Development of All-Male Tilapia Hybrids for Improved Cold Tolerance

COLD TOLERANCE OF GENETICALLY PRODUCED ALL-MALE TILAPIA HYBRIDS (ORECOCHROMIS AUREUS)

Eran Lahav1 and Zivi Ra’anan2

1 Nir David fish Breeding Farm, D.N. Gilboa, Israel

2 Aquaculture Production Technology (Israel) Ltd., P.O.Box 4330, Jerusalem, Israel

Abstract

The performance of all-male fry progeny of O. aureus was examined under temperate conditions, in comparison with that of the commercially cultured O. niloticus x O. aureus all-male hybrids. In both cases all males progeny were obtained genetically without use of hormonal sex reversal.

The effects of low temperature was evaluated on three production traits: (a) The onset of spawning at Spring time; (b) The growth rate during interim seasons as well as during summer; and (c) The survival in winter temperature, as a function of both absolute minimal temperatures as well as duration of exposure to that temperature.

It was found that O. aureus begins to reproduce in Spring, at water temperature 3oC lower than the cross of O. niloticus x O. aureus, resulting in the availability of fry for stocking 25 days prior to the appearance of the hybrid fry. When water temperature is lower than 24oC, fry of O. aureus grow significantly faster than the hybrid fry. However, this advantage diminishes during the warm summer season, when the hybrids grow faster. Thus, the relative duration of the cold period determines the extent of the O. aureus advantage in this respect. Regarding cold resistance, O. aureus and the hybrid showed a significant advantage over O. niloticus, and only a slight advantage of O. aureus over the hybrid, which may not be significant. The implications of these finding to commercial production are discussed.

Introduction

Temperature tolerance is a key factor in tilapia production in temperate zones. Most strains of commercially grown tilapia cease feeding at temperatures lower than 18oC. Water temperatures below 14o or 12oC (depending on the species and duration of exposure) can cause mortality (Tave, 1990; Tave et al., 1990). In tropical regions, the preferred species for commercial production is Oreochromisniloticus and its hybrids, due to their high growth rate in tropical climate. However, tilapia aquaculture in temperate, and to some extent in sub-tropical zones, such as Israel and the USA, is characterized by seasonal changes in water temperature. During winter, water temperature may drop to levels that cause severe growth inhibition and sometimes, mortality. The gradual increase and decrease of temperature during the interim seasons of Autumn and Spring, affect the physiology of the fish. Under these conditions, O. niloticus, which is not very tolerant to cold temperatures (Khater and Smitherman, 1988) is at a disadvantage, and a species or a hybrid with greater cold tolerance may significantly improve the profitability of the industry.

Oreochromisaureus is the most cold-resistant species among commercially produced tilapias in Israel, as well as in other countries (Saris, 1969, Behrends & Smitherman, 1984). For this reason, Nir David Fish Breeding Farm, which has been producing the all-male genetic hybrid of O. niloticus x O. aureus, decided to examine the production potential of O. aureus. The method proposed by Shelton et al. (1978) was adopted to established O. aureus broodstock which produces all-male fry without the need for hormonal sex-reversal (Lahav, 1993). O. aureus males, which are believed to be homozygous in their sex chromosomes, were sex-reversed by feeding them with estrogen impregnated fish meal. Sex-reversed females (phenotypic females with a male genotype) were identified by mating them with normal O. aureus males. Pairs which produced 100% male progeny were selected and the females, once identified, were bred again with males from the same O. aureus line. The all-male fry produced were treated with estrogen to produce the female line of the commercial broodstock. Crossing these females with normal O. aureus males resulted in the commercial production of 98-100% male populations.

The tolerance to low temperature of the O. aureus all-male fry was examined on three production traits. First, we examined the initiation of reproduction and spawning activity. This trait is of importance since the earlier the spawning begins in Spring, the longer is the growth period, until the onset of the following winter. Moreover, early spawning means more efficient utilization of the spawning facilities, which, otherwise, may stay idle during the greater part of the year.

The second production trait examined was the growth rate of the O. aureus hybrid, in comparison with that of the commonly produced O. niloticus x O. aureus hybrids. Although it is well established that under optimal water temperatures (24o - 30oC), O. niloticus and its hybrids grow faster than O. aureus (Wohlfarth and Hulata, 1981), it was important to compare their growth during the transition period under conditions of gradual seasonal changes in water temperature (i.e. spring and fall time), as well as during summer.
The third production trait examined was the sensitivity of the fish to exposure to low temperatures. Winter kill of tilapia in temperate zone aquaculture is a serious problem (Snodgrass, 1991). Episodes of cold temperature shocks (where the determining factor is the absolute minimal temperature) which result in tilapia kills (Stauffer, 1988) are rather rare. More often, cold-tolerance is more than just a direct function of the absolute lowest temperature tolerated by the fish. Rather, the resistance to low temperature is determined primarily by the combination of temperature and time of exposure, and is affected also by the rate of decrease in temperature, by the physical condition of the fish in terms of health and nutrition, by water quality and possibly more. This aspect is of utmost importance for the tilapia industry in Israel, since, in most cases, tilapia cannot reach market-size within one season and must be stored in open ponds over-winter. Under such conditions “winter kills” occur quite often, causing losses to the industry.

