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.

N-cadherin、MT1-MMP、ADAM17在甲状腺癌中的表达及其意义研究

目的探讨N-cadherin、MT1-MMP、ADAM17在甲状腺癌中的表达及其意义。方法选取2016年1月-2017年12月期间医院行手术治疗的103例甲状腺癌患者术后甲状腺癌组织标本为观察组,另选取同期在我院行活检的65例结节性甲状腺肿病灶旁正常甲状腺组织标本为对照组,并分别监测观察组和对照组患者N-cadherin、MT1-MMP、ADAM17表达情况,并与甲状腺癌患者临床病理特征进行比较和分析。结果观察组患者N-cadherin、MT1-MMP、ADAM17的阳性表达率明显高于对照组,差异具有统计学意义(P<0.05)。Ⅲ～Ⅳ期患者N-cadherin、MT1-MMP、ADAM17的阳性表达率明显高于Ⅰ～Ⅱ期,差异具有统计学意义(P<0.05)。浸及浆膜患者N-cadherin、MT1-MMP、ADAM17的阳性表达率明显高于未及浆膜,差异具有统计学意义(P<0.05)。分化程度为低分化患者N-cadherin、MT1-MMP、ADAM17的阳性表达率明显高于中、高分化,差异具有统计学意义(P<0.05)。发生肝脏转移患者N-cadherin、MT1-MMP、ADAM17的阳性表达率明显高于未发生肝脏转移,差异具有统计学意义(P<0.05)。发生淋巴结转移患者N-cadherin、MT1-MMP、ADAM17的阳性表达率明显高于未发生淋巴结转移,差异具有统计学意义(P<0.05)。结论 N-cadherin、MT1-MMP、ADAM17在甲状腺癌的发生和发展中发挥重要作用,同时对于判断病情和预后亦具有十分重要的参考价值。

17α-甲基睾酮对棘胸蛙生长发育及性别分化的影响

为研究17α-甲基睾酮(MT)对棘胸蛙生长发育及性别分化的影响,采用乙醇喷雾法处理的含0,10,30,50 mg/kg MT饵料投喂棘胸蛙蝌蚪至60 d,测定体重、体长并记录生长发育情况,继续投喂并记录蝌蚪变态、畸形、死亡数,之后通过组织切片鉴定棘胸蛙性别和性腺发育状况.结果显示:MT对棘胸蛙蝌蚪的生长与正常发育有一定的抑制作用,并可通过影响性腺发育提高棘胸蛙雄性率,对棘胸蛙蝌蚪具有雄性化效应.以上研究对棘胸蛙的高效养殖及单性养殖具有重要意义.

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.

激素诱导斜带石斑鱼(Epinephelus coioides)雄性化的研究

石斑鱼属雌雄同体雌性先成熟鱼类,低龄鱼卵巢先成熟,表现为雌性,高龄鱼才性转变为雄性。本实验通过持续跟踪取样,分析了人工养殖条件下斜带石斑鱼(Epinephelus coioides)在36月龄内的性腺发育过程,发现其性分化完成后,雌性卵巢在9月龄后开始发育。因此,采取投喂含有雄激素甲基睾酮(MT)饲料的方法,可以在石斑鱼卵巢发育之前,成功诱导8月龄的斜带石斑鱼幼鱼向雄性方向发育。结果表明,以MT 10mg/kg饲料的剂量投喂3个月能够有效诱导卵巢未发育的幼鱼往雄性方向发育;但停止投喂MT后,幼鱼精巢又开始出现退化,不会继续往雄性方向发展,而转变为雌性。以上结果表明,斜带石斑鱼性别决定和分化除了受遗传因子和内分泌调控外,还受到年龄、体型和群体、环境等因子的影响,人工投喂MT激素的作用并不能从根本上影响石斑鱼幼鱼的性腺发育的方向,只有在持续剂量的MT激素诱导下,石斑鱼幼鱼才能发育为成熟的雄鱼。