Stegomyia indices and pattern recognition of Aedes aegypti (Diptera: Culicidae) in selected agrogeoclimatic zones of Punjab, Pakistan

Mosquito-borne diseases especially, dengue is gaining currency nowadays in Pakistan. As there is no approved dengue vaccine available worldwide, prevention and control of vector is the only solution amid prevailing circumstances. The present study is a maiden attempt to screen indoor and outdoor breeding containers for the presence of Aedes (Ae.) aegypti larvae from selected study districts of Punjab, Pakistan i.e., Dera Ghazi Khan (DG Khan), Chakwal, and Faisalabad. A total of 384 houses from each study districts were surveyed for a calendar year. Mosquito larvae were collected, preserved, and identified using standard taxonomic keys. House Index (HI), Container Index (CI), and Breteau Index (BI) were estimated. Chi-square analysis was applied to calculate the association between Ae. aegypti larvae and breeding containers. Chakwal was identified with the highest values of Stegomyia indices (HI = 46.61 %, BI = 91.67 %, and CI = 15.28 %) followed by Faisalabad (HI = 34.11 %, BI = 68.75 % and, CI = 13.04 %) and DG Khan (HI = 28.39 %, BI = 68.23 % and, CI = 11.29 %). Earthen jars, tree holes, and water tanks were found significantly (p < 0.05) associated with the abundance of larvae irrespective of the geographical location. However, flower tubs and plastic buckets were found significantly (p < 0.05) associated in Faisalabad and Chakwal while, tyres and plastic bottles were found associated (p < 0.05) with the abundance of Ae. aegypti larvae in Faisalabad and DG Khan. These findings will help the stakeholders to devise appropriate preventive measures in combating the risk of dengue transmission.


Introduction
Mosquitoes, especially Ae. aegypti (Linnaeus) is distributed worldwide.It usually breeds in indoor and outdoor breeding sites either made naturally or artificially e.g., water tanks, tree holes, tyres, earthen jars, clay pots, flower vases, plastic sheets, plastic bottles, plant axils etc.In outdoor settings, breeding sites are created out of neglected areas of construction, and stagnant water either of rain or tap water which create favorable conditions for breeding of mosquitoes (Elsinga et al., 2018).
The risk of mosquito-borne disease transmission is influenced by multiple factors i.e., mobility of goods, human migration, climatic changes, population density, deforestation, urbanization, presence of invasive vectors and pathogens, vectoral capacity, and insufficient control strategies (Manikandan et al., 2023).As an approved vaccine against dengue has not been launched yet in the country, vector surveillance and control remain the only ways to contain dengue transmission at mass level (Nakase et al., 2023).To provide a measurable quantity of dengue virus transmission and spatial distribution, WHO recommends a routine vector surveillance to determine the risk of outbreaks and probable interventions (WHO, 2009).In these quantifiable measures, Stegomyia indices i.e., HI, CI, and BI are used to predict disease outbreaks likely caused by mosquito Ae. aegypti (Borah and Bora, 2022).These indices are estimates of number of mosquito larvae present or absent in potential breeding containers either in indoor or outdoor settings in and around each house.The calculation of these indices is not only helpful in identifying the areas at high risk of Aedes-borne diseases, but also help to formulate appropriate control measures and their implementation in areas at risk.These measures may include release of larvae predators in water holding containers, elimination of breeding habitats and breeding containers from indoor and outdoor settings, and spray of insecticides.
Therefore, to effectively control the population of mosquitoes and harness the mosquito-borne diseases, it is necessary to increase the knowledge of potential breeding habitats of Aedes mosquitoes (Souza et al., 2019).In Pakistan, despite of increase in surveillance of mosquitoes and mosquito-borne diseases, the association of potential breeding containers in indoor and outdoor settings with the abundance of Ae. aegypti larvae has not been established in the selected study areas so far.Therefore, the present investigation is a maiden attempt in establishing the association of potential breeding containers with the abundance of Ae. aegypti larvae and investigation of Stegomyia indices of the selected areas.The evidence generated here will be vital for undertaking early prevention and control interventions.

