scholarly journals A qPCR method for distinguishing biomass from non-axenic terrestrial cyanobacteria cultures in hetero- or mixotrophic cultivations

2020 ◽  
Vol 32 (6) ◽  
pp. 3767-3774 ◽  
Author(s):  
Jakob Walther ◽  
Anna Schwarz ◽  
Marco Witthohn ◽  
Dorina Strieth ◽  
Kai Muffler ◽  
...  
Keyword(s):  
Carbon Source ◽  
Heterotrophic Bacteria ◽  
Quantitative Polymerase Chain Reaction ◽  
Nostoc Muscorum ◽  
Rrna Gene ◽  
Slow Growth Rate ◽  
Terrestrial Cyanobacteria ◽  
Heterotrophic Cultivation ◽  
Biomass Partition

AbstractThe cultivation of cyanobacteria with the addition of an organic carbon source (meaning as heterotrophic or mixotrophic cultivation) is a promising technique to increase their slow growth rate. However, most cyanobacteria cultures are infected by non-separable heterotrophic bacteria. While their contribution to the biomass is rather insignificant in a phototrophic cultivation, problems may arise in heterotrophic and mixotrophic mode. Heterotrophic bacteria can potentially utilize carbohydrates quickly, thus preventing any benefit for the cyanobacteria. In order to estimate the advantage of the supplementation of a carbon source, it is essential to quantify the proportion of cyanobacteria and heterotrophic bacteria in the resulting biomass. In this work, the use of quantitative polymerase chain reaction (qPCR) is proposed. To prepare the samples, a DNA extraction method for cyanobacteria was improved to provide reproducible and robust results for the group of terrestrial cyanobacteria. Two pairs of primers were used, which bind either to the 16S rRNA gene of all cyanobacteria or all bacteria including cyanobacteria. This allows a determination of the proportion of cyanobacteria in the biomass. The method was established with the two terrestrial cyanobacteria Trichocoleus sociatus SAG 26.92 and Nostoc muscorum SAG B-1453-12a. As proof of concept, a heterotrophic cultivation with T. sociatus with glucose was performed. After 2 days of cultivation, a reduction of the biomass partition of the cyanobacterium to 90% was detected. Afterwards, the proportion increased again.

scholarly journals Characterization and Transcriptome Analysis of a Long-Chain n-Alkane-Degrading Strain Acinetobacter Pittii SW-1

2021 ◽  
Vol 18 (12) ◽  
pp. 6365
Author(s):  
Weina Kong ◽  
Cheng Zhao ◽  
Xingwang Gao ◽  
Liping Wang ◽  
Qianqian Tian ◽  
...  
Keyword(s):  
Unsaturated Fatty Acids ◽  
Sulfur Metabolism ◽  
Quantitative Polymerase Chain Reaction ◽  
Deep Ocean ◽  
Rrna Gene ◽  
Alkane Hydroxylase ◽  
Degradation Rates ◽  
Acinetobacter Pittii ◽  
Ocean Environment ◽  
Long Chain

Strain sw-1, isolated from 7619-m seawater of the Mariana Trench, was identified as Acinetobacter pittii by 16S rRNA gene and whole-genome sequencing. A. pittii sw-1 was able to efficiently utilize long-chain n-alkanes (C18–C36), but not short- and medium-chain n-alkanes (C8–C16). The degradation rate of C20 was 91.25%, followed by C18, C22, C24, C32, and C36 with the degradation rates of 89.30%, 84.03%, 80.29%, 30.29%, and 13.37%, respectively. To investigate the degradation mechanisms of n-alkanes for this strain, the genome and the transcriptome analyses were performed. Four key alkane hydroxylase genes (alkB, almA, ladA1, and ladA2) were identified in the genome. Transcriptomes of strain sw-1 grown in C20 or CH3COONa (NaAc) as the sole carbon source were compared. The transcriptional levels of alkB and almA, respectively, increased 78.28- and 3.51-fold in C20 compared with NaAc, while ladA1 and ladA2 did not show obvious change. The expression levels of other genes involved in the synthesis of unsaturated fatty acids, permeases, membrane proteins, and sulfur metabolism were also upregulated, and they might be involved in n-alkane uptake. Reverse transcription quantitative polymerase chain reaction (RT-qPCR) confirmed that alkB expression was significantly induced by C20, C24, and C32, and almA induction extent by C24 and C32 was higher than that with C20. Furthermore, ladA2 expression was only induced by C32, and ladA1 expression was not induced by any of n-alkanes. In addition, A. pittii sw-1 could grow with 0%–3% NaCl or 8 out of 10 kinds of the tested heavy metals and degrade n-alkanes at 15 °C. Taken together, these results provide comprehensive insights into the degradation of long-chain n-alkanes by Acinetobacter isolated from the deep ocean environment.

