47 results found
Furukawa T, van Rhijn N, Chown H, et al., 2022, Exploring a novel genomic safe-haven site in the human pathogenic mould br Aspergillus fumigatus, FUNGAL GENETICS AND BIOLOGY, Vol: 161, ISSN: 1087-1845
Taori S, Rhodes J, Khonyongwa K, et al., 2022, First experience of implementing Candida auris Real Time PCR for surveillance in the UK: detection of multiple introductions with two international clades and improved patient outcomes, Journal of Hospital Infection, ISSN: 0195-6701
Background: Candida auris has been associated with rapid transmission and high mortality. A novelPCR based surveillance programme was initiated at a London teaching hospital from January 2018.The results of this implementation until March 2019 are presented along with the clinical,transmission and phylogenetic characteristics encountered in that setting.Methods: A real time-PCR assay for C auris was developed, validated, and implemented for directuse on skin swabs and urine. Environmental swabs were also tested by PCR as an emergencyoutbreak control measure. Clinical risk factors and outcomes of patients were determined.Environmental dispersal was assessed using 24 h settle plate cultures around 9 colonised patientsfollowed by air sampling around one colonised patient during high and low turbulence activities.Sequencing was performed using Illumina HiSeq and maximum likelihood phylogenies wereconstructed using rapid bootstrap analysis.Results 21 C. auris colonised patients were identified. Median turnaround time of colonisationdetection reduced from 141 h (5.8 d) to approximately 24 h enabling rapid infection controlprecautions. Settle plates detected 70 to 600 CFU/m2 around colonised patients over 24 h and airsampling suggested dispersal during turbulent activities. C. auris DNA was detected from 35.7%environmental swabs. Despite being in a high-risk setting, no patients developed invasive infection.Sequencing analysis of isolates from this centre identified two introductions of the South Asian(Clade I) and one of the South African (Clade III) strain.Conclusion: The PCR offers a rapid, scalable method of screening and supports clinical risk reductionin settings likely to encounter multiple introductions.
Rhodes J, 2022, Population genomics confirms acquisition of drug resistant Aspergillus fumigatus infection by humans from the environment, Nature Microbiology, Vol: 7, ISSN: 2058-5276
Infections caused by the fungal pathogen Aspergillus fumigatus are increasingly resistant to first-line azole antifungal drugs. However, despite its clinical importance, little is known about how susceptible patients acquire infection from drug resistant genotypes in the environment. Here, we present a population genomic analysis of 218 A. fumigatus from across the United Kingdom and Ireland (comprising 153 clinical isolates from 143 patients, and 65 environmental isolates). First, phylogenomic analysis shows strong genetic structuring into two clades (‘A’ and ‘B’) with little interclade recombination and the majority of environmental azole resistance found within Clade A. Secondly, we show occurrences where azole resistant isolates of near identical genotypes were obtained from both environmental and clinical sources, indicating with high confidence the infection of patients with resistant isolates transmitted from the environment. Third, genome-scans identified selective sweeps across multiple regions indicating a polygenic basis to the trait in some genetic backgrounds. These signatures of positive selection are seen for loci containing the canonical genes encoding fungicide resistance in the ergosterol biosynthetic pathway, whilst other regions under selection have no defined function. Lastly, pangenome analysis identified genes linked to azole resistance and novel resistance mechanisms. Understanding the environmental drivers and genetic basis of evolving fungal drug resistance needs urgent attention, especially in light of increasing numbers of patients with severe viral respiratory tract infections who are susceptible to opportunistic fungal superinfections.
Fisher MC, Alastruey-Izquierdo A, Berman J, et al., 2022, Tackling the emerging threat of antifungal resistance to human health, NATURE REVIEWS MICROBIOLOGY, ISSN: 1740-1526
Duong T-MN, Le T-V, Tran K-LH, et al., 2021, Azole-resistant Aspergillus fumigatus is highly prevalent in the environment of Vietnam, with marked variability by land use type, Environmental Microbiology, Vol: 23, Pages: 7632-7642, ISSN: 1462-2912
Azole-resistant environmental Aspergillus fumigatus presents a threat to public health but the extent of this threat in Southeast Asia is poorly described. We conducted environmental surveillance in the Mekong Delta region of Vietnam, collecting air and ground samples across key land-use types, and determined antifungal susceptibilities of Aspergillus section Fumigati (ASF) isolates and azole concentrations in soils. Of 119 ASF isolates, 55% were resistant (or non-wild type) to itraconazole, 65% to posaconazole and 50% to voriconazole. Azole resistance was more frequent in A. fumigatus sensu stricto isolates (95%) than other ASF species (32%). Resistant isolates and agricultural azole residues were overrepresented in samples from cultivated land. cyp51A gene sequence analysis showed 38/56 resistant A. fumigatus sensu stricto isolates carried known resistance mutations, with TR34/L98H most frequent (34/38).
