Southward range extension and transboundary movements of reef manta rays Mobula alfredi along the east African coastline.

Opportunistic in-water and aerial surveys in South Africa and the analysis of contributed citizen science data have extended the known range of reef manta rays Mobula alfredi along the eastern coast of Africa by 140 km (to Mdumbi Beach) and verified the first transboundary movements for the species. Additionally, six new long-range dispersal records have provided evidence of connectivity with the M. alfredi population off the Inhambane coastline of Mozambique. Five of these records captured one-way movements, the longest of which was an individual that travelled 505 km from Praia do Tofo to Sodwana Bay with 301 days between sightings. A single individual made a return trip between Závora, Mozambique and Sodwana Bay, South Africa (a total distance of ~870 km). These findings support the Convention on Migratory Species listing for the species, suggesting regional transboundary management units are warranted for this wide-ranging elasmobranch.

Residency, movement patterns, behavior and demographics of reef manta rays in Komodo National Park.

Background: The reef manta ray (Mobula alfredi) is a globally threatened species and an iconic tourist attraction for visitors to Indonesia's Komodo National Park (NP). In 2013, manta ray fishing was banned in Komodo NP and its surroundings, preceding the nationwide manta ray protection in 2014. Over a decade ago, a previous acoustic telemetry study demonstrated that reef manta rays had high fidelity to sites within the park, while more recent photo-identification data indicated that some individuals move up to 450 km elsewhere. Characterization of manta ray demographics, behavior, and a focused assessment on site use of popular tourism locations within the park is vital to assist the Komodo NP Management Authority formulate appropriate manta ray conservation and management policies. Methods: This study uses a long-term library (MantaMatcher.org) of photo-identification data collected by researchers and citizen scientists to investigate manta ray demographics and habitat use within the park at four sites frequented by tour operators: Cauldron, Karang Makassar, Mawan, and Manta Alley. Residency and movements of manta rays were investigated with maximum likelihood analyses and Markov movement models. Results: A total of 1,085 individual manta rays were identified from photographs dating from 2013 to 2018. In general, individual manta rays displayed a higher affinity to specific sites than others. The highest re-sighting probabilities came from the remote southern site, Manta Alley. Karang Makassar and Mawan are only ~5 km apart; however, manta rays displayed distinct site affinities. Exchange of individuals between Manta Alley and the two central sites (~35.5 km apart) occurred, particularly seasonally. More manta rays were recorded traveling from the south to the central area than vice versa. Female manta rays were more mobile than males. Similar demographic groups used Karang Makassar, Mawan, and Manta Alley for foraging, cleaning, cruising, or courtship activities. Conversely, a higher proportion of immature manta rays used the northern site, Cauldron, where foraging was commonly observed. Fishing gear-related injuries were noted on 56 individuals (~5%), and predatory injuries were present on 32 individuals (~3%). Tourism within the park increased from 2014 to 2017, with 34% more dive boats per survey at Karang Makassar and Mawan. Discussion: The Komodo NP contains several distinct critical habitats for manta rays that encompass all demographics and accommodate seasonal manta ray movements. While the present study has not examined population trends, it does provide foundational data for such work. Continued research into manta ray abundance, long-range movements, and identifying and protecting other critical aggregation areas within the region is integral to securing the species' recovery. We provide management recommendations to limit undue pressure on manta rays and their critical habitats from tourism.

Half a century of global decline in oceanic sharks and rays.

Overfishing is the primary cause of marine defaunation, yet declines in and increasing extinction risks of individual species are difficult to measure, particularly for the largest predators found in the high seas1-3. Here we calculate two well-established indicators to track progress towards Aichi Biodiversity Targets and Sustainable Development Goals4,5: the Living Planet Index (a measure of changes in abundance aggregated from 57 abundance time-series datasets for 18 oceanic shark and ray species) and the Red List Index (a measure of change in extinction risk calculated for all 31 oceanic species of sharks and rays). We find that, since 1970, the global abundance of oceanic sharks and rays has declined by 71% owing to an 18-fold increase in relative fishing pressure. This depletion has increased the global extinction risk to the point at which three-quarters of the species comprising this functionally important assemblage are threatened with extinction. Strict prohibitions and precautionary science-based catch limits are urgently needed to avert population collapse6,7, avoid the disruption of ecological functions and promote species recovery8,9.

Genome-wide SNPs detect no evidence of genetic population structure for reef manta rays (Mobula alfredi) in southern Mozambique.

