Ecology of Anopheles Mosquito Larvae in Different Ecological Zones in Ghana

Background: Understanding the ecology of larval malaria mosquitoes is important in a changing environment is important in developing effective control tools or programmes. This study characterized the breeding habitats of Anopheles mosquitoes in rural communities in different ecological zones in Ghana during the dry and rainy seasons. Methods: The spatio-temporal distribution, species composition, and abundance of larval Anopheles mosquitoes in breeding habitats were studied in 3 ecological zones of Ghana. These were Anyakpor (Coastal Savanna area), Duase (Forest area), Libga, Pagazaa, and Kpalsogu (Sahel Savanna area). Larvae were collected using standard dippers and were raised in the insectary for identication. Results: Out of a total of 7,984 mosquito larvae collected, 2,152 (27.26%) were Anophelines and were more abundant in the rainy season (70.82%) than in the dry season (29.18 %). The Anophelines were made up of 2,128 (98.88%) An. gambiae s.l., 16 (0.74%) An. rupees and 8 (0.37%) An. pharoensis. In Anyakpor and Duase, dug-out wells were the most productive habitat in the dry (1.59 larvae/dip and 1.47 larvae/dip) and rainy seasons (11.28 larvae/dip and 2.05 larvae/dip). The most productive habitats in Kpalsogu were natural ponds in the dry season (0.89 larvae/dip) and swamps in the rainy season (2.57 larvae/dip). In Libga, the most productive habitats were drainage ditches in the dry season (0.30 larvae/dip) and furrows in the rainy season (1.83 larvae/dip). The most productive habitats in Pagazaa were puddles (1.44 larvae/dip). Anopheles coluzii was the most abundant sibling species in all the ecological zones except Libga in the sahel savannah area where An. gambiae s.s was the most abundant. Anopheles melas and An. arabiensis were encountered only in the coastal savannah and the sahel savanna areas respectively. Larval habitat types inuenced the presence of larvae as well as larval densities (p < 0.001). The land-use type affected the presence of Anopheles larvae (p = 0.001), while vegetation cover inuenced larval densities (p < 0.05). Conclusion: The study revealed that the abundance of Anopheles breeding habitats and hence Anopheles larvae are closely associated with anthropogenic activities. Regulating such activities will lead to a signicant reduction in Anopheles breeding habitats. mosquito larvae in different ecological zones in Ghana. The availability of larval habitats, their productivity, and distribution among different zones in Ghana was studied. The spatio-temporal distribution and species composition of larval malaria vectors were investigated. study was conducted in 5 locations in three ecological landscapes of Ghana – the the and the Sahel savannah zones. Anyakpor in the It is a rural coastal about of Ada in southern and has a equatorial with the and maximum reaching 33°C. It has a bimodal rainfall pattern with a long a It has savannah type of for smaller habitats such as footprints and hoof prints were considered to be one dip. A test for normality of the larval density distribution using the Shapiro-Wilk test showed a non-normal distribution. The density of Anopheles mosquito larvae was compared among the various breeding habitats and study sites. The Mann-Whitney U and the Kruskal-Wallis test was used to test the associations between continuous and categorical variables. The Chi-square and Fisher's exact tests were used to test the association between two categorical variables. Logistic regression was used to assess the association between the habitat characteristics with categorical data and the presence of Anopheles larvae. Nested generalized linear mixed models with sites nested within ecological zones were used to model the effect of habitat characteristics on larval densities. All statistical analyses were conducted in STATA v15 (StataCorp. 2017. Stata Statistical Software: Release 15. College Station, TX: StataCorp LLC).


