Halophilic bacteria of salt lakes and saline soils of the Peri-Caspian lowland (Republic of Daghestan) and their biotechnological potential

The article presents the results of studying the biodiversity and biotechnological potential of halophilic microorganisms from the thermal highly mineralized Berikey Lake, the salty Lake Tarumovskoye and saline soils of the Peri-Caspian Lowland (Republic of Daghestan). Denitrifying halophilic bacteria of the genus Halomonas and Virgibacillus were identified using microbiological methods and 16S rRNA gene analysis. A new species Halomonas sp. G2 (MW386470) with a similarity of the nucleotide sequences of the 16S rRNA genes is 95 %. Strain G2 is an extreme halophile capable of growing in the range of 5–25 % NaCl (optimum 25 %) and forming a carotenoid pigment. Mesophil, 30–37 °С (optimum 30 °С); neutrophil, pH 6–8 (optimum 7.2–7.4). Strain G2 chemolithotroph; reduces nitrate or nitrite as electron donors; catalase-, amylase-, protease- and β-galactosidase-positive; lipase-, oxidase- and urease-negative. Not able to hydrolyze inositol, indole; produces lysine, gelatin, ectoine; uses citrate and sodium malate as a source of carbon and energy; does not produce ornitin, H2S or acid from d-mannose, sucrose, glycerol, cellobiose, except for lactose and d-glucose. Susceptible to trimethoprim, ciprofloxacin, ofloxacin, kanamycin, vancomycin, rifampicin, cefuroxime, ampicillin, ceftazidime, fosfomycin, clarithromycin, cefepime, cefaclor. The G+C content in DNA is 67.3 %. A distinctive characteristic of the isolate was the production of industrially significant hydrolytic enzymes such as amylase, protease, β-galactosidase, and oxidoreductase (catalase) at a NaCl concentration of 25 % in the medium. Habitat: saline soils on the territory of the Tersko-Kumskaya lowland (Republic of Daghestan, Russia). The rest of the halophilic isolates of H. ventosae G1 (MW386469), H. elongata G3 (MW386471), V. salinarius B2 (MW386472), and V. salinarius B3 (MW386473) had a high degree of similarity (100 %) with the type strains of H. elongata DSM 2581Т and V. salarius DSM 18441Т; the content of G+C in DNA was 65.8, 66.5, 42.8 and 37.3 %, respectively. The strains had a high biotechnological potential at NaCl concentrations of 5 and 25 % in the medium. The data obtained expanded the understanding of the diversity and ecological significance of denitrifying bacteria in the functioning of arid ecosystems and make it possible to identify strains producing enzymes of industrial importance.

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Polyphase Taxonomy of Antarctic Bacteria

Mikrobiolohichnyi Zhurnal ◽

10.15407/microbiolj83.03.003 ◽

2021 ◽

Vol 83 (3) ◽

pp. 3-13

Author(s):

G.V. Gladka ◽

◽

N.V. Borzova ◽

O.V. Gudzenko ◽

V.M. Hovorukha ◽

...

Keyword(s):

