Table of Content

28 December 2019, Volume 14 Issue 6

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Original articles

Expression of proinflammatory cytokines in mouse dendritic cells stimulated by Sporothrix globosa

WANG Xiao-dong, Dlinuer·Taximaimaiti, Dlidaer·Dlixiati, Hadiliya·Hasimu, Paride Abliz

2019, 14(6):  321-325. 

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Objective To observe the characteristics of the secretion and expression of different inflammatory factors in mouse dendritic cells(DCs) stimulated by the Sporothrix globosa clinical strain in Xinjiang, and to predict the function of the inflammatory factor. Methods The experimental clinical strain was derived from infected lymphatic sporotrichosis, which was configured into different concentrations of fungal suspension (1×10 ⁴ cells/mL to 1×10⁷ cells/mL) to stimulate mouse DC (cell suspension concentration 1×10⁶ cells/mL), cell culture supernatants at concentrations of 6 hours, 24 hours, 48 hours, and 72 hours were collected, and IL-1β, IL-6, IL-4, TNF-α,and IFN-γ were detected by enzyme immunoassay. Results The stimulated DCs secreted IL-1β, IL-6 and TNF-α, and the secretions at different time points were:IL-1β (6 hours:21.26±3.03; 24 hours:24.04±4.25; 48 hours):24.90 ±4.31; 72 hours:27.29 ±6.09), IL-6 (6 hours:44.38 ±3.73; 24 hours:101.72 ±12.28; 48 hours:133.10 ±8.67; 72 hours:180.38 ±13.84), TNF-α (6 hours:860.36 ±20.64; 24 hours:356.03 ±11.46; 48 hours:457.43 ±17.39; 72 hours:1454.53 ±19.46), but no secretory expression was observed for IFN-γ and IL-4.The secretion levels of IL-1β and IL-6 increased gradually with time and dose, but the release of TNF-α was irregularly expressed. Conclusion DCs are involved in the innate immune response of Sporothrix globosa infection. The key inflammatory factors secreted are IL-1β, IL-6, and TNF-α, and the expression levels are TNF-α > IL-6 > IL-1β. The secretion levels of IL-1β and IL-6 are gradually increased in a time- and dose-dependent manner.

Effects of NETs on Trichophyton verrucosum infection

ZHANG Xiu-jun, GUO Yan-yang, ZHU Zhen-lai, XUE Xiao-wen, MA Cui-ling, WANG Gang, FU Meng

2019, 14(6):  326-331. 

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Objective A large number of neutrophils were found in the local lesions of patient with tinea barbae caused by Trichophyton verrucosum (T. verrucosum). Previous literatures reported that neutrophils participate in antifungal immunity through the formation of extracellular traps (NETs), but it was unclear whether T. verrucosum could induce NETs formation and whether NETs were involved in anti-T. verrucosum, so we explored whether neutrophils participate in the killing of T. verrucosum by forming NETs. Methods The patient's clinical data was collected. T. verrucosum was isolated and identified from patient's tissues. NETs in the tissues of patients and T. verrucosum-infected mice were observed by immunofluorescence. T. verrucosum was used to stimulate peripheral neutrophils in normal subjects, immunofluorescence was used to detect the formation of NETs. NETs were incubated with T. verrucosum, and then T. verrucosum DNA was extracted, Real-time quantitative PCR was used to quantitatively detect the killing effect of NETs on T. verrucosum. Results NETs formation could be seen in the tissues of T. verrucosum infected patients and mice. T. verrucosum co-incubation with neutrophils showed NETs formation. The survival rate of T. verrucosum after incubation with NETs was lower than that of the control group (P<0.01). Conclusions T. verrucosum can induce neutrophils to produce NETs, and NETs have a significant killing effect on T. verrucosum.

The study on the relationship between ERG11 gene and azole-resistant isolates of Candida tropicalis

WANG Ying, XIANG Ming-jie, LIU Jin-yan, YE Shu-lai, ZHOU xin

2019, 14(6):  332-337. 

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Objective To investigate the mutation and expression of ERG11 gene in clinical azole-resistant Candida tropicalis isolates. Methods The clinical Candida tropicalis isolates were collected and identified, then antifungal susceptibility tests to fluconazole, voriconazole and itraconazole were performed. ERG11 which was the gene coding antifungal drug target enzyme of cytochrome P450 lanosterol 14a-demethylase, was quantified by real-time RT-PCR and sequenced. The sequencing results were compared with the known standard sequence (M23673) in GenBank. Results There were 92 clinical Candida tropicalis isolates were obtained, of which 29 were resistant to at least one of the above three azole antifungals. Two missense mutations (S154F,Y132F) and five synonymous mutations were found in 40 clinical isolates, including 29 azole-resistant isolates and 11 azole-susceptible isolates. It is worth noting that the two missense mutations were existed in 24 of 29 azole-resistant isolates, however, no missense mutations were found in azole-susceptible isolates. The results of real-time PCR showed that the expression of ERG11 gene in azole-resistant clinical isolates was significantly higher than that in azole-susceptible isolates, and 16 isolates resistant to all three of the azole antifungals had a higher level of ERG11 gene expression than 13 isolates that were only resistant to one or two azole antifungals. Conclusions Azole resistance of clinical Candida tropicalis isolates is associated with the mutations and overexpression of ERG11 gene, and the resistance mechanism of Candida tropicalis to azole antifungals needs further study.

