Kelimpahan dan Aktivitas Bakteri Nitrifikasi, Denitrifikasi, dan Nitrat Amonifikasi di Tambak Udang Sistem Intensif

Abstract

Udang merupakan salah satu komoditas unggulan perikanan budidaya di Indonesia. Ekosistem tambak udang memiliki senyawa nitrogen anorganik utama yang dapat terlarut dalam air, yaitu amonia (NH3), nitrat (NO3-), dan nitrit (NO2-). Salah satu permasalahan yang sering dihadapi dalam pengelolaan ekosistem tambak udang yaitu penurunan kualitas air dan sedimen yang disebabkan oleh polusi senyawa nitrogen anorganik. Polusi senyawa nitrogen di lingkungan ekosistem tambak menimbulkan dampak negatif. Senyawa nitrogen anorganik seperti amonia (NH3) dan nitrit (NO2-) dapat menyebabkan keseimbangan ekologi mikroorganisme di dalam tambak terganggu. Upaya pelestarian ekosistem tambak udang terhadap hal tersebut dapat dilakukan melalui pengendalian mikroorganisme dan aktivitasnya dalam proses transformasi senyawa nitrogen. Penelitian ini dilakukan untuk mengkaji kelimpahan maupun aktivitas bakteri serta mendapatkan isolat bakteri potensial yang memiliki aktivitas nitrifikasi, denitrifikasi, dan nitrat amonifikasi dari air kolam tambak udang sistem intensif. Sampel air diambil mulai dari tahap persiapan tambak hingga tahap panen dengan interval waktu 0 hari, 21 hari, 65 hari, dan 89 hari umur udang. Pengambilan sampel dilakukan secara acak pada 4 titik area kolam tambak dengan kedalaman air 60 cm. Analisis parameter kualitas air meliputi suhu, salinitas, pH, konsentrasi oksigen terlarut (DO), nitrat (NO3-), nitrit (NO2-), amonia (NH3), total amonium (TAN), dan kandungan total bahan organik (TOM). Analisis kelimpahan bakteri nitrifikasi, denitrifikasi, dan nitrat amonifikasi dilakukan menggunakan metode MPN. Laju potensial bakteri ditentukan dengan persamaan kinetika Michaelis-Menten. Isolasi bakteri dilakukan dari medium uji MPN yang positif. Isolat bakteri murni yang diperoleh dikarakterisasi morfologi koloni dan morfologi sel dengan pewarnaan Gram. Isolat murni diuji aktivitas oksidasi amonia dan reduksi nitrat. Isolat dengan aktivitas oksidasi amonia dan reduksi nitrat tertinggi dipilih untuk dianalisis kinetika aktivitas, diamati pola pertumbuhannya, dan diidentifikasi secara molekuler berdasarkan gen 16S rRNA. Hasil penelitian menunjukkan kelimpahan tertinggi bakteri nitrifikasi pada air kolam tambak udang sistem intensif terdapat pada hari ke-65 (3,69 log sel mL-1) dan kelimpahan terendah terdapat pada hari ke-0 (2,33 log sel mL-1). Kelimpahan tertinggi bakteri denitrifikasi terdapat pada hari ke-89 (3,42 log sel mL-1) dan kelimpahan terendah terdapat pada hari ke-0 (1,82 log sel mL-1). Kelimpahan tertinggi bakteri nitrat amonifikasi terdapat pada hari ke-65 (3,08 log sel mL-1) dan kelimpahan terendah terdapat pada hari ke-0 (1,31 log sel mL-1). Laju potensial kelompok bakteri nitrifikasi dominan pada hari ke-0 hingga ke-65. Laju potensial kelompok bakteri denitrifikasi dominan pada hari ke-89. Laju potensial kelompok bakteri nitrat amonifikasi lebih rendah dibandingkan nitrifikasi dan denitrifikasi. Sebanyak 32 isolat terseleksi berhasil diperoleh dari air kolam tambak udang sistem intensif, 15 isolat merupakan kelompok bakteri nitrifikasi, 8 isolat termasuk kelompok bakteri denitrifikasi, dan 9 isolat termasuk kelompok bakteri nitrat amonifikasi. Isolat yang memiliki aktivitas oksidasi amonia dan reduksi nitrat terbaik yaitu KN25, KN45, KD12, KD19, KR5, dan KR9. Kecepatan maksimum (Vmax) oksidasi amonia isolat KN25 dan KN45 masing-masing adalah 0,07 mM jam-1 dan 0,15 mM jam-1. Nilai Km isolat KN25 dan KN45 yaitu 0,23 mM dan 0,08 mM. Kecepatan maksimum (Vmax) reduksi nitrat isolat KD12, KD19, KR5, dan KR9 masing-masing adalah 0,06 mM jam-1, 0,03 mM jam-1, 0,18 mM jam-1, dan 0,08 mM jam-1. Nilai Km masing-masing isolat tersebut yaitu 0,37 mM, 0,46 mM, 0,14 mM, dan 0,47 mM. Hasil identifikasi 16S-rRNA isolat KN25 memiliki kemiripan sebesar 99% dengan Acinetobacter radioresistens, isolat KN45 memiliki kemiripan sebesar 98% dengan Halomonas sp., isolat KD12 memiliki kemiripan sebesar 98% dengan Glutamicibacter soli, isolat KD19 memiliki kemiripan sebesar 98% dengan Microbacterium sp., isolat KR5 memiliki kemiripan sebesar 100% dengan Bacillus safensis, dan isolat KR9 memiliki kemiripan sebesar 99% dengan Shigella sonnei.

