This site needs JavaScript to work properly. Please enable it to take advantage of the complete set of features!
Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

NIH NLM Logo
Log in
Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2011 Jan 13;6(1):e16076.
doi: 10.1371/journal.pone.0016076.

Cryptococcus gattii virulence composite: candidate genes revealed by microarray analysis of high and less virulent Vancouver island outbreak strains

Affiliations

Cryptococcus gattii virulence composite: candidate genes revealed by microarray analysis of high and less virulent Vancouver island outbreak strains

Popchai Ngamskulrungroj et al. PLoS One. .

Abstract

Human and animal cryptococcosis due to an unusual molecular type of Cryptococcus gattii (VGII) emerged recently on Vancouver Island, Canada. Unlike C. neoformans, C. gattii causes disease mainly in immunocompetent hosts, despite producing a similar suite of virulence determinants. To investigate a potential relationship between the regulation of expression of a virulence gene composite and virulence, we took advantage of two subtypes of VGII (a and b), one highly virulent (R265) and one less virulent (R272), that were identified from the Vancouver outbreak. By expression microarray analysis, 202 genes showed at least a 2-fold difference in expression with 108 being up- and 94 being down-regulated in strain R265 compared with strain R272. Specifically, expression levels of genes encoding putative virulence factors (e.g. LAC1, LAC2, CAS3 and MPK1) and genes encoding proteins involved in cell wall assembly, carbohydrate and lipid metabolism were increased in strain R265, whereas genes involved in the regulation of mitosis and ergosterol biosynthesis were suppressed. In vitro phenotypic studies and transcription analysis confirmed the microarray results. Gene disruption of LAC1 and MPK1 revealed defects in melanin synthesis and cell wall integrity, respectively, where CAS3 was not essential for capsule production. Moreover, MPK1 also controls melanin and capsule production and causes a severe attenuation of the virulence in a murine inhalational model. Overall, this study provides the basis for further genetic studies to characterize the differences in the virulence composite of strains with minor evolutionary divergences in gene expression in the primary pathogen C. gattii, that have led to a major invasive fungal infection outbreak.

PubMed Disclaimer

Conflict of interest statement

Competing Interests: The authors have declared that no competing interests exist.

Figures

Figure 1
Figure 1. Melanin production of strains R265 and R272.
Strain R265 produced more melanin on all melanin inducing media, e.g. A) bird seed agar at 48 hr at 30°C and B) caffeic acid agar at 48 h at 37°C. C) Quantitative measurement of the melanin production shows a higher laccase activity for strain R265 with either dopamine (Dopa) or epinephrine (Epi) as substrates (standard error bar +/− 2SE).
Figure 2
Figure 2. Capsule formation of strains R265 and R272.
Yeast cells were grown in DME at 37°C with 5% CO2 and RPMI-1640 medium at 37°C with 5% CO2 for 72 h.
Figure 3
Figure 3. Cell wall integrity tests for strain R265 vs. strain R272 isolates after incubation in YPD medium supplemented with CaCl2, calcofluor white (CFW), caffeine and Congo red.
Figure 4
Figure 4. Mating reactions of the Vancouver Island isolates showing similar mating properties, with typical bacilli shaped basidiospores being observed for both strains .
The dark arrows present basidia and the white arrows identify basidiospores.
Figure 5
Figure 5. Phenotypic study of lac1Δ and cas3Δ mutants.
A) The lac1Δ mutant strain exhibited lower melanin production on caffeic acid agar at 30°C as compared with the wild-type R265 strain and its reconstituted strain (lac1Δ::LAC1). B) No difference in capsule formation was observed in the cas3Δ mutant on 5% CO2 DME at 37°C as compared to the wild-type R265 strain and the reconstituted strain (cas3Δ:: CAS3Δ).
Figure 6
Figure 6. Growth of the wild type R265 stain, mpklΔ mutant and reconstituted mpk1Δ::MPK1 strain under different temperatures and cell wall stress conditions.
Cells were grown on YPD at different temperatures or in the presence of cell wall disturbing agents as indicated.
Figure 7
Figure 7. A) Phenotypic impact of mpklΔ on capsule, melanin, and control of laccase gene transcription.
Capsule production test in DME with 5%CO2 at 37°C, B) Melanin synthesis test on Caffeic acid agar at 37°C, C) MPK1, LAC1 and LAC2 transcription level of the mpk1Δ mutant (white bars) were 0, 0.07 and 0.33 times to the wild type R265 strain, respectively. No significant differences between the transcriptions between the wild type and the mpk1Δ::MPK1 strain.
Figure 8
Figure 8. The organ burden of the wild type R265 strain, the mpk1Δ mutant and the complemented strain in Balb/c mice lungs.
The mpk1 mutant was not detected in any of the lungs (*).

References

    1. Sorrell TC. Cryptococcus neoformans variety gattii. Med Mycol. 2001;39:155–168. - PubMed
    1. Casadevall A, Perfect JR. Washington, DC: ASM press; 1998. Cryptococcus neoformans.
    1. Kwon-Chung KJ, Bennett JE. Epidemiologic differences between the two varieties of Cryptococcus neoformans. Am J Epidemiol. 1984;120:123–130. - PubMed
    1. Datta K, Bartlett KH, Marr KA. Cryptococcus gattii: Emergence in Western North America: Exploitation of a Novel Ecological Niche. Interdiscip Perspect Infect Dis. 2009;2009:176532. - PMC - PubMed
    1. MacDougall L, Kidd SE, Galanis E, Mak S, Leslie MJ, et al. Spread of Cryptococcus gattii in British Columbia, Canada, and detection in the Pacific Northwest, USA. Emerg Infect Dis. 2007;13:42–50. - PMC - PubMed

Publication types

Full text links
Public Library of Science full text link Public Library of Science Free PMC article
Cite

AltStyle によって変換されたページ (->オリジナル) /