An Unusual Case of Blastomycosis and Severe Lung Necrosis in a Hmong Woman With Preexisting Liver Cirrhosis.

Predisposing factors that lead to blastomycosis remain unknown, although like other fungal infections, blastomycosis is an opportunistic infection. Here, we report on an unusual presentation in a Hmong woman with preexisting liver disease. This case highlights genetic and medical factors that may increase susceptibility to blastomycosis.

Effect of thioridazine on experimental cutaneous staphylococcal infections.

BACKGROUND: Some non-antibiotic drugs, such as the phenothiazine antipsychotic agents, may have antimicrobial activity. MATERIALS AND METHODS: We sought to determine the in vivo antimicrobial effects of the phenothiazine thioridazine in two mouse models of Staphylococcus aureus skin infection. RESULTS: Thioridazine significantly suppressed dissemination from skin to spleen and kidney after inoculation of the skin surface. However, the drug did not affect infection parameters in the skin itself. Thioridazine did suppress the size of abscesses produced when the bacteria were injected intradermally. On the other hand, using the cutaneous abscess model we were not able to demonstrate synergistic activity between thioridazine and the ß-lactam drug cefazolin against methicillin-resistant S. aureus, as previously demonstrated in vitro. CONCLUSION: The phenothiazine drug thioridazine has in vivo antimicrobial activity against certain S. aureus skin infections, although the previously-demonstrated reversal of methicillin resistance by this agent may not be readily evident in vivo.

Direct translocation of staphylococci from the skin surface to deep organs.

BACKGROUND: Staphylococcus aureus can invade the bloodstream and cause bacteremic infections, but this organism frequently produces serious deep infections without bacteremia or an identifiable portal of entry. METHODS: We used experimental cutaneous S. aureus infections in mice to determine if the bacteria could reach deep organs without travel through the bloodstream. RESULTS: After skin surface application the bacteria rapidly distributed to lymph nodes, spleen, kidneys and other organs. In these animals, blood cultures were negative, dissemination was more efficient after surface application than injection near dermal blood vessels, and kidney bacterial localization sites were unlike those of bacteremic infections. Whereas normal mice eventually cleared bacteria from the deep sites, those with prolonged immunosuppression became moribund from these infections; they also had negative blood cultures and kidney localization not consistent with hematogenous dissemination. Bacteria were also found in the intervening abdominal wall outside the spleen and kidney sites, suggesting direct movement of the organisms from the skin surface through connecting tissues. CONCLUSIONS: Although capable of hematogenous dissemination, S. aureus can also spread from skin to deep organs by a non-bacteremic process. In this case the bacteria appear to migrate directly from the skin surface to the deep organs below.

Clearance of experimental cutaneous Staphylococcus aureus infections in mice.

Staphylococcal skin infections are quite common in human patients. These infections often clear spontaneously, but may also progress locally and/or disseminate to cause serious and sometimes fatal deep infections. The present studies were undertaken to examine the clearance phase of experimental cutaneous Staphylococcus aureus infections in a mouse model system. Previous work in this system has shown that staphylococci applied to the skin rapidly disseminate to the spleen and kidney. In the present experiments the bacteria were found to persist at the skin infection site at a time (8 days after inoculation) when they had disappeared from the spleen and kidney. Examination of the infected skin at earlier times revealed rapid (within 6 h) invasion into the stratum corneum, stratum Malpighii, and dermis, but subsequent redistribution of bacteria (at 1-2 days) to more superficial sites, particularly crusts located just above the skin surface. The crusts seen in these infections were of two distinct types, which were termed type 1 and type 2. Type 1 crusts appeared first, consisted of bacteria, inflammatory cells, and debris, and developed over an intact epidermis. Type 2 crusts arose from the process of dermal necrosis previously reported to take place at 2 days in this model system. In the latter situation the bacteria were not really cleared from the epidermis and dermis; rather those layers were transformed into a superficial crust that contained the bacteria. Deep hair follicle infections in the dermis were found in these infections, but they did not persist and did not seem to be a reservoir for organisms in the dermis. Resolution of these experimental infections appeared to involve redistribution of invading bacteria to more superficial locations in crusts above the skin surface, marked proliferation of the epidermis, loss of the bacteria-laden crusts from the skin, and eventual healing of the cutaneous damage.