Brackin A, Hemmings S, Fisher M, et al., 2021, Fungal genomics in respiratory medicine: what, how and when?, Mycopathologia, Vol: 186, Pages: 589-608, ISSN: 0301-486X
Respiratory infections caused by fungal pathogens present a growing global healthconcern and are a major cause of death in immunocompromised patients. Worryingly,coronavirus disease-19 (COVID-19) resulting in acute respiratory distress syndrome,has been shown to predispose some patients to fungal co-infection and secondarypulmonary aspergillosis. Aspergillosis is most commonly caused by the fungalpathogen Aspergillus fumigatus and primarily treated using the triazole drug group,however in recent years, this fungus has been rapidly gaining resistance against theseantifungals. This is of serious clinical concern as multi-azole resistant forms ofaspergillosis have a higher risk of mortality when compared against azole-susceptibleinfections. With the increasing numbers of COVID-19 and other classes ofimmunocompromised patients, early diagnosis of fungal infections is critical to ensuringpatient survival. However, time-limited diagnosis is difficult to achieve with currentculture-based methods. Advances within fungal genomics have enabled moleculardiagnostic methods to become a fast, reproducible, and cost-effective alternative fordiagnosis of respiratory fungal pathogens and detection of antifungal resistance. Herewe describe what techniques are currently available within molecular diagnostics, howthey work and when they have been used.
Edwards H, Rhodes J, 2021, Accounting for the biological complexity of pathogenic fungi in phylogenetic dating, Journal of Fungi, Vol: 7, ISSN: 2309-608X
In the study of pathogen evolution, temporal dating of phylogenies provides information on when species and lineages may have diverged in the past. When combined with spatial and epidemiological data in phylodynamic models, these dated phylogenies can also help infer where and when outbreaks occurred, how pathogens may have spread to new geographic locations and/or niches, and how virulence or drug resistance has developed over time. Although widely applied to viruses and, increasingly, to bacterial pathogen outbreaks, phylogenetic dating is yet to be widely used in the study of pathogenic fungi. Fungi are complex organisms with several biological processes that could present issues with appropriate inference of phylogenies, clock rates, and divergence times, including high levels of recombination and slower mutation rates although with potentially high levels of mutation rate variation. Here, we discuss some of the key methodological challenges in accurate phylogeny reconstruction for fungi in the context of the temporal analyses conducted to date and make recommendations for future dating studies to aid development of a best practices roadmap in light of the increasing threat of fungal outbreaks and antifungal drug resistance worldwide.
Soraggi S, Rhodes J, Altinkaya I, et al., 2021, HMMploidy: inference of ploidy levels from short-read sequencing data
<jats:title>Abstract</jats:title><jats:p>The inference of ploidy levels from genomic data is important to understand molecular mechanisms underpinning genome evolution. However, current methods based on allele frequency and sequencing depth variation do not have power to infer ploidy levels at low- and mid-depth sequencing data, as they do not account for data uncertainty. Here we introduce <jats:monospace>HMMploidy</jats:monospace>, a novel tool that leverages the information from multiple samples and combines the information from sequencing depth and genotype likelihoods. We demonstrate that <jats:monospace>HMMploidy</jats:monospace> outperforms existing methods in most tested scenarios, especially at low-depth with large sample size. We apply <jats:monospace>HMMploidy</jats:monospace> to sequencing data from the pathogenic fungus <jats:italic>Cryptococcus neoformans</jats:italic> and retrieve pervasive patterns of aneuploidy, even when artificially downsampling the sequencing data. We envisage that <jats:monospace>HMMploidy</jats:monospace> will have wide applicability to low-depth sequencing data from polyploid and aneuploid species.</jats:p>
Verweij PE, Lucas JA, Arendrup MC, et al., 2020, The one health problem of azole resistance in Aspergillus fumigatus: current insights and future research agenda, FUNGAL BIOLOGY REVIEWS, Vol: 34, Pages: 202-214, ISSN: 1749-4613
Ghosh P, Brookes L, Edwards H, et al., 2020, Cross-Disciplinary Genomics Approaches to Studying Emerging Fungal Infections, Life, Vol: 10, ISSN: 2075-1729
Emerging fungal pathogens pose a serious, global and growing threat to food supply systems, wild ecosystems, and human health. However, historic chronic underinvestment in their research has resulted in a limited understanding of their epidemiology relative to bacterial and viral pathogens. Therefore, the untargeted nature of genomics and, more widely, -omics approaches is particularly attractive in addressing the threats posed by and illuminating the biology of these pathogens. Typically, research into plant, human and wildlife mycoses have been largely separated, with limited dialogue between disciplines. However, many serious mycoses facing the world today have common traits irrespective of host species, such as plastic genomes; wide host ranges; large population sizes and an ability to persist outside the host. These commonalities mean that -omics approaches that have been productively applied in one sphere and may also provide important insights in others, where these approaches may have historically been underutilised. In this review, we consider the advances made with genomics approaches in the fields of plant pathology, human medicine and wildlife health and the progress made in linking genomes to other -omics datatypes and sets; we identify the current barriers to linking -omics approaches and how these are being underutilised in each field; and we consider how and which -omics methodologies it is most crucial to build capacity for in the near future.
