A new DNA extraction method (HV-CTAB-PCI) for amplification of nuclear markers from open ocean-retrieved faeces of an herbivorous marine mammal, the dugong.

Non-invasively collected faecal samples are an alternative source of DNA to tissue samples, that may be used in genetic studies of wildlife when direct sampling of animals is difficult. Although several faecal DNA extraction methods exist, their efficacy varies between species. Previous attempts to amplify mitochondrial DNA (mtDNA) markers from faeces of wild dugongs (Dugong dugon) have met with limited success and nuclear markers (microsatellites) have been unsuccessful. This study aimed to establish a tool for sampling both mtDNA and nuclear DNA (nDNA) from dugong faeces by modifying approaches used in studies of other large herbivores. First, a streamlined, cost-effective DNA extraction method that enabled the amplification of both mitochondrial and nuclear markers from large quantities of dugong faeces was developed. Faecal DNA extracted using a new 'High Volume- Cetyltrimethyl Ammonium Bromide- Phenol-Chloroform-Isoamyl Alcohol' (HV-CTAB-PCI) method was found to achieve comparable amplification results to extraction of DNA from dugong skin. As most prevailing practices advocate sampling from the outer surface of a stool to maximise capture of sloughed intestinal cells, this study compared amplification success of mtDNA between the outer and inner layers of faeces, but no difference in amplification was found. Assessment of the impacts of faecal age or degradation on extraction, however, demonstrated that fresher faeces with shorter duration of environmental (seawater) exposure amplified both markers better than eroded scats. Using the HV-CTAB-PCI method, nuclear markers were successfully amplified for the first time from dugong faeces. The successful amplification of single nucleotide polymorphism (SNP) markers represents a proof-of-concept showing that DNA from dugong faeces can potentially be utilised in population genetic studies. This novel DNA extraction protocol offers a new tool that will facilitate genetic studies of dugongs and other large and cryptic marine herbivores in remote locations.

Diet composition of the African manatee: Spatial and temporal variation within the Sanaga River Watershed, Cameroon.

The present study aimed to investigate the diet of African manatees in Cameroon to better inform conservation decisions within protected areas. A large knowledge gap on diet and seasonal changes in forage availability limits the ability to develop informed local management plans for the African manatee in much of its range. This research took place in the Sanaga River Watershed, which includes two protected areas in the Littoral Region of Cameroon: the Douala-Edea National Park and the Lake Ossa Wildlife Reserve. We analyzed 113 manatee fecal samples and surveyed shoreline emergent and submerged vegetation within the Sanaga River Watershed. We used microhistological analyses to determine the relative contribution of each plant species to African manatee diets and compared across locations and across seasons (wet vs. dry season). We found that the shoreline vegetation is diverse with over 160 plant species, unevenly distributed across space and season, and dominated by emergent vegetation mostly represented by the antelope grass (Echinochloa pyramidalis). We recorded a total of 36 plant species from fecal samples with a spatial and temporal distribution mostly reflecting that of the corresponding shoreline vegetation. African manatees appear to be primarily opportunistically feeding on available vegetation across the seasons and habitat. This work documents the current, but changing, state of plant availability in the Sanaga River Watershed and reports the African manatee diet in Cameroon for the first time. This information can play a critical role in successfully managing the species and these protected areas. If we wish to protect the African manatee and the aquatic ecosystems within the Sanaga River Watershed, we must understand how forage availability changes over time, especially as its waters become nutrient enriched, eutrophic, and exposed to invasive species of plants in a changing world.

First characterization of vocalizations and passive acoustic monitoring of the vulnerable African manatee (Trichechus senegalensis).

Even among the understudied sirenians, African manatees (Trichechus senegalensis) are a poorly understood, elusive, and vulnerable species that is difficult to detect. We used passive acoustic monitoring in the first effort to acoustically detect African manatees and provide the first characterization of their vocalizations. Within two 3-day periods at Lake Ossa, Cameroon, at least 3367 individual African manatee vocalizations were detected such that most vocalizations were detected in the middle of the night and at dusk. Call characteristics such as fundamental frequency, duration, harmonics, subharmonics, and emphasized band were characterized for 289 high-quality tonal vocalizations with a minimum signal-to-noise ratio of 4.5 dB. African manatee vocalizations have a fundamental frequency of 4.65 ± 0.700 kHz (mean ± SD), duration of 0.181 ± 0.069 s, 97% contained harmonics, 21% contained subharmonics, and 27% had an emphasized band other than the fundamental frequency. Altogether, the structure of African manatee vocalizations is similar to other manatee species. We suggest utilizing passive acoustic monitoring to fill in the gaps in understanding the distribution and biology of African manatees.

Using Pooled Local Expert Opinions (PLEO) to Discern Patterns in Sightings of Live and Dead Manatees (Trichechus senegalensis, Link 1785) in Lower Sanaga Basin, Cameroon.

We aimed at unveiling patterns in live and dead manatee sightings in the Lower Sanaga Basin, Cameroon. For this purpose, the expert opinions of 133 local fishers were collected during in-person interviews, distilled using categorical data analysis, and checked against scientific literature. The five main results are as follows: manatees were sighted averagely once a week in lakes, rivers, and the coast & estuaries, mostly in group sizes of 2-3; the odds of sighting live manatees (respectively dead manatees) decreased (respectively increased) from inland lakes to estuaries and the coast, via rivers; manatee carcasses were reported in all habitats, albeit more frequently in rivers; a distribution map based on fishers' reports show two manatee concentration areas: Lake Ossa and the Malimba-Mbiako section of River Sanaga; the number of manatees was perceived as increasing despite incidental and directed catches. Thus, our findings corroborate earlier assessments of the Lower Sanaga Basin as being a major manatee conservation area. Additionally, from these results and the literature, we identified three hypotheses about local manatee persistence: deep pools such as lakes offer year round sanctuaries, not just dry-season refugia; seasonality of specific habitat variables determine manatee occurrence patterns; and local variability in habitat encroachment mediate the meta-population dynamics of manatee in the Lower Sanaga Basin. Finally, we examine the implications for data requirements in light of the small ecological scale at which the surveyed fishers ply their trade. Thus, consonant with the Malawi principles for the ecosystem approach to management (www.cbd.int/ecosystem), we recommend collecting data preferably at landscape scale, through a participatory monitoring program that fully integrates scientific and traditional knowledge systems. This program should include, amongst others, a standardised necropsy protocol for collecting mortality and biological data together with sonar and radio-telemetry technology to discern manatee use and movements between critical habitat components.