Phenotypic Methods for Determining Genomovar Status of the Burkholderia cepacia Complex

Abstract

Recent taxonomic advances have demonstrated that Burkholderia cepacia is a cluster of at least seven closely related genomic species (or genomovars) collectively referred to as the B. cepacia complex, all of which may cause infections among cystic fibrosis patients and other vulnerable individuals. Thus, it is important for clinical microbiologists to be able to differentiate genomovars. Prior to this study, 361 B. cepacia complex isolates and 51 isolates easily confused with B. cepacia complex previously had been identified using a polyphasic approach, and in this study, a comparison of phenotypic and biochemical tests was carried out. It was determined that Burkholderia multivorans and Burkholderia stabilis could reliably be separated from other members of the B. cepacia complex by phenotypic methods. A combination of phenotypic and molecular tests such as recA PCR and 16S rRNA RFLP are recommended for differentiation among the genomovars of the B. cepacia complex. A biochemical reaction scheme for the identification of B. gladioli, Pandoraea species, and Ralstonia pickettii and the differentiation of these species from the B. cepacia complex is also presented.Burkholderia cepacia has been recognized as a major pathogen in patients with cystic fibrosis (CF) since the late 1970s (15). Recent taxonomic advances have demonstrated that B. cepacia is actually a cluster of at least seven closely related genomic species (or genomovars) now called the B. cepacia complex (8, 8a, 11, 31, 33). Genomovars II, IV, and V are now formally named Burkholderia multivorans, Burkholderia stabilis, and Burkholderia vietnamiensis, respectively (with the species designation B. cepacia being reserved for genomovar I) (11,31,33); genomovar III has not been formally named, pending the availability of differential diagnostic tests. Genomovar VI has recently been described and is closely related to B. multivorans (8). Genomovar VII has also been described, and the name Burkholderia ambifaria (used herein) has been proposed (8a). These two newly proposed genomovars, Burkholderia fungorum, and Pandoraea species were initially described when B. cepacia-like organisms were submitted for whole-cell protein and amplified fragment length polymorphism (AFLP) analyses and were shown to be different from the presently recognized five genomovars (5,25). Although all the genomovars thus far described can cause infections in patients with CF (21, 31), the prevalence of each genomovar has not been fully determined and may vary among different patient populations. Examination of large collections of isolates demonstrates that genomovar III is the predominant species isolated from patients with CF and that B. multivorans may frequently be recovered (22,31). Genomovars I and VI, B. vietnamiensis, and B. ambifaria appear to be less commonly found in CF patients (8, 8a, 19, 31).Commercial bacterial identification systems are not able to differentiate among the genomovars nor accurately confirm the identification of B. cepacia complex isolates while differ...

Prevalência de bactérias potencialmente patogênicas em espécimes respiratórios de fibrocísticos do Recife

“…aeruginosa, a bactéria mais freqüentemente encontrada, foi obtida de 96 espécimes (65, (9) permitiu incluir determinada cultura no complexo "B. cepacia". Porém não foi capaz de distinguir as várias espécies dentro do complexo (resultados não-apresentados).…”

“…De interesse aos laboratórios clínicos, que não dispõem de metodologia molecular para a identificação de isolados pertencentes ao complexo "B. cepacia", foi a constatação do bom desempenho do esquema de identificação fenotípica empregado (9) . Esse esquema foi capaz de incluir no ou de excluir do complexo bactérias gram-negativas não-fermentadoras, porém sem determinar a espécie.…”

“…As que apareceram no BCSA e que, ao serem transferidas para o meio tríplice açúcar ferro, cresceram apenas na parte inclinada do meio, isto é, mostraram-se não-fermentadoras da glicose, foram identificadas usando-se uma bateria simples de testes bioquímicos tradicionais (9) . As culturas que apresentaram fenótipo compatível com "B. cepacia" foram enviadas ao Professor Anthony Hart, da Faculdade de Medicina de Liverpool, para confirmação diagnóstica através da análise molecular do gene recA, como anteriormente publicado (3) .…”

Taxon K, a complex within the Burkholderia cepacia complex, comprises at least two novel species, Burkholderia contaminans sp. nov. and Burkholderia lata sp. nov.

“…A biochemical characterization was performed as described previously (Henry et al, 1997(Henry et al, , 2001). …”

“…In general, species of the Bcc are phenotypically nearly indistinguishable, making their differentiation quite difficult, even with an extended panel of biochemical tests (Henry et al, 2001). The variable nature of biochemical results for Bcc strains may be due to their unusual genomes, which are among the largest seen in Gramnegative bacteria (6-9 Mb) and which are divided into a minimum of two chromosomal replicons (Lessie et al, 1996;Mahenthiralingam & Drevinek, 2007).…”