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Articles by issue : Vol 2, No 1 (2008): April 2008
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Identity and Sequence Diversity of Begomovirus Associated with Yellow Leaf Curl Disease of Tomato in Indonesia

SANTOSO, TRI JOKO ( Indonesian Center for Agricultural Biotechnology and Genetic Resources Research and Development ) , HIDAYAT, SRI HENDRASTUTI ( Bogor Agricultural University ) , DURIAT, ATI SRI ( Indonesian Vegetable Research Institute ) , HERMAN, MUHAMMAD ( Indonesian Center for Agricultural Biotechnology and Genetic Resources Research and Development ) , SUDARSONO, . ( Bogor Agricultural University )

Microbiology Indonesia Vol 2, No 1 (2008): April 2008
Publisher : Indonesian Society for microbiology

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Abstract

Infection of tomato by Begomovirus is known to cause serious disease and yield losses. Samples of tomato plants showing typical symptoms of begomovirus infection were collected from eight locations in Java and Sumatra. Amplification of a putative AV1 gene was performed using AV1 specific primers for Geminivirus, total nucleic acid isolated from tomato samples exhibiting leaf curl disease as the template, and the PCR technique. Direct sequencing of PCR product was carried out, followed by nucleotide and predicted amino acid sequence analysis using the BLAST program. Positive results were obtained, the PCR amplification proved that diseased tomato samples collected from eight locations in Java and Sumatra were infected with Begomovirus. When nucleic acid and amino acid sequences of the eight isolates were compared to other begomovirus’s sequences present in the GenBank it was found that the isolates determined in this research were Indonesian isolates of AYVV. Further phylogenetic analysis of eight Begomovirus isolates identified in this study indicated they belonged into two different clades. Results of this research also suggest that the existence of Begomovirus genetic diversity in various regions in Indonesia needs further investigation. Moreover, the prevalence of distinct Begomovirus species or isolates also need investigation.

Isolation and Identification of Ice-Nucleating-Active Bacteria from Indonesian Edible Leafy Plant Poh-Pohan (Pilea glaberina)

WATURANGI, DIANA ELIZABETH ( Universitas Katolik Indonesia Atma Jaya ) , MEICY, VICKY ( Universitas Katolik Indonesia Atma Jaya ) , SUWANTO, ANTONIUS ( Institut Pertanian Bogor )

Microbiology Indonesia Vol 2, No 1 (2008): April 2008
Publisher : Indonesian Society for microbiology

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Abstract

Two ice-nucleating-active (INA) bacteria (isolates C and 6) were isolated from poh-pohan (Pilea glaberina), an Indonesian edible leafy plant (lalaban). The maximum nucleation temperature of aqueous suspensions of the two isolates is -5 °C. They were classified as a type II ice nucleator. Microscopic and morphological determination showed that these isolates had yellow pigmentation, rod shape, and were Gram negative. Biochemical analysis indicated that the isolates were exhibited catalase activity, but negative in oxidase and indole assays. DNA sequencing of 16SrRNA gene of isolate A3 showed a 94% similarity to Pseudomonas sp. while isolate A4 showed a 97% similarity to Xanthomonas campestris. To our knowledge, this is the first report of INA bacteria isolated from a tropical edible leafy plant.

The Effect of Mutation at Thr 295 of Saccharomyces cerevisiae eRF1 on Suppression of Nonsense Codons and eRF1 Structure

SUSILOWATI, PRIMA ENDANG, ADITIAWATI, PINGKAN, MADAYANTI, FIDA, AKHMALOKA, .

Microbiology Indonesia Vol 2, No 1 (2008): April 2008
Publisher : Indonesian Society for microbiology

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Abstract

The termination of translation in Saccharomyces cerevisiae is controlled by two interacting polypeptide chain release factors, eRF1, and eRF3. Two regions in eRF1, at position 281-305 and 411-415, were proposed to be involved on the interaction to eRF3. In this study we have constructed and characterized eRF1 mutants at position 295 from threonine to alanine and serine residues resulting in eRF1(T295A) and eRF1(T295S) respectively. The mutations did not affect the viability or temperature sensitivity of the cells. The stop codons readthrough of the mutants were analyzed in vivo using PGK-stop codon-LACZ gene fusion and the results showed that thesuppression of the mutants was increased in all of the codon terminations. The suppression of the UAG codon was the high for both mutants, with a 7-fold increased for eRF1(T295A) and a 9 fold increase for eRF1(T295S). The suppressor activity of eRF1(T295S) was higher compared to that of eRF1(T295A), suggesting that the accuracy of translational termination in eRF1(T295S) was lower than that of eRF1(T295A). Computer modeling analysis using Swiss-Prot and Amber version 9.0 programs revealed that the overall structure of eRF1 mutants has no significant difference with the wild type. However, substitution of threonine to serine on eRF1(T295S) triggered a secondary structure change on the other motif of the C-terminal domain of eRF1. This observation did not occur for on eRF1(T295A). This suggests that the high stop codon suppression on eRF1(T295S) is probably due to the slight modification of the structure of the C terminal motif.

