Mitigating bycatch in Mediterranean trammel net fisheries using species-specific gear modifications.

Small-scale fisheries (SSF) use static gear which are thought to interact with marine ecosystems more benignly than towed gear. Despite this, trammel nets, one of the most extensively used type of fishing gear in the Mediterranean SSF, generate large amounts of discards, which can account for 25% or more of the captured biomass. Discarded organisms may include endangered or threatened species such as elasmobranchs, as well as non-commercial invertebrates that damage fishing gear or cause disentanglement delays. We evaluated various trammel-net gear modifications, including (i) the use of a guarding net attached to the footrope, (ii) increasing the length of the rigging twine between the footrope and the netting panel, and (iii) decreasing the mesh size of the outer panels. The last two modifications were successful in lowering captures of the marbled electric ray Torpedo marmorata, which is commonly discarded in the study area. Both sorts of modifications are relatively simple, their manufacturing does not represent an added cost to implement, and most importantly they do not negatively affect the catch of the target species. The current study shows that prior evaluation of the discard profile of distinct métiers is essential to accomplish species-specific gear modifications and underlines the importance of collaboration among scientists, fishers and gear manufacturers.

Oogonial proliferation and early oocyte dynamics during the reproductive cycle of two Clupeiform fish species.

Although oogonial proliferation continues in mature females in most teleosts, its dynamics and the transformation of oogonia to early meiotic oocytes during the reproductive cycle have received little attention. In the present study, early oogenesis was examined throughout the reproductive cycle in two Clupeiform fishes, the Mediterranean sardine, Sardina pilchardus, and the European anchovy, Engraulis encrasicolus. Observations using confocal laser scanning microscopy (CLSM) provided extensive information on markers of oogonial proliferation (mitotic divisions, oogonia nests) and meiotic prophase I divisions of oocyte nests (leptotene, zygotene, pachytene, diplotene) in ovaries of different reproductive phases. In sardine, oogonial proliferation persisted throughout the entire reproductive cycle, whereas in anchovy, it was more pronounced prior to (developing ovaries) and after (resting ovaries) the spawning period. Anchovy exhibited a higher rate of meiotic activity in developing ovaries, whereas sardine exhibited a higher rate in resting ovaries. The observed differences between the two species can potentially be attributed to different seasonal patterns of energy allocation to reproduction and the synchronization between feeding and the spawning season.

Balbiani body formation and cytoplasmic zonation during early oocyte development in two Clupeiform fishes.

The Balbiani body (Bb) was examined in primary growth phase oocytes for the first time in two clupeoid fish species, the Mediterranean sardine, Sardina pilchardus, and the European anchovy, Engraulis encrasicolus, which belong to different families, Clupeidae and Engraulidae, respectively. Cytoplasmic morphological changes of early secondary growth oocytes were also investigated using confocal laser scanning microscopy, light and transmission electron microscopy. The ultrastructural observations showed that the two species develop a distinct spherical Bb. However, differences in the cytoplasm, mainly in the perinuclear area, were observed. Briefly, in sardine the Bb coexists with a thick perinuclear ring containing mitochondria, nuage, endoplasmic reticulum and small vesicles, while in anchovy this perinuclear ring is thinner, consisting of complexes of nuage and mitochondria. After the disassembly of the Bb, a prominent cytoplasmic zonation develops in the secondary growth oocytes of sardine and anchovy, although with different organelle distribution between the two species. Sardine oocytes exhibit a thick zone of endoplasmic reticulum around the nucleus, whereas in those of anchovy, a thick mitochondria-rich ring surrounding the nucleus was observed. The cytoplasmic characteristics, such as the perinuclear ring in primary oocytes in sardine and the mitochondria-rich ring of early secondary oocytes in anchovy, are also discernible in histological sections by standard procedures and could thus be used as indicators of maturity or imminent spawning period in routine light microscopy observations, providing a valuable tool for applied fisheries biology.

Ultrastructural changes in mitochondria during oogenesis in two phylogenetically close fish species.

