Comparing Long-Read Assemblers to Explore the Potential of a Sustainable Low-Cost, Low-Infrastructure Approach to Sequence Antimicrobial Resistant Bacteria With Oxford Nanopore Sequencing.
Comparing Long-Read Assemblers to Explore the Potential of a Sustainable Low-Cost, Low-Infrastructure Approach to Sequence Antimicrobial Resistant Bacteria With Oxford Nanopore Sequencing.
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DOI：
10.3389/fmicb.2022.796465
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发表时间：
2022
影响因子：
5.2
通讯作者：
Sands K
中科院分区：
文献类型：
--
作者：
Boostrom I;Portal EAR;Spiller OB;Walsh TR;Sands K
长读段测序（LRS）可以解析重复区域，这是短读段（SR）数据的限制。成本和仪器尺寸的降低导致了诊断和研究中LRS的稳步增长。在这里，我们重新对2018年至2021年之间的FAST 5数据进行了测序，并在一个大型数据集（n = 200）中分析了与gDNA相关的数据，该数据集涵盖了广泛的GC含量（25-67%）。我们检查了重新碱基调用的数据是否会改善杂交组装，并且对于较小的队列，在抗菌素耐药（AMR）基因和移动的遗传元件的背景下比较了长读段（LR）组装。在比较SR和LR工具时，我们进行了成本分析。我们比较了R9和R10化学，并报告了R9流动池不仅产量更高，而且读取质量更高。在LR组装中，ARG存在/不存在和/或变体检测经常存在差异。基于果蝇的组装通常在检测染色体和质粒上ARG的存在方面是有效的。Raven的速度更快，但回收的小质粒不一致，特别是携带blaKPC的15-kb Col样质粒。卡努组装是最碎片化的，基因组大小大于预期。LR组装件未能一致地测定Unicycler参考文献所鉴定的相同ARG的多个拷贝。即使对ONT化学和碱基判定进行了改进，长读段组装也可能导致对数据的误解。如果当前依赖LR数据，则有必要执行多个组装，尽管这是资源（计算）密集型的并且还不容易获得/使用。
Long-read sequencing (LRS) can resolve repetitive regions, a limitation of short read (SR) data. Reduced cost and instrument size has led to a steady increase in LRS across diagnostics and research. Here, we re-basecalled FAST5 data sequenced between 2018 and 2021 and analyzed the data in relation to gDNA across a large dataset (n = 200) spanning a wide GC content (25–67%). We examined whether re-basecalled data would improve the hybrid assembly, and, for a smaller cohort, compared long read (LR) assemblies in the context of antimicrobial resistance (AMR) genes and mobile genetic elements. We included a cost analysis when comparing SR and LR instruments. We compared the R9 and R10 chemistries and reported not only a larger yield but increased read quality with R9 flow cells. There were often discrepancies with ARG presence/absence and/or variant detection in LR assemblies. Flye-based assemblies were generally efficient at detecting the presence of ARG on both the chromosome and plasmids. Raven performed more quickly but inconsistently recovered small plasmids, notably a ∼15-kb Col-like plasmid harboring blaKPC. Canu assemblies were the most fragmented, with genome sizes larger than expected. LR assemblies failed to consistently determine multiple copies of the same ARG as identified by the Unicycler reference. Even with improvements to ONT chemistry and basecalling, long-read assemblies can lead to misinterpretation of data. If LR data are currently being relied upon, it is necessary to perform multiple assemblies, although this is resource (computing) intensive and not yet readily available/useable.
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DOI：
10.1093/jac/dkaa345
发表时间：
2020-12-01
期刊：
The Journal of antimicrobial chemotherapy
影响因子：
--
作者：
Bortolaia V;Kaas RS;Ruppe E;Roberts MC;Schwarz S;Cattoir V;Philippon A;Allesoe RL;Rebelo AR;Florensa AF;Fagelhauer L;Chakraborty T;Neumann B;Werner G;Bender JK;Stingl K;Nguyen M;Coppens J;Xavier BB;Malhotra-Kumar S;Westh H;Pinholt M;Anjum MF;Duggett NA;Kempf I;Nykäsenoja S;Olkkola S;Wieczorek K;Amaro A;Clemente L;Mossong J;Losch S;Ragimbeau C;Lund O;Aarestrup FM
通讯作者：
Aarestrup FM
影响因子：
12.3
作者：
Huang YT;Liu PY;Shih PW
通讯作者：
Shih PW
影响因子：
4.6
作者：
Castro-Wallace SL;Chiu CY;John KK;Stahl SE;Rubins KH;McIntyre ABR;Dworkin JP;Lupisella ML;Smith DJ;Botkin DJ;Stephenson TA;Juul S;Turner DJ;Izquierdo F;Federman S;Stryke D;Somasekar S;Alexander N;Yu G;Mason CE;Burton AS
通讯作者：
Burton AS
影响因子：
9.2
作者：
Nicholls, Samuel M.;Quick, Joshua C.;Loman, Nicholas J.
通讯作者：
Loman, Nicholas J.
QualiMap 2：高通量测序数据的高级多样本质量控制。
DOI：
10.1093/bioinformatics/btv566
发表时间：
2016-01-15
期刊：
Bioinformatics (Oxford, England)
影响因子：
--
作者：
Okonechnikov K;Conesa A;García-Alcalde F
通讯作者：
García-Alcalde F