….
DETERMINATION OF BREEDING SEASON OF BLACK POMFRET (Parastromateus niger) FROM
THE BAY OF BENGAL, BANGLADESH
Md. Abirul Islam
Roll No.: 0118/02 Registration No.: 0573 Session: 2018-2019
A thesis submitted in the partial fulfillment of the requirements for the degree of Master of Science in Fish Biology and Biotechnology
Department of Fish Biology and Biotechnology Faculty of Fisheries Chattogram Veterinary and Animal Sciences University Chattogram-4225, Bangladesh
JUNE 2019
Authorization I hereby declare that I am the sole author of the thesis. I also authorize the Chattogram Veterinary and Animal Sciences University (CVASU) to lend this thesis to other institutions or individuals for the purpose of scholarly research. I further authorize the CVASU to reproduce the thesis by photocopying or by other means, in total or in part, at the request of other institutions or individuals for the purpose of scholarly research.
I, the undersigned, and author of this work, declare that the electronic copy of this thesis provided to the CVASU Library, is an accurate copy of the print thesis submitted, within the limits of the technology available.
Md. Abirul Islam
June 2019
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DETERMINATION OF BREEDING SEASON OF BLACK POMFRET (Parastromateus niger) FROM
THE BAY OF BENGAL, BANGLADESH
Md. Abirul Islam
Roll No.: 0118/02 Registration No.: 0573 Session: 2018-2019
This is to certify that we have examined the above Master’s thesis and have found that is complete and satisfactory in all respects, and that all revisions required by the thesis examination committee have been made.
Md. Moudud Islam Dr. Helena Khatoon
Supervisor Co-supervisor
Department of Fish Biology and Department of Aquaculture Biotechnology
------Md. Moudud Islam Chairman of the Examination Committee
Department of Fish Biology and Biotechnology Faculty of Fisheries Chattogram Veterinary and Animal Sciences University Chattogram-4225, Bangladesh
JUNE 2019
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Dedicated To my
Beloved parents
Acknowledgement All praises are due to the Almighty Allah for blessing me with the strength, aptitude, patience and enabled me to pursue higher education and to complete the thesis for the degree of Masters of Science (MS) in Fish Biology and Biotechnology.
First of all I want to pay heartily gratitude to Professor Dr. Goutam Buddha Das, Vice-Chancellor, Chattogram Veterinary and Animal Sciences University (CVASU) for giving special opportunity and providing such research facilities.
I would like to pay my sincere regards and thanks to Prof. Dr. M. Nurul Absar Khan, Dean, Faculty of Fisheries, Chattogram Veterinary and Animal Sciences University, who introduced Master’s program in the Faculty of Fisheries and provided update instrument and laboratory for conducting any kind of research.
With great pleasure, I would like to express my deepest sense of gratitude, sincere appreciation, profound regards and immense indebtedness to my honorable teacher and research supervisor Md. Moudud Islam, Assistant Professor and Head, Dept. of Fish Biology and Biotechnology, Chattogram Veterinary and Animal Sciences University for giving the opportunity to do research and providing invaluable guidance and continuous support throughout this research. His dynamism, vision, sincerity and motivation have deeply inspired him a lot. It was a great privilege and honor to work and study under his guidance. I am extremely grateful to him and would also like to thank him for his friendly cooperation, empathy, and great sense of humor.
I feel proud in expressing my regard and immense gratitude to my co- Supervisor Dr. Helena Khatoon, Assisstant Professor, Department of Aquaculture, Chattogram Veterinary and Animal Sciences University for her kind co-operation, valuable suggestions and constructive criticism in improving the quality of the research work.
I expresses my deepest sense of gratitude, indebtedness, sincere appreciation and profound regards to my honorable teacher Md. Mahiuddin Zahangir, Assistant Professor, Dept. of Fish Biology and Biotechnology, Chattogram Veterinary and Animal Sciences University for his scholastic guidance, cordial support, constant encouragement, valuable suggestions, punctuality and constructive criticism throughout the course of this research work and during the write up of the thesis. His
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guardianship, counsel, simplicity and contribution will remain as indelible memories forever.
I am greatly indebted to Fatema Akhter, Lecturer, Dept. of Fish Biology and Biotechnology (CVASU) for her valuable advice, scholastic guidance, suggestions and inspiration throughout the research.
I acknowledge the consistent academic support, co-operation and guidance of all the teachers of Faculty of Fisheries during my one and half years study period at Chattogram Veterinary and Animal Sciences University, Chattogram.
Finally, I would like to express my cordial thanks to my loving friends, employees and laboratory attendants of the Department of Fish Biology and Biotechnology for their active assistance during the whole study period. I also express my heartfelt gratitude and sincere appreciation and profound indebtedness to all the people who have supported to complete the research work directly or indirectly.
At last my heartfelt respects and thanks to my beloved parents for their ultimate understanding, inspirations, moral support, kindness and blessings, forbearance and endless love to complete this study. Thank you.
The Author
June 2019
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CONTENTS CHAPTER TITLE PAGE NO. AUTHORIZATION i ACKNOWLEDGEMENT iii- iv LIST OF TABLES ix LIST OF FIGURES x- xi LIST OF PLATES xii- xiii LIST OF APPENDICES xiv LIST OF ABBREVIATIONS xv ABSTRACT xvi 1 INTRODUCTION 01- 04
1.1 Aim and objectives of the research 04
2 REVIEW OF LITERATURE 05- 08
2.1 Length-weight relationship 05- 07
2.2 Reproductive biology and spawning season 07- 08
3 MATERIALS AND METHODS 09- 22
3.1 Sampling site and sample collection 09- 10
3.2 Recording of length-weight data and 10- 11 determination of length-weight relationship
3.3 Determination of condition factor 11
3.4 Determination of relative condition factor 11- 12
3.5 Collection and preservation of gonad and liver 12- 13
3.6 Determination of hepato-somatic index (HSI) and 13- 14 gonado-somatic index (GSI)
3.7 Size at first sexual maturity (Lm) 14
3.8 Estimation of fecundity 14- 15
3.9 Estimation of oocyte diameter 15
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3.10 Histological analysis of gonad 15- 22
3.10.1 Fixation 15- 16
3.10.2 Washing of fixed tissue 16
3.10.3 Dehydration 16- 17
3.10.4 Cleaning 17
3.10.5 Infiltration 18
3.10.6 Embedding 18
3.10.7 Trimming 18- 19
3.10.8 Sectioning 19
3.10.9 Floating of section in water bath 19- 20
3.10.10 Attachment of section on glass slide and 20 drying
3.10.11 Staining 20- 21
3.10.12 Mounting 22
3.10.13 Microscopic examination of the gonad tissue 22 sections
3.11 Statistical analysis of data 22
4 RESULTS 23- 45
4.1 Length-weight relationship 23- 25
4.2 Condition factor and relative condition factor 25- 28
4.3 Hepato-somatic index (HSI) 29- 30
4.4 Gonado-somatic index (GSI) 30- 32
4.5 Oocyte diameter 33- 34
4.6 Size at first Sexual Maturity (Lm) 34- 37
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4.7 Fecundity 37
4.8 Gonadal maturation stages in female 37- 41
4.8.1 Pre-vitellogenic stage 37- 38
4.8.2 Early primary vitellogenic stage 38- 39
4.8.3 Advanced primary vitellogenic stage 39
4.8.4 Secondary vitellogenic stage 40
4.8.5 Ripe stage 41
4.8.6 Spent or regressing stage 41
4.9 Gonadal maturation stages in male 41- 45
4.9.1 Immature stage 41- 42
4.9.2 Developing stage 42- 43
4.9.3 Pre-spawning stage 43
4.9.4 Ripe stage 43-44
4.9.5 Spawning stage 44
4.9.6 Post-spawning stage 44- 45
5 DISCUSSION 46- 52
5.1 Length-weight relationship 46- 47
5.2 Condition factor and relative condition factor 47- 48
5.3 Hepato-somatic index (HSI) 48- 49
5.4 Gonado-somatic index (GSI) 49- 50
5.5 Oocyte diameter 50
5.6 Size at first Sexual Maturity (Lm) 50
5.7 Fecundity 51
5.8 Histological observation of gonads 51- 52
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6 CONCLUSIONS 53
7 RECOMMENDATION AND FUTURE 54 PERSPECTIVES
References 55- 58
Appendices 59- 66
Biography 67
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LIST OF TABLES TABLE TITLE PAGE NO. NO. 1 The dehydration schedule 17 2 The cleaning schedule 17 3 The infiltration schedule 18 4 The staining schedule 21 5 Condition factor and relative condition factor data of 26 P. niger of 12 months
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LIST OF FIGURES FIGURE TITLE PAGE NO. NO. 1 Logarithmic relationship between the standard length and 24 weight based on pooled data of P. niger 2 Logarithmic relationship between the total length and 24 weight of female P. niger 3 Logarithmic relationship between the total length and 25 weight of male P. niger 4 Condition factor based on pooled data of P. niger of 12 27 months 5 Relative condition factor based on pooled data of P. niger 27 of 12 months 6 Condition factor (K) of male and female of P. niger 28 7 Relative condition factor (Kn) of male and female of P. 28 niger 8 Hepato-somatic index (HSI) of male and female P. niger 29 9 Hepato-somatic index (HSI) and condition factor (K) of P. 30 niger based on pooled data 10 Monthly variations of gonado-somatic index (GSI) in 31 female P. niger 11 Monthly variations of gonado-somatic index (GSI) in male 31 P. niger 12 Monthly variations of gonado-somatic index in male and 32 female P. niger 13 Monthly variation of gonado-somatic index (GSI) and 32 hepato-somatic index (HSI) of P. niger based on pooled data 14 Oocyte diameter of P. niger collected from Bay of Bengal. 33 Data are presented as mean ± standard deviations 15 Oocyte diameter and GSI of P. niger collected from Bay of 34 Bengal. Values are presented as mean ± standard deviations 16 Relationship between gonado-somatic index (GSI %) with 34 total length of female P. niger 17 Relationship between modified gonado-somatic index 35 (MGSI%) with total length of female P. niger
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18 Relationship between Dobriyal index (DI) with total length 35 of female P. niger 19 Relationship between gonado-somatic index (GSI%) with 36 total length of male P. niger 20 Relationship between modified gonado-somatic index 36 (MGSI %) with total length of male P. niger 21 Relationship between Dobriyal index (DI) with total length 37 of male P. niger
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LIST OF PLATES PLATE TITLE PAGE NO. NO. 1 The Black Pomfret, P. niger 2 2 The map of sample collection site 09
3 Recording of length-weight data of P. niger 10 4 Collection and preservation of gonad and liver 13 5 Counting of eggs 15 6 Measurement of oocyte diameter (µm) 15 7 Sample preservation in Bouin’s fixative 16 8 Dehydration of tissues in alcohol 16 9 Cleaning of tissues 17 10 Embedded sample 18 11 Trimming unwanted paraffins 19 12 Sectioning by using rotary microtome 19 13 Floating of section in water bath 20 14 Attachment of section on glass slide and drying 20 15 Mounting of stained sample slide 22 16 Microscopic observation of gonad tissue 22 17 Different stages of ovarian development of P. niger. Pre- 38 vitellogenic stage (A, B, C and D) 18 Early primary vitellogenic stages (E and F) and advanced primary 39 vitellogenic stages (G and H) of ovarian development of P. niger 19 Secondary vitellogenic stage (I), Ripe stage (J and K) and Spent 40 or regressing stage (L) of P. niger 20 Gonadal maturation cycle of testes of P. niger. Immature stage 42 (A, B, C and D) 21 Gonadal maturation cycle of testes of P. niger. Developing stage 43 (E and F) 22 Gonadal maturation cycle of testes of P. niger. Pre-spawning 43 stage (G and H) 23 Ripe stage (I) and Spawning stage (J) of testes of P. niger 44
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24 Gonadal maturation cycle of testes of P. niger. Post-spawning 45 stage (K and L)
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LIST OF APPENDICES APPENDIX TITLE PAGE NO. NO.
