Murine Model of Disseminated Fusariosis: Evaluation of the Fungal Burden by Traditional CFU and Quantitative PCR

Abstract

Systemic disease is the most severe clinical form of fusariosis, and the treatment involves a challenge due to the refractory response to antifungals. Treatment for murine Fusarium solani infection has been described in models that employ CFU quantitation in organs as a parameter of therapeutic efficacy. However, CFU counts do not precisely reproduce the amount of cells for filamentous fungi such as F. solani. In this study, we developed a murine model of disseminated fusariosis and compared the fungal burden with two methods: CFU and quantitative PCR. ICR and BALB/c mice received an intravenous injection of 1 × 10(7) conidia of F. solani per mouse. On days 2, 5, 7, and 9, mice from each mice strain were killed. The spleen and kidneys of each animal were removed and evaluated by qPCR and CFU determinations. Results from CFU assay indicated that the spleen and kidneys had almost the same fungal burden in both BALB/c and ICR mice during the days of the evaluation. In the qPCR assay, the spleen and kidney of each mouse strain had increased fungal burden in each determination throughout the entire experiment. The fungal load determined by the qPCR assay was significantly greater than that determined from CFU measurements of tissue. qPCR could be considered as a tool for quantitative evaluation of fungal burden in experimental disseminated F. solani infection.

Consensus guidelines for the treatment of invasive mould infections in haematological malignancy and haemopoietic stem cell transplantation, 2014

“…Imaging techniques are non‐specific. Blood cultures and tissue biopsy are the most specific diagnostic tests, and pan‐fungal polymerase chain reaction, galactomannan and β(1,3)‐D‐glucan assays may all be positive …”

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“…Thus, although fungal burden is a useful parameter for assessing fungal dissemination in the host, it fails to provide detailed information on the impact of filamentous pathogens on the infected organs. It was reported previously that during F. solani or A. fumigatus infection, quantitative culture led to underestimation of absolute fungal burden as compared to non-culture-based methods such as quantitative PCR or determination of galactomannan levels by enzyme immunoassays (EIAs) [32], [33], [34], [35]. In an inhalational rat model of invasive pulmonary aspergillosis (IPA), both real-time nucleic acid sequence-based amplification (NASBA) and qPCR showed a progressive increase of fungal biomass in lung tissue, whereas CFU counts were stable over time [36].…”

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“…Symptoms of immunosuppressed mice are also remarkably similar to those reported in humans. Fusarium species, such as F. oxysporum, can disseminate and persist in the organs of immunocompetent animals, and these latent infections can lead to lethal systemic fusariosis if the host is later subjected to immunosuppressive treatment [106][107][108][109][110][111][112][113][114][115].…”