Designing Aedes (Diptera: Culicidae) Mosquito Traps: The Evolution of the Male Aedes Sound Trap by Iterative Evaluation.

Effective surveillance of Aedes aegypti (Linnaeus, Diptera: Culicidae) is critical to monitoring the impact of vector control measures when mitigating disease transmission by this species. There are benefits to deploying male-specific traps, particularly when a high level of catch-specificity is desired. Here, the rationale behind the developmental process of an entirely new trap which uses a sound lure to capture male Ae. aegypti, the male Aedes sound trap (MAST), is presented as a target product profile with findings from developmental trials of key trap components and performance. Trial results suggest that the presence of a black base associated with the trap influenced male catches as did variations in size of this base, to a degree. Trap entrance shape didn't influence catch rates, but entrance size did. No significant differences in catch rates were found when sound lures were set to intermittent or continuous playbacks, at volumes between 63-74 dB or frequencies of 450 Hz compared to 500 Hz. Additionally, adult males aged 3 days post-eclosion, were less responsive to sound lures set to 500 Hz than those 4 or 6 days old. Lastly, almost no males were caught when the MAST directly faced continual winds of 1.5 ms-1, but males were captured at low rates during intermittent winds, or if the trap faced away from the wind. The developmental process to optimising this trap is applicable to the development of alternate mosquito traps beyond Aedes sound traps and provides useful information towards the improved surveillance of these disease vectors.

Genetic and environmental influences on cortisol reactivity to a psychosocial stressor in adolescents and young adults.

Individuals vary in their response to psychological and physiological stressors, and this reactivity can be captured using measures of cortisol. Previous research suggests cortisol reactivity is under some degree of genetic control; however, the measures used have varied widely. This study (N = 524) examined potential differences in heritability across varying cortisol metrics of stress reactivity following the Trier Social Stress Test (TSST) and whether these measures are genetically or environmentally interrelated. Participants included twins aged 15-20 years (56% female). Cortisol reactivity to the TSST was assessed via serial salivary cortisol samples collected pre- and post-TSST. Modest to moderate heritability estimates (12% [95CI: 1-36%] - 45% [95CI: 16-69%]) were observed across measures purported to capture stress reactivity (peak, area under the curve [AUC], baseline-to-peak change). Findings also demonstrate both shared and unique genetic and environmental influences between baseline cortisol and cortisol reactivity. Minimal to no additional genetic innovations above and beyond the contributions of peak cortisol were found for other measures of cortisol reactivity such as AUC. This study is one of the largest twin-based samples to examine the heritability of cortisol reactivity, and results suggest that simpler measures of cortisol reactivity demonstrate higher heritability compared to more complex measurements.

Outcomes from international field trials with Male Aedes Sound Traps: Frequency-dependent effectiveness in capturing target species in relation to bycatch abundance.

Aedes aegypti and Aedes albopictus vector dengue, chikungunya and Zika viruses. With both species expanding their global distributions at alarming rates, developing effective surveillance equipment is a continuing priority for public health researchers. Sound traps have been shown, in limited testing, to be highly species-specific when emitting a frequency corresponding to a female mosquito wingbeat. Determining male mosquito capture rates in sound traps based on lure frequencies in endemic settings is the next step for informed deployment of these surveillance tools. We field-evaluated Male Aedes Sound Traps (MASTs) set to either 450 Hz, 500 Hz, 550 Hz or 600 Hz for sampling Aedes aegypti and/or Aedes albopictus and compared catch rates to BG-Sentinel traps within Pacific (Madang, Papua New Guinea) and Latin American (Molas, Mexico and Orange Walk Town, Belize) locations. MASTs set to 450-550 Hz consistently caught male Ae. aegypti at rates comparable to BG-Sentinel traps in all locations. A peak in male Ae. albopictus captures in MASTs set at 550 Hz was observed, with the lowest mean abundance recorded in MASTs set to 450 Hz. While significantly higher abundances of male Culex were sampled in MASTs emitting lower relative frequencies in Molas, overall male Culex were captured in significantly lower abundances in the MASTs, relative to BG-Sentinel traps within all locations. Finally, significant differences in rates at which male Aedes and Culex were positively detected in trap-types per weekly collections were broadly consistent with trends in abundance data per trap-type. MASTs at 550 Hz effectively captured both male Ae. aegypti and Ae. albopictus while greatly reducing bycatch, especially male Culex, in locations where dengue transmission has occurred. This high species-specificity of the MAST not only reduces staff-time required to sort samples, but can also be exploited to develop an accurate smart-trap system-both outcomes potentially reducing public health program expenses.

