scholarly journals Identification by molecular techniques of halophilic bacteria producing important enzymes from pristine area in Campeche, Mexico

2023 ◽  
Vol 83 ◽  
Author(s):  
L. A. Can-Herrera ◽  
C. D. Gutierrez-Canul ◽  
M. A. A. Dzul-Cervantes ◽  
O. F. Pacheco-Salazar ◽  
J. D. Chi-Cortez ◽  
...  
Keyword(s):  
Bacillus Subtilis ◽  
Microbial Community ◽  
16S Rrna ◽  
Relative Abundance ◽  
Halophilic Bacteria ◽  
Molecular Techniques ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Pristine Area ◽  
Blast Program

Abstract Isla Arena is located in the coordinate 20° 70´ N - 90° 45´ W, from Campeche, Mexico. In these estuaries, the ocean mixes with fresh water, and ecosystems are concentrated where petenes and pink flamingos proliferate. Crustaceans and mollusks abound in the sea. Despite its enormous marine wealth, there are no studies carried out on which halophilic microorganisms are present in these waters. In this work, the diversity and structure of the microbial community was investigated through a metagenomics approach and corroborated for sequencing of 16S rRNA genes. It was found that the phylum Fimicutes predominates with more than 50%, in almost the same proportion of the class Bacilli and with almost 41% of relative abundance of the order Bacillales. The sequencing results showed that one of the samples presented a high percentage of similarity (99.75%) using the Nucleotide BLAST program with a peculiar microorganism: Bacillus subtilis. This microorganism is one of the best characterized bacteria among the gram-positive ones. Our results demonstrate that B. subtilis can be an efficient source of proteases, lipases and cellulases, from halophilic microbial communities located in poorly explored areas.

scholarly journals An inter-laboratory study to investigate the impact of the bioinformatics component on microbiome analysis using mock communities

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Denise M. O’Sullivan ◽  
Ronan M. Doyle ◽  
Sasithon Temisak ◽  
Nicholas Redshaw ◽  
Alexandra S. Whale ◽  
...  
Keyword(s):  
Microbial Community ◽  
16S Rrna ◽  
16S Rrna Gene ◽  
Relative Abundance ◽  
Amplicon Sequencing ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Rrna Gene ◽  
Microbiome Analysis ◽  
The Impact

AbstractDespite the advent of whole genome metagenomics, targeted approaches (such as 16S rRNA gene amplicon sequencing) continue to be valuable for determining the microbial composition of samples. Amplicon microbiome sequencing can be performed on clinical samples from a normally sterile site to determine the aetiology of an infection (usually single pathogen identification) or samples from more complex niches such as human mucosa or environmental samples where multiple microorganisms need to be identified. The methodologies are frequently applied to determine both presence of micro-organisms and their quantity or relative abundance. There are a number of technical steps required to perform microbial community profiling, many of which may have appreciable precision and bias that impacts final results. In order for these methods to be applied with the greatest accuracy, comparative studies across different laboratories are warranted. In this study we explored the impact of the bioinformatic approaches taken in different laboratories on microbiome assessment using 16S rRNA gene amplicon sequencing results. Data were generated from two mock microbial community samples which were amplified using primer sets spanning five different variable regions of 16S rRNA genes. The PCR-sequencing analysis included three technical repeats of the process to determine the repeatability of their methods. Thirteen laboratories participated in the study, and each analysed the same FASTQ files using their choice of pipeline. This study captured the methods used and the resulting sequence annotation and relative abundance output from bioinformatic analyses. Results were compared to digital PCR assessment of the absolute abundance of each target representing each organism in the mock microbial community samples and also to analyses of shotgun metagenome sequence data. This ring trial demonstrates that the choice of bioinformatic analysis pipeline alone can result in different estimations of the composition of the microbiome when using 16S rRNA gene amplicon sequencing data. The study observed differences in terms of both presence and abundance of organisms and provides a resource for ensuring reproducible pipeline development and application. The observed differences were especially prevalent when using custom databases and applying high stringency operational taxonomic unit (OTU) cut-off limits. In order to apply sequencing approaches with greater accuracy, the impact of different analytical steps needs to be clearly delineated and solutions devised to harmonise microbiome analysis results.

scholarly journals Effects of Supplement of Marichromatium gracile YL28 on Water Quality and Microbial Structures in Shrimp Mariculture Ecosystems

