The larval sites used by An. albimanus are characterised across its range as open, sunlit and containing clear water  (Table 3). The species can be found in natural and man-made habitats where these characteristics exist. For example, it occurs in recently planted rice fields, or in older fields with sunlit areas in between the rice plants  (Table 4). Anopheles albimanus has been associated with floating mats of blue-green algae [76, 78, 79], which are often found in sunlit waters. The larvae of this ubiquitous species tolerate a wide variation in water chemistry and are able to exploit diverse food sources , enabling them to survive in both fresh water (e.g. irrigation channels, small ponds, marshes, slow flowing streams and river margins [73, 80–83]) and brackish water (e.g. mangrove swamps [78, 83, 84]) (Tables 3-5).
Anopheles albimanus is predominantly exophagic with exophilic resting behaviour [77, 80, 85, 86] (Table 6), however there is some indication that in the northern reaches of its distribution (Mexico, Central America), this species exhibits a preference for resting indoors after feeding [87, 88]. In a mark-recapture study that ultimately influenced changes in the vector control regime in southern Mexico , Bown et al.  examined An. albimanus resting behaviour and found that 80% landed indoors after feeding. Anopheles albimanus bites in the evening and during the night [77, 80, 85, 86, 90, 91]. It appears to show a tendency for zoophily, but this is dependent upon location [86, 89, 91, 92]. In Colombia, Solarte et al.  described An. albimanus as exhibiting "a high degree of anthropophilic activity". In contrast, Loyola et al.  described this species as highly zoophilic in Mexico, however they also pointed out that host availability and other ecological conditions influence the host choice of this species.
Anopheles albitarsis complex
The An. albitarsis species complex includes An. albitarsis (formerly Sp. A), An. albitarsis Sp. B and Sp. E, An. marajoara (formerly Sp. C) and An. deaneorum (formerly Sp. D). Anopheles marajoara is discussed separately below. Overall, members of this complex exhibit larval habitat preferences similar to An. albimanus, occupying sunlit, clear, fresh water [93, 94] (Rubio-Palis, unpub. obs.) (Table 3), although there are examples of it being found in atypical conditions, Da Silva-Vasconcelos et al.  found relatively high numbers, compared to other species, in brick pits containing turbid water mixed with clay particles . Typical habitats are fresh, still water bodies such as lagoons, lakes or rice fields [74, 93, 96] (Table 4). Indeed, members of the An. albitarsis complex are closely associated with human rice cultivation [97, 98], although, like An. albimanus, immature stages tend to be found only in fields with early stages of rice growth . Adults are generally exophilic in their resting behaviour [99, 100], but will bite both indoors and outdoors and appear to show little host preference, biting humans and animals indiscriminately (dependent on location), during the evening and at night [95, 99–105] (Table 6).
Grillet  described An. aquasalis as an opportunistic species whose individuals "may be poor competitors or may develop few anti-predator defences". In areas where An. aquasalis and An. albimanus co-exist, An. albimanus will dominate (Rubio-Palis, unpub. obs.) suggesting that the inability of An. aquasalis to out-compete other dominant Anopheles species may be highly influential in defining its ecological requirements (as its name implies, it is generally only found in coastal areas).
Anopheles aquasalis is found in sunlit habitats containing emergent vegetation, in both brackish and fresh water [93, 106, 107] (Table 3). It is considered to "prefer" clear, still, non-polluted water such as stream pools, mangrove swamps, grass swamps, lagoons and ditches [108, 109], although there are examples of it being found in turbid, slow flowing water bodies, in relatively high numbers (e.g. ) (Tables 4 & 5).
Adults, once again, are opportunistic, feeding indoors or outdoors on animal or human hosts, but generally resting outdoors before and after feeding [100, 110–112]. Biting tends to begin at dusk, peaking early in the night, and tailing off as the night progresses [111, 112] (Table 6). The time of the biting peak and bias towards endo-or exophagy depends on the location, for example in Maranhão, Brazil, Xavier & Rebelo  observed a tendency to bite at dusk, indoors. Berti et al.  measured biting behaviour in two villages in Sucre State, Venezuela, where An. aquasalis predominantly bit outdoors, and found that biting peaked earlier (7 pm) in the village of Guayana, where greater numbers of An. aquasalis were found, than in Santa Fe, where the peak occurred between 8 and 9 pm. During the study, two females were collected biting during the day.
