IDENTIFICATION OF A HOMOLOGUE OF A HOUSE DUST MITE ALLERGEN IN A CDNA LIBRARY FROM SARCOPTES SCABIEI VAR. HOMINIS AND EVALUATION OF ITS VACCINE POTENTIAL IN A RABBIT/S. SCABIEI VAR. CANIS MODEL PEARLY HARUMAL, MARJORIE MORGAN, SHELLEY F. WALTON, DEBORAH C. HOLT, JURGEN RODE, LARRY G. ARLIAN, BART J. CURRIE, AND DAVID J. KEMP Menzies School of Health Research, Darwin, Northern Territory, Australia; Wright State University, Dayton, Ohio; The Queensland Institute of Medical Research, Australian Centre for International and Tropical Health and Nutrition, The University of Queensland, Brisbane, Australia; and Royal Darwin Hospital, Darwin, Northern Territory, Australia Abstract. Sarcoptes scabiei (“itch mite”) causes scabies, a disease of considerable human and veterinary significance. Little work has been done at the molecular level because of the difficulty of obtaining mites. We have used mites in skin from the bedding of crusted scabies patients for the construction of a library of 10 5 cDNAs from S. scabiei var. hominis cloned in the vector pGEX4T-2. We describe the isolation by immunoscreening of 2 clones, one of which (Ssag1) is homologous to and cross-reactive with the house dust mite Euroglyphus maynei allergen M-177, an apolipoprotein from hemolymph. Immunohistochemistry revealed that it is located around the internal organs and cuticle of the mite and in eggs. Although it was not found to be protective in a challenge trial, the rabbits did not exhibit typical crust character- istics. This work shows that it is now possible to conduct such challenge trials with cloned scabies antigens. INTRODUCTION Scabies (“itch mite,” or Sarcoptes scabiei) is a common infestation in the developing world, 1–4 with about 300 million people affected. Although it can be treated, 5 it is endemic among some disadvantaged populations, such as members of remote northern and central Australian Aboriginal commu- nities, 6 in which 50% of children may be infested with S. scabiei. These infestations decrease the quality of life by caus- ing intense itching and often lead to bacterial skin infections followed by serious complications, including septicemia and renal damage. 7 Secondary infections with Group A strepto- cocci have been associated with outbreaks of rheumatic heart disease. 8 Scabies is also a problem in other crowded situa- tions, such as nursing homes and day care centers. Histori- cally, rapid increases in incidence have been noted during times of war and social upheaval. A vaccine is the most de- sirable long-term solution because of limitations of drug de- livery and compliance. In most cases of scabies, the mite burden is self-limiting in humans, suggesting that it is controlled immunologically. Mel- lanby 9 determined that in experimental infestations, the av- erage number of mites on adults was 11 to 13. Reinfection was more difficult, and the parasite rate was even lower. A few people develop the serious alternative form, crusted scabies, characterized by an extremely high burden of mites. Out- breaks of scabies in hospitals after admission of such patients show that this is caused by the same variety of mite that causes normal scabies. Crusted scabies commonly occurs in patients with acquired immunodeficiency syndrome and other immunosuppressive conditions, but it also occurs in people with no known immunodeficiency. 10 It has been postulated that there is a specific immune deficit that results in a few individuals developing this hyperinfestation. 11 Scabies pro- gresses to the crusted state in most animals if it is untreated, however. Mites ingest rabbit antibodies during feeding; it has been shown by immunologic staining of sections that rabbit anti- bodies are present in the gut of the mite (LG Arlian et al., unpublished data). We have postulated that these antibodies in the mite gut may be protective. 11 Immunity after pri- mary infestations has been shown in an animal model, 12,13 and experimental immunization with mite extracts can induce protective immunity. 14 Taken together, these observa- tions suggest that antibodies may play a role in the develop- ment of protective immunity. Mechanical removal of mites by scratching also can play an important role in control of mite numbers. The epidemiology of scabies in populations suggests partial immunity is acquired with age and expo- sure. It may be possible to control scabies in children using a vaccine. Until more recently, there has been no literature on the molecular biology of scabies, owing to the difficulty of ob- taining sufficient mites. A system of growing dog-derived mites on the ears of New Zealand white rabbits, which pro- vides access to S. scabiei var. canis and allows the testing of potential vaccines, has been developed. 14,15 At the Royal Darwin Hospital, about 10 patients per year are admitted with crusted scabies. Shed skin from the bedding of these patients provides a noninvasive source of up to 4,000 mites per gram of skin, and we previously have cloned EcoRI genomic DNA fragments of S. scabiei var. hominis from this material. 16 Mi- crosatellite clones from this library were used to show that S. scabiei var. canis and S. scabiei var. hominis are distinct populations regardless of their geographic origin. 17 Sequence analysis of the second internal transcribed spacer of the ribo- somal RNA complex 18 nevertheless suggested that Sarcoptes mites are the same species. This suggestion was supported by sequencing a mitochondrial ribosomal RNA gene, 19 although both these studies were limited. A library of cDNA clones from S. scabiei var. vulpes has been constructed from mites obtained from red foxes, and a cDNA clone encoding paramyosin has been characterized. 20 The sequences of many other cDNAs from this library have been placed in GenBank (accession numbers BG817579-BG817974). We report here the construction of cDNA libraries from S. scabiei var. hominis. We describe the isolation by immuno- screening of 2 clones, one of which is the S. scabiei homologue of a major house dust mite allergen. Although these clones showed no protective efficacy in the rabbit/S. scabiei var. ca- nis model, the work shows that it is now possible to test cloned scabies antigens for vaccine potential. Future applica- tions of these new resources are discussed. Am. J. Trop. Med. Hyg., 68(1), 2003, pp. 54–60 Copyright © 2003 by The American Society of Tropical Medicine and Hygiene 54