Xylanopectinolytic enzymes by marine actinomycetes from sediments of Sarena Kecil, North Sulawesi: high potential to produce galacturonic acid and xylooligosaccharides from raw biomass.

BACKGROUND: Actinomycetes isolated from marine habitats are known to have the potential for novel enzymes that are beneficial in the industry. In-depth knowledge is necessary given the variety of this bacterial group in Indonesia and the lack of published research. Actinomycetes isolates (BLH 5-14) obtained from marine sediments of Sarena Kecil, Bitung, North Sulawesi, Indonesia, showed an ability to produce pectinase and xylanase that have equal or even higher potential for pectic-oligosaccharides (POS) and xylooligosaccharides (XOS) production from raw biomass than from commercial substrates. This study's objective was to characterize both enzymes to learn more for future research and development. RESULTS: Pectinase had the highest activity on the 6th day (1.44±0.08 U/mL) at the optimum pH of 8.0 and optimum temperature of 50 °C. Xylanase had the maximum activity on the 6th day (4.33±0.03 U/mL) at optimum pH 6.0 and optimum temperature 60 °C. Hydrolysis and thin layer chromatography also showed that pectinase was able to produce monosaccharides such as galacturonic acid (P1), and xylanase was able to yield oligosaccharides such as xylotriose (X3), xylotetraose (X4), and xylopentaose (X5). BLH 5-14 identified as the genus Streptomyces based on the 16S rDNA sequences and the closely related species Streptomyces tendae (99,78%). CONCLUSIONS: In the eco-friendly paper bleaching industry, Streptomyces tendae has demonstrated the potential to create enzymes with properties that can be active in a wide range of pH levels. The oligosaccharides have the potential as prebiotics or dietary supplements with anti-cancer properties. Further research is needed to optimize the production, purification, and development of the application of pectinase and xylanase enzymes produced by Actinomycetes isolates.

Chemical Composition and Sensory Profiles of Fermented Cocoa Beans Obtained from Various Regions of Indonesia.

The chemical composition and sensory profile of cocoa beans are essential factors determining the quality of cocoa-based products. In this study, cocoa bean samples were collected from various regions of Indonesia, including Aceh, Banten, Bali, East Java, West Sumatra, West Sulawesi, East Kalimantan, and Yogyakarta. The cocoa beans were fermented and sun-dried according to the producers' protocols and local practices. The sensory profile, fat content, total phenolic content, and the composition of sugars, organic acids, and amino acids of the cocoa bean samples were analyzed. The results revealed that the chemical composition and sensory profiles of the samples were diverse. The sensory profiles of cocoa liquor samples were described by low intensities of cocoa notes with the occurrence of fruity, floral, spicy, and sweet notes. The concentration of acetic acid, lactic acid, and some amino acids (glutamic acid, proline, and methionine) was associated with fresh fruit, browned fruit, and roasted note of the cocoa liquor, respectively. The variation in the environmental conditions and postharvest practices contributed to the diversity of cocoa beans' chemical and sensory characteristics.

Harnessing originally robust yeast for rapid lactic acid bioproduction without detoxification and neutralization.

Acidic and chemical inhibitor stresses undermine efficient lactic acid bioproduction from lignocellulosic feedstock. Requisite coping treatments, such as detoxification and neutralizing agent supplementation, can be eliminated if a strong microbial host is employed in the process. Here, we exploited an originally robust yeast, Saccharomyces cerevisiae BTCC3, as a production platform for lactic acid. This wild-type strain exhibited a rapid cell growth in the presence of various chemical inhibitors compared to laboratory and industrial strains, namely BY4741 and Ethanol-red. Pathway engineering was performed on the strain by introducing an exogenous LDH gene after disrupting the PDC1 and PDC5 genes. Facilitated by this engineered strain, high cell density cultivation could generate lactic acid with productivity at 4.80 and 3.68 g L-1 h-1 under semi-neutralized and non-neutralized conditions, respectively. Those values were relatively higher compared to other studies. Cultivation using real lignocellulosic hydrolysate was conducted to assess the performance of this engineered strain. Non-neutralized fermentation using non-detoxified hydrolysate from sugarcane bagasse as a medium could produce lactic acid at 1.69 g L-1 h-1, which was competitive to the results from other reports that still included detoxification and neutralization steps in their experiments. This strategy could make the overall lactic acid bioproduction process simpler, greener, and more cost-efficient.

Expression of Codon-Optimized Gene Encoding Murine Moloney Leukemia Virus Reverse Transcriptase in Escherichia coli.

