Purification of Biotransformation Products of Cis-Isoflavan-4-Ol by Biphenyl Dioxygenase of Pseudomonas pseudoalcaligenes Kf707 Strain Expressed in Escherichia coli

Isoflavone has multiple beneficial effects on human health, especially as antioxidant and anticancer. Biotransformation of two enantiomers (CE1 and CE2) of cis-isoflavan-4-ol by E. coli JM109 (pJHF108) carrying a biphenyl dioxygenase gene from P. pseudoalcaligenes KF707 produced two products and they were designated as CM1 and CM2. They had retention time at 11.9 and 14.6 min, respectively, and same absorption peaks at 204, 220 and 275 nm. CM1 and CM2 had [M-H2O+H]+  at m/z 225. Based on the molecular mass, hydrolysis products, and previous report, this study proposed that epoxidation occurred on cis-isoflavan-4-ol. Chloroform extraction was done to improve the stability of CM1 and CM2

Isolation and Characterization of Nitrogen Fixing Endophytic Bacteria Associated with Sweet Sorghum (Sorghum bicolor)

Sweet sorghum (Sorghum bicolor) has high adaptability to the dryland and resistant to drought. Many endophytic bacteria live in association with their host and play crucial role in health and development of plant. The aims of this work were to isolate and characterize nitrogen-fixing endophytic bacteria associated with sorghum. Sorghum samples were collected from Cibinong Science Center, Cibinong, Bogor, Indonesia. Sorghum werecultivated in four different treatment (compost and inoculants (CI), inoculants (TI), compost without addition inoculants (CN), without compost and inoculants (TN). The samples were sterilized by surface sterilization method and isolation of endophytic bacteria was carried out using spread plate method on TSA medium 1/10. Characterization of endophytic bacteria to fix nitrogen were done using Jensen’s media and tested by specific primers to amplify nifH gene. A total of 89 isolates were successfully isolated from root and stem. Sixteen isolates were able to grow on Jensen’s media. Based on PCR amplification of nifH gene was observed the expected size of the nifH region

IDENTIFICATION OF ENDOPHYTIC BACTERIA FROM Curcuma zedoaria BASED ON PROTEIN PROFILE USING MALDI-TOF MASS SPECTROMETRY

Valid identification of microorganisms is a vital information to establish culture collections. Currently, molecular approach based on 16S rDNA is widely used for bacterial identification. This approach  is however, time consuming and expensive. Matrix assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF MS) allows the identification of microorganisms directly from colonies and it only takes some few minutes. The interest of this study was to identify endophytic bacteria associated with Curcuma zedoaria based on protein profile using MALDI-TOF MS system and compare with 16S rDNA sequence results. Endophytic bacteria were isolated from part of medicinal plant C. zedoaria collected from Bogor, West Java Indonesia. The identification of selected bacteria was performed by protein profile using MALDI-TOF MS. A total of 66 endophytic bacteria from C. zedoaria plant, were selected for identification. The result of MALDI-TOF MS analysis showed that eleven isolates (16.67%) were correctly identified to the species level and 23 isolates (34.85%) matched on genus level of molecular approach. These results demonstrates that the MALDI-TOF system is suitable and feasible approach for the bacterial identification, mainly for screening and grouping of large number isolates

Keragaman Bakteri Endofit Penghasil L-Asparaginase Bebas L-Glutaminase

Endophytes are potential as a source of active compound producer. Endophytes that is isolated from tropical medicinal plants has been getting attention due to its high biodiversity and active compound producing ability. L-asparaginase is the first enzyme used as chemotherapeutic agent for leukemia. The aim of this study is to collect the endophytic bacteria associated with tropical medicinal plants from Sumba Island and investigate the activity of L-asparaginase without L-glutaminase from endophytic bacteria isolates. The samples Gliricidia sepium, Pittosporum moluccanum, Clerodendrum buchanani, and Zingiberaceae are collected from Wanggameti, East Sumba, NTT. Samples are sterilized using surface-sterilization method and endophytic bacteria are isolated using plant piece method on R2A media. Selected endophytic bacteria are identified by 16S rDNA sequences. L-asparaginase screening is conducted using modified R2A with addition of L-asparagine and phenol red as colour indicator. A total of 34 isolates of endophytic bacteria were collected from 5 samples. A total of 14 genus consisted of 17 different bacterial species were obtained from 34 selected isolates. Endophytic bacteria of P. stutzeri strains of SMKL1 and R. radiobacter strains of SMKW2 from the Kahili plant were needed as L-glutaminase-free L-asparaginase and were potential to be candidate of leukemia cancer chemotherap

