Impact of visual features on capture of Aedes aegypti with host decoy traps (HDT).

The host decoy trap (HDT) is a surveillance trap that presents a combination of heat, visual and odour stimuli to attract bloodmeal-seeking mosquitoes. Here we employed a semi-field study to demonstrate the role of the visual attributes present on the HDT on the effectiveness of Aedes aegypti capture. Our results show that the HDT is an effective means of capturing Ae. aegypti mosquitoes in semi-field conditions, with a per trial capture rate of up to 69% across four visually distinct HDTs. The solid black coloured HDT captured more mosquitoes than HDTs with black-white stripes, black-white checkerboard patches or solid white colour by a factor of 1.9, 1.7 and 1.5, respectively. In all cases, mosquito capture was not evenly distributed on the HDT surface, with captures on the HDT's outer downwind half, away from the odour delivery, exceeding captures on the inner upwind half. We conclude that the solid black surface of the original HDT design is more effective than the other surfaces (white or black/white patterns) for the capture of Ae. aegypti. Our results demonstrate that mosquito attraction to the thermal and odorant cues of the HDT is modulated by visual information.

The characterization of the Phlebotomus papatasi transcriptome.

As important vectors of human disease, phlebotomine sand flies are of global significance to human health, transmitting several emerging and re-emerging infectious diseases. The most devastating of the sand fly transmitted infections are the leishmaniases, causing significant mortality and morbidity in both the Old and New World. Here we present the first global transcriptome analysis of the Old World vector of cutaneous leishmaniasis, Phlebotomus papatasi (Scopoli) and compare this transcriptome to that of the New World vector of visceral leishmaniasis, Lutzomyia longipalpis. A normalized cDNA library was constructed using pooled mRNA from Phlebotomus papatasi larvae, pupae, adult males and females fed sugar, blood, or blood infected with Leishmania major. A total of 47 615 generated sequences was cleaned and assembled into 17 120 unique transcripts. Of the assembled sequences, 50% (8837 sequences) were classified using Gene Ontology (GO) terms. This collection of transcripts is comprehensive, as demonstrated by the high number of different GO categories. An in-depth analysis revealed 245 sequences with putative homology to proteins involved in blood and sugar digestion, immune response and peritrophic matrix formation. Twelve of the novel genes, including one trypsin, two peptidoglycan recognition proteins (PGRP) and nine chymotrypsins, have a higher expression level during larval stages. Two novel chymotrypsins and one novel PGRP are abundantly expressed upon blood feeding. This study will greatly improve the available genomic resources for P. papatasi and will provide essential information for annotation of the full genome.

Positional cloning of rp2 QTL associates the P450 genes CYP6Z1, CYP6Z3 and CYP6M7 with pyrethroid resistance in the malaria vector Anopheles funestus.

Pyrethroid resistance in Anopheles funestus is threatening malaria control in Africa. Elucidation of underlying resistance mechanisms is crucial to improve the success of future control programs. A positional cloning approach was used to identify genes conferring resistance in the uncharacterised rp2 quantitative trait locus (QTL) previously detected in this vector using F6 advanced intercross lines (AIL). A 113 kb BAC clone spanning rp2 was identified and sequenced revealing a cluster of 15 P450 genes and one salivary protein gene (SG7-2). Contrary to A. gambiae, AfCYP6M1 is triplicated in A. funestus, while AgCYP6Z2 orthologue is absent. Five hundred and sixty-five new single nucleotide polymorphisms (SNPs) were identified for genetic mapping from rp2 P450s and other genes revealing high genetic polymorphisms with one SNP every 36 bp. A significant genotype/phenotype association was detected for rp2 P450s but not for a cluster of cuticular protein genes previously associated with resistance in A. gambiae. QTL mapping using F6 AIL confirms the rp2 QTL with an increase logarithm of odds score of 5. Multiplex gene expression profiling of 15 P450s and other genes around rp2 followed by individual validation using qRT-PCR indicated a significant overexpression in the resistant FUMOZ-R strain of the P450s AfCYP6Z1, AfCYP6Z3, AfCYP6M7 and the glutathione-s-transferase GSTe2 with respective fold change of 11.2, 6.3, 5.5 and 2.8. Polymorphisms analysis of AfCYP6Z1 and AfCYP6Z3 identified amino acid changes potentially associated with resistance further indicating that these genes are controlling the pyrethroid resistance explained by the rp2 QTL. The characterisation of this rp2 QTL significantly improves our understanding of resistance mechanisms in A. funestus.

