BioluminescentAspergillus fumigatus, a New Tool for Drug Efficiency Testing and In Vivo Monitoring of Invasive Aspergillosis

Abstract

Aspergillus fumigatus is the main cause of invasive aspergillosis in immunocompromised patients, and only a limited number of drugs for treatment are available. A screening method for new antifungal compounds is urgently required, preferably an approach suitable for in vitro and in vivo studies. Bioluminescence imaging is a powerful tool to study the temporal and spatial resolutions of the infection and the effectiveness of antifungal drugs. Here, we describe the construction of a bioluminescent A. fumigatus strain by fusing the promoter of the glyceraldehyde-3-phosphate dehydrogenase gene from A. fumigatus with the luciferase gene from Photinus pyralis to control the expression of the bioluminescent reporter. A. fumigatus transformed with this construct revealed high bioluminescence under all tested growth conditions. Furthermore, light emission correlated with the number of conidia used for inoculation and with the biomass formed after different incubation times. The bioluminescent strains were suitable to study the effectiveness of antifungals in vitro by several independent methods, including the determination of light emission with a microplate reader and the direct visualization of light emission with an IVIS 100 system. Moreover, when glucocorticoid-treated immunosuppressed mice were infected with a bioluminescent strain, light emission was detected from infected lungs, allowing the visualization of the progression of invasive aspergillosis. Therefore, this new bioluminescence tool is suitable to study the in vitro effectiveness of drugs and the disease development, localization, and burden of fungi within tissues and may also provide a powerful tool to study the effectiveness of antifungals in vivo.

In vitro antifungal activity and cytotoxic effect of essential oils and extracts of medicinal and aromatic plants against Candida krusei and Aspergillus fumigatus

“…To obtain oils and extracts active against C. krusei and A. fumigatus is important because C. krusei has been recognized as a potentially multidrug-resistant pathogen and A. fumigatus is the main cause of invasive aspergillosis in immunocompromised patients; moreover, only a limited number of drugs are available (Brock et al, 2008, Pfaller et al, 2008. Additionally, both C. krusei and A. fumigatus, have the ability to form biofilms in biomedical devices that come in contact with skin, mucosal or inert surfaces (Quindós et al, 2009;Seidler et al, 2008).…”

Citral and carvone chemotypes from the essential oils of Colombian Lippia alba (Mill.) N.E. Brown: composition, cytotoxicity and antifungal activity

“…This finding is important because C. krusei has been recognised as a potential multidrug-resistant pathogen due to its intrinsic fluconazole resistance and recent reports showing a decreased susceptibility of the pathogen to both flucytosine and amphotericine B (Pfaller et al 2008). Aspergillus fumi-gatus is the main cause of invasive aspergillosis in immunocompromised patients and only a limited number of drugs are available for treatment (Brock et al 2008). In addition, recent studies have reported emergence of azole resistance in this species (Snelders et al 2008).…”

Duality of liver and kidney lesions after systemic infection of immunosuppressed and immunocompetent mice withAspergillus fumigatus

“…A. fumigatus strain C3. The bioluminescent A. fumigatus strain C3 was cultured on malt extract agar slants as described earlier 10 and bioluminescence of the strain was confirmed in vitro by determination of light emission from cultures grown for 8 h in RPMI medium by using an IVIS 100 system (Xenogen Corporation).…”

“…10 In vivo bioluminescence imaging allowed studying the progression of infection in individual animals. Significant differences in the temporal and spatial manifestation of organ infections were observed in dependence of the immune status of infected animals.…”