Introduction

The Haemophilus influenzae MLST scheme has been developed for the unambiguous characterisation of both typeable (encapsulated) and non-typeable (non-capsulated) isolates.   The initial database contains the allelic profiles of 131 isolates from the paper Meats et.al. (see below) and further isolates are being added at intervals.   Those carrying out MLST on this species are encouraged to submit their data to the curator so that the strain details can be added to the database. In this way the MLST database becomes an increasingly useful resource for the H. influenzae community.

The MLST scheme and initial MLST database were developed by Emma Meats (e.meats@ic.ac.uk) and Ed Feil (e.feil@bath.ac.uk now at the University of Bath; ) in the laboratory of Brian Spratt (b.spratt@ic.ac.uk) in the Department of Infectious Disease Epidemiology, Imperial College London, with help from Suzanna Stringer (stringer@molbiol.ox.ac.uk) at the PHLS Haemophilus Reference Laboratory, John Radcliffe Hospital, Oxford.  The initial strains were kindly provided by Richard Moxon, Simon Kroll, Richard Goldstein and Tanja Popovic.

REF: Emma Meats, Edward J. Feil, Susan Stringer, Alison Cody, Richard Goldstein, J. Simon Kroll, Tanja Popovic and Brian G. Spratt. 2003. Characterization of Encapsulated and Noncapsulated Haemophilus influenzae and Determination of Phylogenetic Relationships by Multilocus Sequence Typing. J Clin Microbiol 2003 April;41(4):1623-1636.

Obtaining an allelic profile and comparing your strains with those in our database

The allelic profile of an H. influenzae strain is obtained by sequencing internal fragments of seven house-keeping genes. The primers for the amplification and sequencing of these gene fragments can be obtained here The sequences must be obtained on both strands, and they must be 100% accurate, since even a single error may convert a known allele into a novel allele.

The sequences have to be trimmed so that they correspond exactly to the region that we use to define the alleles. The sequences of the seven loci from a typical H. influenzae can be obtained here and can be used to ensure that your sequences have been trimmed correctly.

You then need to access our databases, which involves a simple registration process, that allows us to inform you of new developments by e-mail.

For a query isolate, the sequences at the seven loci have to be compared with those in our database. Select the H. influenzae database, and the  locus query drop-down menu.  Select single locus and paste the correctly trimmed sequence into the box.  Select the appropriate locus in the drop-down menu, followed by submit query.

The software will check that the sequence is the correct length and that it does not contain any unrecognised characters. A check is also made to see if the submitted sequence is at least 70% similar to another allele at that locus (in case you have cut and pasted a sequence into the wrong box).   If the sequence corresponds to a known allele, the allele number will be returned.  If the sequence appears to be a new allele it should be compared with the most similar locus (or loci) to check that any sequence differences are real.  If you are convinced you have a new allele, the forward and reverse traces from the sequencer should be sent to the curator who will check their quality and if OK will assign a new allele number.

After you have obtained the allele numbers at each locus for your query strain, you select profile query and in the drop-down menu select allelic and enter the seven allele numbers in the appropriate boxes and submit.  If the allelic profile is in the database, the sequence type assigned to this allelic profile will be returned.  Otherwise the most similar allelic profile will be returned. You can then search for isolates that have allelic profiles that are similar to yours. For example, isolates that have at least 4/7, 5/7 or 6/7 matches to the submitted allelic profile and can show the relationships between your query strain and these strains by using the Tree button.

Further details about strains that are identical, or similar, to the submitted strain can be obtained by clicking on the strain names.

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