Genomics ofCryptococcus neoformans

Abstract

27C. neoformans var. grubii (C. neoformans) is an environmentally acquired pathogen causing 181 000 28 HIV-associated deaths each year. We used whole genome sequencing (WGS) to characterise 699 29 isolates, primarily C. neoformans from HIV-infected patients, from 5 countries in Asia and Africa. We 30 found that 91% of our clinical isolates belonged to one of three highly clonal sub-clades of VNIa, 31 which we have termed VNIa-4, VNIa-5 and VNIa-93. Parsimony analysis revealed frequent, long 32 distance transmissions of C. neoformans; international transmissions took place on 13% of VNIa-4 33 branches, and intercontinental transmissions on 7% of VNIa-93 branches. The median length of 34 within sub-clade internal branches was 3-6 SNPs, while terminal branches were 44.5-77.5 SNPs. The 35 short median internal branches were partly driven by the large number (12-15% of internal 36 branches) of polytomies in the within-sub-clade trees. To simultaneously explain our observation of 37 no apparent molecular clock, short internal branches and frequent polytomies we hypothesise that 38 C. neoformans VNIa spends much of its time in the environment in a quiescent state, while, when it 39 is sampled, it has almost always undergone an extended period of growth. Infections with VNIa-93 40 were associated with a significantly reduced risk of death by 10 weeks compared with infections 41 with VNIa-4 (Hazard Ratio = 0.45, p = 0.003). We detected a recombination in the mitochondrial 42 sequence of VNIa-5, suggesting that mitochondria could be involved in the propensity of this sub-43 clade to infect HIV-uninfected patients. These data highlight the insight into the biology and 44 epidemiology of pathogenic fungi which can be gained from WGS data. 45 46 3

Identification of pathogen genomic differences that impact human immune response and disease duringCryptococcus neoformansinfection

“…Strain background can 96 significantly influence human disease, and there is tremendous disparity in strain frequency; 97 some strain groups are much more common than others. ST93 is common in Uganda, but is also 98 5 the most frequently isolated ST strain from HIV-infected patients in Brazil (85% [25,26]) and 99 India (71% [27,28]). Sequence type prevalence also has a clear geographic component as 100 different ST groups are dominant in other well-sampled countries (e.g., China, Thailand, 101 Vietnam, Indonesia, Botswana, France [27-29]).…”

“…Given these limitations, we anticipate additional studies will uncover more genes from our study 382 with an impact on pathogenesis. It would also of course be of general interest to reconstruct a 383 knockout collection in a strain background more representative of typical clinical strains [14,28]. 384 We purposefully chose to focus our study on strains from ST93, which was the most 385 prevalent ST group from strains we sampled from participants in the COAT trial (~63% of all 386 strains).…”