scholarly journals Optimization of Analytical Parameters for Inferring Relationships among Escherichia coli Isolates from Repetitive-Element PCR by Maximizing Correspondence with Multilocus Sequence Typing Data

2006 ◽  
Vol 72 (9) ◽  
pp. 6049-6052 ◽  
Author(s):  
Tony L. Goldberg ◽  
Thomas R. Gillespie ◽  
Randall S. Singer
Keyword(s):  
Escherichia Coli ◽  
Multilocus Sequence Typing ◽  
Large Scale ◽  
Repetitive Element ◽  
Phylogenetic Analyses ◽  
Epidemiological Studies ◽  
Genetic Distances ◽  
Accurate Information ◽  
Data Types ◽  
Wide Range

ABSTRACT Repetitive-element PCR (rep-PCR) is a method for genotyping bacteria based on the selective amplification of repetitive genetic elements dispersed throughout bacterial chromosomes. The method has great potential for large-scale epidemiological studies because of its speed and simplicity; however, objective guidelines for inferring relationships among bacterial isolates from rep-PCR data are lacking. We used multilocus sequence typing (MLST) as a “gold standard” to optimize the analytical parameters for inferring relationships among Escherichia coli isolates from rep-PCR data. We chose 12 isolates from a large database to represent a wide range of pairwise genetic distances, based on the initial evaluation of their rep-PCR fingerprints. We conducted MLST with these same isolates and systematically varied the analytical parameters to maximize the correspondence between the relationships inferred from rep-PCR and those inferred from MLST. Methods that compared the shapes of densitometric profiles (“curve-based” methods) yielded consistently higher correspondence values between data types than did methods that calculated indices of similarity based on shared and different bands (maximum correspondences of 84.5% and 80.3%, respectively). Curve-based methods were also markedly more robust in accommodating variations in user-specified analytical parameter values than were “band-sharing coefficient” methods, and they enhanced the reproducibility of rep-PCR. Phylogenetic analyses of rep-PCR data yielded trees with high topological correspondence to trees based on MLST and high statistical support for major clades. These results indicate that rep-PCR yields accurate information for inferring relationships among E. coli isolates and that accuracy can be enhanced with the use of analytical methods that consider the shapes of densitometric profiles.

scholarly journals Additional Og-Typing PCR Techniques Targeting Escherichia coli-Novel and Shigella-Unique O-Antigen Biosynthesis Gene Clusters

2020 ◽  
Vol 58 (11) ◽  
Author(s):  
Atsushi Iguchi ◽  
Hironobu Nishii ◽  
Kazuko Seto ◽  
Jiro Mitobe ◽  
Kenichi Lee ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Strong Association ◽  
Gene Clusters ◽  
Epidemiological Studies ◽  
Content Type ◽  
E Coli ◽  
O Antigen ◽  
Wide Range ◽  
Biosynthesis Gene ◽  
Almost All

ABSTRACT The O-serogrouping of pathogenic Escherichia coli is a standard method for subtyping strains for epidemiological studies and controls. O-serogroup diversification shows a strong association with the genetic diversity in some O-antigen biosynthesis gene clusters. Through genomic studies, in addition to the types of O-antigen biosynthesis gene clusters (Og-types) from conventional O-serogroup strains, a number of novel Og-types have been found in E. coli isolates. To assist outbreak investigations and surveillance of pathogenic E. coli at inspection institutes, in previous studies, we developed PCR methods that could determine almost all conventional O-serogroups and some novel Og-types. However, there are still many Og-types that may not be determined by simple genetic methods such as PCR. Thus, in the present study, we aimed to develop an additional Og-typing PCR system. Based on the novel Og-types, including OgN32, OgN33, and OgN34, presented in this study, we designed an additional 24 PCR primer pairs targeting 14 novel and 2 diversified E. coli Og-types and 8 Shigella-unique Og-types. Subsequently, we developed 5 new multiplex PCR sets consisting of 33 primers, including the aforementioned 24 primers and 9 primers reported in previous studies. The accuracy and specificity of the PCR system was validated using approximately 260 E. coli and Shigella O-serogroup and Og-type reference strains. The Og-typing PCR system reported here can determine a wide range of Og-types of E. coli and may help epidemiological studies, in addition to the surveillance of pathogenic E. coli.

scholarly journals A large-scale forest fragmentation experiment: the Stability of Altered Forest Ecosystems Project

