Validation of an enzyme immunoassay for the non-invasive measurement of faecal androgen metabolites in spinifex hopping mice (Notomys alexis)
Kelly S. Williams-Kelly
A B * , Kylie A. Robert A B , Rupert Palme C and Kerry V. Fanson A
+ Author Affiliations
- Author Affiliations
A Discipline of Animal Physiology and Health, School of Agriculture, Biomedicine and Environment, La Trobe University, Melbourne, Vic. 3086, Australia.
B Centre for Future Landscapes, La Trobe University, Melbourne, Vic. 3086, Australia.
C Department of Biomedical Sciences, University of Veterinary Medicine, 1210 Vienna, Austria.
Australian Mammalogy 45(2) 192-198 https://doi.org/10.1071/AM22025
Submitted: 3 August 2022 Accepted: 22 October 2022 Published: 15 November 2022
© 2023 The Author(s) (or their employer(s)). Published by CSIRO Publishing on behalf of the Australian Mammal Society.
Abstract
Androgens may play a key role in shaping the unique reproductive traits of male spinifex hopping mice (Notomys alexis), but little is known about the reproductive endocrinology of this species. Measurement of faecal androgen metabolites (FAMs) offers a non-invasive tool for monitoring testicular activity. Before applying this tool, physiological validation is required to demonstrate that changes in FAMs reflect changes in circulating testosterone for a given species. The goal of this study was to validate an enzyme immunoassay for monitoring FAMs in Notomys alexis. We compared the performance of two different assays (UVM-T and R156/7) for monitoring changes in FAMs following the administration of human chorionic gonadotropin (hCG) to stimulate androgen production by the testes. Both assays detected a significant increase in FAMs and had similar patterns. The UVM-T assay was more sensitive to changes in testicular activity, with a shorter excretion lag time and greater response magnitude. These findings indicate that we can reliably monitor testicular activity using faecal hormone metabolite analysis in Notomys alexis and can now utilise FAM measurements to better understand the species’ unusual reproductive biology.
Keywords: EIA, faecal metabolites, hormone, Muridae, rodent, sex steroids, testicular activity, testosterone.
References
Allen, C. D., McKinnon, A. J., Lisle, A. T., D’Occhio, M. J., and Johnston, S. D. (2006). Use of a GnRH agonist and hCG to obtain an index of testosterone secretory capacity in the koala (Phascolarctos cinereus). Journal of Andrology 27, 720–724.| Use of a GnRH agonist and hCG to obtain an index of testosterone secretory capacity in the koala (Phascolarctos cinereus).Crossref | GoogleScholarGoogle Scholar |
Auer, K. E., Kußmaul, M., Möstl, E., Hohlbaum, K., Rülicke, T., and Palme, R. (2020). Measurement of fecal testosterone metabolites in mice: replacement of invasive techniques. Animals 10, 165.
| Measurement of fecal testosterone metabolites in mice: replacement of invasive techniques.Crossref | GoogleScholarGoogle Scholar |
Bauer, M., and Breed, W. G. (2006). Variation of sperm head shape and tail length in a species of Australian hydromyine rodent: the spinifex hopping mouse, Notomys alexis. Reproduction, Fertility and Development 18, 797–805.
| Variation of sperm head shape and tail length in a species of Australian hydromyine rodent: the spinifex hopping mouse, Notomys alexis.Crossref | GoogleScholarGoogle Scholar |
Bauer, M., and Breed, W. G. (2008). Testis mass of the spinifex hopping mouse and its impact on fertility potential. Journal of Zoology 274, 349–356.
| Testis mass of the spinifex hopping mouse and its impact on fertility potential.Crossref | GoogleScholarGoogle Scholar |
Berris, K. K., Breed, W. G., and Carthew, S. M. (2020a). What can we deduce about the reproductive condition of spinifex hopping mice (Notomys alexis) from external examination? Australian Mammalogy 42, 11–15.
