Louisiana State University LSU Digital Commons LSU Doctoral Dissertations Graduate School 2007 Detection of the bloodmeal origin of mosquitoes collected at alligator farms, and West Nile Virus activity in mosquitoes collected from alligator farms, sentinel chicken surveillance sites, and larval habitats Isik Unlu Louisiana State University and Agricultural and Mechanical College, [email protected] Follow this and additional works at: https://digitalcommons.lsu.edu/gradschool_dissertations Part of the Entomology Commons Recommended Citation Unlu, Isik, "Detection of the bloodmeal origin of mosquitoes collected at alligator farms, and West Nile Virus activity in mosquitoes collected from alligator farms, sentinel chicken surveillance sites, and larval habitats" (2007). LSU Doctoral Dissertations. 184. https://digitalcommons.lsu.edu/gradschool_dissertations/184 This Dissertation is brought to you for free and open access by the Graduate School at LSU Digital Commons. It has been accepted for inclusion in LSU Doctoral Dissertations by an authorized graduate school editor of LSU Digital Commons. For more information, please [email protected]. DETECTION OF THE BLOODMEAL ORIGIN OF MOSQUITOES COLLECTED AT ALLIGATOR FARMS, AND WEST NILE VIRUS ACTIVITY IN MOSQUITOES COLLECTED FROM ALLIGATOR FARMS, SENTINEL CHICKEN SURVEILLANCE SITES, AND LARVAL HABITATS A Dissertation Submitted to the Graduate Faculty of the Louisiana State University and Agricultural and Mechanical College in partial fulfillment of the requirement for the degree of Doctor of Philosophy In The Department of Entomology By Isik Unlu B. S., Hacettepe University, Ankara, 1997 M.S., Hacettepe University, Ankara, 2000 December 2007 Dedicated to the late Dr. Michael Perich (the wind beneath my wings) and my husband Emre Unlu ii ACKNOWLEDGEMENTS I would like to express my gratitude to my major professor, Dr. Lane Foil, for his support, patience, and encouragement throughout my graduate studies. I extend my appreciation to Dr. Wayne Kramer, without whose support and guidance the culmination of this thesis would not have been possible. My deepest gratitude also goes out to the Chair of the Entomology Department of Louisiana State University, Timothy Schowalter for his collaboration and constant encouragement. I would like to thank my committee members; Drs. Alma Roy, Mark Mitchell, Claudia Husseneder, and Todd Monroe for their valuable insights and guidance. To the late Dr. Michael Perich, who believed in me and inspired me to be a medical entomologist, I will never forget you. I thank Audrey Perich for unceasing generosity and friendship. I extend my appreciation to the director of East Baton Rouge Parish Mosquito Abatement and Rodent Control (EBRMARC), Matthew Yates for providing with support and resources so critical to my research. Additionally, I am very grateful for the friendship and help of all of the members of EBRMARC, especially Daniel Garrett, Randy Veath, Steve Hahn, Larry Mercier and Fred Augustine, with whom I worked closely during my field studies. Many thanks for the financial assistance from Louisiana Mosquito Control Association whose support made this research possible. I would also like to thank all of the members of the Louisiana Animal Disease and Diagnostic Laboratory (LADDL) especially Tarra Harden, Heather Lampinen, and Kim Bowles. iii The others who I wish to recognize for helping throughout my research endevours; Cole Younger, Gerardo Boquin, Dea Colby, Tom Mascari, Andrew Mackay, Stephanie Gil, Latha Bommireddy, Asli Dogan, Jack Tourr, and student workers. Although it was really hard to be separated from my family and loved ones, I always felt their love. My parents, Fatma and Gurtekin Oguzoglu, my sister Neslihan Apaydin, my brothers Harun Apaydin and Oguz Oguzoglu receive my deepest gratitude and love for their dedication and the many years of support during my undergraduate studies that provided the foundation for this work. The friendship of mother- in-law Aysel Unlu is much appreciated. Even though I was away from my family I was fortunate enough to have close friends who I consider to be my Louisiana family. They have been very supportive and encouraging and without their love I would not have been able to accomplish this graduate program. I especially thank Dr. Margo Abadie Gerald, Michael Gerald, Elena and Kyle Keegan, Karen and Lyndsay Mcdonough, Kerry Cooly ‘Cool Daddy’, Andy Venuto, Claire Bateman, Bunni Perich, Sarah Perich, Rita and Charles Myers, Sadik and Ann Marie Artunc for opening their hearts and homes to me. Last, but not least, I would like to thank my husband Emre for his understanding and love during the past four years. He helped me with my field work and every possible way he can help. His support and encouragement was in the end what made this dissertation possible. iv TABLE OF CONTENTS DEDICATION………………………………………………………...………………….ii ACKNOWLEDGEMENTS............................................................................................... iii LIST OF TABLES ............................................................................................................ vii LIST OF FIGURES ......................................................................................................... viii ABSTRACT ....................................................................................................................... ix INTRODUCTION .............................................................................................................. 1 CHAPTER 1. LITERATURE REVIEW ............................................................................ 3 Taxonomy of Mosquitoes ............................................................................................... 3 Important Mosquito Transmitted Disease Agents .......................................................... 4 West Nile Virus .............................................................................................................. 6 West Nile Virus Historical Data ..................................................................................... 8 West Nile Virus Virology ............................................................................................. 11 West Nile Virus Replication ......................................................................................... 12 West Nile virus Epidemiology ...................................................................................... 12 Vector Incrimination and Vector Competence ............................................................. 14 West Nile Virus Transmission Cycle ............................................................................ 16 Surveillance Methods for West Nile Virus ................................................................... 19 Virus Detection Protocols for West Nile Virus in Mosquito Specimens and Vertebrate Hosts ........................................................................................................... 21 CHAPTER 2: EVALUATION OF SURVEILLANCE METHODS FOR DETECTION OF WEST NILE VIRUS ACTIVITY IN EAST BATON ROUGE PARISH, LOUISIANA, 2004-2006 ................................................................................................. 26 Introduction ................................................................................................................... 26 Materials and Methods .................................................................................................. 27 Experimental Design ................................................................................................. 27 Mosquito Pool Testing .............................................................................................. 29 Statistical Analysis .................................................................................................... 30 Results ....................................................................................................................... 30 Discussion ................................................................................................................. 36 CHAPTER 3: IDENTIFICATION OF BLOODMEAL ORIGINS FOR MOSQUITOES COLLECTED AT ALLIGATOR FARMS IN LOUISIANA .......................................... 41 Introduction ................................................................................................................... 41 Materials and Methods .................................................................................................. 43 Experimental Design ................................................................................................. 43 Mosquito Processing ................................................................................................. 44 v DNA Extraction for Sequence .................................................................................. 44 PCR Primers and Conditions .................................................................................... 45 Sequencing of Ctb-Derived PCR Products of Known Vertebrates .......................... 45 Results ........................................................................................................................... 45 Discussion ..................................................................................................................... 52 CHAPTER 4: DETECTION OF WEST NILE VIRUS RNA FROM MOSQUITOES COLLECTED AT ALLIGATOR FARMS IN LOUISIANA, 2004-2006 ....................... 56 Introduction ..................................................................................................................