Genes ◽  
2020 ◽  
Vol 12 (1) ◽  
pp. 40
Author(s):  
Liang Cui ◽  
Bitong Zhu ◽  
Xiaobo Zhang ◽  
Zhuhua Chan ◽  
Chungui Zhao ◽  
...  
Keyword(s):  
Water Quality ◽  
16S Rrna ◽  
Relative Abundance ◽  
Animal Health ◽  
Denitrifying Bacteria ◽  
Nitrite Reduction ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Sequencing Analysis ◽  
Shrimp Culture

The elevated NH3-N and NO2-N pollution problems in mariculture have raised concerns because they pose threats to animal health and coastal and offshore environments. Supplement of Marichromatium gracile YL28 (YL28) into polluted shrimp rearing water and sediment significantly decreased ammonia and nitrite concentrations, showing that YL28 functioned as a novel safe marine probiotic in the shrimp culture industry. The diversity of aquatic bacteria in the shrimp mariculture ecosystems was studied by sequencing the V4 region of 16S rRNA genes, with respect to additions of YL28 at the low and high concentrations. It was revealed by 16S rRNA sequencing analysis that Proteobacteria, Planctomycete and Bacteroidetes dominated the community (>80% of operational taxonomic units (OTUs)). Up to 41.6% of the predominant bacterial members were placed in the classes Gammaproteobacteria (14%), Deltaproteobacteria (14%), Planctomycetacia (8%) and Alphaproteobacteria (5.6%) while 40% of OTUs belonged to unclassified ones or others, indicating that the considerable bacterial populations were novel in our shrimp mariculture. Bacterial communities were similar between YL28 supplements and control groups (without addition of YL28) revealed by the β-diversity using PCoA, demonstrating that the additions of YL28 did not disturb the microbiota in shrimp mariculture ecosystems. Instead, the addition of YL28 increased the relative abundance of ammonia-oxidizing and denitrifying bacteria. The quantitative PCR analysis further showed that key genes including nifH and amoA involved in nitrification and nitrate or nitrite reduction significantly increased with YL28 supplementation (p < 0.05). The supplement of YL28 decreased the relative abundance of potential pathogen Vibrio. Together, our studies showed that supplement of YL28 improved the water quality by increasing the relative abundance of ammonia-oxidizing and denitrifying bacteria while the microbial community structure persisted in shrimp mariculture ecosystems.

scholarly journals Ultra-accurate microbial amplicon sequencing with synthetic long reads

Microbiome ◽  
2021 ◽  
Vol 9 (1) ◽  
Author(s):  
Benjamin J. Callahan ◽  
Dmitry Grinevich ◽  
Siddhartha Thakur ◽  
Michael A. Balamotis ◽  
Tuval Ben Yehezkel
Keyword(s):  
Microbial Community ◽  
16S Rrna ◽  
Amplicon Sequencing ◽  
Species Level ◽  
Full Length ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Strain Identification ◽  
Long Reads ◽  
Long Read

Abstract Background Out of the many pathogenic bacterial species that are known, only a fraction are readily identifiable directly from a complex microbial community using standard next generation DNA sequencing. Long-read sequencing offers the potential to identify a wider range of species and to differentiate between strains within a species, but attaining sufficient accuracy in complex metagenomes remains a challenge. Methods Here, we describe and analytically validate LoopSeq, a commercially available synthetic long-read (SLR) sequencing technology that generates highly accurate long reads from standard short reads. Results LoopSeq reads are sufficiently long and accurate to identify microbial genes and species directly from complex samples. LoopSeq perfectly recovered the full diversity of 16S rRNA genes from known strains in a synthetic microbial community. Full-length LoopSeq reads had a per-base error rate of 0.005%, which exceeds the accuracy reported for other long-read sequencing technologies. 18S-ITS and genomic sequencing of fungal and bacterial isolates confirmed that LoopSeq sequencing maintains that accuracy for reads up to 6 kb in length. LoopSeq full-length 16S rRNA reads could accurately classify organisms down to the species level in rinsate from retail meat samples, and could differentiate strains within species identified by the CDC as potential foodborne pathogens. Conclusions The order-of-magnitude improvement in length and accuracy over standard Illumina amplicon sequencing achieved with LoopSeq enables accurate species-level and strain identification from complex- to low-biomass microbiome samples. The ability to generate accurate and long microbiome sequencing reads using standard short read sequencers will accelerate the building of quality microbial sequence databases and removes a significant hurdle on the path to precision microbial genomics.

