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      • Record: found
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      Is Open Access

      Cultivation and sequencing-free protocol for Serratia marcescens detection and typing

      research-article

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          Summary

          Serratia marcescens is an opportunistic pathogen that survives in inhospitable environments causing large outbreaks, particularly in neonatal intensive care units (NICUs). Genomic studies revealed that most S. marcescens nosocomial infections are caused by a specific clone (here “Infectious clone”). Whole genome sequencing (WGS) is the only portable method able to identify this clone, but it requires days to obtain results. We present a cultivation-free hypervariable-locus melting typing (HLMT) protocol for the fast detection and typing of S. marcescens, with 100% detection capability on mixed samples and a limit of detection that can reach the 10 genome copies. The protocol was able to identify the S. marcescens infectious clone with 97% specificity and 96% sensitivity when compared to WGS, yielding typing results portable among laboratories. The protocol is a cost and time saving method for S. marcescens detection and typing for large environmental/clinical surveillance screenings, also in low-middle income countries.

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          Highlights

          • •

            Most Serratia marcescens clinical infections are due to a specific clone

          • •

            We developed a PCR/HRM protocol for fast Serratia marcescens detection and typing

          • •

            The protocol has high sensitivity and sensibility in identify the infectious clone

          • •

            The protocol is highly reproducible and gives portable results

          Abstract

          Biological sciences; Microbiology; Environmental Biotechnology; Biological sciences research methodologies

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          Most cited references45

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          RAxML version 8: a tool for phylogenetic analysis and post-analysis of large phylogenies

          Alexandros Stamatakis (2014)
          Motivation: Phylogenies are increasingly used in all fields of medical and biological research. Moreover, because of the next-generation sequencing revolution, datasets used for conducting phylogenetic analyses grow at an unprecedented pace. RAxML (Randomized Axelerated Maximum Likelihood) is a popular program for phylogenetic analyses of large datasets under maximum likelihood. Since the last RAxML paper in 2006, it has been continuously maintained and extended to accommodate the increasingly growing input datasets and to serve the needs of the user community. Results: I present some of the most notable new features and extensions of RAxML, such as a substantial extension of substitution models and supported data types, the introduction of SSE3, AVX and AVX2 vector intrinsics, techniques for reducing the memory requirements of the code and a plethora of operations for conducting post-analyses on sets of trees. In addition, an up-to-date 50-page user manual covering all new RAxML options is available. Availability and implementation: The code is available under GNU GPL at https://github.com/stamatak/standard-RAxML. Contact: alexandros.stamatakis@h-its.org Supplementary information: Supplementary data are available at Bioinformatics online.
          Article 0 comments Cited 7703 times – based on 0 reviews     Review now
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            SPAdes: a new genome assembly algorithm and its applications to single-cell sequencing.

            Anton Bankevich, Sergey Nurk, Dmitry Antipov … (2012)
            The lion's share of bacteria in various environments cannot be cloned in the laboratory and thus cannot be sequenced using existing technologies. A major goal of single-cell genomics is to complement gene-centric metagenomic data with whole-genome assemblies of uncultivated organisms. Assembly of single-cell data is challenging because of highly non-uniform read coverage as well as elevated levels of sequencing errors and chimeric reads. We describe SPAdes, a new assembler for both single-cell and standard (multicell) assembly, and demonstrate that it improves on the recently released E+V-SC assembler (specialized for single-cell data) and on popular assemblers Velvet and SoapDeNovo (for multicell data). SPAdes generates single-cell assemblies, providing information about genomes of uncultivatable bacteria that vastly exceeds what may be obtained via traditional metagenomics studies. SPAdes is available online ( http://bioinf.spbau.ru/spades ). It is distributed as open source software.
            Article 0 comments Cited 6657 times – based on 0 reviews     Review now
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              MUSCLE: multiple sequence alignment with high accuracy and high throughput.