2011 ◽  
Vol 366 (1582) ◽  
pp. 3292-3302 ◽  
Author(s):  
Robert M. Ewers ◽  
Raphael K. Didham ◽  
Lenore Fahrig ◽  
Gonçalo Ferraz ◽  
Andy Hector ◽  
...  
Keyword(s):  
Forest Fragmentation ◽  
Large Scale ◽  
Forest Cover ◽  
Forest Ecosystems ◽  
Field Experiments ◽  
Spatial Scales ◽  
Ecological Impacts ◽  
Data Types ◽  
Wide Range ◽  
The Stability

Opportunities to conduct large-scale field experiments are rare, but provide a unique opportunity to reveal the complex processes that operate within natural ecosystems. Here, we review the design of existing, large-scale forest fragmentation experiments. Based on this review, we develop a design for the Stability of Altered Forest Ecosystems (SAFE) Project, a new forest fragmentation experiment to be located in the lowland tropical forests of Borneo (Sabah, Malaysia). The SAFE Project represents an advance on existing experiments in that it: (i) allows discrimination of the effects of landscape-level forest cover from patch-level processes; (ii) is designed to facilitate the unification of a wide range of data types on ecological patterns and processes that operate over a wide range of spatial scales; (iii) has greater replication than existing experiments; (iv) incorporates an experimental manipulation of riparian corridors; and (v) embeds the experimentally fragmented landscape within a wider gradient of land-use intensity than do existing projects. The SAFE Project represents an opportunity for ecologists across disciplines to participate in a large initiative designed to generate a broad understanding of the ecological impacts of tropical forest modification.

scholarly journals What can we learn from over 100,000 Escherichia coli genomes?

2019 ◽  
Author(s):  
Kaleb Abram ◽  
Zulema Udaondo ◽  
Carissa Bleker ◽  
Visanu Wanchai ◽  
Trudy M. Wassenaar ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Large Scale ◽  
Phylogenetic Analyses ◽  
Bacterial Species ◽  
Single Copy ◽  
Gene Gain ◽  
E Coli ◽  
Phylogenetic Profiles ◽  
Genomic Studies ◽  
Gain Loss

ABSTRACTThe explosion of microbial genome sequences in public databases allows for large-scale population genomic studies of bacterial species, such as Escherichia coli. In this study, we examine and classify more than one hundred thousand E. coli and Shigella genomes. After removing outliers, a semi-automated Mash-based analysis of 10,667 assembled genomes reveals 14 distinct phylogroups. A representative genome or medoid identified for each phylogroup serves as a proxy to classify more than 95,000 unassembled genomes. This analysis shows that most sequenced E. coli genomes belong to 4 phylogroups (A, C, B1 and E2(O157)). Authenticity of the 14 phylogroups described is supported by pangenomic and phylogenetic analyses, which show differences in gene preservation between phylogroups. A phylogenetic tree constructed with 2,613 single copy core genes along with a matrix of phylogenetic profiles is used to confirm that the 14 phylogroups change at different rates of gene gain/loss/duplication. The methodology used in this work is able to identify previously uncharacterized phylogroups in E. coli species. Some of these new phylogroups harbor clonal strains that have undergone a process of genomic adaptation to the acquisition of new genomic elements related to virulence or antibiotic resistance. This is, to our knowledge, the largest E. coli genome dataset analyzed to date and provides valuable insights into the population structure of the species.

scholarly journals Molecular Epidemiology, Genetic Diversity, and Antifungal Susceptibility of Major Pathogenic Dermatophytes Isolated From Human Dermatophytosis

2021 ◽  
Vol 12 ◽  
Author(s):  
Zahra Salehi ◽  
Masoomeh Shams-Ghahfarokhi ◽  
Mehdi Razzaghi-Abyaneh
Keyword(s):  
Genetic Diversity ◽  
Phylogenetic Analyses ◽  
Antifungal Susceptibility ◽  
Elongation Factor ◽  
Trichophyton Mentagrophytes ◽  
Epidemiological Studies ◽  
Genetic Distances ◽  
Mlst Scheme ◽  
Low Genetic Diversity ◽  
Starting Point

