THE SCIENTIFIC BASIS OF MATURATION CULTURE AND TECHNOLOGY OF WHISKER CATFISH REPRODUCTION (micronema bleekeri)

The study of whisker catfish rearing from fry to fingerling Experiment 1: The study of whisker catfish rearing to 30 th day using live food... The experiment was completely randomized

CAN THO UNIVERSITY

-♣♣♣ -NGUYEN VAN TRIEU

THE SCIENTIFIC BASIS OF

MATURATION CULTURE AND TECHNOLOGY OF WHISKER CATFISH

REPRODUCTION (Micronema bleekeri)

Specialization: Freshwater Aquaculture Field of Study Code: 62 62 60 01

Dissertation of Doctor Degree in Aquaculture

Cantho, 2014

Can Tho University

Supervisors:

Ass/Prof Dr Nguyen Anh Tuan

Ass/Prof Dr Duong Nhut Long

Examiner 1:

Examiner 2:

The dissertation will be defended at the university committee in … at: ….… hr ….… date …… month …… year 2014

The disseratation is available in Libraries:

Learning Resource Center of Can Tho University

College of Aquaculture, Can Tho University

AN OVERVIEW OF THE THESIS

1 Imperative thesis

Fresh water aquaculture which is a position of strength in the Mekong Delta includes mainly objects such as tra catfish, climbing perch, snakehead,… However, in recent years, aquaculture has had difficulties because prices which are not usually stable Therefore, farmers have found some new culture species As assessment of several farmers both An Giang and Dong Thap provinces that

whisker catfish is a species which has had high economic values and

prospects for developmental aquaculture, especially in cages culture Fingerlings which have been depending on nature do not guarantee quality and quantity These study results about whisker catfish have provided basic data about biological characteristics and technology of artificial reproduction The contribution of building the process of

whishker catfish reproduction which is: “The scientific basis of maturation culture and technology of whisker catfish

reproduction (Micronema bleekeri Gunther, 1864)” was carried

out

2 General objectives of thesis

To provide scientific data about the technology of gonadosomatic conditioning and whisker catfish reproduction, contributing to production of fingerlings initiatively both quality and quantity for farmers, and objecting diversification and sustainable development of aquaculture in the Mekong Delta

3 Significance of scientific and reality

In the scientific aspect, these studies have provided scientific information about the effective feed to martunity of whiker catfish

Besides, thesis has also studied the capacity of M bleekeri spawning

by different hormones and technology of whisker catfish rearing from fry to fingerling stages

4 New results of thesis: The first in identifing:

Whisker catfish which was conditioned in ponds by trash tiny shrimp will have been maturity in May and June with gonadosomatic index

(about 3.8 ± 0.08%) in females and relative fecundity (about 110 ± 9.1 eggs/g female) Through conditioning process that Vitellogenin (Vg) in plasma of female will change proportional with the development stages of gonad Vg content increased the highest when gonad has changed from III to IV stage and had closely correlation between Vg content and the development of egg diameter in times

Spawning stimulation of M bleekeri by pituitary with 3.5 mg/kg

female; LRHa + Dom (70 µg) + 3.5 mg/kg female; and Ovaprim which was used with 0.3 mL/kg so that fish has had spawning, besides the rates of fecundity, fertilization and hatching was relatively high Whereas, spawning stimulation by HCG (4,000 – 6,000 UI/kg female) so that 100 % of fish did not ovulate

M bleekeri started first feeding after two days From 2 to 5 days, fish

was fed by rotifera and copepoda larvae, through 6 – 30 days fish

was fed by Cladocera and Copepoda The rearing of M bleekeri by

tubifex with 3.5 individuals/L so that daily weight gain (20.2 ± 0.25 mg/day) and survival rate (88.9 ± 3.2 %) were relatively high Whisker catfish was also used well home-made feed at 7th day

Protein requirement of M bleekeri fingerlings (sizes of fish about

269 mg) was 43.2 % The rearing of whisker catfish by pellet (36 % crude protein) with density of 3.5 fingerlings/L at 60 days; which has achieved survival rate and daily weight gain were 81.2 ± 3.5 % and 25.9 ± 0.25 mg/day, respectively

THE STUDY METHODS 3.1 The times and location of study

The study was carried out from 2007 to 2011 at the College of Aquaculture and Fisheries, Can Tho University