Study area and survey sites
In this proposal, three districts of Punjab (Chakwal, Faisalabad, and Dera Ghazi Khan) representative of the three different agro geo-climatic zones (North, Central and South respectively) of Punjab province were selected for sampling of Ae. aegypti larvae (Fig. 1).A pilot survey was conducted in the selected study districts to identify the potential breeding sites of the Ae.aegypti (Thrusfield et al., 2018).

Development of questionnaire
The questionnaire was developed and refined through formal and informal testing.Intra-observer variation was dealt through variability measurement through standard deviation.The data obtained was shifted to excel data sheets and statistical tools were applied to investigate any association between the prevalence of Ae. aegypti larvae and associated risk factors (Thrusfield et al., 2018).

Classification of water containers
Based on pilot survey, all containers were divided into two major categories (i) indoor containers (located inside the houses screened) and (ii) outdoor containers (located outside of the houses or in the field area near by human or animal population.The list of containers is mentioned in Table 1:

Examination of the water containers
All containers inside the houses as well as outside area were investigated for the presence of Ae. aegypti larvae.The respondents were asked about the source and use of water within each container.Any outdoor container having water without nearby water source was considered rainwater.Stegomyia indices including House Index (HI), Breteau Index (BI), Container Index (CI), and their 95 % confidence interval were calculated.The following formulae were used: The areas were considered at high risk of mosquito-borne disease transmission, if the threshold of HI and BI found more than 5 % and of CI found more than 3 %.

Collection of mosquitoes
A total of 384 houses from each study districts were surveyed for a calendar year 2018-19.A systematic random sampling technique was used.First house was randomly selected for screening.Thereafter, every 10th house was screened for the presence of potential breeding habitats (Thrusfield et al., 2018).All the contents of the containers were shifted to large trays.The immatures were picked through plastic cup.The larvae collected were transferred to the laboratory for rearing purpose.

Rearing and taxonomic identification of Aedes aegypti
Mosquito larvae were raised at temperature of 27 • C with relative humidity of 80 % and a day/night period of 12 h.The adults emerged from pupae were identified based on arrangement of white stripes using identification guides under stereomicroscope (Harrison, 2005;Rueda, 2007).

Statistical analysis
The data obtained from survey was statistically analyzed through SPSS statistical software package version 25.0 (Corp, 2017).The information collected from field survey was added to the questionnaire and shifted to the excel data sheets.The population of Ae. aegypti was considered as a dependent variable while, different types of breeding containers (plastic bottles, bags, sheets, drums, buckets, flower tubs, refrigerator trays, tree holes, plant axils, earthen jar, water tanks, clay pots and tyres) were considered as an independent variable.Chi-square test was applied to calculate the Odds Ratio (OR).The value of OR > 1 was considered significant and showed the sampling area is at high risk while OR < 1 was considered non-significant and showed no significant association between the breeding containers and population of Aedes immatures.The probability value (P-value) of less than 0.05 % was considered effective to accept or reject the hypothesis with a 95 % Confidence Interval.

Results
The presence of water holding containers indoor and outdoor at the selected study sites permits larvae of Ae. aegypti to breed which ultimately results in increased population of Aedes and transmission of arbovirus.In all the selected study districts, the Stegomyia indices HI, BI and CI were higher than the threshold level indicated by WHO (Anonymous, 2009).The HI, BI and CI of district Faisalabad were 34.11 %, 68.75 % and 13.04 % respectively.Similarly, HI, BI and CI for district DG Khan were 28.39 %, 68.23 % and 11.29 % respectively.In Chakwal, these larval indices were recorded highest among all the study districts i. e., 46.61 %, 91.67 % and 15.28 % respectively.The Stegomyia indices mapped through GIS are shown in Fig. 9.

