Abstract:
:Characterization of commercial microbial consortia products for human and environmental health risk assessment is a major challenge for regulatory agencies. As a means to develop an approach to assess the potential environmental risk of these products, research was conducted to compare four genomics methods for characterizing bacterial communities; (i) Denaturing Gradient Gel Electrophoresis (DGGE), (ii) Clonal-Restriction Fragment Length Polymorphism (C/RFLP), (iii) partial 16S rDNA amplification, cloning followed by Sanger sequencing (PRACS) and (iv) Next-Generation Sequencing (NGS) based on Ion Torrent technology. A commercially available microbial consortium, marketed as a remediation agent for degrading petroleum hydrocarbon contamination in soil and water, was assessed. The bacterial composition of the commercial microbial product was characterized using the above four methods. PCR amplification of 16S rDNA was performed targeting the variable region V6 for DGGE, C/RFLP and PRACS and V5 for Ion Torrent sequencing. Ion Torrent technology was shown to be a promising tool for initial screening by detecting the majority of bacteria in the consortium that were also detected by DGGE, C/RFLP and PRACS. Additionally, Ion Torrent sequencing detected some of the bacteria that were claimed to be in the product, while three other methods failed to detect these specific bacteria. However, the relative proportions of the microbial composition detected by Ion Torrent were found to be different from DGGE, C/RFLP and PRACS, which gave comparable results across these three methods. The discrepancy of the Ion Torrent results may be due to the short read length generated by this technique and the targeting of different variable regions on the 16S rRNA gene used in this study. Arcobacter spp. a potential pathogenic bacteria was detected in the product by all methods, which was further confirmed using genus and species-specific PCR, RFLP and DNA-based sequence analyses. However, the viability of Arcobacter spp. was not confirmed. This study suggests that a combination of two or more methods may be required to ascertain the microbial constituents of a commercial microbial consortium reliably and for the presence of potentially human pathogenic contaminants.
journal_name
J Microbiol Methodsjournal_title
Journal of microbiological methodsauthors
Samarajeewa AD,Hammad A,Masson L,Khan IU,Scroggins R,Beaudette LAdoi
10.1016/j.mimet.2014.11.013subject
Has Abstractpub_date
2015-01-01 00:00:00pages
103-11eissn
0167-7012issn
1872-8359pii
S0167-7012(14)00328-5journal_volume
108pub_type
杂志文章abstract::We evaluated the use of mixed oligonucleotide probes hybridized to metagenomic clones spotted on high density membranes. The pooled probes included oligonucleotides designed for genes associated with denitrification, antibiotic resistance, and dehalogenation among others. Pyrosequence comparison between the clones and...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/j.mimet.2008.09.009
更新日期:2009-01-01 00:00:00
abstract::A simplified amplified-fragment length polymorphism (AFLP) method was developed and applied to genotype 52 Mycobacterium tuberculosis isolates. This method can be carried out using only one restriction enzyme (XhoI), one double strand adapter, and one PCR primer. The amounts of DNA and DNA polymerase, and the concentr...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/j.mimet.2009.07.007
更新日期:2009-09-01 00:00:00
abstract::A quantitative real time reverse transcription polymerase chain reaction (qRT-PCR) assay was utilized to quantify the expression of ElHDO in the fungus Exophiala lecanii-corni during the biodegradation of ethylbenzene and other volatile organic pollutants. The assay was applied to measure the impact of pollutant mixtu...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/j.mimet.2006.03.018
更新日期:2006-11-01 00:00:00
abstract::DNA probes specific for rRNA of selected target species were utilised for the detection of beer spoilage bacteria of the genera Pectinatus and Megasphaera using a hybridisation protection assay (HPA). All the probes were modified during synthesis by addition of an amino linker arm at the 5' end or were internally modi...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/j.mimet.2013.10.014
更新日期:2014-01-01 00:00:00
abstract::To establish a simple and rapid high-resolution melting curve (HRM) method, 5 different strains of Salmonella were identified by adding DNA denaturants at different concentrations into the HRM system to change the characteristics of DNA melting and to obtain different Tm (dissolving temperature) values of DNA from dif...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/j.mimet.2019.105806
更新日期:2020-05-01 00:00:00
abstract::Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) and chromogenic media are widely used in clinical microbiology laboratories to facilitate the rapid selection and identification of pathogens. The aim of this study was to evaluate whether usage of chromogenic media limits the ...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/j.mimet.2017.03.001
更新日期:2017-05-01 00:00:00
