Internet-Accessible DNA Sequence Database for Identifying Fusaria from Human and Animal Infections

Abstract

Because less than one-third of clinically relevant fusaria can be accurately identified to species level using phenotypic data (i.e., morphological species recognition), we constructed a three-locus DNA sequence database to facilitate molecular identification of the 69 Fusarium species associated with human or animal mycoses encountered in clinical microbiology laboratories. The database comprises partial sequences from three nuclear genes: translation elongation factor 1␣ (EF-1␣), the largest subunit of RNA polymerase (RPB1), and the second largest subunit of RNA polymerase (RPB2). These three gene fragments can be amplified by PCR and sequenced using primers that are conserved across the phylogenetic breadth of Fusarium. Phylogenetic analyses of the combined data set reveal that, with the exception of two monotypic lineages, all clinically relevant fusaria are nested in one of eight variously sized and strongly supported species complexes. The monophyletic lineages have been named informally to facilitate communication of an isolate's clade membership and genetic diversity. To identify isolates to the species included within the database, partial DNA sequence data from one or more of the three genes can be used as a BLAST query against the database which is Web accessible at FUSARIUM-ID (http://isolate.fusariumdb.org) and the Centraalbureau voor Schimmelcultures (CBS-KNAW) Fungal Biodiversity Center (http://www.cbs.knaw.nl/fusarium). Alternatively, isolates can be identified via phylogenetic analysis by adding sequences of unknowns to the DNA sequence alignment, which can be downloaded from the two aforementioned websites. The utility of this database should increase significantly as members of the clinical microbiology community deposit in internationally accessible culture collections (e.g., CBS-KNAW or the Fusarium Research Center) cultures of novel mycosis-associated fusaria, along with associated, corrected sequence chromatograms and data, so that the sequence results can be verified and isolates are made available for future study.

Emergence of fusarioses in a university hospital in Turkey during a 20-year period

“…Tentative identification of species (complexes) was by nuclear ribosomal internal transcribed spacer (ITS) sequencing with primers ITS1 and ITS4 [21] and a part of the translation elongation factor 1-alpha (tef-1α) gene with primers EF1 and EF2 [22]. To determine the exact haplotypes within species, partial sequences of the rpb2 gene [5], the intergenic spacer region IGS [3], the large ribosomal subunit LSU, and the beta-tubulin gene [23] were obtained as needed. For sequence typing of members of the FSSC, LSU and rpb2 [2] were used in addition to ITS and tef-1α, for members of the FDSC and the FFSC, rpb2 and β-tubulin [23], and for the FOSC, IGS and rpb2 [3].…”

“…Strains were identified by BLAST identification of the obtained sequences in GenBank, the CBS database, the Fusarium ID [22] and Fusarium MLST [5] databases. Besides phylogenetic placements with species/haplotypes within species, complexes were checked with the above mentioned available databases specific for the found species complexes.…”

“…Most of the former Fusarium species are now known to be species complexes composed of clusters of cryptic species with little to no morphological differences. Multilocus sequence typing (MLST) is necessary to distinguish between such species [2][3][4][5]. Now that the Bone fungus = one name^rule has been adopted, a proposal was made to conserve the name Fusarium above linked sexual state names like Haemonectria and Gibberella [6], and a plea to keep the taxonomy of clinically important species as stable as possible [7].…”

“…Most of the opportunistic Fusarium pathogens identified in the literature are members of the F. solani species complex (FSSC), the F. oxysporum species complex (FOSC), and the F. fujikuroi species complex (FFSC), and less often of the F. incarnatum-equiseti species complex (FIESC), the F. d i m e r u m s p e c i e s c o m p l e x ( F D S C ) , a n d t h e F. chlamydosporum species complex (FCSC) [5,8,9]. Some species and haplotypes causing infections have a worldwide distribution, while local variation also exists [10].…”

Fusarium fujikuroi causing fusariosis of pineapple in peninsular Malaysia

“…Fusarium fujikuroi is a well-known pathogen of bakanae disease of rice. Besides rice, F. fujikuroi has been reported as saprophytic or endophytic colonisers of vanilla (Pinaria et al 2010) and was isolated from human skin (O'Donnell et al 2010). According to Slippers et al (2005), fungi are more likely to undergo plant host jumps as a result of human movement and weather changes.…”