Date Published: May 23, 2019
Publisher: Public Library of Science
Author(s): Rachel M. Olson, Deborah M. Anderson, Matthew B. Lawrenz.
Yersinia pestis is the causative agent of pneumonic plague, a disease involving uncontrolled bacterial growth and host immunopathology. Secondary septicemic plague commonly occurs as a consequence of the host inflammatory response that causes vasodilation and vascular leakage, which facilitates systemic spread of the bacteria and the colonization of secondary tissues. The mortality rates of pneumonic and septicemic plague are high even when antibiotics are administered. In this work, we show that primary pneumonic or secondary septicemic plague can be preferentially modeled in mice by varying the volume used for intranasal delivery of Y. pestis. Low volume intranasal challenge (10μL) of wild type Y. pestis resulted in a high frequency of lethal secondary septicemic plague, with a low degree of primary lung infection and rapid development of sepsis. In contrast, high volume intranasal challenge (30μL) yielded uniform early lung infection and primary disease and a significant increase in lethality. In a commonly used BSL2 model, high volume challenge with Y. pestis lacking the pigmentation locus (pgm-) gave 105-fold greater deposition compared to low volume challenge, yet moribund mice did not develop severe lung disease and there was no detectable difference in lethality. These data indicate the primary cause of death of mice in the BSL2 model is sepsis regardless of intranasal dosing method. Overall, these findings allow for the preferential modeling of pneumonic or septicemic plague by intranasal dosing of mice with Y. pestis.
Historically Yersinia pestis has caused three pandemics, manifesting as bubonic, pneumonic and septicemic plague. Plague is usually a flea-borne disease and can occur in most mammals. Ecologically important flea-rodent transmission cycles are responsible for maintaining Y. pestis in areas throughout the world, and many years can pass between recognized human or animal outbreaks . Upon deposition in the skin by flea bite, bacteria traffic to the draining lymph node and rapidly multiply. Vascular spread results in bacteremia and colonization of secondary and tertiary immune tissues as well as the lungs and liver. Secondary septicemia progresses rapidly to lethality due to disseminated vascular coagulation, acute respiratory distress syndrome, and multi-organ failure . Although antibiotic treatment of bubonic plague is usually successful, the mortality rate of pneumonic and septicemic plague is high . Even in countries with strong medical infrastructure, such as the US, recently reported annual mortality rates have been as high as 25%, with no age or sex bias in disease susceptibility . Furthermore, reemergence of pneumonic plague has occurred in Madagascar, where in a 2017 outbreak 85% of the 2,500 cases were the pneumonic form . Multi-drug resistant Y. pestis strains have been isolated from plague patients and are believed to evolve naturally in the flea-rodent enzootic cycle [6–9].
In this work, we identified a significant impact of dosing volume on the development of pneumonic or septicemic plague as the likely cause of death following intranasal challenge of mice with wild type Y. pestis. These data have important implications on plague research and the early testing of candidates for new vaccines, therapeutics and diagnostics. Retention in the upper respiratory tract that results from low volume intranasal instillation leads to a high frequency of secondary septicemic plague, with multi-organ failure as evidenced by severe hepatocyte necrosis and elevation of cytokines and liver enzymes in the serum in mice as they succumb to infection. Presumably this occurs via the nasal sinuses when a high degree of bacteria reside in the nasal passages. In contrast, high volume administration results in reproducibly uniform bacterial deposition in the lower respiratory tract and a high frequency of primary pneumonic plague, with bacterial microcolonies but few inflammatory foci in the secondary tissues of moribund mice. Similar pathology in lungs, liver and spleen was reported in moribund mice that had been aerosol challenged indicating high volume intranasal challenge is a reasonable substitute for aerosol . With isoflurane anesthesia, we routinely observed 5–10% deposition using 30μL dosing volume, and even very low doses resulted in high fidelity lower respiratory tract deposition and disease. Furthermore, we found that 20μL dosing volume, a method commonly used in the field, was more similar to 30μL than it was to 10μL, but resulted in more variability, mixed results, and reduced the power of the study.