scholarly journals 1358. A Novel Rapidly Growing Mycobacteria (RGM) Species Causing Soft Tissue and Orthopedic Hardware Infection After Trauma

2019 ◽  
Vol 6 (Supplement_2) ◽  
pp. S492-S492
Author(s):  
David C Nguyen ◽  
Michelle Lisgaris ◽  
Sruthi Vasireddy ◽  
Richard J Wallace ◽  
Federico Perez ◽  
...  
Keyword(s):  
Soft Tissue ◽  
16S Rrna ◽  
Antibiotic Susceptibility ◽  
Molecular Techniques ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Rrna Gene ◽  
Mycobacterium Fortuitum ◽  
Rapidly Growing Mycobacteria ◽  
Maldi Tof

Abstract Background The widespread use of molecular techniques has resulted in increasing numbers of newly characterized rapidly growing mycobacteria (RGM). Many RGM cause soft tissue and orthopedic hardware infection, particularly after trauma. RGM species identification remains challenging with few genetic differences between species. Methods We describe a case involving RGM. We report results of matrix-assisted laser desorption/ionization time of flight (MALDI-TOF) mass spectrometry (Bruker Biotyper), sequencing of rpoB, erm(39), and 16S rRNA genes, and antibiotic susceptibility testing (AST). We review previous reports describing similar RGM infections. Results A 58-year-old male sustained multiple fractures and right thigh compartment syndrome after a motorcycle accident. He underwent fasciotomy and multi-stage surgical fixations. 3 months later, he had wound dehiscence, purulence and multiple fluid collections of his right leg and knee requiring surgical drainage and removal of orthopedic hardware. After 4 days, acid-fast bacilli grew on routine bacterial culture media. MALDI-TOF identified the isolate as Mycobacterium mageritense. In contrast, sequencing of 16S rRNA (100% identity) and erm(39) (> 99% identity) identified the isolate as Mycobacterium houstonense; erm(39) only had 80% similarity with Mycobacterium fortuitum. Sequencing of rpoB showed a 19 bp difference with the M. houstonense type strain, and showed similarity to M. fortuitum (97.64%) than M. houstonense (97.45%). AST demonstrated resistance to clarithromycin only. After initial treatment with imipenem, ciprofloxacin, and doxycycline, definite therapy with ciprofloxacin and doxycycline was successful. In the literature, we found one case each of M. mageritense and M. houstonense infection after trauma. Conclusion This case highlights the importance of RGM other than M. fortuitum as a cause of soft tissue and orthopedic hardware infections, and illustrates the difficulty of identifying them to the species level. Sequencing of erm(39) and 16S rRNA gene identified the isolate as M. houstonense, but the larger difference (>2.5%) in rpoB sequence suggests a novel species. Further characterization is underway. Efforts to determine RGM species and antibiotic susceptibility give important insight into diagnosis and management. Disclosures All authors: No reported disclosures.

scholarly journals Bacterial Diversity and Function of Aerobic Granules Engineered in a Sequencing Batch Reactor for Phenol Degradation

2004 ◽  
Vol 70 (11) ◽  
pp. 6767-6775 ◽  
Author(s):  
He-Long Jiang ◽  
Joo-Hwa Tay ◽  
Abdul Majid Maszenan ◽  
Stephen Tiong-Lee Tay
Keyword(s):  
Microbial Community ◽  
16S Rrna ◽  
Bacterial Diversity ◽  
Sequencing Batch Reactor ◽  
Batch Reactor ◽  
Phenol Degradation ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Aerobic Granules ◽  
And Function