Plastic drums
In Faisalabad district, the association of plastic drums with the abundance of Ae. aegypti larvae throughout the year was found statistically insignificant with Odds Ratio (OR) = 1.73,P-value = 0.6425 as described in Fig. 2 and Table 2. Similarly, plastic drums screened for the presence of Ae. aegypti larvae in district Dera Ghazi Khan have shown insignificant association with the abundance of mosquitoes (OR = 4.54, P-value = 0.2084) as shown in Fig. 3 and Table 3.On the contrary, plastic drums screened for the presence of larvae in district Chakwal have been found significantly associated with the production of larvae of the mosquito (OR = 2.10, P-value = 0.5515) shown in table and Fig. 4. Table 4.

Flower tubs
Statistically significant association has been found among larval population of Ae. aegypti and flower tubs screened throughout the year in district Faisalabad with OR = 2.18, P-value = 0.0165 as shown in table and Fig. 2. Similarly, significant association has been found between prevalence of Ae. aegypti larvae and flower tubs screened in district Chakwal (OR = 15.33,P-value = 0.0016) shown in table and Fig. 4.However, insignificant association has been found between flower tubs screened in Dera Ghazi Khan for the prevalence of Ae. aegypti larvae (OR = 3.74, P-value = 0.2909) as shown in table and Fig. 3.

Plastic buckets
Statistically significant association has been found between abundance of Ae. aegypti larvae in plastic bucket containers in both districts Faisalabad and Chakwal with OR = 7.69, P-value = 0.0529.and OR = 21.23,P-value = 0.0001, respectively shown in table and Figs. 2 and 4, respectively.However, insignificant association has been found with plastic buckets screened for the presence of Ae. aegypti larvae in district Dera Ghazi Khan with OR = 3.71 and P-value = 0.2951 as shown in table and Fig. 3.

Refrigerator trays
The refrigerator trays screened during the year 2018-19 in district Faisalabad were found statistically non-associated with abundance of Ae. aegypti larvae with OR = 2.71, P-value = 0.3677 mentioned in table and Fig. 2.However, significant association was found between refrigerator trays screened for the prevalence of Ae. aegypti larvae in DG Khan and Chakwal districts with OR = 8.26, P-value = 0.0410 and OR = 15.33,P-value = 0.0016, respectively shown in table and Figs. 3 and 4.

Tyres
Irrespective of seasonal variation, a significant statistical association has been found between the prevalence of Ae. aegypti larvae and tyres screened from collection sites in district Faisalabad with OR = 3.70, Pvalue = 0.0001 as shown in table and Fig. 2. Similarly, significant association has been found between tyres screened in district DG Khan and prevalence of Ae. aegypti larvae shown in table and Fig. 3 (OR = 10.56,P-value = 0.0144).However, no significant association has been observed between tyres screened in district Chakwal and abundance of larvae of the Ae.aegypti shown in table and Fig. 4 (OR = 2.56, P-value = 0.4665).

Plastic bottles
The data obtained after screening of plastic bottles from selected collection sites was statistically analyzed and no significant association found among the population of Ae. aegypti larvae and plastic bottles screened throughout the year in district Faisalabad (OR = 1.80,P-value = 0.1480) shown in table and Fig. 2. Similarly, in district Chakwal, no significant association has been observed between the larvae of Ae. aegypti and plastic bottles screened in the study year (OR = 5.16, Pvalue = 1.605) described in table and Fig. 4.However, significant association was found among the plastic bottles screened and Ae.aegypti larvae collected from district DG Khan (OR = 12.60, P-value = 0.056) as shown in table and Fig. 3.

Water tanks
Water tanks screened from collection sites in Faisalabad district were found statistically associated with the population of Ae. aegypti larvae throughout the year 2018-19 (OR = 3.23, P-value = 0.0049) as shown in table and Fig. 2. Similarly, significant association was found among Ae.aegypti larvae and the water tanks screened at selected sites in district DG Khan (OR = 14.17,P-value = 0.0027) and Chakwal (OR = 7.79, P-value = 0.0505) as shown in table and Figs. 3 and 4, respectively.

