Comparative effects of a natural androgen, 11β hydroxyandrostenedione, and a synthetic androgen, 17α-methyltestosterone, on the sex ratios of Oreochromis niloticus Jean-François Baroiller, Aboubacar Toguyeni

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Jean-François Baroiller, Aboubacar Toguyeni. Comparative effects of a natural androgen, 11β hydrox- yandrostenedione, and a synthetic androgen, 17α-methyltestosterone, on the sex ratios of Oreochromis niloticus. The Third International Symposium on Tilapia in Aquaculture, 41, ICLARM, 574 p., 1996, ICLARM Conference Proceedings, 971-8709-42-8. ￿hal-02842698￿

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HAL is a multi-disciplinary open access L’archive ouverte pluridisciplinaire HAL, est archive for the deposit and dissemination of sci- destinée au dépôt et à la diffusion de documents entific research documents, whether they are pub- scientifiques de niveau recherche, publiés ou non, lished or not. The documents may come from émanant des établissements d’enseignement et de teaching and research institutions in France or recherche français ou étrangers, des laboratoires abroad, or from public or private research centers. publics ou privés. 238 Comparative Effects of a Natural Androgen, 11 B-Hydroxyandrostenedione, and a Synthetic Androgen, 17a-Methyltestosterone, on the Sex Ratios of Oreochromls nllotlcus

J.F. BAROILLf.R A. TOGUYENI Programme aquaculture et péche du Centre de coopération internationale en recherche agronomique pour Je développement, Département d'élevage et de médecine vétérinaire (CIRAD-EMVTJ, BP 5095, 34033 Montpell/er Cedex 1, France Laboratoire de physiologie des poissons de l'Institut national de la recherche agronomique (INRA), Campus de Beaulieu 35042 Rennes Cedex, France et Département piscicole de l'Institut des Savanes (IDESSA) BP 621, Bouake 01, Côte d'ivoire

BAROILLER, J.F. and A. TOGUYENI. 1996. Comparative effects of a natural androgen. 11 B­ hydroxyandrostenedione, and a synthetic androgen, 17u-methyltestosterone, on the sex ratios of Oreochromis niloticus, p. 238-245. ln R.S.V. Pullin, J. Lazard, M. Legendre, J.B. Amon Kothias and D. Pauly (eds.) The Third International Symposium on Tilapia in Aquaculture. ICLARM Conf. Proc. 41, 575 p.

Abstract

Synthetic androgens are often considered to be more potent than natural androgens For sex reversai treatments in fish. Preliminary experiments on the natural androgen. 11 B­ hydroxyandrostenedione ( 11 B-OHti.4), recently identified in the gonads of fry of O. nllot/cus ln the early stages of testicular ontogenesis, showed a high masculinizing potential. Fry of 10-14 days post-fertilization (dPF) were produced from single pair matings using untreated females and either normal or sex reversed males, to assess any significant deviation in sex ratio compared to the controls. The fry were then treated during a minimum perlod of 21 days either wlth the natural or synthetic androgen added to the feed at different concentrations. Complete sex reversai was achleved with both steroids. Ali-male populations were produced ln elght groups treated with 1 0-35 µg 1 1 B-OHti.4· g·1 of feed and in two groups treated with 1 7 u-methyltestosterone ( 17 u-MT) at 5 and 20 µg·g·' of feed, respectlvely. There was no significant difference in the potency of the two androgens for concentrations higher or equal to 10 µg·g·•. However. at 5 µg· g·•. only 17u-MT produced 100% male populations whereas 11 B-OHti.4 produced 88.9%. ln contrast, the lowest 1 concentration of 1 1 B-OHti.4 ( 1 µg· g· ) significantly deviated sex ratios whereas 17u-MT had no effect compared to the controls. The natural androgen 11 B-OHti.4 can therefore constltute an alternative treatment to synthetlc sterolds. Low concentration treatments can be optimized by increasing the du ration of treatment.

