Running Title: Acclimation of red tilapia hybrid larvae to increasing salinities

Title: Performance of newly-hatched red tilapia hybrids to increasing salinity levels

Authors: Charlie H. Abayon, Arlene L. Avillanosa* and Christopher Marlowe A. Caipang

Affiliation: College of Fisheries and Aquatic Sciences, Western Philippines University,

Puerto Princesa Campus, Puerto Princesa City 5300, Palawan, Philippines

Corresponding Author: *Arlene L. Avillanosa

College of Fisheries and Aquatic Sciences, Western Philippines

University-Puerto Princesa Campus, Puerto Princesa City 5300,

Palawan, Philippines

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Abstract

The effects on survival, feeding response and growth of newly-hatched red tilapia hybrids

to gradual increasing salinity levels were assessed. Newly-hatched fry were reared in 3-L aquaria

containing either freshwater (Treatment 1) or in water subjected to weekly increments of 5 ppt to

a final salinity of 34-35 ppt (Treatment 2) for 42 days. Each treatment had 5 replicates with a

stocking density of 10 larvae per liter. Survival was relatively similar for both treatment groups.

Likewise, no significant differences (p > 0.05) were observed in the feeding response, weight

gain, specific growth rate (SGR) and feed conversion efficiency (FCE) in both treatments. Taken

together, the results showed that tolerance of red tilapia hybrids to saline waters begins early in

its life history and has a potential to be developed for culture in areas with sufficient supply of

brackishwater or with limited freshwater source. It is recommended that this laboratory-based

study on the performance of red tilapia in saline waters be tested in large-scale brackishwater

ponds or cage culture in marine areas to assess the feasibility of this fish as a candidate for

brackishwater aquaculture or mariculture.

KEY WORDS: fry, growth, hatchery, Oreochromis sp., saline-tolerant, survival

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Introduction

Tilapias are freshwater fish but can tolerate wide fluctuations in water salinity (Ernest et

al., 1991; Watanabe et al., 1992). Among freshwater fish species, tilapia has the fastest growth in

production and widespread appeal in the global market. As a result of its popularity, tilapia is

now known as the ‘aquatic chicken’ due to its robust adaptability to a wide range of

environmental conditions, ease of reproduction in captivity and has a neutral flavor of the meat

that appeals to many consumers (Little, 1998).

Tilapia is one of the best candidates for aquaculture in both brackish- and seawater

environments due to their ability to tolerate wide range of water salinity levels (El-Sayed, 2006).

It was supported by Iqbal et al. (2012) and (Pullin 1982), which reported that tilapia is one of the

important fish species that has several good qualities including good growth in water salinities

ranging 0 to 29 ppt, ability to withstand water temperatures between 8 to 42˚C, and can survive

in low dissolved oxygen (DO). While several studies in the past have demonstrated the

feasibility of culturing tilapias in saline waters, Cnaani and Hulata (2011) stressed that there are

still other approaches that can be utilized to enhance the tolerance of tilapias to saline

environment and the underlying mechanisms that are involved in their improved performance in

saline waters as well as better growth and survival. Examples of these approaches are through

selective breeding (Jaspe and Caipang, 2011a) and hybridization (Lutz, 2001).

The red tilapia hybrids, which are genetic mutants selected from crosses of two species of

the genus Oreochromis, is increasing popularity among fishfarmers and consumers because of its

attractive color, good market price and high salinity tolerance (Carberry and Hanley, 1997;

Gonzalez, 1997; Vadhel et al., 2017). The first reported red tilapia hybrid, known as the

Taiwanese red tilapia, was produced in the late 1960s and was a cross between a mutant reddish-

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orange female O. mossambicus and a normal male O. niloticus (Galman and Avtalion 1983).

Another red tilapia strain, the Florida red tilapia was developed in the 1970s by crossing a

normal colored O. hornorum female with a red-gold male O. mossambicus (Behrends et al.,

1982). A third red tilapia strain was developed in Israel by crossing a red Nile tilapia originating

from Egypt with a wild-type Blue tilapia (Hulata et al. 1995). All these hybrids have been

distributed to different tilapia-producing countries and have been found to be suitable for culture

in both brackish- and seawater (Liao and Chang, 1983; Green, 1997; Watanabe et al., 1997).

