Reply to "Letter to the editor regarding the article 'Taking advantage of seagrass recovery potential to develop novel and effective meadow rehabilitation methods' by Alagna et al., published in Marine Pollution Bulletin, 149: 2019 (110578)" by Calvo et al. Marine Pollution Bulletin, 158:2020 (111395).

Calvo et al. (2020) criticize a new seagrass rehabilitation method proposed by Alagna et al. (2019) and inspired by the Posidonia oceanica spontaneous recovery observed at Capo Feto (Sicily), were recolonization was detected almost exclusively on rubbles deployed to fill a pipeline trench. Calvo et al. (2020) claim that natural recovery occurred consistently also on dead matte along the eastern side of the trench, weakening the assumption on which the method is based. Here we show that the P. oceanica patches reported by these authors as new establishments were already documented in 2003 (Vega Fernandez et al., 2005) and are attributable to the fragmentation of the pristine meadow caused by altered sedimentation rate after an extensive dredging operation. Moreover, we outline the area of applicability of the method tested in Alagna et al. (2019) and provide a point-by-point rebuttal to the complaints of imprecise and misleading contents of the paper.

The perfect microsite: How to maximize Posidonia oceanica seedling settlement success for restoration purposes using ecological knowledge.

The use of sexual propagules to restore seagrass meadows has raised increasing attention in the last years as seed-based strategies avoid impacts on donor beds while preserving genetic diversity in restored populations. However, the availability of suitable microsites for seedling establishment at transplantation locations is crucial in order to achieve positive outcome of restoration actions. In this study we develop ad-hoc holders that act as optimal microsites for Posidonia oceanica seedling establishment. Holders are intended to be transferred in the field for restoration purposes after few months of indoor seedling culture. Seedling ability to self-anchor to rocky substrates via adhesive root hairs was exploited. We tested rocky holders with different designs in order to maximize seedling survival and settlement. The effect of the holder design on seedling anchorage performances was evaluated. Holders were provided with different topographical complexity and substrate slope. Topographical complexity significantly influenced settlement success, as seedlings did not attach to flat holders, while anchorage reached 100% on holders provided with complexity at seed and the root scales. Substrate slope did not affect the percentage of anchored seedlings, conversely it influenced root growth pattern and thus anchorage stability. This study shows how ecological knowledge of species' life history strategies and associated critical traits provides valuable hints to develop alternative approaches to seagrass restoration tailored to the biology of the system under study.

Taking advantage of seagrass recovery potential to develop novel and effective meadow rehabilitation methods.

Seagrasses are among the most threatened biomes worldwide. Until now, seagrass rehabilitation success has reached about 38% overall and more effective approaches to restoration are urgently needed. Here we report a novel method to rehabilitate Posidonia oceanica meadows based on observation of the species' natural recovery after disturbance. Posidonia oceanica rhizomes were transplanted on gabions filled with rocks of selected sizes in order to build a firm substrate with topographic complexity in the relevant scale range to propagules. Five techniques were tested, each involving a different anchoring device. The "slot" technique, which uses a wire-net pocket to retain the cuttings, was the most successful, with survival exceeding 85% after thirty months. Branching allowed final shoot survival to reach 422% of initial planting density. This study shows how an in-depth knowledge of species life history processes provides a suitable foundation for developing effective restoration methods that benefit from species recovery ability.

Anthropogenic impact is negatively related to coral health in Sicily (Mediterranean Sea).

Shallow-water marine organisms are among the first to suffer from combined effects of natural and anthropogenic drivers. The orange coral Astroides calycularis is a shallow-water bioconstructor species endemic to the Mediterranean Sea. Although raising conservation interest, also given its special position within the Dendrophylliidae, information about the threats to its health is scant. We investigated the health status of A. calycularis at five locations in northwestern Sicily along a gradient of cumulative human impact and the most probable origin of the threats to this species, including anthropogenic land-based and sea-based threats. Cumulative human impact appeared inversely related to the performance of A. calycularis at population, colony, and polyp levels. Sea-based human impacts appeared among the most likely causes of the variation observed. The reduction in polyp length can limit the reproductive performance of A. calycularis, while the decrease of percent cover and colony area is expected to impair its peculiar feeding behaviour by limiting the exploitable dimensional range of prey and, ultimately, reef functioning. This endangered habitat-forming species appeared susceptible to anthropogenic pressures, suggesting the need to re-assess its vulnerability status. Creating microprotected areas with specific restrictions to sea-based human impacts could be the best practice preserve these bioconstructions.

Seagrass meadows (Posidonia oceanica) distribution and trajectories of change.

