Melioidosis:
Problems with Current Identification
Methods (API, Vitek and other methods)
Rob Baird
Royal Darwin Hospital
Christoph Weissert, et al Emerging Infec Dis
Volume 15, Number 11—November 2009
or
J Med Microbiol. 2012 Oct;61(Pt 10):1483-4.Epub 2012 Jul 19.
Common phenotypic mis-identifications of B. pseudomallei are primarily due to:
Regional biochemical differences in B. pseudomallei strains
Lack of the regional biochemical profiles in the phenotypic databases
A proliferation of taxonomic change in the genus, occurring well ahead of phenotypic
database updates
and
Ashdown’s media: growth not limited to B. pseudomallei. Coliforms, yeast, S.
maltophilia, Pseud sp. and B. cepacia can all grow.
Can survive the decontamination process of TB and grows quite well on the media used
making isolation of TB difficult.
Mass spectrometry, while an excellent aid in diagnosis, is dependent on the quality of
the protein spectra database of the instrument. Misidentification has been reported in the
literature.
Currently 85 Burkholderia are taxonomically recognised
Burkholderia acidipaludis
Burkholderia ambifaria
Burkholderia andropogonis
Burkholderia anthina
Burkholderia hospita
Burkholderia kururiensis
Burkholderia lata
Burkholderia latens
Burkholderia soli
Burkholderia sordidicola
Burkholderia stabilis
Burkholderia symbiotica
Burkholderia arboris
Burkholderia bannensis
Burkholderia mallei
Burkholderia terrae
Burkholderia terricola
Burkholderia brasilensis
Burkholderia bryophila
Burkholderia caledonica
Burkholderia caribensis
Burkholderia caryophylli
Burkholderia cenocepacia
Burkholderia metallica
Burkholderia mimosarum
Burkholderia multivorans
Burkholderia nodosa
Burkholderia norimbergensis
Burkholderia oklahomensis
Burkholderia thailandensis
Burkholderia cepacia
Burkholderia cepacia complex
Burkholderia oxyphila
Burkholderia vietnamiensis
Burkholderia phenazinium
Burkholderia phenoliruptrix
Burkholderia phymatum
Burkholderia phytofirmans
Burkholderia pickettii
Burkholderia plantarii
Burkholderia xenovorans
Burkholderia zhejiangensis
"Candidatus Burkholderia calva"
"Candidatus Burkholderia crenata"
"Candidatus Burkholderia hispidae"
"Candidatus Burkholderia kirkii"
Burkholderia pseudomallei
"Candidatus Burkholderia mamillata
"Candidatus Burkholderia nigropunctata"
"Candidatus Burkholderia rigidae"
"Candidatus Burkholderia schumannianae"
"Candidatus Burkholderia verschuerenii"
"Candidatus Burkholderia virens"
Burkholderia cocovenenans
Burkholderia contaminans
Burkholderia denitrificans
Burkholderia diazotrophica
Burkholderia diffusa
Burkholderia dolosa
Burkholderia endofungorum
Burkholderia ferrariae
Burkholderia fungorum
Burkholderia ginsengisoli
Burkholderia gladioli
Burkholderia glathei
Burkholderia glumae
Burkholderia graminis
Burkholderia grimmiae
Burkholderia heleia
Burkholderia megapolitana
Burkholderia pyrrocinia
Burkholderia rhizoxinica
Burkholderia sabiae
Burkholderia sacchari
Burkholderia sartisoli
Burkholderia sediminicola
Burkholderia seminalis
Burkholderia silvatlantica
Burkholderia singaporensis
Burkholderia solanacearum
Burkholderia tropica
Burkholderia tuberum
Burkholderia ubonensis
Burkholderia unamae
Burkholderia vandii
Regional differences
Comparison of the VITEK 2 system biochemical profiles of 68
confirmed B. pseudomallei clinical strains from hospitals in Sabah
and Sarawak, Malaysian Borneo with 149 B. pseudomallei and 18
B. cepacia isolates from the Royal Darwin Hospital (RDH) in the
Northern Territory, Australia. One isolate per patient was
analyzed. All isolates were collected between September 2010
and June 2012 except for 17 isolates collected in 1994 from Sabah.
Regional differences
Typical/usual B. pseudomallei profile:
88% of isolates with BNAG identified correctly, only 13% without BNAG identified correctly
B
A
C
D
Figure 1. Nonmetric multidimensional scaling (nMDS) ordination on the Euclidean distance resemblance matrix of the Vitek-2
biochemical profile of 235 B. pseudomallei and B. cepacia isolates from Australia and Malaysian Borneo. (A) Samples were identified as
either B. pseudomallei, B. cepacia, B. pseudomallei misidentified as B. cepacia or isolates with low discrimination. (B) The bubble size
reflects the presence (large) or absence (small) of BNAG substrate in an isolate. (C) Analysis based on isolates from both countries,
Australia and Malaysia. (D) The bubble size reflects the presence (large) or absence (small) of NAGA substrate in an isolate.
Abbreviations: Bps, B. pseudomallei; Bcep, B. cepacia; Bcep misID, B. pseudomallei misidentified as B. cepacia; BNAG, β-N-acetyl-glucosaminidase; NAGA, Nacetylgalactosamine.
Unfamiliarity with B. pseudomallei and problems with inaccurate speciation using some
automated commercial biochemical identification systems have resulted in laboratories
misidentifying the bacterium as a Pseudomonas or other Burkholderia species.
Confirmation of B. pseudomallei identity by real-time
polymerase chain reaction (PCR) of DNA extracted from cultured
bacterial colonies is increasingly the standard for many
laboratories. Various genetic targets have been published for
PCR identification of B. pseudomallei from bacterial cultures and
also for direct detection from clinical samples, with a recent
review showing the TTS1-orf2 assay to be superior in detecting
B. pseudomallei directly from clinical specimens.
Apart from molecular methods, B. pseudomallei from cultures can also be confirmed by
antigen detection assays such as latex agglutination. More recently matrix-assisted
laser desorption ionization-time of flight Mass Spectrometry (Maldi-TOF MS) has been
adapted to identify cultured bacteria based on protein fingerprint profiles.
Common phenotypic mis-identifications of B. pseudomallei are primarily due to:
Regional biochemical differences in B. pseudomallei strains
Lack of the regional biochemical profiles in the phenotypic databases
A proliferation of taxonomic change in the genus, occurring well ahead of phenotypic database
updates
The move towards genotypic and molecular identification, as the gold standard, will
reduce this problem for reference labs, but
local labs still require a high index of clinical and microbiological suspicion, when
identifying Burkholderia isolates from patients with compatible clinical illnesses
and
Ashdown’s media: growth not limited to B. pseudomallei. Coliforms, yeast, S. maltophilia, Pseud sp.
and B. cepacia can all grow.
Can survive the decontamination process of TB and grows quite well on the media used making
isolation of TB difficult.
Mass spectrometry, while an excellent aid in diagnosis, is dependent on the quality of the protein
spectra database of the instrument. Misidentification has been reported in the literature.