JCM Accepted Manuscript Posted Online 15 April 2015 J. Clin. Microbiol. doi:10.1128/JCM.00299-15 Copyright © 2015, American Society for Microbiology. All Rights Reserved.
1
Discrimination of Candida albicans from Candida dubliniensis using the Biofire FilmArray®
2
Blood Culture Identification Panel
3 4
Timothy R. Southerna,b, Aqilah Alelewc, and Peter C. Iwena,b,#
5 6
Department of Pathology and Microbiology, University of Nebraska Medical Center, Omaha,
7
Nebraska, USAa
8
Nebraska Public Health Laboratory, Omaha, Nebraska, USAb
9
King Fahad Specialist Hospital, Dammam, Saudi Arabiac
10 11
#Address correspondence to Peter Iwen, [email protected]
12 13
Short title: Discrimination of Candida albicans and C. dubliniensis
14 15
Keywords: Candidemia, Candida albicans, C. dubliniensis, FilmArray® Blood Culture
16
Identification Panel
17
Candida is the predominant cause of fungemia worldwide (1) including critical care
18
settings (2). Approximately 60% of all fungemia is caused by Candida albicans followed by C.
19
glabrata, C. parapsilosis, C. tropicalis, C. krusei and the emerging pathogen C. dubliniensis (3).
20
C. dubliniensis was originally recovered from the oral cavity of an AIDS patient in Dublin,
21
Ireland in 1995 (4) and is reported to cause candidemia with a frequency estimated to be 0.2-3%
22
(3). Although recent improvements in laboratory methods for Candida identification have been
23
noted (5), the prevalence of C. dubliniensis may still be underestimated (6) due to the phenotypic
24
and genotypic similarity to C. albicans and the difficulty differentiating these two Candida
25
species. This study evaluated the ability of the FilmArray® Blood Culture Identification (BCID)
26
panel (BioFire Diagnostics, Salt Lake City, UT, USA) to discriminate C. albicans and C.
27
dubliniensis from positive blood culture. The FilmArray® BCID panel is a nested multiplexed
28
polymerase chain reaction (nmPCR) test that identifies pathogens directly from positive blood
29
cultures including C. albicans, C. glabrata, C. krusei, C. parapsilosis and C. tropicalis. C.
30
dubliniensis is not included on the panel.
31
Twenty-seven contrived blood cultures were evaluated using the BCID panel to examine
32
cross-reactivity of the panel for C. dubliniensis. Reference isolates used in this study included
33
20 C. dubliniensis and 7 C. albicans (Table 1) whose identities were previously confirmed to
34
species-level using both phenotypic (ie. germ tube test; chlamydoconidia formation and
35
appearance) and genotypic analysis (ie. sequence comparison of the complete internal
36
transcribed spacer 1 and 2 regions of the ribosomal DNA complex) (4, 7, 8, 9). BD BACTEC
37
Plus Aerobic/F culture bottles (BD Diagnostic Systems, Malvern, PA, USA) containing whole
38
blood were each inoculated with a reference isolate. Contrived cultures were incubated in a
39
BACTEC 9240 cabinet until positive followed by BCID panel testing according to the
40
manufacturer’s instructions (10).
41
The BCID panel correctly detected C. albicans (7/7) but did not detect C. dubliniensis
42
(0/20) from contrived blood cultures indicating that the BCID panel can discriminate these two
43
genetically and phenotypically similar species. These results indicated that blood cultures
44
positive for C. albicans by the BCID panel could be reported with confidence as C. albicans.
45
Furthermore, based on BCID analyte specificity for C. albicans, germ-tube-positive and BCID
46
panel-negative yeast can be presumptively identified as “Candida dubliniensis” with
47
confirmatory identification to follow.
48
Direct testing of positive blood cultures using nucleic acid amplification tests (NAATs)
49
provides rapid identification of pathogens and facilitates appropriate antimicrobial therapy.
50
Although NAATs can lack discriminatory power for highly related organisms, this study showed
51
that the FilmArray® BCID panel was capable of distinguishing C. albicans from the genetically
52
related and phenotypically similar C. dubliniensis from positive blood cultures. Future studies
53
will likely support C. dubliniensis as an important cause of human disease and that the addition
54
of this analyte to the BCID panel could help optimize clinical management of patients with
55
candidemia and improve epidemiology of candidemia as it relates to C. albicans and C.
56
dubliniensis.
57
REFERENCES
58 59
1. Delaloye J, Calandra T. 2014. Invasive candidiasis as a cause of sepsis in the critically ill
60
patient. Virulence. 5: 161-169.
61
2. Kollef M, Micek S, Hampton N, Doherty J, Kumar A. 2012. Septic shock attributed to
62
Candida infection: importance of empiric therapy and source control. Clin. Infect. Dis. 54:
63
1739-1746.
