Asymptomatic carriers of Plasmodium spp. as infection source for malaria vector mosquitoes in the Brazilian Amazon

We have described the existence of asymptomatic carriers of Plasmodium vivax and Plasmodium falciparum infections in native Amazon populations. Most of them had low parasitemias, detected only by polymerase chain reaction (PCR). Because they remain symptomless and untreated, we wanted to determine whether they could infect Anopheles darlingi Root, the main Brazilian vector, and act as disease reservoirs. Fifteen adult asymptomatic patients (PCR positive only) were selected, and experimental infections of mosquitoes were performed by direct feeding and by a membrane-feeding system. Seventeen adult symptomatic patients with high parasitemias were used as controls. We found an infection rate in An. darlingi of 1.2% for the asymptomatic carriers and 22% for the symptomatic carriers. Although the asymptomatic group infected mosquitoes at a much lower rate, these patients remain infective longer than treated, symptomatic patients. Also, the prevalence of asymptomatic infections is 4 to 5 times higher than symptomatic infections among natives. These results have implications for the malaria control program in Brazil, which focuses essentially on the treatment of symptomatic patients. © 2005 Entomological Society of America.

The Genome of Anopheles darlingi, the main neotropical malaria vector

Anopheles darlingi is the principal neotropical malaria vector, responsible for more than a million cases of malaria per year on the American continent. Anopheles darlingi diverged from the African and Asian malaria vectors ∼100 million years ago (mya) and successfully adapted to the New World environment. Here we present an annotated reference A. darlingi genome, sequenced from a wild population of males and females collected in the Brazilian Amazon. A total of 10 481 predicted protein-coding genes were annotated, 72% of which have their closest counterpart in Anopheles gambiae and 21% have highest similarity with other mosquito species. In spite of a long period of divergent evolution, conserved gene synteny was observed between A. darlingi and A. gambiae. More than 10 million single nucleotide polymorphisms and short indels with potential use as genetic markers were identified. Transposable elements correspond to 2.3% of the A. darlingi genome. Genes associated with hematophagy, immunity and insecticide resistance, directly involved in vectorhuman and vectorparasite interactions, were identified and discussed. This study represents the first effort to sequence the genome of a neotropical malaria vector, and opens a new window through which we can contemplate the evolutionary history of anopheline mosquitoes. It also provides valuable information that may lead to novel strategies to reduce malaria transmission on the South American continent. The A. darlingi genome is accessible at www.labinfo.lncc.br/index.php/anopheles- darlingi. © 2013 The Author(s).

Seasonal genetic partitioning in the neotropical malaria vector, Anopheles darlingi

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Evidence for temporal population replacement and the signature of ecological adaptation in a major Neotropical malaria vector in Amazonian Peru

The major Neotropical malaria vector, Anopheles darlingi, was reintroduced into the Iquitos, Loreto, Peru area during the early 1990s, where it displaced other anophelines and caused a major malaria epidemic. Since then, case numbers in Loreto have fluctuated, but annual increases have been reported since 2012. The population genetic structure of An. darlingi sampled before and after the introduction of long-lasting insecticidal nets (LLINs) was investigated to test the hypothesis of temporal population change (2006 vs. 2012). Current samples of An. darlingi were used to test the hypothesis of ecological adaptation to human modified (highway) compared with wild (riverine) habitat, linked to forest cover. In total, 693 An. darlingi from nine localities in Loreto, Peru area were genotyped using 13 microsatellite loci. To test the hypothesis of habitat differentiation in An. darlingi biting time patterns, HBR and EIR, four collections of An. darlingi from five localities (two riverine and three highway) were analysed. Analyses of microsatellite loci from seven (2006) and nine settlements (2012-2014) in the Iquitos area detected two distinctive populations with little overlap, although it is unclear whether this population replacement event is associated with LLIN distribution or climate. Within the 2012-2014 population two admixed subpopulations, A and B, were differentiated by habitat, with B significantly overrepresented in highway, and both in near-equal proportions in riverine. Both subpopulations had a signature of expansion and there was moderate genetic differentiation between them. Habitat and forest cover level had significant effects on HBR, such that Plasmodium transmission risk, as measured by EIR, in peridomestic riverine settlements was threefold higher than in peridomestic highway settlements. HBR was directly associated with available host biomass rather than forest cover. A population replacement event occurred between 2006 and 2012-2014, concurrently with LLIN distribution and a moderate El Niño event, and prior to an increase in malaria incidence. The likely drivers of this replacement cannot be determined with current data. The present-day An. darlingi population is composed of two highly admixed subpopulations, which appear to be in an early stage of differentiation, triggered by anthropogenic alterations to local habitat.