Moloney murine leukemia virus reverse transcriptase (MMLV-RT) is the most frequently used enzyme in molecular biology for cDNA synthesis. To date, reverse transcription coupled with Polymerase Chain Reaction, known as RT-PCR, has been popular as an excellent approach for the detection of SARS-CoV-2 during the COVID-19 pandemic. In this study, we aimed to improve the enzymatic production and performance of MMLV-RT by optimizing both codon and culture conditions in E. coli expression system. By applying the optimized codon and culture conditions, the enzyme was successfully overexpressed and increased at high level based on the result of SDS-PAGE and Western blotting. The total amount of MMLV-RT has improved 85-fold from 0.002 g L-1 to 0.175 g L-1 of culture. One-step purification by nickel affinity chromatography has been performed to generate the purified enzyme for further analysis of qualitative and quantitative RT activity. Overall, our investigation provides useful strategies to enhance the recombinant enzyme of MMLV-RT in both production and performance. More importantly, the enzyme has shown promising activity to be used for RT-PCR assay.

The dynamics of circulating SARS-CoV-2 lineages in Bogor and surrounding areas reflect variant shifting during the first and second waves of COVID-19 in Indonesia.

Background: Indonesia is one of the Southeast Asian countries with high case numbers of COVID-19 with up to 4.2 million confirmed cases by 29 October 2021. Understanding the genome of SARS-CoV-2 is crucial for delivering public health intervention as certain variants may have different attributes that can potentially affect their transmissibility, as well as the performance of diagnostics, vaccines, and therapeutics. Objectives: We aimed to investigate the dynamics of circulating SARS-CoV-2 variants over a 15-month period in Bogor and its surrounding areas in correlation with the first and second wave of COVID-19 in Indonesia. Methods: Nasopharyngeal and oropharyngeal swab samples collected from suspected patients from Bogor, Jakarta and Tangerang were confirmed for SARS-CoV-2 infection with RT-PCR. RNA samples of those confirmed patients were subjected to whole genome sequencing using the ARTIC Network protocol and sequencer platform from Oxford Nanopore Technologies (ONT). Results: We successfully identified 16 lineages and six clades out of 202 samples (male n = 116, female n = 86). Genome analysis revealed that Indonesian lineage B.1.466.2 dominated during the first wave (n = 48, 23.8%) while Delta variants (AY.23, AY.24, AY.39, AY.42, AY.43 dan AY.79) were dominant during the second wave (n = 53, 26.2%) following the highest number of confirmed cases in Indonesia. In the spike protein gene, S_D614G and S_P681R changes were dominant in both B.1.466.2 and Delta variants, while N439K was only observed in B.1.466.2 (n = 44) and B.1.470 (n = 1). Additionally, the S_T19R, S_E156G, S_F157del, S_R158del, S_L452R, S_T478K, S_D950N and S_V1264L changes were only detected in Delta variants, consistent with those changes being characteristic of Delta variants in general. Conclusions: We demonstrated a shift in SARS-CoV-2 variants from the first wave of COVID-19 to Delta variants in the second wave, during which the number of confirmed cases surpassed those in the first wave of COVID-19 pandemic. Higher proportion of unique mutations detected in Delta variants compared to the first wave variants indicated potential mutational effects on viral transmissibility that correlated with a higher incidence of confirmed cases. Genomic surveillance of circulating variants, especially those with higher transmissibility, should be continuously conducted to rapidly inform decision making and support outbreak preparedness, prevention, and public health response.

Potential secondary metabolite from Indonesian Actinobacteria (InaCC A758) against Mycobacterium tuberculosis.