Yu L-S, Rodriguez-Manzano J, Moser N, et al., 2020, Rapid detection of azole-resistant Aspergillus fumigatus in clinical and environmental isolates using lab-on-a-chip diagnostic system, Journal of Clinical Microbiology, Vol: 58, Pages: 1-11, ISSN: 0095-1137
Aspergillus fumigatus has widely evolved resistance to the most commonly used class of antifungal chemicals, the azoles. Current methods for identifying azole resistance are time-consuming and depend on specialized laboratories. There is an urgent need for rapid detection of these emerging pathogens at point-of-care to provide the appropriate treatment in the clinic and to improve management of environmental reservoirs to mitigate the spread of antifungal resistance. Our study demonstrates the rapid and portable detection of the two most relevant genetic markers linked to azole resistance, the mutations TR34 and TR46, found in the promoter region of the gene encoding the azole target, cyp51A. We developed a lab-on-a-chip platform consisting of: (1) tandem-repeat loop-mediated isothermal amplification, (2) state-of-the-art complementary metal-oxide-semiconductor microchip technology for nucleic-acid amplification detection and, (3) and a smartphone application for data acquisition, visualization and cloud connectivity. Specific and sensitive detection was validated with isolates from clinical and environmental samples from 6 countries across 5 continents, showing a lower limit-of-detection of 10 genomic copies per reaction in less than 30 minutes. When fully integrated with a sample preparation module, this diagnostic system will enable the detection of this ubiquitous fungus at the point-of-care, and could help to improve clinical decision making, infection control and epidemiological surveillance.
Armstrong-James D, Youngs J, Bicanic T, et al., 2020, Confronting and mitigating the risk of COVID-19 Associated Pulmonary Aspergillosis (CAPA), European Respiratory Journal, Vol: 56, Pages: 1-10, ISSN: 0903-1936
Cases of COVID-19 associated pulmonary aspergillosis (CAPA) are being increasingly reported and physicians treating patients with COVID-19-related lung disease need to actively consider these fungal co-infections.The SARS-CoV-2 (COVID-19) virus causes a wide spectrum of disease in healthy individuals as well as those with common comorbidities . Severe COVID-19 is characterised acute respiratory distress syndrome (ARDS) secondary to viral pneumonitis, treatment of which may require mechanical ventilation or extracorporeal membrane oxygenation (ECMO) . Clinicians are alert to the possibility of bacterial co-infection as a complication of lower respiratory tract viral infection; for example a recent review found that 72% of patients with COVID-19 received antimicrobial therapy . However, the risk of fungal co-infection, in particular COVID-19 associated pulmonary aspergillosis (CAPA), remains underappreciated.Fungal disease consistent with invasive aspergillosis (IA) has been observed with other severe Coronaviruses such as Severe Acute Respiratory Syndrome (SARS-CoV-2003) [4, 5] and Middle East Respiratory Syndrome (MERS-CoV) . From the outset of the COVID-19 pandemic, there were warning signs of secondary invasive fungal infection; Aspergillus flavus was isolated from the respiratory tract from one of 99 patients in the first COVID-19 cohort from Wuhan to be reported in any detail  and Aspergillus spp. were isolated from 2/52 (3.8%) of a subsequent cohort of critically unwell patients from this region . More recently, retrospective case series from Belgium , France , The Netherlands  and Germany  have reported evidence of CAPA in an alarming 20–35% of mechanically ventilated patients.
