Biomedical Engineering Reference
In-Depth Information
on a limited number of antifungals. Resistance to some of them (mainly
azoles) further complicates the therapy. Understanding the mechanisms
that fungi use to cause disease is therefore an important line of research
to reveal novel therapeutic approaches. Research on clinical relevant
fungi has been lagged historically due to limited molecular genetics.
Candida albicans
, the most important fungal pathogen accordingly to the
incidence of diseases that it causes, is diploid and lacks natural plasmids;
sexual recombination has only been recently recognized and its genetic
manipulation has been a bottleneck to understand the mechanisms of
pathogenesis (De Backer et al. 2000). The situation with other important
pathogenic fungi like
Cryptococcus neoformans
,
Aspergillus
spp.,
Histoplasma
capsulatum
and non-
albicans
spp. is rather similar. However, in the last
years, several tools have been developed: gene disruption schemes, more
effi cient transformation protocols and genetic markers are now available.
The analyses of the role of individual genes in key infectious processes of
these fungi are now feasible. In addition, the availability of their genomes
has been also essential in basic research.
The study of microbial pathogenesis is, at least in part, the study of
the regulatory pathways triggered inside -and by- the pathogen during
infection. The fl exibility and adaptability of the microbial cell has enabled
many pathogenic species to face different environmental conditions during
infection (Alonso-Monge et al. 2006, Hube 2004, Justice et al. 2008, Román et
al. 2007). The transcriptional changes that underlie this ability can determine
the success of the pathogen inside the host. Gene reporters are therefore a
valuable tool to understand microbial pathogenesis which has contributed
signifi cantly to our understanding of eukaryotic gene expression and
regulation. Reporter genes have numerous applications, both
in vitro
and
in vivo
, but they are frequently used as indicators of transcriptional activity
in cells to monitor the cellular events associated with signal transduction
and gene expression (Kain and Ganguly 2001). Commonly, a reporter gene is
fused to a specifi c sequence of DNA and the construct is then incorporated
into the genome of host cells. The presence of the reporter is confi rmed
by directly measuring the activity of the reporter protein. Different gene
reporters have been used in the last years. Fluorescent proteins (FPs) (mainly
from the jellyfi sh
Aequorea victoria
) are widely used and can be used to
localize gene products within the cell. Other systems include metabolic
enzymes (β-galactosidases among them), drug resistance genes and several
others (see (Eckert and Muhlschlegel 2009) for an excellent recent review).
Here, we will summarize the main applications of bioluminescent
gene reporters in the study of infections caused by the fungal pathogens
Candida albicans, Aspergillus fumigatus
and
Criptococcus neoformans
, the most
common causes of fungal infections. We will highlight recent fi ndings that