Introduction mis, Sarotherodon and Ti/apla) constitute, with the cyprinids and salmonids, one Today, with an annual global of the three most important groups of production of approximately 500,000 freshwater fish for aquaculture. However, tons ( Lazard 1990). tilapias ( Oreochro- in culture conditions where competition 239 for food is high, the highly efficient human consumption is still prohibited reproduction of the species of in many countries (France and the United Oreochromis (Baroiller and Jalabert 1989) Kingdom, for example) whlch consider and their early sexual maturity lead to that the treatment and the effects of overcrowding and stunting, resulting in the synthetic steroid waste products are limited economic yîelds for fish farms. stîll insufficiently understood, especially A solution to this problem consists in their ecological impact. producing monosex populations. Ali­ ln tilapias, as in ait teleost fish, no male populations are preferred because definite physiological proof supports the theîr growth performance is better hypothesis of Yamamoto ( 1969) that compared to females (Pruginin 1967: steroids are the natural inducers of dif­ Hickling 1968; Hanson et al. 1983). ferentiation (Adkins-Regan 1987). The Currently, two techniques are used modification of the natural process of to produce monosex populations for fish sexual differentiation by exogenous culture (Baroiller and Jalabert t 989): steroids could be due to pharmacological Ma.nuai sexing. Based on the sexual effects (Reinboth 1970). ln fact, few dimorphism observed in the urogenital studies have investigated early papilla, this technique entails the elimi­ steroidogenesis during the gonadal sex nation of ail females, i.e., approximately differentiation of gonochoristic flsh (van half of the initial population as soon as den Hurk et al. 1982: Rothbard et al. possible (after a two to three month nurs­ 1987; Baroiller 1988a and b; Baroiller ing period). Manual sexing, which is used et al. 1988). in Africa, is time-consuming: it also re­ ln Oreochromis niloticus, early steroi­ quires qualified personnel, and includes dogenous potential in male and female 3- 10% errors (Lazard 1980: Chervinski gonads has been analyzed during the and Roth bard 1982) ..ln addition, this first three months of their life; this pe­ technique requires rearing a population riod covers the entire process of testicu­ of fry for two to three months after which lar and ovarian differentiation (Baroiller half (the females) will be eliminated. Al­ et al.. in press). Testosterone can be though manual sexing is simple. it is synthesized by the gonads of both sexes expensive in terms of time and tabor whereas oestradiol is exclusively pro­ and implies the underutilization of farm­ duced by the ovaries (Baroiller 1988b). ing infrastructures and lower feed pro­ ln contrast, certain androgens like t 1 B­ ductivity. hydroxyandrostenedlone ( 1 1 B-OH~4) Hormonal sex reversai. This technique and adrenosterone prove male sex-spe­ consists in masculinizing the entire popu­ cific during the same period (Baroiller lation of fry by adding a steroid to the 1988a and b) and show masculinizing feed for a short period (Guerrero 1982: potentialities (Baroiller 1988b). Hunter and Donald son 1983; Pandian Artificial steroids are generally more and Varadaraj 1987; Baroiller and Jalabert patent in their masculinizing effects than 1989). Hormonal sex reversai has been natural androgens (Hunter and Donaldson commonly used for several decades by 1983). The potency of 17a­ a number of tilapia producing countries methyltestosterone ( 1 7a-MT) is attrib­ such as Israel, Taiwan and the Philip­ uted to the presence of the 1 7 a.-me­ pines. This technique requires the sys­ thyl group which makes its elimination tematic treatment of every new popu­ slower than in natural androgen like lation of fry. However, the use of hor­ testosterone (Fagerlund and McBride mones in the production of animais for 1978; Donaldson et al. 1979). 240