The ultrastructure of oocyte mitochondria and their contribution to the endogenous autosynthesis of the yolk was investigated in two clupeoid species, the Mediterranean sardine, Sardina pilchardus, and the European anchovy, Engraulis encrasicolus. The structure and abundance of mitochondria differ in secondary growth oocytes of the two species, whereas they are similar in chromatin nucleolus and primary growth oocytes. Sardine oocytes show a higher percentage of mitochondria in the cytoplasm as they develop. However, the individual size of each mitochondrion decreases, becoming smaller than those observed in anchovy oocytes. The volume fraction of cristae in mitochondria of sardine oocytes gradually increased throughout the oocyte developmental phases up to the early secondary growth phase and then slightly decreased during the mid-secondary growth phase. In the cytoplasm of early secondary growth oocytes of anchovy, the percentage of mitochondria is larger than in mid-secondary growth oocytes. As oocytes develop, the size of mitochondria diminishes as well. In contrast to the volume fraction of cristae in mitochondria of sardine oocytes, the volume fraction of cristae in anchovy was decreased in early secondary growth oocytes and then it was increased during the mid-secondary growth phase. As a result, based on both cytoplasmic dynamics of each species and mitochondrial alterations, it was assumed that mitochondria in sardine play a role in the formation of yolk granules, whereas mitochondria in anchovy play a role in the lipid synthesis pathway. Both species showed exogenous heterosynthesis of yolk, through the process of pinocytosis in the zona radiata of oocytes.

Reproductive dynamics of the invasive lionfish (Pterois miles) in the Eastern Mediterranean Sea.

The lionfish Pterois miles invaded the Mediterranean Sea in 2012 and spread fast to the entire eastern basin. This study provides evidence of successful spawning and the first detailed analysis of P. miles ovarian dynamics in the Mediterranean Sea. The ovarian reproductive phases of mature females collected from Cyprus (eastern Mediterranean) between September 2017 and August 2018 were analysed, both macroscopically and histologically. The results suggested a prolonged oocyte recruitment and development season, and a spawning season that primarily occurred during summertime, even though spawning-capable females were caught also in autumn. A year-round spawning activity may have been inhibited due to seasonal variations in water temperature. Multiple oocyte developmental stages co-occurred in females at different reproductive phases, indicating that more than one oocyte batches were released per spawning season. The analysis of the spawning batch formation enabled the batch fecundity estimation at 3225-63149 oocytes. The oocyte development pattern described in this study shared characteristics with the indeterminate fecundity type, where new oocytes are recruited to the secondary growth phase in parallel with spawning activity. Climate change is likely to extend the spawning season of lionfish in the Mediterranean Sea and further favour its invasion. The information provided in this study is vital for the design of strategic and effective management plans to restrain the expansion of this highly invasive fish.

Using clustering algorithms for identification of fish oocyte cohorts based on the characteristics of cytoplasmic structures.

In batch spawning fish, secondary growth oocytes (SGO) are recruited and spawned in successive cohorts, and multiple cohorts co-occur in spawning-capable females. So far, histological features such as the prevalence of cortical alveoli or yolk granules are conservatively used to distinguish oocytes in different developmental stages which do not necessarily correspond to different cohorts. In this way, valuable information about spawning dynamics remains unseen and consequently misleading conclusions might be drawn, especially for species with high spawning rates and increased overlapping among oocyte cohorts. We introduce a new method for grouping oocytes into different cohorts based on the application of the K-means clustering algorithm on the characteristics of cytoplasmic structures, such as the varying size and intensity of cortical alveoli and yolk granules in oocytes of different development. The method allowed the grouping of oocytes without the need of using oocyte diameter, and thus, a crucial histological bias dealing with the cutting angle and the orientation of reference points (e.g. nucleus) has been overcome. Using sardine, Sardina pilchardus, as a case study, the separation of cohorts provided new insight into the ovarian dynamics, indentifying successive recruitment of up to five oocyte cohorts between SGO recruitment and spawning. These results verified previous histological indications of the number of cohorts in sardine. Altogether, this method represents an improved tool to study species with complex ovarian dynamics.

Development of a new 'ultrametric' method for assessing spawning progression in female teleost serial spawners.

