A targeted amplicon sequencing panel to simultaneously identify mosquito species and Plasmodium presence across the entire Anopheles genus

Anopheles is a diverse genus of mosquitoes comprising over 500 described species, including all known human malaria vectors. While a limited number of key vector species have been studied in detail, the goal of malaria elimination calls for surveillance of all potential vector species. Here, we develop a multilocus amplicon sequencing approach that targets 62 highly variable loci in the Anopheles genome and two conserved loci in the Plasmodium mitochondrion, simultaneously revealing both the mosquito species and whether that mosquito carries malaria parasites. We also develop a cheap, nondestructive, and high-throughput DNA extraction workflow that provides template DNA from single mosquitoes for the multiplex PCR, which means specimens producing unexpected results can be returned to for morphological examination. Over 1000 individual mosquitoes can be sequenced in a single MiSeq run, and we demonstrate the panel’s power to assign species identity using sequencing data for 40 species from Africa, Southeast Asia, and South America. We also show that the approach can be used to resolve geographic population structure within An. gambiae and An. coluzzii populations, as the population structure determined based on these 62 loci from over 1000 mosquitoes closely mirrors that revealed through whole genome sequencing. The end-to-end approach is quick, inexpensive, robust, and accurate, which makes it a promising technique for very large-scale mosquito genetic surveillance and vector control.     

Report on outcomes of valve-in-valve transcatheter aortic valve implantation and redo surgical aortic valve replacement in the Netherlands

Netherlands Heart Journal - We sought to investigate real-world outcomes of patients with degenerated biological aortic valve prostheses who had undergone valve-in-valve transcatheter aortic valve...     

ATP,an attractive target for the treatment of refractory chronic cough

Purinergic Signalling - Chronic cough is the most common complaint in respiratory clinics. Most of them have identifiable causes and some may respond to common disease-modifying therapies. However,...     

Fluoromethylcyclopropylamine derivatives as potential in vivo toxicophores – A cautionary disclosure

Fluorination of metabolic hotspots in a molecule is a common medicinal chemistry strategy to improve in vivo half-life and exposure and, generally, this strategy offers significant benefits. Here, we report the application of this strategy to a series of poly-ADP ribose glycohydrolase (PARG) inhibitors, resulting in unexpected in vivo toxicity which was attributed to this single-atom modification.     

The Effect of Different Environmental Conditions on the Viability of Naegleria fowleri Amoebae

Naegleria fowleri, a free‐living amoeba found in soil and freshwater environments, is the causative agent of Primary Amoebic Meningoencephalitis. Infection occurs when amoebae enter the nasal cavity, attach to the nasal mucosa and travel along olfactory neurons towards the olfactory bulb. Upon reaching the central nervous system, the amoebae replicate very rapidly and can cause death in 3–10 days. Little is known about the conditions in which the amoeba can survive in the environment. We have tested conditions beyond the known boundaries on the viability of amoebae by introducing them into moderate and extreme salinity, pH, and temperatures. Our data shows that although viability expectedly decreases towards each of these extreme conditions, their tolerance was much greater than anticipated, including viability in moderate salinity, a wide pH range, and temperatures higher than the previously reported 45 °C.     

Identification of common genetic variation that modulates alternative splicing

Alternative splicing of genes is an efficient means of generating variation in protein function. Several disease states have been associated with rare genetic variants that affect splicing patterns. Conversely, splicing efficiency of some genes is known to vary between individuals without apparent ill effects. What is not clear is whether commonly observed phenotypic variation in splicing patterns, and hence potential variation in protein function, is to a significant extent determined by naturally occurring DNA sequence variation and in particular by single nucleotide polymorphisms (SNPs). In this study, we surveyed the splicing patterns of 250 exons in 22 individuals who had been previously genotyped by the International HapMap Project. We identified 70 simple cassette exon alternative splicing events in our experimental system; for six of these, we detected consistent differences in splicing pattern between individuals, with a highly significant association between splice phenotype and neighbouring SNPs. Remarkably, for five out of six of these events, the strongest correlation was found with the SNP closest to the intron–exon boundary, although the distance between these SNPs and the intron–exon boundary ranged from 2 bp to greater than 1,000 bp. Two of these SNPs were further investigated using a minigene splicing system, and in each case the SNPs were found to exert cis-acting effects on exon splicing efficiency in vitro. The functional consequences of these SNPs could not be predicted using bioinformatic algorithms. Our findings suggest that phenotypic variation in splicing patterns is determined by the presence of SNPs within flanking introns or exons. Effects on splicing may represent an important mechanism by which SNPs influence gene function.