ABSTRAKInfeksi white spot syndrome virus (WSSV) dapat menyebabkan kematian massal pada budidaya udang windu Penaeus monodon di Indonesia. Infeksi yang terjadi secara sistematis tersebut disebabkan oleh peran gen nucleocapsid viral protein . Upaya pengembangan gen VP-15 WSSV untuk menginduksi respons imun dan menetralisasi terhadap infeksi WSSV pada udang windu perlu dilakukan. Penelitian ini bertujuan untuk mengisolasi dan merekombinasikan gen penyandi VP-15 WSSV sebagai vaksin dsRNA, serta menganalisis aplikasinya pada udang windu. Gen VP-15 diisolasi dari udang windu yang terinfeksi WSSV, dikloning ke dalam suatu vektor dan ditransformasikan ke sel kompeten (bakteri Escheria coli DH5). Plasmid diisolasi untuk mengonfirmasi insert region gen VP-15 melalui sekuensing nukleotida. Pembuatan vaksin rekombinan dilakukan secara in-vitro menggunakan kit MEGAscript RNAi dan diaplikasikan ke udang windu melalui metode injeksi dengan dosis tunggal 0,2 µg dan kontrol (tanpa injeksi vaksin). Hewan uji yang digunakan berukuran panjang 14,75±3,17 g dan bobot 11,64±0,76 cm; serta dipelihara pada wadah bak fiber volume 250 L dengan kepadatan 10 ekor/bak. Hasil penelitian menunjukkan bahwa gen penyandi VP-15 telah diisolasi dari udang windu dan vaksin rekombinan telah dihasilkan secara in-vitro. Analisis sekuens nukleotida memperlihatkan bahwa sisipan gen DNA VP-15 sebesar 253 bp dan menunjukkan kemiripan yang tinggi (99%) pada GenBank. Penggunaan vaksin rekombinan dsRNA dengan dosis 0,2 µg memperlihatkan sintasan udang windu yang dapat mencapai 40,0% dibandingkan dengan kontrol hanya 3,3% (peningkatan 36,7%). Gambaran histopatologi pada jaringan hepatopankreas udang windu pada perlakuan kontrol menunjukkan adanya kerusakan inti sel, akibat infeksi WSSV. Gene VP-15 berpotensi sebagai bahan vaksin rekombinan dsRNA dalam mencegah infeksi WSSV.
Teknologi RNA interference (RNAi) merupakan salah satu pendekatan yang digunakan untuk meningkatkan resistensi udang windu terhadap infeksi patogen termasuk WSSV. Pengembangan teknologi RNAi melalui aplikasi untai ganda RNA (dsRNA) yang berasal dari gen pengkode viral protein (VP) dari WSSV telah mulai dikembangkan pada udang. Penelitian ini bertujuan untuk mengkaji sintasan dan respons imun udang windu yang diberi VP-15 pasca uji tantang dengan WSSV. Udang windu (panjang 15,21 ± 1,19 cm dan bobot 32,5 ± 1,83 g) diinjeksi dengan 0,2 µg/ekor dsRNA in vitro (A), dsRNA in vivo (B), dan larutan garam/kontrol (C). Setelah tiga hari vaksinasi, udang windu ditantang dengan WSSV dengan dosis 50 µL/ekor. Pengamatan sintasan dilakukan setiap hari, sedangkan respons imun (THC dan aktivitas proPO) dilakukan pada awal dan hari ke-1, ke-3, dan ke-5 pasca uji tantang, serta analisis ekspresi gen antivirus dan histopatologi hepatopankreas dilakukan pada akhir penelitian. Hasil penelitian menunjukkan bahwa aplikasi dsRNA berpengaruh nyata (P<0,05) terhadap sintasan, THC, dan proPO. Sintasan udang windu yang diberi dsRNA VP-15 in vitro dan in vivo memberikan sintasan yang lebih tinggi 75% dibandingkan dengan kontrol. Nilai proPO tertinggi didapatkan pada dsRNA in vivo (0,138); kemudian dsRNA in vitro (0,093); dan terendah kontrol (0,061); sedangkan THC tertinggi (5.704 x 104 sel/mL) pada dsRNA in vivo, kemudian dsRNA in vitro (3.516 x 104 sel/mL) dan terendah pada perlakuan kontrol (3.322 x 104 sel/mL). Ekspresi gen antivirus semakin meningkat dengan semakin lamanya udang windu terpapar dengan WSSV. Jaringan hepatopankreas udang windu pada perlakuan kontrol (tanpa dsRNA) menunjukkan adanya kerusakan sel akibat infeksi virus.RNA interference (RNAi) technology is one of the approaches used to improve tiger shrimp Penaeus monodon resistance against WSSV infection. The development of RNAi technology through double-stranded RNA (dsRNA) isolated from gene encoding viral protein (VP) of WSSV has been applied to shrimp. This study was aimed to assess the survival rate and immune response of injected-VP-15 WSSV tiger shrimp after a challenge with WSSV. The tiger shrimp (15.21 ± 1.19 cm in length and 32.5 ± 1.83 g in weight) were injected with 0.02 µg/shrimp of in vitro dsRNA (A), in vivo dsRNA (b) and saline solution (C). After three days of vaccination, the tiger shrimp were challenged with WSSV using a dosage of 50 µL/shrimp. The survival rate was observed daily. Analyses of immune responses (hemocyte total and PO activity) were performed in several stages: before the challenge test and day-1, day-3, and day-5 post-challenge test. The expression of the antivirus gene and hepatopancreas histophatology were was observed at the end of the experiment. The results showed that the application of dsRNA significantly influenced the shrimp survival rate, THC, and proPO. Tiger shrimp injected with dsRNA VP-15 of in vitro and in vivo exhibited a higher 75% survival rate than the control (P<0.05). The highest proPO activity (0.138) was obtained at dsRNA in vivo, followed by dsRNA in vitro (0.093) and the lowest (0.061) in the control. The highest THC (5,704 x 104 cell/mL) was in vivo dsRNA, then in vitro dsRNA (3,516 x 104 cell/mL), and the lowest in the control (3,322 x 104 cell/mL). The longer the exposure with WSSV, the higher the antivirus gene expression. Histopathology analysis showed some damages to the hepatopancreas cells in the control shrimp (without dsRNA) caused by the virus infection.
