1. Could you begin by introducing the work of
the Pathogenic Yeast Research Group?
The Pathogenic Yeast Research Group focuses
on diseases caused by yeasts, especially those
belonging to the Candida and Cryptococcus
genera. My focus is the role of bioactive lipids
produced by Candida species. I am interested
in understanding how fatty acids, including
those obtained from the host, are used by
these yeasts to produce lipid metabolites
(oxylipins) that contribute to their survival
in the host, as well as to tissue damage
caused by the yeast during infection.
What is the immunological role of bioactive
lipids in hosts of these diseases?
Certain oxylipins can modulate the mammalian
immune system and often have several different
functions in distinct cell types. One of the
best-studied oxylipins produced by both the
host and pathogenic yeasts is prostaglandin E2
(PGE2
), which can shift host immune responses
in favour of the pathogen. It can inhibit the type
of immune response needed to protect against
Candida infection and induce an immune
response that may lead to associated chronic
or disseminated diseases. It can also stimulate
another type of immune response that may
cause uncontrolled inflammation and damage
to the host. Thus the production of PGE2
during infections can benefit the pathogen.
Another important effect caused by PGE2
is
tissue eosinophilia leading to tissue damage,
which is characteristic of some chronic
fungal infections. Interestingly, it is not only
the host that is affected: PGE2
has biological
activity on yeast by stimulating germ tube
formation in Candida species. This change in
morphology, from unicellular to filamentous, is
considered the start of biofilm formation and is
associated with an increased ability to invade
tissue. Interestingly, biofilms also produce
more PGE2
than unicellular yeast forms.
Can you highlight some of the roles that lipids
have as ‘inter-kingdom’ signalling molecules
in yeast-associated multispecies infections?
This is an exciting new research focus for us.
It is increasingly being recognised that many
infections are not caused by one organism.
Interaction between pathogen and host may be
influenced by other non-pathogenic organisms.
Although it is known that lipid metabolites
can act as signals between organisms – even
those belonging to different kingdoms – very
little work has been done on the bioactive
lipids produced by mixed communities. An
example of a multispecies interaction is the
biofilms consisting of Candida albicans and
the Pseudomonas aeruginosa bacterium in
the lungs of patients with cystic fibrosis.
Are there particularly advanced
imaging techniques that you
incorporate in your studies?
Most studies of different antimicrobial effects
start with the production of standardised
biofilms, which are notoriously resistant
to antifungal treatment. To assess the
effectiveness of potential antifungal drugs,
biofilms are stained for viability and visualised
using a confocal laser scanning microscope.
The potential of drugs to decrease biofilm
viability or metabolic activity is assessed in an
XTT-assay based on the ability of mitochondrial
dehydrogenases to convert a substrate to a
coloured formazan. The effect of compounds
on the ultrastructure – including surface
characteristics and organelle structure
– is examined by electron microscopy.
Our group pioneered the use of nano-scanning
Auger microscopy (Nano-SAM) on yeast
cells. With Nano-SAM, nanometre-thin slices
of a yeast cell enable observation of its 3D
structure. The cell elements can also be
mapped. Similarly, time-of-flight secondary
ion mass spectrometry (TOF-SIMS) is used to
map the distribution of certain compounds,
including oxylipins, in cells. We also use
various assays to measure oxidative stress.
Has collaboration with other research
groups aided your research?
My own speciality is enhanced by that of Dr
Olihile Sebolai who focuses on Cryptococcus
species and immunology. Dr Chantel Swart
is an expert in ultrastructural analyses and
was a leading member of the group that
pioneered the application of Nano-SAM to
yeast research. In this, we also collaborate
with our university’s physics department. The
fourth member of the group is Professor J
Albertyn, who is an expert in yeast molecular
biology and yeast signal transduction.
To date, has your work drawn any
interesting conclusions?
An obvious conclusion is that the lipids of
pathogenic yeasts are much more than
structural compounds or reserve material; like
mammalian and plant lipids, they have other
functions. It has also become obvious that much
of the knowledge regarding metabolic pathways
and enzymes responsible for production of
mammalian oxylipins does not translate directly
to yeasts. With the completion of each project,
we end up with more questions – the study of
these bioactive compounds is always exciting.