Rhodes J, Fisher MC, 2019, Global epidemiology of emerging Candida auris, Current Opinion in Microbiology, Vol: 52, Pages: 84-89, ISSN: 1369-5274
The discovery in 2009 of a new species of yeast, Candida auris, heralded the arrival of a novel emerging human infectious disease. This review highlights the unique characteristics of C. auris that have lled to it being of public health concern worldwide, namely public health concern, namely its global emergence, its ability to cause nosocomial outbreaks in healthcare settings, its innate and emerging resistance to multiple antifungal drugs and its resilience in the face of hygiene and infection control measures. Genomic epidemiology has identified four emergences of C. auris marked by four clades of the pathogen. These clades of C. auris are genetically dissimilar and are associated with differential resistance to antifungal drugs, suggesting that they will continue to phenotypically diverge into the future. The global emergence of C. auris testifies to the unmapped nature of Kingdom Fungi, and represents a new nosocomial threat that will require enhanced infection control across diverse healthcare and community settings.
Rhodes J, 2019, Rapid worldwide emergence of pathogenic fungi, Cell Host & Microbe, Vol: 26, Pages: 12-14, ISSN: 1931-3128
The recent coverage in the mainstream media of global Candida auris outbreaks has provided the general public with an awareness of the unprecedented emergence rate of pathogenic fungi. Scientific communities need to collaboratively address this threat to public health.
Sewell TR, Zhang Y, Brackin AP, et al., 2019, Elevated prevalence of azole resistant Aspergillus fumigatus in urban versus rural environments in the United Kingdom., Antimicrob Agents Chemother
Azole resistance in the opportunistic pathogen Aspergillus fumigatus is increasing, dominated primarily by two environmentally-associated resistance alleles: TR34/L98H and TR46/Y121F/T289A. By sampling soils across the South of England we assess the prevalence of azole resistant A. fumigatus (ARAf) in samples collected in both urban and rural locations. We characterise the susceptibility profiles of the resistant isolates to three medical azoles, identify the underlying genetic basis of resistance and investigate their genetic relationships. ARAf was detected in 6.7% of the soil samples, with a higher prevalence in urban (13.8%) compared to rural (1.1%) locations. Twenty isolates were confirmed to exhibit clinical breakpoints for resistance to at least one of three medical azoles, with 18 isolates exhibiting resistance to itraconazole, six to voriconazole and two showing elevated minimum inhibitory concentration to posaconazole. Thirteen of the resistant isolates harboured the TR34/L98H resistance allele and six isolates carried the TR46/Y121F/T289A allele. The 20 azole-resistant isolates were spread across five csp1 genetic subtypes, t01, t02, t04B, t09 and t18 with t02 the predominant subtype. Our study demonstrates that ARAf can be easily isolated in the South of England, especially in urban city centres, which appear to play an important role in the epidemiology of environmentally-linked drug resistant A. fumigatus.
Sewell T, Zhu J, Rhodes J, et al., 2019, Non-random distribution of azole resistance across the global population of Aspergillus fumigatus, mBio, Vol: 10, ISSN: 2150-7511
The emergence of azole resistance in the pathogenic fungus Aspergillus fumigatus has continued to increase, with the dominant resistance mechanisms, consisting of a 34-nucleotide tandem repeat (TR34)/L98H and TR46/Y121F/T289A, now showing a structured global distribution. Using hierarchical clustering and multivariate analysis of 4,049 A. fumigatus isolates collected worldwide and genotyped at nine microsatellite loci using analysis of short tandem repeats of A. fumigatus (STRAf), we show that A. fumigatus can be subdivided into two broad clades and that cyp51A alleles TR34/L98H and TR46/Y121F/T289A are unevenly distributed across these two populations. Diversity indices show that azole-resistant isolates are genetically depauperate compared to their wild-type counterparts, compatible with selective sweeps accompanying the selection of beneficial mutations. Strikingly, we found that azole-resistant clones with identical microsatellite profiles were globally distributed and sourced from both clinical and environmental locations, confirming that azole resistance is an international public health concern. Our work provides a framework for the analysis of A. fumigatus isolates based on their