**2. Technical considerations for the establishment of whitebait cultivation**

*Galaxias maculatus* develops part of its life cycles in freshwater, specifically in rivers and littoral zones of lakes as well as in estuarine environments (Vila et al., 2006). The first stages of its life cycle are carried out in freshwater and it comprises (a) conditioning of crystalline juveniles to obtain marine reproducers; (b) spawning; and (c) fertilization. The second stage is realized under brackish water conditions and comprises (a) incubation and (b) larval culture (Mitchell, 1989; Vega et al., 1996; Valdebenito & Vega, 2003; Mardones et al., 2008; Hicks et al., 2010).

The most important elements and details to consider for *G. maculatus* culture are: (a) selection of the species, (b) selection of the location and the water source for cultivation, (c) appropriate cultivation systems and (d) selection and training of personnel (Mardones et al., 2008).

For reproduction it is necessary a broodstock proportions of one male: three females, and the average production of one female is approximately 600 fertile eggs (Valdebenito & Vega, 2003), and it is possible close the reproductive cycle in a period of 390 days (Tables 1 and 2; Mardones et al., 2008), and it is possible reach crystalline juveniles with commercial weight and size in 180 days after fertilization (Mitchell, 1989; Mardones et al., 2008).




\* Fertilization: total number of eggs obtained from the spawning, 80 % being adequate for incubation.

\*\* Incubation: number of incubated eggs. \*\*\* One kilogram of product = 3330 crystalline larvae approximately.

Table 2. Survival performance required to culture 10 tonnes of *Galaxias maculatus* (Cf: Mardones et al., 2008)

#### **2.1 The selection of sites**

This procedure is based in two aspects: the first is the determination the availability of optimal water quality and the topography of the terrain. Desirable locations have the following features: (a) close to sea level, (b) availability of water with different salinities including brackish water from 5-11 ppt and (c) coastal gravel or sand material, which will allow the installation of stakes in the sea or the sinking of deep sub-superficial wells for marine water (30 m) and deep wells for freshwater (100 m)(Mardones et al., 2008).

#### **2.2 Overview of the production process**

286 Aquaculture

appropriate cultivation systems and (d) selection and training of personnel (Mardones et al.,

For reproduction it is necessary a broodstock proportions of one male: three females, and the average production of one female is approximately 600 fertile eggs (Valdebenito & Vega, 2003), and it is possible close the reproductive cycle in a period of 390 days (Tables 1 and 2; Mardones et al., 2008), and it is possible reach crystalline juveniles with commercial weight

Table 1. Time of duration of the stages of the life cycle of *Galaxias maculatus* (Cf: Mardones et

Number of males 103237 Number of females 309710 Fertilization\* 185825893 Incubation\*\* 50 50 148660714 Hatching 80 20 74330357 First nourishment 80 20 59464286 Postlarva 70 30 47571429 Crystalline larvae\*\*\* 33300000

\* Fertilization: total number of eggs obtained from the spawning, 80 % being adequate for incubation.

This procedure is based in two aspects: the first is the determination the availability of optimal water quality and the topography of the terrain. Desirable locations have the following features: (a) close to sea level, (b) availability of water with different salinities including brackish water from 5-11 ppt and (c) coastal gravel or sand material, which will allow the installation of stakes in the sea or the sinking of deep sub-superficial wells for

Table 2. Survival performance required to culture 10 tonnes of *Galaxias maculatus* (Cf:

marine water (30 m) and deep wells for freshwater (100 m)(Mardones et al., 2008).

Operation % Survival % Mortality Number of inviduals

and size in 180 days after fertilization (Mitchell, 1989; Mardones et al., 2008).

ID Stages Day 1 Fertilization 1 2 Incubation & hatch 30 3 First feeding 40 4 Crystalline larvae 140 5 Maturation 180 6 Total duration 390

2008).

al., 2008)

\*\* Incubation: number of incubated eggs.

Mardones et al., 2008)

**2.1 The selection of sites** 

\*\*\* One kilogram of product = 3330 crystalline larvae approximately.

