Using osteological measurements to estimate body length in Amazonian manatees

Body length is an important parameter in morphological, ecological and behavioral studies of a species and contributes to the understanding of the body condition of individuals. This parameter is essential for conservation and management strategies by informing studies evaluating growth rates, physical maturity and classification of individuals into age groups, promoting better accuracy for the biological parameters of the species. The aim of this study was to identify predictors of body length for Amazonian manatees (Trichechus inunguis) using metric characters of osteological materials. Eleven linear measurements of skull, jaw, scapulae and humeri were collected from 41 Amazonian manatee skeletons of different age classes (calf, juvenile and adult). Data were analyzed by simple linear regression. The condylobasal length was the best predictor of body length for the species (R = 0.943), however, all bones evaluated showed at least one measurement with the capacity to predict body size (R2 > 0.9). Results of this study are useful for inferring body length of Amazonian manatees using bones deposited in museums and biological collections, expanding the informative potential of these materials.


INTRODUCTION
Body length is an important parameter in morphological, ecological and behavioral studies of a species (Amaral et al. 2010) and essential for determining growth curves and age categorizations (Hildebrandt et al. 2007;Mumby et al. 2015). Additionally, body length is used for the analysis of the body condition of individuals, helping the management and conservation of numerous species of mammals (Rosas et al. 2009). Therefore, body length should be evaluated as accurately as possible given its use in ecological interpretations (Rosas et al. 2009;Sarko et al. 2010).
Several studies have shown efficacy in predicting body length based on morphometric characteristics in aquatic mammals, (Sousa-Lima and Groch 2010; Calderón et al. 2018), including correlations of skull bones or other bones with body length (Itoo and Inoué 1993;Perrin et al. 2005;Sarko et al. 2010;Castelblanco-Martinez et al. 2014;Churchill et al. 2014;Danilewicz 2018). VOL. 51(2) 2021: 156 -161 ACTA AMAZONICA Among sirenians, body lengths of dugongs (Dugong dugon Müller, 1776) have been estimated from cranial measurements by Spain and Heinsohn (1974), Sarko et al. (2010) and Nganvongpanit et al. (2017). Both Sarko et al. (2010) and Castelblanco-Martinez et al. (2014) estimated body length using skull measurements for Florida manatees (Trichechus manatus latirostris Harlam, 1824) and Antillean manatees (Trichechus manatus manatus Linnaeus, 1758), respectively, and demonstrated the strong relationship between cranial measurements and body length for these species. Nganvongpanit et al. (2017) evaluated the use of mandible and scapular measurements in addition to cranial measurements to predict total length in dugongs, and observed lower correlations of mandible and scapular structures to body length compared with those of cranial measurements. Although this body length estimation technique has been validated in different sirenian species, as shown by Sarko et al. (2010), there are no reports of body length estimates for Amazonian manatees (Trichechus inunguis Natterer, 1883) using bone measurements, and none of these existing equations have been applied to Amazonian manatees.
The Amazonian manatee is the smallest sirenian and the only species that occurs exclusively in freshwater (Rosas 1994). Predatory hunting of Amazonian manatees, although illegal, currently persists throughout its distribution and, consequently, it is common to find incomplete skeletons deposited in biological collections (Rosas 1994;Franzini et al. 2013;Souza 2015). Thus, important biological information, such as gender and body length, frequently cannot be determined accurately (Vergara-Parente et al. 2010), preventing the use of these specimens in studies of population structure and hunting pressure, for example. Therefore, this study aimed to identify predictors of body length for Amazonian manatees using different bones available in biological collections.

MATERIAL AND METHODS
We measured skeletons from 41 Amazonian manatees (calves, juveniles and adults), with body lengths from 62.8 cm to 257 cm, deposited in the aquatic mammal collection under the supervision of the Aquatic Mammal Laboratory of Instituto Nacional de Pesquisas da Amazônia (INPA), Manaus, Brazil. Body length data (measured in a straight line from the tip of the snout to the tip of the fluke) were available for all the specimens analyzed. However, most specimens did not have complete skeletons, thus, 29 skulls (14 ♂ and 15 ♀), 41 mandibles (21 ♂ and 20 ♀), 33 scapulae (18 ♂ and 15 ♀) and 22 humeri (13 ♂ and 9 ♀) were measured. Since Amazonian manatees do not present sexual dimorphism (Domning and Hayek 1986;Amaral et al. 2010), data from both sexes were combined and analyzed together.
All materials were measured by one researcher (GCMV) with a TIN 300 mm analog Digimess caliper, São Paulo, SP, Brazil. Skull and jaw measurements were obtained according to Domning and Hayek (1986) and Sarko et al. (2010). Measurements of humeri and scapulae followed Ali et al. (2016) and Nganvongpanit et al. (2017). Linear measurements of the bones are presented in Figure 1 with associated acronyms in Table 1. Paired bones (humeri and scapulae) were compared with paired t-tests to assess whether measurements of the right or left side can be used interchangeably. The relationship between body length (BL) and each measured parameter was obtained by simple linear regression. Subsequently, the parameters in each bone with high correlation to total length (adjusted R 2 > 0.9) were reanalyzed by multiple linear regression. All statistical analyses were performed using R Program version 4.0.2 (2020-06-22) from The R Foundation for Statistical Computing.  Table 1.