microsatellite profile, which we have incorporated into a freely available, user-friendly R Shiny application (AfumID) that provides clinicians and researchers with a method for the fast, automated characterization of A. fumigatus genetic relatedness. Our study highlights the effect that azole drug resistance is having on the genetic diversity of A. fumigatus and emphasizes its global importance upon this medically important pathogenic fungus.IMPORTANCE Azole drug resistance in the human-pathogenic fungus Aspergillus fumigatus continues to emerge, potentially leading to untreatable aspergillosis in immunosuppressed hosts. Two dominant, environmentally associated resistance mechanisms, which are thought to have evolved through selection by the agricultural application of azole fungic
Sewell TR, Zhang Y, Brackin AP, et al., 2019, Elevated prevalence of azole resistant <i>Aspergillus fumigatus</i> in urban versus rural environments in the United Kingdom, Publisher: Cold Spring Harbor Laboratory
<jats:title>Abstract</jats:title><jats:p>Azole resistance in the opportunistic pathogen <jats:italic>Aspergillus fumigatus</jats:italic> is increasing, dominated primarily by two environmentally-associated resistance alleles: TR<jats:sub>34</jats:sub>/L98H and TR<jats:sub>46</jats:sub>/Y121F/T289A. Using an environmental sampling strategy across the South of England we assess the prevalence of azole resistant <jats:italic>A. fumigatus</jats:italic> (AR<jats:italic>Af</jats:italic>) in soil samples collected in both urban and rural locations. We characterise the susceptibility profiles of the resistant isolates to three medical azoles, identify the underlying genetic basis of resistance and investigate their genetic relationships. AR<jats:italic>Af</jats:italic> was detected in 6.7% of the soil samples, with a higher prevalence in urban (13.8%) compared to rural (1.1%) locations. Nineteen isolates were confirmed to exhibit clinical breakpoints for resistance to at least one of three medical azoles, with 18 isolates exhibiting resistance to itraconazole, four to voriconazole, with two also showing additional elevated minimum inhibitory concentration to posaconazole. Thirteen of the resistant isolates harboured the TR<jats:sub>34</jats:sub>/L98H resistance allele and six isolates carried TR<jats:sub>46</jats:sub>/Y121F/T289A allele. The 19 azole-resistant isolates were spread across five <jats:italic>csp1</jats:italic> genetic subtypes, t01, t02, t04B, t09 and t18 with t02 the predominant subtype. Our study demonstrates that AR<jats:italic>Af</jats:italic> can be easily isolated in the South of England, especially in urban city centres, which appear to play an important role in the epidemiology of environmentally-linked drug resistant <jats:italic>A. fumigatus</jats:italic>.</jats:p>
Stone NRH, Rhodes J, Fisher MC, et al., 2019, Dynamic ploidy changes drive fluconazole resistance in human cryptococcal meningitis, Journal of Clinical Investigation, Vol: 129, Pages: 999-1014, ISSN: 0021-9738
Background: Cryptococcal meningitis (CM) causes an estimated 180,000 deaths annually, predominantly in sub-Saharan Africa, where most patients receive fluconazole (FLC) monotherapy. While relapse after FLC monotherapy with resistant strains is frequently observed, the mechanisms and impact of emergence of FLC resistance in human CM are poorly understood. Heteroresistance (HetR) — a resistant subpopulation within a susceptible strain — is a recently described phenomenon in Cryptococcus neoformans (Cn) and Cryptococcus gattii (Cg), the significance of which has not previously been studied in humans.Methods: A cohort of 20 patients with HIV-associated CM in Tanzania was prospectively observed during therapy with either FLC monotherapy or in combination with flucytosine (5FC). Total and resistant subpopulations of Cryptococcus spp. were quantified directly from patient cerebrospinal fluid (CSF). Stored isolates underwent whole genome sequencing and phenotypic characterization.Results: Heteroresistance was detectable in Cryptococcus spp. in the CSF of all patients at baseline (i.e., prior to initiation of therapy). During FLC monotherapy, the proportion of resistant colonies in the CSF increased during the first 2 weeks of treatment. In contrast, no resistant subpopulation was detectable in CSF by day 14 in those receiving a combination of FLC and 5FC. Genomic analysis revealed high rates of aneuploidy in heteroresistant colonies as well as in relapse isolates, with chromosome 1 (Chr1) disomy predominating. This is apparently due to the presence on Chr1 of ERG11, which is the FLC drug target, and AFR1, which encodes a drug efflux pump. In vitro efflux levels positively correlated with the level of heteroresistance.Conclusion: Our findings demonstrate for what we believe is the first time the presence and emergence of aneuploidy-driven FLC heteroresistance in human CM, association of efflux levels with heteroresistance, and the successful suppression of hetero
