Species identification and genetic structure of threatened seahorses in Gran Canaria Island (Spain) using mitochondrial and microsatellite markers

A non-invasive DNA analysis of seahorse populations was carried out after extensive underwater surveys in Gran Canaria Island (Spain). In this geographical area, the presence of two species, Hippocampus hippocampus and H. guttulatus, has been previously reported. Sequencing of 16S ribosomal DNA (16S rDNA) was used for specific identification of live seahorses sampled in situ, as a previous step to evaluate genetic structure based on ten microsatellite markers. Phylogenetic analyses revealed the presence of a single species, H. hippocampus, in the seahorse communities found at Gran Canaria. No evidences of H. guttulatus or interspecific hybrids were found based on 16S rDNA and microsatellite data. The nuclear markers revealed low genetic diversity and lack of population structure across populations of Gran Canaria Island, with evidence of small population sizes. This study provides critical information to support conservation strategies of Gran Canaria seahorses.     

Seahorse (Hippocampinae) population fluctuations in the Ria Formosa Lagoon,south Portugal

Comparisons of three sets of surveys in the Ria Formosa Lagoon, Portugal, over a 13 year period (2001–2002, 2008–2009 and 2010–2013) revealed significant population fluctuations in at least one of the two seahorse (Hippocampinae) species living there, and that those fluctuations were potentially associated with habitat changes in the lagoon. After a significant decline between the first two survey periods (2001–2002 v. 2008–2009), long‐snouted seahorse Hippocampus guttulatus populations increased significantly between 2008–2009 surveys and new 2010–2013 surveys. There were no significant differences in H. guttulatus populations between the 2001–2002 and 2010–2013 surveys. In contrast, there were no significant differences in short‐snouted seahorse Hippocampus hippocampus densities among the 16 sites surveyed throughout the three sampling periods, although the ability to detect any change was hampered by the low densities of this species in all time periods. Fluctuations in H. guttulatus densities were positively correlated with the percentage of holdfast coverage, but with none of the other environmental variables tested. These results highlight the importance of holdfast availability in maintaining stable seahorse populations. While population fluctuations are certainly more promising than a consistent downward decline, such extreme fluctuations observed for seahorses in the Ria Formosa Lagoon could still leave these two species vulnerable to any additional stressors, particularly during low density periods.     

Identifying key environmental variables of two seahorse species (<Emphasis Type="Italic">Hippocampus guttulatus</Emphasis> and <Emphasis Type="Italic">Hippocampus hippocampus</Emphasis>) in the Ria Formosa lagoon,South Portugal

Recent findings reported a significant decrease in abundance of two seahorse species (Hippocampus guttulatus and H. hippocampus) in the Ria Formosa lagoon (South Portugal) and no direct causes have been, so far, clearly identified. This study aimed to describe fluctuations in the local seahorse populations through monthly surveys over a course of a year, in order to identify some of the potential drivers behind the seasonal fluctuations. A total of six sites were chosen based on their habitat characteristics. The highest H. guttulatus abundances were recorded at sites with higher holdfast availability and depth ranging from 3 to 6 m, while H. hippocampus were observed at highest numbers in sites with lower holdfast availability and patchy distribution. In most sites, seahorse density decreased during the summer months (from May to August) and increased from August to December. Holdfast use changed across the surveyed sites, according to the respective habitat characteristics. This study identified environmental variables that influenced the abundance of seahorse population, i.e., holdfast availability, depth and temperature in the Ria Formosa lagoon, underlining the importance of monitoring populations over a course of no less than a year in order to avoid bias due to seasonal fluctuations. Identifying critical habitats will provide valuable information for local authorities in order to implement protective measures towards seahorse conservation.     

Novel microsatellite loci in the threatened European long-snouted seahorse (<Emphasis Type="Italic">Hippocampus guttulatus</Emphasis>) for genetic diversity and parentage analysis

The long-snouted seahorse Hippocampus guttulatus is one of the two European seahorse species. We describe the isolation of the first 12 microsatellite loci in this threatened species. These new markers were tested in non-invasive samples of 32 seahorses from NW Spain. The number of alleles ranged from 2 to 15 (mean: 6.3) and expected heterozygosity from 0.031 to 0.912 (mean: 0.500). All loci conformed to Hardy–Weinberg expectations and no genotypic disequilibrium was observed between any pair of loci. The theoretical exclusion probabilities for this set of loci, when no parental information exists or when one parent is known, were 0.973 and 0.998, respectively. This study indicates the usefulness of these novel loci for population analysis and kinship studies in Hippocampus guttulatus. Their potential application is extended to the other European seahorse species, since all loci were successfully cross-amplified in H. hippocampus.     

