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Item Characterization of a broad-based mosquito yeast interfering RNA larvicide with a conserved target site in mosquito semaphorin-1a genes(Springer Nature, 2019-05-22) Mysore, Keshava; Li, Ping; Wang, Chien-Wei; Hapairai, Limb K.; Scheel, Nicholas D.; Realey, Jacob S.; Sun, Longhua; Severson, David W.; Wei, Na; Duman-Scheel, Molly; Medical and Molecular Genetics, School of MedicineBACKGROUND: RNA interference (RNAi), which has facilitated functional characterization of mosquito neural development genes such as the axon guidance regulator semaphorin-1a (sema1a), could one day be applied as a new means of vector control. Saccharomyces cerevisiae (baker's yeast) may represent an effective interfering RNA expression system that could be used directly for delivery of RNA pesticides to mosquito larvae. Here we describe characterization of a yeast larvicide developed through bioengineering of S. cerevisiae to express a short hairpin RNA (shRNA) targeting a conserved site in mosquito sema1a genes. RESULTS: Experiments conducted on Aedes aegypti larvae demonstrated that the yeast larvicide effectively silences sema1a expression, generates severe neural defects, and induces high levels of larval mortality in laboratory, simulated-field, and semi-field experiments. The larvicide was also found to induce high levels of Aedes albopictus, Anopheles gambiae and Culex quinquefasciatus mortality. CONCLUSIONS: The results of these studies indicate that use of yeast interfering RNA larvicides targeting mosquito sema1a genes may represent a new biorational tool for mosquito control.Item GIS-Enhanced Survey of Potential Aedes aegypti and Aedes albopictus Artificial Oviposition Containers Distributed across Communities in Trinidad, West Indies(MDPI, 2024-10-08) Hapairai, Limb K.; Seeramsingh, Roshan; James, Lester D.; Feng, Rachel S.; Nandram, Naresh; Mohammed, Azad; Duman-Scheel, Molly; Severson, David W.; Medical and Molecular Genetics, School of MedicineDengue and other arboviruses remain a global threat, and enhanced efforts to control the mosquitoes that transmit them are urgently needed. A survey of potential manmade Aedes aegypti (L.) and Aedes albopictus (Skuse) oviposition containers was performed in four communities near the end of the typical dry season in 2018 in Trinidad, West Indies. The purpose was to conduct individual premise surveys and use GIS mapping to visualize premises within communities that had Aedes-positive containers, as this information could be used for the prioritization of mosquito control efforts in potential high risk areas as the wet season progressed. Accessible premises were surveyed following standard inspection protocols used by the Insect Vector Control Division (IVCD), Ministry of Health (MOH). The results indicated that two of the four locations would be at high risk for arbovirus transmission going into the wet season. The GIS mapping of premises with Aedes-positive containers facilitated the identification of potential hot spots for arbovirus transmission risk within communities that should be prioritized for enhanced monitoring and vector control efforts, emphasizing the need to increase community participation in standard surveys by IVCD.Item Identification of Aedes aegypti cis-regulatory elements that promote gene expression in olfactory receptor neurons of distantly related dipteran insects(BMC, 2018-07-11) Mysore, Keshava; Li, Ping; Duman-Scheel, Molly; Medical and Molecular Genetics, School of MedicineBACKGROUND: Sophisticated tools for manipulation of gene expression in select neurons, including neurons that regulate sexually dimorphic behaviors, are increasingly available for analysis of genetic model organisms. However, we lack comparable genetic tools for analysis of non-model organisms, including Aedes aegypti, a vector mosquito which displays sexually dimorphic behaviors that contribute to pathogen transmission. Formaldehyde-assisted isolation of regulatory elements followed by sequencing (FAIRE-seq) recently facilitated genome-wide discovery of putative A. aegypti cis-regulatory elements (CREs), many of which could be used to manipulate gene expression in mosquito neurons and other tissues. The goal of this investigation was to identify FAIRE DNA elements that promote gene expression in the olfactory system, a tissue of vector importance. RESULTS: Eight A. aegypti CREs that promote gene expression in antennal olfactory receptor neurons (ORNs) were identified in a Drosophila melanogaster transgenic reporter screen. Four CREs identified in the screen were cloned upstream of GAL4 in a transgenic construct that is compatible with transformation of a variety of insect species. These constructs, which contained FAIRE DNA elements associated with the A. aegypti odorant coreceptor (orco), odorant receptor 1 (Or1), odorant receptor 8 (Or8) and fruitless (fru) genes, were used for transformation of A. aegypti. Six A. aegypti strains, including strains displaying transgene expression in all ORNs, subsets of these neurons, or in a sex-specific fashion, were isolated. The CREs drove transgene expression in A. aegypti that corresponded to endogenous gene expression patterns of the orco, Or1, Or8 and fru genes in the mosquito antenna. CRE activity in A. aegypti was found to be comparable to that observed in D. melanogaster reporter assays. CONCLUSIONS: These results provide further evidence that FAIRE-seq, which can be paired with D. melanogaster reporter screening to test FAIRE DNA element activity in select tissues, is a useful method for identification of mosquito cis-regulatory elements. These findings expand the genetic toolkit available for the study of Aedes neurobiology. Moreover, given that the CREs drive comparable olfactory neural expression in both A. aegypti and D. melanogaster, it is likely that they may function similarly in multiple dipteran insects, including other disease vector mosquito species.Item Niche Modeling of Dengue Fever Using Remotely Sensed Environmental Factors and Boosted Regression Trees(MDPI, 2017-04) Ashby, Jeffrey; Moreno-Madriñán, Max J.; Yiannoutsos, Constantin T.; Stanforth, Austin; Environmental Health Science, School of Public HealthDengue fever (DF), a vector-borne flavivirus, is endemic to the tropical