scholarly journals The 16S rRNA Analysis and Enzyme Screening of Bacillus from Rhizosphere Soil of Lombok Island

2021 ◽  
Vol 26 (4) ◽  
pp. 582-590
Author(s):  
Tri Ratna Sulistiyani ◽  
Mia Kusmiati ◽  
Gita Azizah Putri
Keyword(s):  
16S Rrna ◽  
Enzyme Activities ◽  
Hydrolytic Enzyme ◽  
Hydrolytic Enzymes ◽  
Distinct Species ◽  
Soil Samples ◽  
Rrna Gene ◽  
Bacillus Sp ◽  
16S Rrna Analysis ◽  
Lombok Island

Bacillus are commonly found in nature, especially in soil and food. It has the ability to produce bioactive compounds as well as the enzyme. This study was aimed to isolate, identify, and screen their enzyme activities. Four soil samples from Mandalika, Lombok Island, West Nusa Tenggara (NTB), were used for isolation. Bacillus was isolated using the heat-shock method and characterized through Gram staining, endospore staining, and morphological phenotype. Bacillus identification was conducted based on 16S rRNA gene sequence. The hydrolytic enzyme activities were checked qualitatively using selective media, and the enzyme tested including amylase, galactosidase, lipase, protease, and cellulase. As many as twenty-two bacteria isolates were obtained from four soil samples and represented 15 distinct species. The member of bacteria genera successfully identified, consisted of Bacillus sp., Brevibacillus sp., and Fictibacillus sp. Bacillus sp. was the most isolated. Some of the isolated bacteria have the ability to produce lipase, protease, and cellulase that potential to be used in biotechnology processes.   Keywords: Bacillus, hydrolytic enzymes, identification, screening

scholarly journals Effect of culture conditions on nitrogen-fixing activity of bacteria isolated from cassava cultivated soils of Vietnam

2021 ◽  
Vol 43 (3) ◽  
pp. 27-35
Author(s):  
Pham Viet Cuong ◽  
Nguyen Phuong Hoa
Keyword(s):  
Bacillus Subtilis ◽  
Carbon Source ◽  
16S Rrna ◽  
Culture Conditions ◽  
Rrna Gene ◽  
Atmospheric Nitrogen ◽  
Bacillus Sp ◽  
Nitrogen Fixing ◽  
The 16S Rrna Gene ◽  
Cultivated Soils

The bacteria capable of fixing atmospheric nitrogen were isolated from cassava cultivated soils of Vietnam. The potential isolates were identified by analyzing the 16S rRNA gene and by morphological, biochemical, cultural characteristics. The selected isolates were assigned to the species Bacillus sp. DQT2 M17, Bacillus subtilis DTAN6 M17, and Bacillus megaterium DSHB I8. The effect of culture conditions on the nitrogen-fixing activity of three selected isolates were studied and the obtained results showed that the highest amount of accumulated ammonia was detected after 6 days of incubation at 35 oC, pH 7.0 with sucrose as a carbon source. The selected strains could be exploited as inoculants for microbial fertilizer production.

Soil aggregate-based nucleic acid analyses of the impact of maize roots and their root hairs on the structural diversity of microbial communities

2021 ◽  
Author(s):  
Christoph Tebbe ◽  
Damini Damini ◽  
Damien Finn ◽  
Nataliya Bilyera ◽  
Minh Ganther ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Root Hair ◽  
Soil Aggregates ◽  
Root Hairs ◽  
Soil Samples ◽  
Plant Roots ◽  
Bulk Soil ◽  
Rrna Gene ◽  
The Impact

