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Seqsphere 3

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SeqSphere+3.5.0 is a software for performing comparative sequence analysis. It provides tools for managing, analyzing, and visualizing genome sequence data.

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SeqSphere+

4 protocols using seqsphere 3

1

Multilocus Sequence Typing for Klebsiella pneumoniae

Cited 1 time
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STs were calculated according to multilocus sequence typing (MLST) schemes of the Institut Pasteur using Ariba v2.6.2 [39 (link)]. A simple diversity index (SDI) [40 (link)] was applied to analyze population diversity. A core genome multilocus sequence typing (cgMLST) that relies on species-specific schemes with a fixed number of chromosomal target genes was applied, consisting of 2538 K. pneumoniae targets provided by SeqSphere+ 3.5.0 (Ridom, Münster, Germany). A relatedness threshold of ≤10 alleles was applied for detecting related isolates, as recommended [10 (link)].
Cañada-García J.E., Ramírez de Arellano E., Jiménez-Orellana M., Viedma E., Sánchez A., Alhambra A., Villa J., Delgado-Iribarren A., Bautista V., Lara N., García-Cobos S., Aracil B., Cercenado E., Pérez-Vázquez M, & Oteo-Iglesias J. (2023). Carbapenemase-Producing Klebsiella pneumoniae in COVID-19 Intensive Care Patients: Identification of IncL-VIM-1 Plasmid in Previously Non-Predominant Sequence Types. Antibiotics, 12(1), 107.
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2

Genomic Analysis of ESBL and CPE in LTCFs

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Next-generation sequencing (NGS) was performed using a standardized protocol. 22 To identify the presence of resistance genes, ResFinder 2.1 23 was used at a threshold of 90% identity and 60% minimum matching length. MLST and whole-genome MLST (wgMLST) of E. coli and K. pneumoniae isolates were performed using SeqSphere 3.5.0 (Ridom GmbH, Mu ¨nster, Germany) using in-house wgMLST schemes. 21 All genotyping data were imported into a BioNumerics database for further analyses (BioNumerics v7.6, Applied Maths, Sint-Martens-Latem, Belgium). The wgMLST allelic profiles were compared in advanced cluster analyses as categorical data, ignoring missing genes. Based on analyses of multiple isolates obtained from the same patient and on isolates from patients in proven transmission events, we considered K. pneumoniae differing in no more than 20 wgMLST genes as the same strain and for E. coli we used a 25 wgMLST gene cut-off. For multiple indistinguishable isolates obtained from the same resident, only one isolate was included in further analyses.
ESBL and CPE carriage in Dutch long-term care facilities
van Kleef E., Wielders C.C.H., Schouls L.M., Feenstra S.G., Hertogh C.M.P.M., Bonten M.J.M., van Weert Y., Tostmann A., van der Lubben M, & de Greeff S.C. (2021). National point prevalence study on carriage of multidrug-resistant microorganisms in Dutch long-term care facilities in 2018. The Journal of antimicrobial chemotherapy, 76(6).
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3

Multilocus Sequence Typing of S. aureus

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Sequence types (STs) were calculated according to the multilocus sequence typing (MLST) scheme of the Public databases for molecular typing and microbial genome diversity (PubMLST)3 using Ariba v2.6.2 (Hunt et al., 2017 (link)). A simple diversity index (SDI; Gastmeier et al., 2006 (link)) was applied to analyze population diversity. Core genome MLST (cgMLST), consisting of 1861 targets for S. aureus provided by SeqSphere+3.5.0 (Ridom, Münster, Germany), was performed with the 137 sequenced isolates. Additionally, ST398 isolates from this study were analyzed using cgMLST together with a collection of 239 S. aureus of this ST downloaded from the NCBI database using “chromosome” and “complete” as filtering criteria for assembly level and the absence of mec genes as the genotypic criterion.
El Mammery A., Ramírez de Arellano E., Cañada-García J.E., Cercenado E., Villar-Gómara L., Casquero-García V., García-Cobos S., Lepe J.A., Ruiz de Gopegui Bordes E., Calvo-Montes J., Larrosa Escartín N., Cantón R., Pérez-Vázquez M., Aracil B, & Oteo-Iglesias J. (2023). An increase in erythromycin resistance in methicillin-susceptible Staphylococcus aureus from blood correlates with the use of macrolide/lincosamide/streptogramin antibiotics. EARS-Net Spain (2004–2020). Frontiers in Microbiology, 14, 1220286.
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4

Phylogenetic Analysis of Streptococcus pyogenes

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Sequence types (STs) were calculated according to the multilocus sequence typing (MLST) scheme of the Public databases for molecular typing and microbial genome diversity (https://pubmlst.org/organisms/streptococcus-pyogenes) using Ariba v2.6.2 (22 (link)). Core genome MLST (cgMLST), consisting of 1,168 genes for S. pyogenes provided by SeqSphere+3.5.0 (Ridom, Münster, Germany), was performed. A simple diversity index (SDI) was applied to analyze population diversity (23 (link)).
Additionally, emm1 isolates from this study were analyzed as described by Linskey et al. (7 (link)) in comparison with a collection of 377 M1global and 247 M1UK hypervirulent S. pyogenes isolates reported in the previous reference. After removing all high single nucleotide polymorphisms (SNP) density regions (24 (link)), a core genome alignment with 6,411 SNPs was used to build a maximum-likelihood tree.
Ramírez de Arellano E., Saavedra-Lozano J., Villalón P., Jové-Blanco A., Grandioso D., Sotelo J., Gamell A., González-López J.J., Cervantes E., Gónzalez M.J., Rello-Saltor V., Esteva C., Sanz-Santaeufemia F., Yagüe G., Manzanares Á., Brañas P., Ruiz de Gopegui E., Carrasco-Colom J., García F., Cercenado E., Mellado I., del Castillo E., Pérez-Vazquez M., Oteo-Iglesias J, & Calvo C. (2024). Clinical, microbiological, and molecular characterization of pediatric invasive infections by Streptococcus pyogenes in Spain in a context of global outbreak. mSphere, 9(3), e00729-23.
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