A Low-Powered and Highly Selective Trap for Male Aedes (Diptera: Culicidae) Surveillance: The Male Aedes Sound Trap.

As Aedes aegypti (Linnaeus, Diptera: Culicidae) expands its global distribution and vectors a range of debilitating arboviruses there is an increased need for enhanced mosquito surveillance. Consequently, we developed a Male Aedes Sound Trap (MAST) that requires minimal power and is highly species-specific. Two different versions of the MAST were developed, one that uses synthetic pyrethroid to kill captured mosquitoes (MAST Spray) and another which has an internal divider to create a killing chamber in which a sticky panel can be placed to capture mosquitoes (MAST Sticky). We compared weekly capture rates of male Ae. aegypti and bycatch from the two MAST versions to those from BG-Sentinel (BGS) traps and Sound-producing BG-Gravid Aedes Traps (SGATs) throughout Cairns, northern Australia. Weekly mean male Ae. aegypti catches did not significantly differ between trap types. However, the rate of positive weekly detections of male Ae. aegypti was lower for the MAST Sticky than the other three trap types. The MASTs sampled significantly fewer mosquitoes other than male Ae. aegypti, than either the BGS trap or the SGAT. Also, the MASTs and SGATs all caught significantly less non-Culicidae bycatch than the BGS traps. Consequently, we have developed a versatile male Ae. aegypti trap which is potentially of great benefit to Ae. aegypti surveillance programs.

Author Correction: Efficient production of male Wolbachia-infected Aedes aegypti mosquitoes enables large-scale suppression of wild populations.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Efficient production of male Wolbachia-infected Aedes aegypti mosquitoes enables large-scale suppression of wild populations.

The range of the mosquito Aedes aegypti continues to expand, putting more than two billion people at risk of arboviral infection. The sterile insect technique (SIT) has been used to successfully combat agricultural pests at large scale, but not mosquitoes, mainly because of challenges with consistent production and distribution of high-quality male mosquitoes. We describe automated processes to rear and release millions of competitive, sterile male Wolbachia-infected mosquitoes, and use of these males in a large-scale suppression trial in Fresno County, California. In 2018, we released 14.4 million males across three replicate neighborhoods encompassing 293 hectares. At peak mosquito season, the number of female mosquitoes was 95.5% lower (95% CI, 93.6-96.9) in release areas compared to non-release areas, with the most geographically isolated neighborhood reaching a 99% reduction. This work demonstrates the high efficacy of mosquito SIT in an area ninefold larger than in previous similar trials, supporting the potential of this approach in public health and nuisance-mosquito eradication programs.

Evolutionary superscaffolding and chromosome anchoring to improve Anopheles genome assemblies.

