scholarly journals Diversity, Dynamics, and Activity of Bacterial Communities during Production of an Artisanal Sicilian Cheese as Evaluated by 16S rRNA Analysis

2002 ◽  
Vol 68 (4) ◽  
pp. 1882-1892 ◽  
Author(s):  
Cinzia L. Randazzo ◽  
Sandra Torriani ◽  
Antoon D. L. Akkermans ◽  
Willem M. de Vos ◽  
Elaine E. Vaughan
Keyword(s):  
16S Rrna ◽  
16S Rdna ◽  
Starter Culture ◽  
Lactobacillus Delbrueckii ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Phenotypic Characterization ◽  
Rrna Gene ◽  
Rt Pcr ◽  
16S Rrna Analysis

ABSTRACT The diversity and dynamics of the microbial communities during the manufacturing of Ragusano cheese, an artisanal cheese produced in Sicily (Italy), were investigated by a combination of classical and culture-independent approaches. The latter included PCR, reverse transcriptase-PCR (RT-PCR), and denaturing gradient gel electrophoresis (DGGE) of 16S rRNA genes (rDNA). Bacterial and Lactobacillus group-specific primers were used to amplify the V6 to V8 and V1 to V3 regions of the 16S rRNA gene, respectively. DGGE profiles from samples taken during cheese production indicated dramatic shifts in the microbial community structure. Cloning and sequencing of rDNA amplicons revealed that mesophilic lactic acid bacteria (LAB), including species of Leuconostoc, Lactococcus lactis, and Macrococcus caseolyticus were dominant in the raw milk, while Streptococcus thermophilus prevailed during lactic fermentation. Other thermophilic LAB, especially Lactobacillus delbrueckii and Lactobacillus fermentum, also flourished during ripening. Comparison of the rRNA-derived patterns obtained by RT-PCR to the rDNA DGGE patterns indicated a substantially different degree of metabolic activity for the microbial groups detected. Identification of cultivated LAB isolates by phenotypic characterization and 16S rDNA analysis indicated a variety of species, reflecting to a large extent the results obtained from the 16S rDNA clone libraries, with the significant exception of the Lactobacillus delbrueckii species, which dominated in the ripening cheese but was not detected by cultivation. The present molecular approaches combined with culture can effectively describe the complex ecosystem of natural fermented dairy products, giving useful information for starter culture design and preservation of artisanal fermented food technology.

Molecular methods for identification of Lactobacillus delbrueckii subsp. bulgaricus and Streptococcus thermophilus using methionine biosynthesis and 16S rRNA genes

2008 ◽  
Vol 75 (4) ◽  
pp. 392-398 ◽  
Author(s):  
Aysun Cebeci ◽  
G Candan Gürakan
Keyword(s):  
16S Rrna ◽  
Restriction Analysis ◽  
Starter Culture ◽  
Streptococcus Thermophilus ◽  
Lactobacillus Delbrueckii ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Rrna Gene ◽  
Methionine Biosynthesis ◽  
Difficult Time

Yoghurt and starter culture producers are still searching strains of Lactobacillus delbrueckii subsp. bulgaricus and Streptococcus thermophilus to produce healthier yogurt with longer shelf life, better texture, taste and quality. However, selective identification of Lb. delbrueckii subsp. bulgaricus and Strep. thermophilus from a mixed population using microbiological and biochemical methods is difficult, time consuming and may not be accurate. In this study, a quick, sensitive and accurate method is proposed to identify both Lb. delbrueckii subsp. bulgaricus and Strep. thermophilus using PCR. The method is comprised of two parts. In the first part, methionine biosynthesis genes, known to be present in both species were partially amplified by designed primers (cysmet2F and cysmet2R). Partial amplification of the methionine biosynthesis gene which gives 700 bp fragment resulted in selective identification of Lb. bulgaricus and Strep. thermophilus. All 16 Lb. bulgaricus and 6 Strep. thermophilus isolates assessed by this method gave the expected amplification. On the other hand, further analysis of other closely related species with the same primers have indicated that the same product was also amplified in two more lactobacilli namely, Lb. delbrueckii subsp. lactis and Lb. helveticus species. Thus, in the second part of the method, further differentiation of Lb. delbrueckii subsp. bulgaricus and Strep. thermophilus from each other and these species was achieved using restriction analysis of 16S rRNA gene with EcoRI.

