Aktinomiset Rizosfer Tanaman Kubis sebagai Agens Pengendali Hayati Penyakit Busuk Hitam (Xanthomonas campestris pv. campestris)

Abstract

Aktinomiset merupakan bakteri Gram-positif yang umum ditemukan di berbagai lingkungan tanah dan berperan sebagai pengurai bahan organik. Kemampuan bakteri ini dalam menghasilkan metabolit sekunder, seperti antibiotik dan enzim hidrolitik, berpotensi penting dalam pengendalian hayati patogen tanaman. Penelitian ini bertujuan untuk mengisolasi dan mengidentifikasi aktinomiset dari rizosfer kubis yang berpotensi mengendalikan Xanthomonas campestris pv. campestris bakteri penyebab penyakit busuk hitam kubis. Penelitian terdiri dari empat tahap utama: Isolasi, seleksi, karakterisasi dan identifikasi isolat potensial. Isolat aktinomiset yang diperoleh diuji keamanan hayati melalui uji reaksi hipersensitivitas dan uji hemolisis pada agar darah. Isolat hasil seleksi kemudian diuji antagonismenya secara in vitro menggunakan metode cross-streak, dan melalui uji in vivo pertumbuhan dan perkecambahan benih kubis. Metode analytical hierarchy process (AHP) digunakan untuk menentukan tiga isolat terbaik dan selanjutnya diuji keefektifannya terhadap X. campestris pv. campestris secara in planta pada tanaman kubis. Mekanisme induksi ketahanan dianalisis melalui aktivitas enzim peroksidase pada bibit kubis yang telah diberi perlakuan aktinomiset. Seluruh pengujian dilakukan secara in vitro dan in vivo dengan rancangan acak lengkap (RAL). Tiga isolat aktinomiset terpilih juga diidentifikasi secara molekuler menggunakan PCR berdasarkan gen 16S rRNA dengan sepasang primer universal aktinomiset 27F dan 16Sact1114R dan perunutan nukleotidanya. Penelitian ini menghasilkan 55 isolat aktinomiset dari tanaman kubis asal Garut, Cianjur dan Bandung. Sebanyak 16 isolat di antaranya bersifat non patogenik bagi tumbuhan dan mamalia. Ke-16 isolat tersebut mampu menghambat pertumbuhan X. campestris pv. campestris sebesar 3,96%-20,83%. Pertumbuhan dan perkecambahan benih kubis setelah diberi perlakuan aktinomiset menunjukkan indeks vigor sebesar 41,7%-88,3% dan daya kecambah 50,83%-93,33%., Tiga isolat terpilih paling potensial sebagai agen biokontrol terhadap X. campestris pv. campestris berdasarkan AHP adalah CPT5, CPT8 dan SNT10. Ketiga isolat tersebut menunjukkan kemampuan terbaik sebagai agen biokontrol in vivo dan telah teruji memiliki aktivitas enzim peroksidase. Berdasarkan karakterisasi morfologi dan identifikasi secara molekuler dengan PCR-sequencing isolat CPT5, CPT8 dan SNT10 diidentifikasi berturut-turut sebagai Streptomyces polychromogenes, Streptomyces vinaceus, dan Streptomyces rochei. Penelitian ini berhasil menentukan aktinomiset rizosfer tanaman kubis yang paling berpotensi dalam mengendalikan X. campestris pv. campestris.

Actinomycetes are Gram-positive bacteria commonly found in various soil environments and play a role in decomposing organic matter. Their ability to produce secondary metabolites, such as antibiotics and hydrolytic enzymes, is potentially important in the biological control of plant pathogens. The objective of this study was to isolate and identify actinomycetes from the cabbage rhizosphere that have the potential to control Xanthomonas campestris pv. campestris, pathogenic bacteria that cause black rot disease in cabbage. The research consisted of four main stages: Isolation, selection, characterization, and identification of potential actinomycete isolates. The actinomycete isolates obtained from cabbage rhizosphere were examined for biosafety through hypersensitivity reaction (HR) and hemolysis in blood agar tests. The selected isolates were then in vitro tested for antagonism using the cross-streak method on agar medium, and through in vivo growth and germination of cabbage seeds testing. Analytical hierarchy process (AHP) method was used to determine the best three isolates of actinomycetes. Their effectiveness against X. campestris pv. campestris was determined through in planta test in cabbage plants. The mechanism of resistance induction was analyzed through the activity of the peroxidase enzyme in cabbage seedlings treated with actinomycetes. All in vitro and in vivo experiments were conducted with a completely randomized design (CRD). Three selected actinomycete isolates were also identified molecularly using PCR based on the 16S rRNA gene with universal actinomycete primer pair 27F and 16Sact1114R and nucleotide sequencing. This research yielded 55 actinomycetes isolates from cabbage plants from Garut, Cianjur, and Bandung. Sixteen of these isolates were non-pathogenic to plants and mammals. All 16 isolates were able to inhibit the growth of X. campestris pv. campestris of 3,96%-20,83%. The growth and germination of cabbage seeds after being treated with actinomycetes showed a vigor index of 41,7%-88,3% and a germination level of 50,83%-93,33%. The best three isolates that had the most potential as biocontrol agents against X. campestris pv. campestris were selected based on the AHP method. All three isolates showed the highest ability as biocontrol agents and peroxidase enzyme activity. Based on morphological characterization and molecular identification by PCR-sequencing, isolates CPT5, CPT8, and SNT10 were identified respectively as Streptomyces polychromogenes, Streptomyces vinaceus, and Streptomyces rochei. This research succeeded in determining the rhizosphere actinomycetes of cabbage plants that have the most potential in controlling X. campestris pv. campestris.