1 Data of total length, standard length, body weight of 59- 64 P. niger
2 Month wise hepato-somatic index (HSI) and gonado- 65 somatic index (GSI) of P. niger (both male and female)
3 Condition factor and relative condition factor data of 66 P. niger of 12 months (both male and female)
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LIST OF ABBREVIATIONS
DoF Department of Fisheries
FAO Food and Agriculture Organization g Gram cm Centimeter
µm Micrometer
MT Metric Ton
% Percent sq Square km Kilometer
M Meter
°C Degree Celsius
HSI Hepato-somatic index
GSI Gonado-somatic index
S. D. Standard deviation
U.A.E United Arab Emirates
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ABSTRACT
The reproductive biology of the black pomfret, Parastromateus niger in Bay of Bengal of Bangladesh were investigated from March, 2018 to February, 2019. Analyses of morphometric parameters revealed the exponent ‘b’ value of the standard length-weight relationship of P. niger was 2.78 for both sexes, 2.78 for females and 2.80 for males, all indicating negative allometric growth (b< 3). The average mean condition factor (K) and average mean relative condition factor (Kn) values were 3.20±0.13 and 1.00±.03 respectively which indicating a good health condition of the fish. The maximum mean values of HSI were found both in February, for males (1.36±0.19) and for females (1.51±0.19). The mean GSI value ranged from 0.12±0.09 (August) to 4.02±1.30 (April) in female and 0.07±0.04 (August) to 0.98±0.26 (March) in male. High value of GSI was also observed in October in female (0.52±0.59) and in male GSI continued to increase from October up to December, indicating a chance of second spawning. Maximum oocyte diameter was found in April (285.53±35.09µm) and minimum oocyte diameter was found in July (28.32±5.28µm). The comparative study between oocyte diameter and GSI evidently showed that the oocyte started development as GSI increased from January and gradually become ripe in March, finally spawns in April and May. Based on gonado-somatic index, modified gonado- somatic index, and Dobriyal index, the size at first sexual maturity (Lm) was calculated as 28 cm TL for female and 26.50 cm TL for male. The fecundity was determined from randomly collected gravid female fish samples and found to vary from 114615 (23.70cm SL and 392 g weight) to 127605 (24.10cm SL and 436.20 g weight) eggs. Along with the data of GSI and oocyte diameter histological observation of gonadal maturity stages revealed that the relationship in gonadal maturity in both male and females were in a synchronize manner in this species indicating the spawning period from March to May with a minor peak in October. Findings of the study will be useful for artificial propagation of P. niger as well as to determine ban period of catching marine species of the Bay of Bengal, Bangladesh.
Key Words: Condition factor, Fecundity, HSI, GSI, Histology of gonad, P. niger
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CHAPTER-1
INTRODUCTION
CHAPTER-1
INTRODUCTION
Fish are economically very important to man. They are important as- food, they give by-product, can control diseases and are a source of income and employment to many developing regions and in low-income food-deficit countries. As an affordable animal source of protein in some of the poorest countries, fish is the primary source of nutrition, creating growing demand for this staple. Total fish production in world was 170.9 million tons, where the contribution of marine capture was 79.3 million tons in 2016. Major species showing reduced catches between 2015 and 2016. Many millions of people around the world find a source of income and livelihood in the fisheries and aquaculture sector (FAO, 2018).
Bangladesh is a riverine country blessed with many inland water bodies covering a huge area of water resources of 4.70 million hectares. Besides, there is a huge marine fisheries resources expanding over an exclusive economic zone (EEZ) of 1, 66,000 sq. km (DoF, 2018a). The fisheries sector plays a very important role in the national economy, contributing 3.57% to GDP and 25.30% to agricultural GDP and more than 11% of the population depends directly and indirectly on the fisheries sector for their livelihood. Fish supplements about 60% of Bangladeshi people’s daily animal protein intake (DoF, 2018b).
According to FAO report (The State of World Fisheries and Aquaculture 2018) Bangladesh ranked 3rd in inland open water capture production and 5th in world aquaculture production. In 207-2018, total fishery production of Bangladesh was 42.77 lakh metric tons, whereas inland open water (capture) contributes 28.45% (12.17 lakh metric tons) and inland closed water (culture) contributes 56.24% (24.05 lakh metric tons). On the other hand marine fisheries production is 6.55 lakh MT and its contribution to total fish production is 15.31 percent with growth rate 2.71 percent (DoF, 2018b). The country has limited access to marine fisheries resources. Only demarsal fish and shrimp are being trapped from here. Other potential marine resources are yet to be exploited on commercial scale (DoF, 2018a).
Fish and seafood products, have a high nutritional value regarding beneficial amounts of protein, lipids as well as essential micronutrients. Aquatic animal foods are a rich
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source of protein and have a lower caloric density, and have a high content of omega 3 long chain polyunsaturated fatty acids (n-3 LC PUFA) compared to land living animals Tilami and Sampels (2017). Pomfrets form one of the commercially important fishery all along the Indian coast due to their soaring market price and good foreign exchange value (Mazumdar et al. 2012).
The black pomfret, Parastromateus niger, is a species of carangid family native to reefs of the Indian Ocean and the western Pacific Ocean, where it is found at depths from 15 to 105 m (49 to 344 ft.), though it is rarely found deeper than 40 m (130 ft.). This species grows to 75 cm (30 inch) in total length and is very important to local commercial fisheries. Often occurs in large schools, and not uncommonly observed swimming on their sides. Usually found in 15 to 40 m depth, generally over muddy bottoms. It feeds on zooplankton. A large parasitic isopod is often present clinging to the tongue. This species is the only known member of its genus. They are found in India, Malay Archipelago, China; Bay of Bengal, Eastern Sea of India, Africa, Japan (Shafi and Quddus, 2005).
Black pomfret is a mid-water pelagic species that occur over muddy bottoms in coastal areas. Having similarities in body shape, the black pomfret is often thought to be related to the Chinese (Pampus chinensis) and white (Pampus argenteus) pomfrets. However, the black pomfret belongs to the family Carangidae and the white and Chinese pomfrets to the family Stromateidae. In fact, the black pomfret is unusual for members of the Carangidae (Tan, 2009).
Plate- 1: The Black Pomfret, P. niger
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Both the white and Chinese Pomfrets lack pelvic fins at all stages of life, whereas the black pomfret has a pelvic fin in younger specimens shorter than 100 mm in total length. In addition, the small scutes along the caudal peduncle of the black pomfret are absent on white and Chinese pomfrets.
Living black pomfret is very different in coloration from dead market specimens. Live specimens have a silvery, steel-grey body, with four narrow white bars spaced at regular intervals along the flanks, with the first bar located just behind the opercle. The belly area is whitish. The dorsal fin is yellow and the anal fin is pale yellow and both fins having a narrow black margin. The pectoral fin is hyaline and the caudal fin is whitish (Tan, 2009).
Fish stocks are renewable natural resources which are replenished by reproduction (Olusegun, 2011). To determine the precise spawning season and frequency in a breeding season study of gonadal development cycle and histological examination of gonads are important (Conover, 1992). By histological examination of the gonad one can easily determine the different stages of ovary and can also identify when the ovum becomes mature. It provides an overall idea on the ovum diameter, fecundity and peak spawning season of the fishes. Knowledge on different stages of fish gonadal maturation helps in prohibition of fishing during spawning season, which ultimately allows the fish stock to recover.
To determine the spawning period gonado-somatic index (GSI) also plays a major role as there is a cyclic change in gonad weight in relation to total body weight (Nieland and Wilson, 1993; Jons and Miranda, 1997; Smith, 2008). An increasing GSI suggests that spawning season is approaching, where decreasing GSI suggests spawning has occurred.
This study is necessary to identify the precise spawning season of P. niger, which in result will help to determine the ban period of catching this species from the Bay of Bengal of Bangladesh to replenish the stock.
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1.1 Aim and objectives of the research:
The aim of this study is to describe the spawning season of the P. niger in the Bay of Bengal. The specific objectives of the proposed research are as follows:
To determine the length-weight relationship, condition factor, relative condition factor, fecundity, oocyte diameter, hepato-somatic index (HSI) and gonado-somatic index (GSI) of P. niger To identify the spawning season through studying gonadal maturation cycle of P. niger
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CHAPTER-2
REVIEW OF LITERATURE
CHAPTER-2
REVIEW OF LITERATURE
For understanding the basic biology of fish, length-weight relationship studies are considered as an important tool. Evaluation of the conditions at different body lengths can give valuable information regarding the maturation and spawning in the life span of the fish, whereas a close look at the condition during different months can give definite clues regarding the breeding seasons. This chapter is about the review of length-weight relationship, GSI, length at first maturity, oocyte diameter, cyclic changes in the gonadal development, fecundity of P. niger species. The following information was collected to design the present research and validation of the new findings.
2.1 Length-weight relationship
Dadzie et al. (2008) analyzed the morphometric parameters of the black pomfret, P. niger in the Kuwaiti waters of the Arabian Gulf. During a 44-month study revealed the following length-length relationships: TL (cm) = 0.63 + 1.15SL (females), TL (cm) = 0.94 + 1.14SL (males) and TL (cm) = 0.75 + 1.15SL (both sexes including juveniles). The 95% condition index (CI) of the exponent in length-weight relationship varied from 2.788 to 2.796 in females, 2.726 to 2.732 in males, 2.779 to 2.784 for both sexes and 2.590 to 2.613 in juveniles, all indicating an allometric relationship.
Saleh and Soegianto (2017) carried out a study in the eastern region of Java Sea at north of Madura Island (6°00′– 6°50′S, 112°50′– 114°10′E), Indonesia. A total of 2461 specimens belonging to five families including P. niger, and found negative allometric growth and the coefficient of determination (R2) values 0.936 where, a = 0.045, b = 2.729 for this species.
Abdurahiman et al. (2004) studied the length distribution analysis of P. niger off southern coast of Karnataka, India for 1999 to 2001, based on 23 male individuals of 17.2- 35 cm TL range and 12 female individuals of 19.3– 36.3 cm TL range, and found out that the length-weight relationship of the species followed the equations of W = 0.053* L2.655 for males with correlation co-efficient of 0.98, and W = 0.069*
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L2.573 for females with correlation co-efficient of 0.96, which indicated the allometric growth in the species.
