Shifts in Microbial Diversity and Predictive Functions in Biofloc Aquaculture System Rearing Whiteleg Shrimp, Litopenaeus vannamei

Abstract

A microorganism-based aquaculture technique, biofloc aquaculture technology (BFT) was first devised in the early 1970s and this method has become a good alternative for conventional aquaculture methods. Also, indoor BFT systems provide producers with continuous production in all seasons without complex processes and multiple facilities. With these advantages, indoor BFT systems have been successfully commercialized in many countries including South Korea. The BFT system is relatively independent of external environments but productivity in the system could be affected by the deviation of internal factors such as temperature and water quality, which are managed by a temperature maintainer, sodium bicarbonate and carbon sources. Another important factor in the BFT system is microorganisms that are the core component in the water purification process and sustainable BFT operation. Though many studies have already evaluated various modified and novel physical or chemical elements to increase the productivity and establish stable BFT systems, the microbial diversity in the BFT system has not been extensively studied so far. In the present study, we constructed a lab-scale biofloc aquaculture system to evaluate the effects of two internal factors, temperature and carbon source (wheat flour) on not only water quality and productivity but also microbial community including its functional property in the shrimp BFT system. For the study, nine aquaculture conditions were created by ap-plying different temperature levels (25 ℃, winter; 29 ℃, optimal; 34 ℃, summer) and carbon source treatment (groups L, late treated; N, not treated; E, early treated) to a lab-scale shrimp BFT system. In the results, the water quality was maintained suitably and impaired rearing water effectively recovered by wheat flour without sodium bicarbonate. Also, wheat flour remarkably reduced nitrate which is rarely removed in aerobic BFT rearing water. The deterioration and recovery speed of rearing water and productivity were significantly different depending on temperature. Though the optimal temperature (29 ℃) showed the best productivity, water quality deteriorated faster and recovered slower at 29 ℃. For investigating microbial diversity in the shrimp BFT system, 16S rRNA gene amplicon sequencing was performed using next-generation sequencing (NGS) technique and the raw data was analyzed using statistical tools. Through the analysis, a total of 26 phyla and 340 OTUs at the genus level were identified in rearing water, shrimp stomach, and intestine. Five phyla including 13 bacterial genera were identified as the core bacterial groups that found in all samples and conditions. Correlation analysis revealed that temporal and habitational differences significantly affected bacterial diversity and composition more than temperature and wheat flour. CCA analysis estimated that ammonia and wheat flour were the most significant environmental factors affecting the bacterial communities, followed by culture period, nitrate, pH, alkalinity, and nitrite in the rearing water, while culture period and shrimp weight had larger impact than wheat flour and temperature in the shrimp digestive tract. In addition to microbial diversity, investigating microbial functions is also important to understand the metabolic structure of bacterial communities. A functional prediction pipe-line, PICRUSt was used to reveal the functional diversity in the BFT system and showed various bacterial functions related to genetic processing, environmental processing, metabolisms of amino acid, carbohydrate, cofactor and vitamin, lipid, and nucleotide. Correlation analysis estimated the distinct positive relationships of Proteobacteria and amino acid metabolism, Planctomycetes and lipid metabolism, Actinobacteria and carbohydrate metabolism, Verrucomicrobia and glycan metabolism, and Bacteroidetes and vitamin, cofactor, and nucleotide metabolisms. Finally, the environmental variables that cause the major shifts of bacterial and functional structures in the shrimp BFT system were determined. The shifts of bacterial diversity and predictive functions were significantly affected by two environmental factors, ammonia concentration in the rearing water and culture period in all habitats, and the functional composition was mainly affected by the compositional changes between Proteobacteria groups and the rest of core bacterial phyla. Overall, the present study revealed the major and core bacterial diversity in the shrimp BFT system and estimated ecological relationships between bacterial communities including their functional property and surrounding environment. This study will provide useful and fundamental information for understanding the microbial ecology in the BFT system.