Background
Anopheles mosquitoes are important vectors that transmit diseases, including malaria, lymphatic lariasis among others [1]. The distribution and abundance of adult Anopheles mosquitoes are predicated on the presence and productivity of larval breeding habitats [2]. Species of Anopheles gambiae complex prefer to breed in shallow water collections that are open to sunlight [3]. Their breeding habitats can be varying sizes of water bodies that are natural or man-made, permanent or temporary, freshwater, or saline [2,4]. Anopheles funestus, on the other hand, prefer to breed in shady permanent or semi-permanent water bodies, usually with oating or emergent vegetation such as found in swamps, marshes, and edges of streams [5,6].
The choice of oviposition sites of mosquitoes is in uenced by a myriad of environmental factors, which include climatic components such as temperature, rainfall, vegetation, salinity and turbidity of the water, the size of the habitat, and the amount of sunlight [2]. The temperature of larval habitats can in uence larval development, pupation rate, and time, as well as larval survivorship [2,7]. Variation in rainfall patterns or seasonal changes can also affect the availability of larval habitats as well as productivity [3].
Vector control is key to the elimination of vector-borne diseases such as malaria [8,9]. Even though the most used vector control methods, long-lasting insecticide nets (LLINs) and indoor residual spraying (IRS) have reduced the transmission of malaria in Africa [10,11], these methods have not been successful in malaria eradication because of the emergence and rapid spread of resistance in mosquitoes [12][13][14]. Also, the use of LLINs and IRS which target indoor biting and indoor resting mosquitoes have driven behavioural changes in the Anopheles mosquito from indoor, late-night biting to early biting times when humans might be unprotected outside [15,16]. Nevertheless, larval source management or source control could provide an additional valuable tool for the control of malaria vectors [17]. To assess the feasibility of larval control or larval source management, there is the need to assess the abundance of different types of habitats, measure the productivity in each habitat type [18], and also know how these different habitat types are formed, and how they interact with the society.
The presence and densities of mosquito larvae and consequently, the number of competent adult malaria vectors are regulated by a variety of ecosystem processes interacting at different levels and spatio-temporal scales. These include the presence of water, aquatic plants that protect larvae from predators and serve as detritus that support microbial communities, which, in turn, serves as food for mosquito larvae [19]. Changes in the structure of the ecosystem can have a considerable impact on mosquito populations and species distribution. As such, studies on the ecology of larval habitats should include a landscape context [2,3,20,21]. Landscape features such as topography, land cover and land use in uence the formation, distribution, and microclimate conditions of larval habitats which, in turn, in uence the distribution of adult Anopheles vectors [2,[22][23][24][25]. Human activities can affect habitat distribution and stability through landscape changes such as deforestation, irrigations, and agricultural practices [2].
There are three main ecological zones in Ghana -the Coastal Savannah in southern Ghana, the Forest in the middle of Ghana, and the Sahel Savannah in northern Ghana. These ecological zones affect the distribution of habitats and importantly, species composition [26]. The coastal savannah and forest zone have a bimodal rainfall pattern, allowing for two peaks of malaria transmission while the Sahel zone has a unimodal rainfall pattern making malaria transmission seasonal.
The ecology of larval mosquitoes has implications for vector control; hence there is the need to understand the productivity and dynamics of larval habitats in the changing environment in efforts to model and predict the abundance of adult mosquitoes and ultimately develop effective control tools or programmes [2,27]. The aim of this study, therefore, was to investigate the ecology of Anopheles mosquito larvae in different ecological zones in Ghana. The availability of larval habitats, their productivity, and distribution among different zones in Ghana was studied. The spatio-temporal distribution and species composition of larval malaria vectors were investigated.

Study sites
The study was conducted in 5 locations in three ecological landscapes of Ghana -the coastal savannah, the forest, and the Sahel savannah zones. Anyakpor (5° 46'51.96 "N 0° 35'12.84 "E) was the site in the coastal savannah area. It is a rural coastal community about 5 km west of Ada Foah, in southern Ghana, and has a dry equatorial climate with temperatures ranging from 23°C -28°C throughout the year and maximum temperatures reaching 33°C. It has a bimodal rainfall pattern with a long rainy season from April to June and a short rainy season from September to November. It has coastal savannah type vegetation.
Duase (6° 32'3.05 "N 1° 14'42.22 "W) was the site located in the forest zone. It is a rural community close to Konongo. It has a wet semi-equatorial climate characterized by two distinct rainy seasons with a long rainy season from May to July and a short rainy season from September to November. The mean annual temperature and relative humidities are 26°C and 77% respectively. This study was undertaken during the rainy and dry seasons of 2019.

