Yantyati Widyastuti

Asaia bogorensis gen. nov., sp. nov., an unusual acetic acid bacterium in the alpha-Proteobacteria.

Eight Gram-negative, aerobic, rod-shaped and peritrichously flagellated strains were isolated from flowers of the orchid tree (Bauhinia purpurea) and of plumbago (Plumbago auriculata), and from fermented glutinous rice, all collected in Indonesia. The enrichment culture approach for acetic acid bacteria was employed, involving use of sorbitol medium at pH 3.5. All isolates grew well at pH 3.0 and 30 degrees C. They did not oxidize ethanol to acetic acid except for one strain that oxidized ethanol weakly, and 0.35% acetic acid inhibited their growth completely. However, they oxidized acetate and lactate to carbon dioxide and water. The isolates grew well on mannitol agar and on glutamate agar, and assimilated ammonium sulfate for growth on vitamin-free glucose medium. The isolates produced acid from D-glucose, D-fructose, L-sorbose, dulcitol and glycerol. The quinone system was Q-10. DNA base composition ranged from 59.3 to 61.0 mol% G + C. Studies of DNA relatedness showed that the isolates constitute a single species. Phylogenetic analysis based on their 16S rRNA gene sequences indicated that the isolates are located in the acetic acid bacteria lineage, but distant from the genera Acetobacter, Gluconobacter, Acidomonas and Gluconacetobacter. On the basis of the above characteristics, the name Asaia bogorensis gen. nov., sp. nov. is proposed for these isolates. The type strain is isolate 71T (= NRIC 0311T = JCM 10569T).

Importance of lactobacilli in food and feed biotechnology

The genus Lactobacillus is a heterogeneous group of lactic acid bacteria (LAB) with important implications in food fermentation. The ability to colonize a variety of habitats is a direct consequence of the wide metabolic versatility of this group of LAB. Consequently, lactobacilli have been used for decades in food preservation, as starters for dairy products, fermented vegetables, fish and sausages as well as silage inoculants. Lactobacilli have also been proposed as probiotics and microbial cell factories for the production of nutraceuticals. However, a wide range of applications of lactobacilli in food biotechnology remains potential, whereas a number of important strains still need to be discovered and characterized. This article provides an overview of the taxonomy of lactobacilli and describes four of the most significant case studies on the application of this group of LAB in food and feed biotechnology, including their use as probiotics, dairy starters, silage inoculants, and microbial cell factories. The importance of access to and exchange of biological material within and between different strain collections as a crucial step in expanding the range of different biotechnological applications of lactobacilli is also emphasized.

Kozakia baliensis gen. nov., sp. nov., a novel acetic acid bacterium in the alpha-proteobacteria.

Four bacterial strains were isolated from palm brown sugar and ragi collected in Bali and Yogyakarta, Indonesia, by an enrichment culture approach for acetic acid bacteria. Phylogenetic analysis based on 16S rRNA gene sequences showed that the four isolates constituted a cluster separate from the genera Acetobacter, Gluconobacter, Acidomonas, Gluconacetobacter and Asaia with a high bootstrap value in a phylogenetic tree. The isolates had high values of DNA-DNA similarity (78-100%) between one another and low values of the similarity (7-25%) to the type strains of Acetobacter aceti, Gluconobacter oxydans, Gluconacetobacter liquefaciens and Asaia bogorensis. The DNA base composition of the isolates ranged from 56.8 to 57.2 mol% G+C with a range of 0-4 mol%. The major quinone was Q-10. The isolates oxidized acetate and lactate to carbon dioxide and water, but the activity was weak, as with strains of Asaia bogorensis. The isolates differed from Asaia bogorensis strains in phenotypic characteristics. The name Kozakia baliensis gen. nov., sp. nov., is proposed for the four isolates. Strain Yo-3T (= NRIC 0488T = JCM 11301T = IFO 16664T = DSM 14400T) was isolated from palm brown sugar collected in Bali, Indonesia, and was designated as the type strain.