scholarly journals Antioxidant or Apoptosis Inhibitor Supplementation in Culture Media Improves Post-Thaw Recovery of Murine Spermatogonial Stem Cells

Antioxidants ◽  
2021 ◽  
Vol 10 (5) ◽  
pp. 754
Author(s):  
Sang-Eun Jung ◽  
Hui-Jo Oh ◽  
Jin-Seop Ahn ◽  
Yong-Hee Kim ◽  
Bang-Jin Kim ◽  
...  
Keyword(s):  
Stem Cells ◽  
Functional Activity ◽  
Culture Media ◽  
Quantitative Polymerase Chain Reaction ◽  
Spermatogonial Stem Cells ◽  
Ros Generation ◽  
Apoptosis Assay ◽  
Polymerase Chain ◽  
Apoptosis Inhibitor

We postulated that supplementation of antioxidant or apoptosis inhibitor in post-thaw culture media of spermatogonial stem cells (SSCs) alleviates reactive oxygen species (ROS) generation and apoptosis. Our aim was to develop an effective culture media for improving post-thaw recovery of SSCs. To determine the efficacy of supplementation with hypotaurine (HTU), α-tocopherol (α-TCP), and Z-DEVD-FMK (ZDF), we assessed the relative proliferation rate and SSC functional activity and performed a ROS generation assay, apoptosis assay, and western blotting for determination of the Bax/Bcl-xL ratio, as well as immunocytochemistry and real-time quantitative polymerase chain reaction (RT-qPCR) for SSC characterization. The relative proliferation rates with HTU 400 μM (133.7 ± 3.2%), α-TCP 400 μM (158.9 ± 3.6%), and ZDF 200 μM (133.1 ± 7.6%) supplementation were higher than that in the DMSO control (100 ± 3.6%). ROS generation was reduced with α-TCP 400 μM (0.8-fold) supplementation in comparison with the control (1.0-fold). Early apoptosis and Bax/Bcl-xL were lower with α-TCP 400 μM (2.4 ± 0.4% and 0.5-fold) and ZDF 200 μM (1.8 ± 0.4% and 0.3-fold) supplementation in comparison with the control (5.3 ± 1.4% and 1.0-fold) with normal characterization and functional activity. Supplementation of post-thaw culture media with α-TCP 400 μM and ZDF 200 μM improved post-thaw recovery of frozen SSCs via protection from ROS generation and apoptosis after cryo-thawing.

scholarly journals Determination of important enzymes and antimicrobial resistances of gram-positive haloalkaliphilic bacteria isolated from Salda Lake

2021 ◽  
Vol 38 (3) ◽  
pp. 375-382
Author(s):  
Pınar Çağlayan
Keyword(s):  
Bacillus Subtilis ◽  
Soda Lakes ◽  
Soda Lake ◽  
16S Rrna Gene Sequencing ◽  
Extreme Environment ◽  
Rrna Gene ◽  
Sequencing Analysis ◽  
Haloalkaliphilic Bacteria ◽  
Rrna Gene Sequencing

As an extreme environment, soda lakes harbor various haloalkaliphilic microorganisms. Salda Lake is one of the natural soda lake (pH˃9) in Turkey. Haloalkaliphiles are unique microorganisms in their ability to live in high alkaline and high saline conditions, and play an important role in biodegradation and bioremediation of hydrocarbons. Hence, the aims of this study were to isolate haloalkaliphilic bacteria from water sample of Salda Lake, to identify these isolates by both conventional and molecular methods, to screen their industrially important enzymes, and to investigate their antimicrobial resistance profiles. Six isolates were identified as Bacillus horneckiae, Bacillus subtilis, Bacillus paramycoides, Bacillus pumilus, Staphylococcus epidermidis, Bacillus haynesii according to 16S rRNA gene sequencing analysis. The industrially important enzymes (amylase, cellulase, pullulanase, lipase, urease, protease, caseinase, oxidase, catalase) were produced by haloalkaliphilic isolates. These enzymes maybe used in alkaline and saline industrial processes. Although Bacillus subtilis was susceptible to all antibiotics, other isolates showed resistance to at least one antibiotic. The resistance against antibiotics were found as ampicillin/sulbactam 83%, amoxycillin/clavulanic acid 83%, ampicillin 67%, mupirocin 67%, chloramphenicol 50%, tetracycline 50%, imipenem 50%, meropenem 50%, cefadroxil 17%. These bacteria may have develope resistance to antibiotics that entering their natural environment in different ways.

