Degenerate primers as biomarker for gene-targeted metagenomics of the catechol 1, 2-dioxygenase-encoding gene in microbial populations of petroleum-contaminated environments

Nafian, Fatemeh; Gharavi, Sara; Soudi, Mohammad Reza

doi:10.1007/s13213-016-1197-3

Cited by 7 publications

(2 citation statements)

References 47 publications

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“…Therefore, it is necessary to carefully select primers that are adequate for the desired application, making sure that they allow for the precise detection and identification of the target pathogens. Degenerate primers can increase the detected number of taxa, and also allow for metabarcoding studies using non-standard marker genes [ 79 , 80 ], but the risk of primer slippage can cause a variation in amplicon length, introducing a bias in experiments [ 81 ]. Another risk is the potential amplification of host sequences during metabarcoding analyses.…”

Section: High-throughput Sequencing (Hts)mentioning

confidence: 99%

Metagenomics Approaches for the Detection and Surveillance of Emerging and Recurrent Plant Pathogens

et al. 2021

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Globalization has a dramatic effect on the trade and movement of seeds, fruits and vegetables, with a corresponding increase in economic losses caused by the introduction of transboundary plant pathogens. Current diagnostic techniques provide a useful and precise tool to enact surveillance protocols regarding specific organisms, but this approach is strictly targeted, while metabarcoding and shotgun metagenomics could be used to simultaneously detect all known pathogens and potentially new ones. This review aims to present the current status of high-throughput sequencing (HTS) diagnostics of fungal and bacterial plant pathogens, discuss the challenges that need to be addressed, and provide direction for the development of methods for the detection of a restricted number of related taxa (specific surveillance) or all of the microorganisms present in a sample (general surveillance). HTS techniques, particularly metabarcoding, could be useful for the surveillance of soilborne, seedborne and airborne pathogens, as well as for identifying new pathogens and determining the origin of outbreaks. Metabarcoding and shotgun metagenomics still suffer from low precision, but this issue can be limited by carefully choosing primers and bioinformatic algorithms. Advances in bioinformatics will greatly accelerate the use of metagenomics to address critical aspects related to the detection and surveillance of plant pathogens in plant material and foodstuffs.

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Section: High-throughput Sequencing (Hts)mentioning

confidence: 99%

Metagenomics Approaches for the Detection and Surveillance of Emerging and Recurrent Plant Pathogens

et al. 2021

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“…Metagenomics avoids the separation and culture of microbes and increases the probability of utilizing microbe resources; it provides research strategies for finding new functional genes and biological catalysts (enzymes). To cover the extant diversity, gene‐targeted metagenomics is currently the method of choice for PCR‐based targeting . This approach not only provides insight into microbial gene diversity but also the means to detect genes that could be important in environmental processes.…”

Section: Introductionmentioning

confidence: 99%

Genetic diversity of catechol 1,2-dioxygenase in the fecal microbial metagenome

Xiong

Deng

et al. 2017

J. Basic Microbiol

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Catechol 1,2-dioxygenase is the key enzyme that catalyzes the cleavage of the aromatic ring of catechol. We explored the genetic diversity of catechol 1,2-dioxygenase in the fecal microbial metagenome by PCR with degenerate primers. A total of 35 gene fragments of C12O were retrieved from microbial DNA in the feces of pygmy loris. Based on phylogenetic analysis, most sequences were closely related to C12O sequences from Acinetobacter. A full-length C12O gene was directly cloned, heterologously expressed in Escherichia coli, and biochemically characterized. Purified catPL12 had optimum pH and temperature pH 8.0 and 25 °C and retained 31 and 50% of its maximum activity when assayed at 0 and 35 °C, respectively. The enzyme was stable at 25 and 37 °C, retaining 100% activity after pre-incubation for 1 h. The kinetic parameters of catPL12 were determined. The enzyme had apparent K of 67 µM, V of 7.3 U/mg, and k of 4.2 s for catechol, and the cleavage activities for 3-methylcatechol, 4-methylcatechol, and 4-chlorocatechol were much less than for catechol, and no activity with hydroquinone or protocatechuate was detected. This study is the first to report the molecular and biochemical characterizations of a cold-adapted catechol 1,2-dioxygenase from a fecal microbial metagenome.

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Highly efficient phenol degradation in a batch moving bed biofilm reactor: benefiting from biofilm-enhancing bacteria

Irankhah

Ali

Soudi

et al. 2018

World J Microbiol Biotechnol

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In this study, the efficiency improvement of three moving bed biofilm reactors (MBBRs) was investigated by inoculation of activated sludge cells (R1), mixed culture of eight strong phenol-degrading bacteria consisted of Pseudomonas spp. and Acinetobacter spp. (R2) and the combination of both (R3). Biofilm formation ability of eight bacteria was assessed initially using different methods and media. Maximum degradation of phenol, COD, biomass growth and also changes in organic loading shock were used as parameters to measure the performance of reactors. According to the results, all eight strains were determined as enhanced biofilm forming bacteria (EBFB). Under optimum operating conditions, more than 90% of initial COD load of 2795 mg L −1 was reduced at 24 HRT in R3 while this reduction efficiency was observed in concentrations of 1290 mg L −1 and 1935 mg L −1 , in R1 and R2, respectively. When encountering phenol loading shock-twice greater than optimum amount-R1, R2 and R3 managed to return to the steady-state condition within 32, 24 and 18 days, respectively. SEM microscopy and biomass growth measurements confirmed the contribution of more cells to biofilm formation in R3 followed by R2. Additionally, established biofilm in R3 was more resistant to phenol loading shock which can be attributed to the enhancer role of EBFB strains in this reactor. It has been demonstrated that the bacteria with both biofilmforming and contaminant-degrading abilities are not only able to promote the immobilization of other favorable activated sludge cells in biofilm structure, but also cooperate in contaminant degradation which all consequently lead to improvement of treatment efficiency.

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Degenerate primers as biomarker for gene-targeted metagenomics of the catechol 1, 2-dioxygenase-encoding gene in microbial populations of petroleum-contaminated environments

Cited by 7 publications

References 47 publications

Metagenomics Approaches for the Detection and Surveillance of Emerging and Recurrent Plant Pathogens

Metagenomics Approaches for the Detection and Surveillance of Emerging and Recurrent Plant Pathogens

Genetic diversity of catechol 1,2-dioxygenase in the fecal microbial metagenome

Highly efficient phenol degradation in a batch moving bed biofilm reactor: benefiting from biofilm-enhancing bacteria

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