A genotyping array for the globally invasive vector mosquito, Aedes albopictus

Background: Although whole-genome sequencing (WGS) is the preferred genotyping method for most genomic analyses, limitations are often experienced when studying genomes characterized by a high percentage of repetitive elements, high linkage, and recombination deserts. The Asian tiger mosquito (Aedes albopictus), for example, has a genome comprising up to 72% repetitive elements, and therefore we set out to develop a single-nucleotide polymorphism (SNP) chip to be more cost-effective. Aedes albopictus is an invasive species originating from Southeast Asia that has recently spread around the world and is a vector for many human diseases. Developing an accessible genotyping platform is essential in advancing biological control methods and understanding the population dynamics of this pest species, with significant implications for public health. Methods: We designed a SNP chip for Ae. albopictus (Aealbo chip) based on approximately 2.7 million SNPs identified using WGS data from 819 worldwide samples. We validated the chip using laboratory single-pair crosses, comparing technical replicates, and comparing genotypes of samples genotyped by WGS and the SNP chip. We then used the chip for a population genomic analysis of 237 samples from 28 sites in the native range to evaluate its usefulness in describing patterns of genomic variation and tracing the origins of invasions. Results: Probes on the Aealbo chip targeted 175,396 SNPs in coding and non-coding regions across all three chromosomes, with a density of 102 SNPs per 1 Mb window, and at least one SNP in each of the 17,461 protein-coding genes. Overall, 70% of the probes captured the genetic variation. Segregation analysis found that 98% of the SNPs followed expectations of single-copy Mendelian genes. Comparisons with WGS indicated that sites with genotype disagreements were mostly heterozygotes at loci with WGS read depth \u3c 20, while there was near complete agreement with WGS read depths \u3e 20, indicating that the chip more accurately detects heterozygotes than low-coverage WGS. Sample sizes did not affect the accuracy of the SNP chip genotype calls. Ancestry analyses identified four to five genetic clusters in the native range with various levels of admixture. Conclusions: The Aealbo chip is highly accurate, is concordant with genotypes from WGS with high sequence coverage, and may be more accurate than low-coverage WGS. Graphical Abstract: (Figure presented.) © The Author(s) 2024

Grain and seed protein functionality

Editado por Jiménez-López, José Carlos (CSIC

Infection of Anopheles aquasalis from symptomatic and asymptomatic Plasmodium vivax infections in Manaus, western Brazilian Amazon

BACKGROUND: Asymptomatic individuals are one of the major challenges for malaria elimination programs in endemic areas. In the absence of clinical symptoms and with a lower parasite density they constitute silent reservoirs considered important for maintaining transmission of human malaria. Studies from Brazil have shown that infected individuals may carry these parasites for long periods. RESULTS: Patients were selected from three periurban endemic areas of the city of Manaus, in the western Brazilian Amazon. Symptomatic and asymptomatic patients with positive thick blood smear and quantitative real-time PCR (qPCR) positive for Plasmodium vivax were invited to participate in the study. A standardised pvs25 gene amplification by qPCR was used for P. vivax gametocytes detection. Anopheles aquasalis were fed using membrane feeding assays (MFA) containing blood from malaria patients. Parasitemia of 42 symptomatic and 25 asymptomatic individuals was determined by microscopic examination of blood smears and qPCR. Parasitemia density and gametocyte density were assessed as determinants of infection rates and oocysts densities. A strong correlation between gametocyte densities (microscopy and molecular techniques) and mosquito infectivity (P < 0.001) and oocysts median numbers (P < 0.05) was found in both groups. The ability to infect mosquitoes was higher in the symptomatic group (41%), but infectivity in the asymptomatic group was also seen (1.42%). CONCLUSIONS: Although their infectivity to mosquitoes is relatively low, given the high prevalence of P. vivax asymptomatic carriers they are likely to play and important role in malaria transmission in the city of Manaus. The role of asymptomatic infections therefore needs to be considered in future malaria elimination programs in Brazil

Proposal of an automatic method to measure the cell free layer in microchannels with bifurcations

Ao longo dos anos, a espessura da camada de plasma tem sido determinada com o auxílio de métodos manuais. Apesar destes métodos serem bastante fiáveis, estes são morosos e podem introduzir erros do utilizador nos dados. No presente trabalho, foi desenvolvido um método automático de processamento de imagem para a determinação da espessura camada de plasma de uma forma automática

Cell-free layer measurements in a bifurcation microchannel : comparison between a manual and automatic methods