Abdolrasouli A, Scourfield A, Rhodes J, et al., 2018, High prevalence of triazole resistance in clinical Aspergillus fumigatus isolates in a specialist cardiothoracic centre, International Journal of Antimicrobial Agents, Vol: 52, Pages: 637-642, ISSN: 0924-8579
OBJECTIVES: To evaluate the prevalence of triazole-resistant Aspergillus fumigatus and common molecular cyp51A polymorphisms amongst clinical isolates in a specialised cardiothoracic centre in London, UK. METHODS: All A. fumigatus isolates were prospectively analysed from April 2014 to March 2016. Isolates were screened with a four-well VIPcheck™ plate to assess triazole susceptibility. Resistance was confirmed with a standard microbroth dilution method according to European Committee on Antimicrobial Susceptibility Testing (EUCAST) guidelines. Triazole-resistant A. fumigatus isolates were subjected to a mixed-format real time polymerase chain reaction (RT-PCR) assay (AsperGenius®) to detect common cyp51A alterations. RESULTS: We identified 167 clinical A. fumigatus isolates from 135 patients. Resistance to at least one azole antifungal drug was confirmed in 22/167 (13.2%) of isolates from 18/135 (13.3%) patients, including 12/74 (16.2%) patients with cystic fibrosis (CF). The highest detection rate of azole-resistant A. fumigatus was among the 11- to 20-y age group. All triazole-resistant isolates (n = 22) were resistant to itraconazole, 18 showed cross-resistance to posaconazole and 10 displayed reduced susceptibility to voriconazole. No pan-azole-resistant A. fumigatus was identified. TR34/L98H was identified in 6/22 (27.3%) of azole-resistant isolates and detectable in 5/12 (42%) patients with CF. CONCLUSIONS: In our specialist cardiothoracic centre, the prevalence of triazole-resistant A. fumigatus is alarmingly high (13.2%). The majority of azole-resistant isolates were from patients with CF. We found a higher prevalence of the environmentally driven mutation TR34/L98H in our A. fumigatus isolates than in published UK data from other specialist respiratory centres, which may reflect differing patient populations managed at these institutions.
Farrer R, Ford C, Rhodes J, et al., 2018, Transcriptional heterogeneity of Cryptococcus gattii VGII compared with non-VGII lineages underpins key pathogenicity pathways, mSphere, Vol: 3, ISSN: 2379-5042
Cryptococcus gattii is a pathogenic yeast of humans and other animals, which causes disease predominantly in immunocompetent hosts. Infection begins when aerosolized yeast or spores enter the body, triggering an immune response, including engulfment by macrophages. To understand the early transcriptional signals in both the yeast and its mammalian host, we performed a time-course dual RNA-seq experiment for four lineages of C. gattii (VGI-IV) interacting with mouse macrophages at 1hr, 3hr and 6hr post infection. Comparison of in vitro to ex vivo gene expression indicates lineage VGII is transcriptionally divergent to non-VGII lineages, including differential expression of genes involved in capsule synthesis, capsule attachment and ergosterol production. Various paralogs demonstrate sub-functionalisation between lineages including an upregulation of capsule biosynthesis-related gene CAP2, and downregulation of CAP1 in VGIII. Isolates also compensate for lineage-specific gene-losses by over-expression of genetically similar paralogs, including an over-expression of capsule gene CAS3 in VGIV having lost CAS31. Differential expression of one in five C. gattii genes was detected following co-incubation with mouse macrophages; all isolates showed high induction of oxidative-reduction functions and a downregulation of capsule attachment genes. We also show that VGII switches expression of two laccase paralogs (from LAC1 to LAC2) during co-incubation of macrophages. Finally, we found that mouse macrophages respond to all four lineages of C. gattii by upregulating FosB/Jun/Egr1 regulatory proteins at early time points. This study highlights the evolutionary breadth of expression profiles amongst the lineages of C. gattii and the diversity of transcriptional responses at this host-pathogen interface.
