Genetic Mouse Models of Pneumocystis Pneumonia

Hoving, J. Claire; Munyonho, Ferris T.; Kolls, Jay K.

doi:10.1007/978-1-0716-3199-7_13

“…Susceptibility to infection was investigated in wild-type mice from three different genetic backgrounds: BALB/c, C57BL/6 and Sv129. As previously described for other fungi, all mice received a non-invasive intratracheal inoculation of between 0.5 to 1x10 5 yeast cells/50 μl [ 12 ]. Survival curves showed that all wild-type mice were susceptible to this high-dose infection ( Fig 1A ).…”

Section: Resultsmentioning

confidence: 99%

The pathogenesis of experimental Emergomycosis in mice

Höft,

Duvenage,

Salie

et al. 2024

PLoS Negl Trop Dis

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Emergomyces africanus is a recently identified thermally-dimorphic fungal pathogen that causes disseminated infection in people living with advanced HIV disease. Known as emergomycosis, this disseminated disease is associated with very high case fatality rates. Over the last decade, improved diagnostics and fungal identification in South Africa resulted in a dramatic increase in the number of reported cases. Although the true burden of disease is still unknown, emergomycosis is among the most frequently diagnosed dimorphic fungal infections in Southern Africa; and additional species in the genus have been identified on four continents. Little is known about the pathogenesis and the host’s immune response to this emerging pathogen. Therefore, we established a murine model of pulmonary infection using a clinical isolate, E. africanus (CBS 136260). Both conidia and yeast forms caused pulmonary and disseminated infection in mice with organisms isolated in culture from lung, spleen, liver, and kidney. Wild-type C57BL/6 mice demonstrated a drop in body weight at two weeks post-infection, corresponding to a peak in fungal burden in the lung, spleen, liver, and kidney. An increase in pro-inflammatory cytokine production was detected in homogenized lung supernatants including IFN-γ, IL-1β, IL-6, IL12-p40 and IL-17 at three- and four-weeks post-infection. No significant differences in TNF, IL-12p70 and IL-10 were observed in wild-type mice between one and four-weeks post-infection. Rag-1-deficient mice, lacking mature T-and B-cells, had an increased fungal burden associated with reduced IFN-γ production. Together our data support a protective T-helper type-1 immune response to E. africanus infection. This may provide a possible explanation for the susceptibility of only a subset of people living with advanced HIV disease despite hypothesized widespread environmental exposure. In summary, we have established a novel murine model of E. africanus disease providing critical insights into the host immune components required for eliminating the infection.

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“…Susceptibility to infection was investigated in wild-type mice from three different genetic backgrounds: BALB/c, C57BL/6 and Sv129. As previously described for other fungi, all mice received a non-invasive intratracheal inoculation of between 0.5 to 1x10 5 yeast cells/50 l (Hoving, Munyonho et al 2023). Survival curves showed that all wildtype mice were susceptible to this high-dose infection (Fig 1A As many gene-deficient mice have been created on a C57BL/6 background, we focussed most of our subsequent studies on this strain, starting with a time-kinetic study of progression of disease.…”