Materials and Methods

a.Onset of spawning activity

A comparison was made between females of O. niloticus crossed with O. aureus males, and those of O. aureus phenotypic females and males. Four spawning ponds of 40 m3 were allocated for this purpose, two were stocked with 7 females and 2 males of thehybrid parents, and two were stocked with identical number of parents of O. aureus.

Water flow rate was 5 m3/hour, from a nearby spring. The experiment was carried out from March 27 to April 30, year 1992. During this period, water temperature increased gradually from 18oC to 23oC. Temperature was measured daily in all ponds, early in the morning.

Daily observations for early detection of fry were carried out until each pond produced three spawning cycles (after which, it was impossible to distinguish between old and new batches of fry). Fish were fed according to a standard feeding table, corrected according to water temperature.

b.Growth rate at different temperature ranges

In this experiment, growth rate of O. aureus was compared to that of O. niloticus x O. aureus hybrids (the common hybrid commercially produced in Israel). Both fry populations were all- males.

The growth trial was carried out in two stages of 40 days each: (a) from March 15 to April 25, during which temperature increased from 20o to 23oC degrees, representing the period of seasonal change; and (b) from May 25 to July 5, during which temperature increased from 26o to 30oC, representing summer.

Each trial was carried out in three replicates, in six 2 m3 tanks with central drainage. The temperature of the stream water, pumped through the tanks, changed according to season. Each tank was stocked with 30 fish. Whenever a fish was found dead, another fish of identical size and source replaced it (stand-by populations of both lines were maintained under similar conditions, for this purpose). Altogether, 14 fish, randomly distributed among the tanks, had died and were replaced (4% of the total number of fish). Fish were weighed upon stocking, to minimize stress. Fish were fed 30% protein pellets, according to a standard feeding table. Feed was manually administered twice per day.

c.Cold resistance

Cold resistance of three groups of fish was examined, including O. niloticus (males only), O. aureus (all male), and all male F1 hybrids of O. niloticus x O. aureus. Three replicates were examined for each group. Each replicate included 40 fish, at an average weight of 100 g. In order to equalize the physical status of the fish participating in the study, they were all grown together in a large communal tank, under optimal conditions, for one month prior to the stocking of the trial groups.

The tanks used for this trial are similar to those described above. A recirculating water system was used. Water from all tanks drained into a common external sump, from which it was pumped back into the tanks, using a submersible pump. Fresh make-up water was added to the common sump as required. This arrangement ensured that all tanks were kept at exactly the same temperature. Gradual temperature drop was allowed to occur naturally, according to the cooling during the onset of winter. Tanks were aerated by airlifts.

The trial was carried out for 77 days, from November 15 to January 31. During this period, water temperature naturally dropped from 20o to 8.5oC. Temperatures were recorded every morning at sunrise.

Tanks were fed 30% protein pellets, calculated according to 0.5% biomass. Cleaning was done by flushing water through the central drainage. Sludge accumulating in the common external pit was removed manually. When water temperature dropped below 17oC, feeding (and cleaning) was stopped altogether.

A fish was considered dead upon loosing balance and sinking to the bottom, lying on its side. Dead fish were collected and counted daily.

Results

a.Onset of spawning activity

Onset of spawning activity in each pond was determined by the appearance of the first three batches of fry (each representing a single spawning). Results are presented in Figure 1.

It can be clearly seen that the first three batches of fry produced in the two groups of O. aureus parents were observed earlier (between the 2nd and 9th of April) than those of the O. niloticus parents (between the 24th and 30th of April). Onset of spawning of the O. aureus group occurred at lower temperature, as compared with the O. niloticus x O. aureus group (20.9 and 24.8oC, respectively).

Figure 1.Onset of spawning in 2 groups of O. aureus and 2 groups of O. niloticus x O. aureus, as a function of increase in water temperature.

b.Growth rate at different temperature ranges

In Phase I of the growth trial, fish of both groups (O. aureus and O. niloticus x O. aureus hybrid) were stocked at an average weight of 149.0 g. Water temperature at the beginning of the period was 20oC and at the end, 40 days later, 23oC. All fish were weighed upon harvest. Results of Phase I are summarized in Table I.

Table I. Growth rate at water temperatures ranging between 20-23oC

Results indicate that within the water temperature range examined, O. aureus grew by an average of 50.8% faster than the hybrid.

In phase II of the growth trial, fish averaging 135 g, were stocked for a 40-day, at water temperatures ranging from 26 to 30oC. Fish were weighed at the end of the trial. Results are summarized in Table II.

