Animal Laboratory Study On The Pathogenecity Of Geotrichum Silvicola Isolated From Blood

Objective: Geotrichum silvicola, a new strain in Geotrichum, was firstly isolated from Brazil fly and Indian oak silkworm. In 2002, we isolated a strain from a child's kerion-like lesions, which firstly confirmed that Geotrichum silvicola can infect human. Then it was isolated from blood of the patient suffered from toxic epidermal necrolysis in our department after three years, which demonstrated that this strain can cause systemic infection. Geotrichum silvicola was studied about morphology, molecular biology and physiology. Two strains show a little discrepancy in the colonial morphology and gene order.In this research,mice in different immune states were inoculated Geotrichum silvicola isolated from the skin lesion and blood through intradermal, intraperitoneal and intravenous pathways, respectively. By observing the death rate and mean survival time of experimented mice during 28 days , the retro-culture rate and fungi residual burden of heart, liver, spleen, lung, kidney, we want to investigate the influence of immune state and infected pathway on systemic infection with Geotrichum silvicola isolated from blood, and to compare the pathogenecity of two strains. Methods:1 Establition of animal model suffered from systemic infection of Geotrichum silvicola The preserved fungi were activated, then injected into the vena caudalis of mice. We dissected the infected mice and obtained the strain of recovered virulence, then made up conidial suspension and inoculated different immune mice through intradermal, intraperitoneal and intravenous pathways, respectively.2 Observation of mice's survival states and death conditions We observed the survival states and death conditions of mice during four weeks, and calculated the death rate and mean survival time.3 The change of organs Dissected the sacrificed mice, took the heart, liver, spleen, lung and kidney under sterile conditions, and observed their changes of size, color, and so on.4 Fungal retro-culture The infected tissues were grinded and subcultivated onto the media of SDA. After cultured under 37℃for two weeks, we counted the positive numbers. The colonies were subcultivated onto the media of plate culture and microculture. We observed the colonial morphology and shape of hyphae, spores.5 Observation of fungi residual burden of organs The heart, liver, spleen, lung and kidney of mice that inoculated Geotrichum silvicola isolated from blood and the kidney of mice that inoculated Geotrichum silvicola isolated from the skin lesion were homogenated, then cultured on platiculture. The fungi residual burden of every organ was calculated.6 Histopathological Examination The organs of mice that inoculated Geotrichum silvicola isolated from blood were fixed, dehydrated, cleared, embedded, sliced, stained with HE and PAS respectively, then observed under light microscope.Results:1 The survival state of mice The life habits had no obvious change after inoculated with Geotrichum silvicola. The mice that inoculated fungi through intradermal pathway had a nodue on the back. The immunsuppressed mice that inoculated intraperitoneally and intravenously reduced food-intake obviously, and emaciated gradually.2 The death condition of mice The immunsuppressed groups that inoculated intraperitoneally and intravenously had dead mice, but other groups had no death. Comparing the immunsuppressed groups that inoculated intraperitoneally and intravenously with others, the death rate and mean survival time of mice had significant difference. Among other groups, they had no significant difference. In same immune state and inoculated pathway, two strains also had no significant difference.3 The change of organs Mauve intumescent liver and pink lung of the mice can be seen after dissection. The organs of mice inoculated intraperitoneally have some microabscess.4 Fungal retro-culture When visceras were cultured for three days, the fungi began to grow. The colonial morphology and shape of hyphae, spores were identical with that of inoculated strains. The fungal retro-culture rate of the immunsuppressed group that inoculated intradermally was obviously lower than the immunsuppressed groups that inoculated intraperitoneally and intravenously, while the latter groups had no significant difference. In the same group, two strains had no significant difference. In flat plate, the colony presented white power at first, and then became yeast-like type. On its periphery, there were radiate fuzz. In the microculture, there were a lot of branched septate hyphae, rectangular, tubby arthroconidia and round spores.5 Fungal residual burden of organs The kidney fungal residual burden of every group that inoculated Geotrichum silvicola isolated from blood was obviously different. The kidney fungal residual burden of mice that inoculated Geotrichum silvicola isolated from blood was similar to that of mice that inoculated Geotrichum silvicola isolated from the skin lesion. The fungal residual burden of spleen was more than other organs.6 Histopathological Examination HE dyeing showed cell degeneration and necrosis, acute or chronic inflammation, reactive hyperplasia of histocytes. PAS dyeing displayed mauve hypha, arthroconidia, round or oval spores in tela.Conclusion:1 Mice could develop systemic infection when they were injected Geotrichum silvicola through intradermal, intraperitoneal and intravenous pathways, respectively. 2 Geotrichum silvicola isolated from blood was a opportunistic fungus, its virulence was correlative with immune state.3 Infected degree of mice was relevant to inoculated pathway. The infected degree of mice injected intravenously was more serious than others.4 Spleen was the most common involved organs.5 Geotrichum silvicola from different origination showed the same pathogenicity.