Biology subjects

Ashton, P. M.

Publications and source records attributed to Ashton, P. M..

4 recordsLinked to original sources

bioRxiv2018

Genomics of Cryptococcus neoformans

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

microbiology↗

bioRxiv2017

MULTILOCUS SEQUENCE TYPING REVEALS A UNIQUE CO-DOMINANT POPULATION STRUCTURE OF CRYPTOCOCCUS NEOFORMANS VAR. GRUBII IN VIETNAM

Cryptococcosis is amongst the most important invasive fungal infections globally, with cryptococcal meningitis causing an estimated 180,000 deaths each year in HIV infected patients alone. Patients with other forms of immunosuppression are also at risk, and disease is increasingly recognized in apparently immunocompetent individuals. Cryptococcus neoformans var. grubii (serotype A, molecular type VNI) has a global distribution and is responsible for the majority of cases. Here, we used the consensus ISHAM Multilocus Sequence Typing (MLST) for C. neoformans to define the population structure of clinical isolates of Cryptococcus neoformans var. grubii from Vietnam (n=136) and Laos (n=81). We placed these isolates into the global context using published MLST data from 8 other countries (total N = 669). We observed a phylo-geographical relationship in which Laos was similar to its Southeast Asian neighbor Thailand in being dominated (83%) by Sequence Type (ST) 4 and its Single Locus Variant ST6. On the other hand, Vietnam was uniquely intermediate between Southeast Asia and East Asia having both ST4/ST6 (35%) and ST5 (48%) which causes the majority of cases in East Asia. Analysis of genetic distance (Fst) between different populations of Cryptococcus neoformans var. grubii supported the intermediate nature of the population from Vietnam. A strong association between ST5 and infection in apparently immunocompetent, HIV-uninfected patients was observed in Vietnam (OR: 7.97, [95%CI: 3.18-19.97], p < 0.0001). Our study emphasizes that Vietnam, with its intermediate Cryptococcus neoformans var. grubii population structure, provides the strongest epidemiological evidence of the relationship between ST5 and infection of HIV-uninfected patients. Human population genetic distances within the region suggest these differences in CNVG population across Southeast Asia are driven by ecological factors rather than host factors.\n\nAuthor summaryCryptococcus neoformans is a yeast that causes meningitis in people, usually with damaged immune systems. There are >180,000 deaths in HIV-infected patients each year, most occurring where there are the highest HIV/AIDS disease burdens. Vietnam and Laos have contributed significantly to clinical trials aiming to improve the treatment of cryptococcal meningitis, but the relationship of isolates from these countries to the global population is not yet described. Here, we address this knowledge gap by using Multilocus Sequence Typing to study the population of Cryptococcus neoformans var. grubii (CNVG) in Laos and Vietnam, with the specific aim of incorporating these populations into the wider global context. We found that, in most countries, a single lineage (family) of strains was responsible for most disease. The Vietnamese CNVG population was unusual in that 2 main lineages circulated at the same time. The Vietnamese CNVG population occupies a middle ground between Thailand/Laos in the west and China in the east. The differences in population structure moving from West to East are probably due to ecological differences. Disease in HIV uninfected patients was almost always due to members of a single family of strains (ST5).

molecular biology↗

bioRxiv2017

N VITRO AND IN VIVO CHARACTERISATION OF ISOLATES OF CRYPTOCOCCUS NEOFORMANS CAUSING MENINGITIS IN HIV-INFECTED AND UNINFECTED PATIENTS IN VIETNAM

We previously observed a substantial burden of cryptococcal meningitis in Vietnam atypically arising in HIV-uninfected individuals. This disease was associated with a single genotype of Cryptococcus neoformans (Sequence Type (ST)5), which was significantly less common in HIV-infected individuals. Aiming to compare the phenotypic characteristics of ST5 and non-ST5 C. neoformans we selected 30 representative Vietnamese isolates, compared their in vitro pathogenic potential and in vivo virulence. ST5 and non-ST5 organisms exhibited comparable characteristics with respect to in vitro virulence markers including melanin production, replication at 37{degrees}C, and growth in cerebrospinal fluid. However, the ST5 isolates had significantly increased variability in cellular and capsular sizing compared with non-ST5 organisms (p<0.001). Counter-intuitively, mice infected with ST5 isolates had significantly longer survival with lower fungal burdens at day 7 than non-ST5 isolates. Notably, ST5 isolates induced significantly greater initial inflammatory responses than non-ST5 strains, measured by TNF- concentrations (p<0.001). Despite being generally less virulent in the mouse model, we hypothesize that the significant within strain variation seen in ST5 isolates in the tested phenotypes may represent an evolutionary advantage enabling adaptation to novel niches including apparently immunocompetent human hosts.

microbiology↗

bioRxiv2017

Salmonella enterica Serovar Typhimurium ST313 Responsible For Gastroenteritis In The UK Are Genetically Distinct From Isolates Causing Bloodstream Infections In Africa

The ST313 sequence type of Salmonella enterica serovar Typhimurium causes invasive non-typhoidal salmonellosis amongst immunocompromised people in sub-Saharan Africa (sSA). Previously, two distinct phylogenetic lineages of ST313 have been described which have rarely been found outside sSA. Following the introduction of routine whole genome sequencing of Salmonella enterica by Public Health England in 2014, we have discovered that 2.7% (79/2888) of S. Typhimurium from patients in England and Wales are ST313. Of these isolates, 59/72 originated from stool and 13/72 were from extra-intestinal sites. The isolation of ST313 from extra-intestinal sites was significantly associated with travel to Africa (OR 12 [95% CI: 3,53]). Phylogenetic analysis revealed previously unsampled diversity of ST313, and distinguished UK-linked isolates causing gastroenteritis from African-associated isolates causing invasive disease. Bayesian evolutionary investigation suggested that the two African lineages diverged from their most recent common ancestors independently, circa 1796 and 1903. The majority of genome degradation of African ST313 lineage 2 is conserved in the UK ST313 lineages and only 10/44 pseudogenes were lineage 2-specific. The African lineages carried a characteristic prophage and antibiotic resistance gene repertoire, suggesting a strong selection pressure for these horizontally-acquired genetic elements in the sSA setting. We identified an ST313 isolate associated with travel to Kenya that carried a chromosomally-located blaCTX-M-15, demonstrating the continual evolution of this sequence type in Africa in response to selection pressure exerted by antibiotic usage.\n\nThe S. Typhimurium ST313 sequence type has been primarily associated with invasive disease in Africa. Here, we highlight the power of routine whole-genome-sequencing by public health agencies to make epidemiologically-significant deductions that would be missed by conventional microbiological methods. The discovery of ST313 isolates responsible for gastroenteritis in the UK reveals new diversity in this important sequence type. We speculate that the niche specialization of sub-Saharan African ST313 lineages is driven in part by the acquisition of accessory genome elements.

microbiology↗