Anopheles darlingi is considered to be one of the most efficient malaria vectors in the Neotropical region . It is mainly a riverine mosquito, generally confined to rural, lowland forested locations  (Rubio-Palis & Manguin, unpub. obs.). Conversely, however, Vittor et al.  suggest that deforestation and human environmental alteration can create habitats which are favourable to An. darlingi as they found higher densities in areas with limited forest cover than in areas predominated by forest. The larval habitats of An. darlingi can be characterized as: natural water bodies such as lagoons, lakes and particularly slow flowing streams or rivers with shaded, clear water, and associated submersed vegetation such as bamboo roots from overhanging spiny bamboo [72–74, 113] (Tables 3-5). Larvae are encountered most frequently in patches of floating debris along river margins . There are examples of larvae being found in uncharacteristic locations, however, such as in slightly brackish water (Belize, ), in low numbers in turbid, polluted water (brick pits, ) and in abandoned gold mine dugouts in southern Venezuela , further suggesting a level of adaptation to areas altered by humans .
Anopheles darlingi tends to rest outdoors regardless of where it has taken its blood meal [116, 117] (Table 6). Adults will bite throughout the night [95, 117–120] and the degree of endo- and exophagy of this species varies from one place to another as does its host preference  (Table 6). It has been suggested that the biting pattern of An. darlingi may represent an adaptation to human behaviour [117, 121]. Moreno et al. , for example, contend that the all night activity of An. darlingi in the gold mining areas of southern Venezuela is a response to the all night activity of the miners. Furthermore, a number of studies that report exophagy in this species (Table 6) refer to peri-domestic sampling [113, 116, 122] at sites where indoor insecticide spraying is or had recently been used for vector control [113, 117, 122, 123]. Other indications of the adaptability of An. darlingi are emerging, with an ongoing study in Venezuela, close to the Brazilian border (Roraima), recently reporting that specimens have been collected at altitudes above 800 m (Rubio-Palis, unpub. obs: re. Berti et al. at the Instituto de Altos Estudios "Dr. Arnoldo Gabaldon", Ministry of Health).
Studies examining the bionomics of An. freeborni tend to have been performed prior to 1985 limiting the amount of useful data found within the MAP library. Therefore additional searches of older references were needed to develop a more complete picture of the bionomics of this species.
McHugh  suggests that An. freeborni is an opportunistic feeder. In a study where its blood meals were identified, host availability was key in determining blood source. For example, the An. freeborni females collected in cattle areas contained the highest proportion of bovine blood, whereas 35% had fed on dogs at sites of human habitation near a rice production area, and no bovine blood was found. This suggests that the species is zoophilic, although a preference is not indicated in this study. Washino & Tempelis , in a similar study, reported very low levels of human blood in An. freeborni (<1% of mosquitoes tested) even though sampling was conducted in areas of human habitation. They also indicated the opportunistic nature of this species, given the high level of rabbit blood meals found. However, once again, the proportion was highly dependent on rabbit availability. The resting mosquitoes were collected from farmyard sheds, abandoned domestic animal shelters, house porches, artificial shelters, bridges and culverts, indicating an outdoor resting preference.
Orr & Resh  demonstrated a positive association between An. freeborni larval densities and plant cover, indicating that, as with all the other DVS in the Americas, vegetation is a key characteristic of their larval habitats. This is reflected by the species' ready utilization of rice fields, although significantly higher numbers of adult An. freeborni are found in riparian and mixed habitats than in rice and pasture habitats [127, 128] (Table 4).
Anopheles marajoara is a member of the An. albitarsis complex. Previously believed to be a secondary, local vector of minor importance, it was identified as a DVS in the 1990s in a study conducted in Amapa, Brazil , where it was found in high densities and with high levels of Plasmodium infection when compared to An. darlingi. Here, therefore, this species is considered separately from other members of the An. albitarsis complex.
Anopheles marajoara is a lowland species, associated with wetlands, secondary forests and human intervention [115, 129] (Rubio-Palis, unpub. obs.). Moreno et al.  suggest that the recent studies in Brazil identifying An. marajoara as dominant over An. darlingi may be a result of human interventions which favour species that oviposit in open lagoons with abundant macrophytes. Conn et al.  described forest clearance and pollution as reducing the availability of larval sites for An. darlingi whilst increasing the availability of sites such as agricultural ponds and sunlit marshy areas, which are the preferred habitats of An. marajoara. Overall, An. marajoara larval sites are generally sunlit with clear, still water, although there are examples of this species being found in both clear and muddy waters, such as fish ponds  and gold mine dugouts  (Tables 3-5).
In Amapa, An. marajoara is described as exclusively exophilic , which appears to be the case across its range [99, 116, 117] (Rubio-Palis, unpub. obs.), however a study in Colombia examining the indoor resting behaviour of anophelines, reported An. marajoara resting indoors, close to the ground , suggesting some endophilic behaviour in limited areas (Table 6).