Systemic dissemination and cutaneous damage in a mouse model of staphylococcal skin infections.

Serious staphylococcal infections frequently begin in the skin. The present study used a mouse model of such infections to evaluate the ability of Staphylococcus aureus to disseminate from the skin and to determine if cutaneous damage from the infections was required for dissemination. The mice were inoculated with S. aureus onto flank skin prepared by a tape-stripping method that caused minimal disruption of the epidermal keratinocyte layers. After these inoculations the staphylococci were found to disseminate to the spleen and kidneys of almost all animals within 6h. Induction of leucopenia did not affect this process. Cutaneous damage was prominent in these experimental infections and included loss of the epidermis, neutrophil infiltration into the epidermis, and complete necrosis of the dermis. The latter also occurred in cyclophosphamide-treated animals, indicating that the organisms themselves and not the host inflammatory responses were responsible. Dermal necrosis did not develop until 48h after inoculation, a time by which dissemination had already occurred. Therefore, in this mouse model system S. aureus is capable of penetrating the epidermal keratinocyte layers and disseminating rapidly after inoculation; the experimental infections do produce significant dermal damage, but the latter develops after dissemination has already taken place.

Resistance of athymic nude mice to experimental cutaneous Bacillus anthracis infection.

BACKGROUND: Previous studies in a murine cutaneous anthrax model have demonstrated that hairless and haired HRS/J mice are extremely resistant to Bacillus anthracis. Because these mice are relatively thymus deficient, we used C57BL/6 athymic nude and euthymic mice to evaluate the relationship between T cell deficiency and this heightened resistance. METHODS: Animals were epicutaneously inoculated with 1 X 10(7) B. anthracis (Sterne) spores onto abraded skin or injected with the spores intradermally or subcutaneously. The mice were then either monitored for survival or killed for quantitative histological experiments. RESULTS: Athymic mice were found to be markedly resistant to all 3 inoculation routes, compared with euthymic C57BL/6 mice. Athymic mice rendered leukopenic with cyclophosphamide became susceptible. Histological examination demonstrated increased inflammation and absence of organisms in the skin of athymic mice, compared with euthymic ones. The numbers of organisms in the athymic animals increased markedly after cyclophosphamide treatment. Superficial exudate fluids of inoculated skin showed many more neutrophils and ingested bacilli in the athymic mice. CONCLUSIONS: These experiments demonstrate that athymic nude C57BL/6 mice are markedly resistant to experimental cutaneous anthrax, apparently because of a superficial neutrophilic response that clears the inoculated organisms before they can invade the underlying skin.

Progressive and destructive hair follicle infections in a murine cutaneous anthrax model.

Hair follicles may allow pathogen entry because they represent potential barrier defects and because there is immunological privilege within actively growing follicles. Experimental cutaneous Bacillus anthracis infections in mice have previously shown prominent organism invasion and proliferation within hair follicles. For the present study, C57BL/6 mice were inoculated with B. anthracis (Sterne) spores onto abraded skin with either anagen (actively growing) or telogen (inactive) hair follicles; skin samples were evaluated by histologic methods and electron microscopy. The infections were found to progress similarly in either anagen or telogen hair follicles, with bacilli occasionally invading deeper sites in anagen hair follicles. The infections progressed from the surface inward, rather than growing outward from within the follicles. Infecting bacilli destroyed the hair follicle keratinocytes and were initially not contacted by inflammatory cells within the follicles. However, at 3-4 days after inoculation, inflammatory cells did contact and disperse the massed follicle bacilli and led to apparent resolution of the follicle infections. Therefore, in this model system B. anthracis initially attacks superficial sites in active or inactive hair follicles and then progresses inward, producing destructive infections of the hair follicles; these infections clear when the massed bacilli are eventually contacted and dispersed by inflammatory cells.

Superficial exudates of neutrophils prevent invasion of Bacillus anthracis bacilli into abraded skin of resistant mice.

