A general method for longitudinal CT imaging and quantification of lung pathologies in mouse models of respiratory fungal infections, including aspergillosis and cryptococcosis, using low-dose high-resolution CT is described.
Aspergillus fumigatus and Cryptococcus neoformans infections represent significant and life-threatening fungal hazards for immunocompromised individuals. Selleck 2′,3′-cGAMP Elevated mortality rates are associated with acute invasive pulmonary aspergillosis (IPA) and meningeal cryptococcosis, which represent the most severe presentations in patients, even with current treatment options. The current state of understanding concerning these fungal infections is far from complete, prompting a vital need for additional research, not only within clinical applications but also under tightly regulated preclinical experimental frameworks. This is crucial for enhancing our comprehension of their virulence, host-pathogen relationships, infection development, and suitable treatment options. Animal models in preclinical studies are potent instruments for deeper understanding of certain requirements. Nonetheless, the measurement of disease severity and fungal load in murine models of infection is often restricted by techniques that are less sensitive, single-time, invasive, and prone to variability, such as colony-forming unit counting. Bioluminescence imaging (BLI), performed in vivo, can alleviate these problems. The fungal burden's dynamic, visual, and quantitative longitudinal evolution, tracked by the noninvasive tool BLI, shows its presence from infection onset, possible spread to various organs, and throughout the entire disease process in individual animals. This paper presents an entire experimental procedure, from initiating infection in mice to obtaining and quantifying BLI data, allowing for non-invasive, longitudinal tracking of fungal load and spread throughout infection progression. It is an important tool for preclinical studies of IPA and cryptococcosis pathophysiology and treatment strategies.
The development of novel therapeutic approaches for fungal infections has benefited greatly from the use of animal models, which provide crucial insight into the disease's pathogenesis. A low incidence rate does not diminish the fact that mucormycosis frequently proves fatal or debilitating. Mucormycoses arise from diverse fungal species, each potentially entering the body through unique infection pathways, while affecting patients with varying underlying diseases and risk profiles. Therefore, animal models suitable for clinical research utilize distinct methods of immunosuppression and infection routes. It elaborates upon the intranasal application methods for the purpose of creating pulmonary infections, in addition. In summary, the last part focuses on clinical variables applicable for creating scoring systems and identifying humane end points in mouse trials.
Pneumonia is a frequent manifestation of Pneumocystis jirovecii infection in individuals with compromised immunity. In the context of both drug susceptibility testing and understanding host/pathogen interactions, Pneumocystis spp. presents a significant and multifaceted challenge. Their in vitro existence is not sustainable. The inability to maintain continuous culture of the organism imposes significant constraints on the process of identifying novel drug targets. Because of this constraint, mouse models of Pneumocystis pneumonia have demonstrated exceptional value to researchers. Selleck 2′,3′-cGAMP An overview of selected methods used in mouse infection models is offered in this chapter, detailing in vivo Pneumocystis murina propagation, transmission routes, available genetic mouse models, a P. murina life form-specific model, a mouse model of PCP immune reconstitution inflammatory syndrome (IRIS), and the pertinent experimental factors.
In the global context, dematiaceous fungal infections, specifically phaeohyphomycosis, are emerging, presenting diverse clinical pictures. Mimicking human dematiaceous fungal infections, the condition of phaeohyphomycosis can be usefully studied using the mouse model as a research tool. Our laboratory's creation of a mouse model of subcutaneous phaeohyphomycosis displayed noteworthy phenotypic differences between Card9 knockout and wild-type mice. This finding mirrors the enhanced susceptibility to infection seen in CARD9-deficient human populations. The construction of a mouse model exhibiting subcutaneous phaeohyphomycosis, and the subsequent experiments, are presented here. We anticipate that this chapter will prove advantageous to the study of phaeohyphomycosis, thereby fostering the development of novel diagnostic and therapeutic methodologies.
The fungal infection coccidioidomycosis, resulting from the dimorphic fungi Coccidioides posadasii and Coccidioides immitis, is a prevalent disease in the southwestern United States, Mexico, and parts of Central and South America. The mouse is prominently featured in studies concerning disease pathology and immunology as a model organism. Research on the adaptive immune responses in mice necessary for controlling coccidioidomycosis is hampered by their extreme susceptibility to Coccidioides spp. To model asymptomatic infection with controlled, chronic granulomas, and a slowly progressive, ultimately fatal infection mirroring the human disease's kinetics, we detail the process of infecting mice here.