Role of Bacteria in Tempe Bitter Taste Formation: Microbiological and Molecular Biological Analysis Based on 16S rRNA Gene

BARUS, TATI, SUWANTO, ANTONIUS, WAHYUDI, ARIS TRI, WIJAYA, HANNY

Microbiology Indonesia Vol 2, No 1 (2008): April 2008
Publisher : Indonesian Society for microbiology

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Abstract

Tempe is traditional Indonesian food. It has a variety of tastes, sometimes with a hint of bitterness, which may differ in intensity. The cause of bitterness in tempe has never been reported previously. In this study, the aim is to identify whether bacteria play a role in the formation of bitter tastes in tempe. Sensory tests were carried out in order to determine the scoresof bitter-taste-intensity in tempe. The sensory test on EMP, WJB, CLR, DRG, and MLB tempe shows that EMP tempe has the highest score (2.3) and WJB has the lowest (1.3). It is revealed that the processing method has no impact on the formation of the bitter taste in tempe. Plating analysis, showed that EMP soaking water contained a higher number of Enterobacteria groupbacteria, approximately 103-104 CFU ml-1 and spore-forming bacteria groups, 102 CFU ml-1, compared to WJB. Similarly, other bacteria groups in fresh EMP tempe was 102 CFU g-1 higher than those in fresh WJB tempe. Based on sequencing the16S rRNA gene, the dominant bacteria on PCA media in EMP tempe are Acetobacter indonesiensis, Klebsiella pneumoniae, Bacillus subtilis, and Flavobacterium sp. On the other hand those in WJB tempe were Klebsiella sp., Brevundimonas sp., Bacillus sp., Pseudomonas putida, and Acinetobacter sp. Bacillus, a group of proteolytic bacteria was found 105 CFU m-1 higher in the soaking water of EMP compared to WJB. Nevertheless, the types and numbers of fungi were not significantly different betweentempe types. Accordingly, it is concluded that the difference in the number and the types of bacteria involved in the tempe production process leads to the difference in the bitter taste intensity in both EMP and WJB tempe.

Growth Responses of External Hyphae of Arbuscular Mycorrhizal Fungi to Acidic Soil Conditions and their Effects on Cowpea Growth

ROHYADI, AGUS

Microbiology Indonesia Vol 2, No 1 (2008): April 2008
Publisher : Indonesian Society for microbiology

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Abstract

The effectiveness of arbuscular mycorrhizal (AM) fungi has often been attributed to growth of their external hyphae, whilst the hyphae themselves may be subjected to the effects of severe soil conditions. The growth of external hyphae of Gigaspora margarita and Glomus etunicatum and their functions in cowpea growth have been studied at low soil pH using a pot system making is possible for the hyphae to grow separately from their host’s roots. Pots had two compartments, one for roots (RC) and one for hyphae (HC). The RC was a cylindrical bag made of 30 ìm nylon mesh that retains the roots but allows the hyphae to pass through, placed centrally and surrounded by the HC. Initially, the RC was filled with 120 g of a soil/sand mixture (pH 5.3), inoculated with G. margarita, G. etunicatum or free fungal inoculants. A pre-germinated cowpea seed was grown in the compartment for two weeks before the HC was filled with 580 g of the mix in which the pH had been adjusted to 4.6, 4.9 or 5.2. Growth of the plants and of the fungal hyphae in the HC was assessed 6 weeks later. The two fungi differed in their responses to soil pH levels in their growth of external hyphae although they colonized plant roots in the same way. At pH 4.6, the hyphae of G. etunicatum grew more weakly than those of G. margarita. Increasing the pHenhanced the growth of G. etunicatum’s hyphae but reduced G. margarita’s. In relation to their external hyphal functions, G. margarita was able to improve its shoot dry weight and P uptake of cowpea plants higher than G. etunicatum. These findings highlight the ability of developing an extensive external hyphal network under adverse conditions of excessive H+ ions as an important characteristic for theeffectiveness of AM fungi in acidic soils.

Cell Lysis Method Affects Assessment of Microbial Diversity Based on Ribotyping Analysis

YOHANDINI, HENI, MADAYANTI, FIDA, ADITIAWATI, PINGKAN, AKHMALOKA, .

Microbiology Indonesia Vol 2, No 1 (2008): April 2008
Publisher : Indonesian Society for microbiology

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Abstract

The microbial community in Kawah Hujan, Kamojang, West Java, Indonesia, was analyzed using 16S-rRNA-gene-sequencing combining with denaturing-gradient-gel electrophoresis (DGGE) technique. Two different cell lysis methods, enzymatic-based, and physical treatment-based DNA extraction, were used to isolate chromosomal DNA for 16S rDNA gene-fragment amplification. The DGGE profiles showed some differences in banding pattern that were obtained from both cell lysis methods. The DNA sequence analysis of the individual DGGE bands revealed that most of the band sequences obtained by physical treatment were close to 16S rRNA gene fragments from the bacterial domain, while most of band sequences performed by enzymatic method had high homology with 16S rRNA gene fragments from archaeal domain. Further analysis of the sequences from both methods performed by comparisons with the Ribosomal Database Project showed that some of DGGE sequences from Kawah Hujan consisted unique 16S rDNA sequences.

