Probiotics for ammonium and nitrite treatment in commercial shrimp culture

30/09/2022

Currently, in the face of a serious decline in catches, the global aquaculture industry is growing strongly to meet the increasing demand for aquatic food. According to FAO statistics, the most farmed shrimp species are black tiger shrimp (Penaeus monodon), whiteleg shrimp (Litopenaeus vannamei) and upland shrimp (Penaeus chinensis), these three species account for over 86% of the world's farmed shrimp production. However, the rapid development of intensive shrimp farms in tropical countries has adversely affected the ecological environment, and the quality of the culture environment has deteriorated. Some studies have shown that to produce 1 kg of seafood, 1-2 kg of dry protein-rich feed is required, with the nitrogen component accounting for 56-88 g/kg of feed. But only 25% of this nitrogen is converted into biomass for livestock, 75% of the nitrogen component will be excess in the aquatic environment. Thus, every year according to the calculation, the total amount of nitrogen discharged into the environment from aquaculture activities exceeds 2.8 million tons. This is the main cause of nitrogen pollution, which directly affects aquatic animal health.

The main nitrogen compounds that exist in the aquatic environment are ammonium, nitrite and nitrate. They are compounds that are toxic to fish, molluscs, crustaceans and extremely toxic to shrimp. These inorganic nitrogen compounds, if left untreated, will increase exponentially during the shrimp culture period. With only a concentration of 0.425 mg/L, ammonium (NH4+) in the aquatic environment can be toxic to shrimp, fish and other aquatic animals. When the temperature and pH in the environment increase, NH4+ (ammonium) will turn into NH3 (ammonium) - which is very toxic to shrimp larvae even with a very small dose of 0.1 mg / L. If NH3 is high, the mortality rate will increase because it is easy to infect pathogenic bacteria such as Vibrio alginolicus and no longer has the ability to resist. The nitrite content is usually formed by 2 processes (the first step of nitrification and the first step of denitrification) which is also toxic to shrimp larvae, reducing growth, if the threshold is exceeded, which will reduce the transport of oxygen in the blood, leading to shrimp stress, shrimp weakness and death. Although nitrite is thought to be less toxic than ammonia, prolonged exposure is toxic. The effect of nitrite varies by species: for many shrimp species, the LC50 value after 48 hours is 12.1 mM nitrite, for the fish Macrobrachium rosenbergii it is 0.71 mM. Nitrate is formed by the oxidation of nitrite in the ponds, although it is not directly toxic to shrimp and fish because the concentration is usually lower than the permissible standard, nitrate is only toxic to shrimp and fish when the concentration is > 50 mg. /L, but high concentration will cause eutrophication.

In order to reduce nitrogenous compounds that cause pollution in the shrimp culture environment and ensure the quality of farmed shrimp, biological methods are given special attention due to their environmental friendliness and the protection of the quality and safety of aquatic food sources. The technology applied is mainly nitrification - denitrification technology with the participation of groups of autotrophic, heterotrophic bacteria and the formation of biological products to supplement the shrimp culture environment.
In recent years, in Vietnam, many research works by universities, research institutes, aquatic research centers on probiotics have been used in environmental improvement, disease prevention and treatment in many shrimp farms. But probiotics are still not stable in removing nitrogen compounds in aquaculture ponds, so there are imported products on the market, though many are not effective in Vietnam due to different weather conditions, hydrological, or breed origin, while others have effective treatment but are costly.

On that basis, Institute of Biotechnology has carried out a grassroots research project "Research on production of nitrification preparations for aquatic environment treatment", code CS15-03. Initially, a nitrification product was created for the purpose of treating nitrogen-contaminated aquaculture water and was also tested in practice in a shrimp pond in Soc Trang province, the results removed up to 99% of the nitrogen components during shrimp culture without changing water. The results of the project have been granted a utility solution number 2027 and the head of the training program has been granted a creative labor license from the Vietnam General Confederation of Labor. However, this probiotic only solves the nitrification process, to complete the product, a second process is required, which is denitrification, which includes denitrification, thus creating a preparation. complete biology, thorough removal of ammonium and nitrite.Therefore, the project "Improving the technology of producing nitrification - denitrifying products for aquatic environment treatment" at Vietnam Academy of Science and Technology was implemented.

This probiotic is formed with the participation of nitrifying and denitrifying bacteria/nitrite bacteria in the treatment of an aquaculture environment contaminated with nitrogen compounds. In particular, the nitrifying/nitriding bacteria that the research team isolated can perform under aerobic conditions, so they are completely unaffected by the oxygen supply to aquatic animals during the rearing process.

Figure 1. Nitrification capacity under anaerobic (A) and aerobic (B) conditions of selected nitrifying bacteria

The isolates of nitrifying/nitriding bacteria are capable of aerobic nitrification, of which ST26 has the strongest activity, achieving a nitrite conversion efficiency of 99% in just 2 days and no nitrite is detected on the next day. By the fourth day, the denitrification efficiency of ST20 reached 92.6%, BL5 reached 87.5%. The efficiency of nitrite removal under aerobic conditions was reduced by ½ of the time compared to anaerobic conditions (Figure 1). These three strains of nitrifying bacteria have been identified and belong to the genera Pseudomonas and Bacillus, when they combine with nitrifying bacteria of the genera Nitrosomonas and Nitrobacter (previously studied) to create a complete treatment preparation and thoroughly inorganic nitrogen compounds in commercial shrimp ponds.

When compared with the Pondplus product on the market that people often use to treat nitrogen, the nitrification-reducing nitrate product of this study has high and stable ammonium and nitrite synchronous treatment efficiency at different nitrogen concentrations from 20, 50 and 100 mg N/L, reaching 97 - 98%, while the ammonium removal capacity of Pondplus preparation was only about 60% (Figure 2).

Figure 2. Nitrogen treatment capacity of CP PondPlus (A) and CP studied (B) in the 50L system. Note: [ammonium] = [nitrite] first day of each period

The product was tested in the whiteleg shrimp pond on an area of 3000 m2 in Tra Vinh of Maya farm joint stock company during 3 shrimp farming seasons (90 days/crop) with a stocking density of 300 shrimp/m2. The results showed that the ammonium and nitrite levels in the shrimp culture environment were below the allowable threshold (according to QCVN 38:2011/BTNMT, the ammonium content was not more than 1 mgN/L). The average harvest yield was 3367 kg/crop, the mortality rate was low, and the revenue increased by more than 10% for farmers compared to shrimp farming without using research products. It is hoped that the nitrification-reducing product of this study will be one of the commercial products with broad prospects in improving the aquaculture environment in Vietnam (Figure 3).

Figure 3. Nitrogen treatment efficiency of the inoculum studied on the shrimp farming model in Tra Vinh of 3 shrimp farming seasons

The result of the TNTP project has been patented and accepted as a valid application; published 01 international paper in the list of SCI-E and was assessed as "Excellent" by the Acceptance Council on June 28, 2022 at Vietnam Academy of Science and Technology level.

Figure 4. Nitrification - denitrification preparations

Translated by Phuong Ha
Link to Vietnamese version

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