Little is known about the extent of genetic connectivity along continuous coastlines in manta rays, or whether site visitation is influenced by relatedness. Such information is pertinent to defining population boundaries and understanding localized dispersal patterns and behaviour. Here, we use 3057 genome-wide single-nucleotide polymorphisms (SNPs) to evaluate population genetic structure and assess the levels of relatedness at aggregation sites of reef manta rays (Mobula alfredi) in southern Mozambique (n = 114). Contrary to indications of limited dispersal along the southern Mozambican coastline inferred from photo-identification and telemetry studies, our results show no evidence of population structure (non-significant FST < 0.001) for M. alfredi along this coast. We also found no evidence that individuals sampled at the same site were more related than expected by chance for males, females or across both sexes, suggesting that kinship may not influence visitation patterns at these sites. We estimated the effective population size (Ne) of this population to be 375 (95% CI = 369-380). Comparison to a distant eastern Indian Ocean site (Western Australia, n = 15) revealed strong genetic differentiation between Mozambique and Western Australia (FST = 0.377), identifying the Indian Ocean basin as a barrier to dispersal. Our findings show that genetic connectivity in M. alfredi extends for several hundred kilometres along continuous coastlines. We therefore recommend that the population in Mozambique be considered a discrete management unit, and future conservation plans should prioritize integrated strategies along the entire southern coastline.

It's not all black and white: investigating colour polymorphism in manta rays across Indo-Pacific populations.

Intraspecific colour polymorphisms have been the focus of numerous studies, yet processes affecting melanism in the marine environment remain poorly understood. Arguably, the most prominent example of melanism in marine species occurs in manta rays (Mobula birostris and Mobula alfredi). Here, we use long-term photo identification catalogues to document the frequency variation of melanism across Indo-Pacific manta ray populations and test for evidence of selection by predation acting on colour morph variants. We use mark-recapture modelling to compare survivorship of typical and melanistic colour morphs in three M. alfredi populations and assess the relationship between frequency variation and geographical distance. While there were large differences in melanism frequencies among populations of both species (0-40.70%), apparent survival estimates revealed no difference in survivorship between colour morphs. We found a significant association between phenotypic and geographical distance in M. birostris, but not in M. alfredi. Our results suggest that melanism is not under selection by predation in the tested M. alfredi populations, and that frequency differences across populations of both species are a consequence of neutral genetic processes. As genetic colour polymorphisms are often subjected to complex selection mechanisms, our findings only begin to elucidate the underlying evolutionary processes responsible for the maintenance and frequency variation of melanism in manta ray populations.

Spotting the "small eyes": using photo-ID methodology to study a wild population of smalleye stingrays (Megatrygon microps) in southern Mozambique.

BACKGROUND: The smalleye stingray (Megatrygon microps) is a large and rare dasyatid ray, patchily distributed across the Indo-West Pacific. Free-swimming individuals have regularly been recorded in Southern Mozambican coastal waters utilizing different inshore environments. Distinctive features of the species include latitudinal rows of white spots on the dorsal surface of their pectoral disc. METHODS: This study aimed to determine if the natural spot patterns on M. microps are sufficiently unique and stable to use in photo-identification studies of wild populations. Research dive logs were combined with opportunistic photographs from local dive centers and recreational divers to create a photographic database from the Inhambane Province coastline. RESULTS: Seventy different individuals were identified over a 15-year period, all exhibiting uniquely identifiable patterns. Stingrays were easily identifiable over a period of six years with multiple re-sightings of the same individuals recorded. Analysis of encounters across the Inhambane coastline revealed that individual rays regularly use inshore reefs along a 350 km stretch of coastline. Fifteen stingrays were re-sighted during the study period, including one showing a 400 km return movement between Tofo Beach and the Bazaruto Archipelago, which is the longest distance traveled by a dasyatid ray on record. Several presumably pregnant females have also been recorded in the Bazaruto Archipelago National Park.

Microplastics: No Small Problem for Filter-Feeding Megafauna.

Microplastic pollution can impact filter-feeding marine megafauna, namely mobulid rays, filter-feeding sharks, and baleen whales. Emerging research on these flagship species highlights potential exposure to microplastic contamination and plastic-associated toxins. Research and its wide communication are needed to understand the magnitude of the issue and improve marine stewardship.

Novel signature fatty acid profile of the giant manta ray suggests reliance on an uncharacterised mesopelagic food source low in polyunsaturated fatty acids.

