In their examination of the influence of aminoguanadine treatment of mice on susceptibility toCoccidioidesinfection, the same authors have pointed out that this compound is not a specific inhibitor ofiNOSexpression and the resulting increase in susceptibility to disease, therefore, may not be due directly to reduced NO production as suggested. by an as yet unidentified, secreted fungal factor(s). Heat-, paraformaldehyde- or X ray-treated spherule initials did not show this inhibitory effect. To our surprise, macrophages obtained fromiNOS-deficient mice revealed phagocytic activity and killing efficiency which were comparable to that of macrophages isolated from wild type Felbamate C57BL/6 mice. Although the cultured fungal pathogen can suppress NO production, this oxidative product is apparently not essential forin vitrokilling ofCoccidioidesby activated macrophages. Our results suggest that other unidentified fungicidal mechanisms exist againstCoccidioideswhich are apparently independent of NO production. Keywords:nitric oxide, macrophage suppression,Coccidioides == 1. Introduction == Coccidioidesis the causative agent of coccidioidomycosis or San Joaquin Valley fever, one of the endemic mycoses in the United States with an estimated incidence of respiratory infection of 100,000 people annually [1]. This mycosis is also endemic to other regions of the Western Hemisphere, including Northern Mexico, Guatemala, Honduras, Colombia, Venezuela, Paraguay, and Argentina [2]. Human infection occurs primarily after inhalation of the fungal spores (arthroconidia). The majority of cases of coccidioidomycosis are either asymptomatic or result in self-limited pneumonia. Although this mycosis is rarely life threatening, most patients who do not recover spontaneously develop extrapulmonary infections [1]. Felbamate Coccidioidesencompasses two species,C. immitisandC. posadasiibased on their genetic and biogeographical differences [35]. Felbamate However, laboratory animal studies of the experimental pulmonary disease caused byC. immitisorC. posadasiihave shown no significant differences in the morphogenesis or virulence of the two pathogens [6]. Coccidioidesinteraction with host phagocytes during disease onset is characterized by both an extracellular and intracellular relationship, which is dependent at least in part on the size of the parasitic cells. Arthroconidia are barrel-shaped cells that measure 2.5 to 3 m in width by 3 to 6 m Felbamate in length. They are small enough to reach the alveoli if inhaled and are readily internalized by phagocytic cells. If these fungal cells survive exposure to the host defenses, they germinate by isotropic growth and give rise to small, round cells (spherule initials; approx. 1520 m in diameter) that continue to enlarge and differentiate into coenocytes (spherules). The latter typically range from 40 to 120 m in diameter [7, Alas2 8] and are too large to be engulfed by phagocytes. These multinucleate, parasitic cells subsequently undergo a process of cytoplasmic segmentation, which culminates in the formation of a multitude of endospores each of which are approximately 10 m in diameter at the time of their release from the mature spherules. Endospores are also susceptible to phagocytosis by host innate immune cells. Nitric oxide (NO) is known to be an important component of the arsenal of host oxidative defenses directed against microbial pathogens. NO is produced by enzymatic activity of the inducible isoform of nitric oxide synthase 2 (iNOSorNOS2), which is expressed by phagocytic cells (primarily macrophages). Host cytokines, such as IFN- and tumor necrosis factor (TNF), as well as microbial products, especially LPS, have been shown to activate and induce the synthesis of NO by macrophages [9]. The role of NO in host defense against several fungal pathogens has been previously discussed [1014]. Mice which were genetically engineered to produce high levels of human interleukin (IL)-10 in response toCoccidioidesinfection were shown to be more susceptible to acute coccidioidal peritonitis and pneumonia and expressed lower levels of gamma IFN-, IL-12p40 andiNOSmRNA in their lungs than untreated mice [1]. The authors proposed that the reduced level ofiNOSexpression and increased susceptibility to infection of the transgenic mice implicated NO synthesis as a mechanism of resistance against coccidioidomycosis. In addition, these same investigators revealed that disease-resistant DBA/2 mice treated with aminoguanidine, an inhibitor of NO, increased their susceptibility to coccidioidal infection which further supported their notion that nitric oxide is directly involved in defense against this pathogen. In the present study we report the ability ofCoccidioidesto significantly reduce the level.