The importance of Anopheles albitarsis E and An. darlingi in human malaria transmission in Boa Vista, state of Roraima, Brazil

In several districts of Boa Vista, state of Roraima, Brazil we found Anopheles (Nyssorhynchus) albitarsis E to be the primary vector of human malaria parasites, and during 2001-2002 it was significantly more abundant than An. darlingi (p < 0.001). Other species sampled were An. (Nys.) braziliensis, An. (Ano.) peryassui, An. (Nys.) nuneztovari, An. (Nys.) oswaldoi s.l., and An. (Nys.) triannulatus. As determined by the ELISA technique An. darlingi had a higher overall infection rate (2.1%) compared with An. albitarsis E (1.2%). However a marginally higher proportion of An. albitarsis E was infected with Plasmodium vivax compared with An. darlingi, and the An. albitarsis E biting index was also much higher These results suggest the importance of An. albitarsis E in malaria transmission in a savannah ecoregion of northern Amazonian Brazil, and reconfirm the importance of An. darlingi even if at lower abundance.     

Variability and genetic differentiation among Anopheles (Ano.) intermedius Chagas, 1908 and Anopheles (Ano.) mattogrossensis Lutz & Neiva, 1911 (Diptera: Culicidae) from the Brazilian Amazon

Anopheles (Anopheles) intermedius and Anopheles (Ano.) mattogrossensis are Brazilian anopheline species belonging to the scarcely studied Anopheles subgenus. Few studies have been done on the genetic differentiation of these species. Both species have been found infected by Plasmodium and are sympatric with other anopheline species from the Nyssorhynchus subgenus. Eighteen enzymatic loci were analyzed in larval specimens of An. intermedius and An. mattogrossensis aiming to estimate the variability and genetic differentiation between these species. An. mattogrossensis population showed higher genetic variability (P = 44.4 and Ho = 0.081 +/- 0.031) than that of An. intermedius (P = 33.3 and Ho = 0.048 +/- 0.021). Most analyzed loci showed genotypic frequencies according to Hardy-Weinberg equilibrium, except for LAP1 and LAP2 in An. intermedius, and EST1 and PGM loci in An. mattogrossensis. The genetic distance between these species (D = 0.683) was consistent with the inter-specific values reported for Anopheles subgenus. We verified that the polymorphism and heterozygosity percentile values found in both species and compared to those in the literature, showed no relation between the level of isozyme variability and geographical distribution. The low variability found in these two species is probably more related to the niche they occupy than to their geographic distribution.     

Anopheline species, some of their habits and relation to malaria in endemic areas of Rond?nia State, Amazon region of Brazil

In view of recent studies incriminating several species of anophelines, besides Anopheles darlingi, as malaria vectors in the Brazilian Amazon, we performed an anopheline survey in four localities--Ariquemes, Cujubim, Machadinho and Itapo? do Oeste--in Rond?nia, the most malarious State in the Country. Twenty species were found. An. darlingi was, by far, the dominant species and the only one whose density coincided with that of malaria. On human baits it was more numerous in the immediate vicinity of houses than indoors where, however, it was almost the only species encountered. On both situations it fed mostly at sunset and during the first half of the night. It was less numerous far from houses and scarce inside the forest. Other species (An. triannulatus, An. evansae, An. albitarsis, An. strodei) appeared in appreciable numbers only in Ariquemes, both in areas with and without malaria. The remaining species were scanty. An. darlingi was confirmed as the primary local vector.     