Resisting infection by Plasmodium berghei increases the sensitivity of the malaria vector Anopheles gambiae to DDT

BACKGROUND The evolution of insecticide resistance threatens current malaria control methods, which rely heavily on chemical insecticides. The magnitude of the threat will be determined by the phenotypic expression of resistance in those mosquitoes that can transmit malaria. These differ from the majority of the mosquito population in two main ways; they carry sporozoites (the infectious stage of the Plasmodium parasite) and they are relatively old, as they need to survive the development period of the malaria parasite. This study examines the effects of infection by Plasmodium berghei and of mosquito age on the sensitivity to DDT in a DDT-resistant strain of Anopheles gambiae. METHODS DDT-resistant Anopheles gambiae (ZANU) mosquitoes received a blood meal from either a mouse infected with Plasmodium berghei or an uninfected mouse. 10 and 19 days post blood meal the mosquitoes were exposed to 2%, 1% or 0% DDT using WHO test kits. 24 hrs after exposure, mortality and Plasmodium infection status of the mosquitoes were recorded. RESULTS Sensitivity to DDT increased with the mosquitoes' age and was higher in mosquitoes that had fed on Plasmodium-infected mice than in those that had not been exposed to the parasite. The latter effect was mainly due to the high sensitivity of mosquitoes that had fed on an infected mouse but were not themselves infected, while the sensitivity to DDT was only slightly higher in mosquitoes infected by Plasmodium than in those that had fed on an uninfected mouse. CONCLUSIONS The observed pattern indicates a cost of parasite-resistance. It suggests that, in addition to the detrimental effect of insecticide-resistance on control, the continued use of insecticides in a population of insecticide-resistant mosquitoes could select mosquitoes to be more susceptible to Plasmodium infection, thus further decreasing the efficacy of the control.

Genomic structure and ecdysone regulation of the prophenoloxidase 1 gene in the malaria vector Anopheles gambiae

Prophenoloxidase, a melanin-synthesizing enzyme, is considered to be an important arthropod immune protein. In mosquitoes, prophenoloxidase has been shown to be involved in refractory mechanisms against malaria parasites. In our study we used Anopheles gambiae, the most important human malaria vector, to characterize the first arthropod prophenoloxidase gene at the genomic level. The complete nucleotide sequence, including the immediate 5′ flanking sequence (−855 bp) of the prophenoloxidase 1 gene, was determined. The gene spans 10 kb and is composed of five exons and four introns coding for a 2.5-kb mRNA. In the 5′ flanking sequence, we found several putative regulatory motifs, two of which were identified as ecdysteroid regulatory elements. Electrophoretic mobility gel-shift assays and supershift assays demonstrated that the Aedes aegypti ecdysone receptor/Ultraspiracle nuclear receptor complex, and, seemingly, the endogenous Anopheles gambiae nuclear receptor complex, was able to bind one of the ecdysteroid response elements. Furthermore, 20-hydroxyecdysone stimulation was shown to up-regulate the transcription of the prophenoloxidase 1 gene in an A. gambiae cell line.

Molecular systematics of the Philippine malaria vector Anopheles flavirostris

Allozyme and molecular sequence data from the malaria vector Anopheles flavirostris (Ludlow) (Diptera: Culicidae) were analysed from 34 sites throughout the Philippines, including the type locality, to test the hypothesis that this taxon is a single panmictic species. A finer-scaled allozyme study, of mainly Luzon samples, revealed no fixed genetic differences in sympatric sites and only low levels of variation. We obtained data from partial sequences for the internal transcribed spacer 2 (ITS2) (483 bp), the third domain (D3) (330 bp) of the 28S ribosomal DNA subunit and cytochrome c oxidase subunit I (COI) of mitochondrial DNA (261 bp). No sequence variation was observed for ITS2, only a one base pair difference was observed between Philippine and Indonesian D3 sequences and An. flavirostris sequences were unique, confirming their diagnostic value for this taxon. Sixteen COI haplotypes were identified, giving 25 parsimony informative sites. Neighbour-Joining, Maximum Parsimony, Maximum Likelihood and Bayesian phylogenetic analysis of COI sequences for An. flavirostris and outgroup taxa revealed strong branch support for the monophyly of An. flavirostris, thus confirming that Philippine populations of this taxon comprise a single separate species within the Minimus Subgroup of the Funestus Group. Variation in the behaviour of An. flavirostris is likely to be intraspecific rather than interspecific in origin. © 2006 The Royal Entomological Society.

