Genomic analyses reveal high diversity and rapid evolution of Pichia kudriavzevii within a neonatal intensive care unit in Delhi, India.

Pichia kudriavzevii causes life-threatening infections in immunocompromised hosts, including hospitalized neonates. This pathogen is intrinsically resistant to fluconazole, while uncommon P. kudriavzevii strains resistant to multiple antifungal drugs, including voriconazole, amphotericin B, and echinocandins, have also been reported from healthcare environments. Thus, understanding how P. kudriavzevii spread, persist, and adapt to healthcare settings could help us develop better infection management strategies. In this study, whole genome sequencing identifies multiple outbreaks of bloodstream infections in a single neonatal intensive care unit (NICU) over 5 years caused by genetically diverse strains of P. kudriavzevii. Interestingly, two genetically distinct clusters of P. kudriavzevii strains showed frequent loss of heterozygosity (LOH) events between two temporal samples. The first outbreak cluster (2015-2016) showed LOH at chromosomes 1, 4, and 5, and the other outbreak cluster (2020) exhibited LOH at chromosome 2. The circulation of two separate strain clusters of P. kudriavzevii suggests nosocomial transmission in the NICU in different time periods. Furthermore, we compared the transcriptomic profiles of three isolates of clusters I and II that exhibited distinct fluconazole and itraconazole MICs. While no significant difference in gene expression was found at the azole-target gene ERG11 or the ATP-binding cassette (ABC) transporter genes, such differences were found in genes involved in cell division and filamentation, such as SIR2 (sirtuin deacetylase) and RFA1 (replication factor A). Interestingly, increased filamentation was observed in clade I isolate exhibiting high fluconazole MICs. Together, our study indicates significant diversity, persistence, and rapid evolution of P. kudriavzevii within a single NICU.