Using non-mammalian hosts to study fungal virulence and host defense

doi:10.1016/j.mib.2006.06.004

Current Opinion in Microbiology

Volume 9, Issue 4, August 2006, Pages 346-351

https://doi.org/10.1016/j.mib.2006.06.004 Get rights and content

Non-mammalian hosts have been used to study host–fungal interactions. Hosts such as Drosophila melanogaster, Caenorhabditis elegans, Acathamoeba castellanii, Dictyostelium discoideum, and Galleria mellonella have provided means to examine the physical barriers, cellular mechanisms and molecular elements of the host response. The Drosophila host-response to fungi is mediated through the Toll pathway, whereas in C. elegans the host-response is TIR-1-dependent. Virulence traits that are involved in mammalian infection are important for the interaction of fungi with these hosts. Screening of fungal virulence traits using mutagenized fungi to determine changes in fungal infectivity of non-mammalian hosts has been used to identify novel virulence proteins used to infect C. elegans such as Kin1 (a serine/threonine protein kinase) and Rom2 (a Rho1 guanyl-nucleotide exchange factor) from Cryptococcus neoformans. These heterologous non-mammalian hosts highlight the similarities and differences between different hosts in fungal pathogenesis and they complement studies in mammalian systems and those using other genetic approaches.

Introduction

Recently, model hosts including Drosophila and Caenorhabditis elegans [1, 2, 3, 4, 5] have been used for the study of both bacterial and fungal pathogenesis. The hypothesis is that host-defense mechanisms developed over time because the non-mammalian hosts shared an environment with fungal pathogens and the host-defense mechanisms probably provided a means to survive pathogen attack. From another perspective, the fungal pathogens might have developed virulence factors to survive being consumed by organisms that occupy the same environment. As the fungal microorgansisms move into a susceptible mammalian hosts, they are able to use these same behaviors, which enable fungal survival in non-mammalian hosts, to convert into an opportunistic pathogen.

This report focuses on five recently emerged non-mammalian host models for the study of pathogenic fungi: Drosophila melanogaster, C. elegans, Acathamoeba castellanii, Dictyostelium discoideum and Galleria mellonella. Each of these five model hosts provides benefits for elucidating the host defense and virulence mechanisms of fungal pathogenesis (Table 1). Although there are many similarities between the various mechanisms for defense against fungal pathogens amongst these heterologous hosts, several differences also exist and are explored in this review. Two additional notable non-mammalian host systems that have been used to study cellular defenses against fungal pathogens are the mosquito Culex quinquefasciatus [6], and the German cockroach Blattella germanica (Table 1) [7]. Although not ideal as model hosts, they have both provided some information about phagocytic host defenses, including the means by which nodules are formed around fungal cells [6, 7] and they are not discussed further in this review.

Section snippets

Drosophila and Toll pathway dependent defense

D. melanogaster is an effective model for the study of fungal pathogenesis as several genetic tools are available for this organism, including the sequenced genome [8] and an oligonucleotide DNA microarray of fungal-challenged flies, for detection of immune defense genes [9]. In the laboratory, D. melanogaster can be infected with fungal pathogens using various methods, such as injection, direct spraying of fungal spores onto the flies or by ingestion.

This insect model has provided insight into

C. elegans defense is TIR-1 mediated

Additional information can be gained by using C. elegans as a model host because its genome has been sequenced [17] and many if the functions of its genes are currently being identified through genetic studies. One functional tool that is readily available is an RNAi (RNA interference) library, which uses double-stranded RNA to silence gene expression, that can be used to determine the function of genes of interest [18]. A difficulty arises, however, in the method of fungal delivery. In the

Amoebae use phagocytic defense

A. castellanii is a free-living heterologous host, which has been studied as a model for its phagocytic abilities. As a model fungal host, A. castellanii is able to kill C. albicans and Saccharomyces cerevisiae [23]. Phagocytosis by this amoeba involves enclosure of the fungal cell into a vacuole, similar to the process used by macrophages [24]. Importantly, A. castellanii is killed by C. neoformans. As the C. neoformans cells continue to replicate, they produce polysaccharides that are

Slime mold also uses phagocytic defense

The slime mold D. discoideum is another amoeba used as a model host because it is haploid, has a sequenced genome [26] and mutants are available. It has been used as a general model to aid our understanding of phagocytic processes [27, 28]. Cells can be grown at 24.5 °C, which is agreeable to fungal growth, but cannot survive at temperatures exceeding 27 °C. Available genetic techniques include mutagenesis, RNAi and targeted gene-disruption [29]. This free-living amoeba can phagocytose C.