scholarly journals rDNA analyses of planktonic heterocystous cyanobacteria, including members of the genera Anabaenopsis and Cyanospira The GenBank accession numbers of the 16S rDNA gene sequences reported in this paper are AY038032–AY038037.

Microbiology ◽  
2002 ◽  
Vol 148 (2) ◽  
pp. 481-496 ◽  
Author(s):  
Isabelle Iteman ◽  
Rosmarie Rippka ◽  
Nicole Tandeau de Marsac ◽  
Michael Herdman
Keyword(s):  
16S Rrna ◽  
16S Rdna ◽  
Genomic Sequence ◽  
Sequence Similarity ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
23S Rrna ◽  
Phylogenetic Position ◽  
Rrna Gene ◽  
Heterocystous Cyanobacteria

The taxonomic coherence and phylogenetic relationships of 11 planktonic heterocystous cyanobacterial isolates were examined by investigating two areas of the rRNA operon, the 16S rRNA gene (rrnS) and the internal transcribed spacer (ITS) located between the 16S rRNA and 23S rRNA genes. The rrnS sequences were determined for five strains, including representatives of Anabaena flos-aquae, Aphanizomenon flos-aquae, Nodularia sp. and two alkaliphilic planktonic members of the genera Anabaenopsis and Cyanospira, whose phylogenetic position was previously unknown. Comparison of the data with those previously published for individual groups of planktonic heterocystous cyanobacteria showed that, with the exception of members assigned to the genus Cylindrospermopsis, all the planktonic strains form a distinct subclade within the monophyletic clade of heterocystous cyanobacteria. Within this subclade five different phylogenetic clusters were distinguished. The phylogenetic groupings of Anabaena and Aphanizomenon strains within three of these clusters were not always consistent with their generic or specific assignments based on classical morphological definitions, and the high degree of sequence similarity between strains of Anabaenopsis and Cyanospira suggests that they may be assignable to a single genus. Ribotyping and additional studies performed on PCR amplicons of the 16S rDNA or the ITS for the 11 planktonic heterocystous strains demonstrated that they all contain multiple rrn operons and ITS regions of variable size. Finally, evidence is provided for intra-genomic sequence heterogeneity of the 16S rRNA genes within most of the individual isolates.

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.

Phylogenetic Differentiation of Two Closely RelatedNitrosomonas spp. That Inhabit Different Sediment Environments in an Oligotrophic Freshwater Lake

1999 ◽  
Vol 65 (11) ◽  
pp. 4855-4862 ◽  
Author(s):  
Corinne B. Whitby ◽  
Jon R. Saunders ◽  
Juana Rodriguez ◽  
Roger W. Pickup ◽  
Alan McCarthy
Keyword(s):  
16S Rrna ◽  
16S Rdna ◽  
Freshwater Lake ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Rrna Gene ◽  
Ammonia Oxidizing Bacteria ◽  
Relative Distribution ◽  
Sediment Samples ◽  
Oligonucleotide Hybridization