<p>The deposition of energy rich carbon sources released by plant roots during their growth fuels microbially driven ecosystem processes in soil, but there is a lack of understanding how microorganisms interact and collaborate. The objective of this research was therefore to characterize microbial networks as they assemble under the influence of plant roots. To identify the specific importance of root hairs, we compared the impact of a maize wild-type to a root-air defective mutant (rth3; (1).</p><p>The microbial community structure was analyzed by qPCR and 16S rRNA gene amplicon sequencing from soil DNA. In order to increase the probability of detecting truly interacting microbial partners as a basis for network analyses, we first evaluated a new protocol to obtain DNA from as little as 1 mg instead of the usual 250 mg soil samples, thereby approaching the aggregate level (2). While the diversity of bacterial 16S rRNA gene amplicons of 250-mg samples taken from the same soil was not distinct, DNA analyses from individual aggregates clearly differed from each other underlining that soil aggregates represent distinct microbial habitats.</p><p>Soil column experiments with maize grown in a loam soil (3) revealed distinct communities between rhizosphere and bulk soil. The community composition of individual aggregates showed more differences in bulk soil compared to rhizosphere. Less elaborated networks were seen in bulk soil and a profound effect of root hairs could be unravelled. Null model testing demonstrated that Actinobacteria were equally important for network connectivity independent of the root hair mutation, but for networks of the wildtype, Acidobacteria were essential for synergistic interactions and overall network structure. In contrast, Proteobacteria and Firmicutes connectivity became more important. The observed differences in community composition and interactions suggests carbon cycling, and perhaps other microbially-driven functions, are markedly affected by the presence of root hairs.</p><p>Utilizing maize root soil microcosms for studying soil zymography in the rhizosphere allowed to obtain soil samples from regions with distinct specific enzyme activities. In order to enhance the detection of actively metabolizing bacterial community members, we studied rRNA sequences and compared it to rRNA gene sequences from the same samples. Currently the data are under analysis.</p><p>References</p><p>(1) Wen, T-J, Schnable PS (1994) Analyses of mutants of three genes that influence root hair development in Zea mays (Gramineae) suggest that root hairs are dispensable. Am. J. Bot. 81, 833–842.</p><p>(2) Szoboszlay M, Tebbe CC (2020) Hidden heterogeneity and co-occurrence networks of soil prokaryotic communities revealed at the scale of individual soil aggregates. Microbiol. Open, e1144. DOI: 10.1002/mbo3.1144</p><p>(3) Vetterlein D et al. (2020) Experimental platforms for the investigation of spatiotemporal patterns in the rhizosphere – laboratory and field scale. J. Plant Nutr. Soil Sci., 000, 1–16 DOI: 10.1002/jpln.202000079</p>

scholarly journals Confirmation of Thiomonas delicata (formerly Thiobacillus delicatus) as a distinct species of the genus Thiomonas Moreira and Amils 1997 with comments on some species currently assigned to the genus

2006 ◽  
Vol 56 (11) ◽  
pp. 2553-2557 ◽  
Author(s):  
Yoko Katayama ◽  
Yoshihito Uchino ◽  
Ann P. Wood ◽  
Donovan P. Kelly
Keyword(s):  
16S Rrna ◽  
Dna Hybridization ◽  
Type Strain ◽  
Gene Sequence ◽  
Distinct Species ◽  
Rrna Gene ◽  
Formal Description ◽  
Rrna Gene Sequence ◽  
Physiological Properties ◽  
The Status

The transfer of Thiobacillus delicatus to the genus Thiomonas as a distinct species, Thiomonas delicata (type strain NBRC 14566T), is confirmed by its morphological and physiological properties, DNA–DNA hybridization and the grouping of its 16S rRNA gene sequence with those of other species of the genus. An emended formal description of Thiomonas delicata is given. The status of Thiomonas cuprina DSM 5495T as a member of the genus is reconsidered.

Comparison of 16S ribosomal RNA genes inClavibacter michiganensissubspecies with other coryneform bacteria

1995 ◽  
Vol 41 (10) ◽  
pp. 925-929 ◽  
Author(s):  
Xiang Li ◽  
Solke H. De Boer
Keyword(s):  
16S Rrna ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Rrna Gene ◽  
Clavibacter Michiganensis ◽  
Arthrobacter Simplex ◽  
16S Rrna Analysis ◽  
Coryneform Bacteria ◽  
Clavibacter Michiganensis Subsp ◽  
Complete Sequences

