Genomic background of the Klebsiella pneumoniae NDM-1 outbreak in Poland, 2012–18

2020 ◽  
Vol 75 (11) ◽  
pp. 3156-3162
Author(s):  
R Izdebski ◽  
M Sitkiewicz ◽  
P Urbanowicz ◽  
M Krawczyk ◽  
S Brisse ◽  
...  
Keyword(s):  
Phylogenetic Analysis ◽  
Klebsiella Pneumoniae ◽  
Core Genome ◽  
Illumina Miseq ◽  
Plasmid Analysis ◽  
Homogeneous Cluster ◽  
Oxford Nanopore ◽  
O Antigens ◽  
Oxford Nanopore Technologies ◽  
Plasmid Replicons

Abstract Objectives To characterize genomes of Klebsiella pneumoniae ST11 NDM-1 responsible for a countrywide outbreak in Poland and compare them phylogenetically with other Polish and international ST11 strains. Methods Seventy-one carbapenemase-producing K. pneumoniae ST11 isolates from Poland, including 66 representatives of the NDM-1 epidemic from 2012–18, were sequenced using Illumina MiSeq. Additionally, three outbreak isolates were also sequenced using MinION. The clonality and phylogenetic analysis was done by core-genome MLST and SNP approaches. Resistomes, virulomes, K/O antigens and plasmid replicons were screened for. The detailed plasmid analysis was based on full assemblies using Oxford Nanopore Technologies data. Results Chromosomes of the outbreak isolates formed an essentially homogeneous cluster (though accumulating SNPs gradually with time), differing remarkably from other Polish NDM-1/-5-, KPC-2- or OXA-48-producing K. pneumoniae ST11 strains. The cluster belonged to a clade with 72 additional isolates identified worldwide, including closely related NDM-1 producers from several countries, including organisms from Bulgaria and Greece. All these had KL24 and O2v1 antigens and the chromosomal yersiniabactin locus YbST230 residing in the ICEKp11 element. The specific blaNDM-1-carrying Tn125 transposon derivative, named Tn125A, was located in IncFII/pKPX-1- and/or IncR-like plasmids; however, the IncRs rearranged extensively during the outbreak, contributing to highly dynamic plasmid profiles and resistomes. Conclusions The K. pneumoniae ST11 NDM-1 genotype that has been expanding in Poland since 2012 is largely monoclonal and represents a novel international high-risk lineage that is also spreading in other countries.

scholarly journals De-novo Assembly of Limnospira fusiformis Using Ultra-Long Reads

2021 ◽  
Vol 12 ◽  
Author(s):  
McKenna Hicks ◽  
Thuy-Khanh Tran-Dao ◽  
Logan Mulroney ◽  
David L. Bernick
Keyword(s):  
Phylogenetic Analysis ◽  
Type Strain ◽  
Reference Genome ◽  
De Novo ◽  
Illumina Miseq ◽  
Long Reads ◽  
Oxford Nanopore ◽  
Long Read ◽  
Oxford Nanopore Technologies ◽  
Rdna Analysis

The Limnospira genus is a recently established clade that is economically important due to its worldwide use in biotechnology and agriculture. This genus includes organisms that were reclassified from Arthrospira, which are commercially marketed as “Spirulina.” Limnospira are photoautotrophic organisms that are widely used for research in nutrition, medicine, bioremediation, and biomanufacturing. Despite its widespread use, there is no closed genome for the Limnospira genus, and no reference genome for the type strain, Limnospira fusiformis. In this work, the L. fusiformis genome was sequenced using Oxford Nanopore Technologies MinION and assembled using only ultra-long reads (>35 kb). This assembly was polished with Illumina MiSeq reads sourced from an axenic L. fusiformis culture; axenicity was verified via microscopy and rDNA analysis. Ultra-long read sequencing resulted in a 6.42 Mb closed genome assembled as a single contig with no plasmid. Phylogenetic analysis placed L. fusiformis in the Limnospira clade; some Arthrospira were also placed in this clade, suggesting a misclassification of these strains. This work provides a fully closed and accurate reference genome for the economically important type strain, L. fusiformis. We also present a rapid axenicity method to isolate L. fusiformis. These contributions enable future biotechnological development of L. fusiformis by way of genetic engineering.

scholarly journals Real-Time Sequencing of Mycobacterium tuberculosis: Are We There Yet?