| What can we deduce about the reproductive condition of spinifex hopping mice (Notomys alexis) from external examination?Crossref | GoogleScholarGoogle Scholar |
Berris, K. K., Breed, W. G., Moseby, K. E., and Carthew, S. M. (2020b). Female reproductive suppression in an Australian arid zone rodent, the spinifex hopping mouse. Journal of Zoology 312, 163–173.
| Female reproductive suppression in an Australian arid zone rodent, the spinifex hopping mouse.Crossref | GoogleScholarGoogle Scholar |
Billitti, J. E., Lasley, B. L., and Wilson, B. W. (1998). Development and validation of a fecal testosterone biomarker in Mus musculus and Peromyscus maniculatus. Biology of Reproduction 59, 1023–1028.
| Development and validation of a fecal testosterone biomarker in Mus musculus and Peromyscus maniculatus.Crossref | GoogleScholarGoogle Scholar |
Breed, W. G. (1983). Sexual dimorphism in the Australian hopping mouse, Notomys alexis. Journal of Mammalogy 64, 536–539.
| Sexual dimorphism in the Australian hopping mouse, Notomys alexis.Crossref | GoogleScholarGoogle Scholar |
Breed, W. G., and Leigh, C. M. (2011). Reproductive biology of an old endemic murid rodent of Australia, the spinifex hopping mouse, Notomys alexis: adaptations for life in the arid zone. Integrative Zoology 6, 321–333.
| Reproductive biology of an old endemic murid rodent of Australia, the spinifex hopping mouse, Notomys alexis: adaptations for life in the arid zone.Crossref | GoogleScholarGoogle Scholar |
Breed, W. G., Ding, X., Tuke, J., and Leigh, C. M. (2020). Morphological diversity of the spermatozoon and male reproductive tract in Australian hopping mice, genus Notomys – is it determined by sexual selection? Journal of Zoology 311, 194–203.
| Morphological diversity of the spermatozoon and male reproductive tract in Australian hopping mice, genus Notomys – is it determined by sexual selection?Crossref | GoogleScholarGoogle Scholar |
Brown, J. L., Wildt, D. E., Wielebnowski, N., Goodrowe, K. L., Graham, L. H., Wells, S., and Howard, J. G. (1996). Reproductive activity in captive female cheetahs (Acinonyx jubatus) assessed by faecal steroids. Journal of Reproduction and Fertility 106, 337–346.
| Reproductive activity in captive female cheetahs (Acinonyx jubatus) assessed by faecal steroids.Crossref | GoogleScholarGoogle Scholar |
Busso, J. M., Ponzio, M. F., Dabbene, V., de Cuneo, M. F., and Ruiz, R. D. (2005). Assessment of urine and fecal testosterone metabolite excretion in Chinchilla lanigera males. Animal Reproduction Science 86, 339–351.
| Assessment of urine and fecal testosterone metabolite excretion in Chinchilla lanigera males.Crossref | GoogleScholarGoogle Scholar |
Curry, E., Roth, T.L., Mackinnon, K.M., and Stoops, M.A. (2012). Factors influencing annual fecal testosterone metabolite profiles in captive male polar bears (Ursus maritimus). Reproduction in Domestic Animals 47, 222–225.
| Factors influencing annual fecal testosterone metabolite profiles in captive male polar bears (Ursus maritimus).Crossref | GoogleScholarGoogle Scholar |
de Bruin, P. R., Medger, K., Bennett, N. C., and Ganswindt, A. (2014). Assessment of reproductive function in southern African spiny mice (Acomys spinosissimus) using faeces as hormone matrix. African Zoology 49, 44–53.
| Assessment of reproductive function in southern African spiny mice (Acomys spinosissimus) using faeces as hormone matrix.Crossref | GoogleScholarGoogle Scholar |
Fanson, B., and Fanson, K. V. (2015). hormLong: An R package for longitudinal data analysis in wildlife endocrinology studies. PeerJ Preprints 3, e1546v1.