Larval Habitat Characterization
All larval habitats in each site were classi ed as natural or man-made. Natural habitats included swamps, streams, and natural ponds whiles man-made habitats included drainage ditches, foot and hoof prints. Land-use type was classi ed based on the natural vegetation and activities taking place on the land where the larval habitat was found. These include forest for sites with high canopy cover, farmland for cultivated areas; pasture for grazing areas; shrubland for bushy areas with short trees, roads, swamps, and compound or home for places with human settlement. The length and width of each habitat were measured and recorded in metres. The percentage of vegetation cover was also recorded. The vegetation cover was categorized as follows: zero if vegetation were not present in the habitat, ≤ 24% of surface coverage, 25-49%, 50-74%, and 75-100% of surface coverage [28].

Larval Sampling and Densities
Larval sampling was done for all potential breeding sites by the standard dipping method using the WHO 350 ml standard dipper. The size of each habitat was grouped as ≤ 1 m, >1 m -2 m, >2 m -5 m, >5 m -10 m, >10m -100 m, and > 100 m and a maximum of 2, 4, 6, 10, 50, and 150 dips were taken respectively (i.e., depending on the size of the habitat) as described by Gouagna and Mereta [28,36]. For habitats with much smaller sizes such as hoof prints and footprint, a ladle was used to collect the samples. Larvae collected were classi ed as early instars (L1 and L2) or late instars (L3 and L4). The number of larvae and pupae were recorded, and the larval density was estimated as the ratio of the number of larvae collected per dip [29][30][31][32].

Mosquito Species Identi cation
Anopheline larvae specimens were transported to the insectary of the Department of Medical Microbiology, University of Ghana, where they were bred into adults. The larvae were fed on Tetramin® sh meal and maintained at 27°C ± 2°C. Emerged adult mosquitoes were morphologically identi ed under a stereomicroscope using the taxonomic keys by Gillies and Coetzee [33]. Anopheles gambiae s. l. were further identi ed to sibling species and molecular forms using rDNA PCR [34] and PCR-RFLP [35] analysis, respectively.