Farrer RA, Ford CB, Rhodes J, et al., 2018, Transcriptional Heterogeneity of Cryptococcus gattii VGII Compared with Non-VGII Lineages Underpins Key Pathogenicity Pathways., mSphere, Vol: 3
Cryptococcus gattii is a pathogenic yeast of humans and other animals which causes disease predominantly in immunocompetent hosts. Infection begins when aerosolized yeast or spores enter the body, triggering an immune response, including engulfment by macrophages. To understand the early transcriptional signals in both the yeast and its mammalian host, we performed a time-course dual-transcriptome sequencing (RNA-seq) experiment for four lineages of C. gattii (lineages VGI to IV) interacting with mouse macrophages at 1, 3, and 6 h postinfection. Comparisons of in vitro to ex vivo gene expression levels indicated that lineage VGII is transcriptionally divergent from non-VGII lineages, including differential expression of genes involved in capsule synthesis, capsule attachment, and ergosterol production. Several paralogous genes demonstrated subfunctionalization between lineages, including upregulation of capsule biosynthesis-related gene CAP2 and downregulation of CAP1 in VGIII. Isolates also compensate for lineage-specific gene losses by overexpression of genetically similar paralogs, including overexpression of capsule gene CAS3 in VGIV, which have lost the CAS31 gene. Differential expression of one in five C. gattii genes was detected following coincubation with mouse macrophages; all isolates showed high induction of oxidative-reduction functions and downregulation of capsule attachment genes. We also found that VGII switches expression of two laccase paralogs (from LAC1 to LAC2) during coincubation of macrophages. Finally, we found that mouse macrophages respond to all four lineages of C. gattii by upregulating FosB/Jun/Egr1 regulatory proteins at early time points. This report highlights the evolutionary breadth of expression profiles among the lineages of C. gattii and the diversity of transcriptional responses at this host-pathogen interface.IMPORTANCE The transcriptional profiles of related pathogens and their responses to host-induced stresses underp
Abdolrasouli A, Petrou MA, Park H, et al., 2018, Surveillance for azole-resistant Aspergillus fumigatus in a centralized diagnostic mycology service, London, United Kingdom, 1998-2017, Frontiers in Microbiology, Vol: 9, ISSN: 1664-302X
Background/Objectives: Aspergillus fumigatus is the leading cause of invasive aspergillosis. Treatment is hindered by the emergence of resistance to triazole antimycotic agents. Here, we present the prevalence of triazole resistance among clinical isolates at a major centralized medical mycology laboratory in London, United Kingdom, in the period 1998–2017.Methods: A large number (n = 1469) of clinical A. fumigatus isolates from unselected clinical specimens were identified and their susceptibility against three triazoles, amphotericin B and three echinocandin agents was carried out. All isolates were identified phenotypically and antifungal susceptibility testing was carried out by using a standard broth microdilution method.Results: Retrospective surveillance (1998–2011) shows 5/1151 (0.43%) isolates were resistant to at least one of the clinically used triazole antifungal agents. Prospective surveillance (2015–2017) shows 7/356 (2.2%) isolates were resistant to at least one triazole antifungals demonstrating an increase in incidence of triazole-resistant A. fumigatus in our laboratory. Among five isolates collected from 2015 to 2017 and available for molecular testing, three harbored TR34/L98H alteration in the cyp51A gene that are associated with the acquisition of resistance in the non-patient environment.Conclusion: These data show that historically low prevalence of azole resistance may be increasing, warranting further surveillance of susceptible patients.
Summary:The increase of antifungal drug resistance is a major global human health concern andthreatens agriculture and food security; in order to tackle these concerns, it is important to understandthe mechanisms that cause antifungal resistance. The curated Mycology Antifungal Resistance Database(MARDy) is a web-service of antifungal drug resistance mechanisms, including amino acid substitutions,tandem repeat sequences and genome ploidy. MARDy is implemented on a Linux, Apache, MySQL andPHP web development platform and includes a local installation of BLASTn of the database of curatedgenes.Availability and implementation:MARDy can be accessed at http://www.mardy.net and is free touse. The complete database can be retrieved, ordered by organism, gene and drug. Missing or newmycological antifungal resistance data can be relayed to the development team through a contribute entryform.
Abdolrasouli A, Bercusson AC, Rhodes JL, et al., 2018, Airway persistence by the emerging multi-azole-resistant Rasamsonia argillacea complex in cystic fibrosis, Mycoses, Vol: 61, Pages: 665-673, ISSN: 0933-7407
Infections caused by Rasamsonia argillacea complex have been reported in various clinical settings. Cystic fibrosis (CF) is one of the main underlying conditions. An observational cohort study of CF patients with Rasamsonia in respiratory samples was conducted. Eight isolates from six patients were identified as R. argillacea complex and tested for antifungal susceptibility. All isolates had high MICs to voriconazole and posaconazole and low MECs to echinocandins. Four patients experienced lung function decline in the year preceding first Rasamsonia isolation. This continued in the year following first isolation in three out of four cases. Antifungal therapy was initiated in two patients, to which only one exhibited a clinical response. Three out of six patients died within three years of isolating Rasamsonia. Genotyping suggests that similar genotypes of Rasamsonia can persist in CF airways. Consistent with other fungi in CF, the clinical impact of airway colonization by Rasamsonia is variable. In certain patients, Rasamsonia may be able to drive clinical decline. In others, though a clear impact on lung function may be difficult to determine, the appearance of Rasamsonia acts as a marker of disease severity. In others it does not appear to have an obvious clinical impact on disease progression.
