The interplay of phenotype and genotype in Cryptococcus neoformans disease

Abstract

Cryptococcus neoformans is an opportunistic fungal pathogen that causes life-threatening meningitis primarily in immunocompromised individuals. In order to survive and proliferate during infection, C. neoformans must adapt to a variety of stresses it encounters within the host. Patient outcome depends on the interaction between the pathogen and the host. Understanding the mechanisms that C. neoformans uses to facilitate adaptation to the host and promote pathogenesis is necessary to better predict disease severity and establish proper treatment. Several virulence phenotypes have been characterized in C. neoformans, but the field still lacks a complete understanding of how genotype and phenotype contribute to clinical outcome. Furthermore, while it is known that C. neoformans genotype impacts patient outcome, the mechanisms remain unknown. This lack of understanding may be due to the genetic heterogeneity of C. neoformans and the extensive phenotypic variation observed between and within isolates during infection. In this review, we summarize the current understanding of how the various genotypes and phenotypes observed in C. neoformans correlate with human disease progression in the context of patient outcome and recurrence. We also postulate the mechanisms underlying the genetic and phenotypic changes that occur in vivo to promote rapid adaptation in the host.

Keywords: Cryptococcus neoformans; genotype; microevolution; phenotype.

Figure 1. Pathogenic Cryptococcus species

Relatedness of the (A) Cryptococcus species complex and (B) subpopulations within C. neoformans. (A) There are five proposed species in the C. gattii species complex (yellow), and two species in the C. neoformans species complex (blue) [1,2,20]. (B) There are three evolutionarily distinct subpopulations within C. neoformans, as defined by molecular type: VNI, VNII, and VNB. VNII is very rarely found in clinical isolates. VNB is more diverse, was initially discovered in Botswana, and patients infected with these strains tend to have poor clinical outcomes. VNI causes the most cases of cryptococcosis and has three evolutionarily distinct subpopulations; ST93 and ST5 are in separate subpopulations [21,54,55].

Figure 2. Epidemiology and pathobiology of the Cryptococcus species complex

(A) C. neoformans is the most common species of Cryptococcus to cause infections globally, accounting for 95% of infections, compared with C. deneoformans (4%) and the C. gattii species complex (<1%) (left). In HIV+ patients, C. neoformans causes more than 99% of cases globally (right) [4]. (B) Infections by the C. gattii species complex most commonly occur in immunocompetent patients (64.3%), while infections by C. neoformans species complex primarily occur in patients with an immunocompromising condition (75%) [26–36]. (C) The most common clinical manifestations of cryptococcosis are skin lesions, pneumonia, and meningitis. Skin lesions, while rare, occur most often in patients infected with C. deneoformans, in approximately 14% of infections [41]. Patients with C. neoformans infections have skin lesions in 5% of cryptococcal meningitis cases [44]. Only 3% of C. gattii species complex cases have cutaneous cryptococcosis as a symptom [3]. The second most common manifestation of cryptococcosis is pneumonia. Pneumonia occurs most frequently in infections caused by C. gattii species complex infections, with 54% of C. gattii patients in the United States displaying lung involvement [36,45]. Species in the C. neoformans species complex manifest pulmonary cryptococcosis in 35% of clinical infections [41]. Cryptococcal meningitis occurs most frequently in the C. neoformans species complex, with over 80% of patients displaying meningitis symptoms [42]. Cryptococcal meningitis is less common in the C. gattii species complex with only 49% of patients in the United States developing meningitis [36].

Figure 3. Cell surface alterations and cell size changes in C. neoformans

Model illustrating the cell surface alterations and cell size phenotypes in C. neoformans. The outside of the C. neoformans cell consists of a cell wall (highlighted in pink) and polysaccharide capsule. Typical cells (5–7 μm in diameter) have a capsule predominantly composed of GXM and GalXM (orange box). The cell wall of typical cells contains chitin, chitosan, α-glucan, melanin, and β-glucan (left pink box). Titan cells (>10 μm in diameter) exhibit a thickened cell wall with an increase in chitin, decreased glucan, and a layer of mannan (right pink box) [59]. Micro cells (<1 μm in diameter) also display thickened cell walls [66]. Titanides are 2–4 μm in diameter and are oval in shape. The cell wall of titanides are thinner than typical cells [71]. Abbreviations: GXM, glucuronoxylomannan; GXMGal, glucuronoxylomannogalactin.

Figure 4. Mechanisms of in vivo microevolution

Model displaying possible mechanisms underlying the genetic and subsequent phenotypic changes seen during infection. Individual chromosomes are shown in black, and phenotype changes are represented by change in cell color. (A) During infection, TEs move throughout the genome. Transposon insertion into genes that may result in a range of phenotypes [111]. (B) Mutations in the MMR pathway result in cells with a higher rate of mutations (yellow stars) that may display various phenotypes [114,116]. (C) Polyploid titan cells undergo a reductive division to produce progeny with reduced genome sizes and genomic alterations, including cells with various whole chromosome aneuploidies (purple and green cells). Phenotype changes have also been observed in titan progeny that have no identified genomic alterations (tan cell) [76]. Abbreviation: MMR, mismatch repair.