3.2 Study object was whishker catfish Micronema bleekeri

3.3 Study methods

3.3.1 Effective feed to maturity of whisker catfish

3.3.1.1 Experiment systems

An experiment was carried out in hapas (1 x 1x 1.5 m), sizes of mesh was 0.5 cm; which was put in a pond (500 m2), the depth of water was 1.2 – 1.5 m

3.3.1.2 Fish of experiment: Whisker catfish was equal sizes and

healthy, and the initial weight of fish was 80 – 90 g/fish

3.3.1.3 Feed of experiment: Trash tiny shrimp and fish, which were

crushed and mixed with 1 % of powder of cotton leaves to increase adhisively and pelllets were also wetted by water Feed was put in

”feeding floors” at the bottom of hapas

3.3.1.4 Experimental design

The experiment including three treatments (T) was completely randomized designed with three replicates Every hapa was stocked with 20 whisker catfish (the rate of male and female was 1 : 1) The time of experiment was five months

Table 3.1 Component of home-made feed

Treatment (T) Component of home-made feed

T 1 99% trash tiny shrimp + 1% kapok powder

T 2 49.5% trash tiny shrimp+49.5% trash fish+1% kapok powder

T 3 100% pellet (29.5 % crude protein)

3.3.1.5 Feeding and taking care of: Whishker catfish was fed

demandably, including two times per day at 8 am and 5 pm

2.3.1.6 Result records

Before conducting experimental arrangement, three males and females were chosen contingently to collect the first data Through experimental process, recurrently every treatment will have sampled contingently, including three males and females to check the maturity

of fish through index about the size rates of eggs diameter, the erythrocyte numbers, hemoglobin and vitellogenin contents These indexes were checked at 30; 60; 90; 105; 120 and 150 days after experimental arrangement Gonadosomatic index and fecundity of whishker catfish were checked at the times that gonad of fish was 4thstage

3.3.2 The technological study of artificial spawning of whishker catfish

3.3.2.1 Experimental fish: Healthy whishker catfish with the initial

weight of 50 – 100 g/fish

Female: big belly, concave papilla Whishker catfish has had eggs

diameter which was ≥ 0.8 mm was more than 80%

Male: choosing fish which has had slim body, samll belly and

pointed papilla

3.3.2.2 Experimental design

Table 3.2: Content of hormone in experimental arrangement

Four experiments were completely randomized designed, with four different hormones, every type of hormone has had three content (9 females/repeatedly) (Table 3.2) Male fish was injected by hormone correlatively with each experiment, by 1/3 the content of female

3.3.2.5 Recording of results: The results of artificial breeding of

whishker catfish were evaluated through some indexes: the rate of spawning, the times of effective drugs, fecundity, fertilization rates, and the rates of hatching; which was recorded

3.3.3 The technological study of whisker catfish rearing from fry

to fingerling stages

3.3.3.1 The study of nutritional characteristics of whisker catfish from fry to juvenile

a Experimental pond: The study was carried out in a pond which

has had about 12 m2 (3 x 4 m) in areas, and was coated canvas and muds at bottom about 25 cm, the depth of water was 0.5 m

b Feed: Before stocking, natural food was cultured by haversacks

which have had about 25 µm in mesh and have contained fish meal

in some corners of ponds throughout experiment, avoiding fish was eating directly fish meal

Experiment Hormone Content of hormone

40 + 3.5 0.3

2.5 5,000

70 + 3.5 0.4

3.5 6,000

100 + 3.5 0.5

c Experimental design: Whisker catfish (1 day) which has had

average weight and length were 1.73 mg and 5.02 mm, respectively;

were stocked in the pond about 600 fish/m2 in density and the

experimental time was 30 days

d Sampling and analysis

Sampling: The samplings of zooplankton and phytoplankton and fish

on the day: 1; 2; 3; 4; 5; 6; 8; 10; 15; 20; 25 and 30 after stocking

Fish was sampled contingently about 30 individuals and preserved in

formol solution 10%

The analysis of zooplankton and phytoplankton: qualitative analysis

based on classification documents of Dang Ngoc Thanh (1980), Han

Maosen Shu Yunfang (1995); quantify based on the documents of

Duong Tri Dung (1996); Boyd and Tucker (1992)

Identified the nutritional characteristics of whishker catfish

(1) The rate between intestine and standard length L i /L 0

(2) Identified the sizes of fish mouth

(3) Stomach analysis

Frequency method: Frequency of food is the rate between stomach

which holds this food and the total of stomachs are observed (Hynes,

1950)