| hormLong: An R package for longitudinal data analysis in wildlife endocrinology studies.Crossref | GoogleScholarGoogle Scholar |
Fanson, K. V., Best, E. C., Bunce, A., Fanson, B. G., Hogan, L. A., Keeley, T., Narayan, E. J., Palme, R., Parrott, M. L., Sharp, T. M., Skogvold, K., Tuthill, L., Webster, K. N., and Bashaw, M. (2017). One size does not fit all: monitoring faecal glucocorticoid metabolites in marsupials. General and Comparative Endocrinology 244, 146–156.
| One size does not fit all: monitoring faecal glucocorticoid metabolites in marsupials.Crossref | GoogleScholarGoogle Scholar |
Freeman, H. D., Wood, M., Schook, M. W., Leighty, K. A., Lavin, S. R., Wiebe, S., Blowers, T. E., Daneault, R., Mylniczenko, N., and Wheaton, C. J. (2018). Seasonal dynamics of agonistic behavior and hormones in an ex situ all-male colony of large flying foxes. Zoo Biology 37, 213–222.
| Seasonal dynamics of agonistic behavior and hormones in an ex situ all-male colony of large flying foxes.Crossref | GoogleScholarGoogle Scholar |
Ganswindt, A., Heistermann, M., Borragan, S., and Hodges, J. K. (2002). Assessment of testicular endocrine function in captive African elephants by measurement of urinary and fecal androgens. Zoo Biology 21, 27–36.
| Assessment of testicular endocrine function in captive African elephants by measurement of urinary and fecal androgens.Crossref | GoogleScholarGoogle Scholar |
Jean-Faucher, C., Berger, M., De Turckheim, M., Veyssiere, G., and Jean, C. (1985). Testosterone and dihydrotestosterone levels in epididymis, vas deferens, seminal vesicle and preputial gland of mice after hCG injection. Journal of Steroid Biochemistry 23, 201–205.
| Testosterone and dihydrotestosterone levels in epididymis, vas deferens, seminal vesicle and preputial gland of mice after hCG injection.Crossref | GoogleScholarGoogle Scholar |
Kersey, D. C., and Dehnhard, M. (2014). The use of noninvasive and minimally invasive methods in endocrinology for threatened mammalian species conservation. General and Comparative Endocrinology 203, 296–306.
| The use of noninvasive and minimally invasive methods in endocrinology for threatened mammalian species conservation.Crossref | GoogleScholarGoogle Scholar |
McLennan, H. J., Lüpold, S., Smissen, P., Rowe, K. C., and Breed, W. G. (2017). Greater sperm complexity in the Australasian old endemic rodents (Tribe: Hydromyini) is associated with increased levels of inter-male sperm competition. Reproduction, Fertility and Development 29, 921–930.
| Greater sperm complexity in the Australasian old endemic rodents (Tribe: Hydromyini) is associated with increased levels of inter-male sperm competition.Crossref | GoogleScholarGoogle Scholar |
Möhle, U., Heistermann, M., Palme, R., and Hodges, J. K. (2002). Characterization of urinary and fecal metabolites of testosterone and their measurement for assessing gonadal endocrine function in male nonhuman primates. General and Comparative Endocrinology 129, 135–145.
| Characterization of urinary and fecal metabolites of testosterone and their measurement for assessing gonadal endocrine function in male nonhuman primates.Crossref | GoogleScholarGoogle Scholar |
Mooradian, A. D., Morley, J. E., and Korenman, S. G. (1987). Biological actions of androgens. Endocrine Reviews 8, 1–28.
| Biological actions of androgens.Crossref | GoogleScholarGoogle Scholar |
Morrissey, B. L., and Breed, W. G. (1982). Variation in external morphology of the glans penis of Australian native rodents. Australian Journal of Zoology 30, 495–502.
| Variation in external morphology of the glans penis of Australian native rodents.Crossref | GoogleScholarGoogle Scholar |
Ninnes, C. E., Waas, J. R., Ling, N., Nakagawa, S., Banks, J. C., Bell, D. G., Bright, A., Carey, P. W., Chandler, J., Hudson, Q. J., Ingram, J. R., Lyall, K., Morgan, D. K. J., Stevens, M. I., Wallace, J., and Möstl, E. (2010). Comparing plasma and faecal measures of steroid hormones in Adelie penguins Pygoscelis adeliae. Journal of Comparative Physiology B 180, 83–94.
