scholarly journals A viability qPCR dilemma: Are longer amplicons better?

Author(s):  
Wannes Van Holm ◽  
Justien Ghesquière ◽  
Nico Boon ◽  
Tim Verspecht ◽  
Kristel Bernaerts ◽  
...  

The development of viability qPCR (v-qPCR) has allowed for a more accurate assessment of the viability of a microbial sample by limiting the amplification of DNA from dead cells. Although valuable, v-qPCR is not infallible. One of the most limiting factors for accurate live/dead distinction is the length of the qPCR amplicon used. However, no consensus or guidelines exist for selecting and designing amplicon lengths for optimal results. In this study, a wide range of incrementally increasing amplicon lengths (68-906 bp) was used on live and killed cells of nine bacterial species treated with viability dye (PMA). Increasing amplicon lengths up to approximately 200 bp resulted in increasing quantification cycle (Cq) differences between live and killed cells, while maintaining a good qPCR efficiency. Longer amplicon lengths, up to approximately 400 bp, further increased Cq difference, but at the cost of qPCR efficiency. Above 400 bp, no valuable increase in Cq differences was observed. Importance Viability qPCR (v-qPCR) has evolved to a valuable, mainstream technique for determining the number of viable micro-organisms in samples by qPCR. Amplicon length is known to be positively correlated with the ability to distinguish between live and dead bacteria but is negatively correlated with qPCR efficiency. This trade-off is often not taken into account and might have an impact on the accuracy of v-qPCR data. Currently there is no consensus on the optimal amplicon length. This paper provides methods to determine the optimal amplicon length and suggests an amplicon length range for optimal v-qPCR, taking into consideration the trade-off between qPCR efficiency and live-dead distinction.

2019 ◽  
Vol 5 ◽  
pp. 38-56 ◽  
Author(s):  
Khulod A. Hemida ◽  
Amany M.M. Reyad

Salinity is one of the most dangerous environmental limiting factors of the plant productivity. A wide range of adaptation strategies is required to overcome salinity stress. However, such strategies seem to be long drawn and cost-intensive. It has been confirmed in recent years that plant growth promoting endophytes (PGPEs) that have the ability to further build a symbiotic association with their host to improve host plant salt tolerance. In our investigation try to improve plant salt tolerance using different species of endophytic bacteria. From the total eight endophytic bacterial species were isolated from root, stem, and leaf of Carthamustinctorius (safflower) plant, two isolates were capable of using 1-aminocyclopropane-1-carboxylic acid (ACC) as a sole nitrogen source, and they are of positive results for (ACC) deaminase activity and indole-3-acetic acid (IAA) production. The bacterial isolates were identified using 16S ribosomal DNA technique as Bacillus cereus and Bacillus aerius and had accession numbers MG708176 and MG711593 respectively, by submitting their sequences in GenBank database. This study showed that the bacterial strains B. cereus and B. aerius are valuable biological plant growth promoters that could enhance salt tolerance in Safflower plants under 100, 200, and 300mMNaCl levels resulting in an increase in plant growth and ascorbate-glutathione redox cycle, in comparison with the non-inoculated controls. Our findings reported that the co-inoculation of the two selected endophytic bacteria strains were successfully isolated from Safflower seedlings significantly alleviated the harmful effects of salt stress, promoted plant growth and biomass yield.


2006 ◽  
Vol 274 (1607) ◽  
pp. 225-230 ◽  
Author(s):  
Victoria Furió ◽  
Andrés Moya ◽  
Rafael Sanjuán

Mutation rates should be governed by at least three evolutionary factors: the need for beneficial mutations, the benefit of minimizing the mutational load and the cost of replication fidelity. RNA viruses show high mutation rates compared with DNA micro-organisms, and recent findings suggest that the cost of fidelity might play a role in the evolution of increased mutation rates. Here, by analysing previously published data from HIV-1 reverse transcriptase in vitro assays, we show a trade-off between enzymatic accuracy and the maximum rate of polymerization, thus providing a biochemical basis for the fitness cost of fidelity in HIV-1. This trade-off seems to be related to inefficient extension of mispairs, which increases fidelity at the expense of the polymerization rate. Since in RNA viruses fast replication is critical for survival, this could impose a high cost of fidelity and favour the evolution of high mutation rates.