Nearly complete sequences (97–99%) of the 16S rRNA genes were determined for type strains of Clavibacter michiganensis subsp. michiganensis, Clavibacter michiganensis subsp. insidiosus, Clavibacter michiganensis subsp. sepedonicus, and Clavibacter michiganensis subsp. nebraskensis. The four subspecies had less than 1% dissimilarity in their 16S rRNA genes. Comparative studies indicated that the C. michiganensis subsp. shared relatively high homology with the 16S rRNA gene of Clavibacter xyli. Further comparison with representatives of other Gram-positive coryneform and related bacteria with high G + C% values showed that this group of bacteria was subdivided into three clusters. One cluster consisted of the Clavibacter michiganensis subsp., Clavibacter xyli, Arthrobacter globiformis, Arthrobacter simplex, and Frankia sp.; another cluster consisted of members of the corynebacteria–mycobacteria–nocardia (CMN) group of Mycobacteriaceae including Tsukamurella paurometabolum; and Propionibacterium freudenreichii alone formed a unique cluster, which was remote from other coryneform bacteria analyzed. The three clusters may reflect a systematic rank higher than the genus level among these bacteria.Key words: Clavibacter, coryneform bacteria, phylogeny, 16S rRNA analysis.

scholarly journals Paenibacillus Piscarius Sp. Nov., a Novel Nitrogen-Fixing Species Isolated from the Gut of the Armored Catfish Parotocinclus Maculicauda

2021 ◽  
Author(s):  
Mariana Barbalho Farias da silva ◽  
Ericka Arregue Lemos ◽  
Renata E. Vollú ◽  
Fernanda Abreu ◽  
Alexandre S. Rosado ◽  
...  
Keyword(s):  
16S Rrna ◽  
Bacterial Species ◽  
Distinct Species ◽  
Cellular Fatty Acid ◽  
Rrna Gene ◽  
Nitrogen Fixing ◽  
Closely Related Species ◽  
A Genome ◽  
Encoding Gene ◽  
Encoding Genes

Abstract A gram-positive, nitrogen-fixing and endospore-forming strain, designated P121T, was isolated from the gut of the armored catfish (Parotocinclus maculicauda) and identified as a member of the genus Paenibacillus based on the sequences of the 16S rRNA encoding gene, rpoB, gyrB and nifH genes and phenotypic analyses. The most closely related species to strain P121T were Paenibacillus rhizoplanae DSM 103993T, Paenibacillus silagei DSM 101953T and Paenibacillus borealis DSM 13188T, with similarity values of 98.9%, 98.3% and 97.6%, respectively, based on 16S rRNA gene sequences. Genome sequencing revealed a genome size of 7,513,698 bp, DNA G + C content of 53.9 mol% and the presence of the structural nitrogenase encoding genes (nifK, nifD and nifH) necessary for nitrogen fixation. Digital DNA-DNA hybridization (dDDH) experiments and average nucleotide identity (ANI) analyses between strain P121T and the type strains of the closest species demonstrated that the highest values were below the thresholds of 70% dDDH (42.3% with P. borealis) and 95% ANI (84.28% with P. silagei) for bacterial species delineation, indicating that strain P121T represents a distinct species. Its major cellular fatty acid was anteiso-C15:0 (42.4%), and the major isoprenoid quinone was MK-7. Based on physiological, genomic, biochemical and chemotaxonomic characteristics, we propose that strain P121T represents a novel species for which the name Paenibacillus piscarius sp. nov. is proposed (type strain = DSM 25072 = LFB-Fiocruz 1636).

scholarly journals Uncovering a hidden diversity: optimized protocols for the extraction of bacteriophages from soil

2019 ◽  
Author(s):  
Pauline C. Göller ◽  
Jose M. Haro-Moreno ◽  
Francisco Rodriguez-Valera ◽  
Martin J. Loessner ◽  
Elena Gómez-Sanz
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Ecological Model ◽  
Extraction Process ◽  
Extraction Methods ◽  
Soil Samples ◽  
Rrna Gene ◽  
Optimization Strategy ◽  
Data Set ◽  
Soil Matrix