Kazemi et al. (2013) studied on five important commercial fish species of the Persian Gulf belonging to five families including the species P. niger and estimated co- efficient of determination, = 0.959 which showed a strong correlation between the variables. In length-weight relationship exponent b = 2.919, a = 0.033 and the reported growth is isometric. The mean relative condition factors (Krel) was 1.0299 ±0.021 and it ranged from 1.01±0.08 for P. niger.
Khan et al. (1992) gave the length-weight relationship of P. niger, along the Kerala coast as log W = -4.26513 + 2.7921841 log L with allometric growth pattern.
Hadisubroto and Subani (1994) reported that the length-weight relationship of P. niger from Palau Laut waters, South Kalimantan in the equation of log W = -1.2044 + 2.6415 log L (r = 0.98) with allometric growth type.
Mustafa (1999) established the fork length-weight relationship of P. niger along the Bay of Bengal waters of Bangladesh as W = 0.0211* L3.012, which indicated isometric growth of the species.
Daliri et al. (2012) studied the length-weight relationships (LWR) and relative condition factor for five marine fish species collected by shrimp trawls from Iranian waters of the Persian Gulf and reported isometric growth for P. niger with the value b = 2.9477, a = 0.0342 and co-efficient of determination, R2 = 0.93 which showed a significant correlation between the variables.
Din et al. (2015) studied the Length-weight relationship (LWRs), condition (K), relative condition factor (Kn) for male, female and combined sexes of silver pomfret, P. argenteus from Quetta City of Pakistan. The obtained results of that study revealed that a strong correlation (r>0.80) occurred between the length and weight of the species and was found to be significant at 5% level (p<0.05). The negative allometric growth pattern (b<3.0) were observed, but found to be highly significant (t-test; p<0.05) for male, female and combined sexes, indicating that if the body length of fish increases than weight of body is not increasing accordingly, which might be due to several factors such as, growth, environmental conditions and condition of fish. The values of condition factor (K) showed the difference with increase in size or
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weight of fish, while the values of relative condition factor (Kn) was equal to 1.0, hence indicating that the condition of the environment of this species was favorable for its growth.
2.2 Reproductive biology and spawning season
According to Dadzie et al. (2008) analysis of Fulton’s condition factor (K) and GSI indicate that spawning in black pomfret peaks in May and June even though the condition of fish remains good throughout the spawning season. The Fulton’s condition factor was statistically discriminatory in determining the well-being of P. niger. Variation in average maximum condition was significant between sexes. The mean size at first sexual maturity was 17.5 cm for males and 29 cm for females. Fecundity ranged from 71 305 to 3 895 449 eggs.
The general length–weight relationship equations derived for females, males and juveniles showed that the parameter ‘b’ was significantly lower than three for either sex, juveniles and both sexes, suggesting that specimens become more elongated as they increase in size.
Dadzie et al. (2009) observed the reproductive activities of the male and female black pomfret P. niger in Kuwaiti waters from October 2003 to September 2005. The study revealed that the seasonal variations in the gonado-somatic index (GSI) during the two-year study period revealed high values from February to September, suggesting that the black pomfret has a prolonged spawning season, from February to September. GSI fluctuations correlated positively with rising water temperatures in Kuwait from low values in both parameters in January to high values in February/ March (r = 0.836, p<0.05 for males and r = 0.764, p<0.05 for females), suggesting that temperature plays a role in triggering spawning in both the sexes. Analysis of seasonal distribution of maturity stages for the two years revealed the presence of ripe/running males and females from February to September, thus confirming the spawning periodicity revealed through the analysis of fluctuations in the GSI. Macroscopic and microscopic studies of maturity stages revealed six stages in the males and seven in the females. The logistic function based on pooled data for the two years revealed that the minimum size at sexual maturity was attained at a size of 30.9 cm SL in males (R2 = 0.284) and 36.5 cm SL in females (R2 = 0.355). Total fecundity ranged from 71 305 in a fish measuring 39.8 cm SL and weighing 1 572.5g, to 3 895 449 in a 49 cm SL
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and 2 630g fish, with a mean of 1 216,734 eggs. Positive correlations were found between fecundity and ovary-free body weight, standard length and ovary weight, and a negative one with egg size. The average relative fecundity was 948 eggs/g ovary- free body weights, which was neither a function of fish standard length nor ovary-free body weight.
From the study of Sivaprakasam (1965) it was concluded that P. niger spawn during July to October and the males appear to migrate to the spawning grounds earlier than the females, as indicated by their predominance during April-May. During August- October which is the spawning period, females outnumber the males. It is also found that males are predominant over females in the first size group, become less in number in the next group, and are completely absent in the last group. Most of the females attain first maturity at 32 cm and the males at 30 cm. Unfortunately, enough fish were not available in the group 25-30 cm as the selectivity of the gill-nets was rather high.
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CHAPTER-3
MATERIALS AND METHODS
CHAPTER-3
MATERIALS AND METHODS
This chapter deals with the methods that are followed and materials that are used to achieve the objectives of the study. Methodology is an essential and integral part of any research. In this study a scientific and logical methodology has been followed. The present study was aimed to determine the gonadal maturation cycle of P. niger. The study was based on laboratory work and data were collected for the interpretation of results. To characterize the gonadal maturation cycle of P. niger the following procedures were followed:
3.1 Sampling site and sample collection
The study was conducted to determine the gonadal development cycle of P. niger for a period of one year from March 2018 to February 2019. Fish specimens were collected from the BFDC (Bangladesh Fisheries Development Corporation) fish landing center (Latitude: 21° 45ʹ 19.95ʺ N, Longitude: 91° 96ʹ 80.96ʺ E), Cox’s Bazar, Chattogram, Bangladesh. During this study period about 180 fishes was collected.
Sampling site
Plate 2: The map of sample collection site
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After collecting the fish, they were brought to the Molecular Biology and Biotechnology laboratory of the Faculty of Fisheries, Chattogram Veterinary and Animal Sciences University, Chattogram for further study.
3.2 Recording of length-weight data and determination of length-weight relationship
Total length (TL, is the length of a fish measured from the tip of the snout to the tip of the caudal fin) and standard length (SL, is the length of a fish measured from the tip of the snout to the end of its last vertebrae) of each fish was measured by using a measuring scale to the closest 0.1 centimeter (cm) and the body weight of each fish was measured in gram (g) using an electrical balance (REDWAG, WRT12F1/NV).
Plate 3: Recording of length-weight data of P. niger
The relationship between Length (TL) and weight (W) of fish is usually expressed by the formula based on the exponential equation (Le Cren, 1951) in the form of-
TW = qSLb
Where,
TW is the total weight (expressed in “g”),
SL is the standard length (expressed in “cm”),
“q” is a co-efficient related to body form and
“b” is an exponent indicating isometric growth when equal to “3”
In this equation the parameters ‘q’ and ‘b’ are constant, where the parameter b (also known as the allometry co-efficient) has an important meaning in biological science,
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indicating the rate of weight gain relative to length. This power curve equation was converted into linear form by the use of natural logarithms as (Le Cren, 1951):
Ln TW = Ln q + b Ln SL
This equation is equivalent to regression equation:
Y = a + bX
Where “Y” is equivalent to Ln TW, “a” which represents the intercept of regression line is equivalent to Ln q, “b” is the slope of the line and “X” is equivalent to Ln SL.
After putting the length on X axis and weight on Y axis in excel a scatter diagram was drawn. Then length-weight data was converted to natural logarithms where,
Y-axis = Ln weight (g)
X-axis = Ln standard length (cm)
By using the converted length-weight data again a scatter diagram was plotted. From that diagram the value of q and b was calculated.
3.3 Determination of condition factor
Condition factor (K) is considered as one of the important factors influencing body composition. It is a measurement of the general health condition of fish as calculated by the ratio of body weight to body length. It is also often employed to measure the effects of environmental factors. Condition factor/ Ponderal index/ Fulton’s condition factor (Fulton, 1904) was estimated using formulae:
K= W*100/L3
Where,
W = Weight of fish in gram (g)
L = Length of fish in centimeter (cm)
3.4 Determination of relative condition factor
Information regarding seasonal variations of the condition of a fish in relation to both the internal and external environment becomes an utmost necessity in fishery biological studies. The condition of a fish may be driven to great variations due to
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physiological, environmental, nutritional and biological cycles. Finding of the relative condition factor (Kn) has got great significance for understanding the nutritional and biological cycle of a fish. Variation from the expected weight for length of individual fish or groups of individuals is an indication of condition (Le Cren, 1951) of the fish due to several factors like fatness, general wellbeing or gonad development.
Relative condition factor ‘Kn’ (Le Cren, 1951) was estimated by using the following formula:
Kn = W / ^w
Where,
W = Actual weight of fish in gram (g)
^w = Expected weight [w = (Log W* = log a + b log L)] Where,
W* = Average of W
In every month condition factor and relative condition factor were calculated from the monthly samples, which were used to detect seasonal variations in the condition of fish.
3.5 Collection and preservation of gonad and liver
To characterize the gonadal maturation it is necessary to dissect the fish and collect the gonads from both sexes. After recording length-weight data the fish was cut ventrally to collect the gonad. At first the fish was placed on a tray, and then it was cut from the anus towards the lower jaw by using a scissor. Then the muscle of the abdomen was cut vertically from the anus towards the vertebral column to clearly observe the gonad. After removing the unwanted fat, blood vessels and intestine, the gonad was collected carefully by using a forceps.
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Dissection
Ovary
Liver
Ovary Liver
Plate 4: Collection and preservation of gonad and liver
3.6 Determination of hepato-somatic index (HSI) and gonado-somatic index (GSI)
GSI is the ratio of fish gonad weight to body weight, which is particularly helpful in identifying the spawning seasons, as the ovaries of gravid female increases prior to spawning period. To determine the spawning period gonado-somatic index (GSI) plays a major role as there is a cyclic change in gonad weight in relation to total body weight (Nieland and Wilson, 1993; Jons and Miranda, 1997; Smith, 2008). An increasing in GSI suggests that spawning season is approaching, where decrease in GSI suggests spawning has occurred. To study GSI (gonado-Somatic Index) and HSI (hepato-somatic index) weight of ovary and liver of each specimen were measured by electronic balance (EK600 Dual, U.A.E). The GSI was calculated by month-wise and sex-wise using the equation (Vladykov, 1956).
GSI = Weight of gonad (g)/ Total body weight of fish (g) x 100
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Liver is the vital organ of the body which performs various physiological functions such as glycogenesis, detoxify toxic substances, and destroy the old RBC etc. The weight of liver is related with body weight. The healthy condition of liver is an indicator of healthy life. HSI is the ratio of liver weight to body weight, which gives information about the condition or status of liver and the body. So to know the physiological state of the fish, the hepato-somatic index (HSI) plays a vital role. To know the nutritional state of the fish, the hepato-somatic index (HSI) was calculated on monthly basis by using the equation (Busacker et al. 1990).