An assessment of West African seahorses in fisheries catch and trade

This study provides the first assessment of a heavily traded West African seahorse species, Hippocampus algiricus, and the first information on short‐snouted seahorse Hippocampus hippocampus biology in Africa. A total of 219 seahorses were sampled from fisher catch in Senegal and The Gambia, with estimated height at reproductive activity for H. algiricus (161 mm) larger than mean ± s.d . catch height (150 ± 31 mm). Catch composition, height at reproductive activity and potential biases in fishery retention are discussed with regard to the current Convention on International Trade of Endangered Species (CITES) guidelines.     

Monthly population density and structure patterns of an endangered seahorse Hippocampus capensis: a comparison between natural and artificial habitats

This study investigated aspects of the population ecology of the endangered Knysna seahorse Hippocampus capensis within different habitat types. High densities of H. capensis were found within artificial Reno mattress habitat, within the Knysna Estuary, South Africa. Monthly surveys at three sites were conducted from October 2015 to August 2016 to compare population densities of H. capensis in this artificial habitat with natural eel grass Zostera capensis habitat. Hippocampus capensis densities varied significantly across all sites and highest population densities were consistently observed within the Reno mattress habitat. Hippocampus capensis were also found to be significantly larger within the Reno mattress habitat and pooled data showed that males were significantly larger than females. The overall sex ratio for all three sites was female biased, although this varied across seasons at two sites. The findings suggest that artificial Reno mattresses provide novel habitat for this endangered species and consideration should be given to the usefulness of these structures in future conservation actions.     

First records of Hippocampus algiricus in the Canary Islands (north‐east Atlantic Ocean) with an observation of hybridization with Hippocampus hippocampus

Morphometric and genetic analyses confirmed the first records of the West African seahorse Hippocampus algiricus at Gran Canaria Island (north‐east Atlantic Ocean), and also the first evidence of interspecific hybridization in seahorses. These results provide additional data on the distribution of H. algiricus that may help to establish future conservation strategies, and uncover a new potential sympatric scenario between H. algiricus and Hippocampus hippocampus.     

The endangered White's seahorse Hippocampus whitei chooses artificial over natural habitats

To explore whether the endangered White's seahorse Hippocampus whitei would choose to inhabit artificial over natural habitats, 10 adult H. whitei individuals were put through a series of binary choice trials in aquaria, during which they were offered different paired combinations of natural (different types of macroalga and seagrass) and artificial habitat (panels of swimming-net material). It was found that H. whitei displayed a significant choice for swimming-net material over all other available natural habitats and chose habitats according to the following rankings: (a) Net; (b) Sargassum sp.; (c) Posidonia australis; (d) Zostera muelleri. Hippocampus whitei’s choice of swimming net material over natural habitat suggests that these artificial structures could be a useful conservation measure for seahorses in areas where natural habitat is becoming less favourable due to declines in abundance or quality.     

An endangered seahorse selectively chooses an artificial structure

The development of a residential marina estate within the Knysna estuary, South Africa, introduced Reno mattresses (horizontal wire cages filled with rocks) as a novel habitat for the endangered Knysna seahorse Hippocampus capensis. Consistently high seahorse densities on these artificial structures, despite the availability of seagrass habitat, begged the question of whether this habitat was chosen by the seahorse in preference to natural vegetation. An in situ habitat choice experiment was conducted which focused on the choice made by adult H. capensis between natural vegetation (Zostera capensis) and artificial (Reno mattress) habitat within a choice chamber. Seahorses were significantly more likely to move away from Z. capensis onto a Reno mattress structure or remain on this structure. This study concludes that higher H. capensis densities on Reno mattresses within Thesen Islands Marina are owing to some positive feature of this habitat and the underlying processes responsible for the choice made by this species (additional food, holdfasts, protection) can now be investigated.     