Ontogenetic patterns and genetic variation in Anopheles (Anopheles) intermedius Chagas, 1908 and Anopheles (Anopheles) mattogrossensis Lutz & Neiva, 1911 (Diptera: Culicidae) in the Brazilian amazon

Changes in the expression of genes were observed during development in populations of Anopheles (Anopheles) intermedius and Anopheles (Anopheles) mattogrossensis. Esterase showed seven zones of activity: EST1 was present in all developmental stages of both species; EST2 was observed only in larvae of A. intermedius and larvae and pupae of A. mattogrossensis, with greater activity in pupae; EST3 and EST5 were present in all developmental stages, with greater intensity in larvae; EST4 and EST6 showed weak activity in larvae of A. mattogrossensis and was not found in A. intermedius. Leucine aminopeptidase showed four zones of activity, of which LAP1 and LAP2 were found in all stages of A. intermedius, with highest activity in larvae, and in larvae only of A. mattogrossensis. LAP3 was detected in all stages of A. mattogrossensis and in larvae only of A. intermedius. LAP4 was detected only in larvae and pupae of A. mattogrossensis, with greater intensity in pupae. alpha-Glycerophosphate dehydrogenase showed a single zone of activity, detected in older fourth-instar larvae and becoming more intense from the pupal stage onwards.     

Parity and age composition for Anopheles darlingi root (Diptera: Culicidae) and Anopheles albitarsis Lynch-Arribálzaga (Diptera: Culicidae) of the northern Amazon Basin, Brazil.

Parity and age composition for Anopheles darlingi and Anopheles albitarsis in the northern Amazon Basin, Brazil, were investigated. Anopheline ovaries and ovarioles were examined in order to determine whether hourly and seasonal parity status for the vectors An. albitarsis and An. darlingi would vary in two different landscapes (forest and savanna/forest) where malaria is endemic in the northern Amazon Basin. A total of 1,199 anophelines (535 An. darlingi and 664 An. albitarsis) was dissected for parity status, ovariole dilatations, and follicular stages. The total number of nulliparous and parous females for both species varied by time of collection, locality, and season. During the rainy season for the first two h of collection, more nulliparous An. albitarsis and An. darlingi females were collected in the first hour (18:00-19:00), but during the second hour (19:00-20:00) more parous females of both species were captured. During the dry season in Copaíbas, more parous females of An. albitarsis were observed in the first hour while more nulliparous females were observed in the second hour. Nulliparous and parous females of both species for both hours were not significantly different at Road 19 in the dry season. This location was characterized by a forest malaria pattern of transmission with higher numbers of parous females and population stability in the dry season. In Copaíbas, the density and parity of An. darlingi increased during the rainy season, and it could be classified as an alluvial malaria pattern of transmission. For Copaíbas, control measures would be more successful if adopted at the transition from dry to rainy season. Further investigation on longitudinal spatio-temporal change in longevity and survival rates would help us to clarify differences in vector competence for An. darlingi and An. albitarsis and add to the understanding of differences regarding prevailing landscapes in malaria epidemiology in the northern Amazon Basin.     

Molecular divergence in the timeless and cpr genes among three sympatric cryptic species of the Anopheles triannulatus complex

Anopheles triannulatus s.l. is a malaria vector with a wide geographic distribution, ranging from Argentina-Nicaragua and Trinidad. Here we analysed sequences of two genes, timeless and cpr, to assess the genetic variability and divergence among three sympatric cryptic species of this complex from Salobra, central-western Brazil. The timeless gene sequences did not conclusively differentiate Anopheles halophylus and An. triannulatus species "C". However, a partial separation has been observed between these species and An. triannulatus s.s. Importantly, the analysis of the cpr gene sequences revealed fixed differences, no shared polymorphisms and considerable genetic differentiation among the three species of the An. triannulatus complex. The results confirm that An. triannulatus s.s., An. halophylus and An. triannulatus species C are distinct taxa, with the latter two likely representing a more recent speciation event.     

Tetrahymenidae infection in mosquito populations in a malaria-endemic region of the Amazon

In a field survey performed in a malaria-endemic region of Northern Amazon, Brazil, we encountered ciliate protozoa of the family Tetrahymenidae infecting adults and larvae of the following mosquito species: Culex sp., Anopheles albitarsis l.s., Anopheles strodei, Anopheles mattogrossensis, Anopheles darlingi, and Anopheles oswaldoi l.s. Based on morphological features and life style, we have tentatively identified the parasite as Lambornella sp. The association appears pathogenic for the mosquito. Prevalence of infection in both larvae and adults was higher in the dry season.     