ABSTRACT The population of ammonia-oxidizing bacteria in a temperate oligotrophic freshwater lake was analyzed by recovering 16S ribosomal DNA (rDNA) from lakewater and sediment samples taken throughout a seasonal cycle. Nitrosospira and Nitrosomonas16S rRNA genes were amplified in a nested PCR, and the identity of the products was confirmed by oligonucleotide hybridization.Nitrosospira DNA was readily identified in all samples, and nitrosomonad DNA of the Nitrosomonas europaea-Nitrosomonas eutropha lineage was also directly detected, but during the summer months only. Phylogenetic delineation with partial (345 bp) 16S rRNA gene sequences of clones obtained from sediments confirmed the fidelity of the amplified nitrosomonad DNA and identified two sequence clusters closely related to either N. europaea or N. eutropha that were equated with the littoral and profundal sediment sites, respectively. Determination of 701-bp sequences for 16S rDNA clones representing each cluster confirmed this delineation. A PCR-restriction fragment length polymorphism (RFLP) system was developed that enabled identification of clones containing N. europaea and N. eutropha 16S rDNA sequences, including subclasses therein. It proved possible to analyze 16S rDNA amplified directly from sediment samples to determine the relative abundance of each species compared with that of the other. N. europaea and N. eutropha are very closely related, and direct evidence for their presence in lake systems is limited. The correlation of each species with a distinct spatial location in sediment is an unusual example of niche adaptation by two genotypically similar bacteria. Their occurrence and relative distribution can now be routinely monitored in relation to environmental variation by the application of PCR-RFLP analysis.

scholarly journals Metagenomic Analysis of Slovak Bryndza Cheese Using Next-Generation 16S rDNA Amplicon Sequencing

2016 ◽  
Vol 15 (1) ◽  
pp. 23-34 ◽  
Author(s):  
Matej Planý ◽  
Tomáš Kuchta ◽  
Katarína Šoltýs ◽  
Tomáš Szemes ◽  
Domenico Pangallo ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rdna ◽  
Lactococcus Lactis ◽  
Microbial Population ◽  
Starter Culture ◽  
Amplicon Sequencing ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Next Generation ◽  
16S Rdna Amplicon Sequencing

Abstract Knowledge about diversity and taxonomic structure of the microbial population present in traditional fermented foods plays a key role in starter culture selection, safety improvement and quality enhancement of the end product. Aim of this study was to investigate microbial consortia composition in Slovak bryndza cheese. For this purpose, we used culture-independent approach based on 16S rDNA amplicon sequencing using next generation sequencing platform. Results obtained by the analysis of three commercial (produced on industrial scale in winter season) and one traditional (artisanal, most valued, produced in May) Slovak bryndza cheese sample were compared. A diverse prokaryotic microflora composed mostly of the genera Lactococcus, Streptococcus, Lactobacillus, and Enterococcus was identified. Lactococcus lactis subsp. lactis and Lactococcus lactis subsp. cremoris were the dominant taxons in all tested samples. Second most abundant species, detected in all bryndza cheeses, were Lactococcus fujiensis and Lactococcus taiwanensis, independently by two different approaches, using different reference 16S rRNA genes databases (Greengenes and NCBI respectively). They have been detected in bryndza cheese samples in substantial amount for the first time. The narrowest microbial diversity was observed in a sample made with a starter culture from pasteurised milk. Metagenomic analysis by high-throughput sequencing using 16S rRNA genes seems to be a powerful tool for studying the structure of the microbial population in cheeses.

scholarly journals Assessment of Microbial Diversity in Four Southwestern United States Soils by 16S rRNA Gene Terminal Restriction Fragment Analysis

2000 ◽  
Vol 66 (7) ◽  
pp. 2943-2950 ◽  
Author(s):  
John Dunbar ◽  
Lawrence O. Ticknor ◽  
Cheryl R. Kuske
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
16S Rdna ◽  
Restriction Fragment ◽  
Community Diversity ◽  
Detection Sensitivity ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Rrna Gene ◽  
Clone Libraries

