Interaction of Human Phagocytes with Pigmentless Aspergillus Conidia

Abstract

A defect in the pksP gene of Aspergillus fumigatus is associated with the loss of conidial pigmentation, a profound change of the conidial surface structure, and reduced virulence. The structural change of the conidial surface structure was not observed in similar A. nidulans wA mutants. Our data indicate that the pigment of both species is important for scavenging reactive oxygen species and for protection of conidia against oxidative damage.Aspergillus spp. are the predominant causative agents of invasive pulmonary aspergillosis (IPA), an often lethal infection of the immunocompromised host (4,10,13,16). Since conidia are the infectious agent in IPA, recent studies focused on the elucidation of conidial factors contributing to pathogenicity (8,17). Previously, we and others have shown that conidia lacking pigmentation due to the defective polyketide synthase gene pksP were less resistant to the attack by monocytes in vitro and showed reduced virulence in a murine animal model (8,9,17,18). During these studies, it became apparent that coincubation of human phagocytes with pksP mutant conidia resulted in a marked increase in the release of reactive oxygen species (ROS) compared with wild-type (wt) conidia (8). Since a defective pksP gene not only impaired conidial pigmentation but concomitantly resulted in profound alterations of the conidial surface (8, 9), the question arose as to whether the large amounts of ROS detected after incubation of phagocytes with pksP mutant conidia were due to a change in the activation pattern of the cells or, alternatively, reflected the lack of ROS quenching capacity caused by the loss of conidial pigment. To address this question, conidia of wt strains of both Aspergillus fumigatus and the nonpathogenic fungus Aspergillus nidulans were compared with their respective pigmentless mutant strains.The WA mutant of A. nidulans (strain WG370; wA3 bgaO biA1) lacking the conidial pigment due to a defective polyketide synthase gene (wA) was constructed by a sexual cross of appropriate parental strains (12) using standard genetic techniques (14). The wA gene product might have a function similar to that of the pksP gene product of A. fumigatus, although major differences between the pigment biosyntheses of the two Aspergillus species exist (2, 18; this study).As previously reported, conidia of the A. fumigatus wt strain showed a rough surface; i.e., they had an ornamentation which was lacking in the pksP mutant strain (8, 9) ( Fig. 1A and B). The A. nidulans wt conidia showed a similar ornamentation (Fig. 1C). In contrast to the pigmentless pksP mutant strain of A. fumigatus, however, similar pigmentless conidia of A. nidulans (wA) still exhibited the ornamentation characteristic of wild-type conidia (Fig. 1D). Taken together, the difference in surface structure between the A. fumigatus pksP mutant and the A. nidulans wA mutant further supports the assumption that different pathways exist for either conidial pigment biosynthesis or pigment deposition in the two species (1). As was noted p...

Differential Expression of the Aspergillus fumigatus pksP Gene Detected In Vitro and In Vivo with Green Fluorescent Protein

“…We and others have shown that PKSP is an important factor in A. fumigatus virulence (12,14,28,29). However, although the pigment is able to scavenge ROS, it seems unlikely that this is the major PKSP-dependent virulence determinant (13). This is also supported by preliminary results suggesting that the biosynthesis steps immediately succeeding those catalyzed by PKSP appear to have little impact on virulence (29).…”

“…Conidia of a pksP mutant strain showed reduced virulence in a mouse infection model and an altered surface structure compared with wild-type conidia. Furthermore, wild-type conidia were 10-to 20-fold more resistant against reactive oxygen species (ROS) than pksP mutant conidia and were able to scavenge ROS, presumably thereby detoxifying ROS (12,13).…”

LaeA, a Regulator of Morphogenetic Fungal Virulence Factors

“…However, few null mutants have demonstrated avirulent phenotypes. For example, singleand multiple-gene disruption in a pigment biosynthesis gene cluster has been associated with decreased virulence in mouse models without inducing a concomitant alteration in microbial fitness; this defect is associated with increased microbial susceptibility to oxidative killing by monocytes and polymorphonuclear neutrophils (PMNs) (13,34,37).…”

Fungal melanin stimulates surfactant protein D–mediated opsonization of and host immune response to Aspergillus fumigatus spores

“…In our study, we followed an alternative approach; the inoculation of WT mice with SP-D-opsonized melanin ghosts augmented cytokine levels compared to unopsonized melanin ghosts, confirming the direct immunomodulatory effect of SP-D. In addition, melanin has been shown to quench reactive oxygen species (ROS), one of the defensive mechanisms of the human immune system (30,31), prevents phagocytosis to certain extent and (30) and also binds metals, acting as a physiological redox buffer, thereby contributes as a sink for harmful unpaired electrons (43). However, upon opsonization by SP-D, melanin lost its ROS quenching capacity.…”

“…fumigatus conidial melanin has been shown to quench reactive oxygen species (ROS), one of the defensive mechanisms of the human immune system (30,31). We evaluated ROS quenching property of melanin.…”