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TULISAN PENDEK Deteksi Chytridiomycosis Dengan Jurnal Biologi Indonesia 9(1): 153-157 (2013) TULISAN PENDEK Deteksi Chytridiomycosis dengan Menggunakan Koleksi Museum Zoologicum Bogoriense Pada Katak Asal Taman Nasional Gede-Pangrango, Jawa Barat (Chtridiomycosis detection by using Musem Zoologicum Bogoriense’s frog specimens that were originally collected from Gede-Pangrango National Park, West Java) Hellen Kurniati & Ni Luh Putu Rischa Phadmacanty Pusat Penelitian Biologi-LIPI, Gedung Widyasatwaloka, Jalan Raya Cibinong Km 46, Cibinong 16911, Jawa Barat. Email: [email protected] Memasukkan: September 2012, Diterima November 12 Chytridiomycosis merupakan penyakit tinggi membuat jamur ini mati, akibatnya jamur yang disebabkan oleh jamur chytrid Batracho- ini hidup subur pada daerah dataran tinggi; oleh chytrium dendrobatidis (Bd) yang menginfeksi sebab itu infeksi Chytridiomycosis biasanya terjadi jaringan kulit amfibia; pada banyak kasus terjadi pada jenis-jenis katak yang hidup di dataran ting- pada kelompok katak. Jamur ini menyerang per- gi. Pada tahun 2006 dan 2007, Kusrini dkk mukaan kulit katak yang mengakibatkan rusaknya (2008) mendapatkan beberapa jenis katak yang lapisan keratin pada kulit. Stadium lanjut dari dijumpai di Taman Nasional Gede-Pangrango infeksi chytridiomycosis yaitu jamur akan menge- sudah terinfeksi Bd (Tabel 1). Penemuan ini ber- luarkan substansi toksik yang kemudian akan tanda bahwa penyebaran jamur Bd sudah meluas mengganggu sistem respirasi katak dan berakibat sampai kepada dataran tinggi di Jawa. Untuk pada kematian katak. Kasus chytridiomycosis mengetahui kapan jamur Bd masuk ke Indonesia, ditemukan pada tahun 1998 di Amerika Tengah khususnya pulau Jawa, maka dilakukan sayatan dan Australia sehubungan dengan kematian am- histologi kulit katak yang berasal dari daerah Ci- fibi mereka (http://trushin. wordpress.com/ bodas dan sekitarnya (termasuk Taman Nasional 2011/07/28/batrachochy-trium-dendroba-tidis- Gede-Pangrango) dengan menggunakan koleksi frog-killer). Selain di Australia, jamur ini juga yang disimpan di Museum Zoologicum Bo- ditemukan sudah menyebar di Afrika, Amerika, goriense (MZB). Pengamatan dengan mengguna- Eropa, New Zealand, Oceania dan Asia dan se- kan spesimen museum terbukti dapat mendeteksi bagai penyebab kematian massal, penurunan pop- kapan jamur Bd masuk pertama kali ke dalam ulasi dan kepunahan jenis-jenis amfibi di seluruh suatu negara (Cheng dkk 2011). dunia (Bergera dkk 1998; Catenazzi dkk 2011; Dalam penelitian ini dipakai contoh kulit Krieger & Hero 2007; Kriger dkk 2007; Toledo katak yang hidup di dataran tinggi. Contoh kulit dkk 2006; Une dkk 2008). katak diambil dari koleksi MZB dari tahun 1918 Indonesia memiliki banyak jenis katak yang sampai 1955 yang berasal dari daerah dataran hidup di dataran tinggi maupun di dataran ren- tinggi Cibodas, Taman Nasional Gede- dah. Jamur Chytrid umumnya hidup di daerah Pangrango. Contoh kulit katak tahun 1918 yang dataran tinggi karena jamur ini sensitf pada panas tersedia pada koleksi MZB adalah jenis Huia ma- dan cahaya ultra violet; suhu lingkungan yang sonii, Megophrys montana dan Hylarana chalconota 153 Kurniati & Phadmacanty Tabel 