HSI = Liver weight (g)/ Total body weight of fish (g) x 100
3.7 Size at first Sexual Maturity (Lm)
Size at first maturity (Lm) is the length at which 50% of the fish have reached maturity. The Lm was estimated by using multiple functions such as the relationship of (i) gonado-somatic index, GSI vs total length (TL), (ii) Modified gonado-somatic index, MGSI vs TL and (iii) Dobriyal index, DI vs. TL (Khatun et al. 2019).
The GSI, MGSI, and DI which were calculated by Nikolsky (1963) as
GSI (%) = (GW/BW) × 100;
MGSI (%) = (GW/BW-GW) × 100; and
Dobriyal et al. (1999) as DI =
Where, BW= body weight, GW= gonad weight
3.8 Estimation of fecundity
Gravimetric method was used to estimate the fecundity (Hunter et al. 1989). After the collection of gonad, excess water was removed with the blotting paper. At first 3-4 subsamples from different part of the ovary was cut and then weighed the subsamples. After weighing, the numbers of eggs was counted for each sample and fecundity was determined by the following formula:
Fecundity = (Total ovary weight/ Weight of subsample) x no. of eggs in subsample
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Plate 5: Counting of eggs
3.9 Estimation of oocyte diameter
The diameter of the oocyte was measured every month by ocular microscope in micrometer, (with Optika Vision Lite 2.1 software; Optica B-190 Series, Italy).
Plate 6: Measurement of oocyte diameter (µm)
3.10 Histological analysis of gonad
To determine the specific spawning season and spawning frequency in a breeding season, study of gonadal development cycle and histological examination of gonads are important (Conover, 1992). For histological analysis the gonads were passed through the following procedures:
3.10.1 Fixation
At first the samples were preserved in a vial with Bouin’s fixatives for maximum 24 hours for fixation. Then the samples are washed with tap water to drain out the fixative. Samples were kept in 50% ethanol for 2 or 3 hours and then preserved in
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70% ethanol for future use. Dehydration process of sample was done immediate after the fixation.
Plate 7: Sample preservation in Bouin’s fixative
3.10.2 Washing of fixed tissue
After fixation, excess fixatives were washed out from the tissue to prevent interference with subsequent processes during the histological procedure. Washing was done in running water for about 2 or 3 hours before dehydration.
3.10.3 Dehydration
After washing the gonads were taken out and cut into small pieces (about 1cm) and put into pre labelled cassettes separately. Then the tissue blocks were passed through successive ascending concentration of graded alcohol for dehydration (Table-1). Dehydration is necessary for ideal consistency of tissue for sectioning into thin slice.
Plate 8: Dehydration of tissues in alcohol
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Table 1: The dehydration schedule
Sl no. Solution Time 1 Ethanol 50% 2 hours 2 Ethanol 70% 2 hours
3 Ethanol 80% 2 hours 4 Ethanol 90% 2 hours 5 Ethanol 95% 2 hours 6 Ethanol 100% (I) 2 hours
7 Ethanol 100% (II) 2 hours
3.10.4 Cleaning
After dehydration the blocks were passed through successive changes of xylene until the alcohol from the tissue is replaced by xylene (Table-2).
Plate 9: Cleaning of tissues
Table 2: The cleaning schedule
Sl no. Solution Time
1 Ethanol + xylene (50%+50%) 2 hours 2 Xylene 1st use 2 hours
3 Xylene 2nd use 2 hours
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3.10.5 Infiltration
After clearing, the tissue blocks were placed in melted paraffin in the oven usually at 60℃. Heat causes evaporation of xylene and the space in the tissue become infiltrated with paraffin (Table- 3).
Table 3: The infiltration schedule
Sl no. Solution Time
1 Paraffin + xylene (50%+50%) 2 hours 2 Paraffin 2 hours
3 Paraffin 2 hours 4 Paraffin 2 hours
3.10.6 Embedding
After that each tissue sample was taken from the melted paraffin and put in a block and filled with molten paraffin. Then the embedded blocks were allowed to cool in room temperature.
Plate 10: Embedded sample
3.10.7 Trimming
Trimming is a process in which the unwanted wax layers of the embedded blocks are removed by knife to obtain suitable blocks. Trimming allowed easy sectioning. In this step both side trimming and surface trimming were conducted.
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Plate 11: Trimming of unwanted paraffins
3.10.8 Sectioning
The small embedded paraffin blocks with tissue is sectioned by the hand rotatory microtome (KD-2258 Rotary Microtome, China) at 5µm size. (1µm = meter = 1/1000 cm).
Plate 12: Sectioning by using rotary microtome
3.10.9 Floating of section in water bath
After sectioning, the ribbon likes sections were floated in water bath (42oC) below temperature of melting point of paraffin.
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Plate 13: Floating of section in water bath
3.10.10 Attachment of section on glass slide and drying
The well spread ribbons of section from water bath were transferred to glass slides treated with adhesive (gelatin, egg albumin). The glass slides with section (s) were then dried in a slide warmer at temperature 37 oC for few hours.
Plate 14: Attachment of section on glass slide and drying
3.10.11 Staining
After drying, the slide with tissue section was ready for staining. Following steps (Table- 4) were followed in staining the tissue for hematoxylin and eosin stain:
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Table 4: The staining schedule
Sl. No. Treatment Time Effect 1 Xylene – 1 2 min Deparaffinization 2 Xylene – 2 2 min 3 100% Ethanol -I 2 min 4 100% Ethanol - II 2 min 5 95% Ethanol 2 min Rehydration 6 70% Ethanol 2 min 7 Running water 2 min 8 Hematoxylin 10 min 9 Running water 2 min 10 1% Acid alcohol 2 – 4 dips To differentiate by removing stain (short) from other constituents, check with microscope, Nucleus should be distinct and background very light or colors 11 Running water 5 min 12 Lithium carbonate 4 – 8 dips Alkaline medium, until sections are (as bright blue. required) 13 Running water 10 min 14 1% eosin 1 min 15 70% Ethanol 30 sec 16 95% Ethanol 2 min Redehydration 17 100% Ethanol 2 min 18 100% Ethanol 2 min 19 Ethanol + xylene 2 min (50%+50%) Cleaning and removal of alcohol 20 Xylene- 1 2 min 21 Xylene- 2 2 min
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3.10.12 Mounting
After staining, tissue section with glass slide was protected by thin cover slip. Required drop of DPX was put on each slide followed by attachment of cover slip and mounted slides are allowed to harden.
Plate 15: Mounting of stained sample slide
3.10.13 Microscopic examination of the gonad tissue sections
The mounted slides were observed under microscope (Optika; B-190 Series, Italy), which was connected to computer with digital camera. By the help of this mechanism required photographs were taken at different magnifications.
Plate 16: Microscopic observation of gonad tissue
3.11 Statistical analysis of data
All statistical analysis and recording were performed using computer based software Microsoft Excel, 2013. Data were expressed as mean ± standard deviation (S. D.) of mean.
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CHAPTER-4
RESULTS
CHAPTER-4
RESULTS
The results section presents the finding of an investigation in a systematic, logical, concise and comprehensive manner. This section contains detailed information of the present research work about length-weight relationship, condition index, fecundity, oocyte diameter, HSI, GSI and the histological changes in the gonads of P. niger throughout the year.
4.1 Length-weight relationship
Length-weight relationships are very important in fisheries management for comparison of growth studies. Data value obtained for P. niger in this experiment to calculate the intercept (Log a) and slope (b) of the regression analysis were -1.1957 and 2.7825, respectively. The standard length-weight relationship based on pooled data was W = 0.06372* SL2.7825 (R2 = 0.9562), the logarithmic equation being log W = -1.1957 + 2.7825 logSL (Figure-1). The ‘b’ value of the standard length-weight relationship indicates the negative allometric growth (b < 3) for both sexes of P. niger. The length-weight relationship for different sexes (Male and Female) was best described by the following exponential equations of body weight (W) against total length (TL):
W = 0.038388* TL2.7842 (Females, R2 = 0.9478) and the logarithmic equation being log W = -1.4158+2.7842 logTL (Figure-2). The ‘b’ value of the standard length- weight relationship of female indicates the negative allometric growth (b < 3).
W = 0.036183*TL^2.8022 (Males, = 0.9538) and the logarithmic equation being log W = -1.4415+2.8022 log TL (Figure-3). The ‘b’ value of the standard length- weight relationship of males indicates the negative allometric growth (b < 3).
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Figure 1: Logarithmic relationship between the standard length and weight based on pooled data of P. niger. The ‘b’ value indicates an allometric growth (negative) pattern of P. niger. The coefficient of determination (R2 = 0.9562) revealed that 95.62% of the variation in body weight due to variation in standard length in the sample collected over the period from March 2018 to February 2019
Figure 2: Logarithmic relationship between the total length and weight of female P. niger. The ‘b’ value indicates an allometric growth (negative) pattern of P. niger. The coefficient of determination ( = 0.9478) revealed that 94.78% of the variation in body weight due to variation in total length in the sample collected over the period from March 2018 to February 2019
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Figure 3: Logarithmic relationship between the total length and weight of male P. niger. The ‘b’ value indicates an allometric growth (negative) pattern of P. niger. The coefficient of determination (R2 = 0.9538) revealed that 95.38% of the variation in body weight due to variation in total length in the sample collected over the period from March 2018 to February 2019
4.2 Condition factor and relative condition factor
Condition factor (K) and relative condition factor (Kn) were calculated in every month to observe the overall health condition of the fishes. Condition factor (K), measures the deviation of an individual from the average weight for length and relative condition factor (Kn), measures the deviation from the hypothetical ideal fish. The mean values of condition factor and relative condition factor, ‘K’ and ‘Kn’ for pooled data of P. niger were calculated for each months throughout the year. The monthly mean K values obtained varied from 3.041±0.214 to 3.475±0.187 (Table-5) with an average value of 3.204±0.128 and the monthly mean Kn values obtained varied from 0.949±0.068 to 1.063±0.049 (Table-5) with an average value of 1.000 ±0.036. It was found that, the mean K was minimum (3.041±0.214) during May and maximum (3.475±0.187) during June, whether the mean Kn was minimum (0.949±0.068) during April and maximum (1.063±0.049) during June.
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Table 5: Condition factor and relative condition factor data of P. niger of 12 months. Data are presented as mean ± standard deviations for each month over the period from March 2018 to February 2019
Month Mean K ±S.D. Mean Kn± S.D.
March, 2018 3.100±0.309 0.978±0.084 April, 2018 3.045±0.227 0.949±0.068
May, 2018 3.041±0.214 0.967±0.062 June, 2018 3.475±0.187 1.063±0.049
July, 2018 3.210±0.197 0.988±0.053 August, 2018 3.269±0.357 0.977±0.108
September, 2018 3.287±0.219 1.022±0.065 October, 2018 3.165±0.106 0.978±0.037 November, 2018 3.324±0.201 1.053±0.065
December, 2018 3.095±0.150 0.987±0.045
January, 2019 3.171±0.171 1.001±0.052 February, 2019 3.264±0.146 1.039±0.044 Average 3.204 ±0.128 1.000 ±0.036
Month wise value of condition factor ‘K’ and relative condition factor ‘Kn’ for both sex were calculated and represented in the Figure-4 and Figure-5, respectively. This one year comparison showed that there was a similar trend between condition factor ‘K’ (Figure-6) and relative condition factor ‘Kn’ (Figure-7) of male and female throughout the year.