In the Philippines, several red tilapia strains are available for culture and the strain/stock

is designated based on the name of the farm/research institution where the fish were obtained

(Romana-Eguia and Eguia, 1999). Earlier studies on growth of this species in lakes and ponds

(Galman et al., 1988; Pruginin et al., 1988; Romana-Eguia and Eguia, 1999), as reference species

in evaluation trials for Nile tilapia (Basiao and Doyle, 1990; Eguia and Eguia, 1993) and

breeding performance (Galman et al., 1988; Eguia, 1996) have been conducted. However, the

performance of the red tilapia hybrids during exposure to increasing salinity levels during their

early life stages is not well-documented. The various indicators of performance of the newly-

hatched red tilapia hybrids must be elucidated so that prospective tilapia farmers will have a

reference point in assessing the likely performance of this species during the grow-out phase.

Hence, this study was conducted to determine the survival, growth and feeding activity of the

newly-hatched red tilapia hybrids during exposure to gradual increase in salinity levels.

MATERIALS AND METHODS

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Study site

The study was conducted at the Aquatic Science Laboratory (ASL) of Western

Philippines University (WPU), College of Fisheries and Aquatic Sciences (CFAS) located in

Puerto Princesa Campus, Puerto Princesa City, Palawan, Philippines.

Breeding of tilapia for fry production

Prior to the breeding activity, red tilapia spawners (180-200g) were randomly selected

from the existing broodstock tank of ASL. Female fish were separated from the males by manual

inspection of the genital papilla. Fish were conditioned for two weeks in the conditioning tanks

and were provided with adequate aeration and feeding at 3% of its biomass. Spawning activity

was done upon mixing the male and female spawners at a 1:3 male-to-female sex ratio in 6 units

of 1.0m x 1.0m x 1.0m “hapa” net that was suspended inside the circular tank. On the 4 th day of

pairing, daily monitoring of the fish was done to check for the presence of fry in the tank until

the 14th day. After two weeks, all spawners were returned to the conditioning tanks. Newly-

hatched fry were maintained in the hapa net and subsequently used for the experiment after they

have completed the yolk-sac stage.

Experimental set-up and design

Ten (10) aquaria measuring 20cm x 13cm x 20cm were randomly arranged in a

completely randomized design (CRD). There were two treatments with five replicates each.

Treatment I served as control (freshwater) while Treatment II contained water with increasing

salinity levels in multiples of 5 ppt increment per week until a final salinity of 34-35 ppt.

Adjustments in the salinity followed the protocol described by Talaver (2017) modified from

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Lemarié et al. (2004). Each aquarium has a capacity of 3 liters and was stocked with the newly-

hatched larvae at a density of 10 fry per liter.

Water and feeding management

The quality of water (pH, DO and temperature) and salinity were monitored daily. All

uneaten food and feces that accumulated at the bottom of the aquaria were siphoned and water

was added to maintain the desired water volume in each aquarium.

Commercial fry mass (CP=40%) for tilapia was given at 30% of the fish biomass.

Feeding was done four times a day based on the standard feeding protocol for tilapia fry. The

behavior of the fish was observed four times daily especially during feeding time. Uneaten feeds

were recorded to obtain data on feeding or non-feeding activity.

Zootechnical parameters

Daily monitoring was done to determine the mortality of fry from which the survival rate

was determined (Watanabe et al., 1990). Feeding efficiency (FCE) (Steffens, 1989), specific

growth rate (SGR) (Clark et al., 1990) and weight gain (Watanabe et al., 1990) were calculated

indicators of the feeding behaviour and growth performance of the fry.

Data analyses

Means and standard deviations of all the zootechnical parameters were obtained for the

two treatments. The data in the two groups were compared using Student’s t-test for independent

samples. All statistical computations were performed at the 0.05 probability level. Analyses were

performed using the statistical package of Microsoft Excel 2010.

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RESULTS AND DISCUSSION

Survival of the newly-hatched red tilapia hybrids in freshwater (Treatment 1) and

exposed to increasing levels of salinity (Treatment 2) over a 42-day observation period is shown

in Fig. 1. The mean survival of the red tilapia fry was higher in Treatment 2 (73.3%) than in

Treatment 1 (57.5), although the survival rates in both groups were comparable (p > 0.05). No

mortality in the tilapia fry was observed for 2 weeks. Mortalities were only observed beginning

14th day and lasted until the 35th day. Survival of the fry in both groups stabilized on the 35 th day

until the termination of the experiment.

In terms of feed consumption of the red tilapia fry in both groups, results showed no

significant difference (p > 0.05) in rate of the consumption of feeds during the study period (Fig.

2). This was evident in the feeding behaviour of the fish in both treatment groups. During

feeding time, the red tilapia fry actively swam towards the feed and showed continuous feeding

until satiation.

