Schematic overview of the endocrine and avarian events during the estrous cycle (courtesy Imke Lüders).
LUF= Luteinizing follicles
LH = Luteinizing Hormone
Pm = Progestagens
CL = Corpus Luteum
ovCL = CL from ovulation
acCLs = accessory corpora lutea
Progestagens in elephants include a large group of progesterone-like molecules. The major part of progestagens found in the blood of elephants does not react in the usual human assays. When starting monitoring progestagens, one should always validate first the assay by providing serum samples with known progestagen concentrations of different levels. Even the use of the same hardware, is not always a guarantee that the results are reliable. Companies permanently develop news progesterone assays for the same machine, which are more specific for human progestagens and less for elephant progestagens.
The estrous cycle can be divided in 2 major phases, the luteal phase and the non-luteal or follicular phase. The luteal phase starts after ovulation and lasts on average 10 weeks. It is characterized by elevated progestagen blood levels. The follicular phase starts when the progestagen blood concentration has dropped to what we call in this document the “base line” value.
Progesterone serum levels in mmol/l during the estrous cycle in an elephant. Note the 2 LH-peaks (red arrows).
Many lab assays fail to accurately measure progestagens in the blood during the follicular phase in elephants. Some machines calculate these low concentrations rather then measuring them exactly. Moreover, the progestagen concentrations that are provided by the different laboratories, vary greatly depend on the machine and technique used. It is therefore very important to use the same technique and machine for monitoring the estrous cycle of an individual elephant. Note: progestagen concentrations measured by one assay may differ up to a factor 10 from the results obtained when another assay is used.
Automated analysers than have proven to give reliable results for both species are:
ADVIA Centaur XP (Siemens) with ranges between 0.21 mmol/l and 10 mmol/l.
miniVidas with ranges between 0.79 mmol/l and 15 mmol/l.
During the follicular phase, approximately 18-20 days prior to ovulation, a 1-day LH-peak concentration can be distinguished, which may result in a temporary rise of progesterone. This first LH-peak results in ovarian follicles that will not ovulate. These follicles produce progesterone untill they go into gregression in the luteal phase.
A second LH-peak occurs just prior to ovulation. This LH-peak induced ovulation in one follicle. The remaining corpus luteum will also produce progesterone and maintain the luteal phase for about 10 weeks.
To monitor the estrous cycle in elephants gestagens can be measured in blood
(progesterone), urine (pregnanetriols, Primate Center Göttingen, Germany) or feces (progesterone, Chester Zoo, UK). Frequency of sampling should be once a week for blood and urine and 3 times per week for feces.
Urinary hormone analyses are performed in the Endocrinology Lab at the German Primate Centre in Goettingen, Germany. All facilities in Europe are welcome to send samples on a voluntary basis and assays are run every week. The service for the Asian elephants started in 1994 and that for African elephants in 1996, so comparative data are available for more than 20 years. It is important to note that in urine not progesterone itself but its metabolites are measured. These metabolites differ between the two elephant species. Whilst in Asian elephants pregnanetriol (P3) is the most abundant metabolite in urine, it is 5α-pregnane-3-ol-20–one (5α-P-3-OH) in African elephants. This means that different assays are needed for Asian and African elephants. The pregnanetriol concentration in the urine is always compared with urinary creatinine. If the creatinine level is too low, a new sample should be submitted.
Urine can be collected in many different ways depending on keeping and housing system. Elephants can be trained to urinate on command. Only 2 ml urine are needed for the analysis. It is good to use plastic tubes that close well, best with screw lid. Labelling of the sample is essential! Labels must be waterproof and show the name of animal and the date of collection. If the samples are not sent within 2 days after collection, they need to be frozen soon after collection. See also the Practical guide for urine collection, Ann-Kathrin Oerke).