Adaptation of a Multi-drug Resistant Strain of Plasmodium falciparum from Peru to Aotus lemurinus griseimembra, A. nancymaae, and A. vociferans Monkeys William E. Collins,* Joann S. Sullivan, Patrice Hall, Trenton K. Ruebush II, Allison Williams, Katharine K. Grady, Amy Bounngaseng, Douglas Nace, Tyrone Williams, Curtis Huber, G. Gale Galland, John W. Barnwell, and James J. Sullivan Division of Parasitic Diseases, National Center for Zoonotic, Vector Borne and Enteric Diseases, and Animal Resources Branch, National Center for Preparedness, Detection and Control of Infectious Diseases, Centers for Disease Control and Prevention, U.S. Public Health Service, Department of Health and Human Services, Atlanta, Georgia; Atlanta Research and Education Foundation, Decatur, Georgia; U.S. Agency for International Development, Bureau for Global Health, Washington, District of Columbia Abstract. A strain of Plasmodium falciparum from Peru was adapted to splenectomized Aotus nancymaae and Aotus vociferans monkeys. The Peru 134/CDC strain of P. falciparum was shown to be resistant to treatment with chloroquine in monkeys and partially resistant to mefloquine and malarone. Genetic mutations in crt, dhfr, dhps, and cytochrome b genes conferring drug resistance were also determined for this Peruvian strain of P. falciparum. INTRODUCTION The adaptation of different strains of Plasmodium falci- parum to in vitro culture and New World monkeys has made possible the use of these infections for vaccine trials and drug studies as well as a number of basic biologic studies. Previ- ously, we have reported the adaptation of isolates of P. fal- ciparum from El Salvador, Panama, Haiti, West Africa, East Africa, Cambodia, Indochina, Malaysia, Vietnam, and Ghana. 1–11 These newer isolates are being adapted to provide a range of parasites for heterologous vaccine challenges and for the testing of candidate drug combinations. Of renewed interest has been the adaptation of strains from the Amazon basin to Aotus nancymaae and Aotus vociferans, 2 species of animals currently available for laboratory studies. Our goal has been 1) to obtain parasites representative of parasites currently circulating in endemic areas; 2) to passsage the para- sites in as few animals as possible to determine the predict- ability of parasitemia; and 3) isolate and maintain the para- sites that produce infective gametocytes. In recent years, fewer isolates of P. falciparum have been adapted to nonhu- man primates, primarily because of the lack of Aotus lemuri- nus greiseimembra monkeys. For some reason, these animals are much more susceptible to primary adaptation of human isolates of malaria parasites than other species of Aotus mon- keys. However, once a parasite is established in this host, it generally can then be passaged to another susceptible species of Aotus or other nonhuman primate species. A. l. greiseimem- bra monkeys have not been exported to the United States from Colombia for several decades, and laboratory-bred animals are rarely available in sufficient numbers for these studies. Nevertheless, we report here the adaptation of a strain of multi-drug resistant P. falciparum originating in Peru to New World monkeys. The isolate designated as Peru 01-134 was received from the U.S. Naval Medical Research Center (NMRCD, Lima, Peru). The results of some of the studies conducted with this parasite in different New World monkeys are also reported. MATERIALS AND METHODS A. nancymaae and A. vociferans monkeys were wild-caught animals imported from Peru. A. l. griseimembra monkeys were laboratory-born animals. Upon arrival at the facility, all animals were quarantined for a 2-month conditioning period, weighed, and tested for tuberculosis. Parasitologic and sero- logic examination indicated that the animals were free of in- fection with malaria parasites before inoculation. All mon- keys were splenectomized before exposure to infection. All surgeries were performed in an AAALAC (Association for the Assessment and Accreditation of Laboratory Animal Care, International, Inc.)-approved surgical suite appropriate for aseptic surgery. Protocols were reviewed and approved by the Centers for Disease Control and Prevention Institutional Animal Care and Use Committee, in accordance with proce- dures described in the 1986 U.S. Public Health Policy. Aotus monkeys generally were housed doubly or in some cases singly to avoid injuries caused by fighting with cage mates. Space recommendations for laboratory animals were followed as set forth in the NIH Guide for the Care and Use of Laboratory Animals. All animals were fed a diet that has been proven to provide adequate nutrition and calories in captive Aotus monkeys used in malaria-related research. Feed was free of contaminants and freshly prepared. Daily observations of the animals’ behavior, appetite, stool, and condition were recorded. All were treated as medical condi- tions arose by an attending veterinarian. Anopheles freeborni (F-1 strain originally from California), Anopheles gambiae (originally from The Gambia), Anopheles quadrimaculatus (originally from southeastern United States), and Anopheles stephensi (originally from Delhi, In- dia) were laboratory-reared and maintained at the CDC/DPD insectaries. During periods when gametocytes were present in blood, mosquitoes were allowed to feed on tranquilized mon- keys as previously described. 12 After feeding, mosquitoes were held in an incubator at 25°C until examined 1 week later for the presence of oocysts on their midguts. Blood-stage parasitemia was monitored and quantified by the daily examination of thick- and thin-blood films by the method of Earle and Perez. 13 Infections were terminated by treatment with various combinations of drugs as described. Drugs were administered by oral intubation. DNA was purified by the Qiagen method from Peru 01-134 parasites collected from Aotus monkeys at various times in * Address correspondence to William E. Collins, Mail Stop F-36, Division of Parasitic Diseases, Centers for Disease Control and Pre- vention, 4770 Buford Highway, Chamblee, GA 30341. E-mail: wec1@cdc.gov Am. J. Trop. Med. Hyg., 77(2), 2007, pp. 261–265 261