2017 ◽  
Vol 55 (5) ◽  
pp. 1249-1254 ◽  
Author(s):  
Robyn S. Lee ◽  
Madhukar Pai
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Real Time ◽  
Phylogenetic Trees ◽  
Illumina Miseq ◽  
Turnaround Time ◽  
Epidemiologic Studies ◽  
Content Type ◽  
Leading Role ◽  
Oxford Nanopore ◽  
Oxford Nanopore Technologies

ABSTRACT Whole-genome sequencing has taken a leading role in epidemiologic studies of tuberculosis, but thus far, its real-time clinical utility has been low, in part because of the requirement for culture. In their report in this issue, Votintseva et al. (A. A. Votintseva, P. Bradley, L. Pankhurst, C. del Ojo Elias, M. Loose, K. Nilgiriwala, A. Chatterjee, E. G. Smith, N. Sanderson, T. M. Walker, M. R. Morgan, D. H. Wyllie, A. S. Walker, T. E. A. Peto, D. W. Crook, and Z. Iqbal, J Clin Microbiol 55:1285–1298, 2017, https://doi.org/10.1128/JCM.02483-16 ) present a new method for extracting Mycobacterium tuberculosis DNA directly from smear-positive respiratory samples, making it feasible to generate drug resistance predictions and phylogenetic trees in 44 h with the Illumina MiSeq. They also illustrate the potential for a <24-h turnaround time from DNA extraction to clinically relevant results with Illumina MiniSeq and Oxford Nanopore Technologies MinION. We comment on the promise and limitations of these approaches.

scholarly journals Complete Genome Sequence of the Type Strain Pectobacterium punjabense SS95, Isolated from a Potato Plant with Blackleg Symptoms

2020 ◽  
Vol 9 (32) ◽  
Author(s):  
Sohaib Sarfraz ◽  
Saïd Oulghazi ◽  
Jérémy Cigna ◽  
Shahbaz Talib Sahi ◽  
Kashif Riaz ◽  
...  
Keyword(s):  
Complete Genome ◽  
Type Strain ◽  
Gc Content ◽  
Illumina Miseq ◽  
Blackleg Disease ◽  
Circular Chromosome ◽  
Content Type ◽  
Potato Plants ◽  
Oxford Nanopore ◽  
Oxford Nanopore Technologies

ABSTRACT Pectobacterium punjabense is a newly described species causing blackleg disease in potato plants. Therefore, by the combination of long (Oxford Nanopore Technologies, MinION) and short (Illumina MiSeq) reads, we sequenced the complete genome of P. punjabense SS95T, which contains a circular chromosome of 4.793 Mb with a GC content of 50.7%.

scholarly journals A nationwide genomic study of clinical Klebsiella pneumoniae in Norway 2001-2015: Introduction and spread of ESBL facilitated by CG15 and CG307

2021 ◽  
Author(s):  
Aasmund Fostervold ◽  
Marit A.K. Hetland ◽  
Ragna-Johanne Bakksjo ◽  
Eva Bernhoff ◽  
Kathryn Holt ◽  
...  
Keyword(s):  
Klebsiella Pneumoniae ◽  
Multidrug Resistant ◽  
Sensu Stricto ◽  
Whole Genome Sequence ◽  
Surveillance Program ◽  
Esbl Production ◽  
Resistance Determinants ◽  
Oxford Nanopore ◽  
Genomic Study ◽  
Plasmid Replicons

