Penyakit Mosaik pada Koro Pedang

Recently, a mosaic disease was observed on jack bean cultivation in Dramaga, Bogor,West Java. The symptom on infected plants, i.e. combination of pale and dark green area was more obvious on young leaves. Vein banding, leaf malformation, and growth inhibition was also observed in infected plants. The symptom was not very obvious in older leaves. Serological detection gave positive reaction to Bean common mosaic virus (BCMV) antiserum, but reaction was negative when detected using specific primer to coat protein (CP) of BCMV strain black eye by reverse transcription polymerase chain reaction. DNA fragments were successfully amplified by RT-PCR using universal primer of CP gene and degenerated primer for cylindrical inclusion gen (CI) of Potyvirus. DNA analysis showed the highest homology (86.4–91.5%) to several isolates of BCMV-infecting different crops from several countries based on partial sequence of CP, whereas based on partial sequence of CI the highest homology (87.3%) was to BCMV-RU1-P. This is the first report of BCMV infection on jack bean in Bogor, West Java, Indonesia

Infeksi Bean common mosaic virus pada Umur Tanaman Kacang Panjang yang Berbeda

Bean common mosaic virus (BCMV) is one of the most important virus infecting yard long bean because it can decrease yield and seed transmitted. The aims of research were to determine the effect of plant age infected by BCMV on seed transmission efficiency of the virus, as well as its effect on plant growth. BCMV was mechanically inoculated on yard long bean at 1, 2, 3, and 4 weeks after planting (WAP). Observation was conducted for incubation period, disease incidence and severity, seed transmission efficiency of the virus, plant height, and productivity of the plants. Virus infection was detected using indirect ELISA method. The results showed that the earlier infection of BCMV occurred the shortest incubation period, the most severe symptoms, the highest inhibition of plant growth, and productivity. Disease severity was 94.6, 83.8, 81.1, and 69.6% on plants inoculated at 1, 2, 3, and 4 WAP, respectively. Disease incidence and virus titer was not affected by infection on different plant age. Seed transmission of BCMV was 7, 66, 39, and 24% on plants inoculated at 1, 2, 3, and 4 WAP, respectively. Infection on 2 WAP was considered the critical times for BCMV to be seed-borne on yard long bean

Lima Ekstrak Tumbuhan untuk Menekan Infeksi Bean common mosaic virus pada Tanaman Kacang Panjang

Bean common mosaic virus (BCMV) is one of major virus infecting legumes and is difficult to manage. Utilization of plant extracts as systemic resistance inducer against virus is needed to study. The aim of the research is to evaluate the potency of five leaf extracts, i.e. from pagoda flower, spiny amaranth, four o’clock flower, Chenopodium amaranticolor, and herba andrographitis against BCMV. The effectiveness of leaf extracts were tested by spraying yard long bean leaves. Plants treated by spine spinach shown varied symptoms, while other treatments showed mild mosaic up to symptomless. The highest to lowest of disease incidence was showed by crude leaf extract of spine spinach (70%), four o’clock (10%), herba andrographitis (10%), while C. amaranticolor and pagoda are still uninfected. These results had positive correlation to disease severity and virus inhibition. Four of five tested leaf extracts, except spine spinach, showed their potency as systemic resistance inducer against BCMV.  Key words: BCMV, plant extract, yard long bea

Expression of Recombinant Sugarcane Streak Mosaic Virus Coat Protein Gene in Escherichia coli