ABSTRAKMasalah utama pada budidaya udang intensif adalah menurunnya kualitas air di tambak yang layak selama pemeliharaan dan munculnya penyakit. Upaya mengurangi permasalahan tersebut adalah pemanfaatan bioflok di tambak. Bioflok merupakan campuran dari berbagai mikroba (fitoplankton, zooplankton, protozoa), detritus, dan partikel organik. Teknologi bioflok dapat meningkatkan kualitas air, meminimalkan pergantian air, efisiensi pakan, dan menghambat berkembangnya penyakit selama budidaya. Penelitian ini bertujuan untuk mengetahui pengaruh bioflok terhadap produksi udang vaname intensif. Penelitian dilakukan pada tambak beton ukuran 2.000 m 2 milik masyarakat di Desa Hanura Kecamatan Pasawaran, Lampung. Padat penebaran udang vaname adalah 100 ekor/m 2 . Perlakuan yang dicoba adalah (A) budidaya udang vaname intensif sistem bioflok dan (B) budidaya udang vaname intensif tanpa bioflok. Hasil penelitian menunjukkan bahwa produksi tertinggi diperoleh pada perlakuan bioflok yaitu 10.375 kg/ha dengan bobot udang rata-rata 13,8 g/ekor, sintasan 75%, dan RKP 1,3. Sedangkan tanpa bioflok memperoleh produksi 9.176 kg/ha dengan bobot udang rata-rata 12,0 g/ekor, sintasan 76%, dan RKP 1,6.
The positive effect of organic mineral as a dietary mineral source in aquafeed has been recently reported on several species. Nevertheless, the influence of organic minerals on rabbitfish has not yet been determined. The purpose of the present study was to evaluate the effects of different doses of organic mineral on growth and survival, and mineral content in vertebrae of golden rabbitfish, Siganus guttatus. Three diets were formulated containing 0.5%, 1.0%, and 2.0% organic material (OM). A control diet (OM0) did not contain OM. Instead, it was supplemented with an inorganic mineral mixture at a level of 1%. Three hundred fish were randomly selected and distributed in 12 cages to accommodate the four treatments with triplicates. The stocking density was 20 fish per cage. The initial body weight of fish used was 39.2 ± 0.3 g. Fish were fed the test diets twice a day for 150 days. A significant (P<0.05) cubic effect of the treatments was detected on all dependent parameters analysis, excluding feed intake. The influence of dietary OM was not significant for feed intake, indicating that dietary OM did not negatively affect the appetite of rabbitfish. Mineral (Ca, Mg, Zn) content in the vertebrae was significantly improved when dietary OM was included in the diet up to 1% but decreased at the highest inclusion level of 2%. The optimum level of dietary OM to gain the maximum growth rate of rabbitfish was 0.49% as the reflection of the breakpoint of two regressions fitted on specific growth rate (SGR). It is concluded that dietary OM level significantly affected the growth and vertebral mineral content of golden rabbitfish. The study increases our knowledge of the benefit of utilizing dietary OM in the fish diet.
RNA interference (RNAi) is one of the most recent tools against shrimp viral infection by gene constructs of dsRNA induction. This study aimed to produce dsRNA by in-vivo method and to analyse the effect of its application to tiger shrimp larvae. The dsRNA production was conducted by cloning of genes encoding VP15 and VP24 of WSSV into L4440 vector containing T7 promoter, then transformed to Escherichia coli and grew in LB media for mass production. The bacteria were in-activated using heat-killed bacteria method by immersion at 80°C for 5, 10, and 15 min. Before WSSV-challenge test, tiger shrimp PL-12 were vaccinated by immersion using in-activated bacteria in concentration of 1.3×108 cell/mL for 30 min as a treatment and without vaccination was as a control. The results showed the successful production of VP-dsRNA by in-vivo. In-activated bacteria was effective at 5 min, since dsRNA did not damage. Survival of larvae at 5 days post challenge (dpc) in VP15-dsRNA was relatively higher (28.3%) compared to control (24.4%), and application of VP24-dsRNA also showed higher survival (86.9%) compared to control (83.1%) at 10 dpc. The result implied that the application of dsRNA exhibited a sign of potential way to produce resistant larvae.
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