A study of the masculinizlng poten­ the fertilization date and the marks on tial of 11 B-OH~4 and 17a.-MT was con­ the respective parents, was divided into ducted in O. niloticus to compare the two to five batches of at least 1 OO fry respective performances of both artifi­ reared separately in 2.00-1 aquaria. cial and natural hormones; to test the hypothesis of a possible role of t l B- Hormone Treatment 0Hà4 in the process of testicular dif­ ferentiation; and also to study possi­ Steroids were administered through ble alternatives to traditional sex reversai the feed. A first feeding salmonid feed treatments. (Aqualim) was impregnated with an alcohol solution contalning steroids. Concentrations of 1 to 45 ug of ster­ Materlals and Methods oids per gram of feed were tested. For the contrai batches, the same prepa­ Animais Tested ration was applied except that the feed did not contain any steroid. Two types of male broodfish of O. Fry were fed ad libitum six times dally nllot/cus of the "Bouake" strain (Baroiller using an automatic feeder during the 1988b) were used for the production 12-hour photoperiod, seven days a week. of fry families: normal males (XV) and Fry at 10-15 dPF were treated th us sex reversed males (XX). The latter came for 45 and 21 days. To avoid a poten· from two related families and yielded, tial masculinizing effect from the tem· for most of them, significant propor­ perature (Baroiller et al., in press), the tions of males in their progenies pro­ water, which was filtered and aerated, duced by single pair matings (Baroiller. was maintained at 28± t .5°C. this vol.). Nurslng and Sexlng the Fry Breedlna At the end of the treatment, fry aider Single male broodfish were placed in than 31 dPF were transferred to 1.5- 400-1 aquaria with normal females at a m3 outdoor tanks where they were fed sex ratio of 4: 1. The water in the_breeding the same diet until sexing was possi­ aquaria was filtered and maintained at ble. At 60-90 dPF, when the histologi­ a constant temperature of 27°C. Each cal differentiation of the male and fe· animal was identified by a mark inserted male gonads had already taken place in the dorsal muscles. Reproduction was (Baroiller 1988a and b). ail fry were sexed detected at the onset of maternai by microscopie examination of the gonad mouthbrooding behavior which is char­ squash (x 125). The presence of pre­ acterized by a dilation of the mouth. vitellogenic (auxocytosis) or vitellogenlc On the first day of incubation, ail other oocytes, and the lobular configuration individuals were removed to leave the showed the female and male sexes, re­ mouthbrooding female on her own in spectively. the breeding aquaria. Five days after A x2 test was used ta compare the hatching. i.e., nine days post-fertiliza­ sex ratios of the batches that had re­ tion (dPF). the fry were removed from ceived treatment and the control batches the mouth. Each brood, identlfled by (a .. Q.05). 241

Results OH.6.4) and at 5 µg·g·1 ( l 7a-MT). With concentrations lower or equal to 5 µg After treatment and at the moment of 118-0H.6.4 per gram of feed admin­ of sexing, there was no significant dif­ istered over a period of 21 days, the ference in the survival rate of the fry percentage of males was proportlonal between the contrais and the batches to the concentration used (Fig. t ). that underwent hormonal sex reversai, There was no significant difference in regardless of the treatment duration and masculinizing potency between the two the concentrations used (Table 1 ). hormones for concentrations of t 0-45 1 1 The microscopie examination of the µg·g· • With concentrations of 5 µg·g· gonad squashes of fry treated with l 1 B­ and higher, significant differences were OH.6.4 ( 1 ,63 t sexed animais) or t 7a­ observed in the results of both treat­ 1 MT (4 t 6 fry) did not show any hermaph­ ments: using 5 µg·g- , only 17u-MT rodite characteristics, sterilîty, or struc­ yielded 100% male populations against tural abnormality. The gonads of the a maximum of 88% wîth 1 t B·OH.6.4. ln­ individuals treated with 11 B-OH.6.4 were versely, t 7o.-MT was not effective if used 1 functional, irrespective of theîr geno­ at l µg·g- , whereas l ts-OH.6.4 used at type: functional sex reversed male XX the same concentration signifîcantly and normal males were obtalned and altered the percentage of males com­ identified by progeny testing at the end pared to the contrais. of these treatments (Baroiller, this vol.). Replicates (2-4) were conducted for Of the 1 7 batches treated with l l B­ four different concentrations of 11 B­ 1 OHA4, only one batch showed no de­ OH.:\4 (5, 20, 30 and 35 µg·g- ) (Table viation of sex ratio compared to the 2). There was no signiflcant difference controls (Table 2). The progeny of the in sex ratios between repllcates of the XY4 male underwent hormonal sex re­ same treatment. versai only after treatment 15 dPF whereas the t 3 other families received treatment after 10-14 dPF. Since sex Discussion reversai was produced in the four other 1 batches also treated with 5 µg·g· , this The hormones used for masculiniza­ concentration did not seem to be the tion of flsh are generally artificial mol­ cause of absence of a deviation in sex ecules derived from testosterone: 170.­ ratio. This result could instead indicate MT, 17u-ethynyltestosterone, dihydro­ a critical period of hormonal sensitiv­ testosterone acetate and testosterone îty. propionate. These synthetk androgens Ali treatments using l t 6-0HA4 on are considered more patent than natu­ Fry of Iess than 1 5 dPF signîficantly af­ ral androgens for the hormonal sex re­ fected the sex ratio towards the male versai of gonochoristic teleost fish spe­ sex compared to the controls, regard­ cies (Hunter and Donaldson 1983). less of the concentration used (Table Many authors have also produced 2). l 00% male tilapia popùlations using ln contrast, 17u-MT had masculînizing these steroids, despite a great hetero­ effects only in a range of concentrations genelty ln the experimental conditions: 1 between 5 and 45 µg· g- • especially concentrations varying from The 21-day treatment yielded ail-male l 0 to 240 µg· g-1 of feed (Baroiller and populations using hormonal concentra­ jalabert 1989). The optimum concen­ 1 tions between 10 and 35 µg·g- (1 tB- trations usually suggested are 30 µg·g-1 242