Amazonian malaria vector anopheline relationships interpreted from ITS2 rDNA sequences

Species identification of anopheline mosquitoes (Diptera: Culicidae) can be problematic because many of them belong to complexes of morphologically similar species, often with contrasted ecology, behaviour and vectorial importance. The application of DNA-based diagnostics has proved to be useful for distinguishing between such species. We determined ribosomal DNA sequences of the second internal transcribed spacer (ITS2) from samples of 16 species of Anopheles captured in the Amazon Basin, Brazil. Length of the ITS2 varied from 323 to 410 base pairs, with GC content ranging from 50.7% to 66.5% and sequence identity from 25% to 99% between species. Maximum-likelihood paup analysis separated two distinct groups of species conforming with the recognized subgenera Anopheles (represented by eiseni, mattogrossensis, mediopunctatus and peryassui) and Nyssorhynchus (represented by 12 spp.). For the latter group, the neighbour-joining tree generated from rDNA sequence ITS2 relationships is compatible with the morphological taxonomic key established for these Amazonian species: albitarsis, aquasalis, benarrochi, braziliensis, darlingi, deaneorum, dunhami, evansae, nuneztovari, oswaldoi, rangeli and triannulatus. These ITS2 sequence data proved to be a useful tool for species identification and, potentially, to solve taxonomic problems.     

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 Col?nia Agua 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.     

Mosquito anthropophily: implications on malaria transmission in the Northern Brazilian Amazon

Seasonal variation of adults and larvae of anophelines was studied during 2003 and 2004 in Roraima State, located in the Northern region of Brazilian Amazon. Species diversity increased with distance of capture to human dwellings. Greater diversity was found in extradomiciliary collections than in peridomiciliary or intradomiciliary. A significant association between Anopheles darlingi Root and An. albitarsis (s.l.) Arribálzaga (Diptera: Culicidae) breeding sites and the proximity to human dwellings was observed. Malaria Sporozoite Antigen Panel Assay (VectestTM Malaria) indicated An. albitarsis s.l. as one of the local vectors in the studied area. In this study, an index to describe the anthropophilic behavior of each anopheline species is proposed.     

Incrimination of Anopheles (Nyssorhynchus) rangeli and An. (Nys.) oswaldoi as natural vectors of Plasmodium vivax in Southern Colombia

Malaria transmission in the Southern Colombian state of Putumayo continues despite the absence of traditional vector species, except for the presence of Anopheles darlingi near the southeastern border with the state of Amazonas. In order to facilitate malaria vector incrimination in Putumayo, 2445 morphologically identified Anopheles females were tested for natural infection of Plasmodium vivax by ELISA. Specimens tested included An. apicimacula (n = 2), An. benarrochi B (n = 1617), An. darlingi (n = 29), An. mattogrossensis (n = 7), An. neomaculipalpus (n = 7), An. oswaldoi (n = 362), An. peryassui (n = 1), An. punctimacula (n = 1), An. rangeli (n = 413), and An. triannulatus (n = 6). Despite being overwhelmingly the most anthropophilic species in the region and comprising 66.1% of the mosquitoes tested, An. benarrochi B was not shown to be a vector. Thirty-five An. rangeli and one An. oswaldoi were naturally infected with P. vivax VK210. Sequence data were generated for the nuclear second internal transcriber space region of 31 of these 36 vivax positive mosquitoes (86.1%) to confirm their morphological identification. An. oswaldoi is known to be a species complex in Latin America, but its internal taxonomy remains unresolved. Herein we show that the An. oswaldoi found in the state of Putumayo is genetically similar to specimens from the state of Amapá in Brazil and from the Ocama region in the state of Amazonas in Venezuela, and that this form harbors natural infections of P. vivax. That An. rangeli and this member of the An. oswaldoi complex are incriminated as malaria vectors in Putumayo, is a novel finding of significance for malaria control in Southern Colombia, and possibly in other areas of Latin America.     

