Optimization and Validation of Candida auris Short Tandem Repeat Analysis

Groot, Theun de; Spruijtenburg, Bram; Parnell, Lindsay; Chow, Nancy A.; Meis, Jacques F.

doi:10.1128/spectrum.02645-22

Cited by 9 publications

(8 citation statements)

References 29 publications

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“…The number of SNP differences within each clade is in appendix 1 (p17). In silico STR analysis was performed based on a previously published scheme, and the topology of the dendrogram also demonstrated clustering of Clade VI isolates based on STR profiles, congruent with the observation of a sixth clade using WGS 22,23 (appendix 1 p18). Additional Bayesian phylogenetic analysis suggests that the estimated divergence time of Clade VI from the most closely related Clade IV was at 4831 BCE (HPD 272•4 -311•6 years ago).…”

Section: Resultssupporting

confidence: 54%

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Discovery of the sixthCandida aurisclade in Singapore

Suphavilai,

Ko,

Lim

et al. 2023

Preprint

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SummaryBackgroundThe emerging fungal pathogenCandida aurisposes a serious threat to global public health due to its worldwide distribution, multidrug-resistance, high transmissibility, propensity to cause outbreaks and high mortality rates. We report threeC. aurisisolates detected in Singapore, which are genetically distinct from all known clades (I-V) and represent a new clade (Clade VI).MethodsThree epidemiologically unlinked clinical isolates belonging to the potential newC. aurisclade were whole-genome sequenced and phenotypically characterized. The complete genomes of these isolates were compared to representative genomes of all known clades. To provide a global context, 3,651 international whole-genome sequences (WGS) from the NCBI database were included in the high-resolution single nucleotide polymorphism (SNP) analysis. Antifungal resistance genes, mating type locus, and chromosomal rearrangements were characterized from the WGS data of the Clade VI isolates. We further implemented Bayesian logistic regression models to simulate the automatic detection of Clade V and VI as their WGS data became available.FindingsThe three Clade VI isolates were separated by >36,000 SNPs from all existingC. aurisclades. These isolates had opposite mating type allele and different chromosomal rearrangements when compared to their closest Clade IV relatives. As a proof-of-concept, our classification model was able to flag these outlier genomes as a potential new clade. Furthermore, an independent WGS submission from Bangladesh was found to belong to this new clade.InterpretationThe discovery of a newC. aurisclade in Singapore and Bangladesh, showing close relationship to Clade IV members in South America, highlights the unknown genetic diversity and origin ofC. auris, particularly in under-resourced regions. Active surveillance in clinical settings, along with effective sequencing strategies and downstream analysis, will be essential in the identification of novel strains, tracking of transmission, and containment of adverse clinical impacts caused byC. aurisinfections.FundingThis work was supported by the Singapore National Medical Research Council (NMRC) research training fellowship (MOH-FLWSHP19may-0005), the NCRS Duke-NUS Academic Medical Center Academic Clinical Program grant (09/FY2022/P1/17-A32, GRDUKP003401), and the Genedant-GIS Innovation Program grant.Research in contextEvidence before this studyWe searched PubMed using the search terms “Candida auris” AND “clade”, for papers published between Jan 1, 2009, and July 1, 2023. This search retrieved 115 publications. 60 relevant publications were identified. 28 studies analyzed and discussed the molecular epidemiology ofC. auris, including the description ofC. aurisclades, either in outbreak or surveillance settings. There were 11 case reports ofC. aurisclinical cases that included clade determination. Two studies focused on the detection and clade determination ofC. aurisfrom non-healthcare environments. Clade-specific characteristics were described or analyzed in 14 studies. One study applied machine learning toC. aurisdrug resistance analysis, but not for clade determination. Four studies focused on the description of potentially newC. aurislineages, subclades, or clades. All publications described isolates that belong to one of the five knownC. aurisclades (I-V). All publications found that strains from different clades differed by more than 35,000 SNPs, and that there are clade-specific differences in geographical distribution, phenotypic characteristics, antifungal susceptibility profile, outbreak potential, and clinical manifestations.The NCBI Pathogen Detection system contained 4,506C. aurisgenomes on July 1, 2023. There were ten (0·22%) submissions from Southeast Asian countries and 92 (2·04%) submissions from South Asia and the Indian subcontinent, which are parts of the Indomalayan biogeographic realm.Added value of this studyTo the best of our knowledge, we are the first group to perform hybrid assemblies on three representative isolates in a newC. aurisclade, which is separated from all other existing clades (I-V) by >36,000 SNPs. Whole-genome SNP analysis and phenotypic characterization of these epidemiologically unlinked isolates detected in Singapore suggest that they represent a previously unreported sixth major clade. High-resolution SNP analysis of 3,651 international whole-genome sequences from the NCBI database, which generated the final dataset consisting of more than 6.6 million genome pairs, revealed six distinct genetic clusters representing the five known clades and the new sixth clade (Indomalayan). In addition, we demonstrate that a machine learning approach can be used to flag these outlier genomes for further investigations as soon as they become available, thus providing the earliest possible alert for potential new public health threats.Implications of all the available evidenceDespite the high antimicrobial resistance burden in Southeast Asia and South Asia, these regions are disproportionately underrepresented in terms of genomic surveillance ofC. auris, a multidrug-resistant fungal pathogen. The detection of three epidemiologically unlinkedC. aurisisolates in Singapore belonging to a newC. aurisclade suggests that yet-to-be-reported strains may be circulating in the region. Given the propensity for multidrug resistance, healthcare-associated infection outbreaks, and the associated high mortality, active surveillance and continued vigilance is necessary.