Traditionally, large planktivorous elasmobranchs have been thought to predominantly feed on surface zooplankton during daytime hours. However, the recent application of molecular methods to examine long-term assimilated diets, has revealed that these species likely gain the majority from deeper or demersal sources. Signature fatty acid analysis (FA) of muscle tissue was used to examine the assimilated diet of the giant manta ray Mobula birostris, and then compared with surface zooplankton that was collected during feeding and non-feeding events at two aggregation sites off mainland Ecuador. The FA profiles of M. birostris and surface zooplankton were markedly different apart from similar proportions of arachidonic acid, which suggests daytime surface zooplankton may comprise a small amount of dietary intake for M. birostris. The FA profile of M. birostris muscle was found to be depleted in polyunsaturated fatty acids, and instead comprised high proportions of 18:1ω9 isomers. While 18:1ω9 isomers are not explicitly considered dietary FAs, they are commonly found in high proportions in deep-sea organisms, including elasmobranch species. Overall, the FA profile of M. birostris suggests a diet that is mesopelagic in origin, but many mesopelagic zooplankton species also vertically migrate, staying deep during the day and moving to shallower waters at night. Here, signature FA analysis is unable to resolve the depth at which these putative dietary items were consumed and how availability of this prey may drive distribution and movements of this large filter-feeder.

Manta birostris, predator of the deep? Insight into the diet of the giant manta ray through stable isotope analysis.

The characterization of diet for the giant manta ray Manta birostris has been problematic given their large-scale movement patterns and the difficulty in obtaining stomach contents from this species. The large majority of existing information is based on observational data limited to feeding events at the sea surface during daylight. Recently discovered aggregation sites for the giant manta ray off mainland Ecuador are some of the most accessible to date and provide a unique opportunity for researchers to gather much needed information on this elusive species. To assess how important surface zooplankton is to giant manta ray diet, we conducted stable isotope analysis (15N and 13C) on M. birostris muscle and surface zooplankton. Trophic position estimates placed M. birostris overall at a secondary consumer level of approximately 3.4 but there was large variation in δ15N and δ13C values among individuals. Manta birostris muscle tissue δ13C values were also not consistent with this species feeding predominantly on surface zooplankton and suggest that the majority of dietary intake is of mesopelagic origin. Given the conservative life history and fisheries pressure on large planktivores, knowledge of their trophic role and foraging strategies is essential to better understand their ecology and develop effective conservation measures.

Evaluating manta ray mucus as an alternative DNA source for population genetics study: underwater-sampling, dry-storage and PCR success.

Sharks and rays are increasingly being identified as high-risk species for extinction, prompting urgent assessments of their local or regional populations. Advanced genetic analyses can contribute relevant information on effective population size and connectivity among populations although acquiring sufficient regional sample sizes can be challenging. DNA is typically amplified from tissue samples which are collected by hand spears with modified biopsy punch tips. This technique is not always popular due mainly to a perception that invasive sampling might harm the rays, change their behaviour, or have a negative impact on tourism. To explore alternative methods, we evaluated the yields and PCR success of DNA template prepared from the manta ray mucus collected underwater and captured and stored on a Whatman FTA™ Elute card. The pilot study demonstrated that mucus can be effectively collected underwater using toothbrush. DNA stored on cards was found to be reliable for PCR-based population genetics studies. We successfully amplified mtDNA ND5, nuclear DNA RAG1, and microsatellite loci for all samples and confirmed sequences and genotypes being those of target species. As the yields of DNA with the tested method were low, further improvements are desirable for assays that may require larger amounts of DNA, such as population genomic studies using emerging next-gen sequencing.

Laser photogrammetry improves size and demographic estimates for whale sharks.

Whale sharks Rhincodon typus are globally threatened, but a lack of biological and demographic information hampers an accurate assessment of their vulnerability to further decline or capacity to recover. We used laser photogrammetry at two aggregation sites to obtain more accurate size estimates of free-swimming whale sharks compared to visual estimates, allowing improved estimates of biological parameters. Individual whale sharks ranged from 432-917 cm total length (TL) (mean ± SD = 673 ± 118.8 cm, N = 122) in southern Mozambique and from 420-990 cm TL (mean ± SD = 641 ± 133 cm, N = 46) in Tanzania. By combining measurements of stranded individuals with photogrammetry measurements of free-swimming sharks, we calculated length at 50% maturity for males in Mozambique at 916 cm TL. Repeat measurements of individual whale sharks measured over periods from 347-1,068 days yielded implausible growth rates, suggesting that the growth increment over this period was not large enough to be detected using laser photogrammetry, and that the method is best applied to estimating growth rates over longer (decadal) time periods. The sex ratio of both populations was biased towards males (74% in Mozambique, 89% in Tanzania), the majority of which were immature (98% in Mozambique, 94% in Tanzania). The population structure for these two aggregations was similar to most other documented whale shark aggregations around the world. Information on small (<400 cm) whale sharks, mature individuals, and females in this region is lacking, but necessary to inform conservation initiatives for this globally threatened species.

