Fluorescent staining for the rapid diagnosis of rare invasive fungal infections / 中华微生物学和免疫学杂志

Objective To evaluate the performance of an enhanced fluorescent staining for the rapid diagnosis of invasive mycosis, especially rare cases, considering the traditional culture method always leads to delays in clinical diagnosis for its time consuming. Methods Cases of invasive mycosis identified by fluorescent staining in our hospital from September, 2017 to September, 2018 were retrospectively analyzed. Three rare in-vasive infections were reported in this study. Clinical specimens were pretreated using standard procedures and then smeared on slides along with the enhanced fluorescent dye. Species of the pathogens were identified accord-ing to their morphology under fluorescent microscope. The traditional culture method was used as a standard method to identify the pathogenic species based on their colony morphology, followed by PCR and sequencing analysis for further confirmation. Results Three cases of invasive mycosis caused by rare pathogens of Talaro-myces marneffei, Mucorales and Prototheca were rapidly diagnosed with the fluorescent staining method. Sequen-cing results indicated the species were Talaromyces marneffei, Rhizopus arrhizus and Prototheca wickerhamii. Conclusions Fluorescent staining is a rapid, economic and direct method for the diagnosis of invasive mycosis. The morphology of fungi is clear and easy to identify after fluorescence staining, which could be used for indica-tive diagnosis of highly suspected invasive mycosis and serve as an important complement to the traditional cul-ture method, especially for the diagnosis of rare or uncultured fungal pathogens.

Overexpression of TLR2,TLR4 and MyD88 is associated with inflammation in C3H mice during Chlamydia respiratory infection / 中国免疫学杂志

Objective:To investigate the mechanism of excessive inflammation in the lung of C3H/HeN(C3H) mice following Chlamydia muridarum(Cm) airway infection.Methods:Chlamydial pneumonitis was induced in C3H and C57BL/6(C57) mice by intranasal inoculation with 1×103IFU (inclusion forming unites) of Cm strains.The expression of TLR2,TLR4 and MyD88 mRNA in the lung at different time point post-infection was measured by RT-PCR.Results:Cm infection induced Toll-like receptors expression in two strains of mice.The expression of TLR2 and TLR4 mRNA,especially TLR2 mRNA(P<0.001 or P<0.05),were significantly higher in highly susceptible C3H mice on day 7 and day 14 d post-infection compared with C57 mice.Further studies showed that the expression of MyD88 mRNA was also significantly higher in C3H mice on day 7 post-infection,and maintained high expression untill the day 14.Conclusion:Cm lung infection induced high level of TLR2,TLR4 and MyD88 mRNA expression in C3H mice,which may associate with excessive inflammation in C3H mice.

IL-17 A promotes differentiation and maturation of bone marrow-derived dendritic cells by cooperating with GM-CSF and LPS / 中国免疫学杂志

Objective:To investigate the effect of IL-17A on the differentiation and maturation of murine bone marrow-derived dendritic cells( BMDCs ) . Methods: Murine bone marrow cells were isolated and cultured in RPMI1640 complete medium in the presence of GM-CSF(20 ng/ml) for 8 days to induce differentiation of murine bone marrow cells to DC progenitors. Then these cells were treated with LPS(1 μg/ml) for 36 h which polarized immature DCs into mature DCs. Different concentrations of rmIL-17A(10 or 100 ng/ml) was added to the culture medium at different stages of BMDC differentiation and maturation. Co-stimulatory molecules expression on BMDC were analyzed by flow cytometry,and the culture supernatants were analyzed for IL-12p40 and IL-10 level by ELISA. Results:rmIL-17 could promote co-stimulatory molecules( CD40,CD80,CD86 and MHCⅡ) expression on BMDCs in a does-dependent manner,especially,the expression of CD40 and MHCⅡhad a significant increase in high concentration of rmIL-17A group;rmIL-17A was added while LPS induced maturation of BMDCs. CD40,CD80,CD86 and MHCⅡexpression on BMDC increased sharply in LPS plus rmIL-17A stimulation group,besides,CD86,MHCⅡ showed a higher level expression on BMDC with the increase of con-centration of rmIL-17A. Furthermore,secretion of IL-12p40 and IL-10 increased significantly in the group of DCs treated with LPS plus low concentration of rmIL-17 compared with the group without rmIL-17(P<0. 001). However,high concentration of rmIL-17A group showed significantly higher levels of IL-12p40(P<0. 001),but there was no difference in IL-10. Conclusion:IL-17A promotes the phe-notypic development of BMDC progenitors propagated in GM-CSF and cooperate with LPS to induce BMDC differentiation and matura-tion.

Infiltration and activation of neutrophils in lung tissues during Chlamydia muridarum respiratory tract infection / 中华微生物学和免疫学杂志

Objective To investigate the possible mechanism of aggregation and activation of neu-trophils(polymorphonuclear neutrophils,PMN)in mice with chlamydial pneumonitis. Methods C57BL/ 6 mice were inoculated intranasally with 3×103 inclusion-forming units(IFU)of Chlamydia muridarum(Cm) to induce the murine model of chlamydial pneumonitis. Samples of lung tissues collected at different time points after infection were stained by hematoxylin and eosin for histopathological assessment of inflammation. The levels of myelo-peroxidase(MPO)were detected for the evaluation of PMN aggregation. The mononu-clear cells were isolated from lung tissues. The inflammatory cells were counted with Giemsaˊs staining. CD11b+Gr1+ cell population and CD11b expression in lung mononuclear cells were analyzed by flow cytome-try. The expression of chemokines(MIP-2,LIX,KC and MCP-1)in lung tissues at mRNA level was meas-ured by RT-PCR. Results Chlamydial pneumonitis was induced in mice by intranasal inoculation of 3×103 IFU of Cm. Compared with the mice from control group,large amounts of inflammatory cells including PMN, monocytes and lymphocytes were induced in lung tissues of mice with Cm infection. PMN responded earlier than monocytes to the infection. The levels of MPO were significantly increased in mice with Cm infection and reached the highest level on the 7th day after infection. A decline in MPO levels was observed on the 14th day but the levels were still higher than those on day 0. The percentages and total numbers of CD11b+Gr1+ cells were significantly increased after Cm infection. Moreover,an increased expression of PMN CD11b was also detected by flow cytometry. The expression of chemokines(MIP-2,LIX,KC and MCP-1)was in-creased in lung tissues of mice after Cm infection. The results of the study indicated that Cm infection in-duced the expression of PMN chemoattractants,resulting in the recruitment of PMN. Conclusion The infil-tration and activation of PMN in lung tissues of mice were induced by Cm infection through increasing the ex-pression of chemokines. PMN played an important role in immune responses against Cm infection.