ABSTRACT The ability of terminal restriction fragment (T-RFLP or TRF) profiles of 16S rRNA genes to provide useful information about the relative diversity of complex microbial communities was investigated by comparison with other methods. Four soil communities representing two pinyon rhizosphere and two between-tree (interspace) soil environments were compared by analysis of 16S rRNA gene clone libraries and culture collections (Dunbar et al., Appl. Environ. Microbiol. 65:1662–1669, 1998) and by analysis of 16S rDNA TRF profiles of community DNA. The TRF method was able to differentiate the four communities in a manner consistent with previous comparisons of the communities by analysis of 16S rDNA clone libraries. TRF profiles were not useful for calculating and comparing traditional community richness or evenness values among the four soil environments. Statistics calculated from RsaI, HhaI, HaeIII, and MspI profiles of each community were inconsistent, and the combined data were not significantly different between samples. The detection sensitivity of the method was tested. In standard PCRs, a seeded population comprising 0.1 to 1% of the total community could be detected. The combined results demonstrate that TRF analysis is an excellent method for rapidly comparing the relationships between bacterial communities in environmental samples. However, for highly complex communities, the method appears unable to provide classical measures of relative community diversity.

Effect of Phenylurea Herbicides on Soil Microbial Communities Estimated by Analysis of 16S rRNA Gene Fingerprints and Community-Level Physiological Profiles

1999 ◽  
Vol 65 (3) ◽  
pp. 982-988 ◽  
Author(s):  
Saïd el Fantroussi ◽  
Laurent Verschuere ◽  
Willy Verstraete ◽  
Eva M. Top
Keyword(s):  
16S Rrna ◽  
Microbial Communities ◽  
16S Rdna ◽  
Soil Microbial Communities ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Rrna Gene ◽  
Metabolic Potential ◽  
Soil Microbial ◽  
Enrichment Cultures

ABSTRACT The effect of three phenyl urea herbicides (diuron, linuron, and chlorotoluron) on soil microbial communities was studied by using soil samples with a 10-year history of treatment. Denaturing gradient gel electrophoresis (DGGE) was used for the analysis of 16S rRNA genes (16S rDNA). The degree of similarity between the 16S rDNA profiles of the communities was quantified by numerically analysing the DGGE band patterns. Similarity dendrograms showed that the microbial community structures of the herbicide-treated and nontreated soils were significantly different. Moreover, the bacterial diversity seemed to decrease in soils treated with urea herbicides, and sequence determination of several DGGE fragments showed that the most affected species in the soils treated with diuron and linuron belonged to an uncultivated bacterial group. As well as the 16S rDNA fingerprints, the substrate utilization patterns of the microbial communities were compared. Principal-component analysis performed on BIOLOG data showed that the functional abilities of the soil microbial communities were altered by the application of the herbicides. In addition, enrichment cultures of the different soils in medium with the urea herbicides as the sole carbon and nitrogen source showed that there was no difference between treated and nontreated soil in the rate of transformation of diuron and chlorotoluron but that there was a strong difference in the case of linuron. In the enrichment cultures with linuron-treated soil, linuron disappeared completely after 1 week whereas no significant transformation was observed in cultures inoculated with nontreated soil even after 4 weeks. In conclusion, this study showed that both the structure and metabolic potential of soil microbial communities were clearly affected by a long-term application of urea herbicides.

scholarly journals Serpentinimonas gen. nov., Serpentinimonas raichei sp. nov., Serpentinimonas barnesii sp. nov. and Serpentinimonas maccroryi sp. nov., hyperalkaliphilic and facultative autotrophic bacteria isolated from terrestrial serpentinizing springs

2021 ◽  
Vol 71 (8) ◽  
Author(s):  
Lina J. Bird ◽  
J. Gijs Kuenen ◽  
Magdalena R. Osburn ◽  
Naotaka Tomioka ◽  
Shun’ichi Ishii ◽  
...  
Keyword(s):  
16S Rrna ◽  
Type Strain ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Phenotypic Characterization ◽  
Phylogenomic Analysis ◽  
Rrna Gene ◽  
Content Type ◽  
Link Type ◽  
The Family