O artigo deve também estar associado ao Departamento de Electrotécnia, Tecnologia Mecânica e QuímicaIn the present work, in vitro blood flowing through a bifurcation microchannel was studied. The aim was to measure the Trajectories of the cell-free layer (CFL) by using different methods, i. e., a manual and two automatic methods.The authors acknowledge the financial support provided by: PTDC/ SAU-BEB/108728/2008, PTDC/SAU-BEB/ 105650/2008, PTDC/EMEMFE/099109/2008 and PTDC/SAU ENB/116929/2010 from the FCT (Science and Technology Foundation) and COMPETE, Portugal

Cell-free layer (CFL) measurements in complex geometries: contractions and bifurcations

In this chapter we discuss the cell-free layer (CFL) developed adjacent to the wall of microgeometries containing complex features representative of the microcirculation, such as contractions, expansions, bifurcations and confluences. The microchannels with the different geometries were made of polydimethylsiloxane (PDMS) and we use optical techniques to evaluate the cell-free layer for red blood cells (RBC) suspensions with different hematocrit (Hct). The images are captured using a high-speed video microscopy system and the thickness of the cell free layer was measured using both manual and automatic image analysis techniques. The results show that in in vitro microcirculation, the hematocrit and the geometrical configuration have a major impact on the CFL thickness. In particular, the thickness of the cell-free layer increases as the fluid flows through a contraction-expansion sequence and that this increase is enhanced for lower hematocrit. In contrast, the flow rates tested in this studies did not show a clear influence on the CFL thickness.The authors acknowlcdge the financiaI suppart provided by: PTDC/SAUBEB/l08728/2008, PTDC/SAU-BEB/l05650/2008, PTDC/EME-MFEI099109/2008 and PTDC/SAU-ENB/116929/2010 from FCT (Science and Technnlogy Foundation), COMPETE, QREN and European Union (FEDER). The authors are graterul to Mônica Oliveira for many valuable camments on this study

Assessing the Effects of <i>Aedes aegypti kdr</i> Mutations on Pyrethroid Resistance and Its Fitness Cost

<div><p>Pyrethroids are the most used insecticide class worldwide. They target the voltage gated sodium channel (Na_V), inducing the knockdown effect. In <i>Aedes aegypti</i>, the main dengue vector, the AaNa_V substitutions Val1016Ile and Phe1534Cys are the most important knockdown resistance (<i>kdr</i>) mutations. We evaluated the fitness cost of these <i>kdr</i> mutations related to distinct aspects of development and reproduction, in the absence of any other major resistance mechanism. To accomplish this, we initially set up 68 crosses with mosquitoes from a natural population. Allele-specific PCR revealed that one couple, the one originating the CIT-32 strain, had both parents homozygous for both <i>kdr</i> mutations. However, this pyrethroid resistant strain also presented high levels of detoxifying enzymes, which synergistically account for resistance, as revealed by biological and biochemical assays. Therefore, we carried out backcrosses between CIT-32 and Rockefeller (an insecticide susceptible strain) for eight generations in order to bring the <i>kdr</i> mutation into a susceptible genetic background. This new strain, named Rock-kdr, was highly resistant to pyrethroid and presented reduced alteration of detoxifying activity. Fitness of the Rock-kdr was then evaluated in comparison with Rockefeller. In this strain, larval development took longer, adults had an increased locomotor activity, fewer females laid eggs, and produced a lower number of eggs. Under an inter-strain competition scenario, the Rock-kdr larvae developed even slower. Moreover, when Rockefeller and Rock-kdr were reared together in population cage experiments during 15 generations in absence of insecticide, the mutant allele decreased in frequency. These results strongly suggest that the <i>Ae. aegypti kdr</i> mutations have a high fitness cost. Therefore, enhanced surveillance for resistance should be priority in localities where the <i>kdr</i> mutation is found before new adaptive alleles can be selected for diminishing the <i>kdr</i> deleterious effects.</p> </div

Activity of enzymes related to insecticide metabolic resistance in <i>Aedes aegypti</i> strains.

<p>The cut-offs are (dashed lines) determined by the Rockefeller 99 percentile value of each enzyme (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0060878#pone.0060878-Montella1" target="_blank">[11]</a>). Rockefeller is a reference strain of insecticide susceptibility and vigor. Distributions with less than 15% of individuals beyond the cut-off are considered unaltered. Between 15 and 50% are altered and above 50% are highly altered. CIT-32 is the original <i>kdr</i> strain, derived from a pyrethroid resistant Brazilian <i>Aedes aegypti</i> population. Rock-kdr is the <i>kdr</i> strain, backcrossed for eight generations with Rockefeller in order to reduce the contribution of detoxification enzymes to pyrethroid resistance.</p