OBJECTIVES: This study explored Indonesian Actinobacteria which were isolated from Curcuma zedoaria endophytic microbes and mangrove ecosystem for new antimycobacterial compounds. MATERIALS AND METHODS: Antimycobacterial activity test was carried out against Mycobacterium tuberculosis H37Rv. Chemical profiling of secondary metabolite using Gas Chromatography-Mass Spectroscopy (GC-MS) and High Resolution-Mass Spectroscopy (HR-MS) was done to the ethyl acetate extract of active strain InaCC A758. Molecular taxonomy analysis based on 16S rRNA gene and biosynthetic gene clusters analysis of polyketide synthase (PKS) and non-ribosomal peptide synthetase (NRPS) from InaCC A758 have been carried out. Bioassay guided isolation of ethyl acetate extract was done, then structural elucidation of active compound was performed using UV-Vis, FT-IR, and NMR spectroscopy methods. RESULTS: The chemical profiling using HR-MS revealed that InaCC A758 has the potential to produce new antimycobacterial compounds. The 16S rRNA gene sequencing showed that InaCC A758 has the closest homology to Streptomyces parvus strain NBRC 14599 (99.64%). In addition, InaCC A758 has NRPS gene and related to S. parvulus (92% of similarity), and also PKS gene related to PKS-type borrelidin of S. rochei and S. parvulus (74% of similarity). Two compounds with potential antimycobacterial were predicted as 1) Compound 1, similar to dimethenamid (C12H18ClNO2S; MW 275.0723), with MIC value of 100 µg/ml, and 2) Compound 2, actinomycin D (C62H86N12O16; MW 1254.6285), with MIC value of 0.78 µg/ml. CONCLUSION: Actinomycin D has been reported to have antimycobacterial activity, however the compound has been predicted to resemble dimethenamid had not been reported to have similar activity.

Mining Indonesian Microbial Biodiversity for Novel Natural Compounds by a Combined Genome Mining and Molecular Networking Approach.

Indonesia is one of the most biodiverse countries in the world and a promising resource for novel natural compound producers. Actinomycetes produce about two thirds of all clinically used antibiotics. Thus, exploiting Indonesia's microbial diversity for actinomycetes may lead to the discovery of novel antibiotics. A total of 422 actinomycete strains were isolated from three different unique areas in Indonesia and tested for their antimicrobial activity. Nine potent bioactive strains were prioritized for further drug screening approaches. The nine strains were cultivated in different solid and liquid media, and a combination of genome mining analysis and mass spectrometry (MS)-based molecular networking was employed to identify potential novel compounds. By correlating secondary metabolite gene cluster data with MS-based molecular networking results, we identified several gene cluster-encoded biosynthetic products from the nine strains, including naphthyridinomycin, amicetin, echinomycin, tirandamycin, antimycin, and desferrioxamine B. Moreover, 16 putative ion clusters and numerous gene clusters were detected that could not be associated with any known compound, indicating that the strains can produce novel secondary metabolites. Our results demonstrate that sampling of actinomycetes from unique and biodiversity-rich habitats, such as Indonesia, along with a combination of gene cluster networking and molecular networking approaches, accelerates natural product identification.

Evaluation of crystal violet decolorization assay and resazurin microplate assay for antimycobacterial screening.

The main obstacle in antimycobacterial discovery is the extremely slow growth rates of pathogenic mycobacteria that lead to the long incubation times needed in antimycobacterial screening. Some in vitro testings has been developed and are currently available for antimycobacterial screening. The aim of the study was to compare Resazurin Microplate Assay (REMA) and Crystal Violet Decolorization Assay (CVDA) for testing mycobacteria susceptibility to isoniazid and rifampicin as well as for antimycobacterial screening of natural products (NP). Mycobacterium tuberculosis strain H37Rv and Mycobacterium smegmatis strain mc2 155 were used as tested mycobacteria. Serial two-fold dilutions from 0.0625 to 1.0 µg/mL for the isoniazid and rifampicin and from 6.25 to 100.0 µg/mL for the NP A and B were prepared. Tested mycobacteria were then incubated with tested drugs or NPs in each growth medium at 37 °C for 7 days for M. tuberculosis and 3 days for M. smegmatis. MIC values against M. tuberculosis were interpreted 24-48 h after adding resazurin or at least 72 h after adding crystal violet, whereas MIC values against M. smegmatis were interpreted 1 h after adding resazurin or 24 h after adding crystal violet. The MIC values against M. tuberculosis interpreted by REMA were 0.0625, 0.0625, 6.25, and >100 µg/mL for rifampicin, isoniazid, NP A, and NP B, respectively, and those interpreted by CVDA were 0.0625, 0.0625, 6.25, and >100 µg/mL for rifampicin, isoniazid, NP A, and NP B, respectively. Moreover, the MIC values against M. smegmatis interpreted by REMA were 0.0625, >1, 6.25, and 100 µg/mL for rifampicin, isoniazid, NP A, and NP B, respectively, and those interpreted by CVDA were 0.125, >1, 6.25, and >100 µg/mL for rifampicin, isoniazid, NP A, NP B respectively. In conclusion, REMA is faster and easier than CVDA to interpret MIC values, however CVDA produces higher MIC values than REMA for rifampicin and NP B in M. smegmatis susceptibility testing. Therefore, REMA and CVDA can be used for antimycobacterial screening.