Anopheles marajoara bites both humans and animals [102, 116, 131], both indoors and outdoors throughout the night, with biting tending to peak in the evening [99, 113, 117, 129] (Table 6), though again these characteristics can vary according to location. For example, a study in southern Venezuela reported marked exophagic behaviour, with 74.2% of An. marajoara captured biting outdoors . This contrasts with a study carried out in western Venezuela  that reported indiscriminate indoor and outdoor biting. There is currently some debate, however, about the identity of members of the An. albitarsis complex in Venezuela. Past investigations have only identified the presence of An. marajoara[117, 132], but a recent study suggests the possibility of misidentification and the additional presence of An. janconnae. A consensus for this "discovery" has yet to be reached.
The An. nuneztovari species complex is yet to be fully resolved and there is a need for some clarification of sibling identity. The complex contains either two or three cytological species (A and B/C), with B and C possibly two forms of a single species [134–137] (Rubio-Palis & Manguin, unpub. obs.). Furthermore, Calado et al.  have recently formally resurrected An. goeldii from synonymy with An. nuneztovari A, making the task of truly categorising members of this complex all the more challenging.
Anopheles nuneztovari larvae are found in both sunlit and shaded habitats [74, 138] (Rubio-Palis, unpub. obs.). Sites usually contain fresh, clear, still or flowing water with floating or emergent vegetation [74, 138] (Table 3). Despite this, Nagm et al.  report finding this species in several turbid water bodies, and da Silva-Vasconcelos et al.  found small numbers in brick pits containing very turbid water polluted with brick dust. Indeed, Service  described An. nuneztovari larval habitats as "muddy waters of pools, vehicle tracks, hoof prints, small ponds, especially in and around towns".
Habitats are found in small or large, natural or constructed bodies of water, including lagoons, lakes, slow flowing rivers, fish ponds, gold mine dugouts, rain puddles and temporary or permanent pools [80, 138–140] (Table 4 & 5). Tadei & Thatcher  described An. nuneztovari as a species readily able to colonise and even dominate in altered environments and yet, despite this characteristic, it is not known to breed in rice fields.
Adult behaviour differs depending on the sibling species, most specifically in terms of their biting times, with An. nuneztovari A (Brazil) biting earlier, peaking between 6 and 8 pm [95, 141, 142], and An. nuneztovari B/C (Venezuela and Colombia) biting later and throughout the night, peaking between 10 pm and 2 am . Olano et al.  reported peri-domiciliary biting of An. nuneztovari early in the evening in Buenaventura, Colombia, however this may be the result of low biting densities rather than a true indication of preference. The Brazilian sibling is considered to be a non-vector, possibly due to its behaviour rather than of an inability to transmit malaria [142, 143], given that there is evidence of Plasmodium infection in An. nuneztovari in Amapa, Brazil [144, 145].
Service  suggested that An. nuneztovari mainly feeds on animals, but will bite humans outdoors. Studies in Amapa, Brazil and in Venezuela, which analysed blood meals of An. nuneztovari, suggest zoophilic behaviour, but an accompanying human landing catch in the same area found over 120 mosquitoes/person/night [99, 102, 116]. Exo- and endophagy of members of this species complex vary with location but Tadei & Thatcher  suggested that human behaviour, such as a propensity to stay outdoors late into the evening, or the application of insecticides, may influence biting location. The majority of studies summarised here report exophagic behaviour [80, 99, 116, 139, 141] (Table 6), with only Rubio-Palis & Curtis  reporting both endo- and exophagic behaviour in western Venezuela. Rubio-Palis & Curtis  mentioned the contrasting observations of exo- and endophagy reported for An. nuneztovari and suggested this may be due to collection bias (collector, location or short series of observations) or a result of different behavioural patterns between sympatric siblings, which again highlights a need for further clarification of species identity. All sibling species within the An. nuneztovari complex are highly exophilic, resting outdoors both before and after feeding [99, 116] (Table 6).
