Elizabeth A. Burgess

Swimming performance of hatchling green turtles is affected by incubation temperature

In an experiment repeated for two separate years, incubation temperature was found to affect the body size and swimming performance of hatchling green turtles (Chelonia mydas). In the first year, hatchlings from eggs incubated at 26°C were larger in size than hatchlings from 28 and 30°C, whilst in the second year hatchlings from 25.5°C were similar in size to hatchings from 30°C. Clutch of origin influenced the size of hatchlings at all incubation temperatures even when differences in egg size were taken into account. In laboratory measurements of swimming performance, in seawater at 28°C, hatchlings from eggs incubated at 25.5 and 26°C had a lower stroke rate frequency and lower force output than hatchlings from 28 and 30°C. These differences appeared to be caused by the muscles of hatchlings from cooler temperatures fatiguing at a faster rate. Clutch of origin did not influence swimming performance. This finding that hatchling males incubated at lower temperature had reduced swimming ability may affect their survival whilst running the gauntlet of predators in shallow near-shore waters, prior to reaching the relative safety of the open sea.

Testosterone and tusks: maturation and seasonal reproductive patterns of live, free-ranging male dugongs (Dugong dugon) in a subtropical population

Knowledge of male reproductive status and activity in free-ranging animals is vital to understanding reproductive patterns and population dynamics. Until now, almost all information regarding reproductive behavior of the dugong, a cryptic marine mammal, has relied on post-mortem examination. We examined the relationships between body length, tusk eruption (secondary sexual characteristic), seasonality, and group association on fecal testosterone metabolite concentrations in 322 free-ranging dugongs (159 males, 163 females) in subtropical Moreton Bay, Australia. Fecal testosterone concentrations demonstrated biologically meaningful differences in testicular activity between sexes and across reproductive/age classes, and were correlated with circulating concentrations in serum. Male dugongs have a pre-reproductive period that persists until a body length of 240 cm is achieved. Puberty apparently occurs between 240 and 260 cm body length when fecal testosterone levels increase fourfold (>500 ng/g) over juvenile levels, and is associated with tusk eruption. However, social maturity may be delayed until male dugongs are larger than 260 cm with well-developed tusks. In mature males, the lowest (<500 ng/g) fecal testosterone concentrations occur in the austral autumn months with maximal concentrations in September-October, coincident with the onset of a spring mating season. During spring, solitary mature males had fecal testosterone concentrations double those of mature males sampled within groups, potentially suggesting a mating strategy involving roving of reproductively active males. This study demonstrates that single-point physiological data from individuals across a population have value as indicators of reproductive processes. Our approach provides an efficacious non-lethal method for the census of reproductive status and seasonality in live male dugongs.

Diagnosing pregnancy in free-ranging dugongs using fecal progesterone metabolite concentrations and body morphometrics: A population application

Assessing reproductive status and monitoring reproductive rates is important in the effective management of vulnerable marine mammal species such as the dugong (Dugong dugon). Knowledge of the reproductive physiology of this species is limited, and determining reproductive parameters (e.g., sexual maturation, pregnancy, and reproductive senescence) has been restricted by a lack of non-lethal methods for assessing reproductive status in free-ranging individuals. The aim of this study was to develop a method to identify pregnant individuals in a wild dugong population. Using an enzymeimmunoassay, we quantified concentrations of fecal progesterone metabolites (fP) in 322 dugongs, including confirmed pregnant females (n=10), presumed non-pregnant adult females (n=25), juvenile females (n=24), subadult females (n=41), adult females of unknown pregnancy state (n=63), and males of all sizes (n=159). External body morphometrics of each dugong were measured, and confirmation of pregnancy in adult female dugongs was determined by ultrasonography or observation of subsequent neonates. Concentrations of fP were different between sexes and reproductive size classes (P<0.001), and ∼30-fold higher in confirmed pregnant dugongs (2017-7760 ng/g) compared to presumed non-pregnant females (30-221 ng/g), juvenile females (29-195 ng/g), and males (24-261 ng/g) (P<0.001). Body measures of maximum and anal girths, and teat length were all greater in confirmed pregnant females than presumed non-pregnant females (all P<0.05). We evaluated a Discriminant Function Analysis (DFA) to provide a model for predicting pregnant and non-pregnant dugongs. Cross-validated results showed that the DFA correctly classified 100% of pregnant and non-pregnant females using fP concentrations, body length, fineness ratio (an index of body shape), and teat length (a female reproductive trait). Using the DFA model, we classified the pregnancy status of all female dugongs and identified a total of 30 females as pregnant and 133 females as non-pregnant from the sampled population over the sample period. Pregnant dugongs in the Moreton Bay population are characterized by fecal progesterone metabolite concentrations > 1000 ng/g, body length ≥ 260 cm, maximum girth ≥ 215 cm, anal girth ≥ 126 cm, and teat length ≥ 5 cm long. In summary, analysis of fP concentrations in combination with body morphometrics may be used to diagnose pregnancy in free-ranging dugongs, and provides a new tool to monitor breeding rates of wild sirenian populations.

Sex, scarring, and stress: understanding seasonal costs in a cryptic marine mammal.