BackgroundDermatophytes are a homogeneous group of species with low genetic diversity, and there are still many uncertainties about the boundaries among species.ObjectivesAiming at clarifying the relationships among species in the genus and introducing suitable genes for multilocus sequence typing (MLST), a new MLST scheme approach was developed to characterize the major pathogenic dermatophytes.MethodsWe performed maximum parsimony (MP), MrBayes, RAxML, and eBURST analyses, based on the MLST scheme to scrutinize the evolution within 95 clinical isolates and four reference strains belonging to the four major dermatophytes species. Then, the discriminatory power, pairwise genetic distances, ratio dN/dS, and sequence types (STs) of these isolates were determined. Also, to study taxonomy, sequences of the internal transcribed spacer (ITS), Beta-tubulin (BT2), and translation elongation factor 1-α (TEF-1α) genes of other dermatophytes species available in the GenBank were analyzed.ResultsFindings of the present study indicated that three genes: BT2, ITS, and TEF−1α, which showed the greatest diversity among dermatophyte species, were suitable for MLST. The most prevalent STs were seen among the species of Trichophyton interdigitale. Also, two new genotypes, i.e., XXVII and XXVIII, were introduced for T. interdigitale and Trichophyton mentagrophytes. The least informative sites were found in Epidermophyton floccosum, Trichophyton rubrum, and T. mentagrophytes, while the most informative sites were observed in T. interdigitale. Furthermore, the most informative locus was TEF-1α. The phylogenetic tree, constructed by the combination of the three genes, shows a new topological pattern that confirms the derivation of the anthropophilic and zoophilic genera from the geophilic genus. Also, the phylogenetic analyses and pairwise distances of the combination of the three loci showed that Trichophyton tonsurans and Trichophyton equinum were a species complex, where T. equinum is derived from T. tonsurans.ConclusionsResults of this study showed that MLST is very effective in determining the boundaries between species and taxonomy. Considering that there is no database for MLST dermatophytes, further studies are needed to determine the suitable genes for MLST. Also, the determination of STs in epidemiological studies and raising epidemiological information are helpful. This study was a new starting point to determine the ST and a foundation for a dermatophyte MLST database.

A study of mitochondrial DNA mutations in peripheral lymphocytes in an aging cohort

2003 ◽  
Vol 31 (2) ◽  
pp. 444-446 ◽  
Author(s):  
B. Zhang ◽  
S. Ye ◽  
A.A. Sayer ◽  
S.R. Hammans ◽  
S. Adio ◽  
...  
Keyword(s):  
Mitochondrial Dna ◽  
Mitochondrial Genome ◽  
Somatic Mutation ◽  
Large Scale ◽  
Nuclear Genome ◽  
Epidemiological Studies ◽  
Elderly Subjects ◽  
Long Pcr ◽  
Peripheral Lymphocytes ◽  
Wide Range

Somatic mutation in the mitochondrial genome occurs much more rapidly than in the nuclear genome and is a feature, possibly contributory, of the aging of cells and tissues. Identifying mitochondrial sequence changes in blood DNA of elderly subjects may provide a maker for the epigenetic changes of mitochondrial DNA known to occur in tissues with lower cellular turnover, and would also have implications for immunosenescence. No large-scale epidemiological studies have been reported previously. In this study we have established long-PCR banks of the mitochondrial genome from peripheral lymphocytes for an elderly cohort of 716 individuals with a range of measured aging phenotypes, and we have established assays for three widely reported mutations: the 4977 bp and 8048 bp deletions and point mutation A3243G. No individuals were identified with detectable heteroplasmy for these changes. Implications for tissue and population prevalence are discussed. The mitochondrial long-PCR DNA banks established will be useful for a wide range of studies of somatic mutation and of germline haplotypes in relation to aging.

scholarly journals Consensus Multilocus Sequence Typing Scheme for Pneumocystis jirovecii

2020 ◽  
Vol 6 (4) ◽  
pp. 259
Author(s):  
Lana Pasic ◽  
Lidia Goterris ◽  
Mercedes Guerrero-Murillo ◽  
Laszlo Irinyi ◽  
Alex Kan ◽  
...  
Keyword(s):  
Multilocus Sequence Typing ◽  
Large Scale ◽  
Pathogenic Fungus ◽  
Severe Pneumonia ◽  
Epidemiological Studies ◽  
Discriminatory Power ◽  
Pneumocystis Jirovecii ◽  
Mlst Scheme ◽  
Global Comparison ◽  
Scale Population