AbstractSugarcane streak mosaic virus (SCSMV) is an important virus causing mosaic disease in sugarcane and transmitted through the cutting cane. Commercial antiserum to detect SCSMV and to monitor the disease development is not available. The research was conducted to produce antigen of SCSMV coat protein (SCSMV-CP) through overexpressing it on bacterial expression which will be used for antiserum production. SCSMV-CP was amplified using specific primers for CP gene containing BamHI and HindIII restriction enzyme sites and cloned into pTZ57R/T. Subsequently, the SCSMV-CP was subcloned into pET28a and transformed on Escherichia coli BL21(DE3) and Rosetta-gami(DE3)pLysS. The concentration of isopropyl β-d-thiogalactopyranoside (IPTG), incubation temperature, and bacterial harvesting time after IPTG induction were optimized. SCSMV-CP gene was successfully amplified with size ±855 bp, subcloned into vector expression, and expressed in insoluble fraction either in both bacterial host. Optimal protein expression of SCSMV-CP recombinant was obtained at 25°C with IPTG concentration 0.25–1.00 mM and harvested at 9–12 hours after IPTG induction in E. coli BL21(DE3), and at 30°C with IPTG concentration 0.25–1.00 mM and harvested 3–12 hours after IPTG induction in E. coli Rosetta(DE3)pLysS. SDS-PAGE analysis showed that protein size of SCSMV-CP recombinant was ±35.4 kDa

Utilization of Root-Colonizing Bacteria to Protect Hot-Pepper Against Tobacco Mosaic Tobamovirus

Tobacco Mosaic Tobamovirus (TMV) is one of many important viruses infecting Solanaceous plants including hot pepper in Indonesia. To accomplish and improve the effectiveness of virus management, we used root-colonizing bacteria (rhizobacteria) isolated from healthy hot pepper. Eight rhizobacteria isolates were selected and their capacity in enhancing plant growth and inducing systemic resistance (ISR) against TMV in greenhouse trials were evaluated. The rhizobacteria was applied as seed treatment and soil drench. Bacterized-seedling showed a better growth vigor, fitness and a milder symptom than non-bacterized control plants. The protective effect of rhizobacteria was more pronounced after challenging inoculation by TMV, especially for plants treated by isolates I-6, I-16, and I-35. However, TMV accumulation was slightly affected by bacterial treatment. The rhizobacteria might improved ISR by increasing peroxidase enzyme activity but this depends on the species. Based on whole results, isolate I-35 was the potential plant growth promotion rhizobacteria (PGPR). The I-35 was identified as Bacillus cereus based on morphological characteristics and nucleotide sequences of 16S r-RNA. Key words: root-colonizing bacteria, TMV, IS

Characterization of acyl-homoserine lactonase gene from Brevibacillus brevis strain B37

17-25Acyl-homoserine lactonase (EC 3.1.1.25) is a metallo-betalactamase, specifically hydrolyzed N-acyl-homoserine lactones (AHL) secreted by Gram-negative bacteria. AHL lactonase has been reported as a potential substitute for synthetic anti-bacterial, such as reduce the severity of plant diseases caused by Xanthomonas campestris pv. campestris, and Pectobacterium catrotovorum. The exploration of lactonase producing organisms has been widely reported. AHL-lactonase is produced by Bacillaceae bacteria such as Bacillus thuringiensis, B. cereus, and B. antrachis. AHL-lactonase produced by Bacillaceae bacteria was translated from aiiA gene. In our previous study, aiiA novel gene was detected in Brevibacillus brevis B37 but has not been characterized. This study aimed to clone aiiA gene isolated from B.brevis B37 by polymerase chain reaction (PCR) with a pair of degenerated primers, to reveal homology comparison withothers aiiA genes and amino acids, to express aiiA gene in Escherichia coli BL21 (DE3), and also to assay quorum quencherability. The aiiA gene was successfully isolated with 753 bp and 250 amino acids. The aiiA gene and the AiiA protein fromB.brevis B37 had high similarity with aiiA and AiiA from B. thuringiensis group. The deduced amino acid sequencecontained conserved sequence region 103SHLHFDH109 and 166TPGHTPGH173 as characteristic of the metallo betalactamasefamily. Additionally, the aiiAB37 gene was expressed in E. coli BL21 (DE3) and the expressed AiiA protein could attenuate the expression of violacein produce by Chromobacterium violaceum and decrease the expression of soft rot symptom caused by Dickeya dadantii

