Multilocus sequence typing: A portable approach to the identification of clones within populations of pathogenic microorganisms

Abstract

Traditional and molecular typing schemes for the characterization of pathogenic microorganisms are poorly portable because they index variation that is difficult to compare among laboratories. To overcome these problems, we propose multilocus sequence typing (MLST), which exploits the unambiguous nature and electronic portability of nucleotide sequence data for the characterization of microorganisms. To evaluate MLST, we determined the sequences of Ϸ470-bp fragments from 11 housekeeping genes in a reference set of 107 isolates of Neisseria meningitidis from invasive disease and healthy carriers. For each locus, alleles were assigned arbitrary numbers and dendrograms were constructed from the pairwise differences in multilocus allelic profiles by cluster analysis. The strain associations obtained were consistent with clonal groupings previously determined by multilocus enzyme electrophoresis. A subset of six gene fragments was chosen that retained the resolution and congruence achieved by using all 11 loci. Most isolates from hyper-virulent lineages of serogroups A, B, and C meningococci were identical for all loci or differed from the majority type at only a single locus. MLST using six loci therefore reliably identified the major meningococcal lineages associated with invasive disease. MLST can be applied to almost all bacterial species and other haploid organisms, including those that are difficult to cultivate. The overwhelming advantage of MLST over other molecular typing methods is that sequence data are truly portable between laboratories, permitting one expanding global database per species to be placed on a World-Wide Web site, thus enabling exchange of molecular typing data for global epidemiology via the Internet.

Extra-intestinal pathogenic Escherichia coli (ExPEC): Disease, carriage and clones

“…Multi-locus Sequence Typing (MLST) involves sequencing of selected (often seven) bacterial housekeeping genes and, due to its standardised approach and greater resolution as compared with 'phylogrouping', has allowed more detailed analysis of ExPEC lineages (22,23). It separates the isolates into distinct sequence types (STs), which are defined as isolates with M A N U S C R I P T…”

Molecular tools for the characterisation of antibiotic-resistant bacteria

“…A MultiLocus Sequence Typing (MLST) approach has been proposed that is based upon the sequences of short internal fragments of seven housekeeping genes in order to define sequence types corresponding to alleles at each of the seven loci [37]. MLST has been used with several pathogenic species, including S. pneumoniae [15], Neisseria meningitidis [75], Campylobacter jejuni [13], as well as to type uncultured bacteria directly from cerebrospinal fluid [17], or for the characterisation of methicillin-resistant and methicillin-susceptible clones of Staphylococcus aureus [16].…”

“…However, like the latter method, MLST targets coding regions and uses variation that accumulates very slowly in the population contrary to pulsefield-electrophoresis (PFGE) or arbitrarily primed PCR described above. As a result, MLST is used for long-term epidemiology and for the identification of lineages that have an increased propensity to cause disease [37]. Investigators can determine the allelic profile of their isolates by comparing the obtained sequence with the sequence available in the databases (http://mlst.…”

Clonal distribution and associated characteristics of Escherichia coli clinical and surveillance isolates from a military medical center

“…Minimum inhibitory concentration (MIC) results were interpreted according to the Clinical Laboratory and Standards Institute criteria (Clinical and Laboratory Standards Institute, 2013). In addition, isolates were assessed molecularly for bla CTX-M-15 , major E. coli phylogenetic group (phylogroup) (Clermont et al, 2013), ST or ST complex (STc; as determined by fumC-fimH typing (Weissman et al, 2012), full or partial MLST (Maiden et al, 1998), or STc-specific PCR assays Johnson et al, 2009;Matsumura et al, 2012)), membership in the ST131-H30 clonal subset or its sublineage ST131-H30Rx (Banerjee et al, 2013), O type (O16 and O25b only) (Johnson et al, 2014), and extended virulence genotype (for 50 markers) (Johnson et al, 2015). Resistance scores were defined as the number of antibiotic classes to which an isolate exhibited resistance.…”