BACKGROUND: New sequencing technologies have lowered financial barriers to whole genome sequencing, but resulting assemblies are often fragmented and far from 'finished'. Updating multi-scaffold drafts to chromosome-level status can be achieved through experimental mapping or re-sequencing efforts. Avoiding the costs associated with such approaches, comparative genomic analysis of gene order conservation (synteny) to predict scaffold neighbours (adjacencies) offers a potentially useful complementary method for improving draft assemblies. RESULTS: We evaluated and employed 3 gene synteny-based methods applied to 21 Anopheles mosquito assemblies to produce consensus sets of scaffold adjacencies. For subsets of the assemblies, we integrated these with additional supporting data to confirm and complement the synteny-based adjacencies: 6 with physical mapping data that anchor scaffolds to chromosome locations, 13 with paired-end RNA sequencing (RNAseq) data, and 3 with new assemblies based on re-scaffolding or long-read data. Our combined analyses produced 20 new superscaffolded assemblies with improved contiguities: 7 for which assignments of non-anchored scaffolds to chromosome arms span more than 75% of the assemblies, and a further 7 with chromosome anchoring including an 88% anchored Anopheles arabiensis assembly and, respectively, 73% and 84% anchored assemblies with comprehensively updated cytogenetic photomaps for Anopheles funestus and Anopheles stephensi. CONCLUSIONS: Experimental data from probe mapping, RNAseq, or long-read technologies, where available, all contribute to successful upgrading of draft assemblies. Our evaluations show that gene synteny-based computational methods represent a valuable alternative or complementary approach. Our improved Anopheles reference assemblies highlight the utility of applying comparative genomics approaches to improve community genomic resources.

Investigating Male Aedes aegypti (Diptera: Culicidae) Attraction to Different Oviposition Containers Using Various Configurations of the Sound Gravid Aedes Trap.

Aedes aegypti (Linnaeus), the primary vectors of the arboviruses dengue virus and Zika virus, continue to expand their global distributions. In efforts to better control such species, several mosquito control programs are investigating the efficacy of rearing and releasing millions of altered male Aedes throughout landscapes to reduce populations and disease transmission risk. Unfortunately, little is known about Ae. aegypti, especially male, dispersal behaviors within urban habitats. We deployed Sound-producing Gravid Aedes Traps (SGATs) in Cairns, northern Australia, to investigate male Ae. aegypti attraction to various oviposition container configurations. The traps were arranged to include: 1) water only, 2) organically infused water, 3) infused water and L3 larvae, 4) infused water and a human-scented lure, and lastly 5) no water or olfactory attractant (dry). Our data suggest that males were more attracted to SGATs representing active larval sites than potential larval sites, but were equally attracted to dry SGATs relative to those containing water and/or infusion. Additionally, we found that female Ae. aegypti were equally attracted to wet SGATs, with or without infusion, but not dry ones. These results suggest that male Ae. aegypti within northern Australia are more attracted to active larval sites and equally attracted to dry containers as wet or infused ones. Additionally, female Ae. aegypti are unlikely to enter dry containers. Such findings contribute to our understanding of potentially attractive features for local and released Ae. aegypti throughout the northern Australian urban landscape.

Baseline human gut microbiota profile in healthy people and standard reporting template.

A comprehensive knowledge of the types and ratios of microbes that inhabit the healthy human gut is necessary before any kind of pre-clinical or clinical study can be performed that attempts to alter the microbiome to treat a condition or improve therapy outcome. To address this need we present an innovative scalable comprehensive analysis workflow, a healthy human reference microbiome list and abundance profile (GutFeelingKB), and a novel Fecal Biome Population Report (FecalBiome) with clinical applicability. GutFeelingKB provides a list of 157 organisms (8 phyla, 18 classes, 23 orders, 38 families, 59 genera and 109 species) that forms the baseline biome and therefore can be used as healthy controls for studies related to dysbiosis. This list can be expanded to 863 organisms if closely related proteomes are considered. The incorporation of microbiome science into routine clinical practice necessitates a standard report for comparison of an individual's microbiome to the growing knowledgebase of "normal" microbiome data. The FecalBiome and the underlying technology of GutFeelingKB address this need. The knowledgebase can be useful to regulatory agencies for the assessment of fecal transplant and other microbiome products, as it contains a list of organisms from healthy individuals. In addition to the list of organisms and their abundances, this study also generated a collection of assembled contiguous sequences (contigs) of metagenomics dark matter. In this study, metagenomic dark matter represents sequences that cannot be mapped to any known sequence but can be assembled into contigs of 10,000 nucleotides or higher. These sequences can be used to create primers to study potential novel organisms. All data is freely available from https://hive.biochemistry.gwu.edu/gfkb and NCBI's Short Read Archive.

A chromosome-scale assembly of the major African malaria vector Anopheles funestus.