16S Rrna ◽

Related Species ◽

Nucleotide Sequences ◽

16S Rrna Genes ◽

Rrna Genes ◽

Phenotypic Properties ◽

Bootstrap Analysis ◽

Closely Related Species ◽

Microbiological Methods ◽

The Antarctic

The phylogenetic structure of bacteria is not always consistent with the traditional classification scheme based on the phenotypic properties of bacteria. That is one of the problems of modern bacterial taxonomy. In addition, traditional methods to identify bacteria using phenotypic properties have a number of disadvantages. In recent decades, significant progress was achieved in the investigation of microbial world using molecular methods for fast identification. The aim of the study was to clarify the species status of four strains of bacteria isolated from black lichens of the cliffs of Galindez Island in the Antarctic on the basis of phenotypic and genetic analysis. Methods. Morphological and cultural properties of bacteria were studied according to generally accepted microbiological methods. Physiological and biochemical ones were investigated using test systems API Coryne and API 20E (bioMerieux SA, France), according to the manufacturer’s instructions. Phylogenetic analysis was performed based on the nucleotide sequences of the 16S rRNA gene. To identify closely related species, a comparative analysis of the nucleotide sequences of 16S rRNA genes was performed using the BLAST software package. The phylogenetic position was determined by constructing trees (dendrograms) to show the position of the studied strains among closely related and typical species (programs ClustalX 2.1, Mega 6.06). The tree was constructed applying ClustalX 2.1 by comparing the nearest neighbors with bootstrap analysis (bootstrap NJ tree) using 1000 bootstrap tests (1000 alternative trees). Then the phylogenetic tree was edited by the program Mega v. 6.00. Results. Based on the results of comparative, phylogenetic and phenotypic analysis, the studied Antarctic strains 180n1, 181n2, 188n2, 190n2 were identified as Pseudomonas fluorescens, Microbacterium foliorum, Sporosarcina aquimarina and Rothia sp., respectively. The coefficient of similarity of 16S rRNA genes of strain 180n1 with such a closely related species from the database P. fluorescens NBRC 14160 was 99.5%; 181n2 with M. foliorum P 333/02 – 99.4%; 188n2 with S. aquimarina SW28 – 99.7%. These strains form common clusters with closely related species on phylogenetic dendrograms. The strain 190n2 can be considered as Rothia sp., since has the remote position from closely related strains in the cluster Rothia and a low percentage of similarity (97.3%) with the species Rothia endophytica YIM 67072. These strains belong to the phyla: Firmicutes, Actinobacteria, Proteobacteria. Conclusions. Phylogenetic and phenotypic analyzes allowed determining the taxonomic position of isolated aerobic chemoorganotrophic microbial strains of the Antarctic. Nucleotide sequences of 16S rRNA genes are deposited in the International GenBank database under numbers HG518622, HG518623, HG518625, HG518626.

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

Brazilian Journal of Biology ◽

10.1590/1519-6984.246038 ◽

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.

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Differentiation Of Clarias Batrachus, C. Gariepinus And Heteropneustes Fossilis By Pcr-Sequencing Of Mitochondrial 16s Rrna Gene

Journal of the Asiatic Society of Bangladesh Science ◽

10.3329/jasbs.v41i1.46190 ◽

2015 ◽

Vol 41 (1) ◽

pp. 51-58

Author(s):

Mohammad Shamimul Alam ◽

Hawa Jahan ◽

Rowshan Ara Begum ◽

Reza M Shahjahan

Keyword(s):

16S Rrna ◽

16S Rrna Gene ◽

Fish Larva ◽

16S Rrna Genes ◽

Rrna Genes ◽

Rrna Gene ◽

Valuable Insight ◽

Multiple Sequence ◽

Pcr Rflp ◽

Alternative Means

Heteropneustesfossilis, Clariasbatrachus and C. gariepinus are three major catfishes ofecological and economic importance. Identification of these fish species becomes aproblem when the usual external morphological features of the fish are lost or removed,such as in canned fish. Also, newly hatched fish larva is often difficult to identify. PCRsequencingprovides accurate alternative means of identification of individuals at specieslevel. So, 16S rRNA genes of three locally collected catfishes were sequenced after PCRamplification and compared with the same gene sequences available from othergeographical regions. Multiple sequence alignment of the 16S rRNA gene fragments ofthe catfish species has revealed polymorphic sites which can be used to differentiate thesethree species from one another and will provide valuable insight in choosing appropriaterestriction enzymes for PCR-RFLP based identification in future. Asiat. Soc. Bangladesh, Sci. 41(1): 51-58, June 2015

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

Genes ◽

10.3390/genes12010040 ◽

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.

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Ultra-accurate microbial amplicon sequencing with synthetic long reads

Microbiome ◽

10.1186/s40168-021-01072-3 ◽

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.

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Microdiversity and phylogeographic diversification of bacterioplankton in pelagic freshwater systems revealed through long-read amplicon sequencing

Microbiome ◽

10.1186/s40168-020-00974-y ◽

2021 ◽

Vol 9 (1) ◽

Author(s):

Yusuke Okazaki ◽

Shohei Fujinaga ◽

Michaela M. Salcher ◽

Cristiana Callieri ◽

Atsushi Tanaka ◽

...