Three highly alkaliphilic bacterial strains designated as A1T, H1T and B1T were isolated from two highly alkaline springs at The Cedars, a terrestrial serpentinizing site. Cells from all strains were motile, Gram-negative and rod-shaped. Strains A1T, H1T and B1T were mesophilic (optimum, 30 °C), highly alkaliphilic (optimum, pH 11) and facultatively autotrophic. Major cellular fatty acids were saturated and monounsaturated hexadecenoic and octadecanoic acids. The genome size of strains A1T, H1T and B1T was 2 574 013, 2 475 906 and 2 623 236 bp, and the G+C content was 66.0, 66.2 and 66.1 mol%, respectively. Analysis of the 16S rRNA genes showed the highest similarity to the genera Malikia (95.1–96.4 %), Macromonas (93.0–93.6 %) and Hydrogenophaga (93.0–96.6 %) in the family Comamonadaceae . Phylogenetic analysis based on 16S rRNA gene and phylogenomic analysis based on core gene sequences revealed that the isolated strains diverged from the related species, forming a distinct branch. Average amino acid identity values of strains A1T, H1T and B1T against the genomes of related members in this family were below 67 %, which is below the suggested threshold for genera boundaries. Average nucleotide identity by blast values and digital DNA–DNA hybridization among the three strains were below 92.0 and 46.6 % respectively, which are below the suggested thresholds for species boundaries. Based on phylogenetic, genomic and phenotypic characterization, we propose Serpentinimonas gen. nov., Serpentinimonas raichei sp. nov. (type strain A1T=NBRC 111848T=DSM 103917T), Serpentinimonas barnesii sp. nov. (type strain H1T= NBRC 111849T=DSM 103920T) and Serpentinimonas maccroryi sp. nov. (type strain B1T=NBRC 111850T=DSM 103919T) belonging to the family Comamonadaceae . We have designated Serpentinimonas raichei the type species for the genus because it is the dominant species in The Cedars springs.

scholarly journals Delineation of the genus Actinobacillus by comparison of partial infB sequences

2004 ◽  
Vol 54 (3) ◽  
pp. 635-644 ◽  
Author(s):  
Niels Nørskov-Lauritsen ◽  
Henrik Christensen ◽  
Henrik Okkels ◽  
Mogens Kilian ◽  
Brita Bruun
Keyword(s):  
16S Rrna ◽  
Housekeeping Gene ◽  
Translation Initiation Factor ◽  
16S Rrna Genes ◽  
Initiation Factor ◽  
Rrna Genes ◽  
Rrna Gene ◽  
16S Rrna Sequence ◽  
Sequence Comparisons ◽  
16S Rrna Analysis

A 426 bp fragment of infB, a housekeeping gene that encodes translation initiation factor 2, was sequenced from 59 clinical isolates and type strains of Actinobacillus species and sequences were compared. Partial sequences of 16S rRNA genes were also obtained. By comparing infB sequences, Actinobacillus pleuropneumoniae, Actinobacillus equuli, Actinobacillus suis, Actinobacillus ureae, Actinobacillus arthritidis, Actinobacillus hominis and two unnamed genomospecies showed more than 85 % similarity to the type strain of the type species of the genus, Actinobacillus lignieresii. The taxonomic position of Actinobacillus capsulatus was unresolved; this species is more remotely related to A. lignieresii. The two species A. lignieresii and A. pleuropneumoniae could not be clearly separated by infB sequence analysis. The phylogeny of the genus Actinobacillus based on infB analysis was essentially congruent with relationships inferred from 16S rRNA sequence comparisons and DNA hybridization studies. Discrepancies were encountered with single strains or taxa at the periphery of the genus. Greater intraspecies variation was observed with infB sequences than with 16S rRNA gene sequences, with notable exceptions. The apparent subdivision of some species by 16S rRNA analysis was most likely caused by RNA operon heterogeneity.

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

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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