Characterization of Extracellular Chitinase from Bacterial Isolate 99 and Enterobacter sp. G-1 from Matsue City, Japan

MAHATA, MARIA ENDO, DHARMA, ABDI, RYANTO, IRSAN, RIZAL, YOSE

Microbiology Indonesia Vol 2, No 1 (2008): April 2008
Publisher : Indonesian Society for microbiology

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Abstract

One hundred and twenty isolates of chitosanase producing bacteria were screened from water and soil from localies around Matsue city, Japan. In previous experiments, four isolates (isolates 96, 97, 99, and 100 strain ) were analyzed for their chitosanase characteristics, and one of the isolates (99) was detected as being both a chitosanase and a chitinase producer. Characteristics of the chitinase enzyme were analyzed in this study. Chitinase from bacterial isolate 99 showed higher activities compared to that Enterobacter sp. G-1 (isolated from water in Matsue city, Japan), the activity was 0.039 U/ml and the specific activity was 0.56 U/mg protein, while those from Enterobacter sp. G-1 were 0.029 U/ml and 0.48 U/mg protein respectively. Chitinase from isolate 99 was stable in a pH range between 4-7, while that from Enterobacter sp. G-1 was stable in pH range 3-7. Optimum pH of the chitinase produced by isolate 99 was 5 whereas the chitinase from Enterobacter sp. G-1 it was pH 7. Chitinase from isolate 99 was stable at temperature 20-60°C, while that from Enterobacter sp. G-1 at 20-50°C. Chitinase secreted by isolate 99 showed optimum temperature of 50°C while chitinase from Enterobacter sp. G-1 was optimal at 40°C. Several ions (Fe2+, Ba2+, Co2+) increased the activity of the enzyme from isolate 99 whereas Ca2+ and Co2+ increased activity of the Enterobacter sp. G-1 chitinase..

Vegetative Compatibility Groups within Fusarium oxysporum f. sp. cepae in Hokkaido-Japan

WIDODO, ., KONDO, NORIO, KOBAYASHI, KIROKU, OGOSHI, AKIRA

Microbiology Indonesia Vol 2, No 1 (2008): April 2008
Publisher : Indonesian Society for microbiology

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Abstract

In Hokkaido, Fusarium basal rot, caused by Fusarium oxysporum f. sp. cepae is one of the important constrains since 1973 which contributes to a significant loss in onion production, either in the fields or during storage. Development of resistant cultivars is suggested as one of the effective control measures against the disease, however, this should be accompanied with thebetter understanding of the pathogen’s population dynamics. This study was performed to investigate the population structure of F. oxysporum f. sp. cepae based on vegetative compatibility groupings (VCGs). Vegetative compatibility groups of F.oxysporum f. sp. cepae were characterized using nitrate non-utilizing (nit) mutants. Four VCGs and 2 single self-compatible (SSC) isolates were identified among 48 isolates, designated as VCG 0420 (33 isolates), 0421 (9 isolates), 0422 (2 isolates), 0423 (2 isolates), and 042-(2 isolates). VCG 0420, to which 4 ATCC isolates out of 6 belonged, was the predominant group within the growing region encompassing Hokkaido Japan. VCGs 0421 and artificial VCG 042- were found less frequently. Four isolates from Welsh onion were not compatible with any recovered VCGs and were assigned to 2 distinct VCGs (VCG 0422 and 0423).

Apparent Induction of Xylanase by Bacillus pumilus PU4-2 using Pretreated Substrates

WIDJAJA, SHERLY, PURWADARIA, TRESNAWATI, KETAREN, PIUS PERTUMPUN

Microbiology Indonesia Vol 2, No 1 (2008): April 2008
Publisher : Indonesian Society for microbiology

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Abstract

Bacillus pumilus PU4-2 produces xylanase (â-1,4-D-xylan xylanohydrolase; EC 3.2.1.8) in wheat pollard with high activity. Water and NaOH-soaked pollard were used in this research to enhance the production of assayable enzyme. Enzyme activity was produced in minimal media containing 3% w/v untreated or water or NaOH-soaked pollards in 250 ml flasks incubated on shaker incubator at 30°C and 150 rpm for 36 h. The production was also compared to untreated oatmeal known as an inducer substrate. The highest xylanase activity was obtained by using untreated pollard as a sole carbon source. The enzyme activity was 157 U ml-1 with specific activity at 718 U mg-1. Xylanase production using different soaking time for water pretreated pollard also confirmed that untreated pollard was the best inducer. The production was not influenced by different water soaking times used to remove reducing sugar. Although pretreatment decreased the reducing sugar, the reduction did not enhance assayable enzyme levels. The production was best induced by the soluble oligosaccharides of untreated pollard. We conclude that B. pumilus PU4-2 was able to produce xylanase with reducing sugar content up to 660 ppm present in production medium. With this reducing sugar level, repression of enzyme production was not detected in the production medium.