Objective: We have used the nationwide Norwegian surveillance program on resistant microbes in humans (NORM) to address longitudinal changes in the population structure K. pneumoniae isolates during 2001-15, encompassing the emergence and spread of ESBL-producing Enterobacterales (ESBL-E) in Norway. Material and methods: Among blood (n= 6124) and urinary tract (n=5496) surveillance isolates from 2001-15, we used Illumina technology to whole genome sequence 201 ESBL-producing isolates from blood (n=130) and urine (n=71), and 667 non-ESBL isolates from blood. Complete genomes for four isolates were resolved with Oxford Nanopore sequencing. Results: In a highly diverse collection, Klebsiella variicola ssp. variicola caused a quarter of Klebsiella pneumoniae species complex bacteraemias. ESBL-production was limited to K. pneumoniae sensu stricto (98.5 %). A diverse ESBL population of 57 clonal groups (CGs) were dominated by multidrug resistant CG307 (17%), CG15 (12%), CG70 (6%), CG258 (5%) and CG45 (5%) carrying blaCTX-M-15. Yersiniabactin was significantly more common in ESBL-positive (37.8%) compared to non-ESBL K. pneumoniae sensu stricto isolates (12.7%), indicating convergence of virulence and resistance determinants. Moreover, we found a significant lower prevalence of yersinabactin (3.0 %, 37.8 % and 17.3 %), IncFIB (58.7 %, 87.9 % and 79.4 %) and IncFII plasmid replicons (40.5 %, 82.8 % and 54.2%) in K. variicola ssp. variicola compared to ESBL- and non-ESBL K. pneumoniae sensu stricto, respectively. Conclusion: The increase in Norwegian KpSC ESBLs during 2010-15 was driven by blaCTX-M-15 carrying CG307 and CG15. K. variicola ssp. variicola was a frequent cause of invasive KpSC infection, but rarely carried ESBL.

scholarly journals Diversity of carbapenem-resistant Klebsiella pneumoniae ST14 and emergence of a subgroup with KL64 capsular locus in the Arabian Peninsula

2021 ◽  
Author(s):  
Shaimaa F. Mouftah ◽  
Tibor Pál ◽  
Paul G. Higgins ◽  
Akela Ghazawi ◽  
Youssef Idaghdour ◽  
...  
Keyword(s):  
Saudi Arabia ◽  
Klebsiella Pneumoniae ◽  
United Arab Emirates ◽  
Core Genome ◽  
Arabian Peninsula ◽  
Illumina Miseq ◽  
Carbapenem Resistant ◽  
Distinct Cluster ◽  
Recent Emergence ◽  
Miseq Platform

AbstractTo understand the reasons of successful spread of carbapenem-resistant Klebsiella pneumoniae ST14 (CRKP-ST14) in countries of the Arabian Peninsula, the resistome, capsular locus, carbapenemase carrying plasmid types, and core genome of isolates from the region were compared to global isolates. Thirty-nine CRKP-ST14 strains isolated from 13 hospitals in the United Arab Emirates, Bahrain, and Saudi Arabia were selected for whole genome sequencing on Illumina MiSeq platform based on the variety of carbapenemase genes carried and plasmids bearing these genes. Their resistome, capsular locus, and core genome MLST were compared to 173 CRKP-ST14 genomes available in public databases. The selected 39 CRKP-ST14 produced either NDM-1, OXA-48, OXA-162, OXA-232, KPC-2, or co-produced NDM-1 and an OXA-48-like carbapenemase. cgMLST revealed three clusters: 16 isolates from five UAE cities (C1), 11 isolates from three UAE cities and Bahrain (C2), and 5 isolates from Saudi Arabia (C3), respectively, and seven singletons. Resistance gene profile, carbapenemase genes, and their plasmid types were variable in both C1 and C2 clusters. The majority of CRKP-ST14 had KL2, but members of the C2 cluster and two further singletons possessed KL64 capsular locus. Based on cgMLST comparison of regional and global isolates, CRKP-ST14 with KL64 from four continents formed a distinct cluster, suggesting a recent emergence and spread of this variant. Our findings confirmed clonal transmission coupled with likely horizontal gene transfer in carbapenem-resistant Klebsiella pneumoniae ST14. Dissemination of this genetically flexible, highly resistant clone warrants further monitoring.

scholarly journals Metagenomic data for Halichondria panicea from Illumina and Nanopore sequencing and preliminary genome assemblies for the sponge and two microbial symbionts.