RESULTS
The scapulae and humeri showed no significant difference between the right and left sides for all measurements (P > 0.05; paired t-test; n = 22 scapulae and 22 humeri). Thus, we used the average of measurements from both sides. Only two specimens lacked one humerus and radius each, and measurements from the existing side were used. Of the variables analyzed in this study, the condylobasal length (BSL) was the parameter that showed the highest correlation coefficient (Table 2; Figure 2). However, the other bones showed at least one measurement with a high capacity (adjusted R 2 > 0.9) to predict body length in the Amazonian manatee (mandible: MTL, SPW, and GPW; scapulae: STL; and humeri: GWE) ( Table 2; Figures 2 and 3).
Multiple linear regression using BSL, MTL, SPW, GPW, LNO and STL presented an adjusted correlation value of R 2 = 0.961 (p > 0.0001) and a residual value of 17.5 using the following equation:

DISCUSSION
In this study, the condylobasal length (BSL) was the parameter that best estimated body length of Amazonian manatees. This measurement has also been described as an excellent predictor of body length in other orders of aquatic mammals, such as pinnipeds (Churchill et al. 2014)  The use of mandible measurements to estimate body length in mammals has been reported for manatees, pinnipeds, cetaceans and several terrestrial mammals (Domning and Hayek 1986;Nummela et al. 2004;Churchill et al. 2014;Stansfield 2015). In T. inunguis, development of the mandible and its structures can be correlated with the physical maturity of the animal. Domning and Hayek (1986) described ontogenetic variations in morphometric parameters of the mandible, including the parameters used in the present study (anteroposterior mandible length and mandibular symphysis height). Although these studies did not directly correlate morphometric measurements with the individual body lengths, mandible ontogenetic variation described in Domning and Hayek (1986) supports the effective prediction of body length found in our study when using these mandibular measurements. Nganvongpanit et al. (2017) used linear measurements of D. dugon scapulae to estimate body length in this species, showing that both cranial and scapulae measurements were good predictors of body size in dugongs. In the present study, we used two scapulae measurements used by Nganvongpanit et al. (2017), and the scapulae total length (STL) was satisfactory in determining body length in Amazonian manatees.
The relationship between width of the lateral and medial humeral epicondyles and body length of Amazonian manatees observed in our study may be related to occurrence of pachyostosis in the bones of some tetrapods secondarily adapted to aquatic life (de Buffrénil et al. 2010). The proximal portion of the humerus of Amazonian manatees becomes quite robust as the animal develops, with the medial epicondyle being markedly broader than the lateral epicondyle, making this bone region quite large when compared to the rest of the structure.
Although multiple linear regression showed a high predictive ability for body length, it is not a practical tool to estimate body length since it requires the presence of a skull, a mandible, scapulae and humeri from the same individual in the osteological collection. Therefore, considering the high R 2 values obtained in this study, we recommend the independent use of the skull (BSL), humerus (GWE), mandible (MTL, GPW or SPW) or scapula (STL) measurements, in that order, to provide a better estimate of the body length of the Amazonian manatee.
Considering the lack of body length records in a significant number of specimens deposited in biological collections and the imprecise records of individual body lengths, the equations obtained in this study may be useful to estimate these important biological data. For example, Vergara-Parente et al. (2010) had to use an imprecise body length measurement (estimated using the palmos length of a hunter's hand, converted to 21.71 cm) to calculate growth curves for the Amazonian manatees. This imprecision could be reduced using one of the equations presented herein. Additionally, these results can be used in future studies to evaluate growth rates, physical maturity and classification of individuals into age groups, promoting a better accuracy for the biological parameters of the species.

CONCLUSIONS
Our results indicate that it is possible to use morphometric measurements of skull, humerus, mandible or scapula to estimate the body length of Amazonian manatees when the whole skeleton is not available. Among the parameters evaluated, the condylobasal length was the best predictor of body length.