, place in a solution of Chloramine T and water (5%) and leave it for 7 days. After that, clean the bones carefully with a soft brush. Do not use aggressive
chemicals such as bleach for cleaning the skull and dentition. Bleach has a negative effect
on the condition of the tooth enamel and removes dental calculus, which is of interest to researchers. After one week in chloramine T, the material can be stored under refrigeration in a 2% Chloramine T solution or dried in a ventilated room and stored until shipped.
9. Ensure that you keep the ID label with the remains at all times and that you do not mix the remains of more than one animal together.
If you retrieve more than one individual from the cemetery at the same time, take care to keep the remains of each individual separate. Use separate buckets for the cleaning process.
10. Label the bone material
Once the material is dry, write the full ID code of the individual on the skull with a permanent marker. Label all available large bones as well. These consist of: the femur, tibia and fibula for the legs; the humerus, ulna and radius for the arms; the left and right sides of the pelvis, the sacrum and the left and right shoulder blades (scapula) for the thorax. Collect the other bones by zone: place the hand bones from the left hand in one bag, labelled Left Hand. Place the hand bones of the right hand in another bag labelled Right Hand. Do the same for the bones of the left and right feet, the left and right side of the ribcage, the vertebral column including the hyoid and the sternum. Label the bones with the whole ID code of the
individual, for example 17D1A. Use permanent marker for all labelling and write the ID code on an area of the bone where it is clearly visible. Avoid joint surfaces and areas of muscle attachment when labelling.
150
11. Fill in the skeletal inventory recording sheet
After the bones are cleaned and labelled, fill in the skeletal inventory form (see attachment). This facilitates future research and helps to keep the material organised. Add the skeletal inventory form to the individual’s file, after the autopsy report.
12. Organised storage
Keep the remains of each individual together during storage. Store material in a dry place and avoid contact with moisture. Write the location of storage on skeletal inventory recording sheet. If material is moved to a different location, note this on the skeletal inventory recording sheet.
13. Note that the bone material was collected and stored after death in the individuals file
Notes
1 Collect and store as many skeletal elements as possible.
The long bones of primates are studied in the context of locomotion and its effect on morphology, age estimation, plasticity, influence of diseases, and so on. The fact that the individuals at CIRMF are closely monitored during life and the possibility of collecting their hard tissues after they die creates a unique combination which allows us to address many research questions if material is carefully collected and curated. Knowledge of the individual’s ID is crucial here (see point 1 above). Collect and store skeletons as
completely as possible.
2 Collect the remains of infant and juvenile individuals.
The hard tissues of infants and juveniles allow the study of growth and development in a way that cannot be achieved by only studying the remains of adults. Although the remains of infants are often more difficult to retrieve from the enclosures due to their small size and quick decomposition in combination with the dense forestry, take advantage of any possibility of collecting infant remains. This includes infants and juveniles who die in the volieres or during veterinary procedures. Infant remains are fragile, therefore take extra care during handling.
151
3 Visual documentation of material
If possible, take photographs of the dead individual to record the initial state of the material. This can range from highly decomposed / completely skeletonised in the enclosure to completely intact. Also record the remains after cleaning. These images should accompany the inventory form. When taking photographs, place a clear label close to the remains stating the ID of the individual and the date the photograph was taken.
This protocol was written on the basis of the knowledge of the authors, as well as the knowledge and experience of Dr Barthelemy Ngoubengoye, Dr Anais Herbert, Ivan-Cyr Moussadji Kinga and
152