J Arthropod-Borne Dis, December 2015, 9(2): 253–266 S H Nikookar et al.: Fauna and Larval … 253 Original Article Fauna and Larval Habitat Characteristics of Mosquitoes in Neka County, Northern Iran Seyed Hassan Nikookar 1, *Seyed Hassan Moosa-Kazemi 2, Mohammad Reza Yaghoobi- Ershadi 2, Hassan Vatandoost 2,3, Mohammad Ali Oshaghi 2, Abolfazl Ataei 2, Mehdi Anjamrooz 2 1Health Sciences Research Center, Department of Medical Entomology and Vector Control, Faculty of Health, Mazandaran University of Medical Sciences, Sari, Iran 2Department of Medical Entomology and Vector Control, School of Public Health, Tehran University of Medical Sciences, Tehran, Iran 3Institue for Environmental Research, Tehran University of Medical Sciences, Tehran, Iran (Received 29 May 2012; accepted 12 Oct 2013) Abstract Background: Ecological studies on mosquitoes are very important in vector control programs. There are a few studies about the ecology of mosquitoes in northern Iran. This study was carried out to detect fauna and larval habitat characteristics of mosquitoes. Methods: This study aimed to determine fauna and the ecology of mosquitoes in Neka County, Mazandaran Prov- ince, northern Iran from April to December, 2009. The larval collection was conducted using standard dipper, and the characteristics of larval habitat were investigated based on degree of transparency of water, type of water (stag- nant or running), plant vegetation, sunny or shady, temperature and altitude of the natural or artificial breeding places. Results: The mosquito larvae were collected from 72 habitats and identified using systematic keys. Nine species of mosquitoes were identified: Anopheles claviger (0.31%), An. maculipennis (0.54%), An. plumbeus (10.28%), An. superpictus (0.01%), Culiseta annulata (1.07%), Cs. longiareolata (8.91%), Culex mimeticus (0.03%), Cx. pipiens (63.99%), and Ochlerotatus geniculatus (14.85%). The range of temperature in the larval habitats was 19.6–22.5 °C. Significant difference was observed in the rate of temperature among the species in the larval habitats (P< 0.05). A checklist of mosquitoes including seven genera and 32 species has been provided for Mazandaran Province. Conclusion: The most dominant species were Cx. pipiens. They were collected from the larval habitats like Border Rivers, ponds, rain water pools, discarded tires and tree holes. Culiseta annulata was included to the checklist of mosquitoes in Mazandaran Province. Keywords: Larval habitat, Culicidae, Ecology, Iran Introduction According to the latest classification of Culicidae, the family includes two subfami- lies, 11 tribes, 112 genera and 3537 species. The subfamily Anophelinae has three genera and Culicinae has 109 genera (Harbach 2007). Many of the world's major diseases such as Malaria, West Nile, Sindbis, Dirofilariasis, Dengue fever, Yellow fever, Rift valley fever are transmitted by mosquitoes. West Nile and Sindbis viruses have been reported in northern parts of Iran (Naficy and Saidi 1970, Saidi et al. 1976). The mosquito borne filarial worms, Dirofilaria immitis and D. repens, Setaria (S. labiatopapillosa, S. digitata and S. equina) and Diptalonema evansi (camel filariasis) have been reported in Iran (Siavashi and Massoud 1995, Eslami 1997, Azari-Hamidian et al. 2007, Oryan et al. 2008). Anopheles macu- lipennis and Culex theileri were reported as vectors of Setaria labiatopapillosa and *Corresponding author: Dr Seyed Hassan Moosa- Kazemi, E-mail: moosakazemi@tums.ac.ir http://jad.tums.ac.ir Published Online: March 11, 2015 J Arthropod-Borne Dis, December 2015, 9(2): 253–266 S H Nikookar et al.: Fauna and Larval … 254 Dirofilaria immitis respectively, in Ardebil Province (Azari-Hamidian et al. 2009). Dengue fever cases