Limpet larvae (Patella aspera Röding, 1798), obtained by gonad dissection and fecundation in vitro, settled and metamorphosed on crustose coralline algae

Abstract

The limpet Patella aspera Röding, 1798, is a native species from the Macaronesian region whose fishing is regulated. The early life of limpets, including the settlement process, is poorly known thus far. The current study evaluated different substrates to induce settlement in P. aspera, including microalgae strains (Halamphora coffeaeformis, Navicula incerta and Pavlova sp.) and crustose coralline algae (CCA) obtained from limpet shells. The results showed that gametes obtained by dissection and matured artificially using alkalinized seawater baths can produce viable larvae able to metamorphose to juveniles. Feeding was not required during larval development, suggesting lecithotrophy. Early postlarvae were identified by the shedding of the velum, and juveniles were identified by teleoconch and active grazing behaviour. The presence of CCA shortened the timing for settlement and increased the ratio of juveniles. The type and abundance of CCA can influence settlement success. Moreover, the results suggested that settlement and metamorphosis in true limpets (Patellogastropoda) might be triggered by a two-step mechanism, i.e. a first cue influencing the shift between swimming and crawling activity and a second cue determining settlement and metamorphosis to early postlarvae and juveniles.

Methodologies for Patellid Limpets’ Aquaculture: From Broodstock Management to Juveniles

“…Since the study by Corpuz (1981), it has been known that fertilization success can be enhanced using an artificial maturation treatment consisting of an alkaline seawater bath, generally consisting of NaOH solutions at pH 9.0-9.5 for several hours (Aquino De Souza et al, 2009;Peŕez et al, 2016;Guallart et al, 2020b;Cañizares et al, 2021;Castejoń et al, 2021), or based on NH 4 OH solutions at pH 8.5-9.0 lasting no more than 10 min (Wanninger et al, 1999;Gould et al, 2001;Hodgson et al, 2007;Ribeiro, 2008;Peŕez et al, 2016;Seabra et al, 2019;Nakano et al, 2020). The alkaline treatment promoted chorion removal and the acquisition of a spherical shape and significantly reduced the oocyte diameter to 141 ± 6 µm in P. aspera (Yuen's test, pvalue = 0) and 146 ± 6 µm in P. candei (Yuen's test, p-value = 0) (Figures 5Q, S; assay 5) when using NaOH at pH 8.4-9.0 for 3 h on these species (Cañizares et al, 2021;Castejoń et al, 2021;Castejoń et al, 2022). The duration of the alkaline bath should be controlled to avoid the degradation of the oocytes (Figure 5R) and to limit the ratio of abnormal development (Figures 9J, K) (Cañizares et al, 2021;Castejoń et al, 2021).…”

“…The same type of containers used for the alkaline treatment and the incubation were used successfully for larviculture during several trials. Previous studies used larval densities ranging from 1 larvae ml −1 (Ribeiro, 2008), 3-7 larvae ml −1 (Seabra et al, 2019), to 10-15 larvae ml −1 Castejoń et al, 2022;Ferranti et al, 2022). Nevertheless, particularly good results were obtained when using an initial density of approximately 5 larvae ml −1 .…”

“…Assay 16 studied the settlement in P. candei following the methodology described by Castejoń et al (2022). The pediveligers were obtained from alkalinized oocytes (NaOH, pH 8.9, 3 h) and trochophores cultured in glass beakers (12 ± 4 larvae ml −1 ).…”

“…The fertilization of the oocytes of P. aspera and P. candei was realized by adding the sperm directly to the oocytes at a density of 10 5 -10 6 sperm cells ml −1 (Cañizares et al, 2021;Castejoń et al, 2021;Castejoń et al, 2022), which was within the range used for other patellid limpet species (Hodgson et al, 2007;Ferranti et al, 2018;Guallart et al, 2020b;Ferranti et al, 2022). The fertilization period is highly variable in the literature, from 30 min for Patella ulyssiponensis (Hodgson et al, 2007) to 3 h for Patella vulgata (Peŕez et al, 2016), and intermediate values for other patellids (Gonzaĺez-Novoa, 2014;Seabra et al, 2019;Guallart et al, 2020b).…”

“…The fertilization period is highly variable in the literature, from 30 min for Patella ulyssiponensis (Hodgson et al, 2007) to 3 h for Patella vulgata (Peŕez et al, 2016), and intermediate values for other patellids (Gonzaĺez-Novoa, 2014;Seabra et al, 2019;Guallart et al, 2020b). However, the sperm can be left with the oocytes during the entire incubation period (24 ± 2 h) without adverse effects on the culture, such as increased polyspermy (Cañizares et al, 2021;Castejoń et al, 2021;Castejoń et al, 2022), saving additional rinses and losses of oocytes. The incubation was realized in static conditions, i.e., neither aeration nor stirring was provided.…”