Deteksi Virus Pada Tanaman Mentimun Di Jawa Barat

Gejala infeksi virus banyak ditemukan di pertanaman mentimun di Jawa Barat. Namun, sulit membedakan virus penyebab hanya berdasarkan pengamatan gejala visual. Penelitian ini bertujuan untuk pemutakhiran data virus yang menginfeksi mentimun di Kabupaten Bogor, Karawang dan Subang, Jawa Barat. Sampel tanaman bergejala dikoleksi dari tiap lokasi. Frekuensi virus ditentukan secara serologi dengan metode DIBA (dot immunobinding assay) menggunakan antiserum Cucumber mosaic virus (CMV), Cucurbit aphid borne yellow virus (CABYV), Cucurbit green mottle mosaic virus (CGMMV), Papaya ringspot virus (PRSV), Squash mosaic virus (SqMV), Watermelon mosaic virus (WMV), dan Zucchini yellow mosaic virus (ZYMV). Deteksi asam nukleat dilakukan terhadap virus baru yang dominan dengan reverse transcription polymerase chain reaction (RT-PCR)/PCR. Hasil deteksi menunjukkan bahwa frekuensi virus CMV, CABYV, CGMMV, PRSV, SqMV, WMV, dan ZYMV berturut-turut di Bogor mencapai 98%, 100%, 86%, 0%, 8%, 32%, dan 6%, di Subang mencapai 86%, 100%, 88%, 2%, 8%, 60%, and 46%, dan di Karawang 100%, 100%, 98%, 14%, 72%, 68%, and 64%. CMV, CABYV, and CGMMV were the most common viruses detected in three regencies, while PRSV only detected on samples from Subang and Karawang. RT-PCR/PCR  menggunakan primer spesifik atau universal belum berhasil mendeteksi CGMMV dan WMV, namun berhasil mengonfirmasi keberadaan PRSV, Polerovirus, dan Begomovirus.  Kata kunci: Cucurbitaceae, Geminiviridae, Luteoviridae, Potyviridae, Tobamoviru

Preparasi RNA Virus Mosaik Bergaris dari Tanaman Tebu Menggunakan Metode Tabung PCR

Sugarcane streak mosaic virus (SCSMV) is a new emerging virus infecting sugarcane in Indonesia. The virus can be found almost in every sugarcane plantations in Java and it was known as sett-borne virus. Attempt to get virus-free seedling meet difficulties due to lacking an easy, and accurate detection method. SCSMV antiserum is not available yet commercially. Nucleic acid detection by RT-PCR also hampered difficulties in releasing RNA virus from sugarcane tissues. Here we reported the simple direct tube method with minor modification as the convenient way to provide total RNA template from infected sugarcane leaf, stalk and sheath for RT-PCR detection of SCSMV.Key words: RNA extraction, RT-PCR, SCSMV, tube PC

Identifikasi Begomovirus yang Berasosiasi dengan Penyakit Kuning pada Mentimun di Jawa Barat dan Bali

