The University of Southern Mississippi The Aquila Digital Community Dissertations Fall 12-2016 Epidemiology of White Spot Syndrome Virus in the Daggerblade Grass Shrimp (Palaemonetes pugio) and the Gulf Sand Fiddler Crab (Uca panacea) Muhammad University of Southern Mississippi Follow this and additional works at: https://aquila.usm.edu/dissertations Part of the Animal Diseases Commons, Disease Modeling Commons, Epidemiology Commons, Virology Commons, and the Virus Diseases Commons Recommended Citation Muhammad, "Epidemiology of White Spot Syndrome Virus in the Daggerblade Grass Shrimp (Palaemonetes pugio) and the Gulf Sand Fiddler Crab (Uca panacea)" (2016). Dissertations. 895. https://aquila.usm.edu/dissertations/895 This Dissertation is brought to you for free and open access by The Aquila Digital Community. It has been accepted for inclusion in Dissertations by an authorized administrator of The Aquila Digital Community. For more information, please contact [email protected]. EPIDEMIOLOGY OF WHITE SPOT SYNDROME VIRUS IN THE DAGGERBLADE GRASS SHRIMP (PALAEMONETES PUGIO) AND THE GULF SAND FIDDLER CRAB (UCA PANACEA) by Muhammad A Dissertation Submitted to the Graduate School and the School of Ocean Science and Technology at The University of Southern Mississippi in Partial Fulfillment of the Requirements for the Degree of Doctor of Philosophy Approved: ________________________________________________ Dr. Jeffrey M. Lotz, Committee Chair Professor, Ocean Science and Technology ________________________________________________ Dr. Darrell J. Grimes, Committee Member Professor, Ocean Science and Technology ________________________________________________ Dr. Wei Wu, Committee Member Associate Professor, Ocean Science and Technology ________________________________________________ Dr. Reginald B. Blaylock, Committee Member Associate Research Professor, Ocean Science and Technology ________________________________________________ Dr. Karen S. Coats Dean of the Graduate School December 2016 COPYRIGHT BY Muhammad* 2016 Published by the Graduate School *U.S. Copyright will show Muhammad Muhammad as the author’s name for this document. ABSTRACT EPIDEMIOLOGY OF WHITE SPOT SYNDROME VIRUS IN THE DAGGERBLADE GRASS SHRIMP (PALAEMONETES PUGIO) AND THE GULF SAND FIDDLER CRAB (UCA PANACEA) by Muhammad December 2016 Ever since the first outbreaks of White spot syndrome virus (WSSV), which causes White Spot Disease (WSD), in Asia in the early 1990s, the pathogen has been a major constraint to the profitability of the shrimp aquaculture industry across the globe. WSSV has a broad host range and is routinely detected in wild decapod crustaceans. In the present study, two common species in the tidal salt marsh along the northern coast of the Gulf of Mexico, the daggerblade grass shrimp (Palaemonetes pugio) and the Gulf sand fiddler crab (Uca panacea), were investigated for their role as reservoirs of WSSV and for the possible consequences of WSSV on their population dynamics. From 2013 to 2015, 1884 individuals of P. pugio and 1280 individuals of U. panacea were collected and screened for WSSV. The overall prevalence of WSSV in P. pugio and U. panacea was 7.27% and 12.97%, respectively. From experimental bioassays, the LD50 of WSSV for P. pugio at 5 dpi was 8.24×107 WSSV genome copies g-1 of body weight or 2.45×107 WSSV genome copies/grass shrimp, whereas the LD50 of WSSV for U. panacea at 14 dpi was 1.67×108 WSSV genome copies g-1 of body weight or 2.55×108 WSSV genome copies/fiddler crab. Experimental transmission of WSSV-China isolate to P. pugio -1 occurred when exposed to infected living P. pugio (β1,1 = 0.03 d ), infected living U. -1 -1 panacea (β1,2 = 0.02 d ), infected P. pugio carcasses (χ1,1 = 0.08 d ), and infected U. ii -1 panacea carcasses (χ1,2 = 0.03 d ). However, no WSSV transmission was observed when U. panacea was exposed to infected living or infected carcasses of either of the two -1 species. Virulence of WSSV was higher in P. pugio (α1 = 0.014 d ) than in U. panacea (α2 = 0.00). The decomposition of WSSV infectivity in carcasses of the two species was -1 rapid (δ = 1 d ). The basic reproduction number (R0) as calculated from single-host WSSV epidemic population models was 2.22 for P. pugio and 6.71 for U. panacea. R0 from a two-host WSSV epidemic community model, including both P. pugio and U. panacea, increased to 17.09. iii ACKNOWLEDGMENTS First and foremost, I would like to express my deepest gratitude to my advisor, Dr. Jeffrey M. Lotz for his support, intellectual guidance, understanding and encouragement throughout my study and research, and meaningful contributions throughout the writing process. I would also like to thank members of my committee Dr. Darrell J. Grimes, Dr. Wei Wu and Dr. Reginald B. Blaylock for their support, expert