Running the gauntlet: regional movement patterns of Manta alfredi through a complex of parks and fisheries.

Manta rays (Genus Manta) are economically important for fisheries and tourism in Indonesia. These species have been listed by the International Union for the Conservation of Nature Red List as Vulnerable to extinction; therefore, human exploitation of manta rays must be regulated. A better understanding of the habitat use and movement patterns of manta rays in Indonesia is needed in order to employ effective conservation measures. To gain better insight into the movements of Manta alfredi we used 'Manta Matcher', an online database with an integrated automated matching algorithm, to compare photographs from 2,604 encounters of M. alfredi collected by recreational divers and dive operators throughout Indonesia over a nine-year period. This photographic comparison revealed that manta rays migrated between regional sanctuaries such as Nusa Penida, the Gili Islands, and the Komodo National Park (up to 450 km straight-line distance). The areas between these sanctuaries are heavily fished and trafficked by ships, and when manta rays travel through these regions they risk being fished and injured by ship strikes. These long-range manta ray movements suggest connectivity between M. alfredi populations in neighboring islands and raise concerns about the future management of regional populations. It is recommended that a national conservation strategy be developed to protect the remaining populations in the country.

Stable isotope and signature fatty acid analyses suggest reef manta rays feed on demersal zooplankton.

Assessing the trophic role and interaction of an animal is key to understanding its general ecology and dynamics. Conventional techniques used to elucidate diet, such as stomach content analysis, are not suitable for large threatened marine species. Non-lethal sampling combined with biochemical methods provides a practical alternative for investigating the feeding ecology of these species. Stable isotope and signature fatty acid analyses of muscle tissue were used for the first time to examine assimilated diet of the reef manta ray Manta alfredi, and were compared with different zooplankton functional groups (i.e. near-surface zooplankton collected during manta ray feeding events and non-feeding periods, epipelagic zooplankton, demersal zooplankton and several different zooplankton taxa). Stable isotope δ(15)N values confirmed that the reef manta ray is a secondary consumer. This species had relatively high levels of docosahexaenoic acid (DHA) indicating a flagellate-based food source in the diet, which likely reflects feeding on DHA-rich near-surface and epipelagic zooplankton. However, high levels of ω6 polyunsaturated fatty acids and slightly enriched δ(13)C values in reef manta ray tissue suggest that they do not feed solely on pelagic zooplankton, but rather obtain part of their diet from another origin. The closest match was with demersal zooplankton, suggesting it is an important component of the reef manta ray diet. The ability to feed on demersal zooplankton is likely linked to the horizontal and vertical movement patterns of this giant planktivore. These new insights into the habitat use and feeding ecology of the reef manta ray will assist in the effective evaluation of its conservation needs.

The genetic signature of recent speciation in manta rays (Manta alfredi and M. birostris).

Manta rays have been taxonomically revised as two species, Manta alfredi and M. birostris, on the basis of morphological and meristic data, yet the two species occur in extensive mosaic sympatry. We analysed the genetic signatures of the species boundary using a portion of the nuclear RAG1 (681 base pairs), mitochondrial CO1 (574 bp) and ND5 genes (1188 bp). The assay with CO1 sequences, widely used in DNA barcoding, failed to distinguish the two species. The two species were clearly distinguishable, however, with no shared RAG1 or ND5 haplotypes. The species were reciprocally monophyletic for RAG1, but paraphyletic for ND5 sequences. Qualitative evidence and statistical inferences using the 'Isolation-with-Migration models' indicated that these results were better explained with post-divergence gene flow in the recent past rather than incomplete lineage sorting with zero gene flow since speciation. An estimate of divergence time was less than 0.5 Ma with an upper confidence limit of within 1 Ma. Recent speciation of highly mobile species in the marine environment is of great interest, as it suggests that speciation may have occurred in the absence of long-term physical barriers to gene flow. We propose that the ecologically driven forces such as habitat choice played a significant role in speciation in manta rays.