Kesimpulan Saran Co infection of Infectious Myonecrosis Virus (IMNV) and Vibrio harveyi in Pacific White Shrimp (Litopenaeus vannamei)

V. KESIMPULAN DAN SARAN

5.1 Kesimpulan

Ko-infeksi IMNV dengan berbagai dosis Vibrio harveyi 10 6 , 10 7 dan 10 8 cfuml dapat mempercepat awal mortalitas dan meningkatkan mortalitas dibandingkan dengan infeksi tunggal IMNV. Infeksi V. harveyi 10 7 cfuml tidak menyebabkan mortalitas, dan infeksi tunggal IMNV menyebabkan mortalitas diawali pada hari ke-10 pasca infeksi dengan kumulatif 13.33. Sedangkan ko- infeksi virus IMNV dan V. harveyi 10 7 cfuml menyebabkan awal mortalitas udang lebih cepat hari ke-4 pasca infeksi dengan kumulatif lebih tinggi yaitu 40. Ko-infeksi juga mempercepat pertumbuhan Vibrio koloni berwarna hijau berpendar di tubuh udang, dengan densitas mencapai 108x10 5 ± 27,06x10 5 cfuml pada hari ke-10 pasca infeksi. Tidak ada perbedaan perkembangan gejala klinis visual maupun histopatologi antara infeksi tunggal IMNV dan ko-infeksi IMNV dengan V. harveyi . Infeksi IMNV menunjukkan beberapa gejala yang nampak yaitu nekrosis lebam putih, abnormalitas organ limfoid, terlihat badan inklusi pada sel otot, dan konfirmasi PCR di tubuh udang.

5.2 Saran

Perlu dilakukan penelitian lanjutan untuk mengetahui dampak penanggulangan penyakit IMN melalui pengendalian bakteri Vibrio, misalnya penggunaan bakteri sebagai biokontrol Vibrio. Perlu juga dilakukan kuantifikasi IMNV di tubuh udang menggunakan Real Time PCR. DAFTAR PUSTAKA Alapide-Tendencia EV, Dureza LA. 1997. Isolation of Vibrio spp. from Penaeus monodon Fabricius with red disease syndrome. Aquaculture 154:107– 114. Andrade TPD, Srisuvan T, Tang KFJ, Lightner DV. 2007. Real-time reverse transcription polymerase chain reaction assay using TaqMan probe for detection and quantification of Infectious myonecrosis virus IMNV. Aquaculture 264:9–15. Andrade TPD, Redman RM, Lightner DV. 2008. Evaluation of the preservation of shrimp samples with Davidson’s AFA fixative for infectious myonecrosis virus IMNV in situ hybridization. Aquaculture 278:179– 183. Chrisolite B, Thiyagarajan S, Alavandi SV, Abhilash EC, Kalaimani N, Vijayan KK, Santiago TC. 2008. 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Carcinicultura ameaçada: produtores sofrem com as mortalidades decorrentes do vírus da mionecrose infecciosa IMNV. Pan. Aquic. 14:37–51. OIE Office International Epizootics. 2009. Manual of Diagnostic Tests for Aquatic Animals: Chapter 2.2.3. Infectious Myonecrosis. 96-104p. Phuoc LH, Corteel M, Nauwynck H J, Pensaert MB, Alday-Sanz V, Broeck van den W, Sorgeloos P, Bossier P. 2008. Increased susceptibility of white spot syndrome virus-infected Litopenaeus vannamei to Vibrio campbellii. Environ. Microbiol. 10:2718–2727. Phuoc LH, Corteel M, Nguyen CT, Nauwynck H, Pensaert M, Alday-Sanz V, Broeck van den W, Sorgeloos P, Bossier P. 2009. Effect of dose and challenge routes of Vibrio spp. on co-infection with white spot syndrome virus in Penaeus vannamei. Aquaculture 290:61-68. Pinheiro ACAS, Lima APS, de Souza ME, Neto ECL, Adrião M, Gonçalves VSP, Coimbra MRM. 2007. Epidemiological status of Taura syndrome and Infectious myonecrosis viruses in Penaeus vannamei reared in Pernambuco Brazil. Aquaculture 262:17–22. Poulos BT, Tang KFJ, Pantoja CR, Bonami JR, Lightner DV. 2006. Purification and characterization of infectious myonecrosis virus of penaeid shrimp. J. Gen. Vir. 87:987-996. Rodriguez SAS, Gomez-Gil B, Lozano R. 2010. Density of Vibrios in Hemolymph and Hepatopancreas of Diseased Pacific White Shrimp, Litopenaeus vannamei , from Northwestern Mexico. Journal Of The World Aquaculture Society 41:76-83. Roza D, Zafran, Taufik I, Girsang MA.1997. Pengendalian Vibrio harveyi secara biologis pada larva udang windu Penaeus monodon: Isolasi bakteri penghambat. Jurnal Penelitian Perikanan Indonesia 3:1-10. Rukyani A. 1993. Penanggulangan penyakit udang windu Penaeus monodon. Pusat Penelitian Pengembangan Perikanan, Jakarta. Saulnier D, Haffner P, Goarant C, Levy P, Ansquer D. 2000. Experimental infection models for shrimp vibriosis studies: a review. Aquaculture 191: 133–144. Senapin S, Phewsaiya K, Briggs M, Flegel TW. 2007. Outbreaks of infectious myonecrosis virus IMNV in Indonesia confirmed by genome sequencing and use of an alternative RT-PCR detection method. Aquaculture 266:32-38. Tan Y, Xing Y, Zhang H, Feng Y, Zhou Y, Shi ZL. 2009. Molecular detection of three shrimp viruses and genetic variation of white spot syndrome virus in Hainan Province, China, in 2007. J. Fish. Dis. 32:777-784. Tang J, Ochoa WF, Sinkovits RS, Poulos BT, Ghabrial SA, Lightner DV, Baker TS, Nibert ML. 2008. Infectious myonecrosis virus has a totivirus-like, 120-subunit capsid, but with fiber complexes at the fivefold axes. PNAS 45105:17527-17531. Tang KFJ, Pantoja CR, Poulos BT, Redman RM, Lightner DV. 2005. In Situ Hybridization demonstrates that Lithopenaeus vannamei, L. stylirostris and Penaeus monodon are susceptible to experimental infection with infectious myonecrosis virus IMNV. Dis. Aquat. Org. 63:261-265. Tsai JM, Shiau LJ, Lee HH, Chan PWY, Lin CY. 2002. Simultaneous detection of white spot syndrome virus WSSV and Taura syndrome virus TSV by multiplex reverse transcription-polymerase chain reaction RT-PCR in Pacific white shrimp Penaeus vannamei. Dis. Aquat. Org. 50:9–12. Van de Braak CBT, Botterblom MHA, Taverne N, van Muiswinkel WB, Rombout JHWM, van der Knaap WPW. 2002. The roles of haemocytes and the lymphoid organ in the clearance of injected Vibrio bacteria in Penaeus monodon shrimp. J. Fish. Shell. Immunol. 13:293–309. Vandenberghe J, Li Y, Verdonk L, Li J, Sorgeloos P, Xuh S, Swings J. 1998. Vibrios associated with Penaeus chinensis Crustacea: Decapoda larvae in Chinese shrimp hatcheries. Aquaculture 169:121–132. Walker PJ, Winton JR. 2010. Emerging viral disease of fish and shrimp. Vet. Res. 4151:1-24. Wurmann C, Madrid RM, Brugger AM. 2004. Shrimp farming in Latin America: currents status, opportunities, challenges and strategies for sustainable development. Aquac. Econ. Manag. 8:117–141. Yeh SP, Chen YN, Hsieh SL, Cheng W, Liu CH. 2009. Immune response of white shrimp, Litopenaeus vannamei, after a concurrent infection with white spot syndrome virus and infectious hypodermal and hematopoietic necrosis virus. J. Fish. Shell. Immunol. 26:582-588. Zhang XH, Austin B. 2000. Pathogenicity of Vibrio harveyi to salmonid. J. Fish. Dis. 23: 93-102. Lampiran 1. Pembuatan Media Bakteri SWC dan TCBS.