Data Analysis
Descriptive analysis was done to compare the abundance of the various habitat types and larval densities in the different study sites (ecological zones) and seasons. Larval densities were calculated by dividing the total number of larvae collected by the total number of dips taken. The total number of dips for smaller habitats such as footprints and hoof prints were considered to be one dip. A test for normality of the larval density distribution using the Shapiro-Wilk test showed a non-normal distribution. The density of Anopheles mosquito larvae was compared among the various breeding habitats and study sites. The Mann-Whitney U and the Kruskal-Wallis test was used to test the associations between continuous and categorical variables. The Chi-square and Fisher's exact tests were used to test the association between two categorical variables. Logistic regression was used to assess the association between the habitat characteristics with categorical data and the presence of Anopheles larvae. Nested generalized linear mixed models with sites nested within ecological zones were used to model the effect of habitat characteristics on larval densities. All statistical analyses were conducted in STATA v15
Larval habitat types, the presence, and densities of Anopheles larvae The presence of Anopheles larvae was dependent on the type of habitat present ( 2 = 41. 3651; df = 10; p < 0.0001). Even though there was a signi cant increase in the number of habitats during the rainy season ( 2 = 91.3295; df = 10; p < 0.0001) (Additional le 1) compared to the dry season, the proportion of habitats that had the presence of Anopheles larvae were not signi cantly different between the two seasons ( 2 = 0.0051; df = 1; p = 0.943) (Additional le 2). In the dry season, 29.29% (41/140) of the habitats had Anopheles larvae present while in the rainy season, 29.63 % (72/243) of the habitats were positive for Anopheles larvae. In all, dug-out wells were the most inhabited with Anopheles larvae during the dry season (39.02%, 16/41) followed by man-made ponds (21.95%, 9/41) whereas during the rainy season man-made ponds were the most inhabited habitat (30.56%, 22/41) followed by swamps (16.67% 12/72) ( Table 2).
The type of habitat present was associated with An.gambiae s.l larval density ( 2 = 29.593; d.f =10; p < 0.001) (Additional le 3). Even though the mean An.gambiae s.l larval density in the rainy season (1.49 larvae/dip) was slightly higher than in the dry season (1.15 larvae/dip), the difference was not signi cant (z = -0.232; p = 0.8166). In Anyakpor, the most productive habitat types were dug-out wells during both the dry (1.6 larvae/dip) and rainy (11.28 larvae/dip) seasons ( Table 5). The preferred breeding habitat in Kpalsogou were natural ponds during the dry season (0.89 larvae/dip) and swamps (2.57 larvae/dip) during the rainy season. In Libga, drainage ditches were the only habitat type present in the dry season with the mean larval density of 0.3 larvae/dip, whiles the only productive habitats were furrows in the rainy season (1.83 larvae/dip). There were no productive habitats in Pagazaa during the dry season, but in the rainy season, the most productive habitat type were puddles (1.44 larvae/dip). In Duase, the most productive habitat type was dug-out wells both in the dry (1.47 larvae/dip) and the rainy season (2.05 larvae/dip) ( Table 5).
Abundance and distribution of Anopheles larvae in the different ecological zones A total of 7,894 mosquito larvae were collected during this study. Out of this number, 2,152 (27.26%) were Anophelines whiles 5,742 (72.74%) were culicines.
Anyakpor in the coastal savannah area had the highest abundance of An. gambiae s.l larvae (1,286) with 343 (26.67%) occurring in the dry season and 943 (73.33%) in the rainy season (Table 3). During the dry season in Duase, situated in the forest area 30.17% of An. gambiae s.l larvae were collected, while 69.83% were collected in the rainy season. Kpalsogou and Libga in the Sahel savannah recorded 39.00% and 35.79% of the An. gambiae s.l larvae in the dry season, while the rainy season contributed to 61.00% and 64.21%, respectively. In Pagazaa, also in the Sahel savannah An. gambiae s.l larvae were found only in the rainy season (Table 3) and none during the dry season.
Habitat characteristics, the occurrence, and densities of Anopheles larvae Fifty-ve percent (55%) of all larval habitats found around homes or compounds contained Anopheles larvae. The odds of nding Anopheles larvae in any habitat was twice higher if the vegetation cover was less than 24% (OR = 2.24 [1.02, 4.93]; p = 0.045) ( Table 6). As the vegetation cover increases, the density of Anopheles larvae decreases (B= -0.016 [-0.28, 0.003]; p =0.015) (Additional le 4). The present study, again, showed that Anopheles larvae have the preference to co-habit with Culicine larvae. The likelihood of encountering Anopheles larvae in a breeding habitat was over three times higher when culicines are present (OR = 3.13 [1.75,5.59]; p < 0.01) ( Table 6).