| Comparing plasma and faecal measures of steroid hormones in Adelie penguins Pygoscelis adeliae.Crossref | GoogleScholarGoogle Scholar |
Palme, R. (2005). Measuring fecal steroids: guidelines for practical application. Annals of the New York Academy of Sciences 1046, 75–80.
| Measuring fecal steroids: guidelines for practical application.Crossref | GoogleScholarGoogle Scholar |
Palme, R. (2019). Non-invasive measurement of glucocorticoids: advances and problems. Physiology & Behavior 199, 229–243.
| Non-invasive measurement of glucocorticoids: advances and problems.Crossref | GoogleScholarGoogle Scholar |
Palme, R., and Möstl, E. (1994). Biotin-streptavidin enzyme immunoassay for the determination of oestrogens and androgens in boar faeces. In ‘Proceedings of the 5th Symposium on the Analysis of Steroids’. (Ed. S. Görög.) pp. 111–117. (Akademiai Kiado: Budapest.)
Pribbenow, S., Shrivastav, T. G., and Dehnhard, M. (2017). Measuring fecal testosterone metabolites in spotted hyenas: choosing the wrong assay may lead to erroneous results. Journal of Immunoassay and Immunochemistry 38, 308–321.
| Measuring fecal testosterone metabolites in spotted hyenas: choosing the wrong assay may lead to erroneous results.Crossref | GoogleScholarGoogle Scholar |
R Core Team (2021). ‘R: A language and environment for statistical computing.’ (R Foundation for Statistical Computing: Vienna, Austria.) Available at https://www.R-project.org/
Schradin, C. (2008). Seasonal changes in testosterone and corticosterone levels in four social classes of a desert dwelling sociable rodent. Hormones and Behavior 53, 573–579.
| Seasonal changes in testosterone and corticosterone levels in four social classes of a desert dwelling sociable rodent.Crossref | GoogleScholarGoogle Scholar |
Sipari, S., Ylönen, H., and Palme, R. (2017). Excretion and measurement of corticosterone and testosterone metabolites in bank voles (Myodes glareolus). General and Comparative Endocrinology 243, 39–50.
| Excretion and measurement of corticosterone and testosterone metabolites in bank voles (Myodes glareolus).Crossref | GoogleScholarGoogle Scholar |
Stanley, M. (1971). An ethogram of the hopping mouse, Notomys alexis. Zeitschrift für Tierpsychologie 29, 225–258.
| An ethogram of the hopping mouse, Notomys alexis.Crossref | GoogleScholarGoogle Scholar |
Touma, C., and Palme, R. (2005). Measuring fecal glucocorticoid metabolites in mammals and birds: the importance of validation. Annals of the New York Academy of Sciences 1046, 54–74.
| Measuring fecal glucocorticoid metabolites in mammals and birds: the importance of validation.Crossref | GoogleScholarGoogle Scholar |
Umapathy, G., Deepak, V., Kumar, V., Chandrasekhar, M., and Vasudevan, K. (2015). Endocrine profiling of endangered tropical chelonians using noninvasive fecal steroid analyses. Chelonian Conservation and Biology 14, 108–115.
| Endocrine profiling of endangered tropical chelonians using noninvasive fecal steroid analyses.Crossref | GoogleScholarGoogle Scholar |
Walker, S. L., Waddell, W. T., and Goodrowe, K. L. (2002). Reproductive endocrine patterns in captive female and male red wolves (Canis rufus) assessed by fecal and serum hormone analysis. Zoo Biology 21, 321–335.
| Reproductive endocrine patterns in captive female and male red wolves (Canis rufus) assessed by fecal and serum hormone analysis.Crossref | GoogleScholarGoogle Scholar |