Analysis of infection distribution and drug resistance of invasive yeast in Beijing Hospital

GAO Zhen-xiang, TAO Feng-rong, PIAN Ya-ya, NIE Jing-jing, Guo Li-na, XU Ying-chun, HU Yun-jian

2019, 14(6):  338-341,373. 

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Objective To investigate the infection distribution and to analysis drug sensitivity of invasive yeast in Beijing Hospital from 2012-2013 and 2016-2017. Methods The strains were collected for the clinical detection of invasive yeast. The strain identification and MIC detection were carried out using Komajia chromogenic medium and API20C AUX and VITEK-2 Compact YST card. All strains were also identified by mass spectrometry. Results A total of 62 invasive yeasts were detected in our hospital from 2012-2013, including 36 strains of Candida albicans, accounting for 58.06%; 17 strains of Candida glabrata, accounting for 27.41%; 5 strains of Candida parapsilosis, accounting for 8.06%; 3 strains of Candida tropicalis, accounting for 4.83%; and 1 strain of other Candida, accounting for 1.60%. In 2016-2017, 43 strains of invasive yeast were detected in our hospital, including 21 strains of Candida albicans, accounting for 48.83%; 11 strains of Candida glabrata, accounting for 25.58%; 6 strains of Candida parapsilosis, accounting for 13.95%; 5 strains of Candida tropicalis, accounting for 11.63%. The fungal drug-sensitive results showed that 5-fluorocytosine had different degrees of resistance to Candida albicans, Candida glabrata and Candida tropicalis. Conclusion Candida albicans plays a leading role in invasive yeast infection in our hospital. Although the drug resistance of invasive yeast in our hospital has improved in recent years, further research on fungal drug sensitivity needs to be done.

Clinical analysis for 67 AIDS patients with cryptococcal meningitis

WANG Li-lin, LI Ai-xin, YANG Xue, Wang Wen, Wu Hao

2019, 14(6):  342-345. 

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Objective To investigate the clinical characteristics of AIDS patients with cryptococcal meningitis. Methods We collected and analyzed the clinical data including clinical presentation, laboratory examination, therapeutic regimen and outcome on AIDS patients with cryptococcal meningitis from Beijing You'an Hospital during 1 January 2010 and 31 December 2016. Results There were 67 AIDS patients with cryptococcal meningitis in this study, including 60 male and 7 female. The most common clinical symptoms were headache (74.6%), fever (47.8%), nausea and vomiting (40.3%). Increased intracranial pressure was found in 78.9% patients, the mean pressure was 216.6 (±100.3 mm H₂O), and the baseline CD4⁺ T cell was 53.7 (±79.9)/mm3. The mortality rate was 29.9% (20/67). Conclusion Cryptococcal meningitis with high mortality rate was severe complication of AIDS. Early diagnosis and standard treatment was key to reduce injury of Cryptococcus on brain, and to save lives.

Efficacy and safety of amphotericin B in 124 AIDS-associated cryptococcal meningitis patients

HE Yuan-hong, TAN qing, YANG Tong-tong, HE Sheng-hua, WANG Yin, ZHOU Rui-feng, CAI Lin

2019, 14(6):  346-350. 

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Objective To observe the efficacy and safety of amphotericin B combined with 5-FC in the treatment of AIDS-associated cryptococcal meningitis. Methods We prospectively enrolled 124 AIDS adults with cryptococcal meningitisand received treatment with amphotericin B + 5FC over 2 weeks, who presented to the public health clinical center of Chengdu from January 2013 to July 2018. Results The mean age of 124 patients was (38.85 + 11.22) years old. Mean course of disease was (22.40 + 17.75) days. The most common clinical symptoms were headache (89.52%), fever (47.58), nausea and vomiting (38.71%).The median CD4⁺T lymphocyte count and plasma HIV RNA of 107 ART-naïve patients were 18 cells/μl and 1.38×10⁵ copies/ml. 62.9% patients had a cerebrospinal fluid pressure higher than 250 mmH₂O. AmB infusion treatment time was (4.97 + 1.13) days, and the average treatment time was (33.42 + 13.53) day. The number of cerebrospinal fluid cells and protein content decreased at 2 weeks after treatment compared with that before treatment, while glucose increased. It showed a significant difference (P<0.05). The negative conversion rate of CSF culture was 87.10% at 2 weeks and 96.77% at 4 weeks.2.42% (3/124) patients discontinued AmB due to severe adverse drug reactions. Mortalities were 4.03% at 4 weeks and 12.90% at 12 weeks. Conclusion A short-term use of AmB combined with 5-FC in the treatment of AIDS-associated cryptococcal meningitis was effective and safety. Blood test, liver function, kidney function and electrolyte should be closely monitored during the treatment.