Dr Carolina Pohl describes the work of her research team, one of few groups in the world that are
exploring the roles of bioactive fatty acids in the development of yeast infections
The lipid connection
70 INTERNATIONAL INNOVATION
DR CAROLINA POHL

2. Oxylipins are central elements
in adaptation and survival;
in some instances they
aid organisms to defend
themselves, propagate or
prompt symbiotic relationships,
whereas in other situations they
can induce pathogenesis
A RICH AND diverse population of bacteria
and fungi live in a mutualistic fashion within
the human body. Unfortunately, when their
normal balance is disturbed, their populations
can respond with rapid growth, giving rise to
infection. Because such disturbances typically
follow an event of ill health, the impact of such
infections may have severe consequences,
especially for already immune-compromised
individuals. For example, fungal infections
caused by the Candida yeast are common
among individuals who are HIV-positive and
can inhibit their nutrition. Candida species
infections can also exacerbate infection
by the tuberculosis bacterium or lead to
deterioration in patients with cystic fibrosis
or head and neck cancers. Similarly, infection
by pathogenic Cryptococcus causes serious
fungal infections that can result in pneumonia-
like symptoms, infertility, central nervous
system damage, encephalitis or death.
In the Republic of South Africa – where the
incidence of tuberculosis is high, low birth
weight and infant mortality are prevalent,
sexually-transmitted infections are
widespread and HIV infection is at epidemic
proportions – pathogenic yeasts lie at the
intersection of a number of priority health
issues: women and children’s health, and
infectious and noncommunicable diseases.
ADDRESSING THE PROBLEM
The Pathogenic Yeast Research Group at the
University of the Free State in South Africa is
focused on the role of bioactive lipids, the fatty
molecules fundamental to cellular growth in
both hosts and pathogenic yeasts, with the
aim of furthering development of new drugs
and treatments to counteract yeast infections.
The Group is particularly interested in the
role of these lipids in multispecies infections,
as found in cystic fibrosis, peritonitis and
tuberculosis, and in cancer-related and female
reproductive tract infections. In addition, they
aim to discover more about the vulnerability
of yeasts to antimicrobial compounds, in
order to counteract the growing problem
of multidrug-resistant yeast strains.
This Group was the first to discover yeast
oxylipins, or oxidised fatty acids, including
prostaglandins. One member, Dr Carolina
Pohl, is an expert in lipid metabolism in
yeasts and has been involved in the discovery
of the antifungal effect of acetylsalicylic
acid – an oxylipin that is a mitochondrial
inhibitor and anti-inflammatory compound.
Since making these findings, her work has
mainly concentrated on biofilm formation
and subsequent infection by the Candida
species C. albicans and C. dubliniensis – highly
related strains that behave in similar ways,
which makes them ideal as model systems
of pathogenic yeasts. “These two species
both have the ability to produce the lipid
signalling molecule prostaglandin E2
(PGE2
),
and react in the same way to this chemical
by producing germ tubes,” Pohl explains.
“With both, the formation of biofilms, and
subsequent increase in ability to form PGE2
,
is an important disease-causing mechanism
through the stimulation of inflammation.”
THE ROLE OF OXYLIPINS
Oxylipins have ancient origins as signalling
molecules, elements of which are conserved
across different kingdoms of life. Pohl recently
undertook a review of oxylipin mediation of
diverse inter-organism communication that
leads to beneficial or deleterious interaction:
between plants, animals, fungi and bacteria.
Although the mechanism by which yeasts
‘sense’ the presence of bioactive lipids in their
environment or host remains unclear, the
review has led Pohl to conclude that oxylipins
Research into the molecular mechanisms of common yeasts
and bacteria at the University of the Free State in South Africa
seeks to exploit the roles of lipid molecules as instigators of
disease symptoms towards development of new therapeutics
and antifungal agents
Pathways of infection
Ultrastructure Antifungal drugs
Molecular
biology
Immunology
Bioactive
lipids
www.internationalinnovation.com 71

3. EFFECT OF POLYUNSATURATED FATTY
ACIDS ON PATHOGENICITY OF CANDIDA
ALBICANS AND CANDIDA DUBLINIENSIS
OBJECTIVES
• To understand the metabolism of arachidonic acid
to proinflammatory oxylipins by Candida species
• To understand the signalling pathways of oxylipins
leading to morphological changes in Candida species
• To investigate the lipid metabolisms of
mixed species biofilms and the influence
of the produced oxylipins on the host
KEY COLLABORATORS
Professor J Albertyn; Dr O Sebolai; Dr C Swart,
University of the Free State, South Africa
FUNDING
South African National Research Foundation
CONTACT
Dr Carolina Pohl
Associate Professor
University of the Free State
PO Box 339
Bloemfontein 9300
Republic of South Africa
T +27 824 540 072
E pohlch@ufs.ac.za
twitter.com/PathYeastRes
www.facebook.com/PathYeastRes
www.linkedin.com/pub/carlien-pohl/53/7a7/905
http://bit.ly/RGPohl
DR CAROLINA POHL is an
associate professor in the
Department of Microbial,
Biochemical and Food
Biotechnology at the University
of the Free State, South Africa. Her research focus
is Bioactive Lipids in pathogenic Candida species.