Complete Genome Sequence of the Putative Phosphonate Producer Streptomyces sp. Strain I6, Isolated from Indonesian Mangrove Sediment.

Streptomyces sp. strain I6 is a novel strain isolated from an Indonesian mangrove sediment sample. Bioinformatic analysis of the genome sequence of Streptomyces sp. I6 revealed 23 biosynthetic gene clusters. One of them encodes the synthesis of a putative phosphonate secondary metabolite, a class of underexplored natural compounds with great pharmaceutical potential.

GH-10 and GH-11 Endo-1,4-ß-xylanase enzymes from Kitasatospora sp. produce xylose and xylooligosaccharides from sugarcane bagasse with no xylose inhibition.

A novel strategy for the low-cost, high-yield co-production of xylose and xylooligosaccharides together with no xylose inhibition was developed using a novel heterologous expression of XYN10Ks_480 endo-1,4-ß-xylanase with a ricin-type ß-trefoil type of domain and XYN11Ks_480 endo-1,4-ß-xylanase with a CBM 2 superfamily from the Kitasatospora sp in an actinomycetes expression system. Xylose is the main building block for hemicellulose xylan. Our findings demonstrated high levels of expression and catalytic activity for XYN10Ks_480 during hydrolysis of the extracted xylan of bagasse, and three types of xylan-based substrates were used to produce xylose and xylooligosaccharides. However, hydrolysis by XYN11Ks_480 produced xylooligosaccharides without xylose formation. This study demonstrated how integrating sodium hypochlorite-extracted xylan and enzymatic hydrolysis could provide an alternative strategy for the generation of XOS from lignocellulosic material.

Complete Genome Sequence of Streptomyces sp. Strain SHP22-7, a New Species Isolated from Mangrove of Enggano Island, Indonesia.

Streptomyces sp. SHP22-7 is a novel strain isolated from a mangrove sample on Enggano Island, Indonesia. Here, we present the 7.9-Mbp genome sequence of SHP22-7, which will provide insight into its natural compound biosynthetic potential.

Complete Genome Sequence of Streptomyces sp. Strain BSE7F, a Bali Mangrove Sediment Actinobacterium with Antimicrobial Activities.

The strain Streptomyces sp. BSE7F, a novel Streptomyces strain isolated from Indonesian mangrove sediment, displays antimicrobial activities against Gram-positive bacteria, Gram-negative bacteria, and yeast. Bioinformatic analysis of the genome sequence revealed the occurrence of 22 biosynthetic gene clusters disclosing the secondary metabolite capacity of strain BSE7F.

Xylanase and feruloyl esterase from actinomycetes cultures could enhance sugarcane bagasse hydrolysis in the production of fermentable sugars.

The addition of enzymes that are capable of degrading hemicellulose has a potential to reduce the need for commercial enzymes during biomass hydrolysis in the production of fermentable sugars. In this study, a high xylanase producing actinomycete strain (Kitasatospora sp. ID06-480) and the first ethyl ferulate producing actinomycete strain (Nonomuraea sp. ID06-094) were selected from 797 rare actinomycetes, respectively, which were isolated in Indonesia. The addition (30%, v/v) of a crude enzyme supernatant from the selected strains in sugarcane bagasse hydrolysis with low-level loading (1 FPU/g-biomass) of Cellic® CTec2 enhanced both the released amount of glucose and reducing sugars. When the reaction with Ctec2 was combined with crude enzymes containing either xylanase or feruloyl esterase, high conversion yield of glucose from cellulose at 60.5% could be achieved after 72 h-saccharification.

Mannan endo-1,4-ß-mannosidase from Kitasatospora sp. isolated in Indonesia and its potential for production of mannooligosaccharides from mannan polymers.

Mannan endo-1,4-ß-mannosidase (commonly known as ß-mannanase) catalyzes a random cleavage of the ß-D-1,4-mannopyranosyl linkage in mannan polymers. The enzyme has been utilized in biofuel production from lignocellulose biomass, as well as in production of mannooligosaccharides (MOS) for applications in feed and food industries. We aimed to obtain a ß-mannanase, for such mannan polymer utilization, from actinomycetes strains isolated in Indonesia. Strains exhibiting high mannanase activity were screened, and one strain belonging to the genus Kitasatospora was selected. We obtained a ß-mannanase from this strain, and an amino acid sequence of this Kitasatospora ß-mannanase showed a 58-71% similarity with the amino acid sequences of Streptomyces ß-mannanases. The Kitasatospora ß-mannanase showed a significant level of activity (944 U/mg) against locust bean gum (0.5% w/v) and a potential for oligosaccharide production from various mannan polymers. The ß-mannanase might be beneficial particularly in the enzymatic production of MOS for applications of mannan utilization.