BACKGROUND: Anopheles funestus is one of the 3 most consequential and widespread vectors of human malaria in tropical Africa. However, the lack of a high-quality reference genome has hindered the association of phenotypic traits with their genetic basis in this important mosquito. FINDINGS: Here we present a new high-quality A. funestus reference genome (AfunF3) assembled using 240× coverage of long-read single-molecule sequencing for contigging, combined with 100× coverage of short-read Hi-C data for chromosome scaffolding. The assembled contigs total 446 Mbp of sequence and contain substantial duplication due to alternative alleles present in the sequenced pool of mosquitos from the FUMOZ colony. Using alignment and depth-of-coverage information, these contigs were deduplicated to a 211 Mbp primary assembly, which is closer to the expected haploid genome size of 250 Mbp. This primary assembly consists of 1,053 contigs organized into 3 chromosome-scale scaffolds with an N50 contig size of 632 kbp and an N50 scaffold size of 93.811 Mbp, representing a 100-fold improvement in continuity versus the current reference assembly, AfunF1. CONCLUSION: This highly contiguous and complete A. funestus reference genome assembly will serve as an improved basis for future studies of genomic variation and organization in this important disease vector.

Chronic adolescent stress sex-specifically alters central and peripheral neuro-immune reactivity in rats.

Adversity during development is a reliable predictor of psychiatric disorders such as depression and anxiety which are increasingly recognized to have an immune component. We have previously demonstrated that chronic adolescent stress (CAS) in rats leads to depressive-like behavior in adulthood along with long-lasting changes to the hypothalamic-pituitary-adrenal axis and pro-inflammatory cytokine induction in the hippocampus. However, the mechanisms by which CAS promotes hippocampal inflammation are not yet defined. Here we tested the hypothesis that a history of CAS exaggerates induction of the pro-inflammatory NFκB pathway in the adult rat hippocampus without compromising the peripheral immune response. We also assessed potential sex differences because it is unclear whether females, who are twice as likely to suffer from mood disorders as males, are disproportionally affected by stress-primed inflammation. Male and female adolescent rats underwent a CAS paradigm or received no stress. Six weeks following the last stressor, all rats received a single systemic injection of either lipopolysaccharide or vehicle to unmask possible immune-priming effects of CAS. An NFκB signaling PCR array demonstrated that CAS exaggerated the expression of NFκB-related genes in the hippocampus of both males and females. Interestingly, targeted qPCR demonstrated that CAS potentiated the induction of hippocampal IL1B and REL mRNA in female rats only, suggesting that some immune effects of CAS are indeed sex-specific. In contrast to the hippocampal findings, indices of peripheral inflammation such as NFκB activity in the spleen, plasma IL-1ß, IL-6, TNF-α, and corticosterone were not impacted by CAS in female rats. Despite showing no pro-inflammatory changes to hippocampal mRNA, male CAS rats displayed lower plasma corticosterone response to LPS at 2 h after injection followed by an exaggerated plasma IL-1ß response at 4 h. This potentially blunted corticosterone response coupled with excessive innate immune signaling in the periphery is consistent with possible glucocorticoid resistance in males. In contrast, the effects of CAS manifested as excessive hippocampal immune reactivity in females. We conclude that while a history of exposure to chronic adolescent stress enhances adult immune reactivity in both males and females, the mechanism and manifestation of such alterations are sex-specific.

Mosquito biology. Evolution of sexual traits influencing vectorial capacity in anopheline mosquitoes.

The availability of genome sequences from 16 anopheline species provides unprecedented opportunities to study the evolution of reproductive traits relevant for malaria transmission. In Anopheles gambiae, a likely candidate for sexual selection is male 20-hydroxyecdysone (20E). Sexual transfer of this steroid hormone as part of a mating plug dramatically changes female physiological processes intimately tied to vectorial capacity. By combining phenotypic studies with ancestral state reconstructions and phylogenetic analyses, we show that mating plug transfer and male 20E synthesis are both derived characters that have coevolved in anophelines, driving the adaptation of a female 20E-interacting protein that promotes oogenesis via mechanisms also favoring Plasmodium survival. Our data reveal coevolutionary dynamics of reproductive traits between the sexes likely to have shaped the ability of anophelines to transmit malaria.