She had been part of a research group studying the
distribution and functions of these compounds in
environmental yeasts since completing her PhD in 1999.
She started her own research group in 2006, focusing
on yeasts that are important human pathogens.
INTELLIGENCE
are central elements in adaptation and
survival; in some instances they aid organisms
to defend themselves, propagate or prompt
symbiotic relationships, whereas in other
situations they can induce pathogenesis.
Pohl’s research has shown that some yeast
oxylipins work by mimicking host signalling
molecules to induce an immune response.
They can also bring about structural changes
in the yeast that then assist its colonisation
activities or help it to evade host defences.
Additonally, lipids are involved in cross-talk
between different cellular compartments, such
as cell walls and mitochondria, which can aid
the acquisition of multidrug resistance, but
also underlies their potential as antifungal
agents. Hence Pohl recently conducted an
assessment of the effects of an omega-3 fatty
acid on C. albicans and C. dubliniensis growth.
In this study, the long-chain stearidonic fatty
acid stopped biofilm formation by increasing
reactive oxygen species production and the rate
of apoptosis. The study also suggested that
stearidonic acid can act in synergy with some
antifungal compounds. This was the first finding
that it is not just medium-chain fatty acids that
can stop the growth of pathogenic yeasts.
OXYLIPINS IN MULTISPECIES INFECTION
Pohl’s investigation into the effects of
supplementing levels of an omega-6 fatty acid,
sciadonic acid, in epithelial cells on infection
with C. albicans and C. dubliniensis, showed
changes in PGE2
concentrations, but no
reduction in those of benign anti-inflammatory
omega-3 fatty acids. Furthermore a trial of
the effects of aromatic amine phenothiazine
– a constituent of several types of drugs in
medical and veterinary use – on C. albicans
biofilms, demonstrated significant reductions
in biofilm metabolic activity and biomass
and large reductions in PGE2
production.
Given their abilities to mediate communication
between species, bioactive lipids in single-
species yeast or host contexts are not,
however, Pohl’s only line of
research. Multispecies infections,
particularly concerning the
production of oxylipins and
subsequent reactions of the
host and pathogens in mixed
populations of yeasts and bacteria,
are a current key area of study:
“Finding out more about the types
and levels of oxylipins produced by
different organisms when growing
together may tell us more about
the influence of interaction on each
microbe, and the potential effects
on the host,” she elaborates.
Such information would be particularly
relevant for understanding the oxylipin
role in generating the symptoms of mixed
pathogen diseases, such as cystic fibrosis.
TRANSLATING IN VITRO RESULTS
In a recent project, Pohl explored the principle
of inhibiting the release of arachidonic
acid – a necessary component of the PGE2
production pathway. This was achieved by
substituting sciadonic acid for arachidonic
acid in C. albicans and C. dubliniensis biofilms
grown in vitro. Her team found that the
levels of PGE2
were indeed reduced and
now aim to further validate this result in
experiments using in vivo infection models.
Potential approaches to mitigating yeast
infections currently being tested by Pohl’s
team include preventing the production of
PGE2
in yeast biofilms, so reducing their
proinflammatory effects: “An ideal drug
target may be a metabolic pathway that is
unique to yeasts and not found in the host,”
Pohl muses. She is also working to identify
the pathway that allows Candida species to
react to the presence of oxylipins produced
by either the host or other yeast cells.
Pohl is confident that greater knowledge of
the role of lipids and the mechanisms of lipid
metabolism of pathogenic yeasts will lead to
significant advances in the near future. With
a focus on their structures and their ability to
metabolise fatty acids, she now plans to follow
up several candidate enzymes that her team
have already identified as being implicated
in prostaglandin production and virulence
in Candida. With the support of the rest of
the Pathogenic Yeast Research Group, Pohl
also intends to further explore the influence
of arachidonic acid on gene expression
of pathogenic yeasts, using genomic and
proteomic analysis techniques. She intends for
this to deliver as yet unanticipated antifungal
drug targets and lead to new applications in
diverse fields, from medicine to agriculture.
The Pathogenic Yeast Research Group
72 INTERNATIONAL INNOVATION