Development of a Species-Specific fur Gene-Based Method for Identification of the Burkholderia cepacia Complex

Abstract

Burkholderia is an important bacterial genus with a complex taxonomy that contains species of both ecological and pathogenic importance, including nine closely related species collectively termed the Burkholderia cepacia complex (BCC). Unfortunately, 16S rRNA gene analysis has proven to be not sensitive enough to discriminate between species of the BCC. Alternative species identification strategies such as recA-based PCR followed by restriction fragment length polymorphism analysis, although initially useful, have proven to be inaccurate with the increasing species diversity of the BCC. recA gene sequence analysis is more discriminatory and corroborates other biochemical and polyphasic means of taxonomic differentiation. However, it is limited by the fact that certain BCC species are subdivided into discrete recA sequence subgroups that may confuse clinical diagnoses. In this study, an effective approach is described for the rapid differentiation of BCC species from both environmental and clinical sources by means of a single-locus sequencing and PCR assay using fur as a target gene that provides sequence phylogenies that are species specific and, with few exceptions, not divided into subspecies clusters. This assay is specific and can be used to correctly determine the species status of BCC strains tested following sequencing and amplification of the fur gene by both general and speciesspecific primers. Based on our results, this typing strategy is simpler than and as sensitive as established tests currently in use clinically. This assay is useful for the rapid, definitive identification of all nine current BCC species and potentially novel species groups.

Screening and identification of a highly lipolytic bacterial strain from barbecue sites in Hainan and characterization of its lipase

“…2). Unfortunately, the resolution of the phylogeny was not good enough to distinguish B. cepacia and B. cenocepacia, and both B. cepacia and B. cenocepacia were dispersed in several clades in the phylogenetic tree, with very low bootstrap supports, similar to previous reports (LiPuma et al 1999;Mahenthiralingam et al 2000;Brown and Govan 2007;Lynch and Dennis 2008).…”

“…The fur gene sequence was reported to be more effective in rapid differentiation of BCC species (Lynch and Dennis 2008). Therefore, the fur gene fragment from C737-11 was amplified and sequenced.…”

“…To further genotype the Burkholderia strain, 5′ ATG ACC AAT CCG ACC GAT CTC AA 3′ and 5′ TCA GTG CTT GCG ITN IGG GCA GTT 3′ were used to amplify the ferric uptake regulator (fur) gene fragment in bacteria (Lynch and Dennis 2008).…”

Identification and characterization of ϕH111-1

“…21 Strains from the original and updated BCC experimental strain panels, 22,23 B. cenocepacia K56-2 LPS mutants, 11,12 and clinical isolates from the University of Alberta Hospital Cystic Fibrosis Clinic 13 were used for phage isolation, propagation, and host range testing. Strains were grown aerobically overnight at 30 °C in half-strength Luria-Bertani (½ LB) broth or solid medium (containing agar or, for DNA isolation, agarose).…”

Unusual Distribution of Burkholderia cepacia Complex Species in Danish Cystic Fibrosis Clinics May Stem from Restricted Transmission between Patients

“…One isolate per patient, cultured between 1994 and 2006, was included in the study. Allocation to species within the BCC was performed by partial atpD and recA sequencing (1); occasional isolates with no PCR product from either amplification were subjected to partial sequencing of fur (16). Two independent sequence-based identifications were thus obtained for all BCC isolates.…”