Table 1. Survival of Oreochromi5 nllotlcus fry according to hormone treatment applled for sex reversai.

Treatment Survival Number Survival Number after treatment of batches at sexing of batches (%) tested (%) te:;ted

11 B-0Hii4 83.8 15 55.2 17 17a-MT 85.6 4 56.7 4 Control 79.6 12 50.2 14

Table 2. Sex reversai treatments admlnistered to Oreochromis niloticus fry, the progeny of normal male (XYJ or sex rever$ed male broodfîsh pairs.

Treatment charaderlstlcs Sexing characterlstlcs

Broodfish Steroid Level in Initial age Final Duratlon Number Number Males male type feed of progeny Age day$ of of (%) and no. (µg·g·') (dl'F) (dPF) males females

XYl 1 IB·OHA4 35 12. 57 45 81 0 100 Contrai 0 1 2. 57 45 45 55 45 XYZ 11 B-OHL'.4 35 12 57 45 92 0 100 Co nt roi 0 12 57 45 45 41 52 XX1 1 lB-OHL'.4 35 1 2. 57 45 128 0 100 Control 0 12 57 45 64 56 53 XX2 1 lB-OHL'.4 30 13 55 42 64 0 100 1 IB-OHM lO 13 55 42 51 1 98 Co nt roi 0 13 55 42 0 74 0 XX3 11 B-OH/..4 · 30 14 4Z 28 129 0 lOC 11B-0Htl.4 30 14 35 21 168 0 100 Control 0 14 42 28 47 69 41 XX4 11 B·OHt.4 30 12 54 42 112 2. 98 11B-0Htl.4 ZO 11. 54 4Z 103 99 Contrai 0 12. 54 42 0 86 0 xxs 11B·0Htl.4 20 11 32 21 74 0 100 17cx:-MT 2.0 11 3Z 21 110 0 100 Contrai O 11 3Z 21 23 BO 22. XX6 11 B·OHtl.4 10 10 31 2.1 100 0 100 17a-MT 10 10 31 21 172 99 Control 0 10 31 21 34 118 22 XX7 116-0HM 5 10 31 21 24 3 89 l 7a-MT 5 10 31 21 47 0 100 Contrai 0 10 31 21 6 21 22 xxs 11B-0Htl.4 5 11 32 21 79 30 73 Contrai 0 1 t 32 21 28 96 23 XX9 11B-0Ht.4 11 32 21 71 30 70 Control 0 11 32 21 16 69 19 XY3 l lfS-OHL'.4 5 l 1 32 21 46 20 70 Control 0 11 32. 21 15 70 18 XY4 116-0HM 5 1 5 36 21 79 30 73 Contrnl 0 15 36 21 71 70 70 XX10 115-0Ht.4 13 34 21 41 72 36 17a-MT 13 34 ZI 12 74 14 Control 0 13 34 21 11 59 15 dPF=days post-fertlllzatlon. 243

