title: Multilocus sequence typing of Clostridium difficile.
abstract: A robust high-throughput multilocus sequence typing (MLST) scheme for Clostridium difficile was developed and validated using a diverse collection of 50 reference isolates representing 45 different PCR ribotypes and 102 isolates from recent clinical samples. A total of 49 PCR ribotypes were represented overall. All isolates were typed by MLST and yielded 40 sequence types (STs). A web-accessible database was set up (http://pubmlst.org/cdifficile/) to facilitate the dissemination and comparison of C. difficile MLST genotyping data among laboratories. MLST and PCR ribotyping were similar in discriminatory abilities, having indices of discrimination of 0.90 and 0.92, respectively. Some STs corresponded to a single PCR ribotype (32/40), other STs corresponded to multiple PCR ribotypes (8/40), and, conversely, the PCR ribotype was not always predictive of the ST. The total number of variable nucleotide sites in the concatenated MLST sequences was 103/3,501 (2.9%). Concatenated MLST sequences were used to construct a neighbor-joining tree which identified four phylogenetic groups of STs and one outlier (ST-11; PCR ribotype 078). These groups apparently correlate with clades identified previously by comparative genomics. The MLST scheme was sufficiently robust to allow direct genotyping of C. difficile in total stool DNA extracts without isolate culture. The direct (nonculture) MLST approach may prove useful as a rapid genotyping method, potentially benefiting individual patients and informing hospital infection control.
authors.affiliation: Nuffield Department of Clinical Medicine, Oxford University, John Radcliffe Hospital, Oxford OX3 9DU, United Kingdom.
authors.forename: David
authors.initials: D
authors.lastname: Griffiths
authors.forename: Warren
authors.initials: W
authors.lastname: Fawley
authors.forename: Melina
authors.initials: M
authors.lastname: Kachrimanidou
authors.forename: Rory
authors.initials: R
authors.lastname: Bowden
authors.forename: Derrick W
authors.initials: DW
authors.lastname: Crook
authors.forename: Rowena
authors.initials: R
authors.lastname: Fung
authors.forename: Tanya
authors.initials: T
authors.lastname: Golubchik
authors.forename: Rosalind M
authors.initials: RM
authors.lastname: Harding
authors.forename: Katie J M
authors.initials: KJ
authors.lastname: Jeffery
authors.forename: Keith A
authors.initials: KA
authors.lastname: Jolley
authors.forename: Richard
authors.initials: R
authors.lastname: Kirton
authors.forename: Tim E
authors.initials: TE
authors.lastname: Peto
authors.forename: Gareth
authors.initials: G
authors.lastname: Rees
authors.forename: Nicole
authors.initials: N
authors.lastname: Stoesser
authors.forename: Alison
authors.initials: A
authors.lastname: Vaughan
authors.forename: A Sarah
authors.initials: AS
authors.lastname: Walker
authors.forename: Bernadette C
authors.initials: BC
authors.lastname: Young
authors.forename: Mark
authors.initials: M
authors.lastname: Wilcox
authors.forename: Kate E
authors.initials: KE
authors.lastname: Dingle
mesh_terms: D015373:Bacterial Typing Techniques / Q000379:methods*; D016360:Clostridioides difficile / Q000145:classification* / Q000235:genetics* / Q000302:isolation & purification; D003015:Clostridium Infections / Q000382:microbiology; D016172:DNA Fingerprinting / Q000379:methods*; D005838:Genotype; D006801:Humans; D007223:Infant; D011110:Polymorphism, Genetic; D021521:Ribotyping; D012680:Sensitivity and Specificity; D017422:Sequence Analysis, DNA / Q000379:methods*