The culture performance of fry was determined based on their growth performance as

shown in Table 2. After 42 days of culture, the red tilapia fry exhibited no significant

differences (p > 0.05) in weight gain, specific growth rate (SGR) and feed conversion efficiency

(FCE) in both treatment groups.

Based on the results of the present study, the survival of red tilapia fry exposed to

increasing salinity levels were comparable with the batch that was reared completely in

freshwater conditions. The present study corroborated the results of the recent studies of El-

Sayed (2006), Jaspe and Caipang (2011b), Lawson and Anetekhai (2011) and Iqbal et al. (2012).

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According to El-Sayed (2006), tilapia is an excellent candidate for aquaculture in brackishwater

and seawater due to their ability to tolerate a wide range of salinity. Tilapias are now being

cultured in brackishwater ponds, where salinity levels could reach seawater strength during the

summer months (Jaspe and Caipang, 2011b). The ability of tilapias to withstand a wide range in

salinity levels is dependent on the ability of the body fluids to function in an abnormal range of

internal osmotic and ionic concentrations (Holliday and Jones, 1967; Lawson and Anetekhai,

2011). The fish can regulate the body fluid to restore level of osmotic pressure to near normal

when exposed to increasing salinity levels and this ability is demonstrated early in the life stages

of red tilapias, as what has been observed in an earlier study by Nugon (2003). The weekly

increment of 5 ppt was the appropriate regime in acclimating red tilapia fry to increasing salinity

levels as fish mortality was not drastic. This regime was within the concentration of daily salinity

increments recommended by Lemarié et al. (2004) during acclimation of tilapia juveniles higher

salinity levels.

Red tilapia, particularly the Florida red tilapia strain, are able to reproduce in salinities up

to 36 ppt, however, egg fertilization, hatching, and survival of larvae is highest at 12 g/l and

drops markedly after 18 g/l (Watanabe et al. 1997). Hence, it was not surprising that there was a

considerable drop in survival of the newly-hatched red tilapia larvae when the salinity was 20

ppt, which was on the 28th day of rearing. However, in freshwater conditions, the same pattern of

mortality was also observed. Although, survival rates of the red tilapia fry was comparable in

both freshwater and saline environments, the survival of the fry in increasing salinity was a little

bit higher than those reared in pure freshwater. Watanabe et al. (1997) and Liao and Chang

(1983) also observed equal to or better growth of red tilapias in seawater than in freshwater.

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The restlessness or erratic swimming behaviour of fish that are exposed to high salinities

indicates a fast rate at which the fish are approaching their tolerance limits to salinity (Lawson

and Anetekhai (2011). According to Xu et al. (2005), the feeding behavior of tilapia is altered or

they stay at the bottom of the tank when they are stressed due to their exposure to a wide salinity

gradient. In this study, the red tilapia hybrid fry did not exhibit sluggish or erratic swimming

patterns during gradual changes in the salinity of the rearing water, indicating tolerance to

salinity increments in the aquaria. Moreover, a good feeding behavior displayed by the fish

towards food during feeding time as shown by the high rate of feed consumption is an indication

that the metabolism of the fish was not impaired as a result of these gradual changes in salinity

levels (Lawson and Anetekhai, 2011).

Because of the comparable performance of newly-hatched red tilapia in terms of survival

and growth in both freshwater and in increasing salinity levels, the culture of this fish may be

viewed as a practical alternative to shrimp or milkfish farming particularly in areas where there

are limited sources of freshwater. Further studies are needed to validate the performance of this

hybrid during large-scale operations in ponds or cages in fresh- , brackish- or seawater in order

to confirm these results in the short-term study.

CONCLUSION

In conclusion, this study has shown that the tolerance of red tilapia hybrids to saline

waters begins early in its life history. Survival, growth and feeding activity of the newly-hatched

red tilapia hybrids were comparable in both freshwater and saline water, which may have

implications on their performance during the grow-out phase.

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ACKNOWLEDGEMENTS

This study was partly funded by the CHED-NAFES Extension Project, “Western

Philippines University Learning Environment-friendly Advocacy Farm and Family-based

Education Towards Fisheries Resource Management and Popularization of Aquaculture and

Alternative Agriculture in Western Philippines”. CMA Caipang is supported by the Balik

Scientist Program of the Department of Science and Technology, Philippines. The project staff

of the CHED-NAFES project and some personnel of the College of Fisheries and Aquatic

Sciences of WPU assisted in the conduct of this experiment. Dr. Roger Dolorosa and Dr.

Herminie Palla have significant contributions to improve the manuscript.

REFERENCES:

Basiao, Z.U., Doyle, R.W. 1990. Interaction between test and reference populations when tilapia

strains are compared by the ‘internal control’ technique. Aquaculture, 85: 207–214.