Oleibacter marinus gen. nov., sp. nov., a bacterium that degrades petroleum aliphatic hydrocarbons in a tropical marine environment

Three Gram-negative, motile, mesophilic, aerobic, rod-shaped bacterial strains, designated 2O1(T), 1O14 and 1O18, were isolated from Indonesian seawater after enrichment with crude oil and a continuous supply of supplemented seawater. The strains exhibited high n-alkane-degrading activity, which indicated that the strains were important degraders of petroleum aliphatic hydrocarbons in tropical marine environments. Phylogenetic analyses based on 16S rRNA gene sequences of members of the Gammaproteobacteria showed that the isolates formed a coherent and distinct cluster in a stable lineage containing Oceanobacter kriegii IFO 15467(T) (96.4-96.5 % 16S rRNA gene sequence similarity) and Thalassolituus oleivorans MIL-1(T). DNA G +C content was 53.0-53.1 mol%. The major fatty acids were C(16 : 0), C(16 : 1)ω7 and C(18 : 1)ω9 and the hydroxy fatty acids were C(12 : 0) 3-OH and C(10 : 0) 3-OH. The polar lipids were phosphatidylglycerol, a ninhydrin-positive phospholipid(s) and glycolipids. The major quinone was Q-9 (97-99 %), which distinguished the isolates from Oceanobacter kriegii NBRC 15467(T) (Q-8; 91 %). On the basis of phenotypic, genotypic and chemotaxonomic data, including DNA-DNA hybridization, the isolates represent a novel genus and species, for which the name Oleibacter marinus gen. nov., sp. nov. is proposed. The type strain of Oleibacter marinus is 2O1(T) (=NBRC 105760(T) =BTCC B-675(T)).

Actinokineospora baliensis sp. nov., Actinokineospora cibodasensis sp. nov. and Actinokineospora cianjurensis sp. nov., isolated from soil and plant litter.

Six actinomycete strains isolated from soil and plant-litter samples in Indonesia were studied for their taxonomic position by using a polyphasic approach. Phylogenetically, all the strains were located in the broad cluster of the genus Actinokineospora. Chemotaxonomic data [cell-wall diamino acid, meso-diaminopimelic acid; cell-wall peptidoglycan, type III (A1γ); major sugars, galactose and arabinose; major menaquinone, MK-9(H₄); major fatty acid, iso-C₁₆:₀; major phospholipid, phosphatidylethanolamine] supported the affiliation of all six strains to the genus Actinokineospora. The results of DNA-DNA hybridization with DNA from type strains of Actinokineospora species with validly published names revealed three DNA-DNA relatedness groups. Group I (ID03-0561(T)) showed low relatedness to the other strains studied. The three strains in group II (ID03-0784(T), ID03-0808 and ID03-0809) formed a group with high relatedness (98-100 %) and showed low relatedness to the other strains studied. The two strains in group III (ID03-0810(T) and ID03-0813) showed 58-68 % relatedness to Actinokineospora terrae NBRC 15668(T) and showed low relatedness (2-24 %) to the other strains studied. The description of three novel species is proposed: Actinokineospora baliensis sp. nov., for the single strain in group I (type strain ID03-0561(T) =BTCC B-554(T) =NBRC 104211(T)), Actinokineospora cibodasensis sp. nov., for the strains in group II (type strain ID03-0784(T) =BTCC B-555(T) =NBRC 104212(T)), and Actinokineospora cianjurensis sp. nov., for the strains in group III (type strain ID03-0810(T) =BTCC B-558(T) =NBRC 105526(T)).

Dietzia timorensis sp. nov., isolated from soil.

An actinomycete strain, ID05-A0528(T), was isolated using the SDS-yeast extract pre-treatment method from soil under mahogany (Swietenia mahogani) trees in West Timor, Indonesia, and was examined by using a polyphasic taxonomic approach. Chemotaxonomic and phylogenetic characterizations demonstrated that the novel strain belongs to the genus Dietzia. 16S rRNA gene sequencing studies showed that the strain was related to Dietzia cinnamea (97.2 %). Results of phenotypic and phylogenetic analyses determined that strain ID05-A0528(T) is different from the known species of the genus Dietzia. It is proposed that the isolate should be classified as a representative of a novel species of the genus Dietzia, with the name Dietzia timorensis sp. nov. The type strain is ID05-A0528(T) (=BTCC B-560(T) =NBRC 104184(T)).

Actinophytocola timorensis sp. nov. and Actinophytocola corallina sp. nov., isolated from soil