Anopheles pseudopunctipennis is a complex of at least two species [146, 147] and two forms  (Rubio-Palis & Manguin, unpub. obs.). It can survive and transmit malaria at altitudes higher than many other DVS, with its range extending up to approximately 3000 m [22, 149] (Rubio-Palis & Manguin, unpub. obs.). This species is most frequently found in sun-exposed, shallow, clear and freshwater streams or river pools with abundant filamentous algae , although there are a number of reports of larvae found in turbid, cloudy water [93, 109, 149], including at one site polluted with cow faeces . The majority of larval habitats have fresh water, but about 10% contain brackish or sea water [93, 149] (Table 3). Past studies conducted in Grenada suggested that An. pseudopunctipennis was restricted to still or stagnant water  but more recent investigations on this island indicate that it can survive in slow flowing water bodies, possibly protected against the current by mats of Spirogyra-type green filamentous algae [109, 149]. The presence of such filamentous algae is a key characteristic associated with larval habitats of this species [93, 109, 149, 151–154] (Table 3). Indeed, a study that examined the potential impact of An. pseudopunctipennis control via environmental manipulation demonstrated significant reductions in densities after the removal of filamentous algae from larval sites .
Service  stated that adult An. pseudopunctipennis "...feed almost indiscriminately on humans and domestic animals, indoors or outdoors...". Studies conducted in southern Mexico corroborate his statement, for example Fernandez-Salas et al.  found that a greater proportion of An. pseudopunctipennis were attracted to horse-baited traps than to humans (although significant numbers were captured on humans), however in a previous study in the same four villages , they found that a high proportion of the An. pseudopunctipennis females resting indoors contained human blood. They suggested that host availability was responsible for host selection, rather than this being due to any preference exhibited by the mosquitoes. Lardeux et al. , in a study specifically designed to examine host preference, found some level of choice exhibited by females. They suggested that this species is not highly anthropophilic, but reiterated its opportunistic nature, stating that An. pseudopunctipennis females will bite the first "preferred" host they encounter (in their experiment the feeding preference was ranked as: sheep, goats and donkeys, followed by humans and cows).
Service  also stated that "[An. pseudopunctipennis] rest outdoors after feeding", however the studies above [155, 156], and those of Casas et al. , who used the mark-recapture method to identify resting behaviour of fed and unfed mosquitoes, indicated that a proportion of An. pseudopunctipennis will rest indoors both before and after feeding. However, Loyola et al.  suggested that IRS with DDT has not only increased insecticide resistance in some areas, but also promoted more exophilic behaviour.
Anopheles pseudopunctipennis bites during the night, with small variations in peak activity depending on location and host, for example Fernandez-Salas et al.  demonstrated a uni-modal peak, with indoor biting peaking at 1 am and outdoor biting peaking slightly earlier, at midnight. Interestingly, they also reported a bimodal biting pattern on horse bait, where biting peaked at 7 pm, with a second, smaller peak occurring between midnight and 1 am (Table 6).
Anopheles quadrimaculatus subgroup
The Quadrimaculatus Subgroup of the Maculipennis Group is often mis-reported as a complex (e.g. [19, 69, 70]). Within this subgroup, An. quadrimaculatus (formerly Sp. A) is the most wide-spread species [19, 70], considered the most "important" , and it is the only species within the subgroup identified as a DVS [25–29]. Therefore only this species is given further consideration here, with the caveat that some studies included in the bionomics review do not distinguish species and report An. quadrimaculatus s.l. However, as suggested by Seawright et al. , owing to its abundance, seconded only by An. smaragdinus (formerly An. quadrimaculatus Sp. B), An. quadrimaculatus (and An. smaragdinus) "are probably the species that most researchers have studied in the past".
Anopheles quadrimaculatus is highly associated with rice cultivation [69, 160–166] (Table 4), showing a preference for the oligotrophic conditions found when the rice fields are first flooded . Such conditions reflect those found in the natural larval habitat of this species: generally fresh, still water in relatively large sites such as lakes and marshes with emergent vegetation [69, 167–170] (Tables 3 & 4). Unusual larval sites have been reported including a sewerage retention pond containing highly polluted effluent draining from a pig farm, a small plastic bucket containing rainwater, submerged leaf litter and floating pine needles , and wastewater evaporation-percolation ponds in Florida  (Table 5).
Adults appear to be generally zoophilic, biting and resting outdoors [69, 168, 170] (Table 6), however, amongst all studies summarised, this may be an artefact of the areas sampled and the lack of human hosts rather than an indication of preference [168, 170]. Of the species in this subgroup, however, An. quadrimaculatus appears to show the highest level of anthropophily. Jensen et al.  examined blood meals in An. quadrimaculatus (Sp. A), Sp. B (An. smaragdinus) and Sp. C1 (An. diluvialis) at a campsite where human hosts were available, and in a woodland approximately a mile away, where there was a lower chance of human contact, and found that 10.7% of An. quadrimaculatus sampled at the campsite had fed on humans compared to none in the wooded area. The other two species demonstrated very low or no human biting at both sites. Reinert et al. , in a comprehensive description of the taxonomy and bionomics of the Quadrimaculatus Subgroup, also reported such exophagic and zoophilic behaviour, describing frequent collections of engorged females from horse stables and cattle barns and a number of observations of feeding on large domestic animals.