were reported in Tehran, imported from Malaysia, India and Thailand. In addition, six DF cases were reported from Sistan and Baluchistan Province in southeast Iran (Chinikar et al. 2010, 2012). The systematic key of Anopheles species in Iran was reported by Shahgudian (1960). Many studies have been carried out about the genus of Culex and its medical im- portance (Ghaffary 1956, Lotfi 1970, Lotfi 1973, Lotfi 1976, Zaim et al. 1984a, 1984b, 1985, 1986). The results of these studies have led to the publishing of the Iranian Culicinae checklist (Zaim and Cranston 1986). The role of mosquitoes in malaria transmis- sion and its geographical distribution have been studied by Saebi (1987). The fauna and ecological characteristics of Culicidae with emphasis on their medical importance were conducted in Isfahan and Hormozgan Prov- inces as well as Chabahar County (Mousa- kazemi et al. 2000, Moosa-Kazemi et al. 2005, 2009) and Guilan Province (Azari- Hamidian et al. 2002a). Moreover, the study on the genus of Aedes is very scattered in the northern parts of the Iran. Azari-Hamidian et al. (2002a) stressed the occurrence of Oc. caspius, Oc. echinus, Ae. vexans and Oc. geniculatus in Guilan Province, northern Iran. More than 30 culicidae species reported from Guilan province (Azari-Hamidian et al. 2003a). Distribution of larval habitat of Cx. pipiens was reported by Dehghan et al. (2011) in central Iran. Among the 467 species of Anopheles identified in the world (Harbach 2007), about 70 Anopheles species are ma- laria vectors in which about 40 species are important vectors (Service 1993). Anopheles maculipennis complex, one of the most im- portant vectors of malaria, was reported in northern Iran (Eslami 1997, Azari-Hamidian 2007a). Anopheles maculipennis, and Cx. pipiens were reported as dominant species in Isfahan Province, central Iran (Mousakazemi et al. 2000a) and in East Azarbaijan and Ardebil Provinces (Abai et al. 2007, Azari- Hamidian et al. 2009). By now, seven species of malaria vectors have been reported in Iran (Vatandoost et al. 2009a). They include: An. culicifacies, An. dthali, An. fluviatilis, An. maculipennis, An. sacharovi, An. stephensi, and An. superpictus (Edrissian 2006, Vatandoost et al. 2008, Vatandoost et al. 2009a, Moosa-Kazemi et al. 2010). Anopheles pulcherrimus has been identified as potential vector in Iran using serological method (Zaim et al. 1993). Alt- hough oocyte of Plasmodium was found in the stomach of An. multicolor but sporozoite never found in its salivary gland and not considered as the malaria vector (Eshghy 1977). Anopheles hyrchanus was found as a malaria vector using the PCR method in northern Iran (Dinparast Djadid et al. 2009). At present, 64 species, three subspecies and seven genera of Culicidae have been re- ported from Iran (Azari-Hamidian 2007a). The first study about the mosquitoes in Mazandaran Province was carried out by Gutzevich (1943) and Zolotarev (1945). The occurrence of the species of An. maculipennis, An. melanoon (An. subalpinus), An. Super- pictus, An. sacharovi and An. messeae have been reported by Zolotarev (1945). How- ever, Gutsevich (1943) reported the species of An. pulcherimus, Ae. vexans, Oc. geniculatus and Cq. richiardii in Mazandaran Province. Subsequently, Garrett Jones (1951) reported the species of An. claviger, An. maculipennis, An. plumbeus and An. hyrcanus in Klardasht. Institute of Parasitology and Malariology (1953) reported the species of An. marteri in Ramsar. Dow (1953) reported the species of An. maculipennis, An. melanoon, An. subalpinus (synonym of melanoon), An. hyrcanus, An. pseudopictus, An. superpictus, An. sacharovi, Cx. pipiens, Cx. mimeticus and Cx. tritaeniorhynchus. However, Faghih (1969) stressed the