Plastic sheets
The plastic sheets that are used to cover hay stocks, silage, AC outdoors and other consumables were screened for the presence of Ae. aegypti larvae.Irrespective of the geographical locations, the data analyzed, showed no significant statistical association between the abundance of the mosquito and plastic sheets screened throughout the year in all the selected study districts i.e., Faisalabad (OR = 3.25, Pvalue = 0.4740), DG Khan (OR = 7.15, P-value = 0.0674), and Chakwal (OR = 4.24, P-value = 0.2363) given in table and Figs. 2, 3, and 4, respectively.

Plastic bags
Plastic bags that are thrown aimlessly without proper disposal in garbage, placed on the corners of the floor or roofs of the rooms were screened alongside plastic sheets.The analyses showed significant association between Ae. aegypti larvae and plastic bags screened from collection sites throughout the year in district Faisalabad (OR = 2.83, Pvalue = 0.0128) shown in table and Fig. 2. Similarly, in district Chakwal significant association has been observed between Ae. aegypti larvae and plastic bags screened throughout the study period (OR = 9.79, P-value = 0.0204) given in table and Fig. 4.However, no significant association was found between plastic bags and population of Ae. aegypti larvae in district DG Khan (OR = 5.24, P-value = 0.1549) as shown in table and Fig. 3.

Plant axil
In the field area, mosquito preferred breeding sites were also screened like plant axil.The collected mosquito population was screened for the quantification of Ae aegypti larvae and its association with the breeding site.The statistical analyses showed non-significant association between Ae. aegypti larvae and plant axil screened throughout the year in district Faisalabad (OR = 3.24, P-value = 0.1122) shown in table and Fig. 2. Similarly, no significant association was observed between Ae. aegypti larvae and plant axil screened in district DG Khan (OR = 2.10, P-value = 0.5515) given in table and Fig. 3.However, significant association was observed between plant axil and abundance of Ae. aegypti larvae while screening in district Chakwal (OR = 9.22, P-value = 0.0265) as shown in table and Fig. 4.

Tree holes
Tree holes could be another preferred site for Ae.aegypti mosquito for breeding.To assess the association between Ae. aegypti larvae and tree holes, the breeding material was screened for the presence of mosquito larvae.However, no statistical association has been found between Ae. aegypti larvae and tree holes screened for the year in district Faisalabad (OR = 1.97,P-value = 0.3447) given in table and Fig. 2.However, significant association has been observed among the tree holes and prevalence of Ae. aegypti larvae isolated from both districts DG Khan (OR = 15.83,P-value = 0.0012) and Chakwal (OR = 17.98,P-value = 0.0004) as given in table and Figs. 3 and 4, respectively.

Earthen jars
Earthenware is usually utilized to store fresh drinking water and kitchen gardening etc.These are also preferred breeding sites for the Ae.aegypti due to availability of shade, moisture, humidity, and ambient temperature.The earthen jars were screened for the presence of Ae. aegypti larvae throughout the year in district Faisalabad.The data obtained was statistically analyzed which reveals significant association in all the selected study districts i.e., Faisalabad (OR = 4.63, P-value = 0.0389), DG Khan (OR = 10.88,P-value = 0.0124), and Chakwal (OR = 8.34, P-value = 0.0395) as shown in table and Figs. 2, 3, and 4, respectively.

Clay pots
Clay pots is another earthenware which is preferred by Ae. aegypti for its breeding.The clay pots placed on the roofs with water inside them for the facilitation of birds in extreme summer specifically and throughout year usually also provide a chance to breed and increase the population.However, clay pots screened during the year 2018-19 in district Faisalabad showed no statistical association between the prevalence of Ae. aegypti larvae (OR = 2.09, P-value = 0.5338) given in table and Fig. 2. Similarly, no significant association has been observed between clay pots and Ae.aegypti larvae investigated in district DG Khan throughout the study period (OR = 6.25,P-value = 0.0999) shown in table and Fig. 3.However, significant association was observed between clay pots of Chakwal screened for the abundance of Ae. aegypti larvae throughout study period (OR = 13.97,P-value = 0.0029) as shown in table and Fig. 4. It has been reported that consistent population of the Ae.aegypti larvae was present in the supply lines or inside the sealed containers throughout winter season (Fig. 5a.and Fig. 5b.) when the conditions were not favorable which shows that roof piping might have played the role in seeding water tanks with mosquito larvae during favorable conditions i.e., rainfall etc. (Staunton et al., 2020).The association of other seasons (Summer, Autumn and Spring) has also been shown in Figs.6a, 6b., 7a., 7b., 8a., and 8b.below.