2020 ◽  
Vol 4 (02) ◽  
pp. 34-45
Author(s):  
Naufal Dzikri Afifi ◽  
Ika Arum Puspita ◽  
Mohammad Deni Akbar

Shift to The Front II Komplek Sukamukti Banjaran Project is one of the projects implemented by one of the companies engaged in telecommunications. In its implementation, each project including Shift to The Front II Komplek Sukamukti Banjaran has a time limit specified in the contract. Project scheduling is an important role in predicting both the cost and time in a project. Every project should be able to complete the project before or just in the time specified in the contract. Delay in a project can be anticipated by accelerating the duration of completion by using the crashing method with the application of linear programming. Linear programming will help iteration in the calculation of crashing because if linear programming not used, iteration will be repeated. The objective function in this scheduling is to minimize the cost. This study aims to find a trade-off between the costs and the minimum time expected to complete this project. The acceleration of the duration of this study was carried out using the addition of 4 hours of overtime work, 3 hours of overtime work, 2 hours of overtime work, and 1 hour of overtime work. The normal time for this project is 35 days with a service fee of Rp. 52,335,690. From the results of the crashing analysis, the alternative chosen is to add 1 hour of overtime to 34 days with a total service cost of Rp. 52,375,492. This acceleration will affect the entire project because there are 33 different locations worked on Shift to The Front II and if all these locations can be accelerated then the duration of completion of the entire project will be effective


Author(s):  
Nataliya Stoyanets ◽  
◽  
Mathias Onuh Aboyi ◽  

The article defines that for the successful implementation of an innovative project and the introduction of a new product into production it is necessary to use advanced technologies and modern software, which is an integral part of successful innovation by taking into account the life cycle of innovations. It is proposed to consider the general potential of the enterprise through its main components, namely: production and technological, scientific and technical, financial and economic, personnel and actual innovation potential. Base for the introduction of technological innovations LLC "ALLIANCE- PARTNER", which provides a wide range of support and consulting services, services in the employment market, tourism, insurance, translation and more. To form a model of innovative development of the enterprise, it is advisable to establish the following key aspects: the system of value creation through the model of cooperation with partners and suppliers; creating a value chain; technological platform; infrastructure, determine the cost of supply, the cost of activities for customers and for the enterprise as a whole. The system of factors of influence on formation of model of strategic innovative development of the enterprise is offered. The expediency of the cost of the complex of technological equipment, which is 6800.0 thousand UAH, is economically calculated. Given the fact that the company plans to receive funds under the program of socio-economic development of Sumy region, the evaluation of the effectiveness of the innovation project, the purchase of technological equipment, it is determined that the payback period of the project is 3 years 10 months. In terms of net present value (NPV), the project under study is profitable. The project profitability index (PI) meets the requirements for a positive decision on project implementation> 1.0. The internal rate of return of the project (IRR) also has a positive value of 22% because it exceeds the discount rate.


2020 ◽  
Vol 12 (7) ◽  
pp. 2767 ◽  
Author(s):  
Víctor Yepes ◽  
José V. Martí ◽  
José García

The optimization of the cost and CO 2 emissions in earth-retaining walls is of relevance, since these structures are often used in civil engineering. The optimization of costs is essential for the competitiveness of the construction company, and the optimization of emissions is relevant in the environmental impact of construction. To address the optimization, black hole metaheuristics were used, along with a discretization mechanism based on min–max normalization. The stability of the algorithm was evaluated with respect to the solutions obtained; the steel and concrete values obtained in both optimizations were analyzed. Additionally, the geometric variables of the structure were compared. Finally, the results obtained were compared with another algorithm that solved the problem. The results show that there is a trade-off between the use of steel and concrete. The solutions that minimize CO 2 emissions prefer the use of concrete instead of those that optimize the cost. On the other hand, when comparing the geometric variables, it is seen that most remain similar in both optimizations except for the distance between buttresses. When comparing with another algorithm, the results show a good performance in optimization using the black hole algorithm.


2021 ◽  
Vol 9 (4) ◽  
pp. 862
Author(s):  
Vittoria Catara ◽  
Jaime Cubero ◽  
Joël F. Pothier ◽  
Eran Bosis ◽  
Claude Bragard ◽  
...  

Bacteria in the genus Xanthomonas infect a wide range of crops and wild plants, with most species responsible for plant diseases that have a global economic and environmental impact on the seed, plant, and food trade. Infections by Xanthomonas spp. cause a wide variety of non-specific symptoms, making their identification difficult. The coexistence of phylogenetically close strains, but drastically different in their phenotype, poses an added challenge to diagnosis. Data on future climate change scenarios predict an increase in the severity of epidemics and a geographical expansion of pathogens, increasing pressure on plant health services. In this context, the effectiveness of integrated disease management strategies strongly depends on the availability of rapid, sensitive, and specific diagnostic methods. The accumulation of genomic information in recent years has facilitated the identification of new DNA markers, a cornerstone for the development of more sensitive and specific methods. Nevertheless, the challenges that the taxonomic complexity of this genus represents in terms of diagnosis together with the fact that within the same bacterial species, groups of strains may interact with distinct host species demonstrate that there is still a long way to go. In this review, we describe and discuss the current molecular-based methods for the diagnosis and detection of regulated Xanthomonas, taxonomic and diversity studies in Xanthomonas and genomic approaches for molecular diagnosis.