(4) The food choices of fish

Cofficients of food choices are counted by Ivlve’s Electivity index:

E = (ri-pi)/(ri+pi)

In there, ri which is the percentage of food type i has found in

intestine of fish and pi which is the percentage of food type

correlatively has found in environment on the total food which has

had in environment

3.3.3.2 The study of whisker catfish rearing from fry to

fingerling

Experiment 1: The study of whisker catfish rearing to 30 th day

using live food

a Experimental design: Experiment was arranged in the system of

plastic bucket about 35 L (held 30 L water) The experiment was completely randomized designed, including four treatments (T) with three replicates: (T1) whisker catfish was fed by tubifex, (T2) whisker catfish was fed by zooplankton, (T3) fish was fed by artemia and (T4) fish was fed the combination between tubifex and zooplankton in the same rate

Fish of experiment was whisker catfish which was two days; healthy, not deformities and mean weight of fish was 1.8 mg The density of rearing was 2.5 fingerlings/L in 30 days

b Experimental food: Tubifex is bought by households in Can Tho

province Zooplankton was cultured (was the same with the culture method of live food at the study 3.3.3.1 of section b) After that racket was used to let through for fish which ate gradually through out the experiment Larvae of newly-hatched artemia were collected through the incubation process of cyst artemia about 15 ‰ in salinity for 24 hours

e Feeding and taking care of: Fish was fed by demand, including

two times per day at 8 am and 5 pm

f Result records

Environmental factors such as temperature, oxy, pH; which were identified by 556 YSI – USA, two times per day Before arranging experiment, 30 of fry whishker catfish were scaled to indentify the first weigh When the experiment has finished, most of fish will have scaled by electronic weight to identify: daily weigh gain (DWG, g/ngày), specific growth rate (SGR, %/ngày) and survival rate

Experiment 2: The study of whisker catfish rearing by Tubifex at different densities

a Experimental arrangment: Experiment fish (2 days) was

arranged contingently in plastic bucket about 35 L (held 30 L water), including four treatments that densities were 1.5; 3.5; 5.5 and 7.5 individuals/L, respectively, repeated three times The time of experiment was 30 days

b Food of experiment: Food for fish in this experiment was tubifex which was cut smoothly (the best result in experiment of 1)

Experiment 3: The study of times that whisker catfish has used effectively home-made feed

a Experimental design: Experiment was carried out in plastic

bucket about 35 L (held 30 L water) Whisker catfish (2 days) was arranged in bucket with 2.5 individual/L in density Experiment was carried out about 30 days, including six treatments which were arranged contingently with repeating three times All treatments differed by the first day of feeding home-made, including the day 1; 3; 5; 7; 9 and 11 after arranging experiment

Table 3.5: Nutritional component of home-made feed

b Food of experiment: Components of home-made feed included:

trash fish (100 g), unfatty milk (100g), egg yolk (10 eggs), squid oil (3%) Most materials were husked, steamed, pressed fiber, dried up and kept in cold store Fish was fed three hours per once based on their demands

Experiment 4: The study of whisker catfish rearing by pellets at different densities

a Experiment systems: Experiment was arranged in cement tanks

with 1 m3 (held about 600 L water)

b Fish of experiment: Fish in the experiment was whishker catfish

of 7 days and mean weight was 13.1 mg, weaned by pellets (the study of 3.3.3.4), healthy and not deformities

c Experimental design: Experiment was completely randomized

designed, including four treatments which were 1.5; 3.5; 5.5 and 7.5 individuals/L to equal with 900; 2,100; 3,300 and 4,500 individuals per tank, respectively and repeated three times The time period of the experiment was 60 days

d Feed and taking care of: Feed of experiment was pellet which

has had 36 % in crude protein

* Feeding and taking care of: Fish was fed by demand, including

two times per day at 8 am and 5 pm

* Result records

Environmental factors such as temperature, oxy, pH; which were identified by 556 YSI – USA, two times per day Before arranging experiment, 30 of fry whishker catfish were scaled to indentify the first weigh When the experiment has finished, most of fish will have scaled by electronic weigh to identify: daily weigh gain (DWG, g/ngày), specific growth rate (SGR, %/ngày) and survival rate