BAKTERI ENDOFIT YANG DIISOLASI DARI AKAR Eurycoma longifolia DAN POTENSINYA SEBAGAI PENGENDALI JAMUR PATOGEN TANAMAN

Eurycoma longifolia (pasak bumi) is known as a medicinal plant that contains biologically active compounds. Studies on endophytic bacteria associated with pasak bumi and their biocontrol activities have not been widely reported. The objective of this study is to isolate potential endophytic bacteria associated with E. longifolia possessing biocontrol activity against plant pathogenic fungi, Sclerotium rolfsii, Fusarium solani, F. oxysporum, and Colletotrichum gloeosporioides. Endophytic bacteria were isolated from the roots of E. longifolia using the plant piece method and identified based on 16S rRNA genes analysis. Antagonist test of bacterial isolates was conducted by dual confrontation method. The mechanisms of fungal growth inhibition were evaluated based on their ability to produce hydrolytic enzymes, antibiotic, and volatile organic compounds. Two isolates were obtained and identified as Stenotrophomonas maltophilia (Apb1) and Serratia marcescens (Apb2). Apb2 was able to inhibit the growth of four tested fungi and showed protease, chitinase as well as cellulase activities. The crude extract and volatile organic compound produced by Apb2 were active against F. solani growth.Keywords: biocontrol, endophytic, Eurycoma longifolia, fungi, inhibition mechanism ABSTRAKEurycoma longifolia (pasak bumi) dikenal sebagai tanaman obat yang mengandung beberapa senyawa aktif secara biologis. Penelitian mengenai bakteri endofit yang berasosiasi dengan tanaman pasak bumi berikut aktivitas biokontrolnya belum banyak dilaporkan. Penelitian ini bertujuan mengisolasi bakteri endofit potensial dari tanaman E. longifolia yang memiliki aktivitas biokontrol terhadap empat strain uji jamur patogen tanaman, yaitu Sclerotium rolfsii, Fusarium solani, F. oxysporum, dan Colletotrichum gloeosporioides. Bakteri endofit diisolasi dari akar E. longifola menggunakan metode plant piece dan diidentifikasi berdasarkan analisis gen 16S rRNA. Uji antagonis isolat bakteri dilakukan dengan metode konfrontasi ganda. Mekanisme penghambatan jamur patogen tanaman dievaluasi berdasarkan kemampuannya dalam memproduksi enzim hidrolisis, senyawa antibiotik, dan senyawa organik volatil. Dua isolat bakteri endofit berhasil diperoleh dan teridentifikasi sebagai Stenotrophomonas maltophilia  (Apb1) dan Serratia marcescens (Apb2). Isolat Apb2 mampu menghambat pertumbuhan keempat jamur yang diuji dan menunjukkan aktivitas protease, kitinase dan selulase. Ekstrak kasar dan senyawa organik volatil yang dihasilkan oleh isolat Apb2 aktif menghambat pertumbuhan F. solani

DIVERSITY OF ENDOPHYTIC BACTERIA ASSOCIATED WITH (Curcuma heyneana) AND THEIR POTENCY FOR NITROGEN FIXATION

Endophytic bacteria shows a high biodiversity and some of the species plays important biological roles in agriculture. The aim of this study was to investigate the endophytic bacteria diversity associated with temu giring (Curcuma heyneana) and to evaluate its nitrogen-fixation activity. Temu giring was collected from Bogor Botanic Garden. The isolation of  endophytic bacteria was carried out using two methods (spread plate and plant piece methods) and four different medias (Nutrient Agar (NA), NA contained temu giring extract (NAH), Water Yeast Extract Agar (WYEA), and WYEA contained  temu  giring extract (WYEAH)). The identification of selected isolates were conducted based on 16S rDNA. The ability of selected isolates to fix the nitrogen on Jensen’s media is then being tested. The results revealed that the suitable method and media of endophytic bacteria isolation were spread plate method and NA. Based  on  the  morphological characteristics differentiation, 30 isolates were obtained from rhizome (27%), stem (50%), and leaves (23%). The sequencing result of 16S rDNA showed that community of endophytic bacteria was divided into six clusters, those are Alphaproteobacteria, Betaproteobacteria, Gammaproteobacteria, Flavobacteriia, Actinobacteria, and Firmicutes,which represented 17 genus consisted of Microbacterium, Leclercia, Brevundimonas, Chromobacterium, Enterobacter, Acinetobacter, Novosphingobium, Chryseobacterium, Curtobacterium, Agrobacterium, Sphingomonas, Herbaspirillium, Bacillus, Variovorax, Mycobacterium,  Starkeya, and Rhizobium. A total of eleven isolates could grow in the N free medium. The presence of endophytic bacteria those were able to fix  nitrogen are expected to be applied in agricultural sector as a biological fertilizer