The collection and presentation of accurate reproductive data from wild fish has historically been somewhat problematic, especially for serially spawning species. Therefore, the aim of the current study was to develop a novel method of assessing female spawning status that is robust to variation in oocyte dynamics between specimens. Atlantic cod (Barents Sea stock) were used to develop the new 'ultrametric' method, that is based on the progressive depletion of the vitellogenic oocyte pool relative to the rather constant previtellogenic oocyte (PVO) pool. Fish were subsequently partitioned into one of four categories that accurately reflected changes in their oocyte size frequency distribution characteristics and gonadosomatic index throughout spawning. The ultrametric method overcomes difficulties associated with presence of bimodal oocyte distributions, oocyte tails, lack of clear hiatus region, and presence of free ova, and can be implemented at a single sampling point. Much of the workflow is fully automated, and the technique may circumvent the need for histological analysis depending on the desired outcome. The ultrametric method differs from the traditional autodiametric method in that PVOs can be separated by ultrasonication and then enumerated, and ovarian homogeneity is not a mandatory requirement per se. The method is designed for determinate spawners but might be extended to include indeterminate spawners.

Timing of oocyte recruitment within the ovulatory cycle of Macedonian shad, Alosa macedonica, a batch spawning fish with indeterminate fecundity.

The recruitment of primary growth (PG) oocytes to the secondary growth (SG) phase within the ovulatory cycle, i.e. time interval between two sequential ovulation/spawning events, has rarely been examined in detail in a quantitative manner for batch spawning fishes with indeterminate fecundity. In the present study we analyzed the ovarian dynamics of Macedonian shad, Alosa macedonica, an iteroparous batch spawning clupeid with indeterminate fecundity, with the main goal to define the timing of PG recruitment and relate it to the ovulatory cycle. The latter was classified into four different phases (pre-ovulatory, running, post-ovulatory, intermediate) through postovulatory follicles and presence/absence of hydrated oocytes. Various indices of ovarian dynamics, including the formation and mean diameter of the advanced oocyte batch, the ovarian developmental stage, the oocyte size modality and the gonadosomatic index, varied among the ovulatory phases, evidencing cyclicity. Relative fecundity of newly recruited SG oocytes was used as an index of PG recruitment intensity and was shown to follow a specific pattern within the ovulatory cycle; PG recruitment occurred in a stepwise manner and in parallel with ovulation of the advanced oocyte batch, and synchronization of these two processes kept the ovary in a state of dynamic equilibrium.

Primary and secondary oocyte growth dynamics in anadromous semelparous Allis shad Alosa alosa.

We analysed the ovarian dynamics of the anadromous semelparous allis shad Alosa alosa for which our working hypothesis was that mature pre-spawning females would have very low or even exhausted primary growth (PG) oocyte reserves; semelparity has been linked with the depletion of the pool of PG oocytes. To test this hypothesis, the PG oocytes were enumerated, their recruitment pattern to the secondary growth (SG) phase was analysed and their potential replenishment from the pool of oogonia was examined in females caught very close to the Mondego River mouth, in central Portugal and along the river. The development of the SG oocytes was also analysed, the fecundity (batch, total and annual) values were estimated and the intensity of atresia was quantified. Ovarian samples and histological sections were investigated in parallel. A dynamic recruitment pattern of PG oocytes to the SG phase was revealed, where all PG oocytes were recruited and were not replenished by oogonia. Annual fecundity was subject to down-regulation due to atresia prior to spawning and its size was multiple times higher than the size of batch fecundity. Lack of population synchronicity in ovarian development and spawning migration was also observed. This multifaceted analysis of the ovarian dynamics of this species will contribute to management efforts for this critically endangered and economically important fish throughout its geographical distribution. The results reported in this study will also assist in unravelling the complexity of the early processes of oogenesis in fish.

A field based study of swimbladder adjustment in a physostomous teleost fish.

The present study assesses swimbladder dynamics in natural occurring sardine, Sardina pilchardus, populations with the aim to examine whether this is affected by bathymetric positioning and the physiological state of the individuals. To do so, swimbladder size and shape were modeled in relation to catch depth and the size of various visceral compartments such as gonad, liver, fat and stomach. Swimbladder size was shown to be related to depth in a way that individuals with smaller swimbladders occurred at larger depths. Moreover, evidence is provided that the swimbladder in sardine might have a functional relationship both with the reproductive and the feeding state of individuals, since none of the fish with hydrated gonads and/or large stomachs displayed distended swimbladders.