BackgroundAnthropogenic habitat disturbance is a prime cause in the current trend of the Earth’s reduction in biodiversity. Here we show that the human footprint on the Central African rainforest, which is resulting in deforestation and growth of densely populated urban agglomerates, is associated to ecological divergence and cryptic speciation leading to adaptive radiation within the major malaria mosquito Anopheles gambiae.Methodology/Principal FindingsIn southern Cameroon, the frequency of two molecular forms–M and S–among which reproductive isolation is strong but still incomplete, was correlated to an index of urbanisation extracted from remotely sensed data, expressed as the proportion of built-up surface in each sampling unit. The two forms markedly segregated along an urbanisation gradient forming a bimodal cline of ∼6-km width: the S form was exclusive to the rural habitat, whereas only the M form was present in the core of densely urbanised settings, co-occurring at times in the same polluted larval habitats of the southern house mosquito Culex quinquefasciatus–a species association that was not historically recorded before.Conclusions/SignificanceOur results indicate that when humans create novel habitats and ecological heterogeneities, they can provide evolutionary opportunities for rapid adaptive niche shifts associated with lineage divergence, whose consequences upon malaria transmission might be significant.

Anopheles funestus is a major vector of malaria in Africa. It belongs to a group of sibling species that can be identified morphologically only at certain stages of their development. A diagnostic polymerase chain reaction (PCR)-based tool made it possible to differentiate five species of the group. The assay seems to be applicable over all their distribution area for four of these species: An. funestus, An. leesoni, An. parensis, and An. vaneedenip. The fifth species, An. rivulorum, is the second most abundant species of the group and can be mistaken at its adult stage for the major vector of malaria An. funestus. Molecular and morphologic observations of specimens identified as An. rivulorum from Cameroon and South Africa showed that they belong to two different taxa. The species identified in Cameroon, and named here An. rivulorum-like, might extend to western Africa and central Africa. The species-specific PCR assay is supplemented by a primer specific to An. rivulorum-like and thus makes it possible to differentiate the five species of the An. funestus group and the newly defined taxon.

SummaryDuring extensive sampling in Burkina Faso and other African countries, the Leu-Phe mutation producing the kdr pyrethroid resistance phenotype was reported in both Anopheles gambiae ss and A. arabiensis. This mutation was widely distributed at high frequency in the molecular S form of A. gambiae while it has been observed at a very low frequency in both the molecular M form and A. arabiensis in Burkina Faso. While the mutation in the M form is inherited through an introgression from the S form, its occurrence is a new and independent mutation event in A. arabiensis. Three nucleotides in the upstream intron of the kdr mutation differentiated A. arabiensis from A. gambiae ss and these specific nucleotides were associated with kdr mutation in A. arabiensis. Ecological divergences which facilitated the spread of the kdr mutation within the complex of A. gambiae ss in West Africa, are discussed.

BackgroundLimitations in the ability of organisms to tolerate environmental stressors affect their fundamental ecological niche and constrain their distribution to specific habitats. Evolution of tolerance, therefore, can engender ecological niche dynamics. Forest populations of the afro-tropical malaria mosquito Anopheles gambiae have been shown to adapt to historically unsuitable larval habitats polluted with decaying organic matter that are found in densely populated urban agglomerates of Cameroon. This process has resulted in niche expansion from rural to urban environments that is associated with cryptic speciation and ecological divergence of two evolutionarily significant units within this taxon, the molecular forms M and S, among which reproductive isolation is significant but still incomplete. Habitat segregation between the two forms results in a mosaic distribution of clinally parapatric patches, with the M form predominating in the centre of urban agglomerates and the S form in the surrounding rural localities. We hypothesized that development of tolerance to nitrogenous pollutants derived from the decomposition of organic matter, among which ammonia is the most toxic to aquatic organisms, may affect this pattern of distribution and process of niche expansion by the M form.ResultsAcute toxicity bioassays indicated that populations of the two molecular forms occurring at the extremes of an urbanization gradient in Yaounde, the capital of Cameroon, differed in their response to ammonia. The regression lines best describing the dose-mortality profile differed in the scale of the explanatory variable (ammonia concentration log-transformed for the S form and linear for the M form), and in slope (steeper for the S form and shallower for the M form). These features reflected differences in the frequency distribution of individual tolerance thresholds in the two populations as assessed by probit analysis, with the M form exhibiting a greater mean and variance compared to the S form.ConclusionsIn agreement with expectations based on the pattern of habitat partitioning and exposure to ammonia in larval habitats in Yaounde, the M form showed greater tolerance to ammonia compared to the S form. This trait may be part of the physiological machinery allowing forest populations of the M form to colonize polluted larval habitats, which is at the heart of its niche expansion in densely populated human settlements in Cameroon.

