Gandjariella thermophila gen. nov., sp. nov., a new member of the family Pseudonocardiaceae, isolated from forest soil in a geothermal area.

Six mycelium-forming actinomycete strains were isolated from forest soil near the Cisolok geysers in West Java, Indonesia. The 16S rRNA gene sequences of these strains showed high similarity to members of genera in the family Pseudonocardiaceae with values less than 96.0 %, and most closely related to the genus Thermotunica, T. guangxiensis AG2-7T(94.6-95.2 % similarity). The type strain, designated SL3-2-4T, was aerobic, thermophilic, Gram-stain-positive that formed branched, non-fragmented substrate mycelia and unbranched aerial mycelia with long-chain, oval-shaped spores on International Streptomyces Project (ISP) 3 medium. It produced light-orange substrate mycelia and light-orange diffusible pigments on ISP 3 medium with 2 % gellan gum, grown at 30-55 °C, with optimum growth at 45 °C. The pH range for growth was 4.0-8.0, with optimum growth at pH 7.0. Strain SL3-2-4T was able to hydrolyze casein, esculin, gelatin, guanine, hypoxanthine, starch, L-tyrosine, and xanthine, but not adenine, carboxymethyl-cellulose, cellulose, chitin, Tween 20, or xylan. The major fatty acid was iso-C16 : 0, and the major menaquinone was MK-8 (H4). The detected polar lipids were diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine, phosphatidyl-N-methylethanolamine, unidentified aminophospholipids, unidentified glycolipids, and unidentified phospholipids. The cell wall hydrolysate of SL3-2-4T contained meso-2,4-diaminopimelic acid. The whole cell sugars were arabinose and galactose. The DNA G+C content was 71.6 mol%. Phenotypic features and phylogenetic data differentiated SL3-2-4T from members of the family Pseudonocardiaceae. Therefore, the strain SL3-2-4T is proposed as a representative of a novel species in a novel genus, Gandjariella thermophila gen. nov., sp. nov. The type strain is SL3-2-4T (=UICC B-83T=NRRL B-67478T=InaCC A981T).

Paenibacillus cisolokensis sp. nov., isolated from litter of a geyser.

A Gram-stain-positive, endospore-forming, aerobic and thermophilic bacterium, designated strain LC2-13AT, was isolated from Cisolok geyser, West Java, Indonesia, at 50 °C. The isolate was rod-shaped and motile by means of peritrichous flagella. The major cellular fatty acids were iso-C16 : 0, C16 : 0 and anteiso-C15 : 0 and the major quinone was menaquinone 7. The major polar lipids were diphosphatidylglycerol, phosphatidylglycerol and phosphatidylethanolamine. The genomic DNA G+C content was 56.6 mol% and the major diagnostic diamino acid in the cell-wall peptidoglycan was meso-diaminopimelic acid. Phylogenetic analysis based on the 16S rRNA gene sequence indicated that strain LC2-13AT is related most closely to Paenibacillus kobensis DSM 10249T (94.86 % similarity), Paenibacillus tarimensis SA-7-6T (94.77 %) and Paenibacillus barengoltzii SAFN-016T (94.77 %). On the basis of phenotypic, chemotaxonomic and phylogenetic evidence, strain LC2-13AT is affiliated to the genus Paenibacillus, but could be distinguished from recognized species of this genus. A novel species with the name Paenibacillus cisolokensis sp. nov. is thus proposed. The type strain is LC2-13AT (=UICC B-42T=NRRL B-65368T=DSM 101873T).

Characterization of dengue virus serotype 4 infection in Jakarta, Indonesia.

Dengue hemorrhagic fever has become a worldwide health issue. Heterologous infection by different serotypes may lead to severe forms of dengue infection and even death. In a cohort study in Jakarta from 2009 to 2010 with inclusion criteria of adults with fever of less than 48 hours, 72% were confirmed dengue infection from a total of suspected 190 dengue patients. Using RT-PCR, 16 patients were infected with DENV-4 with or without co-infection with other serotype(s). Dengue fever (DF) patients (73%) were infected with DENV-4 alone, while mixed infections were present in more severe clinical manifestations of DHF grade I and II. The nucleotide sequences of envelope (E) protein gene of 3 isolates of DENV-4 showed that they belonged to genotype II. There were 50 nucleotide substitutions, but only 3 amino acid changes were found at positions 130, 233 and 455, present in E protein isolated from DHF grade II patient.

Unravelling the Molecular Identification and Antifungal Susceptibility Profiles of Aspergillus spp. Isolated from Chronic Pulmonary Aspergillosis Patients in Jakarta, Indonesia: The Emergence of Cryptic Species.

Cryptic species of Aspergillus have rapidly increased in the last few decades. Chronic pulmonary aspergillosis (CPA) is a debilitating fungal infection frequently affecting patients with previous TB. The identification and antifungal susceptibility profiles of different species of Aspergillus are important to support the management of CPA. The aim of this study was to describe the molecular and susceptibility profiles of Aspergillus isolated from CPA patients. The species identity of isolates was determined by combined DNA analyses of internal transcribed space (ITS), partial ß-tubulin genes, and part of the calmodulin gene. We revealed a high (27%) prevalence of cryptic species among previous tuberculosis patients with persistent symptoms. Twenty-nine (49%) patients met the criteria for diagnosis of CPA with 24% containing Aspergillus cryptic species. This is the first report of five cryptic Aspergillus species from clinical isolates in Indonesia: A. aculea tus, A. neoniger, A. brunneoviolacues, A. welwitschiae, and A. tubingensis. Significantly, there was decreased sensitivity against itraconazole in the CPA group (66% susceptible to itraconazole) compared to the non-CPA group (90% susceptible to itraconazole) (p = 0.003). The species-level characterisation of Aspergillus and its antifungal susceptibility tests demands greater attention to better the management of CPA patients.

