Determination of 17<i>α</i>-Methyltestosterone in Freshwater Samples of Tilapia Farming by High Performance Liquid Chromatography

17α-methyltestosterone is used to induce the sex reversal of Tilapia sp. to obtain cultures mono-sex to an economically viable. This practice may lead to environmental contamination and problems in human health. Therefore methods need to be developed to detect residues of 17α-methyltestosterone in aqueous matrices. A simple high-performance liquid chromatographic method using ultraviolet detection (245 nm) and testosterone as internal standard has been developed for the monitoring 17α-methyltestosterone in freshwater samples of tilapia aquaculture. The method described involves limited sample preparation as it includes a filtration followed by a single solid-phase extraction step using C18 cartridge. Validation data indicated that the HPLC-UV method for 17α-methyltestosterone determination in the concentration range of 50 - 2000 μg/L provided good linearity, sensitivity, accuracy and precision. Method performance was efficiently applied to monitoring the freshwater samples of fish ponds and the surrounding aquatic channels.

Residual impact of 17α-methyltestosterone and histopathological changes in sex-reversed Nile tilapia (Oreochromis niloticus)

Objective:To examine sex reversal both by oral and by immersion using 17α-methyltestosterone on the methyltestosterone residual concentration and the organ histopathology of tilapia fish.Methods:This study used oral and immersion treatment methods for sex reversal of tilapia fish and used normal fish as the control and each treatment was repeated 4 times.17α-methyltestosterone at dosages of 60 mg/kg feed and 0.5 mg/L were used for oral and immersion methods,respectively.In the first step,tilapia fry were reared at 100 L aquaria,with a density of 1 fish/L for 2 months.In the next step,male tilapias were reared at happa(net cage)of(2×1×1)m3 size in the controlled pond,with a density of 30 fish/happa for 3 months.The methyltestosterone residual concentrations were analyzed by one-way analysis of variance and Duncan's multiple range tests,while organ histopathology was analyzed by descriptive method.Results:Residual concentrations in the serum of methyltestosterone-treated fish were significantly lower than that in normal fish,especially in 4-and 5-month-old tilapias with averages of less than 5μg/L,while in normal fish was more than 5μg/L.In the flesh,methyltestosterone residual concentrations showed relatively no significant differences between the oral and immersion treatment groups and methyltestosterone-treated fish remained lower compared to normal fish,except in 5-month-old tilapia.Methyltestosterone-treated tilapia exhibited histopathological changes on gill,liver,kidneys,and intestine organs.Conclusions:Sex reversal either by oral or by immersion has methyltestosterone residual concentration,but does not exceed the limits(5μg/L or 5μg/kg)of synthetic steroid on the fish body,although methyltestosterone causes histopathological changes on gill,liver,kidneys,and intestine.

Sexual Reversal with 17α-Methyltestosterone in Oreochromis sp.:Comparison between Recirculation Aquaculture System(RAS)and Biofloc Technology(BFT)

Precocity in tilapia implies the use of several methods of obtaining monosex seed;the most common tends to use masculinizing hormone 17α-methyltestosterone(17αMT),with variable results.Thus the objective of this study was to compare the efficiency of the sexual reversion process using 17αMT,in a recirculation system and in biofloc.In a totally randomized design,three tanks for recirculation(T-RAS)and three tanks for biofloc(T-BIO)with a capacity of 200 L effective volume were taken and filled with 1,056 larvae of Oreochromis sp.,without reversing and with an initial weight of 0.02 g and an initial total length of 1.4 mm.The study was carried out during 65 d,the fish were fed(10%biomass,adjusted every 15 d)by a commercial diet at 45%of crude protein that included 17αMT(60 mg/kg).Water quality,microbiology,zootechnical and gonadal analysis were monitored.Consequently the water quality results showed that just dissolved oxygen(DO),temperature(T-°C)and alkalinity did not show significant differences.Additionally,in the productive parameters there were significant differences in the final length,the gain in length and in K which were better in T-BIO.The microbiological ones did not present significant differences between the treatments.Lastly,the percentage of reversion was significantly better in T-RAS.Then,this study suggests that settleable solids concentrations above 35 cm decrease the efficiency of the sexual reversion for this species.