Skin window procedures in humans have shown rapid accumulation of neutrophils into the exuded fluids above abraded skin. The present study was undertaken to determine if similar epicutaneous neutrophil accumulation might explain the extreme resistance of HRS/J mice, both hairless (hr/hr) and haired (hr/+), to experimental cutaneous Bacillus anthracis Sterne infections on abraded skin. In this study, very early (6 h) biopsies demonstrated a lack of bacilli in skin from the HRS/J hr/hr mice, indicating that the organisms never did invade in these animals as opposed to early skin entry and then efficient clearance by host responses in the tissues. Touch preparations of either the inoculation filter or the skin surface revealed more inflammatory cells, fewer bacilli, and a higher percentage of cell-associated bacilli in the HRS/J hr/hr mice than in comparator strains. In the HRS/J mice, cyclophosphamide treatment or separation of inoculated spores from the inflammatory infiltrates by a second filter below both produced marked increases in the number of bacilli observed. Examination of inoculation filter specimens demonstrated ingestion of spores and bacilli by neutrophils inside the filter at 6 h after inoculation. These findings suggest that an early and vigorous inflammatory cell infiltrate in HRS/J mice attacks the inoculated organisms above the skin surface and does not allow them to invade the tissues below.

Characteristics of spore germination in a mouse model of cutaneous anthrax.

BACKGROUND: Cutaneous infection is the most common form of human anthrax, but little is known about Bacillus anthracis spore germination in these infections. METHODS: We used experimental inoculations of B. anthracis Sterne spores or vegetative bacilli onto intact or abraded mouse flank skin, followed by evaluation of the infections and enumeration of germinating spores and vegetative bacilli. RESULTS: Bacilli developed from a spore inoculum after application onto abraded, but not intact, skin of the mice. Germination appeared to occur extracellularly at the skin surface before the development of a phagocytic response; in fact, vegetative bacilli were seen after inoculation of the spores on top of a filter that separated them from the host phagocytic cells below. Malachite green staining demonstrated that spores began germinating 1-3 h after inoculation onto abraded skin. Vegetative bacilli were found not to be capable of initiating infection in the absence of cutaneous abrasion. CONCLUSIONS: The results indicate that epidermal damage is required for germination of B. anthracis spores in these infections; even so, spore germination by itself is not sufficient to produce infection of undamaged skin. In contrast to events in experimental inhalational anthrax, spore germination in these cutaneous infections appears to occur extracellularly.

Experimental cutaneous Bacillus anthracis infections in hairless HRS/J mice.

Previous studies of experimental Bacillus anthracis cutaneous infections in mice have implicated hair follicles as a likely entry site. Hairless HRS/J mice were used to investigate this possibility because of their non-functional hair follicles that lack penetrating hair shafts. These mice also have diminished macrophage function, increased susceptibility to Listeria, and enhanced neutrophil responses. HRS/J and Balb/c mice were found to be resistant to epicutaneous inoculation with Bacillus anthracis (Sterne) spores onto abraded skin when compared with DBA/2 mice or leucopenic C57BL/6 mice. The HRS/J mice also resisted spore injections that bypassed hair follicles. Haired HRS/J heterozygote mice demonstrated similar reduced susceptibility to B. anthracis spores. Hairless HRS/J mice that were made leucopenic did become susceptible to the epicutaneous spore inoculations. Histologically, the hairless and haired HRS/J mice showed markedly reduced numbers of organisms in hair follicles and the interfollicular dermis when compared even with the resistant Balb/c mice; inflammatory cell infiltrates in the superficial dermis were increased in the HRS/J mice compared with more sensitive strains. Therefore, resistance in the HRS/J mice was apparent at the initial site of epicutaneous inoculation and seemed related to an accumulation of dermal neutrophils rather than to a lack of functional hair follicles.

Analysis of epidermal entry in experimental cutaneous Bacillus anthracis infections in mice.

