Clinical and Pathologic Features of Cynomolgus Macaques (Macaca fascicularis) Infected with Aerosolized Yersinia pestis

Since the anthrax attacks of 2001, the emphasis on developing animal models of aerosolized select agent pathogens has increased. Many scientists believe that nonhuman primate models are the most appropriate to evaluate pulmonary response to, vaccines for, and treatments for select agents such as Yersinia pestis (Y. pestis), the causative agent of plague. A recent symposium concluded that the cynomolgus macaque (Macaca fascicularis) plague model should be characterized more fully. To date, a well-characterized cynomolgus macaque model of pneumonic plague using reproducible bioaerosols of viable Y. pestis has not been published. In the current study, methods for creating reproducible bioaerosols of viable Y. pestis strain CO92 (YpCO92) and pneumonic plague models were evaluated in 22 Indonesian-origin cynomolgus macaques. Five macaques exposed to doses lower than 250 CFU remained free of any indication of plague infection. Fifteen macaques developed fever, lethargy, and anorexia indicative of clinical plague. The 2 remaining macaques died without overt clinical signs but were plague-positive on culture and demonstrated pathology consistent with plague. The lethal dose of plague in humans is reputedly less than 100 organisms; in this study, 66 CFU was the dose at which half of the macaques developed fever and clinical signs (ED₅₀), The Indonesian cynomolgus macaque reproduces many aspects of human pneumonic plague and likely will provide an excellent model for studies that require a macaque model.Yersinia pestis is the causative agent of plague. Likely more people worldwide have died from Y. pestis infections than from any other single infectious disease.^26,27 Bubonic plague, the most common form of the disease, results when the bacterium is inoculated into the skin, typically by means of flea bites. The resulting cutaneous infection spreads to local lymph nodes; the swollen lymph nodes are known as bubos and often serve as a source of systemic infection. Although less common, the bacterium also can spread by aerosol, causing pneumonic plague. Pneumonic plague can result from pulmonary spread of systemic infection or from deliberate dissemination and is associated with nearly 100% human mortality if left untreated. Y. pestis is susceptible to commonly available antibiotics if treatment begins soon after infection. However, depending on the route of infection, the time at which infection is confirmed is often too late for antibiotics to prevent significant morbidity or mortality.¹⁰ Because pneumonic plague is the form most likely to be seen in bioterrorism events,¹⁶ interest in animal models has arisen to support development of vaccines and improved therapeutics.Potential vaccines and therapeutic agents for plague must protect against the pneumonic disease, but contemporary published data regarding disease pathogenesis using aerosolized Y. pestis pathogenesis in nonhuman primates are scant.^4,9,21,23,24 In the United States, when vaccine or antibiotic efficacy cannot be evaluated in humans, an animal species that is reasonably expected to recapitulate human disease must be used.⁹ For many biothreat agents such as plague, a nonhuman primate model often is required. Although some laboratories have examined the cynomolgus macaque model of aerosolized plague briefly,¹ no published reports fully characterize this model. Published studies have examined plague in the African green monkey or vervet (Chlorocebus spp., formerly Cercopithecus aethiops) and rhesus macaque (Macaca mulatta).¹ Vervets reportedly are more sensitive to plague than are macaques,^4,24 such that some vervets are susceptible to infection with vaccine strains, casting some doubt on applicability of this species for plague studies.¹ The disease in rhesus macaques differs from that in humans in that rhesus macaques frequently develop disseminated intravascular coagulation (DIC) and chronic pneumonia as a result of pneumonic plague while humans usually develop acute pneumonia without DIC.^1,7Many participants at a recent symposium sponsored by the Food and Drug Administration and National Institute of Allergy and Infectious Disease endorsed the development of a cynomolgus macaque pneumonic plague model to support plague therapeutic and vaccine studies.⁸ The current study was undertaken to evaluate the Indonesian cynomolgus macaque as a model of aerosolized Y. pestis Colorado 92 (YpCO92) for subsequent vaccine and therapeutic trials. We also sought to determine whether fever development could be used to determine a humane endpoint to the study, as an alternative to LD₅₀ methods.