ABSTRACT Aerobic granules are self-immobilized aggregates of microorganisms and represent a relatively new form of cell immobilization developed for biological wastewater treatment. In this study, both culture-based and culture-independent techniques were used to investigate the bacterial diversity and function in aerobic phenol- degrading granules cultivated in a sequencing batch reactor. Denaturing gradient gel electrophoresis (DGGE) analysis of PCR-amplified 16S rRNA genes demonstrated a major shift in the microbial community as the seed sludge developed into granules. Culture isolation and DGGE assays confirmed the dominance of β-Proteobacteria and high-G+C gram-positive bacteria in the phenol-degrading aerobic granules. Of the 10 phenol-degrading bacterial strains isolated from the granules, strains PG-01, PG-02, and PG-08 possessed 16S rRNA gene sequences that matched the partial sequences of dominant bands in the DGGE fingerprint belonging to the aerobic granules. The numerical dominance of strain PG-01 was confirmed by isolation, DGGE, and in situ hybridization with a strain-specific probe, and key physiological traits possessed by PG-01 that allowed it to outcompete and dominate other microorganisms within the granules were then identified. This strain could be regarded as a functionally dominant strain and may have contributed significantly to phenol degradation in the granules. On the other hand, strain PG-08 had low specific growth rate and low phenol degradation ability but showed a high propensity to autoaggregate. By analyzing the roles played by these two isolates within the aerobic granules, a functional model of the microbial community within the aerobic granules was proposed. This model has important implications for rationalizing the engineering of ecological systems.

Identification of prevalent microbial communities in a municipal solid waste landfill

2006 ◽  
Vol 53 (8) ◽  
pp. 139-147 ◽  
Author(s):  
B. Calli ◽  
S. Durmaz ◽  
B. Mertoglu
Keyword(s):  
16S Rrna ◽  
Municipal Solid Waste ◽  
Microbial Communities ◽  
Solid Waste ◽  
Molecular Techniques ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Fish Analysis ◽  
Municipal Solid Waste Landfill ◽  
Waste Landfill

To identify the microbial communities in Istanbul, Odayeri Municipal Solid Waste Landfill, leachate samples were collected from different sections at different stabilization phases. In identification of microbial communities in leachate samples, molecular techniques such as FISH, DGGE and cloning based on 16S rRNA and mcrA genes were used. As the chemical and microbiological compositions of the samples were compared, obvious correlations were found between the stability of the landfill section and abundance of active methanogens. On the other hand, there were considerable differences between acidogenic and mature leachate samples in DGGE profiles of archaeal and bacterial 16S rRNA genes. Moreover, in acidogenic leachate samples having BOD5/COD ratio of about 0.5 acetate utilizing Methanosarcina and Methanosaeta species were intensively detected in FISH. Although only very few H2-utilizing methanogens were identified with FISH analysis, most of the clones isolated from mature leachate samples clustered within H2-utilizing Methanobacteriales and Methanomicrobiales according to phylogenetic analysis of 16S rRNA and mcrA clones, respectively.

scholarly journals Comparison of Microbial Community Compositions of Two Subglacial Environments Reveals a Possible Role for Microbes in Chemical Weathering Processes

2005 ◽  
Vol 71 (11) ◽  
pp. 6986-6997 ◽  
Author(s):  
Mark Skidmore ◽  
Suzanne P. Anderson ◽  
Martin Sharp ◽  
Julia Foght ◽  
Brian D. Lanoil
Keyword(s):  
Microbial Community ◽  
16S Rrna ◽  
Microbial Communities ◽  
Blot Hybridization ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Solute Flux ◽  
Carbonate Dissolution ◽  
Dot Blot ◽  
Dot Blot Hybridization

ABSTRACT Viable microbes have been detected beneath several geographically distant glaciers underlain by different lithologies, but comparisons of their microbial communities have not previously been made. This study compared the microbial community compositions of samples from two glaciers overlying differing bedrock. Bulk meltwater chemistry indicates that sulfide oxidation and carbonate dissolution account for 90% of the solute flux from Bench Glacier, Alaska, whereas gypsum/anhydrite and carbonate dissolution accounts for the majority of the flux from John Evans Glacier, Ellesmere Island, Nunavut, Canada. The microbial communities were examined using two techniques: clone libraries and dot blot hybridization of 16S rRNA genes. Two hundred twenty-seven clones containing amplified 16S rRNA genes were prepared from subglacial samples, and the gene sequences were analyzed phylogenetically. Although some phylogenetic groups, including the Betaproteobacteria, were abundant in clone libraries from both glaciers, other well-represented groups were found at only one glacier. Group-specific oligonucleotide probes were developed for two phylogenetic clusters that were of particular interest because of their abundance or inferred biochemical capabilities. These probes were used in quantitative dot blot hybridization assays with a range of samples from the two glaciers. In addition to shared phyla at both glaciers, each glacier also harbored a subglacial microbial population that correlated with the observed aqueous geochemistry. These results are consistent with the hypothesis that microbial activity is an important contributor to the solute flux from glaciers.