The optimum dose and period of 17α-methyltestosterone immersion on masculinization of red tilapia(Oreochromis spp.)

This study was carried out to examine the effect of combining factors for immersion technique on masculinization of red tilapia(Oreochromis spp.).Three selected doses(1000,1,500,and 2000μg/L)of 17α-methyltestosterone(17 MT)immersion,2 exposure times(3 and 6 h)once or twice a week,and interaction of these factors were tested.A total of 12 treatment combinations under 3×2×2 factorial experiments in randomized complete block design were conducted to compare with the negative control group(no 17 MT-treated)and positive control group(17 MT-treated feed administration).One hundred red tilapia larvae(14 days post-hatch)were randomly allocated in each treatment and were operated in a 10-L glass jar for 21 days.At the end of the treatment regime,the survival rate was determined.Then,fish were raised in a 150-L glass tank using normal commercial feed for 2 months and sex ratios were determined via the gonad squash technique.None of the immersion treatments tested resulted in a statistically significant(p>0.05)decrease in survival rate.All 17 MT-treated groups produced progenies with a significantly higher(p<0.001)proportion of males than the predicted 1:1 sex ratio.Influences of some factor combinations in the masculinization of red tilapia were observed.The male ratio was significantly affected by the dose-period and dose-frequency treatments(p<0.05).The highest male proportion(95.33%±0.58%)was recorded at 1000μg/L of 17 MT for 6 h once a week which was significantly higher(p<0.05)than other groups.The production cost of the best group was the cheapest(THB0.0066/fish)and decreased from the traditional procedure by about 10.81%.Therefore,this technique could be implemented for decreasing cost and operating process in commercial tilapia farms.

Production of XX neo-male mandarin fish, Siniperca chuatsi, after sexual differentiation by oral administration of 17α-methyltestosterone

A well-known fact about mandarin fish(Siniperca chuatsi)is that females grow faster than males.Mandarin fish that have all-female populations have economic advantages,but traditional neo-male fish are difficult to produce.In this study,indirect feeding of 500 mg/kg 17α-methyltestosterone(MT)to XX mandarin fish(60 dpf,days post fertilization)with completely differentiated ovaries induced secondary sex reversal.At 40 days after MT treatment,14.7%of mandarin fish with complete reversal was produced,which showed for the first time that XX mandarin fishes with differentiated ovaries still have potentially bisexual germ cells and the plasticity of sex reversal.In the MT treatment group,there was no significant difference in survival compared to the control group.The body weight of the MT treatment group was significantly lower than that of the control group 40 days after treatment(dat)(P<0.05).During MT treatment,serum 11-KT and T concentrations increased and serum E2 concentration decreased in the MT treatment group,but there was no significant difference in hormone levels between the MT treatment group and the control group.A masculinization of mandarin fish was also confirmed by the expression patterns of sex-specific genes sox9,dmrt1,cyp19a1a and foxl2.A significant amount of sox9 and dmrt1 was expressed in the fish gonads of the MT treatment group,whereas cyp19a1a and foxl2 were minimally expressed in the fish gonads.

AtMET1基因克隆及化学诱导表达分析

DNA甲基化是植物基因组中普遍存在的一种的重要表观遗传学机制,对植物生长发育及进化起着重要的调节作用[1-2]。植物体DNA甲基化的建立和维持受多个基因的协同调控,其中MET1（DN-MT1-like METHYLTRANSFERASE 1 gene）是在植物中最早分离出来的甲基化相关基因,编码DNA甲基化转移酶1（MET1,Methytransferase1）,该酶主要负责保持CG位点的甲基化,通过DNA甲基化作用影响植物的基因组结构、发育和进化。