abstract::A rapid, sensitive fluorescence measurement method for detecting the bacterial count using CdSe/ZnS as a fluorescence marker was described. High-quality CdSe/ZnS nanocrystals were synthesized and successfully conjugated with bacteria. The fluorescence intensity was proportional to bacterial count in the range of 10(2)...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/j.mimet.2009.09.019
更新日期:2009-12-01 00:00:00
abstract::This paper presents a robust two-step segmentation procedure for the study of biofilm structure. Without user intervention, the procedure segments volumetric biofilm images generated by a confocal laser scanning microscopy (CLSM). This automated procedure implements an anisotropic diffusion filter as a preprocessing s...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/j.mimet.2007.05.022
更新日期:2007-09-01 00:00:00
abstract::Brucellosis is a zoonotic disease transmitted to humans from infected animals. As a systemic disease, it can harm any organ or system of the host body. Human brucellosis presents with various clinical symptoms, which makes diagnosis challenging. Serological diagnosis of brucellosis is based on ELISA or agglutination t...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/j.mimet.2020.105964
更新日期:2020-07-01 00:00:00
abstract::Amplification of high GC content genes by PCR is a major challenge during the creation of recombinant GC-rich DNA constructs. This may be due to the difficulty in DNA denaturation or the possibility of forming secondary structures from DNA templates. Tools have been described to address the technical problems associat...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/j.mimet.2020.106121
更新日期:2020-12-13 00:00:00
abstract::Transposon Integration mediated Mutagenesis (TIM) is a broadly applicable tool for protein engineering. This method combines random integration of modified bacteriophage Mu transposons with their subsequent defined excision employing type IIS restriction endonuclease AarI. TIM enables deletion or insertion of an arbit...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/j.mimet.2008.06.017
更新日期:2008-10-01 00:00:00
abstract::Ruthenium red is a polycationic stain used to visualize acid polysaccharides on the outer surface of cells. Ruthenium red staining followed by electron microscopic analysis was used to demonstrate the presence of an external glycoprotein layer surrounding the spore of both Bacillus anthracis and Bacillus subtilis. Thi...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/j.mimet.2004.02.012
更新日期:2004-07-01 00:00:00
abstract::Two novel modifications of continuous culture with gradual change of dilution rate (A-stat): D-stat and auxo-accelerostat were evaluated in the studies of the effect of changing individual environmental parameters (T, pH, pO(2), substrate concentration, etc.) on growth characteristics of different microorganisms. Comm...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/s0167-7012(03)00143-x
更新日期:2003-10-01 00:00:00
abstract::Phenotypic differentiation between Campylobacter fetus (C. fetus) subspecies fetus and C. fetus subspecies venerealis is hampered by poor reliability and reproducibility of biochemical assays. AFLP (amplified fragment length polymorphism) and MLST (multilocus sequence typing) are the molecular standards for C. fetus s...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/j.mimet.2013.06.005
更新日期:2013-10-01 00:00:00
abstract::We evaluated the performance of two enzyme-immunoassays (EIA) for the detection of naturally occurring, thermophilic Campylobacter spp. found in faecal samples from cattle (n = 21 and n = 26) and swine (n = 43) relative to the standard culture method, and also assuming that none of the tests was the definitive standar...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/s0167-7012(99)00080-9
更新日期:1999-10-01 00:00:00
abstract::We have developed a software tool, GenomeComp, for summarizing, parsing and visualizing the genome sequences comparison results derived from voluminous BLAST textual output. With GenomeComp, the variation between genomes can be easily highlighted, such as repeat regions, insertions, deletions and rearrangements of gen...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/s0167-7012(03)00094-0
更新日期:2003-09-01 00:00:00
abstract::A microplate-based rapid, inexpensive and robust technique is developed by using tetrazolium salt 2,3-bis[2-methyloxy-4-nitro-5-sulfophenyl]-2H-tetrazolium-5-carboxanilide (XTT) and menadione to determine the viability of Mycobacterium tuberculosis, Mycobacterium bovis BCG and Mycobacterium smegmatis bacilli in microp...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/j.mimet.2010.11.013
更新日期:2011-02-01 00:00:00
abstract::Ureaplasma parvum and Ureaplasma urealyticum are recently recognized species of the genus Ureaplasma. In humans, Ureaplasma spp. can be found on mucosal surfaces, primarily in the respiratory and urogenital tracts. They have been implicated in various human diseases such as nongonococcal urethritis, intrauterine infec...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/j.mimet.2004.08.005
更新日期:2005-01-01 00:00:00
abstract::Synthesis and application of reliable nanoscale materials is a progressive domain and the limelight of modern nanotechnology. Conventional physicochemical approaches for the synthesis of metal nanoparticles have become obsolete owing to costly and hazardous materials. There is a need to explore alternative, cost-effec...