2021 ◽  
Author(s):  
Brian W Strehlow ◽  
Astrid Schuster ◽  
Warren R Francis ◽  
Donald E Canfield
Keyword(s):  
Additional Data ◽  
Illumina Miseq ◽  
Metagenomic Data ◽  
Single Individual ◽  
Halichondria Panicea ◽  
Oxford Nanopore ◽  
Long Read ◽  
Microbial Symbionts ◽  
Genome Assemblies ◽  
Oxford Nanopore Technologies

Objectives: These data were collected to generate a novel reference metagenome for the sponge Halichondria panicea and its microbiome for subsequent differential expression analyses. Data description: These data include raw sequences from four separate sequencing runs of the metagenome of a single individual of H. panicea - one Illumina MiSeq (2x300 bp, paired-end) run and three Oxford Nanopore Technologies (ONT) long-read sequencing runs, generating 53.8 and 7.42 Gbp respectively. Comparing assemblies of Illumina, ONT and an Illumina-ONT hybrid revealed the hybrid to be the best assembly, comprising 163 Mbp in 63,555 scaffolds (N50: 3,084). This assembly, however, was still highly fragmented and only contained 52% of core metazoan genes (with 77.9% partial genes), so it was also not complete. However, this sponge is an emerging model species for field and laboratory work, and there is considerable interest in genomic sequencing of this species. Although the resultant assemblies from the data presented here are suboptimal, this data note can inform future studies by providing an estimated genome size and coverage requirements for future sequencing, sharing additional data to potentially improve other suboptimal assemblies of this species, and outlining potential limitations and pitfalls of the combined Illumina and ONT approach to novel genome sequencing.

scholarly journals Identification and Whole-Genome Sequencing of a Monkeypox Virus Strain Isolated in Israel

2020 ◽  
Vol 9 (10) ◽  
Author(s):  
Inbar Cohen-Gihon ◽  
Ofir Israeli ◽  
Ohad Shifman ◽  
Noam Erez ◽  
Sharon Melamed ◽  
...  
Keyword(s):  
West Africa ◽  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
Virus Strain ◽  
Illumina Miseq ◽  
Whole Genome Sequence ◽  
Whole Genome ◽  
Monkeypox Virus ◽  
Oxford Nanopore ◽  
Oxford Nanopore Technologies

We report the whole-genome sequence of a monkeypox virus strain isolated in Israel. The strain was isolated in 2018 from a patient travelling back from West Africa. The virus was fully sequenced on the Illumina MiSeq and Oxford Nanopore Technologies MinION platforms.

scholarly journals Plasmid analysis of NDM metallo-β-lactamase-producing Enterobacterales isolated in Vietnam

PLoS ONE ◽  
2021 ◽  
Vol 16 (7) ◽  
pp. e0231119
Author(s):  
Aki Hirabayashi ◽  
Koji Yahara ◽  
Satomi Mitsuhashi ◽  
So Nakagawa ◽  
Tadashi Imanishi ◽  
...  
Keyword(s):  
Carbapenem Resistance ◽  
Hybrid Plasmid ◽  
Genomic Epidemiology ◽  
Carbapenem Resistant ◽  
Plasmid Analysis ◽  
Oxford Nanopore ◽  
Carbapenemase Gene ◽  
Long Read ◽  
Third Generation Cephalosporins ◽  
Oxford Nanopore Technologies

Carbapenem-resistant Enterobacterales (CRE) represent a serious threat to public health due to the lack of treatment and high mortality. The rate of antimicrobial resistance of Enterobacterales isolates to major antimicrobials, including carbapenems, is much higher in Vietnam than in Western countries, but the reasons remain unknown due to the lack of genomic epidemiology research. A previous study suggested that carbapenem resistance genes, such as the carbapenemase gene blaNDM, spread via plasmids among Enterobacterales in Vietnam. In this study, we characterized blaNDM-carrying plasmids in Enterobacterales isolated in Vietnam, and identified several possible cases of horizontal transfer of plasmids both within and among species of bacteria. Twenty-five carbapenem-nonsusceptible isolates from a medical institution in Hanoi were sequenced on Illumina short-read sequencers, and 13 blaNDM-positive isolates, including isolates of Klebsiella pneumoniae, Escherichia coli, Citrobacter freundii, Morganella morganii, and Proteus mirabilis, were further sequenced on an Oxford Nanopore Technologies long-read sequencer to obtain complete plasmid sequences. Almost identical 73 kb IncFII(pSE11)::IncN hybrid plasmids carrying blaNDM-1 were found in a P. mirabilis isolate and an M. morganii isolate. A 112 kb IncFII(pRSB107)::IncN hybrid plasmid carrying blaNDM-1 in an E. coli isolate had partially identical sequences with a 39 kb IncR plasmid carrying blaNDM-1 and an 88 kb IncFII(pHN7A8)::IncN hybrid plasmid in a C. freundii isolate. 148–149 kb IncFIA(Hl1)::IncA/C2 plasmids and 75–76 kb IncFII(Yp) plasmids, both carrying blaNDM-1 were shared among three sequence type 11 (ST11) isolates and three ST395 isolates of K. pneumoniae, respectively. Most of the plasmids co-carried genes conferring resistance to clinically relevant antimicrobials, including third-generation cephalosporins, aminoglycosides, and fluoroquinolones, in addition to blaNDM-1. These results provide insight into the genetic basis of CRE in Vietnam, and could help control nosocomial infections.

