DNA degradation of Anopheles darlingi collected at high relative humidity and preserved in isopropanol

Anopheles darlingi mosquitoes, exposed to variable relative humidity (RH) in the field, were preserved during transportation to the laboratory using two different methods: 100% isopropanol at ambient temperature or frozen in liquid nitrogen or dry ice. The DNA isolated from samples collected at RH greater than 91% and preserved in isopropanol was degraded, while DNA isolated from insects kept in liquid nitrogen or dry ice maintained its integrity when collected under conditions of up to 95% RH.

Population genetic structure of the dengue mosquito Aedes aegypti in Venezuela.

The mosquito Aedes aegypti is the main vector of dengue in Venezuela. The genetic structure of this vector was investigated in 24 samples collected from eight geographic regions separated by up to 1160 km. We examined the distribution of a 359-basepair region of the NADH dehydrogenase subunit 4 mitochondrial gene among 1144 Ae. aegypti from eight collections. This gene was amplified by the polymerase chain reaction and tested for variation using single strand conformation polymorphism analysis. Seven haplotypes were detected throughout Venezuela and these were sorted into two clades. Significant differentiation was detected among collections and these were genetically isolated by distance.

Population genetic structure of the dengue mosquito Aedes aegypti in Venezuela

The mosquito Aedes aegypti is the main vector of dengue in Venezuela. The genetic structure of this vector was investigated in 24 samples collected from eight geographic regions separated by up to 1160 km. We examined the distribution of a 359-basepair region of the NADH dehydrogenase subunit 4 mitochondrial gene among 1144 Ae. aegypti from eight collections. This gene was amplified by the polymerase chain reaction and tested for variation using single strand conformation polymorphism analysis. Seven haplotypes were detected throughout Venezuela and these were sorted into two clades. Significant differentiation was detected among collections and these were genetically isolated by distance.

Genetic diversity of merozoite surface protein-1 gene of Plasmodium vivax isolates in mining villages of Venezuela (Bolivar State).

The merozoite surface protein-1 gene of Plasmodium vivax is highly polymorphic and so, currently used in epidemiological studies of P. vivax malaria. We sequenced the variable block 5 of the gene from 39 Venezuelan isolates, 18 of which were co-infected with Plasmodium falciparum. We observed a limited variability with 34 isolates belonging to the type Salvador I, none Belem type and only five recombinants. Among the recombinants, only two types of sequences were observed with, respectively, 18 and 21 poly-Q residues. Nucleotide substitutions explained the major differences of the 11 patterns observed. We could evidence neither specific MSP-1 genotype associated with co-infected samples, nor peculiar MSP-1 genotype distribution inside the investigated areas. In comparison with other low endemic regions in the world, our sampling has a lower genetic diversity, which could be mainly explained by the lack of Belem type. In fact, the variable repeats of poly-Q residues involved in the polymorphism of Belem type and recombinant isolates are responsible for a great part of variability observed in MSP-1 block 5.

Detection of dengue viruses in field-caught Aedes aegypti (Diptera: Culicidae) in Maracay, Aragua state, Venezuela by type-specific polymerase chain reaction.

Virological surveillance of dengue viruses in Aedes aegypti populations constitutes a powerful tool for early prediction of dengue outbreaks. We have standardized a protocol for viral RNA extraction from individual and pools of mosquitoes that permits a sensitive detection of dengue virus without RNA degradation or PCR inhibition when we apply a semi-nested RT-PCR. The limit of detection for each dengue serotype was 0.1 PFU. In a prospective field study conducted from November 2000 to December 2001, adult female A. aegypti mosquitoes from several municipalities with high dengue transmission in Maracay, Aragua State, Venezuela were collected and screened for dengue viruses using RT-PCR. We analyzed a total of 296 A. aegypti pools (1,632 mosquitoes); of these, 154 pools (469 mosquitoes) were collected from houses with persons with clinical diagnosis of dengue (dengue houses), and 142 pools (1,163 mosquitoes) from adjacent residences (neighbour houses). From the dengue houses, eight mosquito pools (5.2%) were positive for DENV-1 (0.7%), DENV-3 (3.2%) and DENV-4 (1.3%) viruses. From the neighbour houses, 18 mosquito pools (12.7%) were positive for DENV-3 (12%) and DENV-4 (0.7%) viruses. From these 26 RT-PCR positive mosquito pools (containing 1-25 mosquitoes each), 22 pools (84.6%) were positive for DENV-3. The most prevalent serotype in the 2001 dengue outbreak was also DENV-3. The minimum infection rate in both A. aegypti collections, from dengue houses and neighbour houses was 17 and 15 per 1,000, respectively. The relevance of these results for dengue surveillance is discussed.