Transcription profiling associated with life cycle of Anopheles gambiae.

Complex biological events occur during the developmental process of the mosquito Anopheles gambiae (Giles). Using cDNA expression microarrays, the expression patterns of 13,440 clones representing 8,664 unique transcripts were revealed from six different developmental stages: early larvae (late third instar/early fourth instar), late larvae (late fourth instar), early pupae (< 30 min after pupation), late pupae (after tanning), and adult female and male mosquitoes (24 h postemergence). After microarray analysis, 560 unique transcripts were identified to show at least a fourfold up- or down-regulation in at least one developmental stage. Based on the expression patterns, these gene products were clustered into 13 groups. In total, eight genes were analyzed by quantitative real-time polymerase chain reaction to validate microarray results. Among 560 unique transcripts, 446 contigs were assigned to respective genes from the An. gambiae genome. The expression patterns and annotations of the genes in the 13 groups are discussed in the context of development including metabolism, transport, protein synthesis and degradation, cellular processes, cellular communication, intra- or extra-cellular architecture maintenance, response to stress or immune-related defense, and spermatogenesis.

Introgression as a likely cause of mtDNA paraphyly in two allopatric skippers (Lepidoptera: Hesperiidae).

Gene transfer between species during interspecific hybridization is a widely accepted reality in plants but is considered a relatively rare phenomenon among animals. Here we describe a unique case of mitochondrial DNA (mtDNA) paraphyly in the skipper genus, Erynnis, that involves well-diverged allopatric species. Using molecular evidence from both mitochondrial and nuclear genomes, we found high levels of intraspecific divergence in the mitochondrial genome within E. propertius (over 4% pair-wise sequence divergence) but no such differentiation in the nuclear genome. Sequence comparisons with related Erynnis suggest that past, but recent and infrequent introgression between E. propertius and E. horatius is the most reasonable explanation for the observed pattern of mtDNA paraphyly. This example of putative introgression highlights the complexity of mtDNA evolution and suggests that similar processes could be operating in other taxa that have not been extensively sampled. Our observations reinforce the importance of involving multiple genes with different modes of inheritance in the analysis of population history of congeneric taxa.

SINE insertion polymorphism on the X chromosome differentiates Anopheles gambiae molecular forms.

Polymorphic SINE insertions can be useful markers for assessing population structure and differentiation. Maque is a family of SINE elements which, based on bioinformatic analysis, was suggested to have been active recently in Anopheles gambiae, the major vector of malaria. Here, we report the development of polymorphic Maque insertions as population genetic markers in A. gambiae, and the use of these markers to better characterize divergence on the X chromosome between A. gambiae M and S molecular forms in populations from Burkina Faso and Mali. Our data are consistent with the recent activity of Maque. Phylogenetic analysis suggests that at least two recently active lineages may have a role in mediating genome evolution. We found differences in element insertion frequency and sequence between the M and S populations analysed. Significant differentiation was observed between these two groups across a 6 Mb region at the proximal (centromeric) end of the X chromosome. Locus-specific F(ST) values ranged from 0.14 to 1.00 in this region, yet were not significantly different from zero in more distal locations on the X chromosome; the trend was consistent in populations from both geographical locales suggesting that differentiation is not due to local adaptation. Strong differentiation between M and S at the proximal end of the X chromosome, but not outside this region, suggests the action of selection counteracting limited gene flow between these taxa and supports their characterization as incipient species.

Germ line transformation of the yellow fever mosquito, Aedes aegypti, mediated by transpositional insertion of a piggyBac vector.

Mosquito-vectored diseases such as yellow fever and dengue fever continue to have a substantial impact on human populations world-wide. Novel strategies for control of these mosquito vectored diseases can arise through the development of reliable systems for genetic manipulation of the insect vector. A piggyBac vector marked with the Drosophila melanogaster cinnabar (cn) gene was used to transform the white-eyed khw strain of Aedes aegypti. Microinjection of preblastoderm embryos resulted in four families of cinnabar transformed insects. An overall transformation frequency of 4%, with a range of 0% to as high as 13% for individual experiments, was achieved when using a heat-shock induced transposase providing helper plasmid. Southern hybridizations indicated multiple insertion events in three of four transgenic lines, while the presence of duplicated target TTAA sites at either ends of individual insertions confirmed characteristic piggyBac transposition events in these three transgenic lines. The transgenic phenotype has remained stable for more than twenty generations. The transformations effected using the piggyBac element establish the potential of this element as a germ-line transformation vector for Aedine mosquitoes.

