MORPHOANATOMIC AND MORPHOMETRIC STANDARDS

Morphoanatomic performance

The estimation of the larval rearing performance derived from morphoanatomic criteria refers to the development conformity and/or the chronology of development of specific organs. The assessment of such criteria will therefore be dependent on the existence of standards which accurately:

• describe the normal or abnormal aspect of the specific organs,
• fix the period when the observation can or should be done,
• indicate how it has to be done.

Then, the choice of the criteria depends on the importance of its consequences on the rearing performance (frequency of appearance, effects on growth, on survival, etc.). For seabass and gilthead seabream larvae, morphoanatomic quality principally refers to anomalies affecting urinary bladder, swim bladder and skeleton.

Urinary calculosis

Urinary calculi can be observed in the urethra or urinary bladder. They have the aspect of greyish or yellowish little stones. The colour may sometimes be reddish during the larval stage. They are essentially calcium phosphate crystals, Ca₅(PO₄). Urinary calculi can be observed from hatchlings to juvenile stages. Up to a total larval length of 20 mm, calculi can be observed under the microscope. At a larger size, calculi can only be detected using soft X-rays.

Fig.74.02 Good gilthead seabream fry and juvenile profile (photo M. Caggiano)
Calculi are believed to be related to stress and poor management, but do not seem to be systematically lethal. The frequency of presence of calculi in the fish stocks varies from 0 to 30%. In some particular cases, it may reach 60%.
For larval rearing calculi are the earliest signal of poor population quality, due to many factors like management, water quality, scarce feed etc. For gilthead seabream when the total observation overcome 30% during the first feeding days
(5 to 15) is better to empty the tank and restart the culture with a new population.

Fig.75.01-2 Very large calculi in this case larval survival is frequently very low (photo STM Aquatrade)

Swim-bladder development

The swim bladder is located between the backbone and the anterior part of the digestive tract of the fish larva. The functional organ looks like a refractive bubble. It reaches 20-30% of the total fish length in individuals longer than 40-50 mm. When the swim-bladder is not functional, it looks like a small slightly translucent vesicle whose size does not exceeds 3 to 5% of the total fish length. The initial inflation of the swim-bladder occurs in the early weeks of the larval life, when gilthead seabream larvae measure 4-5 mm and seabass about 5.5-6.5 mm. This organ can be observed by transparency under a microscope up to a larval size if 15 mm. Thereafter, soft X-rays have to be used to detect it.
The initial inflation of the swim-bladder is triggered when the larvae gulp for air at the water surface. The presence of an oily film or an excessive turbulence in the larval tank inhibit its inflation by preventing fish to reach the surface. Hence the introduction of floating skimmers that remove any surface dirt and oily films, and the adoption of a gentle water circulation to prevent excessive turbulence. Without these basic precautions, the percentage of larvae which do not develop a functional swim-bladder can reach 100%. The non-activation of the swim-bladder has serious consequences on fish, such as:

- an important delay in growth in both species, reaching 20-30% (in weight) when larvae measure around 10-15 mm (60 days old) and over 50% in 30-50 g fish;
- deformities of the backbone (lordosis) that appear at a size of about 20 mm, in both species.

Skeletal deformities

The most common skeletal deformities affecting bass and bream larvae, juveniles and adults concern jaws, gill opercula, head and backbone.

Deformities in newly hatched larvae

A fair number of abnormalities can be observed in newly hatched larvae, the most frequent ones being a form of body twisting. Affected larvae do not survive more than few hours, or few days at best, as the affected portion generally necroses. This deformity may affect from a small percentage to the totality of the population. If this percentage is above 10% it may be opportune to discard the entire batch.
The genetic origin of such anomalies is not proven, even if in trout farming it can be induced by inbreeding. On the contrary most authors believe that poor rearing conditions are a likely cause, in particular in relation to:

• nutritional deficiencies in the broodstock during ovogenesis (the most probable);
• inadequate lighting during incubation;
• excessive egg density (leading to mechanical stress and limited oxygen supply);
• handling, salinity or thermal shocks;
• pollutants in the rearing environment;
• a mix of the above mentioned causes.

Jaw and opercula deformities

Deformities can affect both the maxilla and/or the mandible, which can be either incomplete or protruding. A single operculum or both of them may be absent or be incomplete, or even be bent outwards. For larval sizes below 15-20 mm a microscope has to be used to detect them. For larger sizes they can be visually observed. Deformed jaws may be observed in larvae from hatching. Operculum deformities cannot be detected before larvae reach a length of 12 mm.
In both cases frequencies vary from 0 to 80% during the larval stage. Mandible deformities are often lethal as more than 80% of the affected larvae die, most probably due to starvation. The growth of surviving fish, although delayed, is not greatly affected (about 20% less than normal) and no additional mortality is observed later on.
On the contrary, opercula deformities severely affect growth performance (a difference of up to 60% in weight was observed in 7 months old fish) as well as they affect survival rate (over twice the mortality present in normal fish).