tested as masculinizing agents. Among these, two t t -oxygenated derivatlves, 100 administered through the rearing wa­ mi llf3-0H~4 ter and through the feed, respectively, 17"" ·MT 80 0 can modify the differentiation process Ill • Control in tilapia: adrenosterone causes a de­ ~ E struction of the ovarian structures in O. 0 60 niloticus at a concentration of 5 mg- 1- 1 QI Cl (Katz et al. 1976) and 11 ketotes­ 1 ~ tosterone, at 200 µg·g- , inhibits the for­ QI 40 e mation of the ovarian cavity in O. Il."' mossambicus but does not prevent the 20 formation of young oocytes that will sub­ sequently degenerate (Nakamura 1981). 0 However, considering the high levels 20 IO !> used, the hypothesis of toxlc or para­ 1 Hormone levels in feed (JJQ·g- ) doxical effects cannot be discounted (Hunter and Donaldson 1983). ln O. Fig. t. E.fflclency of hormonal sex reversai us­ niloticus, the rnasculinizing effects of ing dlfferent sterolds and different concentra­ adrenosterone have also been demon­ tions ln tllapla feed. strated after treatrnents through the feed at 45 µg·g·1 (Baroiller 1988b). for 17a-MT and 60 µg·g-1 for ethynyltes­ Until then, 11 B-OHLi4 had not been tosterone (Pandian and Varadaraj 1987; used in hormonal sex reversai treatment McGeachin et al. 1987: Rothbard et al. in tilapia. This natural androgen shows 1987; Barolller and Jalabert 1989). For a masculinlzing potency comparable to O. mossambicus, the minimum concen­ that of 1 7 a-MT at concentrations of 10- 1 tration for the production of ail-male 35 µg· g· • ln addition, deviations of sex batches using 17a-MT is 5 µg·g-1 (Pandian ratio are observed at concentrations 1 and Varadaraj 1987). Other artificial an­ lower than 5 µg·g· , which is not the drogens such as 17a-methyl-5- case for 17a-MT. androsten-3B-17B-diol (Varadaraj and The microscopie examination of the Pandian 1987) and rnibolerone (Guerrero gonads of fry treated for 60-90 days and Guerrero, this vol.) have also been dld not show any abnormality in the used. course of gonadal ontogenesis. Moreo­ The present study indicates that in ver, the functional reversai of the go­ O. niloticus, the minimum concentra­ nads was demonstrated by the identi­ tion of 17a-MT yielding a 100% male fication of the sex reversed males af­ population ls also 5 µg· g-1 for a 21-day ter progeny testing of the individuals treatment period; there was no devia­ treated in the present study (Baroiller, tion in sex ratio with concentration of this vol.). 1 µg·g-1. The steroid 1 1B-OHLi4 was identified As the steroidogenesis in the period ln vitro in three species of teleost fish of gonadal differentiation has not been in the early stages of differentiation: this extensively studied in teleost fish, par­ 11-oxygenated steroid can be specifi­ ticularly in tilapia (Baroiller 1988a and cally synthesized by the testes of the b; Baroiller et al. 1988), only a limited rainbow trout Oncorhynchus mykiss (van number of na.tura.I androgens have been den Hurk et al. 1982), the ca.tfish C!arias 244 gariepinus (van den Hurk et al. 1989) 1988a and b). In males, a highly pro­ and O. niloticus (Baroiller 1988b) dur­ gressive multiplication of somatlc cells ing the early gonadal ontogenesis. and spermatogonia is observed during Administered through the feed {60 the same period (Baroiller 1988 a and 1 and 6 µg·g- ) and through the rearing b). Exogenous hormones are therefore 1 water (300 µg·l- ), 11 B-0Htl4 produces administered to the fry before the es­ populations with high percentages of tablishment of the se histological