Candida (Lodderomyces) parapsilosis is a fungal opportunistic pathogen and an important cause of different forms of candidiasis, including candidemia, especially in hospitalized and frail patients. The emergence of resistance to main antifungal classes is a great challenge for its treatment, and the increasing number of reported resistant strains represents a global burden nowadays. Advances in genomics, particularly in whole-genome sequencing, allowed us to have a better understanding of the genetic basis of antifungal resistance, other than investigating the epidemiology and evolution of this pathogen. However, the genomic changes underlying C.paraprilosis adaptation to host environment and drug exposure during persistent infections remain incompletely understood. Analysis of sequential isolates from the same patient taken at different time spots of treatment can help to shed light on within-host antifungal resistance evolution and connect acquired genetic changes with drug sensitivity alterations. In this study, eighteen clinical isolates of C. parapsilosis were collected from blood cultures of eight patients with candidemia at the IRCCS Fondazione Policlinico San Matteo in Pavia, Italy between 2018 to 2024. The isolates were obtained at different time points during patient treatment, and the assessment of sensitivity to different drugs was conducted through the measurement of Minimal Inhibitory Concentration (MIC). For the isolates from individual patients who exhibited changes in MIC over time (within a close time range), whole-genome sequencing of isolates was performed, allowing us to investigate genomic changes occurring during individual infections. Sequencing data were analyzed using a comparative genomics approach to characterize genetic variation among isolates. To ensure MIC changes were related to within-host evolution and not co-infection by multiple isolates, phylogenetic relationships and cluster analysis were performed, highlighting three cases in which the phenotype variation could be possibly explained by acquired genetic changes. Genetic mutations that were identified in these isolates included variants affecting genes potentially involved in cellular adaptation, as well as variants affecting Major Facilitator Superfamily (MFS) proteins and potentially their expression, which can explain the different drug response during the treatment. Additional genomic changes, including changes in ploidy, were detected in selected isolates. Overall, the analysis showed that C. parapsilosis can accumulate genetic changes during persistent infection. The genomic variation occurred along with alterations in antifungal susceptibility and could contribute to these phenotypic changes, highlighting possible novel mechanisms for further investigation of antifungal resistance development.
Candida (Lodderomyces) parapsilosis è un patogeno fungino opportunista e un’importante causa di diverse forme di candidiasi, tra cui la candidemia, soprattutto nei pazienti ospedalizzati e fragili. L’emergere di resistenze alle principali classi di farmaci antifungini rappresenta una grande sfida per il trattamento, e il numero crescente di ceppi resistenti segnalati costituisce oggi un problema di rilevanza globale. I progressi nel campo della genomica, in particolare il sequenziamento dell’intero genoma (whole-genome sequencing), hanno permesso di comprendere meglio le basi genetiche della resistenza agli antifungini, nonché di studiare l’epidemiologia e l’evoluzione di questo patogeno. Tuttavia, i cambiamenti genomici alla base dell’adattamento di C. parapsilosis all’ambiente dell’ospite e all’esposizione ai farmaci durante le infezioni persistenti rimangono ancora poco conosciuti. L’analisi di isolati sequenziali ottenuti dallo stesso paziente in diversi momenti del trattamento può contribuire a chiarire l’evoluzione della resistenza agli antifungini all’interno dell’ospite e a correlare i cambiamenti genetici acquisiti con le variazioni della sensibilità ai farmaci. In questo studio, sono stati raccolti diciotto isolati clinici di C. parapsilosis da emocolture di otto pazienti affetti da candidemia presso l’IRCCS Fondazione Policlinico San Matteo di Pavia, Italia, nel periodo compreso tra il 2018 e il 2024. Gli isolati sono stati ottenuti in diversi momenti durante il trattamento dei pazienti e la sensibilità a diversi farmaci è stata valutata mediante la determinazione della concentrazione minima inibente (MIC). Per gli isolati provenienti dai singoli pazienti che hanno mostrato variazioni della MIC nel tempo, in un intervallo temporale ravvicinato, è stato eseguito il sequenziamento dell’intero genoma, consentendo di studiare i