Behrends, L. L., Nelson, R.G., Smitherman, R.O., Stone, N.M. 1982. “Breeding and Culture of

Red-Gold Color Phase of Tilapia”. Journal of the World Mariculture Society, 13: 210-

220.

Carberry, J., Hanley, F. 1997. Commercial intensive tilapia culture in Jamaica”. Pp. 64-67. In:

Alston, D.E., Green, B.W., Clifford, H.C. (eds.) IV Symposium on Aquaculture in

Central America: Focusing on Shrimp and Tilapia, 22-24 April 1997, Tegucigalpa,

10

Honduras. Asociacion Nacional de Acuicultores de Honduras and the Latin American

Chapter of the World Aquaculture Society.

Clark, A.E., Watanabe, W.O., Olla, B.L., Wicklund, R.I. 1990. Growth, feed conversion and

protein utilization of Florida red tilapia fed isocaloric diets with different protein levels in

seawater pools. Aquaculture, 88:75-85.

Cnaani, A., Hulata, G. 2011. Improving salinity tolerance in tilapias: past experience and future

prospects. Israeli Journal of Aquaculture-Bamidgeh, 63: 1-21.

Eguia, M.R.R., 1996. Reproductive performance of four red tilapia strains in different seed

production systems. Israeli Journal of Aquaculture—Bamidgeh, 48: 10–18.

Eguia, M.R.R., Eguia, R.V. 1993. Growth response of three Oreochromis niloticus strains to feed

restriction. Israeli Journal of Aquaculture—Bamidgeh, 45: 8–17.

El-Sayed, A.M. 2006. Tilapia culture in salt water: Environmental requirements, nutritional

implications and economic potentials. Eighth Symposium on Advances in Nutritional

Aquaculture. November 15–17, Nuevo Leon, Mexico. pp. 95-106.

Ernest, D.H., Watanabe, W.O., Ellingson, L.J., 1991. Commercial-scale production of Florida

red tilapia seed in low- and brackish-salinity tanks. Journal of the World Aquaculture

Society, 22: 36–44.

Galman, O. R., Avtalion, R.R. 1983. A preliminary investigation of the characteristics of red

tilapias from the Philippines and Taiwan. Pp. 291-301. In: Fishelson, L., Yaron, Z. (eds.)

International Symposium on Tilapia in Aquaculture, Tel Aviv University Publisher, Tel

Aviv, Israel.

11

Galman, O.R., Moreau, J., Avtalion, R.1988. Breeding characteristics and growth performance

of Philippine red tilapia. Pp. 169-175. In: Pullin, R.S.V., Bhukaswan, T., Tonguthai, K.,

Maclean, J.L. (eds.) The Second International Symposium on Tilapia in Aquaculture,

ICLARM Conference Proceedings 15, Department of Fisheries, Bangkok, Thailand and

International Center for Living Aquatic Resources Management, Manila, Philippines. 623

p.

Gonzalez, C. E. 1997. La piscicultura en Colombia: technologia de punta en el Departamento

del Valle del Cauca”. Pp. 78-84. In: Alston, D.E., Gree, B.W., Clifford, H.C. (eds.) IV

symposium on aquaculture in Central America: focusing on shrimp and tilapia, 22-24

April 1997, Tegucigalpa, Honduras. Asociacion Nacional de Acuicultores de Honduras

and the Latin American Chapter of the World Aquaculture Society.

Green, B. W. 1997. Inclusion of tilapia as a diversification strategy for penaeid shrimp culture.

Pp. 85-93. In: Alston, D.E., Gree, B.W., Clifford, H.C. (eds.) IV symposium on

aquaculture in Central America: focusing on shrimp and tilapia, 22-24 April 1997,

Tegucigalpa, Honduras. Asociacion Nacional de Acuicultores de Honduras and the Latin

American Chapter of the World Aquaculture Society.

Holliday, F.G.T., Jones, M.P. 1967. Some effects of salinity on the development of eggs and

larvae of the Plaice (Pleuronectes platessa). Marine Biological Association of UK, 47:

39-48.

Hulata, G., Karplus, I., Harpaz, S. 1995. Evaluation of some red tilapia strains for aquaculture:

growth and color segregation in hybrid progeny. Aquaculture Research 26: 765-771.

12

Iqbal, K.J., Qureshi, N.A., Ashraf, M., Rehman, M.H.U., Khan, N., Javid, A., Majeed, H. 2012.

