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Nature Reviews. Microbiology Oct 2013Multilocus sequence typing (MLST) was proposed in 1998 as a portable sequence-based method for identifying clonal relationships among bacteria. Today, in the... (Review)
Review
Multilocus sequence typing (MLST) was proposed in 1998 as a portable sequence-based method for identifying clonal relationships among bacteria. Today, in the whole-genome era of microbiology, the need for systematic, standardized descriptions of bacterial genotypic variation remains a priority. Here, to meet this need, we draw on the successes of MLST and 16S rRNA gene sequencing to propose a hierarchical gene-by-gene approach that reflects functional and evolutionary relationships and catalogues bacteria 'from domain to strain'. Our gene-based typing approach using online platforms such as the Bacterial Isolate Genome Sequence Database (BIGSdb) allows the scalable organization and analysis of whole-genome sequence data.
Topics: Alleles; Bacteria; Bacterial Typing Techniques; Base Sequence; Databases, Genetic; Evolution, Molecular; Genetic Variation; Genome, Bacterial; Genomics; Multilocus Sequence Typing; RNA, Bacterial; RNA, Ribosomal, 16S
PubMed: 23979428
DOI: 10.1038/nrmicro3093 -
Clinical Microbiology Reviews Sep 2020This review provides a state-of-the-art description of the performance of Sanger cycle sequencing of the 16S rRNA gene for routine identification of bacteria in the... (Review)
Review
This review provides a state-of-the-art description of the performance of Sanger cycle sequencing of the 16S rRNA gene for routine identification of bacteria in the clinical microbiology laboratory. A detailed description of the technology and current methodology is outlined with a major focus on proper data analyses and interpretation of sequences. The remainder of the article is focused on a comprehensive evaluation of the application of this method for identification of bacterial pathogens based on analyses of 16S multialignment sequences. In particular, the existing limitations of similarity within 16S for genus- and species-level differentiation of clinically relevant pathogens and the lack of sequence data currently available in public databases is highlighted. A multiyear experience is described of a large regional clinical microbiology service with direct 16S broad-range PCR followed by cycle sequencing for direct detection of pathogens in appropriate clinical samples. The ability of proteomics (matrix-assisted desorption ionization-time of flight) versus 16S sequencing for bacterial identification and genotyping is compared. Finally, the potential for whole-genome analysis by next-generation sequencing (NGS) to replace 16S sequencing for routine diagnostic use is presented for several applications, including the barriers that must be overcome to fully implement newer genomic methods in clinical microbiology. A future challenge for large clinical, reference, and research laboratories, as well as for industry, will be the translation of vast amounts of accrued NGS microbial data into convenient algorithm testing schemes for various applications (i.e., microbial identification, genotyping, and metagenomics and microbiome analyses) so that clinically relevant information can be reported to physicians in a format that is understood and actionable. These challenges will not be faced by clinical microbiologists alone but by every scientist involved in a domain where natural diversity of genes and gene sequences plays a critical role in disease, health, pathogenicity, epidemiology, and other aspects of life-forms. Overcoming these challenges will require global multidisciplinary efforts across fields that do not normally interact with the clinical arena to make vast amounts of sequencing data clinically interpretable and actionable at the bedside.
Topics: Bacteria; Bacterial Infections; Bacterial Typing Techniques; Clinical Laboratory Techniques; Humans; RNA, Ribosomal, 16S
PubMed: 32907806
DOI: 10.1128/CMR.00053-19 -
Current Opinion in Infectious Diseases Aug 2021The advancement of molecular techniques such as whole-genome sequencing (WGS) has revolutionized the field of bacterial strain typing, with important implications for... (Review)
Review
PURPOSE OF REVIEW
The advancement of molecular techniques such as whole-genome sequencing (WGS) has revolutionized the field of bacterial strain typing, with important implications for epidemiological surveillance and outbreak investigations. This review summarizes state-of-the-art techniques in strain typing and examines barriers faced by clinical and public health laboratories in implementing these new methodologies.
RECENT FINDINGS
WGS-based methodologies are on track to become the new 'gold standards' in bacterial strain typing, replacing traditional methods like pulsed-field gel electrophoresis and multilocus sequence typing. These new techniques have an improved ability to identify genetic relationships among organisms of interest. Further, advances in long-read sequencing approaches will likely provide a highly discriminatory tool to perform pangenome analyses and characterize relevant accessory genome elements, including mobile genetic elements carrying antibiotic resistance determinants in real time. Barriers to widespread integration of these approaches include a lack of standardized workflows and technical training.