A cohort study of the effectiveness of insecticide-treated bed nets to prevent malaria in an area of moderate pyrethroid resistance, Malawi.

BACKGROUND: Insecticide-treated bed nets (ITNs) are the cornerstone of malaria control in sub-Saharan Africa but their effectiveness may be compromised by the spread of pyrethroid resistance among malaria vectors. The objective of this investigation was to assess the effectiveness of ITNs to prevent malaria in an area of Malawi with moderate pyrethroid resistance. METHODS: One deltamethrin ITN was distributed in the study area for every two individuals in each household plus one extra ITN for households with an odd number of residents. A fixed cohort of 1,199 children aged six to 59 months was seen monthly for one year and at sick visits to measure malaria infection and use of ITNs. Insecticide resistance among malaria vectors was measured. The effect of ITN use on malaria incidence was assessed, adjusting for potential confounders using generalized estimating equations accounting for repeated measures. RESULTS: There were 1,909 infections with Plasmodium falciparum over 905 person-years at risk (PYAR), resulting in an observed incidence of 2.1 infections per person-year (iPPY). ITNs were used during 97% of the PYAR. The main vector was Anopheles funestus: mortality in WHO tube assays after exposure to 0.05% deltamethrin was 38% (95% confidence interval (CI) 29-47), and resistance was due to elevated oxidase enzymes. After adjusting for potential confounders, the incidence of malaria infection among ITN users was 1.7 iPPY (95% CI 1.5-2.1) and among non-bed net users was 2.6 iPPY (95% CI 2.0-3.3). Use of ITNs reduced the incidence of malaria infection by 30% (rate ratio 0.7; 95% CI, 0.5-0.8) compared to no bed nets. CONCLUSION: ITNs significantly reduced the incidence of malaria infection in children in an area with moderate levels of pyrethroid resistance and considerable malaria transmission. This is the first study to show that ITNs provide protection in areas where pyrethroid-resistant An. funestus is the major malaria vector. Malaria control programmes should continue to distribute and promote ITNs in areas with low to moderate pyrethroid resistance; however, insecticide resistance may intensify further and it is not known whether ITNs will remain effective at higher levels of resistance. There is an urgent need to identify or develop new insecticides and technologies to limit the vulnerability of ITNs to insecticide resistance.

Rehabilitation of Combat-Related Injuries in the Veterans Administration: A Web of Support.

The Department of Veterans Affairs provides acute, subacute, and continuing rehabilitation for veterans using a hub-and-spoke system of hospitals and outpatient facilities. Using traumatic brain injury as an example, this commentary illustrates how this system provides interdisciplinary rehabilitative care to veterans throughout North Carolina.

The evolution of the Anopheles 16 genomes project.

We report the imminent completion of a set of reference genome assemblies for 16 species of Anopheles mosquitoes. In addition to providing a generally useful resource for comparative genomic analyses, these genome sequences will greatly facilitate exploration of the capacity exhibited by some Anopheline mosquito species to serve as vectors for malaria parasites. A community analysis project will commence soon to perform a thorough comparative genomic investigation of these newly sequenced genomes. Completion of this project via the use of short next-generation sequence reads required innovation in both the bioinformatic and laboratory realms, and the resulting knowledge gained could prove useful for genome sequencing projects targeting other unconventional genomes.

Up-regulation of miR-182 expression after epigenetic modulation of human melanoma cells.