Halobium palmae gen. nov., sp. nov., an extremely halophilic archaeon isolated from a solar saltern.

A novel and extremely halophilic archaeon, designated strain 2a_47_2T, was isolated from a solar saltern sample collected in Indonesia. Cells of the strain were Gram-stain-negative, non-motile and pleomorphic and formed orange-red pigmented colonies. Strain 2a_47_2T grew at 20-48 °C (optimum 38-41 °C), pH 6.0-8.5 (optimum pH 7.5), >1.7 M NaCl (optimum 2.6 M) and <0.5 M MgCl2 (optimum 0.3 M). The major polar lipids were phosphatidylglycerol, phosphatidylglycerol phosphate methyl ester, two phospholipids and sulfated diglycosyl diether. The cells mainly contained menaquinone-8. The G+C content in the genomic DNA of the strain was 67.0 mol%. Phylogenetic analysis based on 16S rRNA gene sequences indicated that strain 2a_47_2T represents a member of the family Halorubraceae and is different from any other known halophilic archaea. This finding was also demonstrated by phylogenetic analyses based on deduced RpoB' amino acid sequences. Collectively, these results show that strain 2a_47_2T represents a novel genus and species in the family Halorubraceae, and the name Halobium palmae gen. nov., sp. nov. is proposed. The type strain is 2a_47_2T (=NBRC 111368T=InaCC Ar34T).

Kocuria pelophila sp. nov., an actinobacterium isolated from the rhizosphere of a mangrove.

A novel spherical actinobacterium, designated RS-2-3T, was isolated from the rhizosphere of a mangrove growing on Rambut Island, Indonesia, and its taxonomic position was investigated using a polyphasic approach. Phylogenetic analysis based on 16S rRNA gene sequence comparison revealed that strain RS-2-3T was related to the members of the genus Kocuria. The highest 16S rRNA gene sequence similarity value was observed with Kocuria marina KMM 3905T (97.0 %). The peptidoglycan type of strain RS-2-3T was found to be A3α with an interpeptide bridge comprising l-Ala4-5. The predominant menaquinone was MK-7(H2) and the major fatty acids were anteiso-C15 : 0 and iso-C15 : 0. The DNA G+C content was 71.8 mol%. These characteristics were consistent with those of members of the genus Kocuria. Meanwhile, physiological and biochemical characteristics revealed that strain RS-2-3T differed from the species of the genus Kocuria with validly published names. Therefore, strain RS-2-3T represents a novel species of the genus Kocuria, for which the name Kocuria pelophila sp. nov. is proposed. The type strain is RS-2-3T (=NBRC 110990T=InaCC A704T).

Cellulosimicrobium marinum sp. nov., an actinobacterium isolated from sea sediment.

A novel Gram stain positive actinobacterium, designated RS-7-4(T), was isolated from a sea sediment sample collected in Indonesia, and its taxonomic position was investigated using a polyphasic approach. Strain RS-7-4(T) was observed to form vegetative hyphae in the early phase of growth, but the hyphae eventually fragmented into short rods to coccoid cells. Growth occurred at 15-37 °C, pH 6.0-11.0 and in the presence of 0-7 % (w/v) NaCl. Phylogenetic analysis based on 16S rRNA gene sequence comparisons revealed that strain RS-7-4(T) was closely related to the members of the genus Cellulosimicrobium, with a similarity range of 98.08-99.10 %. The peptidoglycan type of strain RS-7-4(T) was found to be A4α L-Lys-L-Thr-D-Asp. The predominant menaquinone was MK-9(H4), and the major fatty acids were anteiso-C15:0, iso-C15:0 and anteiso-C17:0. The DNA G+C content was 75.6 mol%. These chemotaxonomic features corresponded to those of the genus Cellulosimicrobium. Meanwhile, the results of DNA-DNA hybridization, and physiological and biochemical tests revealed that strain RS-7-4(T) was different from the recognized species of the genus Cellulosimicrobium. Therefore, strain RS-7-4(T) represents a novel species of the genus Cellulosimicrobium, for which the name Cellulosimicrobium marinum sp. nov. is proposed. The type strain is RS-7-4(T) (=NBRC 110994(T) =InaCC A726(T)).

Haloarchaeobius baliensis sp. nov., isolated from a solar saltern.