AbstractBackgroundBacteriophages are the most numerous biological entities on earth and play a crucial role in shaping microbial communities. Investigating the bacteriophage community from soil samples will shed light not only on the yet largely unknown phage diversity, but also may result in novel insights into phage biology and functioning. Unfortunately, the study of soil viromes lags far behind any other ecological model system, due to the heterogeneous soil matrix that rises major technical difficulties in the extraction process. Resolving these technical challenges and establishing a standardized extraction protocol is therefore a fundamental prerequisite for replicable results and comparative virome studies.ResultsWe here report the optimization of protocols for extraction of bacteriophage DNA from soil preceding metagenomic analysis such that the protocol can equally be harnessed for phage isolation. As an optimization strategy, soil samples were spiked with a viral community consisting of phages from different families (106 PFU/g soil): Listeria phage ΦA511 (Myovirus), Staphylococcus phage Φ2638AΔLCR (Siphovirus), and Escherichia phage ΦT7 (Podovirus). The efficacy of bacteriophage (i) elution, (ii) filtration, (iii) concentration, and (iv) DNA extraction methods was tested. Successful extraction routes were selected based on spiked phage recovery and low bacterial 16S rRNA gene contaminants. Natural agricultural soil viromes were then extracted with the optimized methods and shotgun sequenced. Our approach yielded sufficient amounts of inhibitor-free viral DNA for non-amplification dependent sequencing and low 16S rRNA gene contamination levels (≤ 0.2 ‰). Compared to previously published protocols, the number of bacterial read contamination was decreased by 65 %. In addition, 468 novel circularized soil phage genomes in size up to 235 kb were obtained from over 29,000 manually identified viral contigs, promising the discovery of a large, previously inaccessible viral diversity.ConclusionWe have shown a dramatically enhanced extraction of the soil phage community by protocol optimization that has proven robustness in both a culture-depended as well as through metaviromic analysis. Our huge data set of manually curated soil viral contigs roughly doubles the amount of currently available soil virome data, and provide insights into the yet largely undescribed soil viral sequence space.

scholarly journals Molecular cloning gene and nucleotide sequence of the gene encoding an endo-1,4-beta-glucanase from Bacillus sp VLSH08 strain applying to biomass hydrolysis

2012 ◽  
Vol 3 (2) ◽  
pp. 80-86
Author(s):  
Minh Thi Tuyet Phan ◽  
Viet Quoc Nguyen ◽  
Hy Gia Le ◽  
Thoa Kim Nguyen ◽  
Man Dinh Tran
Keyword(s):  
Escherichia Coli ◽  
Bacillus Subtilis ◽  
Amino Acid ◽  
16S Rrna ◽  
Cellulase Activity ◽  
Rrna Gene ◽  
Bacillus Sp ◽  
Biomass Hydrolysis ◽  
Gene Encoding ◽  
Beta Glucanase

Bacillus sp VLSH08 screened from sea wetland in Nam Dinh province produces an extracellular endo-1,4-beta-glucanase. According to the results of the classified Kit API 50/CHB as well as sequence of 1500 bp fragment coding for 16S rRNA gene of the Bacillus sp VLSH 08 strain showed that the taxonomical characteristics between the strain VLSH 08 and Bacillus amyloliquefaciene JN999857 are similar of 98%. Culture supernatant of this strain showed optimal cellulase activity at pH 5.8 and 60°C and that was enhanced 2.03 times in the presence of 5 mM Co2+. Moreover, the gene encoding endo-1,4-beta-glucanase from this strain was cloned in Escherichia coli using pCR2.1 vector. The entire gene for the enzyme contained a 1500-bp single open reading frame encoding 500 amino acids, including a 29-amino acid signal peptide. The amino acid sequence of this enzyme is very close to that of an EG of Bacillus subtilis (EU022560.1) and an EG of Bacillus amyloliquefaciene (EU022559.1) which all belong to the cellulase family E2. A cocktail of enzyme containing this endo-1,4-beta-glucanase used for biomass hydrolysis indicated that the cellulose conversion attained to 72.76% cellulose after 48 hours. Chủng vi khuẩn Bacillus sp VLSH08 được tuyển chọn từ tập hợp chủng vi khuẩn phân lập ở vùng ngập mặn tỉnh Nam Định có khả năng sinh tổng hợp enzyme endo-1,4-beta-glucanase ngoại bào. Kết quả phân loại chủng vi khuẩn Bacillus sp VLSH08 bằng Kit hóa sinh API 50/CHB cũng như trình tự gen mã hóa 16S rRNA cho thấy độ tương đồng của chủng Bacillus sp VLSH08 và chủng Bacillus amyloliquefaciene JN999857 đạt 98%. Dịch lên men của chủng được sử dụng làm nguồn enzyme thô để nghiên cứu hoạt độ tối ưu của enzyme ở pH 5,8 và nhiệt đô 60°C. Hoạt tính enzyme tăng 2,03 lần khi có mặt 5 mM ion Co2+. Đồng thời, gen mã hóa cho enzyme endo-1,4-beta-glucanase cũng được tách dòng trong tế bào Escherichia coli sử dụng vector pCR 2.1. Gen mã hóa cho enzyme này có chiều dài 1500 bp, mã hóa cho 500 axit amin, bao gồm 29 axit amin của chuỗi peptid tín hiệu. So sánh cho thấy trình tự gen endo-1,4-beta-glucanase của chủng Bacillus sp VLSH08 có độ tương đồng cao với enzyme này của chủng Bacillus subtilis (EU022560.1) và của chủng Bacillus amyloliquefaciene (EU022559.1). Tất cả các enzyme nhóm này đều thuộc họ cellulase E2. Enzyme của chủng này cũng đã được phối trộn với các enzyme khác tạo thành cocktail để thủy phân sinh khối cho kết quả cellulose bị thủy phân 72,76% sau 48 giờ.