Population dynamics of the long-snouted seahorse (Hippocampus guttulatus Cuvier, 1829) in the Mar Menor coastal lagoon

A 10-year monitoring program was developed to quantify the population dynamics of the long-snouted seahorse population in the Mar Menor coastal lagoon. Based on 985 underwater visual censuses, we estimated the long-snouted seahorse (Hippocampus guttulatus Cuvier, 1829) population size in the Mar Menor lagoon and its reduction in size in the last decades, as well as the effect of eutrophication crises in 2016 and 2019 on the species. The annual recruitment for the 2013–2020 period was estimated by comparing the relative abundance of early seahorse life stages in the ichthyoplankton. The density ranged from 0.0458 specimens/m³ at the beginning of the sampling period to 0.0004 at the end, showing a statistically significant difference between the three analyzed periods (H_gl=2 = 14.0, p = 0.001). The long-snouted seahorse population from the Mar Menor lagoon exemplifies the impact of fishing activities and human pressure, especially euxinic episodes and habitat destruction. As a result of this, the Mar Menor population has decreased from several million specimens to a few thousand, in only three decades. This species showed considerable resilience, the seahorse population began to recover once fishing activity stopped. In contrast, the long-snouted seahorse showed high vulnerability to habitat loss and an episodic flooding event. Adult seahorses showed preferences for highly complex habitats, especially Caulerpa prolifera–Cymodocea nodosa mixed meadows and habitats of high complexity and anthropogenic origin, such as harbors, jetties, or breakwaters. In contrast, juvenile seahorses preferred monotonous seabeds with low complexity, such as the sandy beds that are characteristic of the Mar Menor lagoon littoral.     

Life history of an unusual marine fish: survival, growth and movement patterns of Hippocampus guttulatus Cuvier 1829

The life history of the long‐snouted seahorse Hippocampus guttulatus was characterized using mark‐recapture data collected within a focal study site and catch data from 53 additional sites in the Ria Formosa coastal lagoon, southern Portugal. Population structure in benthic habitats was characterized by high local densities (0·3–1·5 m^?2), equal sex ratios and few juveniles <70 mm. Adult H. guttulatus maintained small (19·9 ± 12·4 m²), strongly overlapping home ranges during multiple reproductive seasons. Recruited (benthic) juveniles exhibited significantly lower site fidelity than adults. A Ford‐Walford plot of standard length (L_S) at time t against L_S measured during the previous year from tagged juveniles and adults led to estimates of the von Bertalanffy parameters K = 0·571 and L_∞ = 197·6 mm. The growth rate of planktonic juveniles (inferred from previous studies), was greater than predicted by the von Bertalanffy model, providing evidence of an ontogenetic shift in growth trajectory. The instantaneous rate of natural mortality, M, ranged from 1·13 to 1·22 year^?1(annual survival rate = 29·4–32·2%). Sexes did not differ in movement, growth or survival patterns. On average, H. guttulatus measured 12·2 ± 0·8 mm at birth. Planktonic juveniles recruited to vegetated habitat at 96·0 ± 8·0 mm (0·25 years), had mature brood pouches (males only) at 109·4 mm (0·49 years), began maintaining home ranges and reproducing at 125–129 mm (0·85–0·94 years), and lived for 4·3–5·5 years. Early age at maturity, rapid growth rates, and short generation times suggested that H. guttulatus may recover rapidly when direct (e.g. exploitation) and indirect (e.g. by‐catch and habitat damage) effects of disturbance cease, but may be vulnerable to extended periods of poor recruitment.     

Serial monogamy in the European long-snouted seahorse Hippocampus guttulatus

Seahorses (Hippocampus spp.) are non-sex-role-reversed members of the Syngnathidae family that provide extensive brood care. Previous studies of seahorses have revealed monogamy within a single brood, but their longer term mating system had not been comprehensively evaluated. The parental contribution to 29 wild-born broods of Hippocampus guttulatus, sampled from six Portuguese populations with differing seahorse densities and sex ratios, was assessed using microsatellite DNA markers. To assess the longer term genetic mating system of this species parentage was determined in eleven broods sampled from a captive population over two breeding seasons. Genetic data suggest that this socially polygamous seahorse is serially monogamous across breeding seasons, i.e. monogamous within a season but may switch mates between seasons, and that differing population densities and sex ratios do not affect the mating system.     