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.

scholarly journals Disinfection performance of a drinking water bottle system with a UVC LED cap against waterborne pathogens and heterotrophic contaminants

2021 ◽  
Author(s):  
Richard M Mariita ◽  
Sebastien A Blumenstein ◽  
Christian M Beckert ◽  
Thomas Gombas ◽  
Rajul V Randive
Keyword(s):  
Public Health ◽  
16S Rrna ◽  
16S Rrna Gene ◽  
Heterotrophic Bacteria ◽  
Health Concern ◽  
Plate Count ◽  
Rrna Gene ◽  
Public Health Concern ◽  
Worst Case ◽  
The 16S Rrna Gene

Background: The purgaty One systems (cap+bottle) are portable stainless-steel water bottles with ultraviolet subtype C (UVC) disinfection capability. This study examines the bottle design, verifies disinfection performance against Escherichia coli, Pseudomonas aeruginosa, Vibrio cholerae and heterotrophic contaminants and addresses the public health relevance of heterotrophic bacteria. Methods: Bottles were inoculated with bacterial strains and disinfection efficacy examined using colony forming unit (CFU) assay. The heterotrophic plate count (HPC) method was used to determine the disinfection performance against environmental contaminants at day 0 and after 3 days of water stagnation. All UVC irradiation experiments were performed under stagnant conditions to confirm that the preset application cycle of 55 seconds offers the desired disinfection performance under worst-case condition. To determine the effectiveness of purgaty One systems (cap+bottle) in disinfection, inactivation efficacy or log reduction value (LRV) was determined using bacteria concentration between UVC ON condition and controls (UVC OFF). The study utilized the 16S rRNA gene for isolate characterization by identifying HPC bacteria to confirm if they belong to groups that are of public health concern. Results: Purgaty One systems fitted with Klaran UVC LEDs achieved 99.99% inactivation (LRV4) efficacy against E. coli and 99.9% inactivation (LRV3) against P. aeruginosa, V. cholerae and heterotrophic contaminants. Based on the 16S rRNA gene analyses, the study determined that the identified HPC isolates enriched by UVC irradiation are of rare public health concern. Conclusion: The bottles satisfactorily inactivated the target pathogenic bacteria and HPC contaminants even after 3 days of water stagnation.

scholarly journals Alloscardovia omnicolens gen. nov., sp. nov., from human clinical samples

2007 ◽  
Vol 57 (7) ◽  
pp. 1442-1446 ◽  
Author(s):  
Geert Huys ◽  
Marc Vancanneyt ◽  
Klaas D'Haene ◽  
Enevold Falsen ◽  
Georges Wauters ◽  
...  
Keyword(s):  
Gardnerella Vaginalis ◽  
Phenotypic Characterization ◽  
Clinical Samples ◽  
Rrna Gene ◽  
Heat Shock Protein 60 ◽  
The Novel ◽  
Phenotypic Differentiation ◽  
Novel Genus ◽  
Sequence Similarities

The taxonomic position of 12 isolates tentatively assigned to the genus Bifidobacterium on the basis of a limited phenotypic characterization was examined. The isolates were collected between 1978 and 2005 in Belgium, Sweden and Norway, and originated from various human clinical samples, including urine, blood, urethra, oral cavity, tonsil, and abscesses of lung and aortic valve. On the basis of band number and clustering analysis, repetitive DNA element-based PCR fingerprinting using the BOXA1R and (GTG)5 primers indicated that the clinical isolates represented a taxon probably not belonging to the genus Bifidobacterium. Analysis of 16S rRNA gene sequence similarities revealed that the isolates were most closely affiliated to Parascardovia denticolens LMG 18312T (93.0–93.2 %), Scardovia inopinata LMG 18313T (92.9–93.1 %) and other members of the Bifidobacteriaceae, indicating that the isolates belong to a novel genus within that family. This observation was further substantiated by the results of partial sequencing of the heat-shock protein 60 gene (hsp60) and determination of the DNA G+C contents (47.3–48.3 mol%). Members of the novel taxon can be phenotypically distinguished from S. inopinata, P. denticolens and Gardnerella vaginalis by the ability to grow on agar under aerobic conditions and on the basis of positive reactions for acid production from l-arabinose, raffinose, salicin and d-xylose. Unambiguous phenotypic differentiation from Aeriscardovia aeriphila and Bifidobacterium species may be difficult, so phenotypic analyses should be complemented by molecular methods. The values for DNA–DNA binding among four members of the novel genus were in the range of 89–100 %, indicating that the strains should be considered as a single novel species of a novel genus, for which the name Alloscardovia omnicolens gen. nov., sp. nov. is proposed. The type strain of Alloscardovia omnicolens is CCUG 31649T (=LMG 23792T).

scholarly journals Effects of Contemporary Irrigant Activation Schemes and Subsequent Placement of an Interim Dressing on Bacterial Presence and Activity in Root Canals Associated with Asymptomatic Apical Periodontitis