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Figure 4: Condition factor based on pooled data of P. niger of 12 months. Data are presented as mean ± standard deviations for each month over the period from March 2018 to February 2019
Figure 5: Relative condition factor based on pooled data of P. niger of 12 months. Data are presented as mean ± standard deviations for each month over the period from March 2018 to February 2019
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Figure 6: Condition factor (K) of male and female of P. niger. Data are presented as mean ± standard deviations for each month over the period from March 2018 to February 2019
Figure 7: Relative condition factor (Kn) of male and female of P. niger. Data are presented as mean ± standard deviations for each month over the period from March 2018 to February 2019
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4.3 Hepato-somatic index (HSI)
The ratio of liver weight to body weight is an indicator of energy reserves in animal in different environmental conditions. Monthly changes of HSI values for both male and female P. niger were calculated and the results are presented in (Figure-8). The minimum mean value of HSI was 0.512±0.059 in males found in July and 0.589±0.072 in females found in June. The maximum mean values of HSI were found both in February, (1.361±0.186) for males and (1.513±0.194) for females. There was also a second crowning of HSI value in November (Female= 0.960±0.100 and Male= 0.829±0.063). This line chart comparison shows that there was a similar trend between male and female HSI throughout the year and increased significantly before the spawning season (Figure-8).
Though there was a similar trend between male and female HSI throughout the year, there was no apparent correlation among hepato-somatic index (HSI) and condition factor (K) of male and female P. niger (Figure-9).
Figure 8: Hepato-somatic index (HSI) of male and female P. niger. Data are presented as mean ± standard deviations for each month over the period from March 2018 to February 2019
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Figure 9: Hepato-somatic index (HSI) and condition factor (K) of P. niger based on pooled data. Data are presented as mean ± standard deviations for each month over the period from March 2018 to February 2019
4.4 Gonado-somatic index (GSI)
In this study both male and female were taken into consideration for calculation of GSI throughout the year. The gonado-somatic index for each fish was calculated and averaged for each month.
Highest GSI value (4.022±1.299) for female was found in April whereas lowest GSI value (0.122±0.087) was observed in August. To be noted that another high value of GSI was also observed in October (0.523±0.593) which indicates the second phase of ovarian maturation in female. Mean GSI value of female P. niger increased significantly from month of February onwards and reaches at peak at April (Figure- 10). GSI value drastically decreases in of June in female and thereafter months and again slight increased GSI value was observed in the October compared to adjoining months which indicates the second phase of ovarian maturation in female.
In male, the mean GSI value ranged from 0.067±0.036 to 0.977±0.255. GSI value drastically decreases in April in male and thereafter months (Figure-11). Again slight increased GSI value was observed in October in male P. niger. The comparison of male-female GSI suggested a significant correlation between male-female GSI (Figure-12). There was no apparent correlation between GSI and HSI (Figure-13).
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Figure 10: Monthly variations of gonado-somatic index (GSI) in female P. niger. Data are presented as mean ± standard deviations for each month over the period from March 2018 to February 2019
Figure 11: Monthly variations of gonado-somatic index (GSI) in male P. niger. Data are presented as mean ± standard deviations for each month over the period from March 2018 to February 2019
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Figure 12: Monthly variations of gonado-somatic index in male and female P. niger. Data are presented as mean ± standard deviations for each month over the period from March 2018 to February 2019
Figure 13: Monthly variation of gonado-somatic index (GSI) and hepato-somatic index (HSI) of P. niger based on pooled data. Data are presented as mean for each month over the period from March 2018 to February 2019
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4.5 Oocyte diameter
In this study, annual increase in the oocyte diameter of P. niger was observed every month throughout the year and compared with the GSI of females. Data of oocyte diameter shows significant correlation with the GSI changes, which suggests a synchronize development of the gonads with the increasing size of eggs. Maximum oocyte diameter (Figure-14) was found in April (285.529±35.099µm) and minimum oocyte diameter was found in July (28.324±5.279µm) where new eggs initiate development for the following spawning season. Another rise in the oocyte diameter was also observed in October (88.930±62.915µm). The comparative study between oocyte diameter and GSI (Figure-15) evidently shows that the oocyte starts developing as GSI increase from January and gradually becomes ripe in March, finally spawns in April and May.
Figure 14: Oocyte diameter of P. niger collected from Bay of Bengal. Data are presented as mean ± standard deviation
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Figure 15: Oocyte diameter and GSI of P. niger collected from Bay of Bengal. Values are presented as mean ± standard deviation
4.6 Size at first sexual maturity (Lm)
Size at first sexual maturity was measured by identifying the starting point of sudden GSI increase. Size and age at sexual maturity are strongly correlated with growth, maximum size and longevity. Growth of a species also very important to study about the reproductive cycles for better understanding and management in an ecosystem.
Figure 16: Relationship between gonado-somatic index (GSI%) with total length of female P. niger
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The relationship between TL vs GSI, MGSI and DI of female P. niger are shown in (Figure-16, 17 and 18). The GSI and MGSI was low (< 2.5%) for females smaller than 28 cm TL. DI was also low (< 2.2) at the same length. The GSI, MGSI (> 2.5%) and DI (> 2.2) rose sharply around at 28 cm TL in female.
Figure 17: Relationship between modified gonado-somatic index (MGSI%) with total length of female P. niger
Figure 18: Relationship between Dobriyal index (DI) with total length of female P. niger
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In case of male, the GSI and MGSI was low (< 1.1%) for fishes smaller than 26.5 cm TL (Figure-19 and 20). DI value was also low (< 1.1) at the same length (Figure-21). The GSI, MGSI (> 1.1%) and DI (> 1.1) rose sharply around at 26.5 cm TL in male.
Figure 19: Relationship between gonado-somatic index (GSI%) with total length of male P. niger
Figure 20: Relationship between modified gonado-somatic index (MGSI%) with total length of male P. niger
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Figure 21: Relationship between Dobriyal index (DI) with total length of male P. niger
4.7 Fecundity
The fecundity was determined from randomly collected gravid female fish samples. Size of the gravid female ranging from 23.7cm (392 g weight) to 24.1cm (436.2 g weight) in standard length has ovaries weighing from 13.537g to 19.365g respectively. Fecundity was counted by gravimetric methods and was found to vary throughout the spawning season. The fecundity was found to vary from 114615 to 127605 with a mean value of 119433±5340 eggs.
4.8 Gonadal maturation stages in female Gross ovarian stages and monthly ovarian development of the P. niger were studied throughout the year. Each of the histological ovarian sections was studied thoroughly and described the different vitellogenic stages of ovarian maturation.
The following maturity stages of ovarian development were identified in P. niger:
4.8.1 Pre-vitellogenic stage
This stage is often called primary growth phase or immature stage of ovarian development, have found in July (Plate-17A), August (Plate-17B), November (Plate- 17C) and December (Plate-17D). Ovary was small, thread-like and of equal length. It is translucent in immature virgins and reddish in recovering spents due to strong vascularization. This stage is characterized with a number of oogonia (Oo), chromatin
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nucleolus (CN) and early perinucleolus (EPN) and number of primary growth oocytes. Oocytes were very small in diameter and the cytoplasm was densely stained with hematoxylin as evidenced by large and bright nucleus containing peripheral nucleoli surrounded by basophilic cytoplasm.
A B Oo
CN
OL OC BV OL CN Oo
OC 100 µm 100 µm
C D
PO
Oo OL CN CN
Epo OC
100 µm 100 µm
Plate 17: Different stages of ovarian development of P. niger. Pre-vitellogenic stage (A, B, C and D). CN- Chromatin nucleolar, PO- Perinucleoar oocyte, Epo- Early perinucleolarar oocyte , Oo- Oogonia, OC- Ovarian cavity, OL- Ovarian lamellae, BV- Blood vessel
4.8.2 Early primary vitellogenic stage
The early primary vitellogenic stage is sometimes called the early maturing or secondary growth phase of the oocyte. Ovary increases in size, pale-yellow to dark pink in color and occupies about half of the peritoneal cavity. In this stage ovary filled with early perinucleolar oocyte (Epo) and late perinucleolar stage oocytes (Lpo). At the beginning of the vitellogenic stage, the oocyte diameter increased than the previous stage and became transparent and pinkish in color. Blood capillaries in this
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stage were still not clear. This stage have found in January (Plate-18E) and September (Plate-18F).
E F
Lpo Ygr PO CN Lpo
YV CN
Epo 100 µm 100 µm
G H PO
PO Ygr GV GV
ZR Pof YV Ygl Ygr 100 µm 100 µm
Plate 18: Early primary vitellogenic stages (E and F) and advanced primary vitellogenic stages (G and H) of ovarian development of P. niger. GV- Germinal vesicle, CN-Chromatin nucleolar, PO- Perinucleolar oocyte, Epo- Early perinuclear oocyte, Lpo- Late perinucleolar oocyte, Ygr- Yolk granule, Ygl- Yolk globule, YV- Yolk vesicle, ZR- Zona radiate, Pof= Post ovulatory follicle
4.8.3 Advanced primary vitellogenic stage The number and size of the yolk vesicle increased significantly than the prior stages. Zona radiate becomes visible at the periphery at the end of this stage. Some advanced oocytes reach to the primary globular stage. Oocyte appeared more rounded with small vacuole. Ova were visible to naked eye and nucleus was partly hidden by yolk. This is the developing stages of ovary and primarily found in February (Plate-18G) and October (Plate-18H).
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4.8.4 Secondary vitellogenic stage The secondary vitellogenic stage is accompanied with the accumulation of eosinophilic yolk globule in the inner cortex. The cytoplasm was mostly covered with yolk globules sometimes with large oil droplet. The nucleus contains some peripheral nucleoli. The zona radiata has increased its thickness. Ova in this stage were spherical and yellowish or orange in color. Blood vessel was appeared in the surface of the ovary. The number of ova can be counted by necked eye. This stage was found in March (Plate-19I).
I J Ygr ZR ZR Ygl Ygl Ygr
GV OC
100 µm 100 µm
K L Ygr Pof Pso
Oo
Ygl Pof
ZR CN
100 µm 100 µm
Plate 19: Secondary vitellogenic stage (I), Ripe stage (J and K) and Spent or regressing stage (L) of P. niger. YV- Yolk vesicle, Ygl- Yolk globule, ZR- Zona radiata, Ygr- Yolk granule, OC- Oocyte cavity, CN- Chromatin nuclear, Oo- Oogonia, Pof- Post ovulatory follicle, Pso- Partially spent ovary.
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4.8.5 Ripe stage Ovary was swollen, maximally distended and yellowish, occupying majority of the body cavity. Eggs were clearly visible throughout the thin ovary wall. A network of blood vessels surrounds the organ. The oocyte rapidly increased its weight absorbing water to become fully hydrated. Post-ovulatory follicle often co-existed with the yolk granules in this stage. Oil droplets also began to fuse into a single oil droplet, and the nucleus moved towards the animal pole. Ovary was fully large in this stage, yellowish or orange in color and occupied the entire body cavity. Blood vessels were conspicuous. Large ova were often come out from the vent with a slight pressure in the belly. This stage was found in April (Plate-19J) and May (Plate-19K).