Author(s):  
Vincent E. Castillo ◽  
John E. Bell ◽  
Diane A. Mollenkopf ◽  
Theodore P. Stank

Materials ◽  
2021 ◽  
Vol 14 (6) ◽  
pp. 1486
Author(s):  
Eugene B. Caldona ◽  
Ernesto I. Borrego ◽  
Ketki E. Shelar ◽  
Karl M. Mukeba ◽  
Dennis W. Smith

Many desirable characteristics of polymers arise from the method of polymerization and structural features of their repeat units, which typically are responsible for the polymer’s performance at the cost of processability. While linear alternatives are popular, polymers composed of cyclic repeat units across their backbones have generally been shown to exhibit higher optical transparency, lower water absorption, and higher glass transition temperatures. These specifically include polymers built with either substituted alicyclic structures or aromatic rings, or both. In this review article, we highlight two useful ring-forming polymer groups, perfluorocyclobutyl (PFCB) aryl ether polymers and ortho-diynylarene- (ODA) based thermosets, both demonstrating outstanding thermal stability, chemical resistance, mechanical integrity, and improved processability. Different synthetic routes (with emphasis on ring-forming polymerization) and properties for these polymers are discussed, followed by their relevant applications in a wide range of aspects.


2021 ◽  
Vol 8 (1) ◽  
Author(s):  
Jon Kepa Izaguirre ◽  
Leire Barañano ◽  
Sonia Castañón ◽  
José A. L. Santos ◽  
M. Teresa Cesário ◽  
...  

AbstractThe management of municipal solid waste is a major logistic and environmental problem worldwide. Nonetheless, the organic fraction of municipal solid waste (OFMSW) is a valuable source of nutrients which can be used for a variety of purposes, according to the Circular Economy paradigm. Among the possible applications, the bioproduction of a biodegradable polyester, poly(3-hydroxybutyrate) [P(3HB)], using OFMSW as carbon platform is a promising strategy. Here, an economic and environmental assessment of bacterial P(3HB) production from OFMSW is presented based on previously published results. The SuperPro Designer® software was used to simulate P(3HB) production under our experimental parameters. Two scenarios were proposed depending on the fermentation medium: (1) enzymatic hydrolysate of OFMSW supplemented with glucose and plum waste juice; and (2) basal medium supplemented with glucose and plum waste juice. According to our results, both scenarios are not economically feasible under our experimental parameters. In Scenario 1, the low fermentation yield, the cost of the enzymes, the labour cost and the energy consumption are the factors that most contribute to that result. In Scenario 2, the cost of the extraction solvent and the low fermentation yield are the most limiting factors. The possibility of using process waste as raw material for the generation of other products must be investigated to enhance economic feasibility. From an environmental viewpoint, the photochemical oxidation potential (derived from the use of anisole as extraction solvent) and the generation of acid rain and global warming effect (caused by the burning of fuels for power generation) are the most relevant impacts associated to P(3HB) production under our experimental parameters.


Microbiome ◽  
2021 ◽  
Vol 9 (1) ◽  
Author(s):  
Hannes Petruschke ◽  
Christian Schori ◽  
Sebastian Canzler ◽  
Sarah Riesbeck ◽  
Anja Poehlein ◽  
...  

Abstract Background The intestinal microbiota plays a crucial role in protecting the host from pathogenic microbes, modulating immunity and regulating metabolic processes. We studied the simplified human intestinal microbiota (SIHUMIx) consisting of eight bacterial species with a particular focus on the discovery of novel small proteins with less than 100 amino acids (= sProteins), some of which may contribute to shape the simplified human intestinal microbiota. Although sProteins carry out a wide range of important functions, they are still often missed in genome annotations, and little is known about their structure and function in individual microbes and especially in microbial communities. Results We created a multi-species integrated proteogenomics search database (iPtgxDB) to enable a comprehensive identification of novel sProteins. Six of the eight SIHUMIx species, for which no complete genomes were available, were sequenced and de novo assembled. Several proteomics approaches including two earlier optimized sProtein enrichment strategies were applied to specifically increase the chances for novel sProtein discovery. The search of tandem mass spectrometry (MS/MS) data against the multi-species iPtgxDB enabled the identification of 31 novel sProteins, of which the expression of 30 was supported by metatranscriptomics data. Using synthetic peptides, we were able to validate the expression of 25 novel sProteins. The comparison of sProtein expression in each single strain versus a multi-species community cultivation showed that six of these sProteins were only identified in the SIHUMIx community indicating a potentially important role of sProteins in the organization of microbial communities. Two of these novel sProteins have a potential antimicrobial function. Metabolic modelling revealed that a third sProtein is located in a genomic region encoding several enzymes relevant for the community metabolism within SIHUMIx. Conclusions We outline an integrated experimental and bioinformatics workflow for the discovery of novel sProteins in a simplified intestinal model system that can be generically applied to other microbial communities. The further analysis of novel sProteins uniquely expressed in the SIHUMIx multi-species community is expected to enable new insights into the role of sProteins on the functionality of bacterial communities such as those of the human intestinal tract.


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