2020 ◽  
Vol 9 (3) ◽  
pp. 854 ◽  
Author(s):  
Alexandre P. L. Carvalho ◽  
Laura C. L. Nardello ◽  
Fernanda S. Fernandes ◽  
Fernanda P. Bruno ◽  
Luiza R. Paz ◽  
...  
Keyword(s):  
Calcium Hydroxide ◽  
Root Canal ◽  
Quantitative Polymerase Chain Reaction ◽  
Apical Periodontitis ◽  
Universal Primers ◽  
Rrna Gene ◽  
Root Canal Preparation ◽  
Root Canals ◽  
Mechanical Preparation ◽  
Root Canal Disinfection

New tools for activating endodontic irrigants have evolved, yet their impact on root canal disinfection, in comparison to the passive placing of an inter-visit medication, have not yet been fully elucidated. The use of DNA- and rRNA-based methods may cast some new light on this issue, as they allow a comparison to be made between microbial presence and activity. Therefore, the aim of this single-arm intervention trial is to evaluate the antibacterial effect of endodontic procedures using both molecular methods. Root canal samples were obtained from 20 patients with asymptomatic apical periodontitis after each treatment step: access cavity, chemo-mechanical preparation, adjunctive procedures (XP-endo Finisher file and passive ultrasonic irrigation), calcium hydroxide medication, and 2nd-visit root canal preparation. DNA and cDNA from the samples were subjected to quantitative polymerase chain reaction with universal primers for the bacterial 16S rRNA gene. Chemo-mechanical preparation promoted a drastic reduction in bacterial levels and activity, whereas the adjunctive procedures did not make a significant contribution to further disinfection. At the 2nd visit, bacteria were active after the use of calcium hydroxide medication; however, they were significantly reduced after a 2nd-visit preparation. Consequently, the lowest bacterial levels were found at the end of the treatment. This clinical trial, which used an rRNA and rDNA combined approach, confirmed previous studies showing that root canal preparation represents the main strategy for root canal disinfection.

Sandarakinotalea sediminis gen. nov., sp. nov., a novel member of the family Flavobacteriaceae

2006 ◽  
Vol 56 (5) ◽  
pp. 959-963 ◽  
Author(s):  
Shams Tabrez Khan ◽  
Yasuyoshi Nakagawa ◽  
Shigeaki Harayama
Keyword(s):  
Fatty Acids ◽  
16S Rrna ◽  
16S Rrna Gene ◽  
Gene Sequence ◽  
Heterotrophic Bacteria ◽  
Sequence Similarity ◽  
16S Rrna Gene Sequence ◽  
Rrna Gene ◽  
Rrna Gene Sequence ◽  
The Family

Four Gram-negative, orange-coloured, aerobic, heterotrophic bacteria were isolated from sediment samples collected on the Pacific coast of Japan near the cities of Toyohashi and Katsuura. 16S rRNA gene sequence analysis indicated that these strains form a distinct lineage within the family Flavobacteriaceae. The four isolates shared 99.9–100 % 16S rRNA gene sequence similarity with each other and showed 88–90.9 % similarity with their neighbours in the family Flavobacteriaceae. The four strains also shared high DNA–DNA reassociation values of 67–99 % with each other. All the strains grew at 37 °C but not at 4 °C, and degraded gelatin, starch and DNA. The major fatty acids were i-C15 : 0, a-C15 : 0, i-C16 : 0 and i-C17 : 0 3-OH. However, two common fatty acids of members of the Flavobacteriaceae, i-C15 : 1 and a-C15 : 1, were absent in these strains. The DNA G+C contents of the four strains were in the range 35–37 mol%. On the basis of the polyphasic evidence, it was concluded that these strains should be classified as a novel genus and a novel species in the family Flavobacteriaceae, for which the name Sandarakinotalea sediminis gen. nov., sp. nov. is proposed. The type strain of Sandarakinotalea sediminis is CKA-5T (=NBRC 100970T=LMG 23247T).

scholarly journals Observability of anammox activity in single-stage nitritation/anammox reactors using mass balances

2015 ◽  
Vol 1 (4) ◽  
pp. 523-534 ◽  
Author(s):  
Sarina Schielke-Jenni ◽  
Kris Villez ◽  
Eberhard Morgenroth ◽  
Kai M. Udert
Keyword(s):  
Heterotrophic Bacteria ◽  
Single Stage ◽  
Anammox Bacteria ◽  
Mass Balances ◽  
Microbial Kinetics ◽  
Anammox Activity

Theoretically, mass balances based on microbial kinetics allow the determination of the activity of anammox bacteria (AMX) and heterotrophic bacteria (HET). In practise, the variance of the resulting activities is too high.

Sign in / Sign up

Export Citation Format

Share Document