For the purpose of understanding the interplay between a host and a fungus in fungal diseases, experimental rodent models provide a helpful tool. Due to spontaneous cures in animal models, a relevant model for the long-term, chronic disease manifestation in humans, specifically for Fonsecaea sp., a causative agent of chromoblastomycosis, is currently absent. This chapter describes an experimental rat and mouse model using a subcutaneous approach. A critical analysis of the acute and chronic lesions, mimicking human disease, included fungal burden and the examination of lymphocytes.
Commensal organisms, numbering in the trillions, constitute a significant part of the human gastrointestinal (GI) tract's microbial ecosystem. Some of these microbial agents are capable of evolving into pathogenic forms upon modifications to the microenvironment and/or host physiology. Among the organisms inhabiting the gastrointestinal tract is Candida albicans, which typically acts as a harmless commensal, but can also become the cause of severe infection in certain circumstances. Factors like antibiotic use, neutropenia, and abdominal surgery may increase susceptibility to gastrointestinal Candida albicans infections. The transformation of commensal organisms into pathogenic agents warrants significant investigation and research. Utilizing mouse models of fungal gastrointestinal colonization provides a critical platform for exploring the underlying processes of Candida albicans's transition from a benign commensal to a harmful pathogen. This chapter explores a groundbreaking approach to the consistent, long-term colonization of the murine gastrointestinal system by the Candida albicans fungus.
The central nervous system (CNS), including the brain, can be affected by invasive fungal infections, potentially causing fatal meningitis in immunocompromised individuals. Thanks to recent technological advancements, the scope of brain research has broadened from analyses of the brain's inner substance to a deeper understanding of the immune systems in the meninges, the protective covering of the brain and spinal column. Researchers are now able to visualize the structure of the meninges and the cellular components responsible for the inflammatory response within the meninges, using advanced microscopy techniques. The techniques for preparing meningeal tissue mounts for confocal microscopy are illustrated in this chapter.
CD4 T-cells are indispensable for the long-term control and eradication of various fungal infections in humans, including those induced by Cryptococcus species. A comprehensive understanding of the protective mechanisms of T-cell immunity against fungal infections is essential for developing a mechanistic insight into the complex nature of the disease. A protocol for analyzing fungal-specific CD4 T-cell responses in vivo is presented, employing the technique of adoptive transfer with fungal-specific T-cell receptor (TCR) transgenic CD4 T-cells. This protocol, while utilizing a TCR transgenic model responsive to Cryptococcus neoformans peptides, holds adaptable potential for other fungal infection research settings.
The opportunistic fungal pathogen, Cryptococcus neoformans, is a frequent cause of fatal meningoencephalitis in immunocompromised patients. The intracellular fungus evades the host's immune system, establishing a latent infection (latent cryptococcal infection, LCNI), and cryptococcal disease manifests when this latent state is reactivated due to a compromised host immune response. Unraveling the pathophysiology of LCNI is challenging due to the absence of suitable mouse models. This document outlines the established methodologies for LCNI and its subsequent reactivation.
The fungal pathogen, Cryptococcus neoformans species complex, causes cryptococcal meningoencephalitis (CM), which can have a high mortality rate or lead to debilitating neurological sequelae in those who survive, often due to excessive inflammation in the central nervous system (CNS). This is particularly true for those who develop immune reconstitution inflammatory syndrome (IRIS) or post-infectious immune response syndrome (PIIRS). Selleck 2′,3′-cGAMP The capacity of human studies to establish a definitive cause-and-effect relationship for a particular pathogenic immune pathway during central nervous system (CNS) events is hampered; however, the use of mouse models permits the investigation of potential mechanistic links within the CNS's immune system. Specifically, these models assist in the differentiation of pathways primarily associated with immunopathology from those of paramount importance in fungal eradication. The methods for inducing a robust, physiologically relevant murine model of *C. neoformans* CNS infection, outlined in this protocol, accurately reproduce key aspects of human cryptococcal disease immunopathology, enabling subsequent detailed immunological investigation. Investigations leveraging gene knockout mice, antibody blockade, cellular adoptive transfer, and high-throughput methods, such as single-cell RNA sequencing, within this model will unveil intricate cellular and molecular processes pivotal to the pathogenesis of cryptococcal central nervous system diseases, facilitating the development of more effective therapeutic interventions.