Two actinomycete strains, ID05-A0653(T) and ID06-A0464(T), were isolated from soils of West Timor and Lombok island, respectively, in Indonesia. 16S rRNA gene sequence analysis clearly demonstrated that the isolates belonged to the family Pseudonocardiaceae and were closely related to the genus Actinophytocola. Strains ID05-A0653(T) and ID06-A0464(T) exhibited 98.1 and 98.2 % 16S rRNA gene sequence similarity, respectively, with Actinophytocola oryzae GMKU 367(T). The isolates grew well on ISP media and produced white aerial mycelium. Short spore chains were formed directly on the substrate mycelium. The isolates contained meso-diaminopimelic acid, arabinose and galactose as cell-wall components, MK-9(H(4)) as the sole isoprenoid quinone, iso-C(16 : 0) as the major cellular fatty acid and phosphatidylethanolamine as the diagnostic polar lipid. The DNA G+C contents of strains ID05-A0653(T) and ID06-A0464(T) were 69.7 and 71.2 mol%, respectively. On the basis of phenotypic characteristics, DNA-DNA relatedness and 16S rRNA gene sequence comparisons, strains ID05-A0653(T) and ID06-A0464(T) each represent a novel species of the genus Actinophytocola, for which the names Actinophytocola timorensis sp. nov. (type strain ID05-A0653(T)  = BTCC B-673(T)  = NBRC 105524(T)) and Actinophytocola corallina sp. nov. (type strain ID06-A0464(T)  = BTCC B-674(T)  = NBRC 105525(T)) are proposed.

Streptomyces baliensis sp. nov., isolated from Balinese soil

The taxonomic positions of actinomycete strains ID03-0915T and ID03-0825, isolated from soil on the Indonesian island of Bali, were examined using a polyphasic taxonomic approach. The morphological and chemotaxonomic characteristics of these organisms are typical of the genus Streptomyces. Phylogenetic analyses performed using almost-complete 16S rRNA gene sequences demonstrated that the strains were closely related to Streptomyces glauciniger and Streptomyces lilacinus. However, DNA-DNA hybridization and phenotypic characteristics revealed that the strains differed from known Streptomyces species. Therefore, we conclude that strains ID03-0915T and ID03-0825 (=BTCC B-563) represent a novel species of the genus Streptomyces, for which we propose the name Streptomyces baliensis sp. nov. The type strain is strain ID03-0915T (=BTCC B-608T=NBRC 104276T).

Fermentation Characteristics and Microbial Diversity of Tropical Grass-legumes Silages

Calliandra calothyrsus preserved in silage is an alternative method for improving the crude protein content of feeds for sustainable ruminant production. The aim of this research was to evaluate the quality of silage which contained different levels of C. calothyrsus by examining the fermentation characteristics and microbial diversity. Silage was made in a completely randomized design consisting of five treatments with three replications i.e.: R0, Pennisetum purpureum 100%; R1, P. purpureum 75%+C. calothyrsus 25%;, R2, P. purpureum 50%+C. calothyrsus 50%; R3, P. purpureum 25%+C. calothyrsus 75%; and R4, C. calothyrsus 100%. All silages were prepared using plastic jar silos (600 g) and incubated at room temperature for 30 days. Silages were analyzed for fermentation characteristics and microbial diversity. Increased levels of C. calothyrsus in silage had a significant effect (p<0.01) on the fermentation characteristics. The microbial diversity index decreased and activity was inhibited with increasing levels of C. calothyrsus. The microbial community indicated that there was a population of Lactobacillus plantarum, L. casei, L. brevis, Lactococcus lactis, Chryseobacterium sp., and uncultured bacteria. The result confirmed that silage with a combination of grass and C. calothyrsus had good fermentation characteristics and microbial communities were dominated by L. plantarum.

Isolation and Screening of Surfactant-producing Bacteria from Indonesian Marine Environments and Its Application on Bioremediation

Isolation and screening have been undertaken on oil-degrading microbes from Indonesian marine environments. During screening process it has been found many bacterial isolates capable of degrading crude oil. Hence, study has been focused on the biodiversity of biosurfactant-producing bacterial species in Indonesian marine environment and its function for remedial the pollutant in marine and soil areas. A total of 103 out of 463 isolates showed positive surfactant-degrading properties. By means of partial 16S rRNA gene analyses, it has been found that the majority of taxa are related to Alcanivorax, Pseudomonas, Bacillus, Bortetela, Brucella, Acenitobacter, Staphia, Lysobacter, and Talasosophira . Biosurfactant properties assay showed that they were capable of lowering the surface- and interfacial water tension from 74 mN/m to 40-65 mN/m and from 24 mN/m to 6-10 mN/m, respectively. In addition, most of the surfactants were capable of emulsifying hydrocarbon (crude oil) of 0.01 to 0.15 units, comparable to 0.08 units of synthetic surfactant (20% Tween). Further observation showed that the majority of the surfactants were able to degrade a long chain of alkane, but not branched alkane, with a recovering rate of 20-80%. The application of the surfactant towards oil polluted model beach was done in laboratory scale and showing the surfactant obtained from microbial broth cultures capable for recovering the oil pollutant significantly, compared to the control (without addition microbial broth).