Cutaneous infection is the most common form of human anthrax, but little is known of Bacillus anthracis-epidermal interactions. To study the latter, we used experimental inoculations of B. anthracis Sterne spores onto mouse flank skin. In DBA/2 mice (a sensitive strain) 10(7) spores injected intradermally or applied under occlusive dressings to abraded skin produced ipsilateral inguinal edema and rapid death. Epicutaneous application to shaved-only skin produced edema and death in most animals, but at longer times. Mortality after inoculation onto abraded skin was less in C57BL/6 mice (a relatively resistant strain). Inoculations onto shaved-only skin immunized C57BL/6 mice, and they survived later intradermal spore injections. Histology revealed massive organism proliferation in remaining epidermis and hair follicles of inoculated abraded skin, but less growth in the dermis itself. Conversely, no foci could be located by microscopic examination after inoculation onto shaved-only skin. High-dose nonocclusive dressing inoculations onto unshaved skin in DBA/2 mice revealed small numbers of infective foci, all in hair follicles. These results suggest that epidermal damage may increase infection susceptibility to B. anthracis of hair follicle contents and remaining epidermal remnants; the findings also indicate that access may occur through hair follicles and the denuded dermis.

Fungal susceptibility to zinc deprivation.

Calprotectin is a neutrophil-derived antimicrobial protein that competes with microorganisms for zinc. The zinc-specific effect of calprotectin against Candida albicans appears to be related to this organism's marked susceptibility to deprivation of this metal. However, it is not known whether this susceptibility is particular to C albicans or whether it is a characteristic of pathogenic fungi in general. As a means of deciding between these 2 possibilities, we undertook the study reported here to compare the susceptibility to zinc deprivation of 6 other pathogenic fungal species in addition to that of C albicans . We tested the effect of metals in reversing growth inhibition of the 7 fungi against abscess-fluid supernatant (a source of calprotectin) and 3 chemical chelators. Data were expressed as the concentration of metal required to bring about 50% restoration of growth. Zinc was found to be much more potent than the other metals tested in reversing growth inhibition of all the organisms by human abscess fluid and all 3 chemical chelators. Copper and manganese also had some effect. In some cases, chelator stability constants were higher for other metals than for zinc; in particular, although diethylenetriaminopentaacetic acid has a stability constant for iron almost 10(10) times greater than that for zinc, zinc was more effective than iron in reversing growth inhibition by this chelator against all of the organisms. These results suggest that marked susceptibility to zinc deprivation is a general characteristic of pathogenic fungi.

Effect of prolonged fluconazole treatment on Candida albicans in diffusion chambers implanted into mice.

Fluconazole is an azole agent with primarily fungistatic activity in standard in vitro susceptibility tests. The present study was undertaken to develop a diffusion chamber model system in mice in order to study the in vivo effects of prolonged fluconazole treatment on Candida albicans. Chambers containing 100 C. albicans yeast cells were implanted subcutaneously on the flanks of C57BL/6 mice and were then retrieved 6 or 14 weeks later (after fluconazole treatment for 4 or 12 weeks, respectively). Leukocyte counts demonstrated that implantation of the chambers did elicit an inflammatory response but that only small numbers of inflammatory cells were able to enter the chamber interior. Treatment with fluconazole at 10 mg/kg of body weight/day for 12 weeks not only reduced the numbers of viable organisms within the chambers compared to those in untreated mice (mean +/- standard deviation of log(10) CFU of 0.7 +/- 1.2 versus 2.3 +/- 2.0; P < 0.001 by the Bonferroni test) but also increased the numbers of chambers that became sterile over the treatment period (14 of 16 versus 6 of 19; P = 0.0009 by the chi-square test). However, treatment for only 4 weeks had minimal effects on the numbers of chamber CFU, and none of the chambers became sterile during this period. Distribution of retrieved organisms between interior fluid and the chamber filters was approximately equal in all the treatment groups. This model system appears to be useful for evaluating the effects of antifungal drugs over prolonged periods in vivo. Its use in the present study demonstrates that fluconazole can increase the rate of sterilization of C. albicans foci that are protected from the host's inflammatory response.

Blastomycosis: clinical and immunologic aspects

Dr. Basil Varkey: Blastomycosis was first described by Gilchrist in 1894. For some 50 years thereafter, progress in the understanding of the various aspects of this disease evolved very slowly.