Posidonia oceanica meadows are declining at alarming rates due to climate change and human activities. Although P. oceanica is considered the most important and well-studied seagrass species of the Mediterranean Sea, to date there has been a limited effort to combine all the spatial information available and provide a complete distribution of meadows across the basin. The aim of this work is to provide a fine-scale assessment of (i) the current and historical known distribution of P. oceanica, (ii) the total area of meadows and (iii) the magnitude of regressive phenomena in the last decades. The outcomes showed the current spatial distribution of P. oceanica, covering a known area of 1,224,707 ha, and highlighted the lack of relevant data in part of the basin (21,471 linear km of coastline). The estimated regression of meadows amounted to 34% in the last 50 years, showing that this generalised phenomenon had to be mainly ascribed to cumulative effects of multiple local stressors. Our results highlighted the importance of enforcing surveys to assess the status and prioritize areas where cost-effective schemes for threats reduction, capable of reversing present patterns of change and ensuring P. oceanica persistence at Mediterranean scale, could be implemented.

Assessing Posidonia oceanica seedling substrate preference: an experimental determination of seedling anchorage success in rocky vs. sandy substrates.

In the last decades the growing awareness of the ecological importance of seagrass meadows has prompted increasing efforts to protect existing beds and restore degraded habitats. An in-depth knowledge of factors acting as major drivers of propagule settlement and recruitment is required in order to understand patterns of seagrass colonization and recovery and to inform appropriate management and conservation strategies. In this work Posidonia oceanica seedlings were reared for five months in a land-based culture facility under simulated natural hydrodynamic conditions to identify suitable substrates for seedling anchorage. Two main substrate features were investigated: firmness (i.e., sand vs. rock) and complexity (i.e., size of interstitial spaces between rocks). Seedlings were successfully grown in culture tanks, obtaining overall seedling survival of 93%. Anchorage was strongly influenced by substrate firmness and took place only on rocks, where it was as high as 89%. Anchorage occurred through adhesion by sticky root hairs. The minimum force required to dislodge plantlets attached to rocky substrates reached 23.830 N (equivalent to 2.43 kg), which would potentially allow many plantlets to overcome winter storms in the field. The ability of rocky substrates to retain seedlings increased with their complexity. The interstitial spaces between rocks provided appropriate microsites for seedling settlement, as seeds were successfully retained, and a suitable substrate for anchorage was available. In conclusion P. oceanica juveniles showed a clear-cut preference for hard substrates over the sandy one, due to the root system adhesive properties. In particular, firm and complex substrates allowed for propagule early and strong anchorage, enhancing persistence and establishment probabilities. Seedling substrate preference documented here leads to expect a more successful sexual recruitment on hard bottoms compared with soft ones. This feature could have influenced P. oceanica patterns of colonization in past and present time.

Evidences of adaptive traits to rocky substrates undermine paradigm of habitat preference of the Mediterranean seagrass Posidonia oceanica.

Posidonia oceanica meadows are acknowledged as one of the most valuable ecosystems of the Mediterranean Sea. P. oceanica has been historically described as a species typically growing on mobile substrates whose development requires precursor communities. Here we document for the first time the extensive presence of sticky hairs covering P. oceanica seedling roots. Adhesive root hairs allow the seedlings to firmly anchor to rocky substrates with anchorage strength values up to 5.23 N, regardless of the presence of algal cover and to colonise bare rock without the need for precursor assemblages to facilitate settlement. Adhesive root hairs are a morphological trait common on plants living on rocks in high-energy habitats, such as the riverweed Podostemaceae and the seagrass Phyllospadix scouleri. The presence of adhesive root hairs in P. oceanica juveniles suggests a preference of this species for hard substrates. Such an adaptation leads to hypothesize a new microsite driven bottleneck in P. oceanica seedling survival linked to substrate features. The mechanism described can favour plant establishment on rocky substrates, in contrast with traditional paradigms. This feature may have strongly influenced P. oceanica pattern of colonisation through sexual propagules in both the past and present.

The impact of dredge-fill on Posidonia oceanica seagrass meadows: regression and patterns of recovery.

Posidonia oceanica meadows can be severely damaged by dredge-fill operations. We report on the construction of gas pipelines that occurred between 1981 and 1993 in SW Sicily, Italy. A large portion of the meadow was mechanically removed, and the excavated trench was filled with a mosaic of substrates, ranging from sand to consolidated rock debris. Meadow loss and recovery were quantified over 7 years after the end of operations. We recorded an overall loss of 81.20 ha of meadow. Substrate strongly affected recovery as the percent cover by P. oceanica consistently increased on calcareous rubble, reaching values of 44.37 ± 3.05% in shallow sites after 7 years, whereas no significant increase occurred on other substrates. As in the Mediterranean Sea exploitation of coastal areas continues to grow with consequent impacts on P. oceanica meadows, this case study illustrates how artificial rubble-like materials could be employed to support the restoration of damaged meadows.