scholarly journals Complete and Circularized Genome Assemblies of the Kroppenstedtia eburnea Genus Type Strain and the Kroppenstedtia pulmonis Species Type Strain with MiSeq and MinION Sequence Data

2020 ◽  
Vol 9 (44) ◽  
Author(s):  
Christopher A. Gulvik ◽  
Dhwani Batra ◽  
Lori A. Rowe ◽  
Milli Sheth ◽  
Ben W. Humrighouse ◽  
...  
Keyword(s):  
Type Strain ◽  
Sequence Data ◽  
Gene Clusters ◽  
Illumina Miseq ◽  
Putative Gene ◽  
Content Type ◽  
Oxford Nanopore ◽  
Species Type ◽  
Genome Assemblies ◽  
Oxford Nanopore Technologies

ABSTRACT Kroppenstedtia eburnea DSM 45196T and Kroppenstedtia pulmonis W9323T are aerobic, Gram-positive, filamentous, chemoorganotrophic thermoactinomycetes. Here, we report on the complete and circular genome assemblies generated using Illumina MiSeq and Oxford Nanopore Technologies MinION reads. Putative gene clusters predicted to be involved in the production of secondary metabolites were also identified.

scholarly journals Spirochetes isolated from arthropods constitute a novel genus Entomospira genus novum within the order Spirochaetales

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Lucía Graña-Miraglia ◽  
Silvie Sikutova ◽  
Marie Vancová ◽  
Tomáš Bílý ◽  
Volker Fingerle ◽  
...  
Keyword(s):  
Culex Pipiens ◽  
Type Species ◽  
Gc Content ◽  
Illumina Miseq ◽  
Genomic Information ◽  
Orthologous Genes ◽  
The Czech Republic ◽  
Oxford Nanopore ◽  
The Family ◽  
Oxford Nanopore Technologies

Abstract Spirochetal bacteria were successfully isolated from mosquitoes (Culex pipiens, Aedes cinereus) in the Czech Republic between 1999 and 2002. Preliminary 16S rRNA phylogenetic sequence analysis showed that these strains differed significantly from other spirochetal genera within the family Spirochaetaceae and suggested a novel bacterial genus in this family. To obtain more comprehensive genomic information of these isolates, we used Illumina MiSeq and Oxford Nanopore technologies to sequence four genomes of these spirochetes (BR151, BR149, BR193, BR208). The overall size of the genomes varied between 1.68 and 1.78 Mb; the GC content ranged from 38.5 to 45.8%. Draft genomes were compared to 36 publicly available genomes encompassing eight genera from the class Spirochaetes. A phylogeny generated from orthologous genes across all taxa and the percentage of conserved proteins (POCP) confirmed the genus status of these novel spirochetes. The genus Entomospira gen. nov. is proposed with BR151 selected as type species of the genus. For this isolate and the closest related isolate, BR149, we propose the species name Entomospira culicis sp. nov. The two other isolates BR208 and BR193 are named Entomospira nematocera sp. nov. (BR208) and Entomospira entomophilus sp. nov. (BR193). Finally, we discuss their interesting phylogenetic positioning.

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