57Background: The stable fly, Stomoxys calcitrans, is a major blood-feeding pest of livestock that 58 has near worldwide distribution, causing an annual cost of over $2 billion for control and product 59 loss in the United States alone. Control of these flies has been limited to increased sanitary 60 management practices and insecticide application for suppressing larval stages. Few genetic 61 and molecular resources are available to help in developing novel methods for controlling stable 62 flies. 63 64 Results: This study examines stable fly biology by utilizing a combination of high-quality 65 genome sequencing, microbiome analyses, and RNA-seq analyses targeting multiple 66 developmental stages and tissues. In conjunction, manual curation of over 1600 genes was 67 used to examine gene content related to stable fly reproduction, interactions with their host, 68 host-microbe dynamics, and putative routes for control. Most notable was establishment of 69 reproduction-associated genes and identification of expanded vision, chemosensation, immune 70 repertoire, and metabolic detoxification pathway gene families. 71 72 Conclusions: The combined sequencing, assembly, and curation of the male stable fly genome 73 followed by RNA-seq and downstream analyses provide insights necessary to understand the 74 biology of this important pest. These resources and knowledge will provide the groundwork for 75 expanding the tools available to control stable fly infestations. The close relationship of 76 Stomoxys to other blood-feeding (Glossina) and non-blood-feeding flies (medflies, Drosophila, 77 house flies) will allow for understanding the evolution of blood feeding among Cyclorrhapha 78 flies. 79 80 immunity 83 Additional File 1: Supplementary Material; Additional File 2: Supplementary Tables 84 Supplementary Dataset Files: 10 85 109 or a contaminated substrate to a susceptible host, association between specific vector and 110 pathogen is not necessary) of Equine infectious anemia, African swine fever, West Nile, and Rift 111 Valley Viruses, Trypanosoma spp., and Besnoitia spp. (reviewed by [13]). The apparent low 112 vector competence of stable flies implicates the importance of immune system pathways not 113 only in regulating larval survival in microbe-rich environments but also in the inability of 114 pathogens to survive and replicate in the adult midgut following ingestion [14-16]. 115 Stable fly mate location and recognition are largely dependent upon visual cues and contact 116 pheromones [17, 18], and gravid females identify suitable oviposition sites through a 117 combination of olfactory and contact chemostimuli along with physical cues [19, 20]. Since 118 stable flies infrequently associate with their hosts, feeding only 1 to 2 times per day, on-animal 119 5 and pesticide applications are less effective control efforts than those that integrate sanitation 120 practices with fly population suppression by way of traps [21]. Given the importance of 121 chemosensory and vision pathways, repellents have been ident...