4.8.6 Spent or regressing stage
The weight of the ovary drastically became low due to release of eggs. Ovary was reddish and flaccid, shrunken, hollow, sac like structure, small number of immature eggs were present sometimes with denatured ova. Ovary of totally spent females contain numerous post-ovulatory follicles at different stages of degeneration, atretic oocytes and a reserve stock of oogonia and perinucleolar stage oocytes; ovary of partially spent female contain, additionally, oocytes in different vitellogenic stages. Spent fishes were found in June (Plate- 19L). In this species, some mature like eggs were also persisting in some fishes indicating the second chance of spawning.
4.9 Gonadal maturation stages in male:
On the basis of histological observations and degree of spermatogenesis, the annual testicular activity of P. niger was divided into the following developmental stages:
4.9.1 Immature stage
Testes in this stage were small, transparent and pale whitish in color. They occupied a very small portion of the body cavity. The seminal lobule of testes contained a large number of spermatogonia with few number of spermatocyte. Spermatogonia are spherical in shape and stained with hematoxylin. This stage was found in July (Plate- 20A), August (Plate-20B), November (Plate-20C) and December (Plate-20D).
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A B
SG
SC SC
GE E GE SC
100 µm 50 µm
C D
PS SG SC SG GE GE PS SG
L L
50 µm 50 µm
Plate 20: Gonadal maturation cycle of testes of P. niger. Immature stage (A, B, C and D). GE- Germinal epithelium, SG- Spermatogonia, PS- Primary spermatocyte, SC- Spermatocyte, SZ- Spermatozoa L- Lumen
4.9.2 Developing stage
Testes are slightly larger, flat and translucent in color. Germinal epithelium was seen throughout the testes in this stage. Spermatocytes formation started. Primary spermatocyte, secondary spermatogonia, secondary spermatocyte were also seen. Spermatogonium has noticeable basophilic nucleoli and stains very faintly, some become degenerated. No spermatozoa in the lumen. This stage was found in September (Plate-21E) and October (Plate-21F).
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E- SEP F- OCT ST
SG SG SC
ST L
GE E 50 µm 50 µm
Plate 21: Gonadal maturation cycle of testes of P. niger. Developing stage (E and F). SC- Secondary Spermatocyte, SG- Spermatogonia, TL- Testicular Lamellae, GE- Germinal epithelium, ST- Spermatids, L- Lumen
4.9.3 Pre-spawning stage
Testes were in late developing stages with spermatozoa in the lumens of the sperm ducts. All stages of spermatogenesis like spermatogonia, spermatocyte and spermatids were usually seen in this stage, among them spermatocytes were predominant in the sperm tissue. Many cysts containing spermatocytes were seen throughout the seminal lobule. Testes were then looking creamy white. This stage was found in January (Plate-22G) and February (Plate-22H).
G H TL
LL SG SC ST ST SS
SG 50 µm 50 µm
Plate 22: Gonadal maturation cycle of testes of P. niger. Pre-spawning stage (G and H). SC- Secondary Spermatocyte, SG- Spermatogonia, ST- Spermatids, SS- Secondary spermatocyte, TL- Testicular lamellae, LL- Lobular lumen
4.9.4 Ripe stage
The testes became ripe within March (Plate-23I) and were ready for spawning. Sperm did not come out from the vent by gentle pressure on the belly, by which this stage
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can be distinguished from the spawning stage. Increase of spermatids in the seminal lobules and spermatozoa in the lobular lumen and sperm ducts was clear. The number of spermatogonia and spermatocyte decreased with an increment of the spermatids. The spermatozoa had a distinct spherical shaped head; sometimes tails were also visible but not always.
4.9.5 Spawning stage
In the study, the spawning male was found in April (Plate- 23J). Testes in this period were translucent, white with some blood slots. Sperm comes out with gentle pressure upon belly. Testes were large in size and dominated by mature spermatozoa in the peripheral and central sperm ducts and throughout the lumens.
I J
SZ TC
SC ST SZ SZ
SG 50 µm 50 µm
Plate 23: Ripe stage (I) and Spawning stage (J) of testes of P. niger. SC- Spermatocyte, SG- Spermatogonia, SZ- Spermatozoa, ST- Spermatids, TC- Testicular cavity
4.9.6 Post-Spawning stage:
The GSI value drastically reduced and found very minimum in the reproductive cycle. Some SZ present in partially shrunken testis, but empty spaces characterize fully shrunken testis. This stage was found in May (Plate-24K) and June (Plate-24L).
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K L
SG RS TC
RS
50 µm 50 µm
Plate 24: Gonadal maturation cycle of testes of P. niger. Post-spawning stage (K and L). SG- Spermatogonia, RS- Residual Spermatids, TC- Testicular cavity
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CHAPTER-5
DISCUSSION
CHAPTER-5
DISCUSSION
This research work was designed to describe breeding biology of P. niger. This chapter described the overall results comparison, achievement of the study and problems associated to the research work on the reproductive aspects of P. niger. The objectives of the research work were achieved by the study of related reproductive parameters and histological study of gonad throughout a year (March, 2018 to February, 2019). However, information on the breeding biology of P. niger in the Bay of Bengal of Bangladesh does not exist. Information of previous experience from other areas around the world was taken into consideration for breeding biology study of the species.
5.1 Length-weight relationship
The study of length-weight relationship of fishes has great significance with regards to their morphology, biology, growth, general well-being of fishes and also in differentiating the unit stocks, which in turn is useful in fisheries management. In general, growth of fishes or any other animal increases with the increase in body length. Thus, it can be said that length and growth are interrelated. Length-weight relationship is expressed by the ‘cube law’ i.e., the weight of the fish will be proportional to the cube of their length, based on its dimensional equality, which may be mathematically expressed as W = qLb (Allen, 1938; Brody, 1945 and Roy, 1987). The exponential forms of equations derived between standard length and weight of P. niger based on pooled data was W = 0.06372*SL2.7825. The exponential forms of equations derived between standard length and weight of female and male P. niger were W = 0.038388* TL2.7842 ( = 0.9478) and W = 0.036183* TL2.8022 (R2 = 0.9538), respectively. In the present study, the value of ‘b’ in P. niger for pooled data was found 2.7825 and 2.7842 in female, 2.8022 in male, which was lower than 3. If 'b' is equal to 3.0, growth is isometric, meaning shape does not change as fish grow and when the value 'b' greater than 3.0 indicates a population where fish become more rotund as length increases (positive allometric growth) and when the value of 'b' less than 3.0 represents fish becoming less rotund as length increases (negative allometric growth). In the study the slope b value was less than 3.0 which indicates that P. niger
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become more slender as they increase in length and the growth pattern was allometric not isometric.
Dadzie et al. (2008), Pati (1981) have also recorded the allometric growth in Kuwaiti waters and Bay of Bengal in P. niger.
Dadzie et al. (2008) recorded the exponent 'b' value of 2.788 to 2.796 in females, 2.726 to 2.732 in males, 2.779 to 2.784 for both sexes and 2.590 to 2.613 in juveniles, all indicating an allometric relationship for P. niger. The 'b' values recorded by Pati (1981) was 2.5411.
Contradictory to the above results, Daliri et al. (2012) reported isometric growth in Bushehr coastal waters, Northern Persian Gulf. Similarly, Tao et al. (2012) and Kazemi et al. (2013) reported isometric growth for P. niger in with ‘b’ value of 2.9811 (in the Taiwan Strait) and 2.919 (in the Persian Gulf and Oman Sea).
5.2 Condition factor and relative condition factor
Condition factor is an indicator of physiological state and general well-being of fishes involving maturity, spawning, environmental conditions and availability of food (Brown, 1957). Information regarding seasonal variations of the condition of a fish in relation to both the internal and external environment becomes an utmost necessity in fishery biological studies. The condition of a fish may be greatly varied due to physiological, environmental, nutritional and biological cycles. Finding of the relative condition factor has got great significance for understanding the nutritional and biological cycle of a fish. The number of variables that can affect the value of K is however considerable. Le Cren (1951) figured out that the condition factor 'K' was affected by factors correlated with length which affect the condition factor because the fish does not in fact follow the cube law in its length-weight relationship, by selection in sampling length, food supply, degree of parasitism etc., thus making the correlation and interpretation of these results sometimes difficult. To solve the problem regarding the condition factor, Le Cren (1951) suggested the use of relative condition factor 'Kn'. Le Cren (1951) pointed out that Kn was a function of fatness and condition of gonads. The Kn value also depends upon the stage, maturity of the gonads and length of the fish (Chakrabarty and Singh, 1963). According to Le Cren (1951), the value of Kn more than 1 indicates good health of the fishes and Kn less than 1 indicates poor condition of fish.
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The monthly mean K values obtained in this study were varied from 3.041±0.214 (May) to 3.475±0.187 (June) with an average value of 3.204±0.128. The monthly mean relative condition factor (Kn) values obtained during this study varied between 0.949±0.068 (April) to 1.063±0.049 (June) with an average value of 1.00±0.036. The average mean value of relative condition factor ‘Kn’ in this study was 1.00±0.036, which indicates good condition of the fish.
Relative condition values during the months of January, February, June, September and November were found above the mean value which indicates that fish was in good condition during these months. The observed maximum value of Kn (1.063±0.049) in these month might be attributed to the availability of foods, high feeding intensity and favorable environmental conditions in that period. The lowest Kn values during April may be due to the physiological, environmental, nutritional and biological cycles. The low Kn values were found when the fishes were in maturing or matured stage. The condition factor (K) of male and female has fairly similar trend throughout the year with highest value in June (Female = 3.437±0.106, Male = 3.513±0.216) indicating that the fish was in good condition immediate after spawning. Same trend also observed for relative condition factor (Kn) of male and female P. niger with the highest ‘Kn’ value in June, immediate after spawning (Female = 1.055±0.032, Male = 1.072±0.054).
Dadzie et al. (2008) reported that the Fulton’s condition factor was statistically discriminatory in determining the well-being of P. niger and there was no apparent correlation between GSI and mean monthly condition in Kuwaiti waters. They also reported that variation in average maximum condition was significant between sexes. Daliri et al. (2012) observed that the relative condition factor (Krel) did not differ significantly between species (P > 0.05) and ranged from 0.98±0.14 to 1.03±0.17 for five marine fish species including P. niger in Bushehr coastal waters, Northern Persian Gulf. The study of Kazemi et al. (2013) revealed that the mean relative condition factors (Krel) was 1.01±0.08 for P. niger.