Discussion
Understanding the ecology of Anopheline larvae in a changing environment is crucial for the development and successful implementation of targeted control measures [36,37] to supplement current adult vector control tools. In this study, the distribution of Anopheles breeding habitats in rural communities in the different ecological zones of Ghana has been characterized. These communities were Anyakpor in the coastal savannah area and Duase in the Forest area. Three communities were selected in the Sahel savannah area; Kpalsogu which is an IRS site, Libga where IRS was stopped after 2014, and Pagazaa which has never been under the IRS intervention. The study revealed differences in the abundance and distribution of Anopheles breeding habitats in the different ecological zones. Although man-made ponds were the most abundant habitat type, dug-out well the most productive for Anopheles mosquito larvae. Anopheles larvae also preferred to breed in small habitats while increasing vegetation cover reduced Anopheles larval densities.
The common habitat types were man-made ponds, natural ponds, drainage ditches, and swamps. Other habitats such as tyre tracks and puddles were formed usually during the rainy season when rainwater collects on untarred roads [32]. Such habitats are temporal. The formation and abundance of habitat types are greatly in uenced by the nature of the topography of the land as well as the land-use [5]. The habitats encountered were mostly associated with anthropogenic activities. This explains why communities that practice irrigation farming, Anyakpor, and Kpalsogou had the highest number of habitats. Because it is a low-lying area with a high-water table, farmers in Anyakpor in coastal Ghana dig wells and ponds to get underground water for irrigation purposes. As a result, wells and man-made ponds were the only breeding habitats encountered in Anyakpor with man-made ponds being the most abundant. In the dry season, when the water table reduces, some of these wells and ponds which serve as breeding habitats dry up.
In contrast to Anyakpor, large dams have been constructed in Kpalsogu and Libga to provide water for both domestic and irrigation purposes during the severe dry season experienced in the Sahel savannah region of Ghana. During this season, most of these dams and other large ponds dry up forming small collections of open sunlit water bodies which are more conducive for Anopheles breeding. These ndings are similar to previous studies [38][39][40] which reported that during the dry season, drying water bodies have the most abundant mosquito larvae before the water body becomes completely dried up. On the other hand, during the rainy season, water from the dams and ponds over ow and cause huge swamps and marshes on livestock grazing elds making swamps the most abundant breeding habitats in Kpalsogou. Also, in Libga, and Pagazaa, water from natural ponds is diverted through drainage ditches and furrows onto farmlands, usually rice farms.
Duase and Kpalsogou had the most diverse breeding habitat types, which included eight (8) out of the eleven (11) habitat types encountered. The persistence of breeding habitats during both dry and rainy seasons in the forest and coastal savannah areas account for perennial malaria found within these sites while seasonal variations are observed in the Sahel savanna areas [41,42].
In all, the most abundant habitat type was man-made ponds. Natural ponds are the only habitat type found in all the study sites. Hoofprints were found only in the sahel savannah zone where livestock are left to graze on swampy pastures. Most of the habitats were man-made and found on farmlands. This emphasizes the importance of human activities and, for that matter, land-use in the creation of Anopheles breeding habitats and the impact they have on malaria transmission.
The reason why most of the breeding habitats are found on farmland can be attributed to the practice of irrigation. Irrigation provides ideal breeding habitats for Anopheles vectors, and this study corroborates with that of Appawu et al. [43] who found that irrigated elds generate large numbers of mosquitoes. It is important to note that agrochemicals used on these farms end up polluting the water sources which serve as breeding sites on these farms, thereby leading to the development and spread of insecticide resistance by exposing mosquito larvae to high or sub-lethal doses of agrochemicals [44][45][46][47].
Even though 163 of the 383 habitats were inhabited by mosquito larvae, only 29.5% of the had Anopheles larvae were present in them. The presence of Anopheles larvae also varied signi cantly based on the habitat type (p < 0.0001). The variation in the presence of Anopheles larvae may be due to the differences in the physical, chemical, and biological properties as well as the quality of the water present in the various habitats [48]. These properties directly in uence the choice of oviposition sites by gravid females and also in uence the development and survivorship of larvae [49][50][51][52]. Anopheles larvae were mostly present in man-made ponds and dug-out wells. This further establishes the role of human activities on the presence and distribution of Anopheles vectors [50].
Dug-out wells were the preferred breeding habitats in Anyakpor and Duase, both in the dry and rainy seasons with an increased larval density in the rainy season. In the Sahel savannah area, the preferred breeding habitats for Anopheles larvae varied in each site and for each season. With all the sites combined, dug-out wells remained the most productive habitat just as observed by Afrane and co [53]. Dug-out wells were also the most productive habitats both in Anyakpor in the coastal savannah area and Duase in the forest area. In the Sahel savannah area, the most productive habitats were swamps in Kpalsogu, furrows in Libga, and puddles in Pagazaa. It is worthy to note that these habitats that had the highest Anopheles larval densities hence major contributors to the Anopheles population were man-made or associated with anthropogenic activities which as reported by other studies in Ghana [37,53] and other parts of Africa [31,50,54,55].