Section: Wild-type Mice Develop Emergomyces Africanus Infectionmentioning

confidence: 99%

The pathogenesis of experimental Emergomycosis in mice

Höft,

Duvenage,

Salie

et al. 2023

Preprint

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Emergomyces africanusis a recently identified thermally-dimorphic fungal pathogen that causes disseminated infection in people living with advanced HIV disease. Known as emergomycosis, this disseminated disease is associated with very high case fatality rates. Over the last decade, improved diagnostics and fungal identification in South Africa resulted in a dramatic increase in the number of reported cases. Although the true burden of disease is still unknown, emergomycosis is among the most frequently diagnosed dimorphic fungal infections in Southern Africa; and additional species in the genus have been identified on four continents. Little is known about the pathogenesis and the host’s immune response to this emerging pathogen. Therefore, we established a murine model of pulmonary infection using a clinical isolate,E. africanus(CBS 136260). Both conidia and yeast forms caused pulmonary and disseminated infection in mice with fungus subsequently isolated in culture from lung, spleen, liver, and kidney tissue. Wild-type C57BL/6 mice demonstrated a drop in body weight at two weeks post-infection, corresponding to a peak in fungal burden in the lung, spleen, liver, and kidney. An increase in pro-inflammatory cytokine production was detected in homogenized lung supernatants including IFN-γ, IL-1β, IL-6, IL12-p40 and IL-17 at three- and four-weeks post-infection. Rag-1-deficient mice, lacking mature T and B cells, had an increased fungal burden associated with reduced IFN-γ production. Together our data support a protective T-helper type-1 immune response toE. africanusinfection. This may provide a possible explanation for the susceptibility of only a subset of people living with advanced HIV disease despite hypothesized widespread environmental exposure. In summary, we have established a novel murine model ofE. africanusdisease providing critical insights into the host immune components required for eliminating the infection.Author summaryEmergomyces africanusis a thermally-dimorphic fungus that was recently described as the cause of disseminated infections in persons living with advanced HIV disease in South Africa, where emergomycosis is among the most frequently diagnosed dimorphic fungal diseases. Four additionalEmergomycesspecies have been described causing serious infections around the world, and the infection has a high case fatality rate. Despite the seriousness of this infection, the pathophysiology, immunology, diagnosis, and optimal management of emergomycosis remain largely unknown, hampered by the absence of an animal model. Here, we established a mouse model of pulmonary and disseminated emergomycosis using a clinical isolate ofE. africanuswith the aim of investigating the host immune response. We found that infection spread from the lungs to other organs within two weeks. We describe the effector immune responses in immune-competent mice aiding in controlled infection. We also highlight the immune-modulating components lacking in immunocompromised mice that promote susceptibilty to infection. Our study provides new insights into the components of the host immune response required for controllingEmergomycesspp. infection.

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“…Moreover, the failure to culture Pneumocystis spp. outside the mammalian host lung has hampered progress in studying the biology of these fungal pathogens as well as their mechanism of attachment to the host lung cells ( 10 ). Despite the efforts, several attempts to propagate Pneumocystis spp.…”

Section: Introductionmentioning

confidence: 99%

Precision-cut lung slices as an ex vivo model to study Pneumocystis murina survival and antimicrobial susceptibility

Munyonho,

Clark,

Lin

et al. 2024

mBio

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Pneumocystis pneumonia (PCP) is a serious fungal lung infection caused by Pneumocystis jirovecii in immunosuppressed individuals. The lack of viable in vitro/ex vivo PCP models has greatly hindered the progress in studying the biology of these fungi, the host/pathogen interactions, as well as antifungal susceptibility testing. In this study, we show the utility of precision-cut lung slices (PCLS) to support the survival of Pneumocystis murina in vitro . We cultured PCLS tissue derived from wild type and immunocompromised mice with a P. murina inoculum in submerged or air-liquid interface models for up to 14 days. We isolated total RNA from the cultured lung tissues at days 0, 3, 7, 10, and 14 and analyzed for the expression of host lung genes and P. murina genes ( Gsc1 for asci and Sp for trophs) by real-time quantitative PCR. When cultured in media alone, P. murina died gradually within a few days. However, when cultured on PCLS, both the troph and ascus forms survived throughout the incubation period of 2 weeks. Moreover, immunohistochemistry staining of P. murina inoculated PCLS sections using polyclonal anti- Pneumocystis sera and showed evidence of fungal aggregation and possible biofilm formation. Additionally, in vitro (PCLS) antibiotic susceptibility testing using commonly used antifungal drugs confirmed successful targeting of the troph and ascus forms by trimethoprim sulfamethoxazole and the ascus form by echinocandins. IMPORTANCE Our study reveals the potential of precision-cut lung slices as an ex vivo platform to study the growth/survival of Pneumocystis spp. that can facilitate the development of new anti-fungal drugs.

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Genetic Mouse Models of Pneumocystis Pneumonia

Cited by 3 publications

References 32 publications

The pathogenesis of experimental Emergomycosis in mice

The pathogenesis of experimental Emergomycosis in mice

The pathogenesis of experimental Emergomycosis in mice

Precision-cut lung slices as an ex vivo model to study Pneumocystis murina survival and antimicrobial susceptibility

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