countries of the world with nearly 400 million people becoming infected each year and roughly one-third of the world’s population living in areas of risk. The main vector for DF is the Aedes aegypti mosquito, which is also the same vector of yellow fever, chikungunya, and Zika viruses. To gain an understanding of the spatial aspects that can affect the epidemiological processes across the disease’s geographical range, and the spatial interactions involved, we created and compared Bernoulli and Poisson family Boosted Regression Tree (BRT) models to quantify the overall annual risk of DF incidence by municipality, using the Magdalena River watershed of Colombia as a study site during the time period between 2012 and 2014. A wide range of environmental conditions make this site ideal to develop models that, with minor adjustments, could be applied in many other geographical areas. Our results show that these BRT methods can be successfully used to identify areas at risk and presents great potential for implementation in surveillance programs.Item Preparation and Use of a Yeast shRNA Delivery System for Gene Silencing in Mosquito Larvae(Humana Press, 2019) Mysore, Keshava; Hapairai, Limb K.; Wei, Na; Realey, Jacob S.; Scheel, Nicholas D.; Severson, David W.; Duman-Scheel, Molly; Medical and Molecular Genetics, School of MedicineThe mosquito genome projects facilitated research in new facets of mosquito biology, including functional genetic studies in the dengue and Zika virus vector Aedes aegypti and the primary African malaria vector Anopheles gambiae. RNA interference (RNAi) has facilitated gene silencing experiments in both of these disease vector mosquito species and could one day be applied as a new method of vector control. Here, we describe a procedure for the genetic engineering of Saccharomyces cerevisiae (baker’s yeast) that express short hairpin RNA (shRNA) corresponding to mosquito target genes of interest. Following cultivation, which facilitates inexpensive propagation of shRNA, the yeast is inactivated and prepared in a ready-to-use dry tablet formulation that is fed to mosquito larvae. Ingestion of the yeast tablets results in effective larval target gene silencing. This technically straightforward and affordable technique may be applicable to a wide variety of mosquito species and potentially to other arthropods that feed on yeast.Item Spatial‐Temporal Assessment of Environmental Factors Related to Dengue Outbreaks in São Paulo, Brazil(Wiley, 2019-08) Ogashawara, I.; Li, L.; Moreno-Madriñán, M. J.; Environmental Health Science, School of Public HealthDengue fever, a disease caused by a vector‐borne flavivirus, is endemic to tropical countries, but its occurrence has been reported worldwide. This study aimed to understand important factors contributing to the spatial and temporal patterns of dengue occurrence in São Paulo, the largest municipality of Brazil. The temporal assessment of dengue occurrence covered the 2011–2016 time period and was based on climatological data, such as the El Niño indices and time series statistical tools such as the continuous wavelet transformation. The spatial assessment used Landsat 8 data for years 2014–2016 to estimate land surface temperature and normalized indices for vegetation, urban areas, and leaf water. Results from a cross correlation for the temporal analysis found a relationship between the sea surface temperature anomalies index and the number of reported dengue cases in São Paulo (r = 0.5) with a lag of +29 (weeks) between the climatic event and the response on the dengue incidence. This relationship, initially nonlinear, became linear after correcting for the lag period. For the spatial assessment, the linear stepwise regression model detected a low relationship between dengue incidence and minimum surface temperature (r = 0.357) and no relationship with other environmental parameters. The poor relationship might be due to confounding effects of socioeconomic factors as these seem to influence the spatial dynamics of dengue incidence. More testing is needed to validate these methods in other locations. Nevertheless, we presented possible tools to be used for the improvement of dengue control programs.Item Zika and Flavivirus Shell Disorder: Virulence and Fetal Morbidity(MDPI, 2019-11-06) Goh, Gerard Kian-Meng; Dunker, A. Keith; Foster, James A.; Uversky, Vladimir N.; Biochemistry and Molecular Biology, School of MedicineZika virus (ZIKV) was first discovered in 1947 in Africa. Since then, sporadic ZIKV infections of humans have been reported in Africa and Asia. For a long time, this virus was mostly unnoticed due to its mild symptoms and low fatality rates. However, during the 2015–2016 epidemic in Central and South America, when millions of people were infected, it was discovered that ZIKV causes microcephaly in the babies of mothers infected during pregnancy. An examination of the M and C proteins of the ZIKV shell using the disorder predictor PONDR VLXT revealed that the M protein contains relatively high disorder levels comparable only to those of the yellow fever virus (YFV). On the other hand, the disorder levels in the C protein are relatively low, which can account for the low case fatality rate (CFR) of this virus in contrast to the more virulent YFV, which is characterized by high disorder in its C protein. A larger variation was found in the percentage of intrinsic disorder (PID) in the C protein of various ZIKV strains. Strains of African lineage are characterized by higher PIDs. Using both in vivo and in vitro experiments, laboratories have also previously shown that strains of African origin have a greater potential to inflict higher fetal morbidity than do strains of Asian lineage, with dengue-2 virus (DENV-2) having the least potential. Strong correlations were found between the potential to inflict fetal morbidity and shell disorder in ZIKV (r2 = 0.9) and DENV-2 (DENV-2 + ZIKV, r2 = 0.8). A strong correlation between CFR and PID was also observed when ZIKV was included in an analysis of sets of shell proteins from a variety of flaviviruses (r2 = 0.8). These observations have potential implications for antiviral vaccine development and for the design of cancer therapeutics in terms of developing therapeutic viruses that penetrate hard-to-reach organs.