Aedes aegypti is the primary vector of yellow fever (YF) and dengue fever (DF) flaviviruses worldwide. In this review we focus on past and present research on genetic components and environmental factors in Aedes aegypti that appear to control flavivirus transmission. We review genetic relationships among Ae. aegypti populations throughout the world and discuss how variation in vector competence is correlated with overall genetic differences among populations. We describe current research into how genetic and environmental factors jointly affect distribution of vector competence in natural populations. Based on this information, we propose a population genetic model for vector competence and discuss our recent progress in testing this model. We end with a discussion of approaches being taken to identify the genes that may control flavivirus susceptibility in Ae. aegypti.

Abstract. Aedes aegypti from 24 collections in Mexico and the United States were challenged orally with dengue 2 virus JAM1409 (DEN-2 JAM1409). The vector competence (VC) of the populations ranged from 24% to 83%. Mosquito populations from the Yucatan exhibited greater VC than those from other areas of Mexico. The presence or absence of a midgut infection barrier (MIB) and a midgut escape barrier (MEB) was determined for mosquitoes in each population. The percentage of mosquitoes exhibiting an MIB ranged from 14% to 59%, and those exhibiting an MEB ranged from 4% to 43% in the collections. The MIB and MEB were not completely independent as determined by regression analysis. Midgut infection rates were dose dependent.

Abstract. A population genetic analysis of Aedes aegypti was conducted among 38 collections from throughout coastal regions of Mexico. Multiple collections were made within 5 cities to examine local patterns of gene flow. Single-strand conformation polymorphism analysis was used to screen for variation in a 387-bp region of the Nicotinamide Adenine Dinucleotide Dehydrogenase subunit 4 mitochondrial gene (ND4) and 25 haplotypes were detected. Northeastern Mexico collections were genetically differentiated from and had lower genetic diversity than Yucatan and Pacific coastal collections. Yucatan and Pacific collections were genetically homogeneous. Regression analysis of geographic distances and F ST values indicated that collections were genetically isolated by distance in the Pacific and the Yucatan, but not among collections in the northeast. Free gene flow occurred among all collections within 130 km of one another in the northeast and within 180 km in the Yucatan. F ST values were never large among Pacific collections, suggesting extensive gene flow along the Pacific coast.

Background Aedes aegypti is the primary global vector to humans of yellow fever and dengue flaviviruses. Over the past 50 years, many population genetic studies have documented large genetic differences among global populations of this species. These studies initially used morphological polymorphisms, followed later by allozymes, and most recently various molecular genetic markers including microsatellites and mitochondrial markers. In particular, since 2000, fourteen publications and four unpublished datasets have used sequence data from the NADH dehydrogenase subunit 4 mitochondrial gene to compare Ae. aegypti collections and collectively 95 unique mtDNA haplotypes have been found. Phylogenetic analyses in these many studies consistently resolved two clades but no comprehensive study of mtDNA haplotypes have been made in Africa, the continent in which the species originated.Methods and FindingsND4 haplotypes were sequenced in 426 Ae. aegypti s.l. from Senegal, West Africa and Kenya, East Africa. In Senegal 15 and in Kenya 7 new haplotypes were discovered. When added to the 95 published haplotypes and including 6 African Aedes species as outgroups, phylogenetic analyses showed that all but one Senegal haplotype occurred in a basal clade while most East African haplotypes occurred in a second clade arising from the basal clade. Globally distributed haplotypes occurred in both clades demonstrating that populations outside Africa consist of mixtures of mosquitoes from both clades.ConclusionsPopulations of Ae. aegypti outside Africa consist of mosquitoes arising from one of two ancestral clades. One clade is basal and primarily associated with West Africa while the second arises from the first and contains primarily mosquitoes from East Africa

Background: Vector competence refers to the intrinsic permissiveness of an arthropod vector for infection, replication and transmission of a virus. Notwithstanding studies of Quantitative Trait Loci (QTL) that influence the ability of Aedes aegypti midgut (MG) to become infected with dengue virus (DENV), no study to date has been undertaken to identify genetic markers of vector competence. Furthermore, it is known that mosquito populations differ greatly in their susceptibility to flaviviruses. Differences in vector competence may, at least in part, be due to the presence of specific midgut epithelial receptors and their identification would be a significant step forward in understanding the interaction of the virus with the mosquito. The first interaction of DENV with the insect is through proteins in the apical membrane of the midgut epithelium resulting in binding and receptormediated endocytosis of the virus, and this determines cell permissiveness to infection. The susceptibility of mosquitoes to infection may therefore depend on their specific virus receptors. To study this interaction in Ae. aegypti strains that differ in their vector competence for DENV, we investigated the DS3 strain (susceptible to DENV), the IBO-11 strain (refractory to infection) and the membrane escape barrier strain, DMEB, which is infected exclusively in the midgut epithelial cells.