occurrence of the species http://jad.tums.ac.ir Published Online: March 11, 2015 J Arthropod-Borne Dis, December 2015, 9(2): 253–266 S H Nikookar et al.: Fauna and Larval … 255 of An. melanoon, and An. messeae. Minar (1974) reported the species of An. Macu- lipennis, An. pseudopictus, Oc. pulcritarsis. However, Zaim et al. (1984a) cited the species of Ae. vexans, Oc. echinus, Oc. geniculatus and Oc. pulcritarsis. Zaim (1987) cited the species of Ae. vexans, Oc. echinus, Oc. geniculatus, Oc. pulcritarsis, Cx. perexiguss, Cx. theileri, Cx. mimeticus, Cx. Tritaenior- hynchus, Cx. hortensis, Cx. pipiens, Cs. longiareolata and Cs. subochrea. Saebi (1987) stated the species of An. plumbeus, An. hyrchanus, An. psudopictus, An. maculipennis and An. claviger. Momeni (1991) reported the species of An. messeae, An. melanoon, and An. maculipennis. Glick (1992) reported the species of An. psudopictus. Karami (2000) cited the presence of Cx. bitaeniorhynchus, Ur. unguiculata, Oc. caspius. Sedaghat et al. (2003) reported the species of An. Macu- lipennis, An. sacharovi, An. persiensis in Mazandaran Province. Sedaghat and Harbach (2005) stated the occurrence of the species of An. melanoon, An. persiensis, An. pseudopictus. Moreover, Dinparast et al. (2007) cited the species of An. atroparvous and An. persiensis. Dinparast et al. (2009) stressed the presence of the species of An. hyrchanus. Nikookar et al. (2010) reported the species of An. plumbeus, Oc. echinus, Oc. geniculatus, Oc. pulcritarsis, Cx. pipiens and Cs. annulata. Iran is in the malaria pre elimination stage (WHO 2008), thus the bionomic study of mosquitoes in this country is very important. There are scattered studies on the fauna and ecological properties of mosquitoes in Mazandaran Province. Many travelers may travel to Mazandaran Province and Neka County during visiting Iran. This part of Iran holds specific location due to access to the Caspian Sea and to other aspects such as trade, commerce and navigation. This study aimed to determine fauna and the ecology of mosquitoes in Neka County, Mazandaran Province, northern Iran from April to December, 2009. The data obtain in this study will be valuable to develop pro- grams for future planning of mosquito con- trol in this area. Materials and Methods The study area Mazandaran Province is located in the northern Iran between the Caspian Sea and the Alborz Mountains (36°39 N, 53°17 E) (Fig. 1). This area located in the north with Caspian Sea, in the south with Tehran and Semnan Provinces, in the east with Golestan Province and in the west Guilan Province, including 19 Counties (Geography of Mazandaran Province 2013). Maximum and minimum temperatures were 27.6 and 9.1 °C in July and December 2009, respectively. The maximum annual rainfall has been re- ported 204.6 mm in October, and the least was 0.1 mm in July 2009. The most im- portant crop of the region is rice, and Hyrcanian forest area and mild and rainy weather caused this Province to be a suitable mosquito habitat (Mazandaran Local Mete- orological Office 2009). Three fixed villages, namely Darvishan, Chalmardi, and Khairabad, were randomly selected for field survey. Moreover, six vari- able villages (Golbestan, Elmiyeh, Baghearab, Nimchah, Chalehpol, and Khorshid) were selected with regard to existing facilities in Neka County. In each fixed village, six fixed and two variable locations were selected in plain, slope and mountain conditions. Larvae sampling was collected from the natural larval habitats of fixed villages including river margins, ponds and water craters, craters rain water, marsh, tree holes, rice fields, worn tires, irrigation canals, wells, and sewage, using the standard dipper rate of 350 mm capacity every 15 days (WHO 1963). In