Anopheles larvae were always present and also at higher larval densities in habitats less than 10 m 2 in size. They were not found in habitats that had a perimeter greater than 100 m 2 . More than half of the habitats found around human settlements or homes harboured Anopheles larvae and the chances of nding Anopheles larvae in habitats close to human settlements where it was easy to nd the next blood meal source increased threefold (OR = 3.06, p = 0.050). These ndings corroborate with studies from Kenya [56,57] that also suggest that since An. gambiae s.l. are closely associated with humans, they will make use of the closest habitat for oviposition when they become gravid [56,57]. Choosing habitats close to human settlements where An. gambiae may have taken a blood meal is also an evolutionary strategy to conserve energy [48,58].
This study also revealed that Anopheles larvae were predominantly present in breeding habitats with vegetation cover of less than 24%. Similar ndings have been observed in Kenya [59][60][61]. Low vegetation cover allows the habitat to be more exposed to sunlight, a suitable preference for ovipositing mosquitoes [5].
Also, adequate exposure to sunlight warms the water to suitable temperatures as temperature is a key factor that also in uences larval development and survival [2,5,7,62]. Inadequate exposure to sunlight, caused by high vegetation cover affects the photosynthetic e ciency of algae biomass which serves as food for mosquito larvae [63]. Evidently, from this study, as the percentage of vegetation cover increased, the density of Anopheles larvae decreased (B = -0.016, p = 0.015). This is in line with studies conducted in Ethiopia [32,39]. As vegetation cover increases, the amount of sunlight reaching the habitat is limited. The study also showed that Anopheles larvae preferred to live in sympatry with culicine larvae just as observed by Djamouko-Djonkam and colleagues in Mali [52]. This study, again, showed that the abundance of Anopheles larvae increased in the rainy season due to the formation of new breeding habitats [32].
In Anyakpor in the coastal zone, the predominant species was An. coluzzii (60.82 %) similar to what was reported by Kudom [64] and Fossog et al., [65] whose model shows An. coluzzii to dominate the coastal line of Africa. They were mostly found in man-made ponds dug-out wells and concrete wells. Anopheles gambiae s.s contributed to 20.62% of the species in Anyakpor. Compared to An. gambiae s.s., both inland and coastal An. coluzzii are known to have a higher tolerance to salinity [65,66]. Anopheles melas in Anyakpor were found only in dug-out wells and man-made ponds because these habitats are fed by salty underground water which An. melas prefer to breed in, unlike concrete wells which were fed by rainwater. Anopheles coluzzii and An. gambiae s.s were the only species found in Duase, in the forest zone, with An. coluzzii being the dominant species which is contrary to the ndings of other studies [41,67,68]. This could be as a result of the deforestation which is caused by rapid urbanization. Rapid deforestation affects climatic conditions, and this might be causing Duase to become drier, thereby making An. coluzzii to thrive there. Anopheles coluzzii were found in drainage ditches, natural ponds, and tyre tracks. Anopheles gambiae s.s were found in dug-out wells, puddles, tyre tracks, and drainage ditches. Anopheles coluzzii was also the dominant species in Kpalsogou followed by An. gambiae s.s. and An. arabiensis. On the other hand, An. gambiae s.s. was the dominant species in Libga. In Pagazaa, An. coluzzii and An. arabiensis were co-dominant. Anopheles arabiensis were found only in the Sahel savanna area because of its dry sub arid environments [69]. Anopheles arabiensis was predominantly found in swamps and furrows. Furrows were mostly present on rice farms. Anopheles arabiensis prefer to be zoophilic even though they can also be anthropophilic [70] and this can account for the reason they were mostly found in swamps since swamps in the Sahel savannah areas mostly serve for grazing cattle. As shown by this study, An. coluzzii and An. gambiae s.s are known to live in sympatry in most parts of Ghana [64]. However, An. coluzzii predominates in the Coastal and Sahel savannah regions of Ghana [67,68,71]. The dominance of An. coluzzii in the coastal regions of Ghana has been attributed to permanent habitats created on irrigation farms [67,68]. Anopheles. coluzzii is usually seen to be breeding in large permanent habitats, but in this study, they were also found in small temporary habitats such as hoofprints. Studies done by Edillo et al., (2006) showed that An. coluzzii preferred different breeding habitat types to An. gambiae s.s. and An. arabiensis but this study showed otherwise. Anopheles gambiae s.s and An. coluzzii were found in all 11 habitat types.
This study involved eld surveys carried out under natural conditions and external factors. These factors could have in uenced the presence and development of mosquito larvae in the various habitats. Studies on the ecology of Anopheles mosquito larvae are methodologically challenging as such has several limitations. The standard dipping technique adapted for this study was not suitable for sampling in small habitats especially footprints and hoofprints. Consequently, the larval densities in these habitats may have been overestimated in this study. In this study, larval density was used to express the productivity of a larval habitat [20,30] instead of the number of pupae or adult mosquitoes that emerged from the habitat [56,72].
This study provides a baseline insight into the development of integrated larval source management suitable for the larval habitats in the various study sites.
This study revealed that irrigated farms contribute to higher populations of malaria mosquitoes as this system of agriculture creates a lot of habitats suitable for Anopheles mosquito larvae. This made communities that practice irrigation farming have a higher abundance of Anopheles larvae. This study, again, revealed that human activities contribute greatly to the presence and abundance of Anopheles mosquito larvae. This implies that changes in agricultural methods and environmental management would be extremely appropriate in controlling malaria transmission. Considering that An. gambiae s.l prefers to breed in small habitats, habitat modi cation would be a suitable method of larval source reduction. Other integrated larval source management approaches such as water and environmental management and biological control methods are feasible. In communities that rely on the same water collections for domestic purposes, and in situations where the water collections cannot be drained, microbial bio-larvicides may be used.