              R. C. Edgar (2004)
              We describe MUSCLE, a new computer program for creating multiple alignments of protein sequences. Elements of the algorithm include fast distance estimation using kmer counting, progressive alignment using a new profile function we call the log-expectation score, and refinement using tree-dependent restricted partitioning. The speed and accuracy of MUSCLE are compared with T-Coffee, MAFFT and CLUSTALW on four test sets of reference alignments: BAliBASE, SABmark, SMART and a new benchmark, PREFAB. MUSCLE achieves the highest, or joint highest, rank in accuracy on each of these sets. Without refinement, MUSCLE achieves average accuracy statistically indistinguishable from T-Coffee and MAFFT, and is the fastest of the tested methods for large numbers of sequences, aligning 5000 sequences of average length 350 in 7 min on a current desktop computer. The MUSCLE program, source code and PREFAB test data are freely available at http://www.drive5. com/muscle.
              Article 0 comments Cited 6454 times – based on 0 reviews     Review now
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                Author and article information

                Contributors
                Francesco Comandatore
                Journal
                Journal ID (nlm-ta): iScience
                Journal ID (iso-abbrev): iScience
                Title: iScience
                Publisher: Elsevier
                ISSN (Electronic): 2589-0042
                Publication date PMC-release: 01 March 2024
                Publication date Collection: 19 April 2024
                Publication date (Electronic): 01 March 2024
                Volume: 27
                Issue: 4
                Electronic Location Identifier: 109402
                Affiliations
                [1 ]Department of Biomedical and Clinical Sciences, Pediatric Clinical Research Center “Romeo and Enrica Invernizzi”, University of Milan, 20157 Milan, Italy
                [2 ]Department of Biosciences and Pediatric Clinical Research Center "Romeo Ed Enrica Invernizzi", University of Milan, 20133 Milan, Italy
                [3 ]Unit of Microbiology and Clinical Microbiology, Department of Clinical-Surgical, Diagnostic and Pediatric Sciences, University of Pavia, Pavia 27100, Italy
                [4 ]Department of Environmental Health Sciences, Mailman School of Public Health, Columbia University, New York, NY 10032, USA
                [5 ]Biochemistry, Microbiology and Immunology Department, Faculty of Medicine, University of Ottawa, Ottawa, ON K1H 8M5, Canada
                [6 ]Centre for Innovation, Canadian Blood Services, Ottawa, ON K1G 4J5, Canada
                [7 ]Department of Biology and Biotechnology, University of Pavia, 27100 Pavia, Italy
                [8 ]Laboratorio di Microbiologia Clinica, Virologia e Diagnostica delle Bioemergenze, ASST Fatebenefratelli Sacco, 20157 Milan, Italy
                [9 ]Department of Microbiology & Virology, Fondazione IRCCS Policlinico San Matteo, Viale Camillo Golgi 19, 27100 Pavia, Italy
                [10 ]Microbiology Unit, Fondazione IRCCS San Gerardo, 20900 Monza, Italy
                [11 ]Istituto Zooprofilattico Sperimentale della Lombardia e dell'Emilia Romagna (IZSLER), 27100 Pavia, Italy
                [12 ]Laboratory of Microbiology and Virology, Azienda Socio-Sanitaria Territoriale (ASST) Papa Giovanni XXIII, 24127 Bergamo, Italy
                [13 ]Emerging Bacterial Pathogens Unit, Division of Immunology, Transplantation and Infectious Diseases, IRCCS San Raffaele Scientific Institute, 20132 Milan, Italy
                [14 ]Department of Paediatrics, Children’s Hospital "V. Buzzi", 20154 Milano, Italy
                Author notes
                [∗ ]Corresponding author francesco.comandatore@ 123456unimi.it
                [15]

                Lead contact

                Article
                Publisher Item ID: S2589-0042(24)00623-0 Publisher ID: 109402
                DOI: 10.1016/j.isci.2024.109402
                PMC ID: 10952028
                PubMed ID: 38510115
                SO-VID: 64fc57b3-254f-45f4-8061-7cef9a402009
                Copyright © © 2024 The Authors
                License:

                This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).

                History
                Date received : 25 September 2023
                Date revision received : 8 January 2024
                Date accepted : 28 February 2024
                Categories
                Subject: Article

                Keywords: biological sciences,microbiology,environmental biotechnology,biological sciences research methodologies
                Data availability:
                Keywords: biological sciences, microbiology, environmental biotechnology, biological sciences research methodologies

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