5.3 Hepato-somatic index (HSI)
The HSI gives valuable information about the condition of liver and body and also about the impact of water pollution on fish body. HSI also provides an indication on status of energy reserved in fish. In poor water quality fish usually have a smaller
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liver with less energy reserved in the liver. HSI has been reported to decrease in fish exposed to water pollution. In this study, the maximum mean HSI value was found in February (1.45±0.20) which indicates the period of maximum growth in P. niger and minimum HSI value was found in July (0.55±0.13). When the food is available in large amount and conditions are favorable it causes increase in the HSI value. That means highest HSI value in February due to the good water quality, environmental conditions and food availability, whereas in July fishes had lowest HSI or smaller liver which indicates poor environment and loss food supply. HSI value was found highest in February for both male (1.361±0.186) and female (1.513±0.194) which suggests that the fishes were in good condition before the spawning. The relation between hepato-somatic index (HSI) and condition factor (K) was similar throughout the year except in June where HSI increased but ‘K’ value decreased.
Dadzie et al. (2008) observed that the fish (P. niger) remain in good condition throughout the spawning season which supports the present study.
5.4 Gonado-somatic index (GSI)
The term gonado-somatic index (GSI) is used as an indicator of gonadal maturity and hence the spawning season of a fish. In the pre-spawning period a gradual increase of the GSI, reaching a peak during the spawning period and a gradual decrease in the post-spawning period had been observed for P. niger.
In the study, both male and female were taken into consideration for calculation of GSI and the gonadal developmental changes in both male and female fishes were also observed intensively throughout the study time. The highest GSI value was observed in female in April (4.022±1.299) and in male in March (0.977±0.255). A second peak in GSI value was also observed in October (0.523±0.593) in female and continued increase in GSI from October up to the December was observed in male which indicates the second phase of gonadal maturation. The comparison of mean male and female GSI throughout the year was quite similar except in April where the female GSI was highest (4.022±1.299) but the male GSI (0.455±0.149) was low than the previous month. This suggests that the male fish of P. niger became mature earlier than the female. GSI values also indicates the prolonged spawning season of P. niger from February to May.
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After analysis of seasonal variations in the gonado-somatic index (GSI) during the two-year study period, Dadzie et al. (2009) found that the P. niger has a prolonged spawning season from February to September.
5.5 Oocyte diameter
Maximum oocyte diameter (Figure-14) was found in March (231.695±15.637µm), April (285.529±35.099µm) and May (269.435±27.968µm) where oocyte diameter rapidly increased February (172.908±89.899µm) and minimum oocyte diameter was found in July (28.324±5.279µm). This data suggests that the gonad of P. niger matures and spawns in March to May mainly with a second slight spawning in October. The comparison of oocyte diameter and GSI was similar throughout the year indicating that there was a positive trend between oocyte diameter and GSI. No similar study has been found for P. niger.
Jons and Miranda (1997) mentioned that the strong relationship between egg size and GSI was expected because GSI increases with egg maturity.
5.6 Size at first sexual maturity (Lm)
Size at first sexual maturity (Lm ) was estimated by using multiple functions such as the relationship of (i) Gonado-somatic Index, GSI vs.TL; (ii) Modified gonado somatic Index, MGSI vs. TL; and (iii) Dobriyal Index, DI vs. TL. The GSI and MGSI was low (< 2.5%) for females smaller than 28 cm TL. DI was also low (< 2.2) at the same length. The GSI, MGSI (> 2.5%) and DI (> 2.2) rose sharply around at 28 cm TL in female. In case of male, the GSI and MGSI was low (< 1.1%) for fishes smaller than 26.5 cm TL. DI value was also low (< 1.1) at the same length. The GSI, MGSI (> 1.1%) and DI (> 1.1) rose sharply around at 26.5 cm TL in male.
Therefore, the size at first sexual maturity was considered to be 28 cm TL and individuals with a GSI ≥ 2.5% could be roughly defined as mature female. Furthermore, the size at first sexual maturity was considered to be 26..5 cm TL and individuals with a GSI ≥ 1.1% could be roughly defined as mature male P. niger in the Bay of Bengal of Bangladesh.
Dadzie et al. (2008) reported length at first maturity ranged from 15 to 32 cm in males and 20 to 42 cm in females for P. niger. Pati (1983) reported 30 cm SL in female and 28 cm SL in male for P. niger in the Bay of Bengal.
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5.7 Fecundity
Knowledge of the fecundity of a species is an important factor in fish stock management. Fecundity is also one of the most important biological aspects of fish to assess the reproductive potential of a fish stock. The fecundity was determined from randomly collected gravid female fish samples. Size of the gravid female ranging from to 23.7cm (392g weight) to 24.1cm (436.2g weight) in standard length which have ovaries weighing from 13.537g to 19.365g, respectively. Fecundity was counted by gravimetric methods and found variable throughout the spawning season. The fecundity was found to vary from 114615 to 127605.
Dadzie et al. (2008) and Dadzie et al. (2009) found total fecundity of P. niger varied widely even among fishes of the same size, ranging from 71305 (for a 39.8 cm SL female weighing 1572.5g) to 3895449 (for a 49 cm SL female weighing 2630g), with a mean of 1216734 eggs. Pati (1983) reported that the number of eggs present in the ovary of ripe female (32.6 cm SL) was 86250.
5.8 Histological observation of gonad
Histological observations of the gonads showed that, increased number of yolk granules was present in the month from March to May in females. A number of post ovulatory follicles were also observed in the ovaries. In April and May, number of oil droplets were also fused together to form single droplet. Mature eggs were come out upon slight pressure on the belly in April and May which suggest the maturation of ovaries in these following months. In July, a number of atresia and empty spaces in the ovary were seen by histological observations. Few number of oogonia were also began to develop from September. Some yolk granule was also observed in the eggs in October in some fishes besides with chromatin nucleolar or perinucleolar stages. Few post ovulatory follicles were also seen in the ovaries which strongly suggest a second phase of spawning of this species.
In male, a number of mature spermatids were found in March and April which confirmed the maturity in males and also indicates that males mature first than females. In May and June empty ovary with some residual spermatids was observed. Though continued increase in GSI from October up to the December was observed in male, some developmental stages was only observed in September and October. Spermatogonial cells or spermatocytes and primary spermatocyte were observed in
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July, November and December indicating immature stage of testes in male P. niger. The study confirmed the relationship in gonadal maturity in both male and females in a synchronize manner in this species indicating the two spawning periods one in March to May and another in October.
Dadzie et al. (2009) reported that black pomfret has an extended spawning periodicity in the Arabian Gulf, beginning in February and ending in September. The evidence for their conclusion was derived from the presence of both males and females in the ripe/running condition in the samples from February till September. After a quiescent period from October till January, the increase in the GSI in both sexes from February, till their decline in September.
Pati (1983) informed that P. niger from Bay of Bengal has a spawning period from March to June.
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CHAPTER-6
CONCLUSIONS
CHAPTER-6
CONCLUSIONS
Fish reproductive biology is important in fisheries research, stock assessment and management. Sustainable, productive fisheries and aquaculture improve food and nutrition security, increase income and improve livelihoods, promote economic growth and protect environment and natural resources. A sustainable approach to fisheries and aquaculture will help to protect natural resources and ensure that fish stocks are available for future generations. Open water fisheries management greatly relies on the information of different stages of reproductive development of any fish population because reproductive traits of any particular species determine its intrinsic capacity and sustainability of exploitation. Fishing regulations must be ordained by complete understanding of the reproductive cycles and population dynamics of economically important fishes of the area concerned. But information about the reproductive biology of marine fishes of Bay of Bengal of Bangladesh is quite rare because of less research on them. To replenish the stock of P. niger, it is necessary to give enough time to the stock for reproduction. Another important reason to identify the breeding biology is to establish artificial breeding method of any species. The present study has been carried out with an aim to describe the spawning season and life history characteristics of the P. niger in the Bay of Bengal. After completing this study the spawning season of P. niger was identified from March to May with a minor peak in October. This information will be helpful to determine the ban period of catching P. niger from the Bay of Bengal of Bangladesh to replenish the stock as well as to establish artificial breeding.
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CHAPTER-7
RECOMMENDATIONS AND
FUTURE PERSPECTIVES
CHAPTER-7
RECOMMENDATIONS AND FUTURE PERSPECTIVES
The aim of this study was to describe the spawning season and life history characteristics of the P. niger in the Bay of Bengal. This study will be helpful for the fisheries management authority to set the ban period for fishing of P. niger during its breeding season and to maintain a sustainable stock. Also, this study will be helpful for the establishment of artificial breeding. Although a qualitative approach was followed to explore the objective of the research, there are some limitations of the study which can be minimized by following the recommendations:
Different sample collection site should be followed to identify the differences of spawning season based on the geographical location. Samples should be collected from authentic sources because authentic samples lead to a concrete result. Fresh and properly preserved samples give better histological diagram. So, samples should be collected directly from the fisherman immediately after catch. There is a scope of artificial breeding of P. niger which should be established by following their breeding biology.
