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Principal Investigator: Jarrod R. Fortwendel
Organization: UNIVERSITY OF TENNESSEE HEALTH SCI CTR
Fiscal Year: 2024
Award: $656,395
Funding agency: National Institute of Allergy and Infectious Diseases
A critical barrier to overcoming triazole resistance in Aspergillus fumigatus is the significant lack of
understanding of its genetic and molecular basis. We have shown that the known mechanisms of resistance do
not fully explain resistance observed among most clinical isolates. Our long-term goal is to improve antifungal
therapy and ensure the sustained clinical utility of the triazole class for treatment of infections caused by
Aspergillus species. Our central hypothesis is that non-cyp51A-mutation mediated mechanisms are essential
to triazole resistance in clinical isolates of A. fumigatus and involve complex genetic changes altering 1) sterol
biosynthesis and its transcriptional activation, 2) triazole transport and its transcriptional activation, and 3) as yet
unknown mechanisms. Our current objective is to address critical knowledge gaps by identifying the genetic
and molecular determinants of non-cyp51A-mutation mediated resistance. Our preliminary data suggest that
while mutations in cyp51A among triazole resistant clinical isolates are common, their overall contribution to
resistance is minimal. We have observed mutations, unique to resistant isolates in our collection, in genes
encoding sterol sensing proteins, regulators of sterol biosynthesis, and sterol biosynthesis enzymes. We have
also observed clinical isolates that overexpress not only cyp51A, but most genes of the ergosterol biosynthesis
pathway, suggesting its constitutive activation. We have observed several potential transporters that are up-
regulated among triazole resistant isolates in our collection, suggesting a role for triazole efflux and resistance
by these transporters. We have also shown that clinical isolates of A. fumigatus take up triazole antifungals via
facilitated diffusion and we believe that altered triazole import may represent an important mechanism of
resistance. To accomplish our objective we will undertake experiments that will lead to an understanding of what
genetic and molecular determinants influence triazole susceptibility through altered sterol biosynthesis or its
transcriptional activation (Aim 1) and triazole transport and its regulation (Aim 2). In Aim 3, we will also utilize an
unbiased whole genome comparisons, coupled with in vitro evolution experiments, to identify completely novel
mechanisms of resistance in clinical isolates. Our approach is innovative as we will use the latest genetic and
genomic techniques to study and discover novel non-cyp51A-mutation mediated mechanisms of triazole
resistance that are operative in a U.S.-based collection of triazole resistant clinical isolates. The proposed
research is significant as it represents a comprehensive analysis of the molecular and genetic basis of non-
cyp51A-mutation mediated triazole resistance in A. fumigatus and will provide novel insights into ways in which
triazole activity can be improved against this important human pathogen.
Terms: <3-hydroxy-3-methylglutaryl-CoA><3-hydroxy-3-methylglutaryl-coenzyme A><A fumigatus><A. fumigatus><Address><Affect><Alleles><Allelomorphs><Anabolism><Antifungal Therapy><Aspergillosis><Aspergillus><Aspergillus fumigatus><Aspergillus resistance><Automobile Driving><Azole resistance><Azole resistant><Azoles><CRISPR approach><CRISPR based approach><CRISPR method><CRISPR methodology><CRISPR technique><CRISPR technology><CRISPR tools><CRISPR-CAS-9><CRISPR-based method><CRISPR-based technique><CRISPR-based technology><CRISPR-based tool><CRISPR/CAS approach><CRISPR/Cas method><CRISPR/Cas technology><CRISPR/Cas9><CRISPR/Cas9 technology><Cas nuclease technology><Clinical><Clustered Regularly Interspaced Short Palindromic Repeats approach><Clustered Regularly Interspaced Short Palindromic Repeats method><Clustered Regularly Interspaced Short Palindromic Repeats methodology><Clustered Regularly Interspaced Short Palindromic Repeats technique><Clustered Regularly Interspaced Short Palindromic Repeats technology><Collection><Complex><Coupled><DNA Alteration><DNA Sequence Alteration><DNA mutation><Data><Death Rate><Dehydrogenases><Diagnosis><Diffusion><Disease><Disorder><Drug Transport><Ensure><Ergosterol><Evolution><Exhibits><Expression Signature><Filamentous Fungi><Fungal Drug Resistance><Fungus drug resistant><Gene Expression Monitoring><Gene Expression Pattern Analysis><Gene Expression Profile><Gene Expression Profiling><Genes><Genetic><Genetic Alteration><Genetic Change><Genetic analyses><Genetic defect><Genetic mutation><Genome><Genomics><Goals><HMG-CoA><Immunocompromised><Immunocompromised Host><Immunocompromised Patient><Immunosuppressed Host><In Vitro><Individual><Infection><Knowledge><Laboratories><Mediating><Methods><Molds><Molecular><Molecular Analysis><Morbidity><Morbidity - disease rate><Mutation><Outcome><Oxidoreductase><Oxidoreductase Gene><Pathway interactions><Play><Predisposition><Proteins><Reductases><Regulation><Research><Resistance><Resistance development><Resistance profile><Resistant development><Resistant profile><Risk><Role><SEQ-AN><Sequence Alteration><Sequence Analyses><Sequence Analysis><Sterol Biosynthesis><Sterol Biosynthesis Pathway><Sterols><Susceptibility><System><Techniques><Testing><Therapeutic><Therapeutic Fungicides><Transcript Expression Analyses><Transcript Expression Analysis><Transcription Activation><Transcriptional Activation><Translating><Treatment Failure><Voriconazole><Work><analyze gene expression><anti-fungal><anti-fungal agents><anti-fungal drug><anti-fungal drug resistance><anti-fungal drug resistant><anti-fungal resistance><anti-fungal resistant><biosynthesis><clinical predictors><clinical relevance><clinically relevant><developing resistance><differential expression><differentially expressed><diffused><diffuses><diffusing><diffusions><driving><effective therapy><effective treatment><entire genome><enzyme biosynthesis><experiment><experimental research><experimental study><experiments><full genome><fungal infectious disease treatment><fungus drug resistance><gene expression analysis><gene expression assay><gene expression pattern><gene expression signature><gene manipulation><genetic analysis><genetic manipulation><genetically manipulate><genetically perturb><genome analysis><genome mutation><genomic alteration><human pathogen><hydroxymethylglutaryl-CoA><immunosuppressed patient><improved><inhibitor><innovate><innovation><innovative><insight><mortality><mortality rate><mortality ratio><mutant><new approaches><novel><novel approaches><novel strategies><novel strategy><overexpress><overexpression><pathway><patient population><prevent><preventing><public health relevance><resistance in Aspergillus><resistance mechanism><resistance mutation><resistance to anti-fungal><resistance to azole><resistant><resistant Aspergillus><resistant mechanism><resistant mutation><resistant to anti-fungal><resistant to azole><social role><stem><therapy failure><transcriptional differences><transcriptional profile><transcriptional profiling><transcriptional signature><treatment choice><whole genome>