The cysts of brine shrimp Artemia are widely used in the aquaculture industry (Sorgeloos 1980, Sorgeloos et al. 1998, Sorgeloos et al. 2001). They are produced commercially by culturists in many places or harvested from their natural habitat, such as the Great Salt Lake (Van Stappen 2003).
Irrespective of the fact that each species or population has its own unique characteristics, the processing of Artemia cysts is an important factor in price determination. Thus, different definitions have been developed for the evaluation of the hatching quality of Artemia cysts. Those hatching percentage, hatching efficiency, hatching rate, hatching synchrony and hatching output (Sorgeloos et al. 1978, Van Stappen 1996).
- Hatching Percentage: The number of nauplii that can be produced under standard hatching conditions from 100 full cysts; this criterion does not take into account cyst impurities, such as cracked shells, sand or salt, and refers only to the hatching capacity of the full cysts.
- Hatching Efficiency: Number of nauplii that can be produced from 1 g of dry cyst product under standard hatching conditions.
- Hatching Rate: The time period for full hatching from the start of incubation (hydration of cysts) until nauplius release (hatching), at a number of time intervals.
- Hatching Synchrony: Time lapse during which most nauplii hatch, i.e. Ts = T90-T10.
- A high hatching synchrony ensures a maximal number of instar I nauplii available within a short time span; in case of poor synchrony the same hatching tank needs to be harvested several times in order to avoid a mixed instar I-II-III population when harvesting at T90.
- Hatching Output: Dry weight biomass of nauplii that can be produced from 1 gram dry cyst product incubated under standard hatching conditions; best products yield about 600 mg nauplii/g of cysts. The calculation is made as follows: hatching efficiency x individual dry weight of an instar I nauplius.
These concepts are used in Artemia based on research (Vanhaecke et al. 1981,Vanhaecke and Sorgeloos 1982, Drinkwater and Crowe 1991, Triantaphyllidis et al. 1993, Van Stappen et al. 2003, Saygi 2004, Kara et al. 2004, Abatzopoulos et al. 2006) and as quality determining factors for economic purposes (Vanhaecke and Sorgeloos 1983).
All five definitions, but in particular hatching percentage and hatching efficiency, have major roles in cyst quality determination. The major issue is that these definitions can provide a real role in cyst quality only where there is a suitable processing of cysts. Sometimes, cysts have a high percentage of shells and are cracked. Although freshwater-saturated brine technology is used for purification of Artemia (Baert et al. 1996, Treece 2000), in some cases the method is not efficient in separating shells and cracked cysts of from full Artemia2 cysts. Present definitions and calculations are unable to determine the percentages of shell content and cracked cysts in samples.
In determining hatching percentage after decapsulation of cysts, the shells and cysts with undeveloped embryos are deleted. Most of the cracked cysts disintegrate during the first 3-4 hours after hydration, but they are included as part of the cyst batch. Even hatching efficiency is unable to determine quality of cysts, because it depends on their diameter (Asem et al. 2007). Moreover, because shells and cracked cysts are eliminated at the end of determining the hatching percentage, it doesn't seem unexpected for the cyst mass containing shells and cracked cysts to show a high percentage of hatching.
This article delineates six new concepts and definitions for examining the quality and evaluation brine shrimp cysts that can accurately show the hatching percentage, cyst quality and purification effectiveness.
New Concepts and Their Formulas
Since hatching percentage can't clearly provide cyst quality because cracked cysts are disintegrated in the first 2-3 hours after hydration and the shells are finally removed by sodium hypochlorite. Therefore, it is suggested the title percentage of false hatching (FH) should be substituted for hatching percentage.
1. Percentage of Potential hatching (%PH): the number of larvae harvested from 100 shells, perfect and cracked cysts.
2. Percentages of Absolute Hatching (%AH): is the number of larvae harvested from 100 embryo-bearing cysts whether perfect or cracked
3. Percentage of Cyst Purity (%Cyst Purity): percentage of perfect and perfect embryo-bearing cysts out the total embryo-bearing cysts whether perfect or cracked.
4. Shell Percentage (%Shell): the percentage of shells and imperfect embryo-bearing cysts
5. Percentage of Cracked Cyst (%Cracked Cyst): the percentage of cracked embryo bearing cysts out of total embryo bearing cysts
6. Percentage of Hatching Potential Loss (%HPL): the difference between the %FH and %PH that indicates the potential loss due to existence of cracked cysts and sells. The lower the loss percentage and the higher the potential hatching, the better the quality of the cysts.
The abbreviations FH, PH, AH and HPL are used in Figure 1 and Table 2.
Raw Data from the Study
Mean (S.D) of the Results of Cysts Quality (Same Letters in Each Column Show Significant Difference, ANOVA; Tukey, p<0.05)
Methods and Materials
1. One gram of dry cysts are added to of 1,000 cc of 35 ppt salinity water and aerated.
2. After one hour, six samples are taken from replicates.
3. The total number of cysts in each the six samples are counted (including those cracked, fully cyst: hydrated or dehydrated cysts). The mean is calculated for the six samples so the total number of cysts can be calculated.
4. Immediately, the cysts are decapsulated with sodium hypochloride. At this stage shells and the cysts with imperfect embryos are eliminated but the cracked and full cysts remain and are decapsulated. Then, the decapsulated cysts are counted and mean from the replicates is obtained to determine the total number of decapsulated cysts.
5. After 24 hours (hatching time), the percentage of false hatching (%PFH) is calculated by the method of Van Stappen (1996).
After 24 hours (hatching time), the percentage of false hatching (%PFH) is calculated by the method of Van Stappen (1996).
6. Other conditions required for hatching must be observed.
7. All experiments should be repeated three times.
Finally, the value of each defined parameter is computed.
Hatching quality of cysts from three species of Artemia (A. urmiana, A. franciscana and a parthenogenetic population) were evaluated following the new concepts.
Before examination, all cyst samples were purified using freshwater-saturated brine method. Tables 1 and 2 show the raw data and summarize the results of cysts quality. The results are shown in Figure 1.
In Table 2 the percentages of potential hatching (PH), absolutely hatching (AH), false hatching (FH), hatching efficiency (HE) and cyst purity of the A. franciscana were higher than those of A. urmiana and the parthenogenetic population. The lowest percentage of cracked cyst were associated with A. franciscana. Results shown in Table 2 indicate the shell percentage in the three samples have weree not significantly different but the percentage of cracked cysts in the A. franciscana sample was the lowest. This difference can be attributed to the method of processing the cysts and not to the purification method.
Although the three cyst samples were purified by the freshwater-saturated brine method, the results show even if this method separates a considerable amount of the cyst waste, it isn't perfect. It was concluded that because of the low percentage of hatching potential loss and high hatching efficiency, A. franciscana cysts are of higher quality than the other two types evaluated. Finally, it seems that the new concepts can provide another approach to studies to assess the economics brine shrimp cysts.
This study was carried out in the Iranian Artemia Research Center (I.A.R.C). I am grateful to Latif Esmaeili (the previous headmaster of I.A.R.C) and Siyavash Ganji for administrative and laboratory help.
This study was funded by the Artemia and Aquatic Animals Research Institute of Urmia University, Iran