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REFERENCES
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APPENDICES
APPENDICES
Appendix 1: Data of total length, standard length, body weight of P. niger
Month Total Standar Body Log10_TL Log10_SL Log10_W lengt d Length weight t h (cm) (g) (cm) March, 31 25.9 508.92 1.491 1.413 2.707 2018 29.6 24.8 455.38 1.471 1.394 2.658 30.7 25.6 515.3 1.487 1.408 2.712 31.9 26.6 521.68 1.504 1.425 2.717 27.6 23 414.07 1.441 1.362 2.617 32 26.9 504.4 1.505 1.430 2.703 32.6 27.4 575.08 1.513 1.438 2.760 26.7 22.3 401.9 1.427 1.348 2.604 27.9 23.5 426.1 1.446 1.371 2.630 31.2 26.1 552.8 1.494 1.417 2.743 26.9 22.4 402.7 1.430 1.350 2.605 30.5 25.5 523.4 1.484 1.407 2.719 29 24.3 430.17 1.462 1.386 2.634 April, 2018 27.5 22.9 401.4 1.439 1.360 2.604 26.6 22.5 404.4 1.425 1.352 2.607 28.9 24.1 436.2 1.461 1.382 2.640 27.4 23.2 391.4 1.438 1.365 2.593 28.7 24 390 1.458 1.380 2.591 28.3 23.7 382 1.452 1.375 2.582 28.6 24.1 409 1.456 1.382 2.612 28.4 23.9 398.2 1.453 1.378 2.600 28.8 24.1 397.2 1.459 1.382 2.599 29 24.2 475 1.462 1.384 2.677 28.2 23.7 392 1.450 1.375 2.593 26.9 22.6 345.6 1.430 1.354 2.539 27.2 22.8 327.2 1.435 1.358 2.515
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27.8 23.3 374.4 1.444 1.367 2.573 27.8 23.1 389.6 1.444 1.364 2.591 May, 2018 33.1 27.8 652.6 1.520 1.444 2.815 31.4 26.4 560.6 1.497 1.422 2.749 30.6 25.7 462 1.486 1.410 2.665 31.7 26.4 554 1.501 1.422 2.744 30.4 25.5 502 1.483 1.407 2.701 32 26.8 528.2 1.505 1.428 2.723 30 24.8 483.8 1.477 1.394 2.685 29.5 24.6 508.4 1.470 1.391 2.706 30.5 25.6 508.8 1.484 1.408 2.707 31 26.1 530.8 1.491 1.417 2.725 32 26.9 563.6 1.505 1.430 2.751 28 23.7 452.6 1.447 1.375 2.656 June, 2018 27.5 22.9 415 1.439 1.360 2.618 26.5 22.2 385.4 1.423 1.346 2.586 24.6 20.5 299.8 1.391 1.312 2.477 25.8 21.5 349.8 1.412 1.332 2.544 25.5 21.5 359.6 1.407 1.332 2.556 25 20.9 324.6 1.398 1.320 2.511 25.7 21.6 372.4 1.410 1.334 2.571 26.4 22.3 382 1.422 1.348 2.582 23.8 20.1 275 1.377 1.303 2.439 26.8 22.7 404.8 1.428 1.356 2.607 24.4 20.4 325.5 1.387 1.310 2.513 24.9 20.8 313.6 1.396 1.318 2.496 26.7 22.2 371.8 1.427 1.346 2.570 24.8 20.6 302.2 1.394 1.314 2.480 25.3 21 302.4 1.403 1.322 2.481 24.9 20.9 318.2 1.396 1.320 2.503 25.2 20.9 326 1.401 1.320 2.513
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32.5 27.3 592.4 1.512 1.436 2.773 July, 2018 22.9 19.1 239 1.360 1.281 2.378 27.4 22.8 361 1.438 1.358 2.558 25.5 21.3 293 1.407 1.328 2.467 24.2 20.4 263.4 1.384 1.310 2.421 23.6 19.7 245.2 1.373 1.294 2.390 35.9 30.3 824 1.555 1.481 2.916 27.4 22.7 379.6 1.438 1.356 2.579 30.1 24.9 484.6 1.479 1.396 2.685 23.4 19.5 240.6 1.369 1.290 2.381 29.3 24.3 441.2 1.467 1.386 2.645 23.1 19.5 234.2 1.364 1.290 2.370 25.2 21.2 362 1.401 1.326 2.559 24.7 20.6 294.8 1.393 1.314 2.470 25.6 21.3 321.8 1.408 1.328 2.508 25.4 21.1 298.6 1.405 1.324 2.475 24.9 20.7 282 1.396 1.316 2.450 36.5 30.6 879.6 1.562 1.486 2.944 August, 23.6 19.6 243.82 1.373 1.292 2.387 2018 23.6 19.9 235.6 1.373 1.299 2.372 23.7 19.7 258.2 1.375 1.294 2.412 22.3 18.5 206.6 1.348 1.267 2.315 21.8 18.3 206.2 1.338 1.262 2.314 23.1 19.1 228.2 1.364 1.281 2.358 22.3 18.8 220.5 1.348 1.274 2.343 24.2 20.1 279.8 1.384 1.303 2.447 23.1 19.1 182.4 1.364 1.281 2.261 23.4 19.6 228 1.369 1.292 2.358 23.2 19.4 230.4 1.365 1.288 2.362 23.5 19.5 249.2 1.371 1.290 2.397 24.5 20.6 271.6 1.389 1.314 2.434
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22.5 18.7 207.4 1.352 1.272 2.317 23.8 19.8 335.6 1.377 1.297 2.526 September 27.9 23.4 440.6 1.446 1.369 2.644 , 2018 24.8 20.5 273 1.394 1.312 2.436 27.4 23 415.2 1.438 1.362 2.618 27.1 22.7 445.2 1.433 1.356 2.649 27.3 23 398 1.436 1.362 2.600 27.2 22.8 388 1.435 1.358 2.589 30.2 25.3 510 1.480 1.403 2.708 23.8 20 246 1.377 1.301 2.391 26.2 22.1 348 1.418 1.344 2.542 30.7 25.8 504.6 1.487 1.412 2.703 28.6 23.7 454.6 1.456 1.375 2.658 29.1 24.1 461.6 1.464 1.382 2.664 26 21.8 363 1.415 1.338 2.560 33.6 28.2 669.2 1.526 1.450 2.826 27.1 22.8 397 1.433 1.358 2.599 October, 23.2 19.3 224.4 1.365 1.286 2.351 2018 22.7 18.8 215.4 1.356 1.274 2.333 22.2 18.5 219.6 1.346 1.267 2.342 22.6 18.8 205 1.354 1.274 2.312 20.1 16.7 152.2 1.303 1.223 2.182 22.3 18.5 201.6 1.348 1.267 2.304 27.2 22.7 370.4 1.435 1.356 2.569 30.8 25.5 517.4 1.489 1.407 2.714 31.5 26.4 560.2 1.498 1.422 2.748 29.6 24.5 458.2 1.471 1.389 2.661 37.2 30.9 936.6 1.571 1.490 2.972 31.2 25.9 554.6 1.494 1.413 2.744 30.9 25.6 515 1.490 1.408 2.712 31.1 25.9 532.6 1.493 1.413 2.726
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31.6 26.3 575.2 1.500 1.420 2.760 November, 30.2 25 485.6 1.480 1.398 2.686 2018 29.5 24.4 527 1.470 1.387 2.722 29.7 24.6 497.6 1.473 1.391 2.697 30.2 25 502.8 1.480 1.398 2.701 30.5 25.3 521.2 1.484 1.403 2.717 30.2 25 539 1.480 1.398 2.732 29.8 24.9 482.6 1.474 1.396 2.684 30.8 25.7 516.4 1.489 1.410 2.713 31.5 26.1 584 1.498 1.417 2.766 31.4 26 589.6 1.497 1.415 2.771 30.4 25.4 536.8 1.483 1.405 2.730 29.2 24.2 499 1.465 1.384 2.698 31.6 26.2 575 1.500 1.418 2.760 29.7 24.6 477.8 1.473 1.391 2.679 30.4 25.2 545 1.483 1.401 2.736 32.2 26.9 740.8 1.508 1.430 2.870 December, 31.1 26 518.6 1.493 1.415 2.715 2018 30.9 25.7 522.4 1.490 1.410 2.718 30.8 25.7 535.4 1.489 1.410 2.729 31.6 26.5 504.2 1.500 1.423 2.703 30.5 25.4 497 1.484 1.405 2.696 32.4 27.2 642.6 1.511 1.435 2.808 33.5 27.9 688.4 1.525 1.446 2.838 31.4 26.2 527.8 1.497 1.418 2.722 29.8 24.9 507.8 1.474 1.396 2.706 29.4 24.6 489.6 1.468 1.391 2.690 32.5 27.3 633.2 1.512 1.436 2.802 30.1 25.1 493.6 1.479 1.400 2.693 30.5 25.3 499.4 1.484 1.403 2.698 30.5 25.6 516.2 1.484 1.408 2.713
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31.2 26.1 575 1.494 1.417 2.760 January, 30.1 25 476.6 1.479 1.398 2.678 2019 28.8 24.2 463.8 1.459 1.384 2.666 28.5 23.9 437 1.455 1.378 2.640 29.7 24.9 511.6 1.473 1.396 2.709 30.1 25.2 489.2 1.479 1.401 2.689 29.1 24.4 513.4 1.464 1.387 2.710 31 25.7 540.2 1.491 1.410 2.733 31.2 26.1 598.2 1.494 1.417 2.777 30.9 25.8 525.2 1.490 1.412 2.720 28.4 23.6 408.2 1.453 1.373 2.611 29.6 24.8 454.8 1.471 1.394 2.658 29.3 24.6 453.8 1.467 1.391 2.657 28.9 23.9 454.6 1.461 1.378 2.658 31.2 26.2 517.4 1.494 1.418 2.714 30 24.9 493 1.477 1.396 2.693 February, 29.1 24.3 463 1.464 1.386 2.666 2019 31.8 26.7 575.6 1.502 1.427 2.760 29.4 24.5 495 1.468 1.389 2.695 30.8 25.8 603.8 1.489 1.412 2.781 31.8 26.7 619 1.502 1.427 2.792 33.4 27.9 701.8 1.524 1.446 2.846 29.4 24.7 497.8 1.468 1.393 2.697 29.8 24.9 535.6 1.474 1.396 2.729 33.1 27.5 699.4 1.520 1.439 2.845 29.7 24.6 480.6 1.473 1.391 2.682 31.3 26.3 593.8 1.496 1.420 2.774 29.4 24.3 455 1.468 1.386 2.658 33.4 28.2 666 1.524 1.450 2.823 30.2 25.3 519.4 1.480 1.403 2.716 31.4 26.4 619.2 1.497 1.422 2.792
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Appendix 2: Month wise hepato-somatic index (HSI) and gonado-somatic index (GSI) of P. niger (both male and female)
Month HSI (Mean ± S.D.) GSI (Mean ± S.D.) Female Male Female Male March, 2018 1.103±0.099 0.868±0.154 0.511±0.291 0.181±0.089
April, 2018 1.052±0.136 0.703±0.089 1.544±0.795 0.923±0.204
May, 2018 1.000±0.123 0.871±0.154 2.387±1.009 0.977±0.255
June, 2018 0.589±0.072 0.609±0.083 4.022±1.299 0.455±0.149
July, 2018 0.593±0.170 0.512±0.059 1.545±1.158 0.448±0.145
August, 2018 0.746±0.197 0.664±0.106 0.305±0.247 0.074±0.020
September, 0.765±0.139 0.709±0.146 0.237±0.250 0.068±0.057 2018 October, 2018 0.633±0.134 0.605±0.082 0.122±0.087 0.067±0.036
November, 0.960±0.100 0.829±0.063 0.331±0.283 0.069±0.051 2018 December, 0.730±0.079 0.626±0.047 0.523±0.593 0.219±0.142 2018 January, 2019 0.776±0.099 0.731±0.115 0.418±0.209 0.231±0.106
February, 2019 1.513±0.194 0.361±0.186 0.290±0.049 0.251±0.054
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Appendix 3: Condition factor and relative condition factor data of P. niger of 12 months (both male and female)
Month Mean K ±S.D. Mean Kn± S.D. March, 2018 3.100±0.309 0.978±0.084 April, 2018 3.045±0.227 0.949±0.068 May, 2018 3.041±0.214 0.967±0.062 June, 2018 3.475±0.187 1.063±0.049 July, 2018 3.210±0.197 0.988±0.053 August, 2018 3.269±0.357 0.977±0.108 September, 2018 3.287±0.219 1.022±0.065 October, 2018 3.165±0.106 0.978±0.037 November, 2018 3.324±0.201 1.053±0.065 December, 2018 3.095±0.150 0.987±0.045 January, 2019 3.171±0.171 1.001±0.052 February, 2019 3.264±0.146 1.039±0.044 Average 3.204 ±0.128 1.00 ±0.036
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Brief Biography
Brief Biography
Md. Abirul Islam completed B.Sc. in Fisheries (Hon’s) from the Faculty of Fisheries of Chattogram Veterinary and Animal Sciences University (CVASU), Chattogram, Bangladesh with CGPA 3.69 out of 4.00. He has strong passionate in research of fisheries and his research interests are on histology, identification of fish breeding season, development of breeding techniques, fish molecular biology, genetic engineering etc. Now, he is a candidate for the degree of MS in Fish Biology and Biotechnology under the Department of Fish Biology and Biotechnology, Faculty of Fisheries, Chattogram Veterinary and Animal Sciences University (CVASU).
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