The greater wax moth uses the hemolymph system for defense

Whereas Galleria does not offer the classical benefits of a model host (e.g. a sequenced genome) it does offer the unique benefit of being able to be maintained under various temperature conditions ranging from 25 °C to 37 °C. The ability to study pathogens at 37 °C enables the study of temperature-related virulence traits [30]. Notably, survival at mammalian temperatures is not shared by the other non-mammalian host systems presented in this review. Galleria also has the benefit of being facile

Virulence traits identified in non-mammalian hosts

Certain features of fungi have been found to be necessary for virulence. One important example of a virulence trait that transcends host systems is the cryptococcal capsule. When acapsular Cryptococcus mutants are exposed to amoebae, they are phagocytosed at a higher frequency [23], indicating that the capsule protects the fungus against phagocytosis by this heterologous host. The capsule is also a virulence factor against C. elegans, as determined by a slower rate of killing of C. elegans and

Conclusions

Fungal pathogenicity continues to develop in nature, and the continued emergence of new environmentally prevalent fungal pathogens is an indicator of the need for continued research towards the understanding of host defense and pathogen virulence [45]. Pathogenic fungi might have evolved by developing virulence factors in response to the hosts that they encounter; this could help explain the observed patterns in the development of fungal virulence in mammalian and non-mammalian model hosts.

Update

As described in this review, non-mammalian hosts can be used as a means to screen for virulence factors. Recently, it has been shown that non-mammalian hosts can also be used in screening techniques as vessels for in vivo screens in order to identify antifungal compounds. Non-mammalian hosts provide a means to initially screen antifungal compounds that specifically affect the pathogen but do not affect the host. In one example, Tol-deficient D. melanogaster was used to test antifungal activity

References and recommended reading

Papers of particular interest, published within the annual period of review, have been highlighted as:

• of special interest
•• of outstanding interest

Acknowledgements

Financial support was provided by a K08 award AI63084-01 from the National Institutes of Health and a New Scholar Award in Global Infectious Diseases of the Ellison Medical Foundation, to EM.

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Using non-mammalian hosts to study fungal virulence and host defense

Introduction

Section snippets

Drosophila and Toll pathway dependent defense

C. elegans defense is TIR-1 mediated

Amoebae use phagocytic defense

Slime mold also uses phagocytic defense

The greater wax moth uses the hemolymph system for defense

Virulence traits identified in non-mammalian hosts

Conclusions

Update

References and recommended reading

Acknowledgements

J Invertebr Pathol

Genes Dev

Nature

Proc Natl Acad Sci USA

J Cell Sci

FEMS Immunol Med Microbiol

Infect Immun

Infect Immun

Genetics

Are innate immune signaling pathways in plants and animals conserved?

Nat Immunol

Worms and flies as genetically tractable animal models to study host–pathogen interactions

Infect Immun

Genetic models in pathogenesis

Annu Rev Genet

The art of serendipity: killing of Caenorhabditis elegans by human pathogens as a model of bacterial and fungal pathogenesis

Expert Rev Anti Infect Ther

Caenorhabditis elegans: an emerging genetic model for the study of innate immunity

Nat Rev Genet

Aspergillosis in German cockroach Blattella germanica (L.) (Blattoidea: Blattellidae)

Mycopathologia

The genome sequence of Drosophila melanogaster

Science

A genome-wide analysis of immune responses in Drosophila

Proc Natl Acad Sci USA

Identification of Drosophila gene products required for phagocytosis of Candida albicans

PLoS Biol

Immune-deficient Drosophila melanogaster: a model for the innate immune response to human fungal pathogens

J Immunol

The dorsoventral regulatory gene cassette spatzle/Toll/cactus controls the potent antifungal response in Drosophila adults

Cell

Phylogenetic perspectives in innate immunity

Science

A drosomycin-GFP reporter transgene reveals a local immune response in Drosophila that is not dependent on the Toll pathway

EMBO J

Insect immunity. Septic injury of Drosophila induces the synthesis of a potent antifungal peptide with sequence homology to plant antifungal peptides

J Biol Chem