First record of the Pacific seahorse Hippocampus ingens in Guadalupe Island,Mexico

This is the first record of the Pacific seahorse Hippocampus ingens at a northern oceanic island from the eastern Pacific Ocean. The photographic record of the juvenile female H. ingens was made in November 2015 during a cage diving trip at Guadalupe Island, Mexico. The presence of H. ingens in this area could be related to long distance dispersal mechanisms, as has been observed in other species of seahorses.     

Conservation value of Codium tenue habitat for the endangered Knysna seahorse Hippocampus capensis

We investigated the density and size structure of the endangered Knysna seahorse Hippocampus capensis in the macroalga Codium tenue at three sites in a residential marina estate in the Knysna Estuary, South Africa, over an 18 month period from March 2017 to August 2018. Seahorses were present in C. tenue throughout the year. Average (± SE) densities of 0.33 (± 0.03) and 0.23 (± 0.03) seahorses per kg of C. tenue were found for 2017 and 2018 respectively. Density did not vary across sites but varied significantly across months with a decreasing trend from summer to spring. The average (± SE) adult seahorse height was 75.16 mm (± 0.63 mm) and 69.09 mm (± 0.64 mm) and mass was 2.05 g (± 0.04 g) and 1.65 g (± 0.04 g) for males and females, respectively, and an even sex ratio was observed throughout the sampling period. Height varied across months, with significantly smaller seahorses found in June and August of 2017. Tail length was highly correlated with height and therefore can serve as a potential proxy for determining the height of H. capensis. This study showed that C. tenue is consistently utilised by Knysna seahorse and should therefore be considered an important habitat to protect and facilitate the ongoing conservation of this endangered seahorse species.     

Life history and population dynamics of an endangered seahorse (Hippocampus capensis) within an artificial habitat

Species-specific life-history information is critical for successful conservation, particularly in establishing an accurate baseline status. Obtaining such information is challenging for most species, but in particular for rare and threatened marine species. To facilitate future conservation of the endangered Knysna seahorse (Hippocampus capensis) this study aimed to determine important life-history information for this species. Visible implant fluorescent elastomer (VIFE) tags were used to mark 78 seahorses within a residential marina estate in the Knysna estuary, South Africa, in February 2018. Using a mark-resight approach, the size and movement patterns of the population and growth rate of seahorses were determined over a 14-month period. The closed population estimate for H. capensis, within Thesen Islands Marina, was estimated to be 134 (118–152 95% C.I. ) in February 2018 compared to only 72 (48–108 95% C.I. ) in February 2019. The species showed rapid initial growth with males and females having similar rates of growth based on the specialised von Bertalanffy growth function model. The importance of Reno mattresses as a habitat for H. capensis was confirmed based on the high abundance and site fidelity of the population, which emphasises the conservation potential of heavily modified environments for threatened seahorse species. The use of VIFE tags was deemed effective in studying this endangered seahorse and allowed the collection of important information for this species which can be used in future Red List assessments and conservation actions.     

The use of a non‐invasive tool for capture–recapture studies on a seahorse Hippocampus guttulatus population

In this study, the spot pattern in Hippocampus guttulatus was analysed using a computer programme algorithm that allowed individual comparison. This methodology was first tested in a controlled environment using 51 adult and 55 juvenile H. guttulatus. Positive matches were obtained in 86·3 and 83·6% of the adults and juveniles, respectively. In a second experiment, monthly surveys were carried out in five selected locations in the Ria Formosa Lagoon, south Portugal, over the course of a year and a total of 980 photographs were analysed. Photographed H. guttulatus were re‐sighted one to nine times during the course of the survey period with an overall re‐sight record of over 30%. Photo‐identification was therefore shown to be a useful tool for non‐invasive mark–recapture studies that can be successfully used to survey the population abundance of H. guttulatus aged 6 months or older in consecutive years. This could be of great value when considering the assessment of H. guttulatus populations and understanding changes over time.     