SUMMARY
Genomic bacterial strain typing has facilitated a paradigm shift in clinical and molecular epidemiology. The increased resolution that these new techniques provide, along with epidemiological data, will facilitate the rapid identification of transmission routes with high confidence, leading to timely and effective deployment of infection control and public health interventions in outbreak settings.
Topics: Anti-Bacterial Agents; Bacterial Typing Techniques; Disease Outbreaks; Electrophoresis, Gel, Pulsed-Field; Genome, Bacterial; Humans; Molecular Epidemiology; Multilocus Sequence Typing
PubMed: 34039880
DOI: 10.1097/QCO.0000000000000743 -
Journal of Clinical Microbiology Aug 2005A multilocus sequence typing (MLST) scheme was developed for Klebsiella pneumoniae. Sequences of seven housekeeping genes were obtained for 67 K. pneumoniae strains,...
A multilocus sequence typing (MLST) scheme was developed for Klebsiella pneumoniae. Sequences of seven housekeeping genes were obtained for 67 K. pneumoniae strains, including 19 ceftazidime- and ciprofloxacin-resistant isolates. Forty distinct allelic profiles were identified. MLST data were validated against ribotyping and showed high (96%) discriminatory power. The MLST approach provides unambiguous data useful for the epidemiology of K. pneumoniae isolates.
Topics: Bacterial Typing Techniques; Base Sequence; Cross Infection; Drug Resistance, Bacterial; Humans; Klebsiella pneumoniae; Microbial Sensitivity Tests; Molecular Sequence Data; Ribotyping; Sequence Analysis, DNA
PubMed: 16081970
DOI: 10.1128/JCM.43.8.4178-4182.2005 -
Revista Chilena de Infectologia :... Apr 2020
Topics: Bacterial Typing Techniques; Comamonas; DNA, Bacterial
PubMed: 32730480
DOI: 10.4067/s0716-10182020000200147 -
Proteomics. Clinical Applications Apr 2016Identification and typing of bacteria occupy a large fraction of time and work in clinical microbiology laboratories. With the certification of some MS platforms in... (Review)
Review
Identification and typing of bacteria occupy a large fraction of time and work in clinical microbiology laboratories. With the certification of some MS platforms in recent years, more applications and tests of MS-based diagnosis methods for bacteria identification and typing have been created, not only on well-accepted MALDI-TOF-MS-based fingerprint matches, but also on solving the insufficiencies of MALDI-TOF-MS-based platforms and advancing the technology to areas such as targeted MS identification and typing of bacteria, bacterial toxin identification, antibiotics susceptibility/resistance tests, and MS-based diagnostic method development on unique bacteria such as Clostridium and Mycobacteria. This review summarizes the recent development in MS platforms and applications in bacteria identification and typing of common pathogenic bacteria.
Topics: Bacterial Toxins; Bacterial Typing Techniques; Campylobacter jejuni; Clostridium; Escherichia coli; Humans; Listeria monocytogenes; Microbial Sensitivity Tests; Mycobacteriaceae; Peptide Fragments; Proteolysis; Proteomics; Salmonella; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Trypsin
PubMed: 26751976
DOI: 10.1002/prca.201500086 -
Current Issues in Molecular Biology Jul 2021Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) is routinely used for bacterial identification. It would be highly beneficial... (Review)
Review
Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) is routinely used for bacterial identification. It would be highly beneficial to also be able to use the technology as a fast way to detect clinically relevant clones of bacterial species. However, studies to this aim have often had limited success. The methods used for data acquisition, processing and data interpretation are highly diverse amongst studies on MALDI-TOF MS sub-species typing. In addition to this, feasibility may depend on the bacterial species and strains investigated, making it difficult to determine what methods may or may not work. In our paper, we have reviewed recent research on MALDI-TOF MS typing of bacterial strains. Although we found a lot of variation amongst the methods used, there were approaches shared by multiple research groups. Multiple spectra of the same isolate were often combined before further analysis for strain distinction. Many groups used a protein extraction step to increase resolution in their MALDI-TOF MS results. Peaks at a high mass range were often excluded for data interpretation. Three groups have found ways to determine feasibility of MALDI-TOF MS typing for their set of strains at an early stage of their project.
