International Journal of Science and Research (IJSR) ISSN (Online): 2319-7064 Index Copernicus Value (2013): 6.14 | Impact Factor (2013): 4.438 Volume 4 Issue 5, May 2015 www.ijsr.net Licensed Under Creative Commons Attribution CC BY Isolation and Characterization of Internal Bacteria from the Mosquito, Culex pipiens from Egypt Fatma H. Galal 1 , Amany Abu Elnasr 3 , Ibtesam Abdallah 3 , Alaa Eddeen M. Seufi 4 , Omaima Zaki 5 1, 4 Department of Biology, College of Science, Aljouf University and Department of Entomology, Faculty of Science, Cairo University 2, 3, 5 Department of Botany and microbiology, Faculty of Science, HelwanUniversity Abstract: A total of 11 bacterial isolates were isolated from internal body of the mosquito,Culexpipiens(eggs, 4 th larval instars, pupae, adult males and adult females)in one gonotrophic cycle.Two bacterial isolates were Gram +ve stained (Staphyllococcusepidermidis and Bacillussubtilis) and the other nine bacterial isolates were Gram –ve stained. Phenotypic, biochemical biotyping and differential media were used to characterize and identify our isolate. These methods could differentiate our isolates to species level.In addition, many of the obtained strains were proved to have economic and commercial importance. Bacterial antagonism was studied and it was noticed that B. subtiliswas antagonistic to bothE. coli and S. aureus; while S. flexneriwas antagonistic to S. typhimurium;and S. epidermidis was antagonistic to the multi-drug resistant strain,S. aureus. Keywords: Culexpipiens, bacterial isolates,bacterial antagonism. 1. Introduction Insects are considered holobiont units in which the insect host and its microbiota are involved in complex reciprocal multipartite interactions (Rosenberg and Zilber-Rosenberg, 2011). Insects contain large communities of diverse microorganisms that probably exceed the number of cells in the insect itself(Dillon and Dillon, 2004). One of these microorganisms is bacteria that present on integument, inhabit digestive tract and in some cases inhabit unique structures within insect body (Chen et al., 2000 and Fukatsuet al., 2000). The bacterial association in insects plays significant role in host insect morphogenesis, food digestion, antifungal toxin production, pheromone production, pH regulation, vitamin synthesis, temperature tolerance, resistance against parasitoid development and detoxification of noxious compounds as well as nutrition, reproduction, development or protection against enemies (Dillon and Dillon, 2004, Gentaet al.,2006 and Douglas, 2011).In addition to fecundity and viability of insect host the bacterial communities play a role on the establishment of parasites within the host gut (Pumpuniet al., 1993) as well as on maturation of the innate immune system of the host (Weiss et al., 2011). Moreover, symbiotic relationships between mosquitoes and several microorganisms most probably have important implications in mosquitoes’ evolutionary success, including their widespread distribution. Furthermore, the resident microbiota of mosquito vectors may inhibit the development of pathogens they transmit )Cirimotichet.al., 2011(. In this context several microbes may offer opportunities to successfully manipulate the vector competence of mosquitoes to reduce their abilities to transmit human pathogens. The aim of our study is to understand the microbial community structure as a step to enable us to understand the organisms that play significant roles in the maintenance of these communities and to identify interaction between the microbiota communities isolated from both immature and mature stages of the mosquito Culexpipiens. 2. Materials and methods 2.1. Colonization of Cx. pipiens A colony of the mosquito, Cx. pipiens was maintained under controlled laboratory conditions (27 + 2 °C, 60-70% RH and 10L: 14D photoperiod) in the insectary of the Department of Entomology, Faculty of Science, Cairo University, Giza, Egypt up till now. This colony was used for supplying immature stages (eggs, larvae and pupae) during this study. Briefly, eggs were set up to hatch, 1 st instar larvae were seeded into plastic cups 25x35x7 cm 3 containing water at a constant density of 300 individuals per cup. Larvae were provided with activated yeast or tetramin every 2 days until pupation. Water was changed on feeding days to avoid bacterial growth on the water surface. On pupation, cups were placed inside an emergence cage (27x40x35 cm 3 ) and provided with a source of 10% sugar solution for the emerged adults. 2.2. Bacterial isolation 2.2.1.Bacterial isolation from the insect Twenty individuals of Cx. pipiens(egg rafts, fourth instar larvae, pupae, adult male and adult female) were surface sterilized with 70% ethanol for 5 min followed by twice washing in phosphate buffered saline (PBS) to remove possible contamination. The surface sterilized samples were homogenized in Ringer's solution using a glass grinder. 0.1 ml suspension was spread on 20 cm diameter plates containing Nutrient agar, MacConkey agar, Mannitol salt agar, Brilliant green agar and Salmonella-Shigella agar (S.S.) agar media separately using a sterilized scalpel inside laminar air flow hood (Thiery and Frachon, 1997).These plates were incubated at 30 ºC for 1-2 days and then investigated for bacterial isolation and identification. Paper ID: SUB154718 2682