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Section: Resultssupporting

confidence: 54%

“…Antifungal susceptibility testing results are in Table 1. Further results and discussions on antifungal resistance testing are in appendix 1 (pp [22][23][24]. The index isolate from Patient A, isolate A (LIMS ID F3485, Biosample SAMN36753178) was cultured from a combined screening swab (nasal, axillae, and groin) in April 2023.…”

Section: Resultsmentioning

confidence: 99%

Discovery of the sixthCandida aurisclade in Singapore

Suphavilai,

Ko,

Lim

et al. 2023

Preprint

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“…While our results show a clear separation of the major geographic entities Asia, Africa and South America, our PCA analysis and subsequent affinity clustering indicate that particularly the South Asian (I), East Asian (II) and Iranian (V) clades are proteomically closely related, a result contrasted by genomic data [ 3 ] where the African clade (III) with strains from South Africa affiliate to strains from a Spanish outbreak closer to the East Asian clade (II). Similar to the genomic typing schemes [ 37 , 40 ], long branches in the affinity clustering indicate strains that appear as outliers or typical strains relative to those strains centralizing with similar or even distances around a cluster, hence forming a representative central entity. These specific isolates require further attention, regardless of the identification technology if they represent true biological deviants or technical artifacts.…”

Section: Discussionmentioning

confidence: 99%

Fast and Accurate Identification of Candida auris by High Resolution Mass Spectrometry

Jamalian

Freeke

Chowdhary

et al. 2023

JoF

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The emerging pathogen Candida auris has been associated with nosocomial outbreaks on six continents. Genetic analysis indicates simultaneous and independent emergence of separate clades of the species in different geographical locations. Both invasive infection and colonization have been observed, warranting attention due to variable antifungal resistance profiles and hospital transmission. MALDI-TOF based identification methods have become routine in hospitals and research institutes. However, identification of the newly emerging lineages of C. auris yet remains a diagnostic challenge. In this study an innovative liquid chromatography (LC)—high resolution OrbitrapTM mass spectrometry method was used for identification of C. auris from axenic microbial cultures. A set of 102 strains from all five clades and different body locations were investigated. The results revealed correct identification of all C. auris strains within the sample cohort, with an identification accuracy of 99.6% from plate culture, in a time-efficient manner. Furthermore, application of the applied mass spectrometry technology provided the species identification down to clade level, thus potentially providing the possibility for epidemiological surveillance to track pathogen spread. Identification beyond species level is required specially to differentiate between nosocomial transmission and repeated introduction to a hospital.

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Section: Discussionmentioning

confidence: 99%

“…SNP differences between isolates from patients P51, P52, P53, P57 and P65 were extremely low, indicating clonality and a common source that caused a nosocomial outbreak within this hospital, as is commonly observed for C. auris. 33,34 Extensive epidemiological information might uncover the chain of transmission and source, but available retrospective epidemiological data were insufficient. However, based on SNP differences, it appears that both hospitals acquired an independent seeding of C. auris from a different source despite being located a mere 500 m apart from each other.…”

Section: Reduced Echinocandin Susceptibility Is Therapy-inducedmentioning

confidence: 99%

Whole genome sequencing analysis demonstrates therapy‐induced echinocandin resistance in Candida auris isolates

Spruijtenburg,

Ahmad,

Asadzadeh

et al. 2023

Mycoses

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Candida auris is an emerging, multidrug‐resistant yeast, causing outbreaks in healthcare facilities. Echinocandins are the antifungal drugs of choice to treat candidiasis, as they cause few side effects and resistance is rarely found. Previously, immunocompromised patients from Kuwait with C. auris colonisation or infection were treated with echinocandins, and within days to months, resistance was reported in urine isolates. To determine whether the development of echinocandin resistance was due to independent introductions of resistant strains or resulted from intra‐patient resistance development, whole genome sequencing (WGS) single‐nucleotide polymorphism (SNP) analysis was performed on susceptible (n = 26) and echinocandin‐resistant (n = 6) isolates from seven patients. WGS SNP analysis identified three distinct clusters differing 17–127 SNPs from two patients, and the remaining isolates from five patients, respectively. Sequential isolates within patients had a maximum of 11 SNP differences over a time period of 1–10 months. The majority of isolates with reduced susceptibility displayed unique FKS1 substitutions including a novel FKS1M690V substitution, and nearly all were genetically related, ranging from only three to six SNP differences compared to susceptible isolates from the same patient. Resistant isolates from three patients shared the common FKS1S639F substitution; however, WGS analysis did not suggest a common source. These findings strongly indicate that echinocandin resistance is induced during antifungal treatment. Future studies should determine whether such echinocandin‐resistant strains are capable of long‐term colonisation, cause subsequent breakthrough candidiasis, have a propensity to cross‐infect other patients, or remain viable for longer time periods in the hospital environment.

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Optimization and Validation of Candida auris Short Tandem Repeat Analysis

Abstract: The emerging fungal pathogen Candida auris poses a threat to public health, mainly causing outbreaks in intensive care units. Genotyping is essential for investigating potential outbreaks and preventing further spread.

Cited by 9 publications

References 29 publications

Discovery of the sixthCandida aurisclade in Singapore

Discovery of the sixthCandida aurisclade in Singapore

Fast and Accurate Identification of Candida auris by High Resolution Mass Spectrometry

Whole genome sequencing analysis demonstrates therapy‐induced echinocandin resistance in Candida auris isolates

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