1. Sea Water Complete SWC Cair.

Media SWC pada penelitian ini digunakan untuk kultivasi Vibrio harveyi yang akan digunakan untuk perlakuan infeksi. Media SWC cair sebanyak 1000 ml dibuat dari bahan-bahan bacto peptone 5 g, yeast extract 1 g, glycerol 3 ml, air laut 750 ml dan akuades 250 ml. Media SWC 1000 ml tersebut dilakukan dengan mencampurkan semua bahan sesuai takaran di atas. Pencampuran dilakukan di labu erlenmeyer yang juga langsung digunakan sebagai wadah kultivasi. Selanjutnya campuran bahan- bahan dipanaskan di penangas air pada suhu 100 o C sampai larut sempurna. Setelah larut, media selanjutnya disterilkan menggunakan autoklaf pada suhu 121 o C dan tekanan 1 atm selama 15 menit.

2. Thiosulphate Citrate Bile Salt Sucrose TCBS Agar.

Media agar TCBS digunakan untuk menumbuhkan bakteri Vibrio karena media ini bersifat spesifik untuk menumbuhkan bakteri Vibrio. Dalam penelitian ini, TCBS digunakan untuk pengukuran atau penghitungan Vibrio pasca perlakuan infeksi. Untuk pembuatan 1000 ml media ini dilakukan dengan melarutkan bubuk media agar TCBS sebanyak 98 gram dalam 1000 ml akuades steril. Larutan tersebut dipanaskan di penangas air pada suhu 100 o C hingga larut sempurna. Lampiran 2. Prosedur Pembuatan Preparat Histologi Jaringan. Pada penelitian ini dilakukan pembuatan preparat histologi untuk jaringan otot dan organ limfoid udang uji. Dalam pembuatan preparat histologi jaringan dilakukan tahap sebagai berikut:

1. Preparasi Jaringan.

Preparasi jaringan terdiri dari 5 tahap, yaitu: 1. Dehidrasi 2. Clearing 3. Impregnasi 4. Embedding 5. Blocking Proses jaringan ini bertujuan agar jaringan yang umumnya lunak nantinya dapat dipotong setebal 3-5 µm, diwarnai dan dilihat bentuk selnya di bawah mikroskop. Bila sayatan jaringan lebih tebal, maka bentuk sel dari jaringan tersebut tidak jelas karena terjadi penumpukan atau saling tindih menindih satu sel dengan yang lainnya. Untuk mendapatkan sayatan yang tipis, maka jaringan perlu “diikat” dalam parafin. Agar “ikatan” dalam parafin tidak mudah lepas, maka perlu dilakukan beberapa tahapan proses pembuatannya.

a. Proses Dehidrasi

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