Research Projects
Reproduction Control Research

Subcontract No. RD010A-09

Staff

Auburn University, Alabama

Ronald P. Phelps	US Principal Investigator, Project Leader
Ron Carpenter	Graduate Research Assistant (USA; from January 2000; CRSP funded)
Ryan Peasley	Graduate Research Assistant (USA; October 1999 through January 2000; CRSP funded)
Lee Warrington	Graduate Research Assistant (USA; through October 1999; CRSP funded)

Background

Limited knowledge of the reproductive physiology and breeding of culture species was identified as one of the key constraints to aquaculture in the Continuation Plan 1996–2001. Specifically, effective and practical control of reproduction is a major constraint in tilapia culture. Inter- and intraspecific breeding programs can result in populations with highly skewed sex ratios but often give inconsistent results. Interspecific crosses have not proven to be practical due to difficulties in maintaining the parent species integrity. Intraspecific breeding programs have been developed to exploit the sex inheritance mechanism in Nile tilapia, Oreochromis niloticus. The androgenetic approach to developing YY males simplifies the identification of YY males as all males produced should be of the YY genotype.

Broodstock and seed supply was also identified as a major constraint in the Continuation Plan 1996–2001, resulting in reproduction control becoming one of the CRSP research priorities. Another priority of the plan is identification of alternatives to dietary hormone treatment, the standard masculinization method. Much of the CRSP research effort has focused on tilapia, for which management of unwanted reproduction is an essential part of most culture systems. The objectives identified in the Eighth Work Plan include a series of studies which address this issue by determining whether there is an autosomal influence on the sex ratio of Nile tilapia and developing a pure YY line of male Nile tilapia. A Ninth Work Plan study examines the use of within- and cross-family breeding as an alternative method of developing YY tilapia and examines the effect of temperature on gonadal differentiation.

Work Plan Research

The following Eighth Work Plan investigation continued into the current reporting period:

• Methods for contribution from the male and female genome to sex inheritance/8RCR1C. The report submitted for this investigation was a final report.

This subcontract was awarded funding to conduct the following Ninth Work Plan investigations:

• Monosex tilapia production through androgenesis: Selection of individuals for sex inheritance characteristics for use in monosex production/9RCR6A. An abstract was submitted for this investigation.
• Monosex tilapia production through androgenesis: Verification of androgenically produced males, their viability, and the influence of the female on progeny sex ratios/9RCR6B. This investigation was cancelled.
• Monosex tilapia production through androgenesis: Growth of genetically derived males in production settings/9RCR6C. This investigation was cancelled.

Note: The methods for 8RCR1C have been modified. The revised methods will appear in the Third Addendum to the Eighth Work Plan. The decision to terminate 8RCR1D will be documented in the same addendum. Owing to delays, the scope of 9RCR6 was limited to 9RCR6A, Selection of individuals for sex inheritance characteristics for use in monosex production. The decision to terminate 9RCR6B and 9RCR6C will be documented in the Addendum to the Ninth Work Plan.

Publications

Phelps, R.P. and T.J. Popma, 2000. Sex reversal of tilapia. In: B.A. Costa Pierce and J.E. Rakocy (Editors), Tilapia Aquaculture in the Americas, Volume Two. The World Aquaculture Society, Baton Rouge, Louisiana, pp. 34–59.
Warrington, L., 2000. Sex ratio variation and sex determining mechanisms in Oreochromis niloticus. M.S. thesis, Auburn University, Alabama.

Methods for the Contribution from the Male and Female Genome to Sex Inheritance

Eighth Work Plan, Reproduction Control Research 1C (8RCR1C)
Final Report

Ronald P. Phelps and Richard Lee Warrington
Department of Fisheries and Allied Aquacultures
Auburn University, Alabama, USA

Abstract

The variability in the factors affecting the sex ratios of Nile tilapia (Oreochromis niloticus) was studied using pair spawns from the Egypt, Ghana, and Ivory Coast strains. Sex ratios from 129 progeny groups were determined by microscopic inspection of the gonads. Of the 12,450 progeny sexed, 54.14% were males; this differed significantly from a 1:1 sex ratio (P < 0.001). Sex ratios from the 129 progeny groups produced a normal distribution (P > 0.45) but did not reflect a binomial distribution (P < 0.01) as would be expected from a simple monofactorial sex determination process. Therefore, other factors must account for the variation observed in sex ratios. Single spawn sex ratios ranged widely from 16 to 100% male. Chi-square tests revealed weak correlations between strain and progeny gender, individual parent and progeny gender, and male-female parent combinations and progeny gender. Sixty-five percent (11 of 17) of the repeat spawns by the same pairings produced sex ratios that extended beyond a range of 10%. The continuous range of sex ratios within the normal distribution suggests the presence of several minor sex-modifying factors in O. niloticus and raises questions as to whether sex inheritance in tilapia is predictable enough for a YY breeding program to be practical.

Monosex Tilapia Production through Androgenesis: Selection of Individuals for Sex Inheritance Characteristics for Use in Monosex Production

Ninth Work Plan, Reproduction Control Research 6A (9RCR6A)
Abstract

Ronald P. Phelps
Department of Fisheries and Allied Aquacultures
Auburn University, Alabama, USA

Abstract

There is evidence that in domestic stocks of Nile tilapia (Oreochromis niloticus) sex ratios of individual pairs may vary from 50:50. Several authors have found wide variation in sex ratios from one single pair mating to another. The grand mean for the population might average 50:50, but individual pairs would produce sets of progeny where the male frequency may range from 5 to 100%. This variability in sex ratios is a challenge to the development of YY breeding programs, where sex determination is assumed to be controlled by the "Y" chromosome with no other factors involved. For YY production of males to be successful, variability in the percent males produced must be reduced and parent lines that conform to a simple Mendelian inheritance must be established.

Nine families of Nile tilapia based on single pair matings have been selected, each with a sex ratio that was either highly skewed to male or female, or that conformed closely to a 50:50 sex ratio. Matings within the families as well as across families are being conducted to determine the heritability of sex and the factors influencing it. Progeny from representative matings are being cultured in 45-l aquaria at temperatures of 27–28°C and 36°C during the period of gonadal differentiation to determine the effect of temperature on altering sex ratios and how that response may vary by family. A total of 80 within-family and 65 cross-family spawns have been obtained from the nine families. Additional pair spawns were made through 30 July 2000. The sets of progeny obtained to date are being grown to a sexable size to determine the frequency of each sex. Sex ratios from each set will be compared to those of other sibling matings and the frequency from the parent family.