ABSTRACTA survey conducted from several cucumber cultivation area of West Java and Bali found some plants showing yellow mosaic, vein banding, and stunting symptoms, caused by Begomovirus infection. This study aimed to detect and determine incidence of Begomovirus on cucumber plants, and analyze variation of Begomovirus coat protein gene in West Java dan Bali. Leaf samples from 50 plants were taken randomly from each location in Sumedang, Karawang, Sukabumi (West Java) and Tabanan, Gianyar, Klungkung (Bali). Disease incidence was determined based on serological assay using specific antiserum of Tomato leaf curl New Delhi virus (ToLCNDV) dan Squash leaf curl virus (SLCV). Incidence of ToLCNDV and SLCV were 28-100% and 30-80%, respectively. PCR using Begomovirus degenerate primers successfully amplified coat protein gene about ± 550bp. There were three Begomovirus species associated with yellowing disease on cucumber plant i.e Squash leaf curl China virus (SLCCNV), ToLCNDV, and Ageratum yellow vein virus (AYVV). Based on nucleotide sequences analysis, it was found that isolate SLCCNV had highest nucleotide homology with SLCCNV isolate Malaysia (EF197940) about 94.5%, and was considered as a strain “China”, whereas ToLCNDV has highest nucleotide similarity with ToLCNDV isolate Indonesia (AB613825) about 99.4% and was considered as a strain “Indonesia”. The AYVV sequences showed highest nucleotide AYVV isolate Nicotiana benthamiana from Indonesia (AB100305) about 92.1%.Keywords: Cucumis sativus, Dot immunobinding assay, Squash leaf curl China virus, Tomato leaf curl New Delhi virusABSTRAKSurvei yang dilakukan di beberapa pertanaman mentimun di Jawa Barat dan Bali menemukan gejala mosaik kuning, daun keriting, penebalan tulang daun, dan kerdil akibat infeksi Begomovirus. Penelitian ini bertujuan untuk mendeteksi dan menghitung insidensi penyakit yang disebabkan oleh Begomovirus pada tanaman mentimun, serta menganalisis keragaman gen protein selubung Begomovirus di Jawa Barat dan Bali. Sampel daun diambil secara acak sebanyak 50 tanaman dari tiap lokasi pertanaman mentimun di Sumedang, Karawang, dan Sukabumi (Jawa Barat), Tabanan, Gianyar, dan Klungkung (Bali). Deteksi Begomovirus dan insidensi penyakit ditentukan dengan uji serologi DIBA menggunakan antiserum Tomato leaf curl New Delhi virus (ToLCNDV) dan Squash leaf curl virus (SLCV). Insidensi ToLCNDV dan SLCV berturut-turut berkisar 28-80% dan 30-80%. Deteksi dengan teknik PCR menggunakan primer universal Begomovirus berhasil mengamplifikasi gen protein selubung berukuran ± 550 pb. Hasil perunutan nukleotida menunjukkan terdapat tiga spesies Begomovirus yang menginfeksi tanaman mentimun di Jawa Barat dan Bali, yaitu Squash leaf curl China virus (SLCCNV), ToLCNDV, dan Ageratum yellow vein virus (AYVV). Isolat SLCCNV Bali memiliki kesamaan nukleotida dan asam amino terhadap isolat SLCCNV dari negara lainnya berkisar antara 89.8-94.5% dan 94.2-96.3%, dan dikelompokkan ke dalam strain “Cina”. ToLCNDV isolat Jawa Barat dan Bali memiliki kesamaan nukleotida dan asam amino berkisar antara 92.8-99.4% dan 97.3-99.4% dengan isolat ToLCNDV dari negara lainnya, tergolong ke dalam strain “Indonesia”. Gen protein selubung AYVV Bali memiliki kesamaan nukleotida dan asam amino berkisar antara 89.5-92.1% dan 94.7-95.2%, dengan kesamaan tertinggi dengan isolat AYVV asal Indonesia yang menginfeksi Nicotiana benthamiana.Kata kunci: Cucumis sativus, Dot immunobinding assay, Squash leaf curl China virus, Tomato leaf curl New Delhi virus

Isolasi, Seleksi, dan Identifikasi Bakteri Endofit sebagai Agens Penginduksi Ketahanan Padi terhadap Hawar Daun Bakteri

Bacterial leaf blight caused by Xanthomonas oryzae pv. oryzae is one of the major problems in rice production in Indonesia. One of the control measures for the disease is by the utilization of endophytic bacteria. This study was aimed to determine the ability of endophytic bacteria isolated from the roots, stems and leaves of rice in inducing plant resistance to bacterial leaf blight on rice. Screening of endophytic bacteria showed that 370 isolates have good viability and have different colony morphology. Among them, 7 isolates were able to induce resistance and 1 isolate was able to promote the growth of rice in the nursery. However, only 8 isolates did not cause  hypersensitive reaction on tobacco plants. The selected isolates of endophytic bacteria were further examined for their ability to induce resistance of rice in greenhouse experiments.  Observation involved several variables, i.e. PR1 and PBZ1 gene expression, peroxidase enzyme activity, incubation period, and disease progression. Seven isolates of endophytic bacteria were able to induce PR1 and PBZ1 gene expression, 4 isolates were able to increase peroxide enzyme activity, 4 isolates could prolong the incubation period, and 2 isolates can inhibit the development of disease. However, only EB4 451 isolate was consistently able to induce PR1 and PBZ1 gene expression, increased peroxide enzyme activity, prolonged incubation period, and suppressed the progression of the disease. The EB4 451 isolate was identified based on nucleotide sequence as Bacillus subtilis