Using GIS and Remote Sensing Technologies to Identify Environmental Variables of Malaria Vector Breeding Sites in Western Kenya

This study used Landsat 8 satellite imagery to identify environmental variables of households with malaria vector breeding sites in a malaria endemic rural district in Western Kenya. Understanding the influence of environmental variables on the distribution of malaria has been critical in the strengthening of malaria control programs. Using remote sensing and GIS technologies, this study performed a land classification, NDVI, Tasseled Cap Wetness Index, and derived land surface temperature values of the study area and examined the significance of each variable in predicting the probability of a household with a mosquito breeding site with and without larvae. The findings of this study revealed that households with any potential breeding sites were characterized by higher moisture, higher vegetation density (NDVI) and in urban areas or roads. The results of this study also confirmed that land surface temperature was significant in explaining the presence of active mosquito breeding sites (P< 0.000). The present study showed that freely available Landsat 8 imagery has limited use in deriving environmental characteristics of malaria vector habitats at the scale of the Bungoma East District in Western Kenya.

Artificial Lighting as a Vector Attractant and Cause of Disease Diffusion

BACKGROUND: Traditionally, epidemiologists have considered electrification to be a positive factor. In fact, electrification and plumbing are typical initiatives that represent the integration of an isolated population into modern society, ensuring the control of pathogens and promoting public health. Nonetheless, electrification is always accompanied by night lighting that attracts insect vectors and changes people's behavior. Although this may lead to new modes of infection and increased transmission of insect-borne diseases, epidemiologists rarely consider the role of night lighting in their surveys. OBJECTIVE: We reviewed the epidemiological evidence concerning the role of lighting in the spread of vector-borne diseases to encourage other researchers to consider it in future studies. DISCUSSION: We present three infectious vector-borne diseases-Chagas, leishmaniasis, and malaria-and discuss evidence that suggests that the use of artificial lighting results in behavioral changes among human populations and changes in the prevalence of vector species and in the modes of transmission. CONCLUSION: Despite a surprising lack of studies, existing evidence supports our hypothesis that artificial lighting leads to a higher risk of infection from vector-borne diseases. We believe that this is related not only to the simple attraction of traditional vectors to light sources but also to changes in the behavior of both humans and insects that result in new modes of disease transmission. Considering the ongoing expansion of night lighting in developing countries, additional research on this subject is urgently needed.

Malaria vectors on Buka and Bougainville islands, Papua New Guinea

Anophelines were sampled from 82 locations oil Buka and Bougainville islands in Papua New Guinea by larval collections, carbon dioxide-baited Mosquito traps, and human biting catches. Anopheles farauti s.s. was collected in larval Surveys but infrequently in mosquito traps on both islands; on Buka Island this species was readily collected in human biting catches. Anopheles faraunti 2 was commonly collected in larval surveys on both islands however. it was not collected in either mosquito traps or human biting catches. Anopheles punctulatus was found only on Buka Island, where it was commonly collected as larvae, but rarely in human biting catches and mosquito traps. Anopheles lungae was collected Lis larvae from only I site on Bougainville. Anopheles farauti s.s. led consistently throughout the night (1900-0600 h): small peaks at midnight and dawn were not statistically significant. Of 1,156 An. farauti s.s. specimens examined by enzyme-linked immunosorbent assay for malaria sporozoites. 20 were found to be positive: 12 were positive for Plasmodium falciparum and 8 were positive for P. vivax (247 variant = 5: 210 variant = 3). Anopheles farauti s.s. seems to be the major malaria vector on these islands, whereas An. punctulatus may play a minor role on Buka Island. Anophele farauti 2 is unlikely to be involved in malaria transmission on Buka or Bougainville islands.

INSECTICIDE-TREATED BED NETS IN RONDÔNIA, BRAZIL: EVALUATION OF THEIR IMPACT ON MALARIA CONTROL

Mosquito nets treated with long-lasting insecticide (LLINs), when used in compliance with guidelines of the World Health Organization, may be effective for malaria vector control. In 2012, approximately 150,000 LLINs were installed in nine municipalities in the state of Rondônia. However, no studies have assessed their impact on the reduction of malaria incidence. This study analyzed secondary data of malaria incidence, in order to assess the impact of LLINs on the annual parasite incidence (API). The results showed no statistically significant differences in API one year after LLIN installation when compared to municipalities without LLINs. The adoption of measures for malaria vector control should be associated with epidemiological studies and evaluations of their use and efficiency, with the aim of offering convincing advantages that justify their implementation and limit malaria infection in the Amazon Region.