cambiamenti genomici verificatisi nel corso delle singole infezioni. I dati di sequenziamento sono stati analizzati mediante un approccio di genomica comparativa al fine di caratterizzare la variabilità genetica tra gli isolati. Per verificare che le variazioni della MIC fossero correlate all’evoluzione all’interno dell’ospite e non alla presenza di infezioni concomitanti sostenute da isolati multipli, sono state analizzate le relazioni filogenetiche e sono state effettuate analisi di clustering, evidenziando tre casi in cui la variazione fenotipica potrebbe essere spiegata, almeno in parte, da cambiamenti genetici acquisiti. Le mutazioni genetiche identificate in questi isolati includevano varianti a carico di geni potenzialmente coinvolti nell’adattamento cellulare, nonché varianti che interessavano proteine della Major Facilitator Superfamily (MFS) e potenzialmente la loro espressione, che potrebbero spiegare le differenti risposte ai farmaci osservate durante il trattamento. In alcuni isolati sono stati inoltre rilevati ulteriori cambiamenti genomici, comprese variazioni della ploidia. Nel complesso, l’analisi ha mostrato che C. parapsilosis può accumulare cambiamenti genetici durante un’infezione persistente. La variazione genomica si è verificata in concomitanza con alterazioni della sensibilità agli antifungini e potrebbe contribuire ai cambiamenti fenotipici osservati, evidenziando possibili nuovi meccanismi che meritano ulteriori approfondimenti nell’ambito dello sviluppo della resistenza agli antifungini.
Studio dell’evoluzione della resistenza agli antifungini di Candida (Lodderomyces) parapsilosis all’interno dell’ospite mediante sequenziamento dell’intero genoma
SEMENOVA, NATALIA
2025/2026
Abstract
Candida (Lodderomyces) parapsilosis is a fungal opportunistic pathogen and an important cause of different forms of candidiasis, including candidemia, especially in hospitalized and frail patients. The emergence of resistance to main antifungal classes is a great challenge for its treatment, and the increasing number of reported resistant strains represents a global burden nowadays. Advances in genomics, particularly in whole-genome sequencing, allowed us to have a better understanding of the genetic basis of antifungal resistance, other than investigating the epidemiology and evolution of this pathogen. However, the genomic changes underlying C.paraprilosis adaptation to host environment and drug exposure during persistent infections remain incompletely understood. Analysis of sequential isolates from the same patient taken at different time spots of treatment can help to shed light on within-host antifungal resistance evolution and connect acquired genetic changes with drug sensitivity alterations. In this study, eighteen clinical isolates of C. parapsilosis were collected from blood cultures of eight patients with candidemia at the IRCCS Fondazione Policlinico San Matteo in Pavia, Italy between 2018 to 2024. The isolates were obtained at different time points during patient treatment, and the assessment of sensitivity to different drugs was conducted through the measurement of Minimal Inhibitory Concentration (MIC). For the isolates from individual patients who exhibited changes in MIC over time (within a close time range), whole-genome sequencing of isolates was performed, allowing us to investigate genomic changes occurring during individual infections. Sequencing data were analyzed using a comparative genomics approach to characterize genetic variation among isolates. To ensure MIC changes were related to within-host evolution and not co-infection by multiple isolates, phylogenetic relationships and cluster analysis were performed, highlighting three cases in which the phenotype variation could be possibly explained by acquired genetic changes. Genetic mutations that were identified in these isolates included variants affecting genes potentially involved in cellular adaptation, as well as variants affecting Major Facilitator Superfamily (MFS) proteins and potentially their expression, which can explain the different drug response during the treatment. Additional genomic changes, including changes in ploidy, were detected in selected isolates. Overall, the analysis showed that C. parapsilosis can accumulate genetic changes during persistent infection. The genomic variation occurred along with alterations in antifungal susceptibility and could contribute to these phenotypic changes, highlighting possible novel mechanisms for further investigation of antifungal resistance development.| File | Dimensione | Formato | |
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https://hdl.handle.net/20.500.14239/37327