ISOLATION, SELECTION AND MOLECULAR IDENTIFICATION OF ANTIBIOTIC PRODUCING ACTINOMYCETES

Microorganisms is one of the biodiversity that is priceless. One type of microorganisms is actinomycetes. Actinomycetes is a potential source of bioactive compounds producing high comercial value in pharmaceuticals and agricultures. The results showed that 44 Actinomycetes isolates has the ability to produce secondary metabo-lites antibiotics. They were active against M. luteus, S. aureus, E. coli, B. subtilis, S. cerevisiae, and C. albicans. Morphological studied indicated that the A16.1 isolate belonged to the genus Streptomyces. The identifi cation result based on 16S rDNA sequence is S. hygroscopicus subsp. azalomyceticus with 97% similarity level

Antifungal Activity of Endophytic Bacteria Associated with Sweet Sorghum (Sorghum bicolor)

The contribution of endophytic bacteria to the wellbeing of plants as biocontrol agents may be due to endophytic bacteria growing in the same niche as phytopathogens. This work was conducted to study the antagonistic activity of endophytic bacteria recovered from sweet sorghum against Sclerotium rolfsii, Fusarium solani, Fusarium oxysporum, Colletotrichum gloeosporioides in vitro and evaluate the mechanisms of these fungal inhibitions. We selected 78 endophytic bacteria from the stem and root of sweet sorghum plants. They were tested for antagonist activity by direct confrontation method. Antifungal compound production and lytic enzyme activity were examined to determine their mechanisms in inhibiting fungal pathogens. Antifungal compound production was checked by detecting the presence of NRPS and PKS genes. Lytic enzyme activity of the bacteria was evaluated by their ability to produce cellulase, chitinase, and protease. Selected bacteria were identified using molecular analysis based on the 16S rRNA gene. 14 out of the 78 tested isolates showed antagonistic activity and two were able to inhibit all four tested fungal strains. Four bacteria, designated as ACIL1, ACNM4, ACNM6, and ATNM4, produced natural products via NRPS pathway, but only one bacterial extract, designated as ACNM4, showed fungal inhibition. Ten isolates were able to produce hydrolytic enzymes. Endophytic bacteria identified as Burkholderia were revealed to have potential as a biocontrol agent

Antifungal Activity of Bacterial Isolates from Straw Mushroom Cultivation Medium against Phytopathogenic Fungi

Several bacteria were isolated from straw mushroom (Volvariella volvacea) cultivation medium. There are three potential isolates previously characterized and has growth inhibition effect against V. volvacea. This screening result lead to the further study about the inhibition activity against phytopathogenic fungi. The aim of this research is to investigate the antifungal activity of three bacterial isolates against three phytopathogenic fungi and identification of the bacteria. The method used in this study are antifungal assay using co-culture method and disk difussion assay using the filtrate of each bacteria. The profile of antifungal compound was identified using ethyl acetate extract followed by evaporation and gas chromatography (GC-MS) analysis. Identification of each isolates was performed using 16S rDNA amplification and sequencing. Three phytopathogenic fungi i.e Cercospora lactucae (InaCC F168), Colletotrichum gloeosporides (InaCC F304) and Fusarium oxysporum f.sp. cubense (F817) were co-cultured with bacterial isolates C2.2, C3.8, and D3.3. The C3.8 isolate has highest average inhibition activity either using isolate and filtrate. The result relatively consistent against three phytopathogenic fungi. The metabolite profile of C3.8 isolate showed the Bis(2-ethylhexyl) phthalate as the main compound with 97% similarity. Bis(2-ethylhexyl) phthalate has potential effect as antibacterial and antifungal compound. According to EzBioCloud and GeneBank databases, the C2.2 isolate identified as Bacillus tequilensis, C3.8 as Bacillus siamensis and D3.3 as Bacillus subtilis subsp. Subtilis. This study also shows the potential of Bacillus siamensis C3.8 as biocontrol against phytopathogenic fungi

Keanekaragaman Bakteri Tanah dari Teluk kodek Area, Pamenan Lombok Barat

Indonesia is the center of the world's biodiversity with a unique biodiversity and priceless. To explore and get more information about biodiversity, strategic research was needed.  The objective of this reseach was to explore the population diversity of soil microbes in Lombok Island.  Bacterium isolates was identified by molecular 16S rDNA. Soil samples from 5 different sites in Lombok Island showed various bacteria population. The highest population 113 x 106 CFU/g soil was found in soil sample around Plumeria acuminata and the lowest 34 x 106CFU/g soil was found in soil sample around  Tamarindus indica tree.  Fourteen of isolates were identified using molecular identification with homology from 94 – 100%