Infeksi Fusarium oxysporum jamur penyebab penyakit rebah semai pada Acacia mangium dan pengendaliannya menggunakan Trichoderma harzianum

TASIK, Susanti, Prof. Dr. Ir. S.M. Widyastuti, M.Sc

2010 | Tesis | S2 Bioteknologi

Fusarium oxysporum adalah salah satu jamur patogen tular tanah yang penting dan penyebab penyakit rebah semai. Jamur ini menyerang semai pada banyak spesies tanaman termasuk Acacia mangium. Karena cepat tumbuh dan kayunya sesuai untuk bahan baku pulp dan kertas, A. mangium ditanam secara luas di Hutan Tanaman Industri (HTI) di Sumatra dan Kalimantan. Informasi mengenai bagaimana F. oxysporum menginfeksi semai dan respon semai terhadap infeksi jamur patogen sangat diperlukan. Selain itu, mekanisme penghambatan agen pengendali hayati Trichoderma harzianum sebagai salah satu teknik pengendalian yang ramah lingkungan perlu dikaji lebih lanjut. Penelitian ini bertujuan untuk mengetahui: (1) proses infeksi F. oxysporum pada semai A. mangium; (2) respon ketahanan semai A. mangium akibat infeksi F. oxysporum, dan (3) mekanisme penghambatan T. harzianum terhadap F. oxysporum. Jamur patogen diidentifikasi dan selanjutnya diuji patogenisitasnya. Proses infeksi diamati secara makroskopis (berdasarkan gejala dan tanda) dan mikroskopis (berdasarkan perkembangan patogen pada semai). Respon ketahanan semai A. mangium diuji menggunakan pewarna histologis phloroglucinol, aniline blue dan lactophenol trypan-blue. Trichoderma harzianum yang mengekspresikan Green Fluorescent Protein (GFP) digunakan untuk mempelajari efek penghambatan agen pengendali hayati terhadap F. oxysporum. Uji penghambatan secara in vitro oleh T. harzianum dilakukan dengan menggunakan metode dual culture. Pada uji penghambatan secara in planta, semai A. mangium diinokulasi dengan T. harzianum GFP 2 hari sebelum inokulasi F. oxysporum; inokulasi T. harzianum secara bersamaan diinokulasi dengan F. oxysporum; dan T. harzianum GFP yang diinokulasi 2 hari sesudah inokulasi F. oxysporum. Hasil penelitian menunjukkan bahwa secara in planta perkecambahan spora jamur teramati pada 2 hari setelah inokulasi. Pada 4 hari setelah inokulasi, hifa F. oxysporum melakukan penetrasi pada pangkal batang semai A. mangium melalui stomata. Setelah terjadi penetrasi, hifa jamur berkembang melalui ruang antar sel hingga mencapai jaringan pengangkutan. Respon ketahanan berupa reaksi hipersensitif terdeteksi pada stomata. Meskipun demikian, reaksi hipersensitif ini tidak efektif untuk menghentikan infeksi F. oxysporum yang merupakan patogen nekrotropik. Pada penelitian ini juga terdeteksi akumulasi lignin, sedangkan akumlasi kalose tidak ditemukan selama pengamatan. Efek penghambatan dari T. harzianum GFP terlihat pada 7 hari inkubasi dengan adanya penempelan hifa T. harzianum GFP pada hifa F. oxysporum. Selanjutnya hifa T.harzianum GFP menutupi sebagian besar koloni F. oxysporum pada 12 hari inkubasi. Persentase hidup semai A. mangium tertinggi ditunjukkan pada inokulasi T. harzianum GFP 2 hari sebelum inokulasi F. oxysporum (82,22%), sebaliknya persentase terendah ditunjukkan dengan inokulasi T. harzianum GFP 2 hari sesudah inokulasi F. oxysporum (64,44%).

Fusarium oxysporum is one of the most important soil-borne pathogenic fungi and the causal agent of damping-off disease. The fungus attacks seedlings of many plant species, including Acacia mangium. Due to its fast growing property, A. mangium is widely cultivated in many industrial forest plantations in Sumatra and Borneo. In order to effectively control the disease, detailed information on how the fungus infects seedlings of A. mangium and how the plant responds to the fungal infection is needed. In addition, the use of biocontrol agent Trichoderma harzianum should be considered. The objectives of this research were to investigate: (1) the infection process of F. oxysporum on seedlings of A. mangium, (2) the defence response of A. mangium seedling to infection by F. oxysporum and (3) the inhibition mechanism of T. harzianum on F. oxysporum. The fungal pathogen was identified, followed by performance of pathogenicity test. The infection process was followed by macroscopically observation (based on the development of signs and symptoms) as well as microscopically (based on the fungal development on the plant). Host defense responses were examined using histological dyes phloroglucinol, aniline blue and lactophenol trypan-blue. Trichoderma harzianum which constitutively expresses Green Fluorescent Protein (GFP) was used to study the inhibition effect of the biocontrol agent on F. oxysporum. An in vitro inhibition test of T. harzianum was performed using dual culture method. In the in planta inhibition tests, seedlings of A. mangium were inoculated with T. harzianum GFP two days before F. oxysporum; T. harzianum GFP was simultaneously inoculated with F. oxysporum and T. harzianum GFP was inoculated two days after inoculation with F. oxysporum. The result indicated that fungal spore germination was observed at two days post inoculation in planta. At four days post inoculation, hyphae of F. oxysporum had penetrated the stem of A. mangium seedling via stomatal aperture. In addition, fungal hyphae had grown intercellulary into the vascular tissue. Hypersensitive response at was detected at the stomatal aperture. However, this defence mechanism is not effective in stopping the fungus since F. oxysporum is a necrotropic pathogen. In addition, accumulation of lignin, but not callose, was observed. Inhibition effect of T. harzianum GFP was observed at seven days incubation, indicated by attachment of hyphae of T. harzianum to F. oxysporum hyphae. Subsequently, hyphae of T. harzianum GFP covered the majority of F. oxysporum colonies at 12 days incubation. The highest healthy A. mangium seedlings were found on the inoculation of T. harzianum GFP two days before inoculation of F. oxysporum (66.67%), whereas the least healthy A. mangium seedlings were found with the inoculation of T. harzianum GFP two days after inoculation of F.oxysporum (22,67%).

Kata Kunci : Fusarium oxysporum,Penyakit rebah semai,Acacia mangium,Trichoderma harzianum,Green Fluorescent Protein