proc­ males, yielding 76-78% in the trout (van esses. den Hurk and Lambert 1982; van den Hurk and van Oordt 1985) and 77% in References the catfish (van den Hurk et al. 1989), respectively, against 48 and 50% males Adkins-Regan, L 1987. Hormones and sexual in the contrais, respectively. ln C. differentiation, p. 1-29. ln D.O. Norris and (van den Hurk et al. 1989), R.E. Jones (eds.) Hormones and reproduc­ gariepinus tion in fishes, amphibians and reptiles. Ple­ 17a-MT significantly bîases the sex ratio num Publishers, New York. towards males (65%) at concentrations Barolller, J.F. 1988.a. Etude corrélée de l'apparition of 30 µg·t- 1 and towards the female sex des critères morphologiques de la dif­ 1 férenciation de la gonade et de ses at 1OO µg· 1- • ln Oncorhynchus myklss, potentlalltés stéroïdogènes chez Oreo­ testosterone derivatives may not be es­ chromls n/lotlcus. Université Pierre et Marle sential for testicular differentiation: Curie. Paris. 89 p. Ph.D. dissertation. testosterone and its 11-oxygenated Baroiller. J.L 1988b. Etude des processus morphologiques et endocrinologîques de derivative can be synthesized by the la différenciation naturelle du sexe chez testls only stages beyond those at which Oreochromis ni/oticus, appliquée à la pro­ 11 B-0Htl4 has been identified (van den duction de populations monosexes : intérêts Hurk et al. 1982). Furthermore, a treat­ et perspectives. p, 256-266. in G.M. Bernacsek and H. Powles (eds.) Aquaculture ment using cyproterone ac:etate did not research systems in Africa. Proceedings of affect the sex ratio of the batches of a Workshop in Bouake, Côte d'ivoire, 14- trout (van den Hurk and van Oordt 1985) 18 Novem ber 1988. International Devel­ and tilapia fry (Hopkins et al. 1979)_ opment Research Centre, Ottawa, Canada. Baroiller, J.F .. A. Fostierand B. Jalabert. 1988. Pre­ The androstenedione 11-oxygenated de­ cocious steroidogenesis in the gonads of rivatives may be involved in some stages Oreochromis niloticus du ring and after sexual of testicular differentiation ln these three differentiation, p. 137-141. ln Y. Zohar and species. B. Breton (eds.) Reproduction in fish. Basic and applied aspects in endocrinology and ge­ ln O. niloticus, response to the hor­ netics. Les Colloques de !'INRA no. 44. mone treatment is observed during a Baroiller. J.F. and B. Jalabert. 1989. Contribu­ determined critic:al period_ To be effi­ tion of research ln reproductive physiol­ cient, treatment must begin between ogy to the culture of tllaplas. Aquat. Liv­ ing Resour. 2:105-116. nlne and 13 dPF. Beyond this perlod, Baroiller, J.F., X. Rognon, V.C. Yapi and A. Cissé. dlfferentiation seems definitively accord­ Genetie characterization and sex determina­ ing to the genotype; from then on, ît tlon ln tllapla ln Côte d'ivoire. ln A.E. Eknath could only be affected by ex:ogenous and B.O. Acosta (eds.) Present progress. fu­ ture direction and needs in flsh genetlc:s re­ steroid factors. search in Asia, Pacifie and Africa. ICLARM The 2 1-day treatment is therefore ap­ Conf. Proc. 38. (ln press.} plled between 9-30 dPF. From a histo­ Chervinski, J. and S. Rothbard. 1982. An aid in logical perspective. at 27°C, oogonia manually sexlng tllapla. Aquaculture 26:389. Donaldson. E.M .. U.H.M. fagerlund, D.A. Higgs proliferatlon occurs in females between and J.R. McBride. 1979. Hormonal enhance­ 20-28 dPF and is followed by their flrst ment of growth, p. 456-578. ln W.S. Hoar. meiotic prophase at 28-35 dPF (Barolller D.J. Randall and J.R. Brett (eds.) Flsh physlology. 245