addition, larval collection carried out in variable villages randomly. http://jad.tums.ac.ir Published Online: March 11, 2015 J Arthropod-Borne Dis, December 2015, 9(2): 253–266 S H Nikookar et al.: Fauna and Larval … 256 We attempted to collect the data about the biological and physical characteristics of the larval habitats, including running water, residents (permanent or temporary), the type of larval habitats natural or artificial, pres- ence or absence of the plant, type of sub- strate (sand, clay, or rocky), to be sunny (sunny or shady), water status (clear or opaque) and water temperature. The larval stages III and IV were pre- served in 100 % lactophenol solution and later identified morphologically using the keys of Shahgudian (1960), Zaim and Cranston (1986a), Harbach (1985, 1988), and Azari- Hamidian and Harbach (2009). The results were analyzed by SPSS 11.5 software. The abbreviations of the genus names were reg- ulated based on Reinert (2009). Results A total of 32530 larvae were collected: 3625 larvae of anophelinae and 28905 culicinae. Table 1 shows the results of anophelinae and culicinae species in the fixed and variable breeding places and tree holes. Four species of Anopheles were col- lected: An. claviger (0.31%), An. maculipennis (0.54%), An. plumbeus (10.28%), and An. superpictus (0.01%). Anopheles claviger collected 0.31 %, 2.78 % of total culicidae and Anopheles collec- tion, respectively (Table 1). The characteris- tics of the habitats under different tempera- ture and altitude conditions are shown in Ta- ble 2. The minimum and maximum temper- ature of water of the larval habitats was cal- culated as 19 °C and 22 °C, respectively (average 20.5 °C). On the other hand, the maximum and minimum altitude of the hab- itats was 300 and 200 meters, respectively (average 250 meters). Anopheles maculipennis was collected 0.54 %, 4.82 % of the total and Anopheles collection, respectively. Maximum and mini- mum water temperature was reported as 26 °C and 17 °C, respectively (average 22.5 °C). Maximum and minimum altitude was 300 and 180 meters respectively, (average 240 meters). Anopheles plumbeus was collected 10.28 %, 92.27 % of total and Anopheles collec- tion, respectively. Moreover, this species was collected in tree holes and the artificial habitats such as pond water. Maximum and minimum temperature was found as16 °C and 14 °C, respectively (average 15 °C). The minimum and maximum sea level of larval habitat was calculated at least 180 and 250 meters, respectivelly and the average were 215 meters. Anopheles superpictus was collected 0.01 %, 0.13 % of the total and Anopheles collec- tion, respectively. Minimum and maximum temperature of the larval habitat was re- ported as 17 °C and 19 °C, respectively and average was 180 °C. The minimum altitude was 280 meters, and the maximum was 300 (average 290 meters).The optimum of tem- perature and favorable altitudes of other spe- cies are presented in Tables 1 and 2. The majority of An. plumbeus was col- lected in natural, permanent, and stagnant water with vegetation, turbid and transparent water and semi-shade tree holes are shown in Table 3. An. superpictus larvae were col- lected from natural, permanent, stagnant water with vegetation, clear and transparent breeding places. On the other hand, An. claviger larvae were collected mostly in per- manent, and stagnant, transparent water with semi-shade and vegetation. Culex mimeticus was found in the natural habitats such as river and ponds from Permanent River, and stagnant pools with vegetation. The larvae of Cx. pipiens, the most abundant species, were found in artificial habitats and sewage breed- ing places in agricultural lands and low abundance along the