A novel halophilic archaeon, designated strain 2b_61_3T, was isolated from a solar saltern in Indonesia. Cells of the strain were Gram-stain-negative, motile, pleomorphic rods that formed orange-red-pigmented colonies on solid medium. The isolate grew optimally at 42-44 °C, pH 6.5-7.0, and with 2.6 M NaCl, and MgCl2 was required for growth. Strain 2b_61_3T had two differential 16S rRNA genes (rrnA and rrnB), and phylogenetic analysis revealed that the strain belonged to the genus Haloarchaeobius. The rrnA and rrnB sequence similarities between strain 2b_61_3T and species of the genus Haloarchaeobius were 98.4-99.2 % and 98.5-98.8 %, respectively. The findings from the 16S rRNA gene analysis were supported by sequence analysis of rpoB', the B' subunit of RNA polymerase. On the basis of the phenotypic characteristics and phylogenetic analyses, as well as DNA-DNA hybridization experiments with Haloarchaeobius iranensis NBRC 110930T, strain 2b_61_3T represents a novel species of the genus Haloarchaeobius, for which the name Haloarchaeobius baliensis sp. nov. is proposed. The type strain is 2b_61_3T ( = NBRC 110517T = InaCC Ar2T).

Actinoplanes bogoriensis sp. nov., a novel actinomycete isolated from leaf litter.

A novel actinomycete, designated LIPI11-2-Ac043(T), was isolated from leaf litter collected in Indonesia. According to phylogenetic analysis based on the 16S rRNA gene sequence comparisons, strain LIPI11-2-Ac043(T) was closely related to Actinoplanes abujensis A4029(T) (99.3%) and Actinoplanes brasiliensis DSM 43805(T) (98.8%). Spores of strain LIPI11-2-Ac043(T) were motile and the sporangia were spherical. The predominant menaquinone was MK-9(H6) and the principal polar lipids were phosphatidylinositol, phosphatidylethanolamine, phosphatidylglycerol and diphospatidylglycerol. The whole-cell sugars were galactose, glucose and mannose; rhibose, arabinose and xylose were also detected as minor components. The major fatty acids were anteiso-C15:0, iso-C16:0, iso-C15:0 and anteiso-C17:0. These data supported the affiliation of strain LIPI11-2-Ac043(T) to the genus Actinoplanes. Meanwhile, the results of DNA-DNA hybridization and physiological and biochemical tests indicated that strain LIPI11-2-Ac043(T) can be distinguished from its closest related species. Therefore, strain LIPI11-2-Ac043(T) represents a novel species of the genus Actinoplanes, for which the name Actinoplanes bogoriensis sp. nov. is proposed. The type strain is LIPI11-2-Ac043(T) (=InaCC A522(T)=NBRC 110975(T)).

Cryptosporangium cibodasense sp. nov., isolated from leaf litter in Indonesia.

A novel actinomycete strain, designated LIPI11-2-Ac046T, was isolated from a leaf litter sample obtained from Cibodas Botanical Garden, West Java, Indonesia, using the rehydration and centrifugation method. The taxonomic status of this organism was established using a polyphasic approach. Comparative 16S rRNA gene sequence analysis revealed that strain LIPI11-2-Ac046T had the closest sequence similarities with members of the genus Cryptosporangium (97.99-98.90 %). The strain grew well on ISP 4 and ISP 5 media and formed sporangia. Spores of this strain were motile. The strain grew in the presence of 0-2 % (w/v) NaCl and the temperature range of 15-28 8C. The cell-wall hydrolysate contained meso-diaminopimelic acid as the diagnostic diamino acid and the whole-cell hydrolysate contained mannose, glucose, galactose, ribose and xylose, together with one unidentified O-methyl-pentose. The predominant menaquinones were MK-9(H4), MK-9(H6) and MK-9(H8), and the major polar lipid was phosphatidylethanolamine. The major cellular fatty acids were C18 : 1ω9c, iso-C16 : 0, C16 : 0 andC17 : 1ω9c. These phenotypic characteristics corresponded to those of the genus Cryptosporangium. Meanwhile, the results of DNA-DNA hybridization as well as physiological and biochemical analyses distinguished strain LIPI11-2-Ac046T from known members of the genus Cryptosporangium. On the basis of these data, it is proposed that strain LIPI11-2-Ac046T represents a novel species of the genus Cryptosporangium, with the name Cryptosporangium cibodasense sp. nov. The type strain is LIPI11-2-Ac046T (=InaCC A457T=NBRC 110976T).