Molecular Phylogeny of Neotropical Anopheles (Nyssorhynchus) Albitarsis Species Complex (Diptera: Culicidae)

A phylogeny was reconstructed for four species belonging to the Neotropical Anopheles (Nyssorhynchus) albitarsis complex using partial sequences from the mitochondrial cytochrome oxidase I (COI) and NADH dehydrogenase 4 (ND4) genes and the ribosomal DNA ITS2 and D2 expansion region of the 28S subunit. The basis for initial characterization of each member of the complex was by correlated random amplification of polymorphic DNA-polymerase chain reaction (RAPD-PCR) markers. Analyses were carried out with and without an outgroup (An.(Nys.) argyritarsis Robineau-Desvoidy) by using maximum parsimony, maximum likelihood, and Bayesian methods. A total evidence approach without the outgroup, using separate models for "fast" (COI and ND4 position 3) and "slow" (rDNA ITS2 and D2, and COI and ND4 position 1) partitions, gave the best supported topology, showing close relationships of An. albitarsis Lynch-Arribálzaga to An. albitarsis B and An. marajoara Galv?o & Damasceno to An. deaneorum Rosa-Freitas. Analyses with the outgroup included showed poorer support, possibly because of a long branch attraction effect caused by a divergent outgroup, which caused one of the An. marajoara specimens to cluster with An. deaneorum in some analyses. The relationship of the above-mentioned result to a separately proposed hypothesis suggesting a fifth species in the complex is discussed.     

Intragenomic rDNA ITS2 variation in the neotropical Anopheles (Nyssorhynchus) albitarsis complex (Diptera: Culicidae)

We cloned and sequenced the rDNA internal transcribed spacer 2 (ITS2) of 4 species belonging to the neotropical Anopheles (Nyssorhynchus) albitarsis complex, that is, A. albitarsis; A. albitarsis B; Anopheles marajoara, a proven malaria vector; and Anopheles deaneorum, a suspected vector. Even though the ITS2 sequences of these species were very similar (< or =1.17% divergence), we found differences suitable for species identification and intragenomic variation of possible consequence in phylogenetic reconstruction. Variation came from 2 microsatellite regions and a number of indels and base substitutions. The existence of partially correlated subsets of clones in A. albitarsis is hypothesized either to be separate rDNA loci or to be semi-independently evolving portions of a single rDNA locus. No differences were found between males and females, suggesting that similar rDNA arrays exist on both the X and Y chromosomes. In addition, highly variant clones, possibly pseudogenes, were found in A. marajoara from Venezuela.     

基于rDNA-ITS2序列的中国按蚊属塞蚊亚属部分种类的系统发育研究(双翅目:蚊科)

应用rDNA-ITS2序列,采用邻接法(NJ)、最大简约法(MP)和最大似然法(ML),以按蚊亚属Anopheles的中华按蚊An.(An.)sinensis和赫坎按蚊An.(An.)hyrcanus为外群,对采自中国的按蚊属Anopheles塞蚊亚属Cellia 21种蚊进行了系统发育分析.结果表明:rDNA-ITS...     

A cytogenetic analysis of the polytene chromosomes of Anopheles triannulatus (Diptera: Culicidae) from western Venezuela

The larval polytene chromosomes of Anopheles triannulatus are described for the first time in this report. The population from western Venezuela was chosen arbitrarily as the chromosome standard and is the basis for the photomap presented here. Anopheles trannulatus displays whole-arm associations distinct from other members of the subgenus Nyssorhynchus that have been examined cytogenetically. The association pattern for An. triannulatus is 2R-3R and 2L-3L, rather than the more common 2R-2L and 3R-3L. The possible role of whole-arm translocations in speciation is briefly discussed.     

