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Summary
Abstract
Introduction
Protocol
Results
Discussion
Disclosures
Acknowledgements
Materials
References
자동 번역
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생물학

眼柄消融术可增加泥蟹卵巢成熟

Published: March 31, 2023
doi:
10.3791/65039
, , , , ,
1Higher Institution Centre of Excellence (HICoE), Institute of Tropical Aquaculture and Fisheries,Universiti Malaysia Terengganu, 2Centre for Chemical Biology,Universiti Sains Malaysia, 3Department of Aquaculture, Faculty of Fisheries,Kasetsart University, 4School of Biological Sciences,Universiti Sains Malaysia, 5International Research Center for Marine Biosciences, Ministry of Science and Technology,Shanghai Ocean University

Summary

对麻醉的雌蟹进行了两种眼柄消融方案（即烧灼和 手术方法）。泥蟹的眼柄消融加速了卵巢的成熟，而不会降低存活率。

Abstract

泥蟹（Scylla spp.）是具有重要商业价值的甲壳类动物，遍布印度-西太平洋地区。在养殖过程中，诱导卵巢成熟对于满足消费者对成熟泥蟹的需求和加速种子生产非常重要。眼柄消融术是促进泥蟹卵巢成熟的有效工具。然而，泥蟹的眼柄消融没有标准方案。在这项研究中，描述了两种眼柄消融技术：烧灼（使用热金属消融麻醉蟹的眼柄）和手术（使用手术剪刀去除眼柄）。在眼柄消融术之前，性成熟的雌性（CW > 86 mm）使用装有海水的冰袋（-20°C）麻醉。当水温达到4°C时，将冰袋从水中取出。流海水（环境温度：28°C）用于眼柄消融后立即从麻醉中恢复。在眼柄消融过程中或之后没有发生死亡。这里介绍的眼柄消融方案加速了泥蟹的卵巢成熟。

Introduction

属于Scylla属的所有四种泥蟹物种都是水产养殖中具有重要商业价值的甲壳类物种1，2。甲壳类动物（包括泥蟹）的生长及其从早熟（亚成体或青春期）阶段到性成熟（成年）阶段的转变是通过蜕皮过程发生的，该过程涉及较老和较小的外骨骼的周期性脱落。甲壳宽度 （CW）、螯部和腹部瓣形态被广泛用于确定镰刀属的性成熟度。3，4，5.蜕皮过程受各种激素的作用调节，需要大量的能量6。除了正常的蜕皮过程外，肢体的丧失，无论是自愿的还是由外部因素引起的，都会加速螃蟹的蜕皮，而不会影响它们的存活率7，8，9。因此，肢体自体切开术常用于软壳泥蟹养殖业的蜕皮诱导7，9。

单侧或双侧眼柄消融术在淡水虾和海虾中最为流行，用于性腺成熟和苗种生产10，11，12，13。甲壳类动物中常见的眼柄消融技术包括：（i）使用绳子14，15在眼柄底部结扎;（ii）使用热镊子或电灼装置烧灼眼柄16;（iii） 去除或直接捏住眼柄以留下开放性伤口12;（iv）用剃刀切开眼睛的远端部分后，通过切口去除眼柄内容物17.眼柄X器官是甲壳类动物中重要的内分泌器官，因为它们调节甲壳类动物的高血糖激素（CHH），蜕皮抑制激素（MIH）和玻璃体生成抑制激素（VIH）6，18，19，20，21，22。眼柄X器官（或窦腺复合物）合成并释放性腺抑制激素（GIH），也称为玻璃体生成抑制激素（VIH），属于神经肽激素家族6。单侧或双侧眼柄消融术可减少GIH合成，导致刺激激素（即促性腺激素，GSH）占主导地位，并加速甲壳类动物卵巢成熟过程23，24，25，26。没有眼柄消融后GIH的影响，雌性甲壳类动物将精力用于卵巢发育27。已经发现，单侧眼柄消融足以诱导甲壳类动物卵巢成熟11，虾和蟹的消融眼柄在几次蜕皮后可以再生28。Scylla spp.记录了四个卵巢发育阶段：i）未成熟（阶段-1），ii）早熟（阶段-2），iii）早熟（阶段-3）和iv）完全成熟（阶段-4）29，30。未成熟的卵巢阶段见于未成熟的女性。在青春期蜕皮和交配后，未成熟的卵巢开始发育并最终成熟（第 4 阶段），然后产卵31。

眼柄消融方案对于泥蟹亲鱼的发育和苗种生产至关重要。在全球食品市场中，具有完全成熟卵巢（第4阶段）的成熟泥蟹而不是肌肉含量较高的螃蟹受到消费者的青睐，因此具有更高的商业价值，甚至高于大型雄性。泥蟹的眼柄消融没有完整的方案。这项工作中的眼柄消融方案通过使用完全麻醉的螃蟹来最大限度地减少压力，并最大限度地减少螃蟹咬伤对人员的身体伤害。该协议简单且具有成本效益。在这里，我们提出了一种可以诱导性腺成熟的 Scylla spp .眼柄消融方案。测试了两种眼柄消融技术（烧灼和手术），并根据雌性泥蟹的性腺发育速度比较了它们的效率。

Protocol

该协议遵循马来西亚实验动物科学协会概述的马来西亚科学用途护理和使用动物行为守则。实验样品的牺牲是根据美国国立卫生研究院实验动物护理和使用指南（NIH出版物第8023号，1978年修订）进行的。性早熟雌性泥蟹（橙泥蟹 S. olivacea）是从马来西亚Setiu湿地的当地市场（5°66′62′′N，102°72′33′′E）收集的。根据形态特征鉴定泥蟹种类1. 1. ?…