Vol. 8. Academlc Press. New York. on gonadal sex dlfferentlation in Tilapia Fagerlund, U.H.M and J.R. Mc8rlde. 1978. Ois­ mossambica. Bull. ]apan Soc. Flsh. Oceanogr. tributlon and dlsappearance of radloactlv­ 47:1321-1327. ity in blood and tissues of coho salmon Pandlan, T.J. and K. Varadaraj. 1987. Techniques (Oncorhynchus klsutch) after oral admin­ to regulate sex-ratio and breeding ln tllapla. istration of 3H-testosterone. j. Flsh. Res. Curr. Sei. 56(8):337-343. Board Can. 35:893-900. Pruglnln. ]. 1967. Report to the Government of Guerrero, R.D. 1982. Control of Tilapia repro­ Uganda on the experimental fish culture duction, p. 309-316. ln R.S.V. Pullln and proJect ln Uganda, 1965-1966. UNDP/FAO R.H. Lowe-McConnell (eds.) The biology Report No. TA 2446, 16 p. and culture of tilapias. ICLARM Conf. Proc. Relnboth, R. 1970. lntersexuality in fishes. Mem. 7. 432 p. Soc. E.ndocr. 18:515-544. Hanson, T.R., R.O. Smitherman, W.L. Shelton and Rothbard, S., B. Moav and Z. Yaron. 1987. Changes R.A. Dunham. 1983. Growth comparlson in steroid concentrations during sexual of monosex tilapia produced by separation ontogenesis in tilapia. Aquaculture 61 : 59- of sexes, hybridization and sex reversai, 74. p. 570-579. ln L. Fishelson and Z. Yaron van den Hurk. R. and J.G.D. Lambert. 1982. Tem­ (comps.) Proceedings of the First Interna­ perature and steroid effects on gonadal sex tional Symposium on Tllapia in Aquaculture, differentiatlon ln ralnbow trout, p. 69-7Z. Tel Aviv, Israel. ln C.J.J. Richter and H.J.Th. Goos (comps.) Hickling, C.F. 1968. Fish hybridizatlon. FAO Flsh. Proceedings of the International Symposium Rep. 44:1-lt. on Reproduction Physîology of Fish. PUDOC. Hopkins, K.O .. W.L. Shelton and C.R. Engle. 1979. Wageningen. Estrogen sex reversai of T!lapla aurea. van den Hurk, R. and P.G.W.J. van Oordt. 1985. Aquaculture 18:263-268. Effects of natural androgens and cortlcosteroids Hunter, G.A. and E.M. Donaldson. 1983. Hor­ on gonad differentîation ln the ralnbow trout, monal sex control and lts application to Salmo gairdneri. Gen. Comp. Endocrinol. flsh culture, p. 223-303. ln W.S. Hoar, D.J. 57:216-222. Randall and E.M. Donaldson (eds.) Fish van den Hurk. R.. C.J.J. Richter and J. janssen­ physlology. Vol. 9(8). Academic Press, New Dommerholt. 1989. Effects of 17a-methyl­ York. testosterone and 11 &-hydroxyand ros­ Katz, Y., M. Abraham and 8. E.cksteln. 1976. Effects tenedione on gonad differentlatlon in the of adrenosterone on gonadal and body growth Afrlcan catfish, Clarias gariepinus. ln T!lapla n//otlCil (Teleostei, Cichlidae). Gen. Aquaculture 83: 179-19 t. Comp. Endocrlnol. 29:414-418. van den Hurk, R., J.G.D. Lambert and j. Peute. Lazard, J. 1980. Le développement de la 1982. Steroidogenesîs in the gonads of pisciculture intensive en Côte d'ivoire. ralnbow trout fry Salmo ga/rdnerl before Exemple de la ferme pilote de Natio­ and after the onset of gonadal sex differ­ Kobadara (Korhogo). Notes et Documents entlatlon. Reprod. Nutr. Develop. ZZ:413- sur la Pêche et la Pisciculture Z 1: 1-44. 426. Lazard, J. 1990. Transfert de poissons et dévelop­ Varadaraj, K. and T.J. Pandian. 1987. pement de la production piscicole. Exemple Masculinization of Oreochromls mossam­ de trois pays d'Afrique sub-saharienne. Rev. b/cus by administration of 17a.-methyl-5- Hydrobiol. Trop. 23(3):251-265. androsten-3&-17&-dlol through rearing McGeachln, R., E.H. Robinson and W.H. Neill. water. Curr. Sei. 56(9):412-413. 1987. Effect of feedlng hlgh levels of an­ Yamamoto, T. 1969. Sex differentlation, p. drogens on the sexe ratio of Oreochrom/s 117-175. ln W.S. Hoar and D.J. Randall {eds.) aureus. Aquaculture 61 :317-321. Flsh physlology. Vol. 3. Academic Press, Nakamura, M. 1981. Effects of 11-ketotestosterone New York. ,.., ;

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International Center for Living Aquatic era . Resources Management Centre de recherches oceanologiques ) I r";f~~~31 L'lnstitut fran!;ais de recherche scientifique Centre de cooperation internationale en recherche pour Ie developpement en cooperation With the cooperation of agronomique pour Ie developpelTlent ,; =.So- 11.::1 c! 'fbA Cooperation Centre technique francaise de cooperation agricole et rurale ' ~- -. - -~ .,...... JJ.. ;;;;; - ~- - "" ~~...- ~- Tl;IÉ THIRD INTERNATIONAL SYMPOSIUM ON TILAPIA IN AQUACULTURE

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