Biting indices,host-seeking activity and natural infection rates of anopheline species in Boa Vista,Roraima, Brazil from 1996 to 1998

The epidemiology of the transmission of malaria parasites varies ecologically. To observe some entomological aspects of the malaria transmission in an urban environment, a longitudinal survey of anopheline fauna was performed in Boa Vista, Roraima, Brazil. A total of 7,263 anophelines was collected in human bait at 13 de Setembro and Caran? districts: Anopheles albitarsis sensu lato (82.8%), An. darlingi (10.3%), An. braziliensis (5.5%), An. peryassui (0.9%) and An. nuneztovari (0.5%). Nightly 12 h collections showed that An. albitarsis was actively biting throughout the night with peak activities at sunset and at midnight. An. darlingi bit during all night and did not demonstrate a defined biting peak. Highest biting indices, entomological inoculation rates and malaria cases were observed seasonally during the rainy season (April-November). Hourly collections showed host seek activity for all mosquitoes peaked during the first hour after sunset. An. darlingi showed the highest plasmodial malaria infection rate followed by An. albitarsis, An. braziliensis and An. nuneztovari (8.5%, 4.6%, 3% and 2.6%, respectively). An. albitarsis was the most frequently collected anopheline, presented the highest biting index and it was the second most frequently collected infected species infected with malaria parasites. An. albitarsis and An. darlingi respectively, are the primary vectors of malaria throughout Boa Vista.     

Diverse population dynamics of three Anopheles species belonging to the Triannulatus Complex (Diptera: Culicidae)

The Triannulatus Complex of Anopheles (Nyssorhynchus) consists of at least three sibling species, namely Anopheles triannulatus s.s., Anopheles halophylus and a third undescribed member herein referred to as An. triannulatus "C". Sympatric anophelines belonging to species complexes, even though closely related, may exploit different environments such as larval habitats. In this paper we hypothesize that rainfall and seasonal flooding would distinctly influence the availability of larval habitats and consequently the seasonal population dynamics of sympatric members of the Triannulatus Complex. A reflection of this is distinct seasonal biting frequencies exhibited by three members of the Triannulatus Complex at a site in Central Brazil. Population dynamics seem to be influenced by the water level in the local rivers, although biting frequency of all three species was negatively influenced by rainfall. An. triannulatus s.s. was more abundant following the end of the rainy season, but notably 30 to 60 days after flooding. On the other hand, An. halophylus and An. triannulatus C peaked during the middle of the dry season, when water impoundments have no inflow, are somewhat reduced in size and the water becomes brackish. Differences in population dynamics were greater between An. triannulatus s.s. and An. halophylus and An. triannulatus C than between An. halophylus and An. triannulatus C. This might reflect differences in larval habitat exploitation and therefore spatial segregation among these members of the complex.     

The biogeography and population genetics of neotropical vector species

Phylogenetic and population genetic data support the Pliocene or Pleistocene divergences of the co-distributed hematophagous insect vectors, the sand fly Lutzomyia longipalpis s.l., the mosquitoes Anopheles darlingi and A. albitarsis s.l., and the triatomines Rhodnius prolixus and R. robustus. We examined patterns of divergence and distribution in relation to three hypotheses of neotropical diversification: Miocene/Pliocene marine incursion, Pliocene/Pleistocene riverine barriers and Pleistocene refugia. Only R. prolixus has a pattern concordant with the refugia hypothesis, and R. robustus conforms to the marine incursion predictions. A. darlingi partially fits the refugia hypothesis. For L. longipalpis s.l. and A. albitarsis s.l., elements of both incursion and refugia hypotheses seem to fit, suggesting perhaps an interaction of factors determining their distribution patterns.     

Incrimination of Anopheles (Anopheles) intermedius Peryassú, An. (Nyssorhynchus) nuneztovari Gabaldón, An. (Nys.) oswaldoi Peryassú as natural vectors of Plasmodium falciparum in French Guiana

Anopheles darlingi Root is the major vector of human malaria in the Neotropics and has been considered to be the sole malaria vector in French Guiana. The presence of other potential vectors suggests that malaria may be transmitted by other species under certain conditions. From 2006-2011, all anopheline specimens collected from 11 localities were assayed to determine if the Plasmodium circumsporozoite protein was present. In addition to An. darlingi, we found Anopheles oswaldoi, Anopheles intermedius and Anopheles nuneztovari specimens that were infected with Plasmodium sp. Further investigations on the behaviour and ecology of An. oswaldoi, An. intermedius and An. nuneztovari are necessary to determine their role in malaria transmission in French Guiana.