Effect of different salinity levels on growth and survival of Nile tilapia (Oreochromis

niloticus). The Journal of Animal & Plant Science, 22: 919–922.

Jaspe, C.J., Caipang, C.M. 2011b. Small-scale hatchery and larval rearing techniques for local

strains of saline-tolerant tilapia, Oreochromis spp. ABAH Bioflux, 3: 71-77.

Jaspe, C.J., Caipang, C.M. 2011a. Increasing salinity tolerance in tilapias: selective breeding

using locally available strains. AACL Bioflux, 4: 437-441.

Lawson, E.O., Anetekhai, M.A. 2011. Salinity tolerance and preference of hatchery reared Nile

tilapia, Oreochromis niloticus (Linneaus 1758). Asian Journal of Agricultural Sciences,

3: 104-110.

Lemarié, G., Baroiller, J.F., Clota, F.., Lazard, J., Dosdat, A. 2004. A simple test to estimate the

salinity resistance of fish with specific application to O. niloticus and S.melanotheron

Aquaculture, 240: 575–587.

Liao, I.-C., Chang, S. 1983. Studies on the feasibility of red tilapia culture in saline water”. Pp.

524-533. In: Fishelson, L., Yaron, Z. (eds.) International Symposium on Tilapia in

Aquaculture. Tel Aviv University Publisher, Tel Aviv, Israel.

Little, D. 1998. Options in the development of the aquatic chicken. Fish Farmer July/August. pp.

35–37.

Lutz, C.G. 2001. Practical Genetics for Aquaculture. Fishing News Books, Blackwell Sciences,

Oxford, United Kingdom.

Nugon, R.W. 2003. Salinity tolerance of juveniles of four varieties of tilapia. M.Sc. Thesis,

Louisiana State University, USA, 76 p.

13

Pruginin, Y., Fishelson, L., Koren, A. 1988. Intensive tilapia farming in brackishwater from an

Israeli desert aquifer. Pp. 75-82. In: Pullin, R.S.V., Bhukaswan,T., Tonguthai, K.,

Maclean, J.L. (eds.). The Second International Symposium on Tilapia in Aquaculture.

International Center for Living Aquatic Resources Mangement Conference Proceedings.

ICLARM, Makati, Manila, Philippines.

Pullin, R. 1982. The Biology and culture of tilapia. ICLARM, Philippines, 432 p.

Romana-Eguia, M. R. R., Eguia, R.V. 1999. Growth of five Asian red tilapia strains in saline

environments. Aquaculture, 173: 161-170.

Steffens, W. 1989. Principles of Fish Nutrition. Ellis Horwood Ltd., Chichester, West Sussex,

UK.

Vadhel, N.P., Akolkar, N.G., Pathan, J., Tandel, J.T., Lende, S., Shrivastava, V. 2017. Red

tilapia: a candidate euryhaline species for aqua farming in Gujarat. Journal of

FisheriesSciences.com, 11: 48-50.

Watanabe, W. O., Olla, B.L., Wicklund, R.I., Head, W.D. 1997. Saltwater culture of the Florida

red tilapia and other saline-tolerant tilapias: a review. Pp. 55-141. In: Costa-Pierce, B.

A., Rakocy, J.E. (eds.). Tilapia Aquaculture in the Americas, Vol. 1. World Aquaculture

Society, Baton Rouge, Louisiana, USA.

Watanabe, W.O., Clark, J.H., Dunham, J.B., Wicklund, R.I., Olla, B.L. 1990. Culture of Florida

red tilapia in marine cages. the effect of stocking density and dietary protein on growth.

Aquaculture, 90: 123-124.

Watanabe, W.O., Smith, S.J., Wicklund, R.I., Olla, B.L., 1992. Hatchery production of Florida

red tilapia seed in brackish water tanks under natural-mouth brooding and clutch-removal

methods. Aquaculture 102, 77–88.

14

Xu, J., Miao, X., Liu, Y., Cui, S. 2005. Behavioral response of tilapia (Oreochromis niloticus) to

acute ammonia stress monitored by computer vision. Journal of Zhejiang University

Science B, 6: 812–816.

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FIGURE LEGENDS

Figure 1. Survival of the newly-hatched red tilapia hybrids reared in freshwater (Trt 1) and in

gradual increase in salinity (Trt 2) over a period of 42 days. Red tilapia fry reared in Trt 2

were exposed to weekly increments of 5 ppt in water salinity. N=5.

Figure 2. Feed consumption rate (%) of the newly-hatched red tilapia hybrids reared in

freshwater (Trt 1) and in gradual increase in salinity (Trt 2) over a period of 42 days.

N=5.

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