Shrimp is one of the leading aquaculture commodities in Indonesia. Shrimp ponds ecosystem have the main inorganic nitrogen compounds that can dissolve in water such as ammonia (NH3), nitrate (NO3-), dan nitrite (NO2-). One of the problems often faced in the management of shrimp pond ecosystems is a decrease in water quality and sediment caused by pollution of inorganic nitrogen compounds. Nitrogen compound pollution in the pond ecosystem has a negative impact. The toxicity of inorganic nitrogen compounds such as ammonia (NH3) and nitrite (NO2-) can disturb the ecological balance of microorganisms in the pond. Efforts to preserve the shrimp pond ecosystem in this regard can be carried out through controlling microorganisms and their activities in the process of transforming nitrogen compounds. This research was conducted to assess the abundance and activity of indigenous bacteria and to obtain potential bacterial isolates that have nitrification, denitrification, and ammonification nitrate activities from intensive system shrimp pond. Water samples were taken from the pond preparation stage to the harvest stage. Water samples were taken randomly at 4 points in the pond pond area with a depth of 60 cm. Analysis of water quality parameters including temperature, salinity, pH, dissolved oxygen concentration (DO), nitrate (NO3-), nitrite (NO2-), ammonia (NH3), total ammonium (TAN), and total organic matter content (TOM). The abundance of nitrifying, denitrifying, and ammonified nitrate bacteria was analyzed using the MPN method. The potential rate of bacteria was determined by the Michaelis-Menten kinetics equation. Bacterial isolation was carried out from the positive MPN test. The pure bacterial isolates were characterized by colony morphology and cell morphology using Gram stain. Pure isolates were tested for ammonia oxidation activity and nitrate reduction. Isolates with the highest ammonia oxidation activity and nitrate reduction were selected for analysis of activity kinetics, observed growth profile, and molecular identification of 16S rRNA gene. The results showed the highest abundance of nitrifying bacteria in intensive system shrimp pond water was found on the 65 days of cultivation (3,69 log sel mL-1) and the lowest abundance was found on the 0 days of cultivation (2,33 log sel mL-1). The highest abundance of denitrifying bacteria was found on the 89 days of cultivation (3,42 log sel mL-1) and the lowest abundance was found on the 0 days of cultivation (1,82 log sel mL-1). The highest abundance of nitrate-ammonifying bacteria was found on the 65 days of cultivation (3,08 log sel mL-1) and the lowest abundance was found on the 0 days of cultivation (1,31 log sel mL-1). The potential rate of nitrifying bacteria was dominant at 0 to 65 days of cultivation. The potential rate of denitrifying bacteria was dominant at 89 days of cultivation. The potential rate of nitrate-ammonifying bacteria was lower than nitrifying and denitrifying bacteria. A total of 32 selected isolates were obtained from the intensive system shrimp pond water, 15 isolates were from the nitrifying bacteria group, 8 isolates belonged to the denitrifying bacteria group, and 9 isolates belonged to the ammonified nitrate bacteria group. Isolate of KN25, KN45, KD12, KD19, KR5, and KR9 have high ammonia oxidation and nitrate reduction activity. The maximum velocity (Vmax) of ammonia oxidation of the KN25 and KN45 isolates was 0,07 mM hour-1 and 0,15 mM hour-1, respectively. The Km values for the KN25 and KN45 isolates were 0,23 mM and 0,08 mM. The maximum velocity (Vmax) reduction of nitrate isolates KD12, KD19, KR5, and KR9 were 0,06 mM hour-1, 0,03 mM hour-1, 0,18 mM hour-1, and 0,08 mM hour-1, respectively. The Km values for each of these isolates were 0,37 mM, 0,46 mM, 0,14 mM, and 0,47 mM. The identification results of 16S-rRNA, KN25 isolate was 99% closely related to Acinetobacter radioresistens, KN45 isolate was 98% closely related to Halomonas sp., KD12 isolate was 98% closely related to Glutamicibacter soli, KD19 isolate was 98% closely related to Microbacterium sp., KR5 isolate was 100% closely related to Bacillus safensis, and KR9 isolate was 99% closely related to with Shigella sonnei.