Vital Role of Glutamate Dehydrogenase Gene in Ammonia Detoxification and the Association Between its SNPs and Ammonia Tolerance in Sinonovacula constricta.

Gaigai Sun,Hanhan Yao,Zhihua Lin,Changsen Sun,Yinghui Dong

doi:10.3389/fphys.2021.664804

Gaigai Sun, Hanhan Yao + Show 3 more

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https://doi.org/10.3389/fphys.2021.664804

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Abstract

Increasing evidence has revealed accumulated ammonia will cause adverse effects on the growth, reproduction, and survival of aquatic animals. As a marine benthic mollusk, the razor clam Sinonovacula constricta shows better growth and survival under high ammonia nitrogen environment. However, little is known about its adaptation mechanisms to high ammonia stress in an integrated mariculture system. In this study, we analyzed the association between the polymorphism of glutamate dehydrogenase gene (GDH), a key gene involved in ammonia nitrogen detoxification, and ammonia tolerance. The results showed that 26 and 22 single-nucleotide polymorphisms (SNPs) of GDH in S. constricta (denoted as Sc-GDH) were identified from two geographical populations, respectively. Among them, two SNPs (c.323T > C and c.620C > T) exhibited a significant and strong association with ammonia tolerance, suggesting that Sc-GDH gene could serve as a potential genetic marker for molecular marker–assisted selection to increase survival rate and production of S. constricta. To observe the histological morphology and explore the histocellular localization of Sc-GDH, by paraffin section and hematoxylin–eosin staining, the gills were divided into gill filament (contains columnar and flattened cells) and gill cilia, whereas hepatopancreas was made up of individual hepatocytes. The results of immunohistochemistry indicated that the columnar cells of gill filaments and the endothelial cells of hepatocytes were the major sites for Sc-GDH secretion. Under ammonia stress (180 mg/L), the expression levels of Sc-GDH were extremely significantly downregulated at 24, 48, 72, and 96 h (P < 0.01) after RNA interference. Thus, we can speculate that Sc-GDH gene may play an important role in the defense process against ammonia stress. Overall, these findings laid a foundation for further research on the adaptive mechanisms to ammonia–nitrogen tolerance for S. constricta.

Highlights

Some improper management methods such as highdensity stocking and overfeeding lead to gradual ammonia nitrogen accumulation in rearing water, which are extremely toxic to the health of aquatic organisms
In the ammonia challenge experiment, the first dead clam was observed in Zhejiang province (ZJ) population at 24 h after ammonia stress, and the mortality rate was steadily increased and reached its apex level at 88–90 h
Researches on the GDH gene in aquatic animals mainly focus on fish and crustaceans, such as Oncorhynchus mykiss (Wright et al, 2007), Periophthalmodon schlosseri, Boleophthalmus boddaerti (Ip et al, 2005), Clarias gariepinus (Wee et al, 2007), L. vannamei (Qiu et al, 2018), and Macrobrachium rosenbergii (Dong et al, 2020)

Summary

Introduction

Some improper management methods such as highdensity stocking and overfeeding lead to gradual ammonia nitrogen accumulation in rearing water, which are extremely toxic to the health of aquatic organisms. A number of studies have demonstrated that ammonia exposure resulted in histopathologic changes in gill and liver of Oreochromis niloticus (Benli et al, 2008), Cyprinus carpio (Peyghan and Takamy, 2002), and Portunus pelagicus (Romano and Zeng, 2006). The gills of sliver catfish exposed to ammonia levels above LC50 (96 h) resulted in edema and fusion of the secondary lamellae (Miron et al, 2008). A similar pattern was observed in Ruditapes philippinarum showing that ammonia can destroy the stability of lysosomal membrane and cause cell apoptosis of the gills (Cong et al, 2017). The changes of protein expression in the Na+/K+-ATPase and Rhesus glycoprotein C like of sea lamprey (Petromyzon marinus) gill during metamorphosis suggested that the methods of excreting ammonia were fundamentally changed (Sunga et al, 2020). To date, little research focuses on the action site of genes related to detoxification metabolism of benthic mollusks exposed to high ammonia

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