64
3. Falagas M, Roussos N, Vardakas K. 2010. Relative frequency of C. albicans and the
65
various non-albicans Candida spp among candidemia isolates from inpatients in various parts of
66
the world: a systematic review. Inter. J. Infect. Dis. 14: e954-e966.
67
4. Sullivan D, Westerneng T, Haynes K, Bennett D, Coleman D. 1995. Candida
68
dubliniensis sp. nov.: phenotypic and molecular characterization of a novel species associated
69
with oral candidosis in HIV-infected individuals. Microbiology. 141: 1507-1521.
70
5. Pfaller M, Woosley L, Messer S, Jones R, Castanheira M. 2012. Significance of
71
molecular identification and antifungal susceptibility of clinically significant yeasts and moulds
72
in a global antifungal surveillance programme. Mycopathologia. 174: 259-271.
73
6. Khan Z, Ahmad S, Joseph L, Chandy R. 2012. Candida dubliniensis: An appraisal of its
74
clinical significance as a bloodstream pathogen. PLoS One. 7: e32952.
75
7. Ells R, Kock L, Pohl C. 2009. Candida albicans or Candida dubliniensis. Mycoses. 54:
76
1-16.
77
8. Loreto E, Scheid L, Nogueira C, Zeni G, Santurio J, Alves S. 2010. Candida
78
dubliniensis: epidemiology and phenotypic methods for identification. Mycopathologia. 169:
79
431-443.
80
9. Iwen PC, Freifeld AG, Bruening TA, Hinrichs SH. 2004. Use of a panfungal PCR assay
81
for detection of fungal pathogens in a commercial blood culture system. J. Clin. Microbiol. 42:
82
2292-2293.
83
10. FilmArray® Blood Culture Identification Panel Instructions for Use, 2014
Table 1. Analysis of contrived blood cultures containing Candida albicans or C. dubliniensis using the FilmArray® Blood Culture Identification (BCID) panel. Organism
Strain
BCID Panel Result
Candida albicansa
B7560-08 B7706-08 B8627-09 B9005-09 B9010-09 B9028-09 ATCC 10231
Candida albicans Candida albicans Candida albicans Candida albicans Candida albicans Candida albicans Candida albicans
C. dubliniensisa
CU001 CU002 CU003 CU004 CU005 CU006 CU007 CU008 CU009 CU010 CU011 20-070-29 20-070-30 20-070-31 20-070-32 NE TNMC,120905 NE TNMC,121007 NE TNMC,072407 NE TNMC,012608 NE TNMC, 041108
NDa ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND
Abbreviation: ND, not detected a Identification of species included both phenotypic and genetic analyses
1
Discrimination of Candida albicans from Candida dubliniensis using the Biofire FilmArray®
2
Blood Culture Identification Panel
3 4
Timothy R. Southerna,b, Aqilah Alelewc, and Peter C. Iwena,b,#
5 6
Department of Pathology and Microbiology, University of Nebraska Medical Center, Omaha,
7
Nebraska, USAa
8
Nebraska Public Health Laboratory, Omaha, Nebraska, USAb
9
King Fahad Specialist Hospital, Dammam, Saudi Arabiac
10 11
#Address correspondence to Peter Iwen, [email protected]
12 13
Short title: Discrimination of Candida albicans and C. dubliniensis
14 15
Keywords: Candidemia, Candida albicans, C. dubliniensis, FilmArray® Blood Culture
16
Identification Panel
17
Candida is the predominant cause of fungemia worldwide (1) including critical care
18
settings (2). Approximately 60% of all fungemia is caused by Candida albicans followed by C.
19
glabrata, C. parapsilosis, C. tropicalis, C. krusei and the emerging pathogen C. dubliniensis (3).
20
C. dubliniensis was originally recovered from the oral cavity of an AIDS patient in Dublin,
21
Ireland in 1995 (4) and is reported to cause candidemia with a frequency estimated to be 0.2-3%
22
(3). Although recent improvements in laboratory methods for Candida identification have been
23
noted (5), the prevalence of C. dubliniensis may still be underestimated (6) due to the phenotypic
24
and genotypic similarity to C. albicans and the difficulty differentiating these two Candida
25
species. This study evaluated the ability of the FilmArray® Blood Culture Identification (BCID)
26
panel (BioFire Diagnostics, Salt Lake City, UT, USA) to discriminate C. albicans and C.
27
dubliniensis from positive blood culture. The FilmArray® BCID panel is a nested multiplexed
28
polymerase chain reaction (nmPCR) test that identifies pathogens directly from positive blood
29
cultures including C. albicans, C. glabrata, C. krusei, C. parapsilosis and C. tropicalis. C.
30
dubliniensis is not included on the panel.