journal_title:Journal of microbiological methods
pub_type: 杂志文章,评审
doi:10.1016/j.mimet.2019.02.010
更新日期:2019-04-01 00:00:00
abstract::A biofilm reactor was constructed to monitor the long-term growth and removal of biofilms as monitored by the use of a quartz crystal microbalance (QCM) and a novel optical method. The optical method measures the reflectance of white light off the surface of the quartz crystal microbalance electrode (gold) for determi...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/j.mimet.2006.01.016
更新日期:2006-09-01 00:00:00
abstract::Bacillus anthracis, the causative agent of the human disease anthrax, Bacillus cereus, a food-borne pathogen capable of causing human illness, and Bacillus thuringiensis, a well-characterized insecticidal toxin producer, all cluster together within a very tight clade (B. cereus group) phylogenetically and are indistin...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/j.mimet.2003.11.004
更新日期:2004-03-01 00:00:00
abstract::Cryptosporidium parvum is an enteric coccidian parasite that is recognised as a frequent cause of water-borne disease in humans. We report for the first time on use of the in vitro HCT-8 cell culture-quantitative PCR (qPCR) assay and the in vivo SCID-mouse bioassay for evaluating critical factors that reduce or elimin...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/j.mimet.2010.01.017
更新日期:2010-03-01 00:00:00
abstract::As part of a major European research project, a diagnostic PCR assay, including an internal amplification control, was developed and validated in a collaborative trial for the detection of Escherichia coli O157. The assay is based on amplification of sequences of the rfbE O157 gene. The collaborative trial, including ...
journal_title:Journal of microbiological methods
pub_type: 杂志文章,多中心研究
doi:10.1016/j.mimet.2003.08.012
更新日期:2003-12-01 00:00:00
abstract::A simple and reliable method was developed to isolate intact epithelial cells from pig and rabbit ilea and these were used to investigate the adhesion of Yersinia enterocolitica. Hydrophobic interaction was eliminated by treating the bacterial culture with 0.8 M tetramethyl urea (TMU). Virulent strains of Y. enterocol...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/j.mimet.2006.08.009
更新日期:2007-02-01 00:00:00
abstract::The aim of this study was to demonstrate that flow cytometry (FACS) could potentially be employed for rapid viability assessment of probiotic bacteria immobilized or encapsulated in complex matrices. Lactobacillus rhamnosus GG was immobilized within six different protein environments using whey protein isolate (WPI) a...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/j.mimet.2010.07.003
更新日期:2010-09-01 00:00:00
abstract::A real-time PCR (RTm-PCR) assay using fluorescently labeled oligonucleotides (TaqMan probes) was used to detect and quantify the recombinant Rhodococcus sp. strain RHA1(fcb) in soil. One primer and probe set targeted a hypervariable region of the 16S rRNA gene unique to strain RHA1(fcb) and its phylogenetic relatives,...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/s0167-7012(02)00067-2
更新日期:2002-10-01 00:00:00
abstract::A real-time quantitative polymerase chain reaction (Q-PCR) assay was developed for detecting and quantifying Escherichia coli in water samples from agricultural watersheds. The assay included optimization of DNA extraction and purification from water samples, and Q-PCR amplification conditions using newly designed spe...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/j.mimet.2007.02.016
更新日期:2007-06-01 00:00:00
abstract::To date, it has been problematic to accurately quantify multiple nucleic acid sequences, representing microbial targets, in multi-target mixtures using oligonucleotide microarrays, primarily due to nonspecific target binding (i.e., cross-hybridization). While some studies ignore the effects of nonspecific binding, oth...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/j.mimet.2008.05.013
更新日期:2008-09-01 00:00:00
abstract::The analysis of soil fungal communities by molecular fingerprinting and subsequent identification of the underlying populations require the amplification of a phylogenetically informative gene fragment. In this study we tested the reliability and suitability of the previously published fungal primer combination (NS1/F...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/j.mimet.2005.06.014
更新日期:2006-04-01 00:00:00
abstract::Any forms of valorization of microorganisms would require accurate identity recognition to ensure repeatability, reproducibility and quality assurance. This study aimed to evaluate the effectiveness of different primers for identifying cultured eukaryotic microalgae using a simple 18S rDNA approach. A total of 34 isol...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/j.mimet.2020.105890
更新日期:2020-05-01 00:00:00