Pneumocystis jirovecii is an opportunistic human pathogenic fungus causing severe pneumonia mainly in immunocompromised hosts. Multilocus sequence typing (MLST) remains the gold standard for genotyping of this unculturable fungus. However, the lack of a consensus scheme impedes a global comparison, large scale population studies and the development of a global MLST database. To overcome this problem this study compared all genetic regions (19 loci) currently used in 31 different published Pneumocystis MLST schemes. The most diverse/commonly used eight loci, β-TUB, CYB, DHPS, ITS1, ITS1/2, mt26S and SOD, were further assess for their ability to be successfully amplified and sequenced, and for their discriminatory power. The most successful loci were tested to identify genetically related and unrelated cases. A new consensus MLST scheme consisting of four genetically independent loci: β-TUB, CYB, mt26S and SOD, is herein proposed for standardised P. jirovecii typing, successfully amplifying low and high fungal burden specimens, showing adequate discriminatory power, and correctly identifying suspected related and unrelated isolates. The new consensus MLST scheme, if accepted, will for the first time provide a powerful tool to investigate outbreak settings and undertake global epidemiological studies shedding light on the spread of this important human fungal pathogen.

scholarly journals RevGadgets: an R Package for visualizing Bayesian phylogenetic analyses from RevBayes

2021 ◽  
Author(s):  
Carrie M. Tribble ◽  
William A. Freyman ◽  
Jun Ying Lim ◽  
Michael J. Landis ◽  
Joëlle Barido-Sottani ◽  
...  
Keyword(s):  
Open Source ◽  
Phylogenetic Analyses ◽  
R Package ◽  
Epidemiological Studies ◽  
Use Cases ◽  
Software Packages ◽  
Phylogenetic Methods ◽  
Wide Range ◽  
Publication Quality

1. Statistical phylogenetic methods are the foundation for a wide range of evolutionary and epidemiological studies. However, as these methods grow increasingly complex, users often encounter significant challenges with summarizing, visualizing, and communicating their key results. 2. We present RevGadgets, an R package for creating publication-quality figures from the results of a large variety of phylogenetic analyses performed in RevBayes (and other phylogenetic software packages). 3. We demonstrate how to use RevGadgets through a set of vignettes that cover the most common use cases that researchers will encounter. 4. RevGadgets is an open-source, extensible package that will continue to evolve in parallel with RevBayes, helping researchers to make sense of and communicate the results of a diverse array of analyses.

scholarly journals Complete Genome Sequence of Escherichia Coli Strain S9922 Isolated from Meningitis in Calf and Comparative Genomic Analysis with Other E. Coli Pathotypes

2021 ◽  
Author(s):  
Beibei Li ◽  
Jingjing Ren ◽  
Xun Ma ◽  
Qian Qin ◽  
Xinyu Wang ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Cluster Analysis ◽  
Large Scale ◽  
Virulent Strain ◽  
Evolutionary Relationship ◽  
Intestinal Flora ◽  
Genomic Analysis ◽  
Comparative Genomic ◽  
E Coli ◽  
Wide Range

Abstract Background: Extraintestinal pathogenic Escherichia coli (ExPEC) exists in the normal intestinal flora, but can invade and colonize extraintestinal sites and cause a wide range of infections. Genomic analysis of ExPEC has mainly focused on isolates of human, poultry and pig. In recent years, some large-scale dairy farms in Xinjiang broke out cases characterized by neurological symptoms and acute death in newborn calves. To better understand the genomic attributes underlying the pathogenicity of bovine-source ExPEC, a highly virulent strain, which named E. coli S9922 was isolated from cerebral effusion in a calf that died of meningitis, was sequenced and analyzed.Results: Using single-molecule sequencing technology on PacBio and then assembled, the genes were predicted and annotated. The whole genome of E.coli S9922 was consisted of a chromosome and three plasmids containing 5055 genes, and the total length was 5269374 bp and the average G+C content was 50.82%. In addition, 291 host-, 204 virulence-, and 185 resistance-related genes, and 182 T3SS effector proteins were found by comparison with related databases. Comparison of this genome to 16 representative strains of pathogenic E.coli genomic sequences showed that E.coli S9922 had the greatest co-linearity with E.coli 90-9272. In addition, Core genes obtained by cluster analysis of E.coli S9922 homologous genes were classified, a total of 2570, 2780, and 2188 genes were obtained via COG, KEGG, and GO comparisons, respectively. The unique genes identified by homologous cluster analysis were classified 204, 550, 239 genes in COG, KEGG, and GO comparisons, respectively. Evolutionary tree analysis revealed a close evolutionary relationship between E.coli S9922 and E.coli 90-9272, and a distant relationship between E.coli S9922 and UTI89.Conclusions: The study provide dgenomics of E.coli S9922 strain from the cattle that had died of meningitis. It enriched the genome data of E.coli and laid a theoretical foundation for further experimental study of ExPEC. Comparative genomics analysis showed that E.coli S9922 had a close evolutionary relationship with E.coli 90-9272, but far from that of UTI89.