Representative Results

性腺成熟在进行眼柄消融术之前，在100%的解剖女性（n = 6）中发现了乳白色的卵巢组织（未成熟的卵巢，1期）（图2）。与未进行眼柄消融的雌蟹相比，眼柄消融的雌蟹（n = 63;烧灼技术为31只雌蟹，手术技术为32只雌蟹）的性腺成熟率高于未进行眼柄消融的雌蟹（n = 31）（图3）。在眼柄消融的雌蟹中发现的早熟卵巢百分比最高（第3阶段?…

Discussion

该方案是为泥蟹 Scylla spp .的眼柄消融而开发的，可以作为诱导性腺成熟的有效方法。该方案可以很容易地复制用于泥蟹的商业卵巢成熟，并且可以实施以减少泥蟹种子生产中的潜伏期（从一个产卵到另一个产卵的时间）。

甲壳类动物（即淡水虾、海虾）的眼柄消融通常是为了诱导性腺成熟和反季节产卵11，12，<sup cla…

Disclosures

The authors have nothing to disclose.

Acknowledgements

这项研究得到了马来西亚教育部的支持，在马来西亚高等院校卓越中心（HICoE）计划下，获得了马来西亚登嘉楼大学热带水产养殖和渔业研究所的认可（Vot No. 63933和Vot No. 56048）。我们感谢马来西亚登嘉楼大学和Sayap Jaya私人有限公司通过私人伙伴关系研究基金（Vot.No. 55377） 提供 的支持。马来西亚理科大学到Khor Waiho和Hanafiah Fazhan的兼职学术研究员职位也得到了认可。

Materials

Aeration tube  Ming Yu Three N/A aquarium and pet shop
Airstone Ming Yu Three N/A aquarium and pet shop
Autoclave machine HIRAYAMA MANUFACTURING CORPORATION N/A MADE IN JAPAN
Bleaching powder (Hi-Chlon 70%) Nippon Soda Co.Ltd,Japan N/A N/A
Blow torch  MR D.I.Y. Group Berhad N/A N/A
Circular tank (32L) BEST PLASTIC INDUSTRY SDN. BHD.  N/A N/A
Cotton hand gloves (thick)  MR D.I.Y. Group Berhad N/A N/A
Cotton towel MR D.I.Y. Group Berhad N/A N/A
Digital thermometer Hanna Instrument HI9814 Hanna Instruments GroLine Hydroponics Waterproof pH / EC / TDS / Temp. Portable Meter HI9814
Digital Vernier Caliper INSIZE Co., Ltd. N/A
Dissecting tray Hatcheri AKUATROP  N/A Research Center of Universiti Malaysia Terengganu
Dropper bottle/Plastic Pipettes Dropper Shopee Malaysia N/A N/A
Ethanol 70% Thermo Scientific Chemicals 033361.M1 Diluted to 70% using double distilled water
Fiberglass tank (1 ton) Hatcheri AKUATROP  N/A Research Center of Universiti Malaysia Terengganu
Fine sand N/A N/A collected from Sea beach of Universiti Malaysia Terengganu
First Aid Kits Watsons Malaysia N/A N/A
Flat head nickel steel metal rod (Screw driver) MR D.I.Y. Group Berhad N/A N/A
Formaldehyde Thermo Scientific Chemicals 119690010
Gas cylinder (butane gas) for blow  torch MR D.I.Y. Group Berhad N/A N/A
Gas lighter gun (long head) MR D.I.Y. Group Berhad N/A N/A
Glass beaker (100 mL)) Corning Life Sciences 1000-100
Ice bag  Watsons Malaysia N/A N/A
Perforated plastic baskets  Eco-Shop Marketing Sdn. Bhd. N/A N/A
PVC pipe 15mm Bina Plastic Industries Sdn Bhd (HQ) N/A N/A
Refractometer ATAGO CO.,LTD.
Refrigerator Sharp Corporation Japan N/A Chest Freezer SHARP 110L – SJC 118
Scoop net MR D.I.Y. Group Berhad N/A
Seawater Hatcheri AKUATROP  N/A Research Center of Universiti Malaysia Terengganu
Siphoning pipe MR D.I.Y. Group Berhad N/A N/A
Spray bottle Mr. DIY Sdn Bhd N/A N/A
Stainless surgical forceps  N/A N/A N/A
Stainless surgical scissors  N/A N/A N/A
Submersible water pump  AS N/A model: Astro 4000
Tincture of iodine solution  (Povidone Iodine) Farmasi Fajr Sdn Bhd N/A N/A
Tissue paper  N/A N/A
Transparent plastic aquarium Ming Yu Three N/A aquarium and pet shop
Waterproof table Hatcheri AKUATROP  N/A Research Center of Universiti Malaysia Terengganu
DOWNLOAD MATERIALS LIST

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Tags

Eyestalk AblationMud CrabOvarian MaturationStress MinimizationAnesthesiaCauterizationSurgical AblationOvarian Maturation Stages

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Asmat-Ullah, M., Rozaimi, R., Fazhan, H., Shu-Chien, A. C., Wang, Y., Waiho, K. Eyestalk Ablation to Increase Ovarian Maturation in Mud Crabs. J. Vis. Exp. (193), e65039, doi:10.3791/65039 (2023).
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