Conclusion
This study showed that the presence and availability of Anopheles breeding habitats varied among the study site, hence, ecological zones. The abundance of breeding habitats was in uenced by rainfall as well as the topography of the land, as more habitats are created during the rainy season and this, in turn, increases the abundance of Anopheles mosquitoes. Even though man-made ponds were the most abundant breeding habitat types, dug-out wells were the most productive. This established the role of human activities in creating suitable breeding habitats for An. gambiae s.l. Anopheles gambiae s.l larvae preferred to breed in small habitats that also have vegetation cover less than 24%. Anopheles larvae also preferred to live in sympatry with culicine larvae. Anopheles coluzzii was the predominant species in all the study sites. They also were usually in sympatry with An. gambiae s.s.

Declarations
Ethics approval and consent to participate Ethical approval for this study with protocol identi cation number CHS-Et/M2 -5.5/2019-2020 was obtained from the Ethics and Protocol Review Committee of the College of Health Sciences, University of Ghana.

Consent for Publication
Not applicable Availability of data and materials All datasets generated and/or analysed during the current study are available on request.

Competing interests
The authors declare that they have no competing interests.

Funding
This study was supported by a grant from the National Institute of Health (NIH: R01 A1123074). The funders had no role or in uence on the design of this study, the collection, analysis, and interpretation of the data collected as well as in writing this manuscript.
Authors' contributions IAH, SKA, AOF, and YAA conceived and designed the study. IAH and YAA were responsible for designing and coordinating the entomological surveys. IAH was responsible for data collection. YAA, SKA, and AOF supervised the data collection. IAH and BAM were responsible for the data analysis and drafted the rst manuscript. All authors read and approved the nal copy of the manuscript.