Understanding how divergent selection generates adaptive phenotypic and population diversification provides a mechanistic explanation of speciation in recently separated species pairs. Towards this goal, we sought ecological gradients of divergence between the cryptic malaria vectors Anopheles coluzzii and An. gambiae and then looked for a physiological trait that may underlie such divergence. Using a large set of occurrence records and eco-geographic information, we built a distribution model to predict the predominance of the two species across their range of sympatry. Our model predicts two novel gradients along which the species segregate: distance from the coastline and altitude. Anopheles coluzzii showed a ‘bimodal’ distribution, predominating in xeric West African savannas and along the western coastal fringe of Africa. To test whether differences in salinity tolerance underlie this habitat segregation, we assessed the acute dose–mortality response to salinity of thirty-two larval populations from Central Africa. In agreement with its coastal predominance, Anopheles coluzzii was overall more tolerant than An. gambiae. Salinity tolerance of both species, however, converged in urban localities, presumably reflecting an adaptive response to osmotic stress from anthropogenic pollutants. When comparing degree of tolerance in conjunction with levels of syntopy, we found evidence of character displacement in this trait.

The swarming and mating systems of natural populations of An. gambiae M and S forms were investigated through longitudinal surveys conducted between July 2006 and October 2009 in Soumousso and Vallée du Kou (VK7), two rural areas of south-western Burkina Faso where these forms are sympatric. In both sites, the majority of swarms were recorded above visual markers localized within human habitats. In Soumousso, a wooded area of savannah, 108 pairs caught in copula from 205 swarms were sampled; in VK7, a rice growing area, 491 couples from 250 swarms were sampled. In neither site was any spatial segregation observed between the swarm sites used by the two forms of An. gambiae, which shared many of their visual markers. Furthermore, mixed swarms were collected annually in frequencies varying from one © 2014. This manuscript version is made available under the CC-BY-NC-ND 4.0 license http://creativecommons.org/licenses/by-nc-nd/4.0/ 2 site to another, though no mixed inseminations were recorded, corroborating the low hybrid rate previously reported in the field. The occurrence of inter-specific mate-recognition mechanisms, which allow individuals to avoid hybridization, is discussed.

Resistance of Anopheles gambiae to organophosphate and carbamate insecticides was first reported in Côte d'Ivoire, West Africa. Subsequent studies revealed that it resulted from a single point mutation in the oxyanion hole of the acetycholinesterase enzyme (ace-1(R) mutation). We investigated the distribution and prevalence of the ace-1(R) mutation in An. gambiae s.l. populations from seven locations in south-western Burkina Faso. The ace-1(R) mutation was found in both M and S molecular forms of An. gambiae s.s., but it was absent in An. arabiensis. Its frequency ranged from 0.25 to 0.5 in S form and 0.04 to 0.13 in M form, though they were sympatric. The lack of homozygous resistance indicated a strong genetic cost associated with the mutation. These data suggest that organophosphate and carbamate resistance conferred by target site insensitivity is spreading in populations of An. gambiae s.s. from West Africa.

This study reports on the distribution of pyrethroid and DDT resistance and the L1014F knockdown resistance (kdr) mutation in Anopheles gambiae s.l. populations from 21 localities in three different climatic zones of Burkina Faso from August to October 2006. The susceptibility of these populations was assessed by bioassay using DDT (4%), permethrin (1%) and deltamethrin (0.05%). Anophelesgambiae were resistant to both permethrin and DDT in the Sudanian regions but were susceptible in the central and sahelian areas and susceptible to deltamethrin at all sites except Orodara, although mortality values in some populations were close to the resistance threshold. The kdr frequency varied from 0.4 to 0.97 in populations from the Sudanian region and was lower in populations from the Sudano-sahelian and sahelian areas (0.047 to 0.54). Compared to the last survey of kdr in An. gambiae populations conducted in 2000, the kdr frequency did not differ in the S form but had increased in the M form (0.6), with an extended distribution into the Sudano-sahelian region. The frequency of kdr was also found to have increased in An. arabiensis populations (0.28), where it was formerly reported in only a single specimen. These results have practical significance for malaria vector control programs.