3.3.3.3 Identify protein requirement of M bleekeri fingerlings

a Experimental system: Experiment was carried out in the

composite system (20 L per tank) in door facility and was aerated continuously

b Experimental arrangement: Experiment fish which was 269

mg/fish was weaned home-made feed (by the method which has achieved in the best results from experiment 3) Experiment was arranged contingently, including seven treatments, repeated three times with 2.5 fish/L (30 fish per tank) in six weeks Most treatments have had the levels of protein which were 24%; 29%; 34%; 39%; 44%; 49% and 54%, respectively

c Food of experiment: Experiment food has had the same of energy

(4.36 Kcal/g) and fatty (10 %) Food materials were fish meal, soya bean meal, wheat meal, fish oil, vegetable oil, vitamin – mineral,

gelatin and fillers

Table 3.6 Nutritional component of feed

Protein (%) 23,94 28,78 33,76 38,94 43,45 48,47 53,73 Lipid (%) 9,38 9,87 9,67 9,63 9,56 9,76 9,84 Carbohydrad (%) 49,0 42,3 35,6 28,8 22,1 15,4 8,72

d Management and taking care of: Diets were about 3 – 7 % of

weight (counted both at demands and dry weight) and fish is fed every day four times at 7 am, 10.30 am, 2 pm and 5.30 pm

e Results records: To identify protein requirement of whisker

catfish fingerlings to record the fish growth; protein demands; feed conversion ratio, effective protein, survival and herd rates

3.4 The method of data processing

Data were calculated mean and standard deviation One-way ANOVA analysis and DUNCAN test were used to compare difference among treatments Data analysis was conducted by using Statistica 5.0 software

RESULT AND DISCUSSION 4.1 The conditioning results of whisker catfish maturity

4.1.2 The maturity of whisker catfish

4.1.2.1 The fluctuations of eggs diameter ratio through conditioning process

In the first time of May (May 5th 2010) eggs of whisker catfish have had egg diameter which was from 0.3 to 0.4 mm held 33.3 – 40 %, whereas the rate of egg diameter from 0.5 from 0.7 mm was 50 – 53.3 %, and egg diameter which was ≥ 0.8 mm held very lowly about 6.67 – 13.3% In June, the rate of egg diameter < 0.4 mm has not virtually appear in sampling at treatments Whereas, the percentage

of egg diameter ≥ 0.8 mm which increased sharply at most treatments

were 93.3 %; 83.3 % and 66.7 %, respectively (Figure 4.1)

Figure 4.1: The fluctuations of eggs diameter ratio

The highest rate was treatment I (about 93.3 %) and was significantly higher than the other treatments (p < 0.05) Besides, eggs of whisker catfish at treatment I have had egg diameter which was ≥ 0.8 mm held the highest rate (93.3 %); indicating that fish has matured early and better than the other treatments These results indicate that trash tiny shrimp which is an appropriate food for the process of maturity conditioning of whisker catfish Fish which has conditioned in ponds can be mature in May and June

<=0,2 0,3-0,4 0,5-0,7 >=0,8 Eggs diameter (mm)

<=0,2 0,3-0,4 0,5-0,7 >=0,8 Eggs diameter (mm)

<=0,2 0,3-0,4 0,5-0,7 >=0,8 Eggs diameter (mm)

4.1.2.2 The correlation between the sizes of egg diameter (the maturity stage) and Vitellogenin content (Vg)

Figure 4.2: The fluctuations of Vg content and average egg diameter

In March, before the experiment had been arranged, whisker catfish has had egg diameter 0.23 ± 0.08 mm, equal with the gonad of II stage that Vg content in plasma of fish was about 0.21 ± 0.02 µg ALP/mg protein Concurrently with the increasing sharply of egg diameter (0.5 – 0.52 mm), Vg content in plasma (0.83 -1.03 µg ALP/mg protein) also increased dramatically Especially Vg content rose suddenly (2.46 – 2.65 µg ALP/mg protein) after 15 days These results show that, the gonad of fish has developed from III stage (egg diameter was < 0.4 mm) to IV stage (egg diameter was 0.5 – 0.7 mm) and Vg content has also increased sharply In June, egg diameter of fish at most treatments were ≥0.8 mm Concomitantly, Vg content fluctuated from 2.94 to 3.28 µg ALP/mg protein In July, both egg diameter and Vg content also decreased sharply and were not significantly different among treatments (P > 0.05)

4.1.2.3 Effective conditioning food to gonadosomatic index, fecundity and Vg content of whisker catfish