Soo Chan Lee

In recent years, we have seen an increase in the number of immunocompromised cohorts as a result of infections and/or medical conditions, which has resulted in an increased incidence of fungal infections. Although rare, the incidence of infections caused by fungi belonging to basal fungal lineages is also continuously increasing. Basal fungal lineages diverged at an early point during the evolution of the fungal lineage, in which, in a simplified four-phylum fungal kingdom, Zygomycota and Chytridiomycota belong to the basal fungi, distinguishing them from Ascomycota and Basidiomycota. Currently there are no known human infections caused by fungi in Chytridiomycota; only Zygomycotan fungi are known to infect humans. Hence, infections caused by zygomycetes have been called zygomycosis, and the term “zygomycosis” is often used as a synonym for “mucormycosis.” In the four-phylum fungal kingdom system, Zygomycota is classified mainly based on morphology, including the ability to form coenocytic (aseptated) hyphae and zygospores (sexual spores). In the Zygomycota, there are 10 known orders, two of which, the Mucorales and Entomophthorales, contain species that can infect humans, and the infection has historically been known as zygomycosis. However, recent multilocus sequence typing analyses (the fungal tree of life [AFTOL] project) revealed that the Zygomycota forms not a monophyletic clade but instead a polyphyletic clade, whereas Ascomycota and Basidiomycota are monophyletic. Thus, the term “zygomycosis” needed to be further specified, resulting in the terms “mucormycosis” and “entomophthoramycosis.” This review covers these two different types of fungal infections.

Summary: Sexual development is extant in virtually all eukaryotic species, including throughout the kingdom Fungi. Positioned within the opisthokonts along with metazoans, fungi serve as model systems to elucidate the genetics and impact of sexual development. Basal fungal lineages such as the Mucoralean fungi provide a unique basis to study sexual reproduction, in which common ancestral traits found in both animal and fungal lineages may be conserved. This review discusses the sexual development, sex loci, and evolution of the sex locus in the Mucoralean fungi, which sheds light on our understanding of the evolution and functions of sex.

Food-borne pathogens are ongoing problems, and new pathogens are emerging. The impact of fungi, however, is largely underestimated. Recently, commercial yogurts contaminated with Mucor circinelloides were sold, and >200 consumers became ill with nausea, vomiting, and diarrhea. Mucoralean fungi cause the fatal fungal infection mucormycosis, whose incidence has been continuously increasing. In this study, we isolated an M. circinelloides strain from a yogurt container, and multilocus sequence typing identified the strain as Mucor circinelloides f. circinelloides . M. circinelloides f. circinelloides is the most virulent M. circinelloides subspecies and is commonly associated with human infections, whereas M. circinelloides f. lusitanicus and M. circinelloides f. griseocyanus are less common causes of infection. Whole-genome analysis of the yogurt isolate confirmed it as being close to the M. circinelloides f. circinelloides subgroup, with a higher percentage of divergence with the M. circinelloides f. lusitanicus subgroup. In mating assays, the yogurt isolate formed sexual zygospores with the (−) M. circinelloides f. circinelloides tester strain, which is congruent with its sex locus encoding SexP, the (+) mating type sex determinant. The yogurt isolate was virulent in murine and wax moth larva host systems. In a murine gastromucormycosis model, Mucor was recovered from fecal samples of infected mice for up to 10 days, indicating that Mucor can survive transit through the GI tract. In interactions with human immune cells, M. circinelloides f. lusitanicus induced proinflammatory cytokines but M. circinelloides f. circinelloides did not, which may explain the different levels of virulence in mammalian hosts. This study demonstrates that M. circinelloides can spoil food products and cause gastrointestinal illness in consumers and may pose a particular risk to immunocompromised patients. IMPORTANCE The U.S. FDA reported that yogurt products were contaminated with M. circinelloides , a mucoralean fungal pathogen, and >200 consumers complained of symptoms, including vomiting, nausea, and diarrhea. The manufacturer voluntarily withdrew the affected yogurt products from the market. Compared to other food-borne pathogens, including bacteria, viruses, and parasites, less focus has been placed on the risk of fungal pathogens. This study evaluates the potential risk from the food-borne fungal pathogen M. circinelloides that was isolated from the contaminated commercial yogurt. We successfully cultured an M. circinelloides isolate and found that the isolate belongs to the species M. circinelloides f. circinelloides , which is often associated with human infections. In murine and insect host models, the isolate was virulent. While information disseminated in the popular press would suggest this fungal contaminant poses little or no risk to consumers, our results show instead that it is capable of causing significant infections in animals.

Mucor circinelloides is a zygomycete fungus and an emerging opportunistic pathogen in immunocompromised patients, especially transplant recipients and in some cases otherwise healthy individuals. We have discovered a novel example of size dimorphism linked to virulence. M. circinelloides is a heterothallic fungus: (+) sex allele encodes SexP and (−) sex allele SexM, both of which are HMG domain protein sex determinants. M. circinelloides f. lusitanicus (Mcl) (−) mating type isolates produce larger asexual sporangiospores that are more virulent in the wax moth host compared to (+) isolates that produce smaller less virulent sporangiospores. The larger sporangiospores germinate inside and lyse macrophages, whereas the smaller sporangiospores do not. sexMΔ mutants are sterile and still produce larger virulent sporangiospores, suggesting that either the sex locus is not involved in virulence/spore size or the sexP allele plays an inhibitory role. Phylogenetic analysis supports that at least three extant subspecies populate the M. circinelloides complex in nature: Mcl, M. circinelloides f. griseocyanus, and M. circinelloides f. circinelloides (Mcc). Mcc was found to be more prevalent among clinical Mucor isolates, and more virulent than Mcl in a diabetic murine model in contrast to the wax moth host. The M. circinelloides sex locus encodes an HMG domain protein (SexP for plus and SexM for minus mating types) flanked by genes encoding triose phosphate transporter (TPT) and RNA helicase homologs. The borders of the sex locus between the three subspecies differ: the Mcg sex locus includes the promoters of both the TPT and the RNA helicase genes, whereas the Mcl and Mcc sex locus includes only the TPT gene promoter. Mating between subspecies was restricted compared to mating within subspecies. These findings demonstrate that spore size dimorphism is linked to virulence of M. circinelloides species and that plasticity of the sex locus and adaptations in pathogenicity have occurred during speciation of the M. circinelloides complex. Mucormycosis is recognized as an emerging infectious disease. Compared to other fungal infections, mucormycosis results in high mortality: ∼50% of overall infections and >90% in disseminated infections. There is therefore an ongoing need to study these fungal pathogens. However, surprisingly little is known about the pathogenesis of mucormycosis. Our findings reveal a correlation between sporangiospore size and virulence: larger sporangiospores are more virulent than small spores. Larger spores start invasive hyphal growth immediately upon phagocytosis by host immune cells, whereas smaller spores have a long period of isotropic growth. Differential host immune response explains their difference in virulence, whereby larger spores escape host immune cells by germinating inside of macrophages. Our findings revealed an example of adaptation through fungal cell gigantism, which enable pathogenic fungi to survive within and establish infection in the host. Knowledge of the mechanisms of pathogenicity and the molecular basis of the sexual cycle in M. circinelloides will contribute to advance our understanding of pathogenic zygomycetes.