31
Twenty-seven contrived blood cultures were evaluated using the BCID panel to examine
32
cross-reactivity of the panel for C. dubliniensis. Reference isolates used in this study included
33
20 C. dubliniensis and 7 C. albicans (Table 1) whose identities were previously confirmed to
34
species-level using both phenotypic (ie. germ tube test; chlamydoconidia formation and
35
appearance) and genotypic analysis (ie. sequence comparison of the complete internal
36
transcribed spacer 1 and 2 regions of the ribosomal DNA complex) (4, 7, 8, 9). BD BACTEC
37
Plus Aerobic/F culture bottles (BD Diagnostic Systems, Malvern, PA, USA) containing whole
38
blood were each inoculated with a reference isolate. Contrived cultures were incubated in a
39
BACTEC 9240 cabinet until positive followed by BCID panel testing according to the
40
manufacturer’s instructions (10).
41
The BCID panel correctly detected C. albicans (7/7) but did not detect C. dubliniensis
42
(0/20) from contrived blood cultures indicating that the BCID panel can discriminate these two
43
genetically and phenotypically similar species. These results indicated that blood cultures
44
positive for C. albicans by the BCID panel could be reported with confidence as C. albicans.
45
Furthermore, based on BCID analyte specificity for C. albicans, germ-tube-positive and BCID
46
panel-negative yeast can be presumptively identified as “Candida dubliniensis” with
47
confirmatory identification to follow.
48
Direct testing of positive blood cultures using nucleic acid amplification tests (NAATs)
49
provides rapid identification of pathogens and facilitates appropriate antimicrobial therapy.
50
Although NAATs can lack discriminatory power for highly related organisms, this study showed
51
that the FilmArray® BCID panel was capable of distinguishing C. albicans from the genetically
52
related and phenotypically similar C. dubliniensis from positive blood cultures. Future studies
53
will likely support C. dubliniensis as an important cause of human disease and that the addition
54
of this analyte to the BCID panel could help optimize clinical management of patients with
55
candidemia and improve epidemiology of candidemia as it relates to C. albicans and C.
56
dubliniensis.
57
REFERENCES
58 59
1. Delaloye J, Calandra T. 2014. Invasive candidiasis as a cause of sepsis in the critically ill
60
patient. Virulence. 5: 161-169.
61
2. Kollef M, Micek S, Hampton N, Doherty J, Kumar A. 2012. Septic shock attributed to
62
Candida infection: importance of empiric therapy and source control. Clin. Infect. Dis. 54:
63
1739-1746.
64
3. Falagas M, Roussos N, Vardakas K. 2010. Relative frequency of C. albicans and the
65
various non-albicans Candida spp among candidemia isolates from inpatients in various parts of
66
the world: a systematic review. Inter. J. Infect. Dis. 14: e954-e966.
67
4. Sullivan D, Westerneng T, Haynes K, Bennett D, Coleman D. 1995. Candida
68
dubliniensis sp. nov.: phenotypic and molecular characterization of a novel species associated
69
with oral candidosis in HIV-infected individuals. Microbiology. 141: 1507-1521.
70
5. Pfaller M, Woosley L, Messer S, Jones R, Castanheira M. 2012. Significance of
71
molecular identification and antifungal susceptibility of clinically significant yeasts and moulds
72
in a global antifungal surveillance programme. Mycopathologia. 174: 259-271.
73
6. Khan Z, Ahmad S, Joseph L, Chandy R. 2012. Candida dubliniensis: An appraisal of its
74
clinical significance as a bloodstream pathogen. PLoS One. 7: e32952.
75
7. Ells R, Kock L, Pohl C. 2009. Candida albicans or Candida dubliniensis. Mycoses. 54:
76
1-16.
77
8. Loreto E, Scheid L, Nogueira C, Zeni G, Santurio J, Alves S. 2010. Candida
78
dubliniensis: epidemiology and phenotypic methods for identification. Mycopathologia. 169:
79
431-443.
80
9. Iwen PC, Freifeld AG, Bruening TA, Hinrichs SH. 2004. Use of a panfungal PCR assay
81
for detection of fungal pathogens in a commercial blood culture system. J. Clin. Microbiol. 42:
82
2292-2293.
83
10. FilmArray® Blood Culture Identification Panel Instructions for Use, 2014
Table 1. Analysis of contrived blood cultures containing Candida albicans or C. dubliniensis using the FilmArray® Blood Culture Identification (BCID) panel. Organism
Strain
BCID Panel Result
Candida albicansa
B7560-08 B7706-08 B8627-09 B9005-09 B9010-09 B9028-09 ATCC 10231
Candida albicans Candida albicans Candida albicans Candida albicans Candida albicans Candida albicans Candida albicans
C. dubliniensisa
CU001 CU002 CU003 CU004 CU005 CU006 CU007 CU008 CU009 CU010 CU011 20-070-29 20-070-30 20-070-31 20-070-32 NE TNMC,120905 NE TNMC,121007 NE TNMC,072407 NE TNMC,012608 NE TNMC, 041108
NDa ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND
Abbreviation: ND, not detected a Identification of species included both phenotypic and genetic analyses
