Project Title: Investigate development of minimally invasive means to gather demographic information in cetacean: A comprehensive comparative approach
Chief Investigators: A/Professor Mark Hindell and Mr Glenn Dunshea
Telomeres are nucleoprotein structures on the ends of linear chromosomes that have been shown to change with age in some animals. This project aimed to examine telomere dynamics in marine mammals with a view towards assessing the use of telomere change as an age marker for individual animals.
There are various molecular biology methods for assessing telomere length, all of which revolve around detecting the specific repetitive DNA sequence of the telomere (TTAGGGn in vertebrates) and each with different requirements for starting material. It has previously been demonstrated that cetaceans contain interstitial telomere sequence (ITS), which is DNA that has the same sequence as telomeric DNA but is not associated with the end of the chromosome (i.e. the telomere) and hence not bound by telomeric processes. We conducted experiments to assess the effect of this ITS sequence on traditional telomere measurements and found that cetacean ITS sequence can dramatically affect telomere length data gained by traditional telomere measurement protocols (i.e. denaturing terminal restriction fragment length assays) and these should probably be avoided. From these experiments we conclude that the best methodology for measurement of cetacean telomeres is a non-denaturing terminal restriction fragment length protocol, however we found this protocol prohibitive to apply to skin biopsy samples for a number of reasons mostly involving the quantity and quality of DNA that can be extracted from cetacean skin samples.
A quantitative PCR (Q-PCR) technique was employed to look at relative changes in telomere amount within and between individuals of three species, the bottlenose dolphin (Tursiops truncatus), the southern right whale (Eubalaena australis) and the harp seal (Phoca groenlandica). While this technique is also effected by ITS and bound by certain assumptions, it can demonstrate relative changes. In bottlenose dolphins we found no significant change in telomere sequence amount between individuals of different ages, however there were significant changes within individuals at different stages of development but these were not in a consistent direction. We found no relationship with telomere length and age in harp seals using both Q-PCR and terminal restriction fragment assays. In southern right whales we examined relative telomere sequence amounts between mothers and their calves and found that while there were generally significant differences between individuals in a cow-calf pair, the direction and magnitude of these differences were not consistent within and between cow-calf pairs and overall there were no differences between calves and cows.
This work has demonstrated that telomere correlates are likely not a good marker for age of individuals and demographic processes in populations of cetaceans and some pinnipeds. This is partly due to methodological constraints but our data also indicate considerable variation of telomere lengths and telomere dynamic processes within and between individuals. Within individuals telomere dynamic processes appear to differ in relation to developmental stage and between individuals telomere dynamic processes likely differ in relation to a number of intrinsic and extrinsic factors such as heredity and environmentally induced variation in growth and condition. Whilst correlations of telomere length and age are present in the literature, there is currently a poor understanding of processes that effect telomere dynamics and how these vary in space and time in wild animal populations.
This work has provided an advancement in the understanding of methodological considerations of telomere measurement in marine mammals and of comparative cellular aging processes in marine mammals and has proven valuable in guiding future research in this area which is on-going.