advice and insightful comments on this dissertation. I would like to specifically thanks Stephen S. Curran, Ph.D. for helping me with specimen collections. My research would not have been possible without his assistance. I would like to thank Verlee Breland, Jean A Jovonovich Alvillar, and Dr. Robin Overstreet for their endless support and encouragement. I would like to express my gratitude to the Government of Aceh Province of the Republic of Indonesia and Human Resources and Development of Aceh for the USA Graduate Scholarship and their generous support throughout my PhD program. I also thank the North Aceh Department of Marine Affairs and Fisheries and the Government of North Aceh district for their continued support. I would also like to thank my late father Tgk. Drs. H. Abdul Gani El Ahmady and my late mother Juwariah for their endless love, support and encouragement. All the support they have provided me over the years was the greatest gift anyone has ever given me. Thank you for always being proud of me. Special thanks also go to all the family and friends for their generous support and friendship along this eventful journey. iv TABLE OF CONTENTS ABSTRACT ........................................................................................................................ ii ACKNOWLEDGMENTS ................................................................................................. iv LIST OF TABLES .............................................................................................................. x LIST OF ILLUSTRATIONS ............................................................................................ xii CHAPTER I – INTRODUCTION ...................................................................................... 1 General Background ....................................................................................................... 1 Research Objectives ........................................................................................................ 4 Dissertation Organization ............................................................................................... 4 White Spot Syndrome Virus Review .............................................................................. 5 Infectious Disease Modeling......................................................................................... 10 CHAPTER II - PREVALENCE OF WHITE SPOT SYNDROME VIRUS IN THE DAGGERBLADE GRASS SHRIMP (PALAEMONETES PUGIO) AND THE GULF SAND FIDDLER CRAB (UCA PANACEA) .................................................................... 14 Introduction ................................................................................................................... 14 Materials and Methods .................................................................................................. 16 Sample Collection ..................................................................................................... 16 Genomic DNA Extraction......................................................................................... 17 Standard Curve.......................................................................................................... 17 Detection and Quantification of WSSV using TaqMan Real-Time PCR ................. 18 v Data Analysis ............................................................................................................ 19 Results ........................................................................................................................... 19 Discussion ..................................................................................................................... 23 CHAPTER III - PATHOGENICITY OF WHITE SPOT SYNDROME VIRUS IN THE DAGGERBLADE GRASS SHRIMP (PALAEMONETES PUGIO) AND THE GULF SAND FIDDLER CRAB (UCA PANACEA) .................................................................... 27 Introduction ................................................................................................................... 27 Materials and Methods .................................................................................................. 29 Experimental Animals .............................................................................................. 29 WSSV Stock Preparation and Production................................................................. 30 WSSV Experimental Challenge Protocol ................................................................. 31 Genomic DNA Extraction and Quantification .......................................................... 32 WSSV Copy Number Quantification by TaqMan Real-Time PCR ......................... 33 Data Analysis ...........................................................................................................