scholarly journals The Phosphopantetheinyl Transferase Superfamily: Phylogenetic Analysis and Functional Implications in Cyanobacteria

2006 ◽  
Vol 72 (4) ◽  
pp. 2298-2305 ◽  
Author(s):  
J. N. Copp ◽  
B. A. Neilan
Keyword(s):  
Large Scale ◽  
Acyl Carrier Protein ◽  
Sequence Similarity ◽  
Phylogenetic Analyses ◽  
Combinatorial Biosynthesis ◽  
Ex Situ ◽  
Phosphopantetheinyl Transferase ◽  
Phylogenetic Studies ◽  
Degenerate Oligonucleotide ◽  
Wide Range

ABSTRACT Phosphopantetheinyl transferases (PPTs) are a superfamily of essential enzymes required for the synthesis of a wide range of compounds including fatty acid, polyketide, and nonribosomal peptide metabolites. These enzymes activate carrier proteins in specific biosynthetic pathways by the transfer of a phosphopantetheinyl moiety to an invariant serine residue. PPTs display low levels of sequence similarity but can be classified into two major families based on several short motifs. The prototype of the first family is the broad-substrate-range PPT Sfp, which is required for biosynthesis of surfactin in Bacillus subtilis. The second family is typified by the Escherichia coli acyl carrier protein synthase (AcpS). Facilitated by the growing number of genome sequences available for analyses, large-scale phylogenetic studies were utilized in this research to reveal novel subfamily groupings, including two subfamilies within the Sfp-like family. In the present study degenerate oligonucleotide primers were designed for amplification of cyanobacterial PPT gene fragments. Subsequent phylogenetic analyses suggested a unique, function-based PPT type, defined by the PPTs involved in heterocyst differentiation. Evidence supporting this hypothesis was obtained by sequencing the region surrounding the partial Nodularia spumigena PPT gene. The ability to genetically classify PPT function is critical for the engineering of novel compounds utilizing combinatorial biosynthesis techniques. Information regarding cyanobacterial PPTs has important ramifications for the ex situ production of cyanobacterial natural products.

scholarly journals Atypical Clinical Manifestations of Loiasis and Their Relevance for Endemic Populations

2019 ◽  
Vol 6 (11) ◽  
Author(s):  
Kevin G Buell ◽  
Charles Whittaker ◽  
Cédric B Chesnais ◽  
Paul D Jewell ◽  
Sébastien D S Pion ◽  
...  
Keyword(s):  
Large Scale ◽  
Case Reports ◽  
Clinical Manifestations ◽  
Central Africa ◽  
Epidemiological Studies ◽  
True Rate ◽  
Public Health Burden ◽  
Wide Range ◽  
Atypical Manifestations ◽  
Atypical Presentations

Abstract Background Loiasis is mostly considered a relatively benign infection when compared with other filarial and parasitic diseases, with Calabar swellings and eyeworm being the most common signs. Yet, there are numerous reports in the literature of more serious sequelae. Establishing the relationship between infection and disease is a crucial first step toward estimating the burden of loiasis. Methods We conducted a systematic review of case reports containing 329 individuals and detailing clinical manifestations of loiasis with a focus on nonclassical, atypical presentations. Results Results indicate a high proportion (47%) of atypical presentations in the case reports identified, encompassing a wide range of cardiac, respiratory, gastrointestinal, renal, neurological, ophthalmological, and dermatological pathologies. Individuals with high microfilarial densities and residing in an endemic country were at greater risk of suffering from atypical manifestations. Conclusions Our findings have important implications for understanding the clinical spectrum of conditions associated with Loa loa infection, which extends well beyond the classical eyeworm and Calabar swellings. As case reports may overestimate the true rate of atypical manifestations in endemic populations, large-scale, longitudinal clinico-epidemiological studies will be required to refine our estimates and demonstrate causality between loiasis and the breadth of clinical manifestations reported. Even if the rates of atypical presentations were found to be lower, given that residents of loiasis-endemic areas are both numerous and the group most at risk of severe atypical manifestations, our conclusions support the recognition of loiasis as a significant public health burden across Central Africa.

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