Biology subjects

Oliver, J. D.

Publications and source records attributed to Oliver, J. D..

3 recordsLinked to original sources

bioRxiv2021

Resistance profiling of Aspergillus fumigatus to olorofim indicates absence of intrinsic resistance and unveils the molecular mechanisms of acquired olorofim resistance

Olorofim (F901318) is a new antifungal currently under clinical development that shows both in vitro and in vivo activity against a number of filamentous fungi including Aspergillus fumigatus. In this study we screened A. fumigatus isolates for intrinsic olorofim-resistant A. fumigatus and evaluated the ability of A. fumigatus to acquire an olorofim-resistant phenotype. No intrinsic resistance was found in 975 clinical A. fumigatus isolates. However, we found that isolates with increased olorofim MICs (> 8 mg/L) could be selected using a high number of conidia and olorofim exposure under laboratory conditions. Assessment of the frequency of acquired olorofim resistance development of A. fumigatus was shown to be higher than for voriconazole but lower than for itraconazole. Sequencing the PyrE gene of isogenic isolates with olorofim MICs of >8 mg/L identified various amino acid substitutions with a hotspot at locus G119. Olorofim was shown to have reduced affinity to mutated target protein dihydroorotate dehydrogenase (DHODH) and the effect of these mutations were proven by introducing the mutations directly in A. fumigatus. We then investigated whether G119 mutations were associated with a fitness cost in A. fumigatus. These experiments showed a small but significant reduction in growth rate for strains with a G119V substitution, while strains with a G119C substitution did not exhibit a reduction in growth rate. These in vitro findings were confirmed in an in vivo pathogenicity model. ImportanceOlorofim represents an important new treatment option for patients with difficult to treat invasive fungal infections, including triazole-resistant A. fumigatus infection. Our study provides insights into one resistance mechanism and the potential dynamics of olorofim resistance, which will help to prevent and manage resistance selection. Such insights are critical to antifungal stewardship and to safeguard its prolonged use in clinical practice.

microbiology↗

bioRxiv2021

Nanopore adaptive sampling for mitogenome sequencing and bloodmeal identification in hematophagous insects

Blood-feeding insects are important vectors for an array of zoonotic pathogens. Despite significant research focused on well-documented insect vectors of One Health importance, resources for molecular species identification of a large number of hematophagous arthropods are limited. Advancements in next-generation sequencing technologies provide opportunities for targeting mitochondrial genomes of blood-feeding insects, as well as their bloodmeal hosts. This dual approach holds great promise for elucidating complex disease transmission pathways and enhancing the molecular resources for the identification of cryptic insect species. To this end, we leveraged the newly developed Oxford Nanopore Adaptive Sampling (NAS) pipeline to dually sequence the mitogenomes of hematophagous insects and their bloodmeals. Using NAS, we sequenced the entire mitogenomes of Aedes vexans, Culex restuans, Culex territans, and Chrysops niger and successfully identified bloodmeal hosts of Chrysops niger, Culex restuans, and Aedes trivittatus. We show that NAS has the utility to simultaneously molecularly identify blood-feeding insects and characterize disease transmission pathways through bloodmeal host identification. Moreover, our data indicate NAS can facilitate a wide array of molecular systematic studies through novel phylogenetic capture methods. We conclude the NAS approach has great potential for informing global One Health initiatives centered on the mitigation of vector-borne disease through dual vector and bloodmeal identification.

genomics↗

bioRxiv2021

Enabling metagenomic surveillance for bacterial tick-borne pathogens using nanopore sequencing with adaptive sampling

Technological and computational advancements in the fields of genomics and bioinformatics are providing exciting new opportunities for pathogen discovery and surveillance. In particular, single-molecule nucleotide sequence data originating from Oxford Nanopore Technologies (ONT) sequencing platforms can be bioinformatically leveraged, in real-time, for enhanced biosurveillance of a vast array of zoonoses. The recently released nanopore adaptive sampling (NAS) pipeline facilitates immediate mapping of individual nucleotide molecules (i.e., DNA, cDNA, and RNA) to a given reference as each molecule is sequenced. User-defined thresholds then allow for the retention or rejection of specific molecules, informed by the real-time reference mapping results, as they are physically passing through a given sequencing nanopore. Here, we show how NAS can be used to selectively sequence entire genomes of bacterial tick-borne pathogens circulating in wild populations of the blacklegged tick vector, Ixodes scapularis. The NAS method provided a two-fold increase in targeted pathogen sequences, successfully enriching for Borrelia (Borreliella) burgdorferi s.s.; Borrelia (Borrelia) miyamotoi; Anaplasma phagocytophilum; and Ehrlichia muris eauclairensis genomic DNA within our I. scapularis samples. Our results indicate that NAS has strong potential for real-time sequence-based pathogen surveillance.

microbiology↗