Backbone deformities

The most frequent skeletal deformities affect 2 to 6 vertebrae of the backbone. Scoliosis, kyphosis and fusion of several vertebrae are frequently observed, but lordosis remains the most diffused type of backbone deformity. When fish are affected, the backbone shows a typical V shape with a more o less pronounced angle. In fish without a functional swim-bladder, lordotic deformities are mainly located at the 15^th vertebra (counting from the tail), and at 9^th vertebra in other cases. As muscles involved in swimming act mechanically on the spine, in a fish with an abnormally developed spine they induce deformities in the area where the swim-bladder should be.
The first spinal deformities can be observed by transparency in larvae measuring around 15-20 mm, which corresponds to a stage in which bone calcification is sufficiently advanced. For larger fishes, soft X-rays have to be used.
The frequency of lordosis in the stock is directly linked to its origin:

• in fish without a functional swim-bladder it appears in both species. The percentage is equal to that of the non functional swim-bladder;
• in fish having a functional swim-bladder it may range from 0 to 100%.

The effects of lordosis on fish also vary according to the origin of the deformity:

• in 1 g seabass with a functional swim-bladder, the lordosis was associated to retarded growth (not well quantified), but no mortality was apparently induced by this deformity, whose angle decreased as fish grew, without disappearing completely.
• in fish without a functional swim-bladder, the lordosis is associated to growth delays and to the mortality previously described. These deformities are irreversible, even in case of late inflation of the swim-bladder (e.g., between 7 and 54 g in gilthead seabream).

In both cases, environmental conditions that force swimming during the nursery stage, which may be due to an excessively strong water circulation, increase lordosis frequency. In fish without a functional swim-bladder, these conditions also increase the lordosis angle.
The criteria described above are now commonly used to assess the quality of hatchery-produced fish during the rearing process. Morphoanatomic criteria are more frequently used when marketing fingerlings. Actually, the quality assessment of seabass and gilthead seabream, both as fingerling and market size fish, is based on the percentage of fish lacking a functional swim-bladder or affected by jaw, operculum or spinal deformities.
Essential information available on the origin of such anomalies, which concerns both fresh water and marine species, has led to formulate an hypothesis of a common osteogenic origin of the different skeletal anomalies at operculum, jaw or spine level. In case of lordotic deformities, the hydrodynamism of the larval rearing tanks would represent a condition that highlights this problem.

Fig.76.01 Gilthead seabream fry with incomplete operculum (photo STM Aquatrade)
A review of osteogenesis related troubles in fish, at the origin of their skeletal deformities, showed that they are mainly induced by nutritional deficiencies or by the toxicity of some ingested elements. They affect bone texture, mainly modifying collagen metabolism or altering calcium and phosphorous fixation. Examples of toxicity-induced deformities are also numerous, involving heavy metals and pesticides, as well as an excess of some metabolites or vitamins.
At first glance, the choice among the possible causes of skeletal deformities in seabass and gilthead seabream is ample. Nevertheless, mechanisms involving heavy metal toxicity, environmental disturbances or the involvement of pathogens should be excluded because they would be too much site related when compared to the relatively large diffusion of this problem. Actually such skeletal deformities are observed in too many different farms and in different rearing conditions to believe that they are only due to some exceptional circumstances.

Fig.77.01 Lordotic gilthead seabream fry (photo STM Aquatrade)
Therefore, there should be a common factor that acts on a variety of environmental situations and rearing conditions. Some nutritional deficiency seems the more realistic hypothesis because rearing techniques (feeding sequences, preys, artificial feed, etc.) are more or less the same in most hatcheries. Looking at nutrition more in detail, a deficiency in vitamin C and/or a toxic excess of vitamin D could represent the most probable candidates. Vitamin C is one of the main agents in the collagen metabolism, an essential component of the bone tissue. Vitamin C deficiency can easily be explained by its high solubility in water.
The hypothesis of hypervitaminosis D is also attractive because it takes into account the appearance of urinary calculosis. This vitamin is present in large amounts in the fish oil used to enrich rotifers and brine shrimps for larval fish, as well as in the fish viscera, a component of artificial feeds used in the nursery stage. Tuna liver oil, for example, may contain up to 200 000 IU of vitamin D and cod liver oil up to 500 IU per gram (1 International Unit = 0.025 µg of vitamin D2 as crystalline form).
In human beings, the non-active D3 form is transformed into the active form 25-1-hydroxycolecalcipherol in the kidney. If this transformation does not take place for any pathological reason, a functional deficiency appears, even if the vitamin is abundant in the assumed food. Unfortunately, the active form of the vitamin D in fish is still unknown.