This study demonstrates that fecal glucocorticoid levels are a useful measure of diverse stressors in dugongs (a vulnerable cryptic marine mammal), including cold temperature and other seasonal factors, intraspecific aggression/injury, and pregnancy. The intrinsic underlying glucocorticoid patterns of dugongs are different between sexes, across reproductive maturity states, and vary seasonally in response to reproductive patterns and environmental conditions.

Variation in the hindgut microbial communities of the Florida manatee, Trichechus manatus latirostris over winter in Crystal River, Florida

The Florida manatee, Trichechus manatus latirostris, is a hindgut-fermenting herbivore. In winter, manatees migrate to warm water overwintering sites where they undergo dietary shifts and may suffer from cold-induced stress. Given these seasonally induced changes in diet, the present study aimed to examine variation in the hindgut bacterial communities of wild manatees overwintering at Crystal River, west Florida. Faeces were sampled from 36 manatees of known sex and body size in early winter when manatees were newly arrived and then in mid-winter and late winter when diet had probably changed and environmental stress may have increased. Concentrations of faecal cortisol metabolite, an indicator of a stress response, were measured by enzyme immunoassay. Using 454-pyrosequencing, 2027 bacterial operational taxonomic units were identified in manatee faeces following amplicon pyrosequencing of the 16S rRNA gene V3/V4 region. Classified sequences were assigned to eight previously described bacterial phyla; only 0.36% of sequences could not be classified to phylum level. Five core phyla were identified in all samples. The majority (96.8%) of sequences were classified as Firmicutes (77.3 ± 11.1% of total sequences) or Bacteroidetes (19.5 ± 10.6%). Alpha-diversity measures trended towards higher diversity of hindgut microbiota in manatees in mid-winter compared to early and late winter. Beta-diversity measures, analysed through PERMANOVA, also indicated significant differences in bacterial communities based on the season.

Small intestinal volvulus in a free-ranging female dugong (Dugong dugon).

An adult female dugong (Dugong dugon) was found dead and floating in Moreton Bay, Queensland, Australia. This animal was found to have a 360° mesenteric volvulus with infarction of the associated segment of small intestine, and fibrinous peritonitis. Mortality was attributed to the volvulus and its sequelae. The cause was not apparent on gross or histological examination.

Reproductive hormone monitoring of dugongs in captivity: Detecting the onset of sexual maturity in a cryptic marine mammal

Determining the reproductive status of long-term captive animals is essential because the onset of sexual maturity and reproductive activity may necessitate changes in husbandry requirements. This study reports on the first multi-year reproductive hormone monitoring program for captive dugongs of both sexes using feces. Fecal samples were collected from one male (Pig) over 9 years (4-13.2y of age; n=288 samples, 0.8±0.1 samples per week from July 2007 to February 2012) and one female (Wuru) over 7 years (from neonate to 6.9 y; n=171 samples, 0.5±0.1 samples per week from July 2007 to February 2012), and from one solitary female dugong (Gracie) over 10 months (10.5-11.3y of age; n=54 samples, 1.1±0.2 sample per week from September 2008 to June 2009). Using enzyme-immunoassay, fecal progesterone (fP) and estradiol-17β (fE) concentrations were assayed in the two captive females, and testosterone (fT) concentration in the captive male, and compared these to concentrations in wild dugongs. Female Wuru exhibited increasing fP concentrations at 5+ y, indicating early onset of ovarian cycling typical of non-pregnant adult females. Female Gracie maintained basal fP concentrations consistent with wild immature dugongs, indicating that she had not reached puberty by 11y. Nutritional plane may account for differences in age at sexual maturity in these female dugongs. At age 3-4y, Wuru had fE concentrations 1.4 times greater than maximum concentrations recorded in all wild females, and these concentrations were coincident with a period of rapid weight gain. For the male Pig, increasing fT concentrations at 9y provided early indications of puberty. Pigs tusks erupted by 11y, and sexual maturity (indicated by spermatic semen) was confirmed by 12.8y. Identification of sexual maturation prompted two trials of a male contraceptive treatment using the GnRH agonist, deslorelin (9.4mg administered in 2010 and 15.6mg in 2011). Testosterone production was not significantly suppressed by these dosages, and treatment did not terminate sperm production at week 10-11 post-implantation, even at the larger dose tested. Routine analysis of fecal hormones was helpful for making reproductive management decisions regarding individual captives and in guiding the long-term captive management of this cryptic species.

Methods to Examine Reproductive Biology in Free-Ranging, Fully-Marine Mammals

Historical overexploitation of marine mammals, combined with present-day pressures, has resulted in severely depleted populations, with many species listed as threatened or endangered. Understanding breeding patterns of threatened marine mammals is crucial to assessing population viability, potential recovery and conservation actions. However, determining reproductive parameters of wild fully-marine mammals (cetaceans and sirenians) is challenging due to their wide distributions, high mobility, inaccessible habitats, cryptic lifestyles and in many cases, large body size and intractability. Consequently, reproductive biologists employ an innovative suite of methods to collect useful information from these species. This chapter reviews historic, recent and state-of-the-art methods to examine diverse aspects of reproduction in fully-aquatic mammals.