Sewell TR, Rhodes JL, Hagen F, et al., 2018, AfumID: An R Shiny application for Aspergillus fumigatus genotyping, Publisher: OXFORD UNIV PRESS, Pages: S65-S65, ISSN: 1369-3786
Abdolrasouli A, Rhodes JL, Sewell TR, et al., 2018, Occurrence of triazole resistance in Aspergillus fumigatus among respiratory patients in a specialized cardio-thoracic centre, London, United Kingdom, Publisher: OXFORD UNIV PRESS, Pages: S76-S76, ISSN: 1369-3786
Rhodes J, Abdolrasouli A, Farrer RA, et al., 2018, Genomic epidemiology of the UK outbreak of the emerging human fungal pathogen Candida auris (vol 7, pg 43, 2018), EMERGING MICROBES & INFECTIONS, Vol: 7, ISSN: 2222-1751
Rhodes J, Fisher MC, 2018, Breaching pathogeographic barriers by the bat white-nose fungus, mBio, Vol: 9, ISSN: 2150-7511
Bat white-nose syndrome has become associated with unparalleled mortality in bat species across the United States since 2006. In a recent article, Drees and colleagues (mBio 8:e01941-17, 2017, https://doi.org/10.1128/mBio.01941-17) utilized both whole-genome sequencing and microsatellite data to explore the origin and spread of the causative agent of bat white-nose syndrome, Pseudogymnoascus destructans The research by Drees et al. supports the hypothesis that P. destructans was introduced into North America from Europe, with molecular dating suggesting a divergence from European isolates approximately 100 years ago. The approaches described in this study are an important contribution toward pinpointing the origins of this infection and underscore the need for more rigorous international biosecurity in order to stem the tide of emerging fungal pathogens.
Nash A, Rhodes JL, 2018, Simulations of CYP51A from Aspergillus fumigatus in a model bilayer provide insights into triazole drug resistance, Medical Mycology, Vol: 56, Pages: 361-373, ISSN: 1460-2709
Azole antifungal drugs target CYP51A in Aspergillus fumigatus by binding with the active site of the protein, blocking ergosterol biosynthesis. Resistance to azole anti-fungal drugs is now common, with a leucine to histidine amino acid substitution at position 98 the most frequent, predominantly conferring resistance to itraconazole, although cross-resistance has been reported in conjunction with other mutations. In this study, we create a homology model of CYP51A using a recently published crystal structure of the paralog protein CYP51B. The derived structures, wild type and L98H mutant, are positioned within a lipid membrane bilayer and subjected to molecular dynamics simulations in order improve the accuracy of both models. The structural analysis from our simulations suggests a decrease in active site surface from the formation of hydrogen bonds between the histidine substitution and neighbouring polar side chains, potentially preventing the binding of azole drugs. This study yields a biologically relevant structure and set of dynamics of the A. fumigatus Lanosterol 14 alpha-demethylase enzyme and provides further insight into azole antifungal drug resistance.
Rhodes JL, Abdolrasouli A, Farrer R, et al., 2018, Genomic epidemiology of the UK outbreak of the emerging human fungal pathogen Candida auris, Emerging Microbes and Infections, Vol: 7, ISSN: 2222-1751
Candida auris was first described in 2009, and has since caused nosocomial outbreaks, invasive infections and fungaemia across at least 19 countries in five continents. An outbreak of C. auris occurred in a specialised cardiothoracic London hospital between April 2015 and November 2016, which to date has been the largest outbreak within the UK, involving a total of 72 patients. To understand the genetic epidemiology of C. auris infection, both within this hospital and within a global context, we sequenced the outbreak isolate genomes using Oxford Nanopore Technologies and Illumina to detect antifungal resistance alleles and to reannotate the C. auris genome. Phylogenomic analysis placed the UK outbreak in the India/Pakistan clade, demonstrating an Asian origin: the outbreak showed similar genetic diversity to that of the entire clade and limited local spatiotemporal clustering was observed. One isolate displayed resistance to both echinocandins and 5-flucytosine; the former was associated with a serine to tyrosine amino acid substitution in the gene FKS1, and the latter was associated with a phenylalanine to isoleucine substitution in the gene FUR1. These mutations add to a growing body of research on multiple antifungal drug targets in this organism. Multiple differential episodic selection of antifungal resistant genotypes has occurred within a genetically heterogenous population across this outbreak, creating a resilient pathogen and making it difficult to define local-scale patterns of transmission as well as implementing outbreak control measures.
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