1. Jenis-jenis kodok yang terinfeksi Bd di Gunung Gede-Pangrango berdasarkan Kusrini dkk (2008) dengan menggunakan tes PCR. Setara Sex/tahap SVL Berat Jumlah Tanggal Jenis Zoospora/ umur (mm) (g) sumuran koleksi contoh Telaga Biru Rhacophorus javanus Jantan 41.66 7.15 385 3 1-1- 2007 Rana chalconota Juvenil 42.60 2.50 72 3 1-1-2007 Ciwalen Leptobrachium hasseltii Berudu - - 22 3 17-1-2007 Limnonectes Juvenil 19.62 0.75 3 3 17-1-2007 microdiscus Cibeureum Leptophryne cruentata Juvenil 23.72 1.30 =1 1 31-12-2006 masing-masing satu individu; contoh kulit tahun parafin 1, kemudian parafin 2 dan terakhir par- 1930 meliputi dua individu H. chalconota, contoh afin 3 masing-masing selama satu jam, kemudian kulit tahun 1952 meliputi M. montana dan Lim- contoh kulit ditanam dalam parafin beku dengan nonectes microdiscus masing-masing satu individu, orientasi untuk potongan melintang. Parafin contoh kulit tahun 1954 meliputi Polypedates leu- beku yang berisi contoh kulit dipotong dengan comystax dan Rhacophorus margaritifer masing- ketebalan 5 μm dengan menggunakan mikrotom; masing satu individu, dan contoh kulit tahun hasil potongan lalu derekatkan pada gelas 1955 meliputi R. margaritifer dan M. montana preparat; kemudian potongan contoh tersebut masing-masing satu individu. diwarnai dengan hematoxilin-eosin. Proses Bagian kulit katak yang dipakai sebagai pewarnaan diawali dengan proses deparafinasi, contoh adalah bagian abdomen, sebab bagian ini yaitu merendam potongan contoh kulit didalam merupakan bagian yang paling mudah terinfeksi larutan xilene selama 1 jam; setelah itu dilanjut- oleh jamur chytrid, karena langsung bersentuhan kan dengan proses hidrasi dengan alkohol dengan substrat habitat katak. Bagian kulit abdo- bertingkat yaitu dari alkohol absolut 96% hingga men disayat dengan ukuran 1x1 cm, kemudian alkohol 70%, kemudian dilanjutkan dengan pe- proses pembuatan preparat histologi sampai pada rendaman di dalam aquades selama satu menit, pewarnaan jaringan dilakukan dengan memodifi- kemudian dicelupkan dalam larutan hematoxilin kasi metode yang dilakukan Kiernan (1990). Per- selama 30 detik, lalu contoh kulit direndam da- tama kali sayatan kulit difiksasi didalam larutan lam air mengalir selama minimal 10 menit atau alkohol 70%, setelah itu dilakukan pencucian setelah contoh kulit terlihat berwarna keunguan. dengan menggunakan alkohol 70% lalu didehid- Proses selanjutnya contoh kulit dicelupkan dalam rasi dengan menggunakan alkohol bertingkat yai- aquades dan alkohol 70%, setelah itu dilanjutkan tu alkohol 80%-absolut masing-masing selama 12 dengan proses pewarnaan menggunakan larutan jam. Setelah proses dehidrasi, contoh kulit diren- eosin dengan cara dicelup selama 10 detik, dam dalam larutan xilene selama 24 jam, setelah kemudian contoh kulit dicuci dalam alkohol itu dilakukan proses infiltrasi dalam oven pada 70%, dan dilakukan didehidrasi dalam alkohol suhu 560C, lalu direndam dalam larutan campu- bertingkat hingga alkohol absolut 96%. Perla- ran xilene-parafin dengan perbandingan 1:1, kuan pembersihan preparat dengan meren- setelah itu contoh kulit direndam dalam larutan damnya dalam larutan xilene selama minimal 15 154 Deteksi Chytridiomycosis dengan Menggunakan Koleksi Museum Zoologicum menit. Setelah itu dilakukan proses mounting hasil sayatan histologi kulit perut tersebut terlihat pada preparat, ditutup dengan kaca penutup, dan spora jamur ini yang berbentuk pundi (Gambar diletakkan di atas hot plate hingga kering. 