Discussion
The Stegomyia indices HI, BI and CI in all the study districts were higher than the threshold level indicated by WHO.These findings are in accordance with the study done at Dire Dawa Eastern Ethiopia where these indices were recorded as 69.10 %, 134.55 % and 54 %, respectively (Getachew et al., 2015).On the contrary, the findings are different from another study conducted somewhere else (Dom et al., 2016) which found the larvae of Ae. albopictus breeding in domestic containers.The higher prevalence of Ae. aegypti in the present study may be attributed to the presence of appropriate water holding containers for breeding of Ae. aegypti and availability of organic material in the water for larval feeding which help maintaining the life cycle (Roy et al., 2021) and ultimately enhanced the risk of dengue virus transmission in the subjected population.The findings are also in line with a recent study published in India where a positive relation of breeding containers has been reported with the prevalence of Ae. aegypti in urban areas (Devi et al., 2023).However, contrary to the findings of the present study no significant association has been reported between the breeding containers and prevalence of Ae. aegypti in Bangladesh establishing multiple reasons e.g., obtaining data at single time interval and neglecting the outdoor breeding habitats which might aided in continuous presence of dengue larvae in the human population (Haque et al., 2023).
The association of plastic drums with the abundance of Ae. aegypti larvae throughout the year was found statistically insignificant (p > 0.05).The lower productivity of plastic containers has also been observed in West Indies, which attributed the production and survival of Ae. aegypti larvae in natural habitats to variation in the presence of micronutrients in water stored in natural habitats like tree holes as compared to artificial habitats like plastic containers (Hemme et al., 2020).On the contrary positive association of plastic drums have also been reported in Canada which stated that higher productivity of plastic containers may be attributed to the presence of excessive vegetation which ultimately mixed in the plastic containers thereby providing nutrients rich in nitrate and phosphorous responsible for growth and survivability of the larvae (Dom et al., 2016).
Insignificant association (p > 0.05) of flower tubs screened in DG Khan may be attributed to the reason that rural backgrounds or less developed areas have less interest in keeping plants indoor (Hemme et al., 2020).However, increased prevalence of Ae. aegypti larvae in developed areas i.e., Faisalabad, Chakwal may be attributed to the reason that lower numbers of larvae have been found from numerous flowerpots screened from the selected areas, making it negligible for people to recognize them as a source of mosquito larvae (Walker et al., 2018;Rahman et al., 2021).
Statistically significant (p < 0.05) association of plastic buckets in both districts Faisalabad and Chakwal, respectively may be attributed to the use of these buckets as source of water storage when water supply has been cut off from government (Colombani et al., 2016;Williams et al., 2019).Similar findings have been reported in Brazil, where significant association has been found between plastic buckets and abundance of Ae. aegypti larvae.The insignificant association of plastic buckets screened in DG Khan may be attributed to the water having higher concentrations of NaCl 2 , responsible for larval killing due to high salinity (Akhter et al., 2017).Statistically insignificant (p > 0.05) association of refrigerator trays with the abundance of Ae. aegypti larvae in Faisalabad district is due to unavailability of refrigerator trays or lower numbers of refrigerators available in the screened population (Dhar-Chowdhury et al., 2016).However, significant association (p < 0.05) in DG Khan and Chakwal, respectively, which is in line with the findings of Elsinga et al. (2018) and Williams et al. (2019) attributed the higher productivity of refrigerator trays to human behavior and practice.Nowadays, affording refrigerators is very economical.Moreover, the provision of refrigerators is easy on the payment of that item in installments (Islam et al., 2019).
Significant statistical association (p < 0.05) of Ae. aegypti larvae and tyres in district Faisalabad and DG Khan may be attributed to the type of water retained in the tyres i.e., rainwater or pond water.The scientists reported that mosquitoes present in tyres having rainwater proliferate more actively as compared to pond water (Getachew et al., 2015;Ferreira-de-Lima and Lima-Camara et al., 2018).Insignificant association (p > 0.05) in district Chakwal may be attributed to the screening of tyres having water source other than rainwater, place of tyres either in shades or sunlight because Ae. aegypti preferred to breed in shady areas (Malla et al., 2020).
Insignificant association (p > 0.05) among the population of Ae. aegypti larvae and plastic bottles in district Faisalabad and Chakwal may be attributed to enough availability of water management resources and measures adopted to keep the water containers covered or dry after use (Aprisa et al., 2018;Ishak et al., 2020;Edillo et al., 2022).However, significant association (p < 0.05) at district DG Khan may be attributed to the domestic storage of tap and rainwater by the residents due to intermittent availability of drinking water supply and low rainfall.It has also been observed that people preferred rainwater for its use for laundry purpose rather tap water (Lin et al., 2018;Chumsri et al., 2018).