Topics: Animals; Bacteria; Bacterial Proteins; Bacterial Typing Techniques; Humans; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
PubMed: 34294671
DOI: 10.3390/cimb43020054 -
Genome Research Feb 2019The routine use of genomics for disease surveillance provides the opportunity for high-resolution bacterial epidemiology. Current whole-genome clustering and multilocus...
The routine use of genomics for disease surveillance provides the opportunity for high-resolution bacterial epidemiology. Current whole-genome clustering and multilocus typing approaches do not fully exploit core and accessory genomic variation, and they cannot both automatically identify, and subsequently expand, clusters of significantly similar isolates in large data sets spanning entire species. Here, we describe PopPUNK (ulation artitioning sing ucleotide -mers), a software implementing scalable and expandable annotation- and alignment-free methods for population analysis and clustering. Variable-length -mer comparisons are used to distinguish isolates' divergence in shared sequence and gene content, which we demonstrate to be accurate over multiple orders of magnitude using data from both simulations and genomic collections representing 10 taxonomically widespread species. Connections between closely related isolates of the same strain are robustly identified, despite interspecies variation in the pairwise distance distributions that reflects species' diverse evolutionary patterns. PopPUNK can process 10-10 genomes in a single batch, with minimal memory use and runtimes up to 200-fold faster than existing model-based methods. Clusters of strains remain consistent as new batches of genomes are added, which is achieved without needing to reanalyze all genomes de novo. This facilitates real-time surveillance with consistent cluster naming between studies and allows for outbreak detection using hundreds of genomes in minutes. Interactive visualization and online publication is streamlined through the automatic output of results to multiple platforms. PopPUNK has been designed as a flexible platform that addresses important issues with currently used whole-genome clustering and typing methods, and has potential uses across bacterial genetics and public health research.
Topics: Bacteria; Bacterial Infections; Bacterial Typing Techniques; Genetic Variation; Genome, Bacterial; Genomics; Software
PubMed: 30679308
DOI: 10.1101/gr.241455.118 -
FEBS Letters Nov 2016An increasing body of evidence indicates the relevance of microbiota for pulmonary health and disease. Independent investigations recently demonstrated that the lung... (Review)
Review
An increasing body of evidence indicates the relevance of microbiota for pulmonary health and disease. Independent investigations recently demonstrated that the lung harbors a resident microbiota. Therefore, it is intriguing that a lung microbiota can shape pulmonary immunity and epithelial barrier functions. Here, we discuss the ways how the composition of the microbial community in the lung may influence pulmonary health and vice versa, factors that determine community composition. Prominent microbiota at other body sites such as the intestinal one may also contribute to pulmonary health and disease. However, it is difficult to discriminate between influences of lung vs. gut microbiota due to systemic mutuality between both communities. With focuses on asthma and respiratory infections, we discuss how microbiota of lung and gut can determine pulmonary immunity and barrier functions.
Topics: Bacterial Typing Techniques; Host-Pathogen Interactions; Humans; Intestines; Lung; Microbiota; Respiratory Tract Infections
PubMed: 27637588
DOI: 10.1002/1873-3468.12421 -
Current Issues in Molecular Biology 2017Within less than a decade matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) has become a gold standard for microbial... (Review)
Review
Within less than a decade matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) has become a gold standard for microbial identification in clinical microbiology laboratories. Besides identification of microorganisms the typing of single strains as well as the antibiotic and antimycotic resistance testing has come into focus in order to speed up the microbiological diagnostic. However, the full potential of MALDI-TOF MS has not been tapped yet and future technological advancements will certainly expedite this method towards novel applications and enhancement of current practice. So, the following chapter shall be rather a brainstorming and forecast of how MALDI-TOF MS will develop to influence clinical diagnostics and microbial research in the future. It shall open up the stage for further discussions and does not claim for overall validity.
Topics: Bacterial Typing Techniques; Clinical Laboratory Techniques; Forecasting; Humans; Microbiological Techniques; Reproducibility of Results; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
PubMed: 28504240
DOI: 10.21775/cimb.023.017