river. Moreover, Oc. geniculatus, the most dominant species, was found in permanent water with vegetation in slow running with clay substrate water, semi- http://jad.tums.ac.ir Published Online: March 11, 2015 J Arthropod-Borne Dis, December 2015, 9(2): 253–266 S H Nikookar et al.: Fauna and Larval … 257 shade in natural breeding places. The larvae of Cs. longiareolata were found in most the larval habitats. The larval habitat charac- teristics of the other species are shown in Table 3. Checklist of Culicidae in Mazandaran Province Family culicidae Subfamily Anophelinae Subgenus Anopheles Meigen 1- An. (Ano.) algeriensis 2- An. (Ano.) claviger 3- An. (Ano.) marteri 4- An. (Ano.) maculipennis 5- An. (Ano.) atroparvus 6- An. (Ano.) melanoon 7- An. (Ano.) messeae 8- An. (Ano.) persiensis 9- An. (Ano.) sacharovi 10- An. (Ano.) plumbeus 11- An. (Ano.) hyrcanus 12- An. (Ano.) pseudopictus Subfamily Culicinae Subgenus Cellia Theobald 13- An. (Cel.) pulcherrimus 14- An. (Cel.) superpictus Subgenus Aedimorphus Theobald 15- Ae. (Adm.) vexans Subgenus Finlaya 16- Oc. (Fin.) echinus 17- Oc. (Fin.) geniculatus Subgenus Ochlerotatus 18- Oc. (Och.) caspius 19- Oc. (Och.) pulcritarsis Subgenus Culex 20- Cx. (Cux.) pipiens 21- Cx. (Cux.) perexiguss 22- Cx. (Cux.) territans 23- Cx. (Cux.) theileri 24- Cx. (Cux.) mimeticus 25- Cx. (Cux.) tritaeniorhynchus 26- Cx. (Cux.) hortensis Subgenus Allotheobaldia 27- Cs. (All.) longiareolata Subgenus Culiseta 28- Cs. (Cus.) annulata 29- Cs. (Cus.) subochrea Subgenus Coquillettidia 30- Cq. (Coq.) richiardii Subgenus Pseudoficalbia 31- Ur. (Pfc.) unguiculata Fig. 1. Map of Iran and Neka County, northern Iran http://jad.tums.ac.ir Published Online: March 11, 2015 J Arthropod-Borne Dis, December 2015, 9(2): 253–266 S H Nikookar et al.: Fauna and Larval … 258 Table 1. Abundance of larval mosquitoes (density per 10 dip) in Neka County, Mazandaran Province, northern Iran, 2009 %n Villages Species VariableKhairabadChalmardiDarvishan 0.311000262An. claviger 0.5417503.5410An. maculipennis 10.283345033301.5An. plumbeus 0.015000.50An. superpictus 1.0735008252Cs. annulata 8.9129001620221.550.5Cs. longiareolata 0.031000.50.50Cx. mimeticus 63.992081569991494527.5Cx. pipiens 14.854830048300Oc. geniculatus Table 2. Species composition of mosquito larvae collected from different temperatures and altitudes in Neka County, northern Iran, 2009 AltitudeTemperature C◦ MeanMaximumMinimumMeanMaximumMinimumSpecies 25030020020.52219An. claviger 24030018022.52617An. maculipennis 215250180151614An. plumbeus 290300280181917An. superpictus 240300180172212Cs. annulata 24030018018.52512Cs. longiareolata 25030020012.51312Cx. mimeticus 24030018018.52512Cx. pipiens 24025023012.51411Oc.geniculatus Table 3. Larval habitat characteristics in Neka County, Mazandaran Province, northern Iran, 2009 A n . claviger A n . m acu lipen n is A n .plu m beu s A n . su perpictu s C s.an n u lata C s. lon giareolata C x. m im eticu s C x.pipien s O c.gen icu latu s Habitat status Permanent 91 95.3 91.3 66.2 82.5 75.1 100 71.6 99 Temporary 9 4.7 8.7 33.8 17.5 24.9 0 28.4 1 Water Slow running 10 16 20 29.5 10 2 3 30 0 Stagnant 90 84 80 70.5 90 98 87 70 100 Vegetation Without vegetation 2 2 7.6 20 5 29 19.6 13 0 With vegetation 98 98 92.4 80 95 71 80.4 87 100 Type of bed Clay 47 1 84.5 50 98 88 66 56 94 Sand 3 17 15.5 20 2 5 30 33 6 Stone or cement 50 82 0 30 0 5 4 11 0 http://jad.tums.ac.ir Published Online: March 11, 2015 J Arthropod-Borne Dis, December 2015, 9(2): 253–266 S H Nikookar et al.: Fauna and Larval … 259 Water status Opaque 1 40 50 9.1 50 40 40 45 50 Transparent 99 60 50 90.9 50 60 60 55 50 Status Light Sunny 1 33.2 0 14.7 0 2.26 26.7 37.4 0 Shaded 34 28.4 5 26.1 5 26.9 19.4 18.9 5 Semi-shade 65 38.4 95 59.2 95 70.5 54.9 43.7 