ANALISIS RESISTENSI UDANG WINDU (Penaeus monodon Fabricius, 1798) HASIL PERSILANGAN SELEKTIF TERHADAP White spot syndrome virus BERDASARKAN EKSPRESI GEN PmVRP15

AUSHIA TANZIH AL HAQ, Dr. Rarastoeti Pratiwi, M.Sc.; Dr. Ir. Murwantoko, M.Si.

2015 | Tesis | S2 Biologi

White spot syndrome virus (WSSV) dapat menyebabkan infeksi berdampak serius dengan tingkat kematian dan morbiditas yang tinggi pada udang. Infeksi WSSV pada udang penaeid mempengaruhi sistem kekebalan tubuh dan mengubah tingkat ekspresi protein, termasuk ekspresi gen PmVRP15 yang berfungsi sebagai media propagasi WSSV. Ekspresi PmVRP15 pada hemosit tersebut mengalami peningkatan pada fase akut infeksi WSSV. Penelitian ini bertujuan untuk menganalisis tingkat ekspresi gen PmVRP15 pada populasi udang windu (Penaeus monodon Fabricius) hasil persilangan selektif untuk dikaitkan dengan resistensinya terhadap WSSV. Hasil persilangan populasi dibatasi pada populasi hasil inbreeding yang disebut G8 dan populasi hasil outbreeding yang disebut G7-Aceh yang terdapat di tambak National Shrimp Broodstock Center (NSBC) Balai Besar Perikanan Budidaya Air Payau (BBPBAP) Jepara. Dua populasi tersebut diuji tantang dengan WSSV secara injeksi intramuskular pada konsentrasi inokulum 10-2. Hemolimfe diambil dalam rentang waktu 48 jam setelah infeksi untuk isolasi hemosit. Pengamatan kematian kumulatif dilakukan dengan interval 6 jam. Konfirmasi infeksi WSSV dilakukan dengan nested PCR. Kuantifikasi ekspresi PmVRP15 pada hemosit antar populasi udang windu dilakukan melalui qPCR dengan normalisasi menggunakan β-actin. Median lethal time (LT50) G8 lebih rendah secara signifikan daripada G7-Aceh; mengindikasikan bahwa G8 kurang resisten terhadap infeksi WSSV dibandingkan G7-Aceh. Metode comparative CT melalui qPCR juga membuktikan bahwa tingkat ekspresi relatif PmVRP15 pada G8 lebih tinggi daripada G7-Aceh. Hasil penelitian menunjukkan bahwa populasi G7-Aceh lebih resisten terhadap WSSV daripada populasi G8. Tingkat ekspresi PmVRP15 terbukti dapat digunakan sebagai parameter untuk menyeleksi populasi antara G8 dan G7-Aceh yang lebih tahan terhadap penyakit.

　　　White spot syndrome virus (WSSV), the most serious viral disease, causes high mortality and morbidity in shrimps. Infection of WSSV into penaeid shrimps affects their immune defense and changes their gene expression levels. PmVRP15 has been reported as a part of WSSV propagation pathway which is highly up-regulated in hemocytes at acute phase of WSSV infection. This study was aimed to analyze the expression level of PmVRP15 gene among selective breeding populations of giant tiger shrimp (Penaeus monodon Fabricius) to be associated with its resistance to WSSV. The selective breeding populations which were collected from National Shrimp Broodstock Center (NSBC) ponds of Main Center for Brackishwater Aquaculture Development in Jepara consisted of inbreeding population called G8 and outbreeding population called G7-Aceh. Those selective breeding populations were challenged using intramuscularly injection at 10-2 WSSV extract concentration. Hemolymph was sampled at 48 h post-infection (hpi) and separated from hemocytes. Cumulative mortality was observed at 6 hours interval. The hemocytes were then used as samples to measure the expression levels of PmVRP15 through relative quantification using qPCR which was normalized with β-actin. Confirmation of WSSV infection was conducted using nested PCR. Median lethal time (LT50) of G8 was significantly lower than G7-Aceh, indicating that G8 was less resistant to WSSV than G7-Aceh. Comparative threshold cycle (CT) methods of qPCR also showed that PmVRP15 transcripts level of G8 was higher than G7-Aceh. The result of this study indicated that G7-Aceh was more WSSV-resistant than G8. PmVRP15 expression level could be also used as parameter for disease resistance in G8 and G7-Aceh broodstock selection.

Kata Kunci : PmVRP15, resistensi, Penaeus monodon, WSSV, real time PCR/ PmVRP15, resistance, Penaeus monodon, WSSV, real time PCR