PURPOSE: We sought to investigate the epigenetic regulation of microRNAs (miRNAs) in melanoma. METHODS: We treated two highly metastatic human melanoma cell lines, C8161.9 and WM266-4, with the demethylating agents DAC (5-aza-2'-deoxycytidine) and trichostatin A. Locked nucleic acid-based miRNA expression profiling was utilized to examine the differential expression of miRNAs before and after treatment. RESULTS: We found that miR-182, a miRNA with oncogenic properties, was significantly up-regulated in human melanoma cells after epigenetic modulation. Genome sequence analysis revealed the presence of a prominent CpG island 8-10 kb upstream of mature miR-182. Methylation analysis showed that this genomic region was exclusively methylated in melanoma cells but not in human melanocytes, skin, or peripheral blood mononuclear cells. DISCUSSION: These results indicate that an epigenetic mechanism is likely involved in modulating the expression level of miR-182 in melanoma, and increased expression of oncogenic-like miR-182 could be a concern for melanoma patients after epigenetic therapy.

Aquaporins mediate the chemoresistance of human melanoma cells to arsenite.

The integral membrane channel protein aquaporin (AQP) is aberrantly expressed with oncogenic characteristics in various human cancers. In this study, we analyzed the expression pattern of all subtypes of AQPs, and found that 8 out of 13 AQPs expressed in melanoma cells. To understand the role of aberrant expression of AQP in this disease, we over-expressed AQP3 and AQP9 in human melanoma WM266.4 cells and found that both AQPs significantly increased the chemoresistance of WM266.4 cells to arsenite. Functional studies showed that AQP3 and AQP9 can inhibit cell apoptosis induced by arsenite through down-regulating p53 and up-regulating Bcl-2 and XIAP. Our data suggest the implication of APQ in melanoma progression and that the over-expression of AQP3 and AQP9 contributes to the chemoresistance of melanoma to arsenite.

Overexpression of the progestagen-associated endometrial protein gene is associated with microphthalmia-associated transcription factor in human melanoma.

BACKGROUND: We recently reported that the progestagen-associated endometrial protein (PAEP) gene is overexpressed and promotes tumor proliferation and metastasis in human melanoma. METHODS: To identify the molecules that regulate its expression and oncogenic properties, we analyzed the gene microarray profiling of melanoma samples of serial clinical stage. RESULTS: We found that the expression profile of the PAEP gene parallels that of microphthalmia-associated transcription factor (MITF, r â€Š=  0.86), a master regulator of melanocyte development and melanoma progression. This parallelism was further confirmed with semiquantitative reverse transcriptase polymerase chain reaction analysis of melanoma-derived daughter cells. Transfection of melanoma cells with MITF small interfering RNA (siRNA) specifically diminishes PAEP gene expression, whereas PAEP siRNA transfection has no effect on MITF. Furthermore, knockdown of either the MITF or PAEP gene reveals a significant inhibition of tumor cell migration. CONCLUSIONS: Our data indicate that PAEP expression is regulated in part by MITF and may thus play a role in MITF-mediated cell migration in human melanoma.

Osteoporosis treatment following hip fracture: how rates vary by service.

OBJECTIVES: Osteoporosis is a prevalent condition among older people. It is often undiagnosed until patients suffer fragility fractures. Previous studies have shown low rates of initiating osteoporosis treatment during the acute hip fracture hospitalization. It is not clear if this varies by the treating service. We compared the rates of instituting osteoporosis treatment during the acute hospitalization for fragility hip fractures. METHODS: Rates of initiating treatment among previously untreated patients were compared between the orthopedic, medicine, and rehabilitation services using retrospective cross-sectional chart review at an academic medical center. Between January 2005 and August 2008, 191 patients admitted with a fragility hip fracture survived to be discharged from the hospital. RESULTS: There were 67 (35%) patients who were started on some form of osteoporosis treatment during their acute hospital stay. Factors statistically associated with starting treatment included having a discharge diagnosis of osteoporosis (P < 0.0001) and treating service (P < 0.0001). Orthopedics was the least likely of the 3 treating services to initiate treatment, while medicine was the most likely. CONCLUSIONS: Overall rates of osteoporosis treatment initiation were low at 35% of the 191 patients' records surveyed. Efforts to increase adherence during the acute hospital stay should be explored. A promising intervention includes instituting an osteoporosis consultative service to improve the likelihood of starting osteoporosis treatment post fragility fracture.