95 Habitat type Natural 99 55 98 89 98 97 70 41 100 Artificial 1 45 2 11 2 3 30 59 0 Discussion In the present study, An. plumbeus larvae were collected from tree holes and artificial habitats such as permanent and stagnant water. However, the maximum and minimum of temperature of the larval habitat was as 16 °C and 14 °C, respectively. In parallel, Azari- Hamidian (2011a) found this species in tree holes and discarded tire. The most favorable temperature is 15–18 °C; however, the spe- cies has tolerated to freeze (up to -6° C to -8 °C) in the laboratory (Horsfall 1955). How- ever, Garrett Jones reported the occurrence of this species in Kelardasht of Mazandaran Province (Garrett Jones 1951). In our investigation, Cs. annulata was found in natural, permanent, stagnant, semi- shade habitats with vegetation. This species has been reported in Guilan Province, Alborz and Zagros Mountains, East Bushehr and Robat Tork of Isfahan Province (Azari- Hamidian et al. 2003b). Zaim and Cranston (1986) included Cs. subochrea in their sys- tematic keys and Cs. annulata in their check- list. The taxonomy and distribution of Cs. annulata, Cs. subochrea and Cs. alaskaensis need to be more investigated in Iran (Azari- Hamidian 2005). Record of Cs. annulata in our research was based on adults, whereas Cs. subochrea were not found in our collec- tion, and neither were the adults of this spe- cies (Table1). Culiseta subochrea and Cs. annulata were reported in Ardebil Province (Azari-Hamidian et al. 2009). The larval habitats of Cs. annulata have also been reported from various places except tree holes (Azari-Hamidian et al. 2003b). However, An. plumbeus and Oc. geniculatus species have been reported in tree holes (Nikookar et al. 2010). In our study, the larvae of Cs. longiareolata were collected from all larval habitats except tree holes, which comprised 8.91 %, 10.03 % of the total and Culicinae collection, re- spectively. This species was reported first time in north of the country by Gutzevich (1943). The occurrence of this species was also reported by the previous study in Mazandaran Province (Zaim 1987). The larval habitat of this species has been re- ported from many parts of Iran (Zaim 1987). At the same study, the larval habitat has been reported as contaminated with organic materials, in old pit located in outdoor places and rice fields (Kitron and Pener 1986, Mousakazemi et al. 2000, Azari-Hamidian 2005). It has also been reported from the North Khorasan and Qom Provinces (Azari- Hamidian et al. 2011, Saghafipour et al. 2012). There is scatter information about the distribution and taxonomy of the three spe- cies of Cs. alaskaensis, Cs. annulata, and Cs. subochrea in Iran (Zaim et al. 1986, Azari-Hamidian et al. 2003b, Azari-Hamidian 2005). Moosa Kazemi et al. (2010) found the species of Cs. subochrea and Cs. longiareolata http://jad.tums.ac.ir Published Online: March 11, 2015 J Arthropod-Borne Dis, December 2015, 9(2): 253–266 S H Nikookar et al.: Fauna and Larval … 260 in Sanandaj County of Kurdistan Province, northwestern Iran. In our research, An. claviger was found in permanent, stagnant water with vegetation habitat, as well as in the clay and stone sub- strate of water. In parallel, Macan (1950) has reported the existence of this species in the sunny/shadowy springs, and slow running pools water temperature of 14–16 °C in Iraq and western Iran. Dow (1953) reported the larval habitat of this species in shallow and small pools with little vegetation. It was also found within the larval habitats with temper- ature of 12–15 °C (Horsfall 1955). The oc- currence of this species in Mazandaran was reported previously by Garrett Jones (Garrett Jones 1951). Zaim (1987) collected An. claviger larvae from