[Fortification of fluid milk for the prevention and treatment of iron deficiency anemia in children under 4 years of age]. / Fortificação do leite fluido na prevenção e tratamento da anemia carencial ferropriva em criancas menores de 4 anos.

The effectiveness of the use of chelate amino acid iron fortified fluid milk in the treatment of iron deficiency in children under four years of age was studied. The 269 children included in this trial received 1 liter/day of fluid milk fortified with 3 mg of chelate amino acid iron and were evaluated at six monthly intervals. At the beginning of the study 62.3% of the children presented anemia. After 6 months, this percentage had decreased to 41.8% and at the end of one year to 26.4%. The greatest decreases occurred in the groups comprising the subjects who were of 12 to 23 months of age and those under one year of age. Among the children who presented initial hemoglobin levels under 9.5 g/dl, 59.3% were free of anemia after one year of follow-up. Of those presenting initial hemoglobin levels between 9.5 and 10.9 g/dl, 66.7% recovered from their anemia. There was also greater hematological improvement in the children that ingested over 750 ml/day of fortified milk in those families that did not share the supply of supplement among their other members and in those families that had only one child under five years of age. These findings led to the conclusion that the fortification of fluid milk is a viable and effective method for the treatment of iron deficiency in pre-school children.

[The effect of the use of milk fortified with iron and vitamin C on hemoglobin levels and nutritional status of children under 2]. / Efeito do uso de leite fortificado com ferro e vitamina C sobre os niveis de hemoglobina e condicao nutricional de criancas menores de 2 anos.

The impact of the use of fortified powdered whole milk (9 mg of iron and 65 mg of vitamin C/100 g of milk) on the hemoglobin levels of children under 2 years of age was evaluated, over a period of 6 months, in 107 children enrolled in municipal Day Care Centers (DCC) and in 228 seen at a Basic Health Care Unit (BHCU). Before the beginning of the intervention, 66.4% of the children in the DCC and 72.8% of those seen at the BHCU had hemoglobin levels under 11.0 g/dl. After 6 months of fortified milk intake, these percentages fell to 20.6% and 18.0% respectively. The average hemoglobin before the intervention was 10.3 g/dl in the DCC and 10.5 in the BHCU. After 6 months these increased to 11.6 g/dl in the populations studied. Concerning the nutritional condition, evaluated according to Gomez's criteria, 57% of the DCC children presented an improvement, 41.1% showed changes and only 1.9% became worse. In the BHCU, 11.4% presented better condition, 70.6% remained the same and 18% worsened, which demonstrated differences in response regarding improvement of nutritional condition, when fortified milk was used in closed and open environments. The authors conclude that the utilization of enriched foods is an excellent alternative in the treatment of iron deficiency in populations of children under 2 years of age.

PIP: The impact of the use of fortified powdered whole milk (9 mg of iron and 65 mg of vitamin C per 100 g of milk) on the hemoglobin levels of children under 2 years of age was evaluated, over a period of 6 months, in 107 children enrolled in municipal day care centers (DCC) and in 228 seen at a Basic Health Care Unit (BHCU). Before the beginning of the intervention, 66.4% of the children in the DCC and 72.8% of those seen at the BHCU had hemoglobin levels under 11.0 g/dl. After 6 months of fortified milk intake, these percentages fell to 20.6% and 18.0%, respectively. The average hemoglobin level before the intervention was 10.3 g/dl in the DCC and 10.5 g/dl in the BHCU. After 6 months these increased to 11.6 g/dl in the populations studied. Concerning the nutritional condition, evaluated according to Gomez's criteria, 57% of the DCC children showed improvement, 41.1% showed no change, and only 1.9% grew worse. In the BHCU, 11.4% showed improvement, 70.6% remained the same, and 18% worsened, which demonstrated differences in response regarding improvement of nutritional condition when fortified milk was used in closed and open environments. The authors conclude that the utilization of enriched foods is an excellent alternative in the treatment of iron deficiency in populations of children under 2 years of age. (author's)