The production process begins with the getting of wild crystalline juveniles at the time of their migratory returns in September-October, that corresponding to southern spring (Allibone & Townsend, 1997; Valdebenito & Vega, 2003; Boy et al., 2006). They are transported to the culture facilities, where they are held up to six months to reach sexual maturity and as first step, the live specimens are placed in quarantine and given a prophylactic treatments to eliminate bacterial and parasite diseases brought from wild environment. Once mature broodstock are stripped by abdominal pressure and the eggs are fertilized and incubated over a period of 25-30 days (Valdebenito & Vega, 2003).

Once incubation occurs, the small larvae remain in the first feeding stage for 38-40 days, during which time they are fed ad libitum. Subsequently the larvae continue to a fattening stage where they remain for 140 days until they reach the weight and size of the commercial crystalline juveniles to be harvested and processed. A fraction of animals of this group is retained to be conditioned as reproducers for the next cycle. The total duration of the first production cycle late approximately 480 days, it begins from the purchase of the natural fry through conditioning as breeders until harvest and processing of 10 tonnes of crystalline juveniles (Valdebenito & Vega, 2003).

#### **2.3 Purchase and transport of wild juveniles**

Captures of wild juveniles are not constant as they are very dependent on the climate and other factors, fluctuating between 0.5 to 20 tonnes/season, this period requires approximately 90 days, and once, approximately 8 kg of juveniles are captured daily and the juveniles are placed in two tanks of 8 m3 each one, until sufficient amounts are reached to realize the transport to the culture centre, where they will be conditioned (Valdebenito & Vega, 2003). The purchase price at this stage is approximately USD 20/kg (Mardones et al., 2008). The carrying density is 12 kg\*m-3, and it is used tanks capacity of 3 m3 the mortality ranges from 0 to 5 % during transportation but is less when adults are captured (Valdebenito & Vega, 2003). The juveniles easily adapt to the captivity, and the total mortality from the arrival of the fish at the culture centre until pre-spawning is 60 %. The total capture of wild juveniles required to ensure sufficient reproducers in the first year considers: (a) each juvenile weighs approximately 0.3 g; (b) mortality due to transport is 0 %; (c) mortality up to pre-spawning is 60 %; (d) 30 % of the animals will not reproduce; and (e) the sex ratio at purchase is 1:1 as it is only possible to determine the sex once the reproducers are mature (Mardones et al., 2008). The table 3 shows the number and total biomass of juveniles that need to be purchased.

#### **2.4 Broodstock conditioning**

Once they arrived the juveniles are held in twelve 2 m3 tanks for approximately 270 days until they become in optimal reproductive conditions (Valdebenito & Vega, 2003). In according to literature (Valdebenito & Vega, 2003; Mardones et al., 2008) the water quality parameters at this stage are 10-15° C, 0 ppm salinity, a density of 7 kg/m3 and a daily exchange rate of 2. There are two main periods: (1) quarantine period and (2) period of sexual maturity.


Table 3. Number of crystalline juvenile required. Cf: Mardones et al., (2008)

A single quarantine it is necessary when wild fish enter the culture center. This process consists of periodic application of prophylactic treatments from the day following entry to the hatchery until 14 days in order to eliminate potential ectoparasites and external bacteria. The prophylactic treatments are calculated based on saline and oxytetracycline baths and oxytetracycline oral administration (Table 4; Valdebenito & Vega, 2003; Mardones et al., 2008).


Table 4. Types of prophylactic treatments applied and their specifications. Cf: Mardones et al., (2008)

The juveniles are fed with beef liver in the first week; 75 % liver and 25 % oil-free salmonid starter diet in the second week; 50 % liver and 50 % starter in the third week; 25 % liver and 75 % starter in the fourth week, and 100 % starter in the fifth week. Once quarantine period finished, the juveniles are conditioned for 140 days to become reproducers and during this period they are fed a pelletized freshwater salmon diet that provides good growth and disease resistance (Table 5). The diet ration is at 2 % body weight daily in 10 daily rations by automatic feeders, depending on the biomass and the growth of the fish (Table 6; Valdebenito & Vega, 2003; Mardones et al., 2008).