2). Prepararat kemudian diamati di bawah mikroskop Jenis kodok Rana catesbeiana adalah penye- dengan perbesaran 400 kali. Preparat diamati bar utama jamur Bd di dunia (Garner et al. 2006); untuk menentukan struktur kulit yang normal perdagangan kodok secara internasional juga dan terinfeksi oleh jamur chytrid. Adanya infeksi memberi andil meluasnya penyebaran jamur ini dapat diketahui melalui kerusakan pada bagian (Fisher & Garner 2007). Penyebaran Bd di Jawa stratum korneum epidermis kulit katak (Gambar Barat kemungkinan besar dibawa oleh kodok R. 1). catesbeiana (Kusrini dkk 2008) yang masuk tahun Hasil sayatan histologi pada enam jenis 1982 sebagai program pemerintah Indonesia un- katak asal dataran tingi Cibodas membuktikan tuk meningkatkan ekspor daging kodok. Bila bahwa jamur B. dendrobatidis sudah menginfeksi melihat hasil histologi dari spesimen MZB, jenis katak jenis R. margaritifer pada koleksi MZB yang R. margaritifer yang dikoleksi tahun 1954 telah diambil tahun 1954 dan 1955 (Tabel 2); dari terinfeksi jamur Bd; bukti ini mengindikasikan Gambar 2. Spora Batrachochytrium dendrobatidis (Bd) yang berbentuk pundi (lingkaran dan panah Gambar 1. Epidermis kulit katak yang terinfeksi merah) pada kulit perut katak R. margaritifer jamur chytrid Batrachochytrium dendrobatidis yang dikoleksi dari Cibodas, Taman Nasional (Bd). Sel berbentuk pundi adalah spora jamur Gunung Gede-Pangrango pada tahun 1954 Bd (Pessier & Mendelson 2010). (Amph. 351). (kiri pembesaran 100 kali dan kanan pembesaran 400 kali. Tabel 2. Jenis-jenis katak dari koleksi MZB asal Cibodas, Taman Nasional Gede-Pangrango, Jawa barat yang di- ambil sayatan histologi kulit abdomen. (+) terinfeksi Bd; (-) tidak terinfeksi Bd. Nomor koleksi Tahun Jenis katak Lokasi Positif infeksi Bd (+) MZB koleksi Amph.94 Huia masonii Cibodas, Gunung Gede Pangrango 1918 - Amph.230 Megophrys montana Cibodas, Gunung Gede Pangrango 1918 - Amph.689 Hylarana chalconota Cibodas, Gunung Gede Pangrango 1918 - Amph.680 Hylarana chalconota Cibodas, Gunung Gede Pangrango 1930 - Amph.684 Hylarana chalconota Cibodas, Gunung Gede Pangrango 1930 - Amph.315 Megophrys montana Cibodas, Gunung Gede Pangrango 1952 - Amph.305 Limnonectes microdiscus Cibodas, Gunung Gede Pangrango 1952 - Amph.331 Polypedates leucomystax Cibodas, Gunung Gede Pangrango 1954 - Amph.333 Rhacophorus margaritifer Cibodas, Gunung Gede Pangrango 1954 + Amph.351 Rhacophorus margaritifer Cibodas, Gunung Gede Pangrango 1955 + Amph.346 Megophrys montana Cibodas, Gunung Gede Pangrango 1955 - 155 Kurniati & Phadmacanty jamur Bd telah menyebar di Jawa Barat sebelum Manu National Park, Southeastern Peru. tahun 1982. Berdasarkan Kusrini dkk (2008), Conservation Biology 25(2): 382–391. prefalensi infeksi jamur Bd pada kodok R. mar- Cheng, TL., SM. Rovitob, DB. Wakeb & VT. garitifer paling tinggi dari pada empat jenis Vredenburga. 2011. Coincident mass extir- lainnya, tetapi hasil sayatan histologi dari koleksi pation
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