scholarly journals Interpreting Proteobacteria diversity through 16S rRNA analysis in Manasbal Lake, Kashmir

2021 ◽  
Vol 1 (01) ◽  
pp. 7-16
Author(s):  
Sana Shafi ◽  
Suhaib A. Bandh ◽  
Nowsheen Shameem
Keyword(s):  
16S Rrna ◽  
Biochemical Characterization ◽  
Bacterial Species ◽  
Bacterial Community Composition ◽  
Open Water ◽  
Freshwater Ecosystem ◽  
Rrna Gene ◽  
16S Rrna Analysis ◽  
Manasbal Lake ◽  
Spatio Temporal

Bacterial community composition in aquatic ecosystems is of renewed interest for its health as well as the people living in the adjoining areas, and the current work on a freshwater ecosystem in Kashmir Himalayas–the Manasbal Lake was carried out in this backdrop. Water samples were collected from various sampling stations, selected as the zones of special ecological interest and according to the degree of difference in the anthropogenic intrusions in the lake. For identification, the isolated bacterial colonies were subjected to morphological and biochemical characterization, which were further confirmed by targeting their 16S rRNA gene, using 27F and 1492R as the universal bacterial primers. Major bacterial phylum, thus isolated, was proteobacteria with 15 different bacterial species belonging to class alpha-proteobacteria, beta-proteobacteria, and gammaproteobacteria. However, the most diverse class isolated was Alphaproteobacteria comprising seven species followed by Betaproteobacteria comprising six species and Gamma-proteobacteria comprising only two species. The distribution of the bacterial group was seen influenced on a Spatio-temporal scale with the maximum density observed during summer and minimum during winter, further the load was highest for littoral sampling stations in comparison to the open water sites. Proteobacteria are important since they perform basic functions in global transformations of elements. But at the same time, they show close interaction with eukaryotes, both as pathogens and as symbionts.

scholarly journals Quantification of Hyphomicrobium Populations in Activated Sludge from an Industrial Wastewater Treatment System as Determined by 16S rRNA Analysis

2000 ◽  
Vol 66 (3) ◽  
pp. 1167-1174 ◽  
Author(s):  
A. C. Layton ◽  
P. N. Karanth ◽  
C. A. Lajoie ◽  
A. J. Meyers ◽  
I. R. Gregory ◽  
...  
Keyword(s):  
Wastewater Treatment ◽  
16S Rrna ◽  
Activated Sludge ◽  
16S Rdna ◽  
Industrial Wastewater ◽  
Bacterial Species ◽  
Treatment Plant ◽  
Rrna Gene ◽  
16S Rrna Analysis ◽  
Industrial Wastewater Treatment