Water tanks screened from all the selected study districts were found statistically associated (p < 0.05).It has been reported that consistent population of the Ae.aegypti larvae was present in the supply lines or inside the sealed containers throughout winter season when the conditions were not favorable which shows that roof piping might have played the role in seeding water tanks with mosquito larvae during favorable conditions i.e., rainfall etc. (Jorge et al., 2019;Staunton et al., 2020).The role of gutters as breeding sites for Ae.aegypti larvae has also been reported previously.These gutters have high level of organic materials and stagnant water which is responsible for egg laying of mosquitoes and if somehow, while sealing the gutters, due to improper covering or sealing, small holes in seals, openings of mesh typical of water tanks having rainwater remained open would convert them to highly productive breeding habitats (Dieng et al., 2018;Chandrasiri et al., 2019).
The plastic sheets showed no significant association (p > 0.05) in any of the selected study districts.The factors responsible for lower numbers of larvae may include small surface area (Ross et al., 2019), small amount of water contained in the pits, exposed surface area to sunlight, daily cleaning of sheets by the residents, husk or hay stocks covered in a sloppy way to avoid retention of rainwater (Sinti-Hesse et al., 2019).On the contrary, positive association may be attributed to the larger surface areas surrounded by trees, retention of water, plastic sheets that were unattended by the residents, and improper cleaning (Britch et al., 2016).
Plastic bags that are thrown aimlessly, placed on the corners of the floor or roofs of the rooms or at the picnic places i.e., parks etc. were screened.The researchers attributed the significant association i.e., Faisalabad and Chakwal to social and behavioral practices of the people and level of hygienic measures adopted by the management authorities in the public places which remain unattended by the cleaning staff which served as potential source of breeding (Sultana et al., 2017).However, non-significant association i.e., DG Khan may be attributed to proper hygienic measures and continuous cleaning of public places (Cruvinel et al., 2020).Non-significant association (p > 0.05) between Ae. aegypti larvae and plant axil in district Faisalabad and DG Khan may be attributed to the scarcity of rainfall leading to smaller volume of water retention in the plant axils (Brown et al., 2019), gravid females of Ae. aegypti mosquitoes also prefer to lay eggs in more than one batch (also called "skip oviposition") and in more than one number of breeding places.However, significant association (p < 0.05) in district Chakwal may be attributed to the overlapping oviposition by multiple gravid females in same breeding container at one time (Suman et al., 2018;Gunathilaka et al., 2020).
Non-significant association (p > 0.05) between Ae. aegypti larvae and tree holes in district Faisalabad may be attributed to the presence less tree holes, more exposure to sunlight and small amount of water retention (Amarasinghe et al., 2019).However, significant association (p < 0.05) at both districts DG Khan and Chakwal may be attributed to the presence of higher number of trees (Walker et al., 2018), and the tree holes in vertical position having less exposure to sunlight as compared to horizontal position.The retention of rainwater for longer period has also been involved in aiding the larval habitat of the Ae.aegypti (Souza et al., 2019).
Earthen wares are also preferred breeding sites for the Ae.aegypti due to availability of shade, moisture, humidity, and ambient temperature.Significant association (p < 0.05) is observed in all the selected study districts.The researchers attributed the significant association of earthen jars to the characteristics of water present in the jar i.e., turbidity of water, alkalinity, pH and presence of macro and micronutrients in the water (Dom et al., 2016;Souza et al., 2019).The presence of vegetation and placement of jars in shady area have also been reported to be the probable reason for population of larvae.The nonsignificant association has also been reported from some researchers.The probable reason of non-significant association may include presence of heavy water (D 2 O), and low amount of oxygen present in the water (Kinamot et al., 2020).Clay pots in district Faisalabad and DG Khan showed no statistical association (p > 0.05).The probable reasons of non-significant association may be attributed to the clay pots having direct sunlight exposure, water replacement by residents on daily basis, frequent use of clay pots by birds (Elsinga et al., 2018;Malla et al., 2020).However, significant association (p < 0.05) was observed in Chakwal whaich may be attributed to the clay pots that were kept in shady areas, unattended, and there was retention of rainwater in it (Souza et al., 2019;Gunathilaka et al., 2020).