scholarly journals Different Active Microbial Communities in Two Contrasted Subantarctic Fjords

2021 ◽  
Vol 12 ◽  
Author(s):  
Claudia Maturana-Martínez ◽  
Camila Fernández ◽  
Humberto E. González ◽  
Pierre E. Galand
Keyword(s):  
Climate Change ◽  
Microbial Community ◽  
16S Rrna ◽  
Microbial Communities ◽  
Standing Stock ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Biogeochemical Processes ◽  
Biogeographic Patterns ◽  
Group I

Microorganisms play a crucial role in biogeochemical processes affecting the primary production and biogeochemical cycles of the ocean. In subpolar areas, the increment of the water temperature induced by climate change could lead to changes in the structure and activity of planktonic microbial communities. To understand how the structure of the microbial community in Chilean Patagonian fjords could be affected by climate change, we analyzed the composition of the prokaryotic community (bacteria-archaea) in two fjords (Pia and Yendegaia) with contrasting morphological and hydrological features. We targeted both the standing stock (16S rRNA genes) and the active fraction (16S rRNA transcripts) of the microbial communities during two consecutive austral winters. Our results showed that in both fjords, the active community had higher diversity and stronger biogeographic patterns when compared to the standing stock. Members of the Alpha-, Gamma-, and Deltaproteobacteria followed by archaea from the Marine Group I (Thaumarchaeota) dominated the active communities in both fjords. However, in Pia fjord, which has a marine-terminating glacier, the composition of the microbial community was directly influenced by the freshwater discharges from the adjacent glacier, and indirectly by a possible upwelling phenomenon that could bring deep sea bacteria such as SAR202 to the surface layer. In turn, in the Yendegaia, which has a land-terminating glacier, microbial communities were more similar to the ones described in oceanic waters. Furthermore, in Yendegaia fjord, inter-annual differences in the taxonomic composition and diversity of the microbial community were observed. In conclusion, Yendegaia fjord, without glacier calving, represents a fjord type that will likely be more common under future climate scenarios. Our results showing distinct Yendegaia communities, with for example more potential nitrogen-fixing microorganisms (Planctomycetes), indicate that as a result of climate change, changing planktonic communities could potentially impact biogeochemical processes and nutrient sources in subantarctic fjords.

scholarly journals A Comparative Study: Taxonomic Grouping of Alkaline Protease Producing Bacilli

2017 ◽  
Vol 66 (1) ◽  
pp. 39-56
Author(s):  
Nilgun Tekin ◽  
Arzu Coleri Cihan ◽  
Basar Karaca ◽  
Cumhur Cokmus
Keyword(s):  
16S Rrna ◽  
Alkaline Protease ◽  
Phylogenetic Analyses ◽  
Molecular Techniques ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Protease Production ◽  
Rrna Gene ◽  
Wide Range ◽  
Its Pcr

Alkaline proteases have biotechnological importance due to their activity and stability at alkaline pH. 56 bacteria, capable of growing under alkaline conditions were isolated and their alkaline protease activities were carried out at different parameters to determine their optimum alkaline protease production conditions. Seven isolates were showed higher alkaline protease production capacity than the reference strains. The highest alkaline protease producing isolates (103125 U/g), E114 and C265, were identified as Bacillus licheniformis with 99.4% and Bacillus mojavensis 99.8% based on 16S rRNA gene sequence similarities, respectively. Interestingly, the isolates identified as Bacillus safensis were also found to be high alkaline protease producing strains. Genotypic characterizations of the isolates were also determined by using a wide range of molecular techniques (ARDRA, ITS-PCR, (GTG)5-PCR, BOX-PCR). These different techniques allowed us to differentiate the alkaliphilic isolates and the results were in concurrence with phylogenetic analyses of the 16S rRNA genes. While ITS-PCR provided the highest correlation with 16S rRNA groups, (GTG)5-PCR showed the highest differentiation at species and intra-species level. In this study, each of the biotechnologically valuable alkaline protease producing isolates was grouped into their taxonomic positions with multi-genotypic analyses.

Sign in / Sign up

Export Citation Format

Share Document