Anopheles quadrimaculatus bites throughout the night, showing higher activity at dusk and dawn . Resting behaviour, as with all other DVS described here, is exophilic, and includes sites in holes and rot cavities in trees, livestock barns, outdoor latrines, under bridges and under the eaves of buildings [69, 170, 173, 174] (Table 6).
This study has focused on describing the distribution and bionomics of the main malaria vectors in the Americas. However, there are numerous secondary or local vectors that may also play an important yet often forgotten role in malaria transmission. In the Americas, secondary vectors include the various species of the subgenus Kerteszia (e.g. An. cruzii, An. bellator and An. neivai), whose larvae characteristically inhabit water contained in bromeliads, as well as An. (Anopheles) vestitipennis, An. (Ano.) neomaculipalpus, An. (Nyssorhynchus) braziliensis, An. (Nys.) triannulatus, An. (Nys.) strodei, An. (Ano.) intermedius and members of the An. (Nys.) oswaldoi complex. Their "secondary" vector status is a factor of location, distribution, vectorial capacity and, occasionally, history (i.e. a species once considered primary, but now relegated to a secondary role through changes in the local environment). Circumstantial evidence, or the identification of Plasmodium circumsporozoite proteins in females, can lead to a species being incriminated as a vector, where in reality it may have little to no impact . However, some "secondary" species are proven and potent vectors within their local range.
Both An. cruzii and An. bellator are identified by White  as main malaria vectors, yet their larval habitats (bromeliads) restrict their role in malaria transmission to areas where such plants are abundant, for example in the rainforests of Brazil where these two species are considered to be primary local vectors. Deforestation may have reduced the availability of habitats for these species but they ought not to be overlooked as their range extends along the eastern coast of South America, from Guyana to the southernmost tip of Brazil. Moreover, there are reports of these species being found in bromeliads and artificial containers in urban and peri-urban sites [175, 176] with An. cruzii specifically described as an aggressive biter throughout the day and night . Anopheles neivai also makes use of bromeliads during its larval stage and is an important vector of human malaria in the Pacific coastal areas of Colombia [178, 179]. It has been responsible for outbreaks of malaria in the Venezuelan Andes at altitudes above 1000 m [180, 181].
Anopheles vestitipennis is considered to be a secondary vector species of major importance within its range. In Belize, it has been described as a primary vector, ousting An. albimanus which plays a more secondary role there [182–185]. Achee et al.  found it to be positive for both P. falciparum and P. vivax and Loyola et al.  found it to be the most abundant species (>80% of those collected on human bait) in their study area within the Lacandon rainforest of Chiapas, Mexico, and the only species in the area to be positive for the P. vivax antigen. They also demonstrated that it readily bites humans both indoors and outdoors. Anopheles vestitipennis is also found in the Caribbean islands where it may have been involved in malaria outbreaks in Cuba  and Haiti (Rubio-Palis, unpub. obs.).
Malaria parasites have also been detected in several other species. De Oliveira Ferreira et al.  found natural infections of Plasmodium in An. triannulatus, An. braziliensis, An. strodei and An. oswaldoi in Rondonia, Brazil. Anopheles triannulatus and An. strodei were only infected with P. vivax, however the very small sample sizes (the largest being five specimens of An. triannulatus), prevents the drawing of any conclusions regarding any refractory characteristics.
Anopheles oswaldoi has been confirmed as a malaria vector in Brazil [119, 188–190], and it is the principal vector in the State of Acre in the Brazilian Amazon where it has been found in large densities and with relatively high sporozoite rates for P. falciparum (3.41%), P. vivax-210 (2.26%), P. vivax-247 (1.22%) and P. malariae (0.42%) . Anopheles oswaldoi has also been incriminated as a vector of P. vivax in Colombia , Peru  and Venezuela .
The role of An. neomaculipalpus in malaria transmission is not clear but it has recently been found to be positive for P. vivax in Venezuela [194, 195] and positive for P. falciparum in Colombia [195, 196] and it is considered to be a highly anthropophilic species (). Moreno et al. suggested that this species may play a role as a secondary vector of "frontier malaria" in areas of forest subject to recent human activity that can increase vector diversity with the creation of new larval sites. De Oliviera Ferreira et al.  reiterated this hypothesis and suggested that the presence of infection amongst these secondary vectors may be linked to the extensive environmental changes that may reduce the populations of sylvatic animals in an area whilst simultaneously increasing the number of humans, which will inevitably cause a higher level of vector-human contact.