Kashan, central Iran. In our study, An. maculipennis was found in permanent, transparent, semi-shady natu- ral breeding places with vegetation habitat, as well as in the cement or stone substrate of water. In parallel, Azari-Hamidian (2011) reported the presence of this species in the stagnant, gravel substrate, sunny springs, and pools with stagnant water. Anopheles maculipennis complex formally comprises 12 Palearctic members, including An. atroparvus, An. beklemishevi, An. labranchiae, An. maculipennis, An. martinius, An. melanoon, An. messeae, An. sacharovi, An. persiensis, An. daciae, An. lewisi and An. artemievi (Dinparast Djadid et al. 2007). Anopheles maculipennis species associated by An. hyrcanus, An. claviger from Mazandaran Province previously (Nikookar et al. 2010). This species has been identified in Guilan (Azari-Hamidian et al. 2004), Mazandaran and Golestan Provinces (Zaim et al. 1986). Anophelesmesseae and Anopheles melanon reported from Guilan Province, and An. sacharovi and An. hyrcanus from Golestan Province (Saebi 1987). Anopheles sacharovi has been reported from Mazandaran Prov- ince (Sedaghat et al. 2003). Dow reported the species of An. subalpinus in Sari, Babolsar, Chalus and Astaneh cities in Mazandaran Province. He also reported An. melanoon in Astaneh city, Guilan Province (Dow 1983). Anopheles subalpinus is a Synonyms An. melanoon. Initial studies mainly focused on identification of the mosquitoes and their role in disease transmission (Gutsevich 1943, Macan 1950, Dow 1953, Minar 1974). In our investigation, An. superpictus was found as 0.01 %, 0.13 % of the total and Anopheles collection, respectively. This spe- cies was found in permanent, stagnant, with clay substrate, transparent water, semi-shade, natural with vegetation habitat. Studies in different parts of Mazandaran Province con- firmed the occurrence of this species (Zolotarev 1945, Dow 1953). Moreover, Azari-Hamidian (2011) stated its presence in stagnant, transient, mud substrate, fully sunlight places with vegetation in natural habitats. Further support for our results comes from a study by Mousakazemi et al. (2000) in Zarrin-Shahr and Mobarakeh areas of Isfahan Province. Some information on the larval stage of An. superpictus was re- ported by Yaghoobi-Ershadi et al. (2001) in Ardebil Province, northwestern Iran. Some ecological aspects of this species have been in Rasht County of Guilan Province (Azari- Hamidian et al. 2002b). Ghanbari et al. (2005) reported the larval breeding places of mos- quitoes including anophelines in Iranshahr, southeastern Iran. However, the edges of stony streams with sunlight and semi- shallow have been mentioned as the main breeding places of this species in western Iran (Macan 1950). Three genotypes named X, Y, and Z within An. superpictus have been reported in Iran by Oshaghi et al. (2008). There are no reports about the gen- otypes of this species in Mazandaran Prov- ince, however, it needs to be studied by the future investigations. In our study, Culex pipiens was found as 63.99 % of the total and 72.01 % of Culicinae larvae collection in natural and http://jad.tums.ac.ir Published Online: March 11, 2015 J Arthropod-Borne Dis, December 2015, 9(2): 253–266 S H Nikookar et al.: Fauna and Larval … 261 artificial larval habitats such as tree holes. In addition, hollow pipes, hot water, holes Hungarian sewage, stagnant water, and toilet field were reported as the breeding places of this species (Jupp 1970, Kulanin 1980, Zaim 1987, Vinogradova 2000, Mousakazemi et al. 2000). This species were reported in Nat- ural habitat (rain pool, river edge) with Stand- ing water Azari-Hamidian (2007b). The species of Cx. pipiens and Cx. quinquefasciatus are very similar. There are no