ABSTRACT The bacterial community structure of the activated sludge from a 25 million-gal-per-day industrial wastewater treatment plant was investigated using rRNA analysis. 16S ribosomal DNA (rDNA) libraries were created from three sludge samples taken on different dates. Partial rRNA gene sequences were obtained for 46 rDNA clones, and nearly complete 16S rRNA sequences were obtained for 18 clones. Seventeen of these clones were members of the beta subdivision, and their sequences showed high homology to sequences of known bacterial species as well as published 16S rDNA sequences from other activated sludge sources. Sixteen clones belonged to the alpha subdivision, 7 of which showed similarity to Hyphomicrobium species. This cluster was chosen for further studies due to earlier work onHyphomicrobium sp. strain M3 isolated from this treatment plant. A nearly full-length 16S rDNA sequence was obtained fromHyphomicrobium sp. strain M3. Phylogenetic analysis revealed that Hyphomicrobium sp. strain M3 was 99% similar to Hyphomicrobium denitrificans DSM 1869T inHyphomicrobium cluster II. Three of the cloned sequences from the activated sludge samples also grouped with those ofHyphomicrobium cluster II, with a 96% sequence similarity to that of Hyphomicrobium sp. strain M3. The other four cloned sequences from the activated sludge sample were more closely related to those of the Hyphomicrobium cluster I organisms (95 to 97% similarity). Whole-cell fluorescence hybridization of microorganisms in the activated sludge with genus-specificHyphomicrobium probe S-G-Hypho-1241-a-A-19 enhanced the visualization of Hyphomicrobium and revealed thatHyphomicrobium appears to be abundant both on the outside of flocs and within the floc structure. Dot blot hybridization of activated sludge samples from 1995 with probes designed forHyphomicrobium cluster I and Hyphomicrobiumcluster II indicated that Hyphomicrobium cluster II-positive 16S rRNA dominated over Hyphomicrobium cluster I-positive 16S rRNA by 3- to 12-fold. Hyphomicrobium 16S rRNA comprised approximately 5% of the 16S rRNA in the activated sludge.

scholarly journals Diversity and nitrogen fixation efficiency of rhizobia isolated from nodules of Centrolobium paraense

2014 ◽  
Vol 49 (4) ◽  
pp. 296-305 ◽  
Author(s):  
Alexandre Cardoso Baraúna ◽  
Krisle da Silva ◽  
Gilmara Maria Duarte Pereira ◽  
Paulo Emílio Kaminski ◽  
Liamara Perin ◽  
...  
Keyword(s):  
Nitrogen Fixation ◽  
16S Rrna ◽  
Vigna Unguiculata ◽  
Gene Sequencing ◽  
Rpob Gene ◽  
Soil Samples ◽  
Symbiotic Efficiency ◽  
16S Rrna Analysis ◽  
Rhizobium Species ◽  
Nitrogen Treatment

The objective of this work was to isolate and characterize rhizobia from nodules of Centrolobium paraense and to evaluate their symbiotic efficiency. Soil samples collected from four sites of the Roraima Cerrado, Brazil, were used to cultivate C. paraense in order to obtain nodules. Isolates (178) were obtained from 334 nodules after cultivation on medium 79. Twenty-five isolates belonging to six morphological groups were authenticated using Vigna unguiculata and they were characterized by 16S rRNA. Isolates identified as Bradyrhizobium were further characterized using rpoB gene sequencing. A greenhouse experiment was carried out with C. paraense to test the 18 authenticated isolates. Approximately 90% of the isolates grew slowly in medium 79. The 16S rRNA analysis showed that 14 authenticated isolates belong to the genus Bradyrhizobium, and rpoB indicated they constitute different groups compared to previously described species. Only four of the 11 fast-growing isolates nodulated V. unguiculata, two of which belong to Rhizobium, and two to Pleomorphomonas, which was not previously reported as a nodulating genus. The Bradyrhizobium isolates ERR 326, ERR 399, and ERR 435 had the highest symbiotic efficiency on C. paraense and showed a contribution similar to the nitrogen treatment. Centrolobium paraense is able to nodulate with different rhizobium species, some of which have not yet been described.

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