No abstract

During the course of invasive intestinal amebiasis, Entamoeba histolytica actively penetrates the mucosa and submucosa of the large intestine (1-6). Electron microscopy studies of intestinal epithelium of in vivo and in vitro infected tissues in autopsies from humans and experimental animals reveal that the basal lamina and the connective tissue of the submucosa are primarily damaged (3,5,6). Since collagen is a major component of the basal lamina and the extracellular matrix of the intestine (7-9), we studied the presence of collagenolytic activity in E. histolytica when plated on native type I or III human collagen. Our results show that Entamoebae possesses a membrane-bound proteolytic enzyme that digests native type I and type III collagen fibers inside the helix when incubated at neutral pH at 37°C. The collagenase was more active against type I collagen. Materials and MethodsEntamoeba Cultures. E. histolytica HMI:IMSS was cultured in TYI-S-33 medium (10). All the experiments were done using trophozoites harvested during logarithmic phase of growth.Collagens. Collagen types I and III were extracted and purified following the procedure described by Rojkind et al. (11). A human placenta was sliced and incubated with 500 ml of 0.25 M acetic acid containing pepsin (0.25 mg/ml) at 4°C for 24 h. After centrifugation at 16,000 g for 40 min, the clear supernatant was decanted and neutralized to pH 7.0 to deactivate the pepsin. This digestion process was repeated twice, and the three supernatants pooled for purification of collagen types. Type III and I collagens precipitated out of solution by dialysis at 4°C against 0.05 M Tris-HCl buffer containing 1.7 M NaCI and 2.5 M NaC1, respectively. Precipitates were dissolved and dialyzed in 0.25 M acetic acid, lyophilized, and stored at 4°C until used.Collagen Substrates. Collagen in 0.25 M acetic acid (15 ml) was dialyzed for 1 h against 1 liter of 0.05 M Tris-HCl buffer, pH 7.2, containing 0.005 M CaC12, and then the pH of the solution adjusted to 6.9 with 0.1 N NaOH. Reconstituted collagen gels for digestion assays were prepared on Falcon dishes (35 mm Diam, Falcon Labware, Becton, Dickinson & Co., Oxnard, Calif.) with 0.5 ml of collagen type I (10 mg/ml) and incubated at 37°C for 1 h, to allow fibrillar polymerization. The gels were incubated overnight at room temperature under an ultraviolet light source placed 20 em above the dishes. To test cell adhesion, films were prepared as described above, except that 0.1 ml of a 1 mg/ml collagen solution was used per dish, and dried after polymerization at room temperature overnight.Adherence of Amebas to Collagen Films. Films of collagen types I or III were covered with 2 ml of modified TYI-S-33 medium (without serum and L-cysteine), and 1 ml of the same medium containing 1 × 105 trophozoites was added. Cell adherence was evaluated after different incubation times at 37°C. Afterwards, unattached amebas were washed twice in modified TYI-S-33-medium, and adherent amebas were removed after incubation in 0.15 M cold NaCI for I0 ...

Background Aedes aegypti is the main mosquito vector of the four serotypes of dengue virus (DENV). Previous population genetic and vector competence studies have demonstrated substantial genetic structure and major differences in the ability to transmit dengue viruses in Ae. aegypti populations in Mexico.Methodology/Principal FindingsPopulation genetic studies revealed that the intersection of the Neovolcanic axis (NVA) with the Gulf of Mexico coast in the state of Veracruz acts as a discrete barrier to gene flow among Ae. aegypti populations north and south of the NVA. The mosquito populations north and south of the NVA also differed in their vector competence (VC) for dengue serotype 2 virus (DENV2). The average VC rate for Ae. aegypti mosquitoes from populations from north of the NVA was 0.55; in contrast the average VC rate for mosquitoes from populations from south of the NVA was 0.20. Most of this variation was attributable to a midgut infection and escape barriers. In Ae. aegypti north of the NVA 21.5% failed to develop midgut infections and 30.3% of those with an infected midgut failed to develop a disseminated infection. In contrast, south of the NVA 45.2% failed to develop midgut infections and 62.8% of those with an infected midgut failed to develop a disseminated infection.ConclusionsBarriers to gene flow in vector populations may also impact the frequency of genes that condition continuous and epidemiologically relevant traits such as vector competence. Further studies are warranted to determine why the NVA is a barrier to gene flow and to determine whether the differences in vector competence seen north and south of the NVA are stable and epidemiologically significant.