Table II. Growth rate at water temperatures ranging between 26-30oC

Results indicate that at the higher water temperature range, the hybrids grew, on the average, 13.9% faster than O. aureus.

c.Cold resistance

As water temperature gradually decreased, fish from the three groups started to die. Dead fish were counted daily. The rate of mortality is plotted against the drop in water temperature (Figure 2). Each curve is the summary of three groups and represents 120 fish.
The curve shows that mortality in the O. niloticus group started on Day 36, when water temperature was about 12-13oC. Within the next 13 days, all fish in this group died. At the same time, water temperature further dropped to 10oC. Mortality in the other two groups started 10-11 days later. Temperature was 10-11oC at the onset of mortality and dropped further to 8.5oC. Temperature was not allowed to go further down since this is the minimal temperature observed under Israeli winter conditions, in most areas where tilapia is cultivated. The comparison between the hybrid and O. aureus show quite similar results, with some advantage to O. aureus, mainly with respect to resistance period. Whereas mortality in the hybrid group started on Day 46 and the continued for 17 days, the mortality in the O. aureus group started on Day 49 and continued for 20 days. This is a slight advantage, which may not be statistically significant.

Figure 2. The rate of mortality of fish exposed to decreasing water temperature

Discussion

The main purpose of this work was to compare between the performance of the commercial hybrid, O. niloticus x O. aureus, and all-male O. aureus, under temperate climatic conditions, such as those prevailing in Israel. Our results show that O. aureus has an advantage over the hybrid, in all aspects examined.

Early spawning allows prolongation of the growout season, since fry are stocked in the ponds earlier. Moreover, it increases the efficiency of spawning pond utilization, which would otherwise stay idle for the interim period between the two main seasons. In fact,
during the last two years, a commercial broodstock capable of producing all-male O. aureus without hormonal treatment is already used. Results of commercial activities demonstrate that O. aureus fry are available for stocking as early as the beginning of May, approximately 25-30 days before the hybrid fry are obtained (late May, beginning of June).

The cold tolerance which results in faster growth potential during interim periods, when water temperature is rather low (>24oC), allows the early stocked fry to start gaining weight and continue to do so while the growth of the hybrid is still inhibited by the cold water. However, since the hybrid grows faster during the period when water temperature is higher than 24oC, the relative advantage of O. aureus depends on the relative duration of the interim periods and the summer season. Therefore, this aspect should be evaluated for each case separately, based on site-specific conditions. In Nir David commercial ponds, O. aureus fry averaging 1 g, were stocked on March 15 (fry obtained from greenhouse covered spawning ponds) and reached an average weight of 440g within 245 days (average daily growth of 1.8 g/day). During the same season, hybrid fry were stocked at an average weight of 1 g on April 4, and grew under similar conditions, to an average weight of 410 g, within 220 days (average daily growth of 1.86 day). It can be clearly seen that although the average daily growth rate of the hybrid, over the entire season, was higher, O. aureus reached a larger size at harvest. This is due to the prolonged growout season. In other locations, where the cold season is longer, the relative advantage of O. aureus may be more pronounced.

Regarding cold resistance, our results correspond with those presented in the literature (Tave, 1990), showing that O. niloticus is the least tolerant to cold temperatures, whereas the hybrid is between the two parental lines, closer to the O. aureus. Although we do not yet have concrete data regarding over-wintering fish in commercial ponds, the O. aureus stock seems to be in better shape and less susceptible to various kinds of parasites, which infect the other fish during winter.

References

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Khater, A. A. and R. O. Smitherman, 1988. Cold tolerance and growth of three strains of Oreochromis niloticus. The 2nd International Symposium on Tilapia in Aquaculture, Bangkok (Thailand). No. 15 pp. 215-218.

Lahav, E., 1993. Use of sex-reversed females to produce all-male tilapia (Oreochromis aureus) fry. The Israeli Journal of Aquaculture - Bamidgeh, 45(3):131-136.

Sarig, S., 1969. Winter storage of tilapia. FAO Fish Culture Bulletin, 2:8-9.

Shelton, W. L., Hopkins, K. D. and G. L. Jensen, 1978. Use of hormones to produce monosex tilapia for aquaculture. Pp. 10-33. In: R. O. Smitherman, W. L. Shelton and J. H. Grover (eds). Culture Exotic Fishes Symp. Proc. Fish Culture Section, Am. Fish Soc., Auburn, Alabama.

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Tave, D., 1990. Cold tolerance in tilapia. Aquaculture Magazine, 4:86.

Tave D., Jayaprakas V. and R. O. Smitherman. 1990. Effects of intraspecific hybridization in Tilapia nilotica on survival under ambient winter temperature in Alabama. J. World Aquaculture Society, 21:201-204.

Wohlfarth, G. W. and G. Hulata, 1981. Applied Genetics of Tilapias. ICLARM Studies and Reviews 6. International Center for Living Aquatic Resources Management, Makati, Metro Manila. 26 pp.

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