Keyword(s):

16S Rrna ◽

Regional Scale ◽

Scale Up ◽

Amplicon Sequencing ◽

Freshwater Ecosystems ◽

16S Rrna Genes ◽

Rrna Genes ◽

Rrna Gene ◽

Metagenomic Sequencing ◽

Long Read

Abstract Background Freshwater ecosystems are inhabited by members of cosmopolitan bacterioplankton lineages despite the disconnected nature of these habitats. The lineages are delineated based on > 97% 16S rRNA gene sequence similarity, but their intra-lineage microdiversity and phylogeography, which are key to understanding the eco-evolutional processes behind their ubiquity, remain unresolved. Here, we applied long-read amplicon sequencing targeting nearly full-length 16S rRNA genes and the adjacent ribosomal internal transcribed spacer sequences to reveal the intra-lineage diversities of pelagic bacterioplankton assemblages in 11 deep freshwater lakes in Japan and Europe. Results Our single nucleotide-resolved analysis, which was validated using shotgun metagenomic sequencing, uncovered 7–101 amplicon sequence variants for each of the 11 predominant bacterial lineages and demonstrated sympatric, allopatric, and temporal microdiversities that could not be resolved through conventional approaches. Clusters of samples with similar intra-lineage population compositions were identified, which consistently supported genetic isolation between Japan and Europe. At a regional scale (up to hundreds of kilometers), dispersal between lakes was unlikely to be a limiting factor, and environmental factors or genetic drift were potential determinants of population composition. The extent of microdiversification varied among lineages, suggesting that highly diversified lineages (e.g., Iluma-A2 and acI-A1) achieve their ubiquity by containing a consortium of genotypes specific to each habitat, while less diversified lineages (e.g., CL500-11) may be ubiquitous due to a small number of widespread genotypes. The lowest extent of intra-lineage diversification was observed among the dominant hypolimnion-specific lineage (CL500-11), suggesting that their dispersal among lakes is not limited despite the hypolimnion being a more isolated habitat than the epilimnion. Conclusions Our novel approach complemented the limited resolution of short-read amplicon sequencing and limited sensitivity of the metagenome assembly-based approach, and highlighted the complex ecological processes underlying the ubiquity of freshwater bacterioplankton lineages. To fully exploit the performance of the method, its relatively low read throughput is the major bottleneck to be overcome in the future.

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Microbiological and Geochemical Heterogeneity in an In Situ Uranium Bioremediation Field Site

Applied and Environmental Microbiology ◽

10.1128/aem.71.10.6308-6318.2005 ◽

2005 ◽

Vol 71 (10) ◽

pp. 6308-6318 ◽

Cited By ~ 172

Author(s):

Helen A. Vrionis ◽

Robert T. Anderson ◽

Irene Ortiz-Bernad ◽

Kathleen R. O'Neill ◽

Charles T. Resch ◽

...

Keyword(s):

16S Rrna ◽

16S Rrna Gene ◽

16S Rrna Genes ◽

Rrna Genes ◽

Acid Volatile Sulfide ◽

Rrna Gene ◽

Gene Sequences ◽

16S Rrna Gene Sequences ◽

Geochemical Heterogeneity

ABSTRACT The geochemistry and microbiology of a uranium-contaminated subsurface environment that had undergone two seasons of acetate addition to stimulate microbial U(VI) reduction was examined. There were distinct horizontal and vertical geochemical gradients that could be attributed in large part to the manner in which acetate was distributed in the aquifer, with more reduction of Fe(III) and sulfate occurring at greater depths and closer to the point of acetate injection. Clone libraries of 16S rRNA genes derived from sediments and groundwater indicated an enrichment of sulfate-reducing bacteria in the order Desulfobacterales in sediment and groundwater samples. These samples were collected nearest the injection gallery where microbially reducible Fe(III) oxides were highly depleted, groundwater sulfate concentrations were low, and increases in acid volatile sulfide were observed in the sediment. Further down-gradient, metal-reducing conditions were present as indicated by intermediate Fe(II)/Fe(total) ratios, lower acid volatile sulfide values, and increased abundance of 16S rRNA gene sequences belonging to the dissimilatory Fe(III)- and U(VI)-reducing family Geobacteraceae. Maximal Fe(III) and U(VI) reduction correlated with maximal recovery of Geobacteraceae 16S rRNA gene sequences in both groundwater and sediment; however, the sites at which these maxima occurred were spatially separated within the aquifer. The substantial microbial and geochemical heterogeneity at this site demonstrates that attempts should be made to deliver acetate in a more uniform manner and that closely spaced sampling intervals, horizontally and vertically, in both sediment and groundwater are necessary in order to obtain a more in-depth understanding of microbial processes and the relative contribution of attached and planktonic populations to in situ uranium bioremediation.

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