Anthrax is one of the most ancient and lethal human diseases caused by the virulent (toxigenic and encapsulated) strains of Bacillus anthracis
. In humans, anthrax may take three forms depending upon the route of infection, namely, pulmonary (inhalational), cutaneous, and gastrointestinal, with pulmonary being the most lethal and difficult to treat. While the naturally acquired pulmonary anthrax is relatively rare (occurring only ~5% as often as cutaneous anthrax), this form has a high potential for misuse as a weapon of bioterrorism, considering that environmental dissemination is the most expected mode of release of the agent in mass attacks (26
). Pulmonary anthrax often proves fatal, with mortality approaching 100% if not treated early. Anthrax spores have long been considered a potential agent of biological warfare (25
). Prior to the 11 cases of pulmonary anthrax that occurred via the U.S. mail delivery system in 2001 (29
), fatal cases of pulmonary anthrax in the United States and other countries have been associated with occupational and accidental exposures (37
). But the bioterrorism attack in 2001 exposed a new cause for serious concern for future and more widespread attacks on military and civilian populations.
The etiological agent for anthrax, Bacillus anthracis
, is a Gram-positive bacterial pathogen that belongs to category A in the CDC list of select agents. The size of its spores (1 to 2 μm) makes them optimal for deposition in the alveolar spaces following inhalation. To be virulent, B. anthracis
requires two major components: the capsule and the anthrax toxin. These components are encoded by the plasmids pXO2 and pXO1, respectively, in virulent strains such as Ames. Strains such as Sterne, which lack the capsule (pXO2), are attenuated for human infection and are therefore used for vaccine applications. However, these attenuated strains can cause serious infections in mice. While there is increasing information on the pathogen virulence factors in B. anthracis
), the mechanisms of host defense and pathogenesis are poorly understood for anthrax.
Although anthrax primarily affects herbivores, virtually all mammals are susceptible to various degrees. Anthrax susceptibility depends on host species (31
), strain (69
), and the route of infection (35
). Interindividual differences in disease onset, survival, and treatment response have been observed in human cases of pulmonary anthrax. In the 2001 anthrax letter attacks, only 5 of the 11 individuals infected by the aerosolized spores succumbed to the disease. The causes underlying these differences in susceptibility are not known but conceivably involve differences in host factors. It is intriguing that some animals (e.g., rats) are very sensitive to B. anthracis
toxins yet are difficult to infect by spores, while other animals (e.g., guinea pigs) are more resistant to the toxins but can be killed with relatively few spores (31
). Previous investigations on the genetic susceptibility to B. anthracis
have mainly focused on the role of anthrax lethal toxin (LT) exposure in vitro
) and in vivo
) in laboratory rodent strains. The toxin-based studies indicated that inbred mice vary in their sensitivity to anthrax LT and that host genetic factors underlie the differences in LT susceptibility (4
). More importantly, previous observations have revealed a reciprocal trend for murine susceptibility to purified anthrax LT-induced pathology versus spore-induced infection (36
). Since spore-induced anthrax better represents a real world infection scenario for bioterrorism, there is an immediate need to fully understand the basis of host defense and susceptibility to B. anthracis
spores. Knowledge on host susceptibility to lethal infection during pulmonary anthrax can be a key initiator in the development of new countermeasures.
Lethal anthrax infection caused by B. anthracis
spores is a multistep process expected to require spore germination, pathogen multiplication, and systemic dissemination, culminating in death (15
). Mechanisms of host resistance could interfere with any of these steps. Hence, there is a need to understand the genetic loci conferring host resistance/susceptibility to anthrax infection. In this context, there is also a continuing need to identify and characterize more appropriate animal models of resistance or susceptibility for dissecting the role of these host factors. Initial investigations in this direction have examined questions of pathogenesis and host resistance in inbred mice by using spores from avirulent anthrax strains, such as Sterne or 34F2 (55
). While inbred mice offer a valuable tool to understand host susceptibility and response to anthrax infection, little has been done in mice in experiments with the fully virulent strains of B. anthracis
. Limited initial studies of mice either used fewer numbers of randomly selected inbred strains (35
) or were based on nonpulmonary routes (69
) of infection, and they did not identify host genomic regions governing the underlying susceptibility. Whether males and females differ in pulmonary anthrax susceptibility is also not known.
The overall goals of this study were to investigate the role of host genetic background and sex in susceptibility to pulmonary anthrax infection by using the Ames strain and to identify putative genomic loci underlying differential susceptibility by quantitative trait locus (QTL) analysis. The QTL approach, based on associating genetic variation with phenotypic variation, has proven useful for identification of genomic loci involved in many complex diseases, including infectious diseases (16
). To begin to examine the role of host genetics, we screened a phylogenetically representative panel of 14 inbred mice strains and F1 progeny from select sensitive-resistant strain breeder pairs. Initial studies established mouse models of host susceptibility and also revealed evidence of a parent-of-origin effect and a differential sex response to pulmonary anthrax. To identify genetic loci responsible for B. anthracis
susceptibility, two strategies were used. First, genome-wide single-nucleotide polymorphism (SNP) analysis was performed on a panel of BXD advanced recombinant inbred (ARI) mice. The BXD ARI lines were derived from the female C57BL/6J and male DBA/2J parent strains (48
), which represent a progenitor strain pair identified in this study for modeling differential genetic susceptibility. These homozygous, inbred, and genetically distinct ARI lines approximate the genetic diversity in human populations and have been used successfully to map QTLs associated with various clinical conditions and diseases (2
). Second, we performed QTL analysis on an F2 population generated from the sensitive DBA/2J and resistant BALB/cJ strains. This work represents the first report of sex bias and a parent-of-origin effect in a murine model of spore-induced pulmonary anthrax infection. These initial genome-wide analyses using two separate mouse models of differential susceptibility, coupled with spore-induced anthrax infection, identified multiple genomic loci associated with pulmonary anthrax outcome. These studies lay the groundwork for follow-on studies to further characterize these susceptibility QTLs to identify the relevant genes.