reports in regarding the existence of Cx. quiquefasciatus in Mazandaran Province. The shape of siphon of larvae in the tip, middle and the base has been mentioned to iden- tification of two recent species (Zaim and Cranston 1986). However, Azari-Hamidian and Harbach (2009) mentiond the existence of two branches of seta 1 on abdominal seg- ment III-VI for Cx. pipiens and one branch for Cx. quinquefasciatus . The common characteristics among the species of Cx. vagans, Cx. torrentium, and Cx. pipiens, were reported as the presence of two pairs of long seta on the base of siphon (Zaim and Cranston 1986). The existence of seta 1 with 4–5 branches and seta 1-X with two branches is attributed to Cx. torrentium and two branches of seta 1 on abdominal segment III-V mentioned to Cx. vagans (Harbach 1985). In our study, Ochlerotatus geniculatus was collected as comprising 14.85 %, 16.7 % of the total and Culicinae larvae collections, respectively. In addition, the most favorable temperature was 11–14 °C. Maximum and minimum altitude of the larval habitat was 250 meters and 230 meters, respectively and the average was 240 meters. Further support of this result comes from some previous studies (Gutsevich 1943, Zaim et al. 1984a, Zaim 1987, Azari-Hamidian et al. 2001, Nikookar et al. 2010). This species is very similar to Oc. echinus as reported in a recent study (Nikookar et al. 2010). Ochlerotatus echinus is distributed in Palaearctic, Europe, north Africa, and south east Asia, and in Iran, it has been reported from Guilan Prov- ince in the north (Knight and Stone 1977, Azarai-Hamidian et al. 2002). In addition, this species were collected by janbakhsh in Sari County and reported by Zaim (1984a). Studies from some parts of Mazandaran Prov- ince showed the occurrence of this species (Zaim et al. 1984a, Zaim 1987, Nikookar et al. 2010). Moreover, the distribution of this species has been reported from both Guilan, Mazandaran Provinces (Azari-Hamidian et al. 2002a), as well as from Gulestan (Gutsevich 1943), and Ardebil (Azari-Hamidian et al. 2009). In our study, Culex mimeticus was found as comprising 0.03 % of the total larvae collection in permanent, stagnant, transpar- ent water, clay substrate, semi-shade natural breeding places. In parallel, Zaim (1987) stressed the permanent, slow running water, full sunlight, mud substrate, transient and fresh water with vegetation in natural stream bed pools and rice fields as the breeding places of this species. Studies from different parts of Mazandaran confirmed the occurrence of this species (Dow 1953, Zaim 1987). Conclusion Our results indicate that the presence of the dominant species of Cx. pipiens in larval habitat natural and artificial Oc. geniculatus and An. plumbeus in natural such as tree hole. These species dramatically increased and reach to peak in warm season. The re- cent species could be creating nuisance and health risks in this area. Acknowledgements The authors are grateful to Dr Azari- Hamidian for giving the useful comments, Dr Davood Nikzad, Mr Hamid Ravesh, Mr Mojtaba Asghari, Mr Mirramazan Nikookar, http://jad.tums.ac.ir Published Online: March 11, 2015 J Arthropod-Borne Dis, December 2015, 9(2): 253–266 S H Nikookar et al.: Fauna and Larval … 262 Mr Mehrdad Badangiz, and Mr Mohammad Badangiz (Environmental Health Director of Neka) for supporting the investigation. We also would like to express our appreciation to the people of Darvishan, Khairabad and Chelmardi villages in Neka County for their kind cooperation during the study. This study was financially supported by Tehran University of Medical Sciences (No: 10513- 27-01-89). The authors declare that there is no conflict of interests. 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