Molecular Epidemiology Reveals Genetic Diversity amongst Isolates of the Cryptococcus neoformans/C. gattii Species Complex in Thailand

Abstract

To gain a more detailed picture of cryptococcosis in Thailand, a retrospective study of 498 C. neoformans and C. gattii isolates has been conducted. Among these, 386, 83 and 29 strains were from clinical, environmental and veterinary sources, respectively. A total of 485 C. neoformans and 13 C. gattii strains were studied. The majority of the strains (68.9%) were isolated from males (mean age of 37.97 years), 88.5% of C. neoformans and only 37.5% of C. gattii strains were from HIV patients. URA5-RFLP and/or M13 PCR-fingerprinting analysis revealed that the majority of the isolates were C. neoformans molecular type VNI regardless of their sources (94.8%; 94.6% of the clinical, 98.8% of the environmental and 86.2% of the veterinary isolates). In addition, the molecular types VNII (2.4%; 66.7% of the clinical and 33.3% of the veterinary isolates), VNIV (0.2%; 100% environmental isolate), VGI (0.2%; 100% clinical isolate) and VGII (2.4%; 100% clinical isolates) were found less frequently. Multilocus Sequence Type (MLST) analysis using the ISHAM consensus MLST scheme for the C. neoformans/C. gattii species complex identified a total of 20 sequence types (ST) in Thailand combining current and previous data. The Thai isolates are an integrated part of the global cryptococcal population genetic structure, with ST30 for C. gattii and ST82, ST83, ST137, ST141, ST172 and ST173 for C. neoformans being unique to Thailand. Most of the C. gattii isolates were ST7 = VGIIb, which is identical to the less virulent minor Vancouver island outbreak genotype, indicating Thailand as a stepping stone in the global spread of this outbreak strain. The current study revealed a greater genetic diversity and a wider range of major molecular types being present amongst Thai cryptococcal isolates than previously reported.

Advances in the understanding of the Cryptococcus neoformans and C. gattii species complexes and cryptococcosis

“…In addition, in both species, the same genotypes have been found to be recovered from patients and from the patients' environment 19,22 , which supports the theory of the acquisition of the infection from environmental sources and reaffirms the saprophytic and arboreal sources of these yeasts. Both species have also been isolated frequently from air, and swabs from trees, from decaying wood in hollows of many tree species, either individually or together 14,23 .…”

“…Unlike C. neoformans, which predominantly affects immunocompromised hosts, C. gattii most often affects patients with no apparent risk factors 1,[18][19][20][21] . In addition, in both species, the same genotypes have been found to be recovered from patients and from the patients' environment 19,22 , which supports the theory of the acquisition of the infection from environmental sources and reaffirms the saprophytic and arboreal sources of these yeasts.…”

“…Both species have also been isolated frequently from air, and swabs from trees, from decaying wood in hollows of many tree species, either individually or together 14,23 . Regarding the epidemiology of clinical cases of cryptococcosis, male patients are more commonly reported, as it is thought that male immune response may be less efficient in controlling cryptococcal infection 24 , whilst cases in children and older patients are still rare in most countries 18,19 . In domestic animals from North America, C. neoformans molecular type VNI has been found to affect mostly dogs whereas C. gattii …”

Cryptococcus gattii Infections

“…More recent studies support the proposal that C. gattii molecular type VGII originated in South America (46,72) and that Australia and Thailand are possible "stepping stones" in the global spread of VGIIb from South to North America (88,89). The introduction of VGIIb from its ancestral origin in South America, via different parts of the world, has recently been confirmed by using WGST (54).…”

Multilocus sequence typing analysis reveals that Cryptococcus neoformans var. neoformans is a recombinant population

“…In addition, data obtained from previous studies of 30 C. neoformans var. grubii isolates (11 VNB, 10 VNI, and 9 VNII) were included in the analysis as outgroup strains (Khayhan et al, 2013; Litvintseva et al, 2006; Sanchini et al, 2014; Cogliati et al, 2013; Meyer et al, 2009; Kaocharoen et al, 2013; Choi et al, 2010; Umeyama et al, 2013; Wiesner et al, 2012) (Table S1). …”