SynopsisThe structure of an Hfq homolog from the deep-branching thermophilic bacterium Aquifex aeolicus, determined to 1.5-Å resolution both in apo form and bound to a uridine-rich RNA, reveals a conserved, pre-organized RNA-binding pocket on the lateral rim of the Hfq hexamer.AbstractThe host factor Hfq, as the bacterial branch of the Sm family, is an RNA-binding protein involved in post-transcriptional regulation of mRNA expression and turnover. Hfq facilitates pairing between small regulatory RNAs (sRNA) and their corresponding mRNA targets by binding both RNAs and bringing them into close proximity. Hfq homologs self-assemble into homo-hexameric rings, with at least two distinct surfaces that bind RNA. Recently, another binding site—dubbed the ‘lateral rim’—has been implicated in sRNA•mRNA annealing; the RNA-binding properties of this site appear to be rather subtle, and its degree of evolutionary conservation is unknown. An Hfq homolog has been identified in the phylogenetically deep-branching thermophile Aquifex aeolicus (Aae), but little is known about the structures and functions of Hfq from basal bacterial lineages such as the Aquificae. Thus, we have cloned, overexpressed, purified, crystallized, and biochemically characterized Aae Hfq. We have determined the structures of Aae Hfq in space-groups P1 and P6, both to 1.5 Å resolution, and we have discovered nanomolar-scale binding affinities for uridine- and adenosine-rich RNAs. Co-crystallization with U6 RNA reveals that the outer rim of the Aae Hfq hexamer features a well-defined binding pocket that is selective for uracil. This Aae Hfq structure, combined with biochemical and biophysical characterization of the homolog, reveals deep evolutionary conservation of the lateral RNA-binding mode, and lays a foundation for further studies of Hfq-associated RNA biology in ancient bacterial phyla.
Effective mosquito surveillance and control relies on rapid and accurate identification of mosquito vectors and confounding sympatric species. As adoption of modified mosquito (MM) control techniques has increased, the value of monitoring the success of interventions has gained recognition and has pushed the field away from traditional ‘spray and pray’ approaches. Field evaluation and monitoring of MM control techniques that target specific species require massive volumes of surveillance data involving species-level identifications. However, traditional surveillance methods remain time and labor-intensive, requiring highly trained, experienced personnel. Health districts often lack the resources needed to collect essential data, and conventional entomological species identification involves a significant learning curve to produce consistent high accuracy data. These needs led us to develop MosID: a device that allows for high-accuracy mosquito species identification to enhance capability and capacity of mosquito surveillance programs. The device features high-resolution optics and enables batch image capture and species identification of mosquito specimens using computer vision. While development is ongoing, we share an update on key metrics of the MosID system. The identification algorithm, tested internally across 16 species, achieved 98.4 ± 0.6% % macro F1-score on a dataset of known species, unknown species used in training, and species reserved for testing (species, specimens respectively: 12, 1302; 12, 603; 7, 222). Preliminary user testing showed specimens were processed with MosID at a rate ranging from 181-600 specimens per hour. We also discuss other metrics within technical scope, such as mosquito sex and fluorescence detection, that may further support MM programs.
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