Life history and ecology of the elusive European short‐snouted seahorse Hippocampus hippocampus

To improve the understanding of the life history and ecology of one of Europe's most elusive fishes, the short‐snouted seahorse Hippocampus hippocampus, data from wild populations in a shallow coastal lagoon in southern Portugal were analysed. The data were collected from 17 tagged seahorses on a focal‐study grid as well as from >350 seahorses encountered during underwater visual surveys and a fishery‐independent study using beach seines. These populations of settled juveniles and adults had a mean population density of 0·009 m^?2. During the study period (2000–2004), reproduction peaked in July and August. Juveniles recruited to the lagoon at c. 66 mm standard length (L_S) and 0·5 years of age and established small home ranges (0·8 to 18·2 m²). First reproduction was estimated at 100 mm and 1 year of age. Based on a fitted von Bertalanffy model, H. hippocampus grew quickly (growth coefficient K = 0·93) to a maximum theoretical size L_∞ = 150 mm and have a maximum lifespan of c. 3·2 years. Courtship behaviours were consistent with the maintenance of pair bonds and males brooded multiple batches of young per year. Estimated annual reproductive output averaged 871 young (±632). Together these analyses provide the first life‐history parameters for this species and indicate that H. hippocampus bears characteristics of opportunist and intermediate strategists. Such populations are predicted to exhibit large fluctuations in abundance, making them vulnerable to extended periods of poor recruitment.     

New records of the Japanese seahorse Hippocampus mohnikei in Southeast Asia lead to updates in range,habitat and threats

New records of the Japanese seahorse Hippocampus mohnikei from Cambodia, Malaysia, Thailand and Vietnam, along with recently published studies from India and Singapore, have greatly expanded the known range of H. mohnikei within Southeast Asia. These new records reveal novel habitat preferences and threats to H. mohnikei in the region. Although the global conservation status of H. mohnikei is classified as Data Deficient according to the IUCN Red List of Threatened Species, new sightings indicate that this species is found in similar habitats and faces similar threats as other Hippocampus species that are considered Vulnerable.     

Ontogenetic and sex‐based differences in habitat preferences and site fidelity of White's seahorse Hippocampus whitei

The aim of this study was to determine and compare habitat preferences for male and female adult and juvenile White's seahorse Hippocampus whitei and assess their movements and site fidelity over 4 years. Data were collected from three sites along 1·5 km of estuarine shoreline in Port Stephens, New South Wales, Australia, from 2006 to 2009 using H. whitei that had been tagged with visible implant fluorescent elastomer. Relative availability of 12 habitats and habitat preferences of H. whitei was determined, based on the habitat that H. whitei used as a holdfast. Hippocampus whitei occurred in nine different habitats; adults preferred sponge and soft coral Dendronephthya australis habitats with no difference between male and female habitat preferences whilst juveniles preferred gorgonian Euplexaura sp. habitat. There was a significant preference by adults for D. australis colonies with height >40 cm and avoidance of colonies <20 cm. Neither adults nor juveniles used sand or the seagrasses Zostera muelleri subsp. capricorni and Halophila ovalis. Hippocampus whitei showed cryptic behaviour with c. 50% of adult sightings cryptic and c. 75% for juveniles with crypsis occurring predominantly in Sargassum sp. for adults and Euplexaura sp. habitat for juveniles. Within sites, females moved significantly longer distances (maximum of 70 m) than males (maximum of 38 m) over 20 months. Strong site fidelity was displayed by H. whitei with males persisting at the same site for up to 56 months and females for 49 months and no H. whitei moved between sites. The longest period that an H. whitei was recorded on the same holdfast was 17 months for a male and 10 months for a female. As this species displays strong site fidelity, specific habitat preferences and has a limited distribution, future management needs to minimize the risk of habitat disturbance as loss of key habitats could have a negative effect on species abundance and distribution.     

Specific authentication of Hippocampus based on SNP markers

Seahorses, which are used as traditional Chinese medicine, have large market demands and premium price. Because of overfishing, many seahorse species are endangered and listed in IUCN Red List. The purpose of this study was to develop a simple, rapid and reliable molecular method for specific authentication of seahorse species. Cytochrome c oxidase subunit I (COI) sequences were analyzed for molecular authentication of seahorses in this study. Many single nucleotide polymorphism (SNP) sites were detected among COI sequences of nine seahorse species. Based on the SNP sites, four pairs of modified specific primers were designed respectively for authentication of Hippocampus kuda, Hippocampus japonicus, Hippocampus trimaculatus and Hippocampus kelloggi using allele-specific PCR. Moreover, a multiplex allele-specific PCR for simultaneous authentication of H. kuda, H. japonicus and H. trimaculatus was also developed. The study showed that allele-specific PCR required no sequencing and electrophoresis, and the only necessity is to add fluorochrome into PCR products. Hence the whole authentication procedure only takes about 1.5 h. This work provides an effective method for rapid and reliable identification of seahorse species.