The broodstock are reared in 8 m3 fibreglass tanks 3 m in diameter. They remain in these tanks during vitellogenesis and they are transferred avoiding excess manipulation, only during the spawning period for separation according to their maturity level (Valdebenito & Vega, 2003). The broodstock are spawned three times during their lifetime and so a program of annual replacement and genetic management is required in order to maintain the reproductive cycle. The broodstock are replaced from juvenile produced, as well as from those fish that are not sexually mature from the previous year (Mardones et al., 2008).

Sex can be differentiated by colour and abdomen size. The state of maturity, once reached, is classified by visual check of the external morphology of the specimens and abdominal pressure on a monthy, biweekly basis according to the descriptions of (Valdebenito & Vega, 2003). This maturity stage is determined weekly for the stages mature 2 and hatched 2 with

A single quarantine it is necessary when wild fish enter the culture center. This process consists of periodic application of prophylactic treatments from the day following entry to the hatchery until 14 days in order to eliminate potential ectoparasites and external bacteria. The prophylactic treatments are calculated based on saline and oxytetracycline baths and oxytetracycline oral administration (Table 4; Valdebenito & Vega, 2003; Mardones et al.,

Treatment Concentration Application Saline bath 6 g L-1 Two times per week until 40

Oxytetracycline – bath 35 mg L-1 Day 2, bath of 30 min. Oxytetracycline – oral 75 mg kg-1 Day 3-14 incorporated into

Table 4. Types of prophylactic treatments applied and their specifications. Cf: Mardones et

The juveniles are fed with beef liver in the first week; 75 % liver and 25 % oil-free salmonid starter diet in the second week; 50 % liver and 50 % starter in the third week; 25 % liver and 75 % starter in the fourth week, and 100 % starter in the fifth week. Once quarantine period finished, the juveniles are conditioned for 140 days to become reproducers and during this period they are fed a pelletized freshwater salmon diet that provides good growth and disease resistance (Table 5). The diet ration is at 2 % body weight daily in 10 daily rations by automatic feeders, depending on the biomass and the growth of the fish (Table 6;

The broodstock are reared in 8 m3 fibreglass tanks 3 m in diameter. They remain in these tanks during vitellogenesis and they are transferred avoiding excess manipulation, only during the spawning period for separation according to their maturity level (Valdebenito & Vega, 2003). The broodstock are spawned three times during their lifetime and so a program of annual replacement and genetic management is required in order to maintain the reproductive cycle. The broodstock are replaced from juvenile produced, as well as from those fish that are not sexually mature from the previous year (Mardones et al., 2008).

Sex can be differentiated by colour and abdomen size. The state of maturity, once reached, is classified by visual check of the external morphology of the specimens and abdominal pressure on a monthy, biweekly basis according to the descriptions of (Valdebenito & Vega, 2003). This maturity stage is determined weekly for the stages mature 2 and hatched 2 with

day, bath of 30 min.

feed.

Items Ammount Number of reproducers required for spawning 412946 Number of exemplary in a reproductive state 619420 Virginal reproducer (30 %) 884885 Mortality until pre-spawning (60 %) 2212213 Mortality (0 %) 2212213 Number of wild alevins to buy 2212213 Kilograms alevins 664

Table 3. Number of crystalline juvenile required. Cf: Mardones et al., (2008)

2008).

al., (2008)

Valdebenito & Vega, 2003; Mardones et al., 2008).


Table 5. Ongrowing diet sequence. Cf: Mardones et al., (2008)


Table 6. Characteristics of the fish and feed supplement quantities. Cf: Mardones et al., (2008)

the purpose of reducing the stress level of reproducers. During the post-spawning period for females and males are transferred to unoccupied tanks and a series of additional tanks are required for the juveniles obtained to replenish the broodstock (Mardones et al., 2008).