The effect of the use of commercial tempeh starter on the diversity of Rhizopus tempeh in Indonesia.

At present, only a single Rhizopus species, R. microsporus, can be found in fresh tempeh produced in Java, Indonesia. The loss of diversity of Rhizopus in tempeh has been associated with the widespread use of commercial tempeh starter in Indonesia since the 2000s. However, the identities of the previous Rhizopus strains associated with tempeh, which have been preserved in a culture collection in Indonesia, have not been verified. The present study aimed to verify the identities of 22 Rhizopus strains isolated from tempeh produced using the traditional tempeh starters from the 1960s to the 2000s. Phylogenetic analysis based on the ITS regions in the rRNA gene sequence data, revealed that the Rhizopus strains belonged to the species R. arrhizus (five strains); R. delemar (14 strains); and R. microsporus (three strains). Verification of the identities of these Rhizopus strains in the present study confirmed the loss of diversity of Rhizopus species in tempeh produced in Indonesia, particularly in Java. Our findings confirmed that the morphological changes in Rhizopus species isolated from tempeh as a result of centuries of domestication.

Insecticidal Activity of Ethyl Acetate Extracts from Culture Filtrates of Mangrove Fungal Endophytes.

In the search for novel potent fungi-derived bioactive compounds for bioinsecticide applications, crude ethyl acetate culture filtrate extracts from 110 mangrove fungal endophytes were screened for their toxicity. Toxicity tests of all extracts against brine shrimp (Artemia salina) larvae were performed. The extracts with the highest toxicity were further examined for insecticidal activity against Spodoptera litura larvae and acetylcholinesterase (AChE) inhibition activity. The results showed that the extracts of five isolates exhibited the highest toxicity to brine shrimp at 50% lethal concentration (LC50) values of 7.45 to 10.24 ppm. These five fungal isolates that obtained from Rhizophora mucronata were identified based on sequence data analysis of the internal transcribed spacer region of rDNA as Aspergillus oryzae (strain BPPTCC 6036), Emericella nidulans (strains BPPTCC 6035 and BPPTCC 6038), A. tamarii (strain BPPTCC 6037), and A. versicolor (strain BPPTCC 6039). The mean percentage of S. litura larval mortality following topical application of the five extracts ranged from 16.7% to 43.3%. In the AChE inhibition assay, the inhibition rates of the five extracts ranged from 40.7% to 48.9%, while eserine (positive control) had an inhibition rate of 96.8%, at a concentration of 100 ppm. The extracts used were crude extracts, so their potential as sources of AChE inhibition compounds makes them likely candidates as neurotoxins. The high-performance liquid chromatography profiles of the five extracts differed, indicating variations in their chemical constituents. This study highlights the potential of culture filtrate ethyl acetate extracts of mangrove fungal endophytes as a source of new potential bioactive compounds for bioinsecticide applications.

New species of the genus Metschnikowia isolated from flowers in Indonesia, Metschnikowia cibodasensis sp. nov.

A novel species, Metschnikowia cibodasensis, is proposed to accommodate eight strains (ID03- 0093(T), ID03-0094, ID03-0095, ID03-0096, ID03-0097, ID03-0098, ID03-0099, and ID03-0109) isolated from flowers of Saurauia pendula, Berberis nepalensis, and Brunfelsia americana in Cibodas Botanical Garden, West Java, Indonesia. The type strain of M. cibodasensis is ID03- 0093(T) (= NBRC 101693(T) =UICC Y-335(T) = BTCC-Y25(T)). The common features of M. cibodasensis are a spherical to ellipsoidopedunculate shaped ascus, which contains one or two needleshaped ascospores, and lyse at maturity. Asci generally develop directly from vegetative cells but sometimes from chlamydospores. The neighbor-joining tree based on the D1/D2 domain of nuclear large subunit (nLSU) ribosomal DNA sequences strongly supports that M. cibodasensis (eight strains) and its closest teleomorphic species, M. reukaufii, are different species by a 100% bootstrap value. The type strain of M. cibodasensis, ID03-0093(T), differed from M. reukaufii NBRC 1679(T) by six nt (five substitutions and one deletion) in their D1/D2 region of nLSU rDNA, and by 18 nt (five deletions, four insertions, and nine substitutions) in their internal transcribed spacer regions of rDNA, respectively. Four strains representative of M. cibodasensis (ID03-0093(T), ID03-0095, ID03-0096, and ID03-0099) showed a mol% G+C content of 44.05 ± 0.25%, whereas that of M. reukaufii NBRC 1679(T) was 41.3%. The low value of DNADNA homology (5-16%) in four strains of M. cibodasensis and M. reukaufii NBRC 1679(T) strongly supported that these strains represent a distinct species.