Culex saltanensis Dyar, 1928: natural vector of Plasmodium juxtanucleare in Rio de Janeiro, Brazil

Searching for the natural vector of Plasmodium juxtanucleare in an enzootic locality: Granjas Calábria (33% of the chickens infected), Jacarepaguá, in Rio de Janeiro, Brazil, 13 comparative captures of mosquitoes were carried out, simultaneously on man (out-doors) and on chiken (in a poultry-yard), between 6 and 9 p.m., from September to March 1989. Culex saltanensis was the most frequent species in captures on chicken, accounting for 41.7% of the mosquitoes collected on this bait, showing to be highly ornithophilic (90% captured on chicken versus 10% on man). Seven specimens of Cx. saltanensis were found naturally infected in granjas Calábria: five with mature pedunculate oocysts and two with sporozoites (on in the haemocoele and one in the salivary glands). These sporozoites porudced an infection by P. juxtanucleare in a chick, which had parasitemia on day 41 after inoculation. One Cx. coronator was found with mature pedunculate oocysts. Culex saltanensis was regarded as primary vector of P. juxtanucleare in Rio de Janeiro for being highly ornithophilic and in enough density to maintain the transmission, having been found with infective sporozoites in its salivary glands, and being susceptible to the parasite and able to transmit experimentally it by the bite.

Transmission factors in malaria epidemiology and control in Africa

Genetic and environmental components of factors contributing in malaria transmission are reviewed. Particular attention is given to density dependent regulation of vector populations in relation to the survival rate anophelines. The expectation of vector activities are different according to the epidemiological characteristics of malaria, mainly its stability. In areas with perennial and high transmission (stable malaria) vector control could reduce malaria related morbidity and mortality, whithout any effect on the endemicity. However this need further investigations. In areas where the transmission period is very short (unstable malaria), vector control will have an important impact on the disease and the endemicity. Control projects using indoor spraying with insecticide and impregnated bed nets are discussed.

Malaria vectors in the municipality of Serra do Navio, State of Amapá, Amazon Region, Brazil

We conducted a survey to determine the vectors of malaria in six localities of Serra do Navio municipality, State of Amapá, from 1990 to 1991. Malaria infection rates of 29.3%, 6.2% and 20.4% were detected by human blood smears in Colônia Água Branca, Porto Terezinha and Arrependido, respectively. There was no malaria infection detected in Serra do Navio. Fifteen species were identified among 3,053 anopheline mosquitoes collected by human bait and 64.4% were identified as Anopheles albitarsis s.l., 16.7% An. braziliensis, 9.5% An. nuneztovari and 5.8% An. triannulatus. An. darlingi, the main vector of malaria in the Amazon region of Brazil, was scare. Using enzyme-linked immunosorbent assay (ELISA), a total positive rate of 0.8% (23/2876) was found for six species: fifteen An. albitarsis s.l., four An. nuneztovari, and one of each: An. braziliensis, An. triannulatus, An. oswaldoi and An. rangeli. Nine of 23 positive mosquitoes were infected with Plasmodium malariae, eight with P. vivax VK210, three with P. vivax VK247 and three with P. falciparum. Since An. albitarsis s.l. was collected feeding on humans, was present in the highest density and was positive by ELISA for malaria sporozoites, it probably plays an important role in malaria transmission in this area.

An ecoregional classification for the state of Roraima, Brazil: the importance of landscape in malaria biology

Understanding the different background landscapes in which malaria transmission occurs is fundamental to understanding malaria epidemiology and to designing effective local malaria control programs. Geology, geomorphology, vegetation, climate, land use, and anopheline distribution were used as a basis for an ecological classification of the state of Roraima, Brazil, in the northern Amazon Basin, focused on the natural history of malaria and transmission. We used unsupervised maximum likelihood classification, principal components analysis, and weighted overlay with equal contribution analyses to fine-scale thematic maps that resulted in clustered regions. We used ecological niche modeling techniques to develop a fine-scale picture of malaria vector distributions in the state. Eight ecoregions were identified and malaria-related aspects are discussed based on this classification, including 5 types of dense tropical rain forest and 3 types of savannah. Ecoregions formed by dense tropical rain forest were named as montane (ecoregion I), submontane (II), plateau (III), lowland (IV), and alluvial (V). Ecoregions formed by savannah were divided into steppe (VI, campos de Roraima), savannah (VII, cerrado), and wetland (VIII, campinarana). Such ecoregional mappings are important tools in integrated malaria control programs that aim to identify specific characteristics of malaria transmission, classify transmission risk, and define priority areas and appropriate interventions. For some areas, extension of these approaches to still-finer resolutions will provide an improved picture of malaria transmission patterns.