Conclusion
The study was a maiden attempt to investigate the prevalence of Ae. aegypti immatures and their preferred breeding containers.The values of risk indices higher than threshold may be attributed to various factors i. e., the climatic conditions of the area like excessive rainfall or availability of natural or artificial source of water.Various biotic and abiotic factors might be overlooked while collecting the larvae or biasness in information on predesigned questionnaire and most importantly, water quality in the breeding containers which is not evaluated in the present study.Apart from these limitations, the study provided he baseline data regarding the presence of Ae. aegypti vector in the population of selected

Fig. 1 .
Fig. 1.Physical map of Punjab showing sampling sites (red spots) in Chakwal, Faisalabad and DG Khan Districts of Punjab, Pakistan.

Fig. 2 .
Fig. 2. Association of different factors with prevalence of Aedes aegypti in Faisalabad isolated during different seasons.

Fig. 3 .
Fig. 3. Association of different factors with prevalence of Aedes aegypti in DG Khan isolated during different seasons.

Fig. 4 .
Fig. 4. Association of different factors with prevalence of Aedes aegypti in Chakwal isolated during different seasons.

Fig. 5a .Fig. 5b .
Fig. 5a.Percent association of winter season with the outdoor breeding containers for the presence of aedes aegypti in selected agrogeoclimatic zones of punjab, Pakistan.

Fig. 6a .Fig. 6b .
Fig. 6a.Percent association of summer season with the outdoor breeding containers for the presence of Aedes aegypti in selected agrogeoclimatic zones of Punjab, Pakistan.

Fig. 7a .Fig. 7b .
Fig. 7a.Percent association of autumn season with the outdoor breeding containers for the presence of aedes aegypti in selected agrogeoclimatic zones of punjab, Pakistan.

Fig. 8a .Fig. 8b .Fig. 9 .
Fig. 8a.Percent association of spring season with the outdoor breeding containers for the presence of aedes aegypti in selected agrogeoclimatic zones of punjab, Pakistan.

Table 1
Classification of breeding containers based on their locations i.e., indoor/ outdoor.

Table 2
Association of different factors with prevalence of Ae. aegypti in Faisalabad isolated during different seasons in the year 2018-19.

Table 3
Association of different factors with prevalence of Ae. aegypti in Dera Ghazi Khan isolated during different seasons in the year 2018-19.

Table 4
Association of different factors with prevalence of Ae. aegypti in Chakwal isolated during different seasons in year 2018-19.