Analysis on pathogenic characteristics of dermatophyte infection and in vitro drug susceptibility of 85 strains of dermatophyte

LIU Jie, LIU Xiao-yun, YU Bo, HU Xiao-ping

2019, 14(6):  351-356. 

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Objective To analyze the pathogenic characteristics of dermatophyte infection in the out-patient department of a teaching hospital in Shenzhen area and to investigate in vitro activity of 7 antifungal agents against 85 strains of dermatophyte. Methods Skin scales (or nail scraps, hairs) were collected from suspected patients with dermatophyte infection for KOH direct microscopic examination and culture. The species of dermatophyte were identified by sequencing the ribosomal DNA internal transcribed spacer(ITS). The antifungal susceptibility testing was performed following Clinical and Laboratory Standards Institute (CLSI) M38-A2 reference broth microdilution methods. Seven antifungal agents were evaluated against 85 strains of dermatophytes. Results Among 161 suspected patients with dermatophyte infection, the positive rate of KOH direct microscopic examination of dermatophyte was 67.7%, and the positive rate of fungal culture of dermatophyte was 52.8%. A total of 85 isolates of positive fungal culture were identified as 68 strains of Trichophyton rubrum, 7 strains of Microsporum canis, 3 strains of Microsporum gypseum, 5 strains of Trichophyton interdigitale, 1 strain of Trichophyton violaceum and 1 strain of Trichophyton tonsurans. In vitro drug susceptibility test results revealed that terbinafine (GM MIC,0.032μg·mL^-1, MIC range, 0.001~0.125μg·mL^-1) had the strongest in vitro antifungal activity against 6 dermatophytes. And voriconazole (GM MIC,0.041μg·mL^-1, MIC range, 0.032~0.125μg·mL^-1)had the lowest MIC among the azole drugs. Conclusions Trichophyton rubrum was the most common pathogen of dermatophyte infection in our hospital. The standardized antifungal susceptibility testing confirmed that terbinafine and voriconazole had strong in vitro antifungal activity against isolated dermatophyte and were suitable for the treatment of dermatophyte infection in this area.

Analysis the pathogen distribution and drug susceptibility of yeast bloodstream infection in Peking Union Medical College Hospital from 2014 to 2018

LIU Wen-jing, SUN Hong-li, ZHANG Xiao-jiang, YANG Wen-hang, XIAO Meng, WANG Yao, YANG Qi-wen, DOU Hong-tao, WANG He, ZHAO Ying, GUO Li-na, LIU Ya-li, ZHANG Li, ZHU Ren-yuan, XU Ying-chun

2019, 14(6):  357-361. 

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Objective To analysis the pathogen distribution and drug susceptibility of yeast blood stream infection, to provide the basis for clinical diagnosis and treatment. Methods Clinical data of Candida blood stream infection was retrospectively analyzed in in Peking Union Medical College Hospital from January 2014 to December 2018. Results A total of 206 non-repetitive yeast strains were isolated from blood culture specimens. The positive isolation rate of yeast from Bactec Mycosis IC/F bottle, aerobic and anaerobic blood culture bottle were 1.32% (79/6003), 0.43%(371/85855)and 0.06% (47/77168) respectively. The top three Candida species were Candida albicans (72, 34.95%), Candida tropicalis (42, 20.39%) and Candida glabrata (39, 18.93%). The uncommon Candida species included Candida guilliermondii, Pichia pastoris, Candida lusitaniae, Saccharomyces cerevisiae, Candida lambica, Candida nivalis, Rhodotorula and Candida Haemulonii. In our hospital,candidemia usually developed in patients aged >60 years (42.23%) and was mostly in men (115, 55.83%). Infected patients were mainly from the emergency department (41, 19.90%), surgery intensive care unit and internal intensive care unit (both were 36, 17.48%). The positive alarm times from long to short was:Candida glabrata, Candida parapsilosis, Candida albicans and Candida tropicalis. The susceptibility rates of Candida albicans, Candida glabrata, Candida parapsilosis and Candida tropicalis to fluconazole were 98.61%、97.44%、89.66% and 71.43%, respectively. Conclusion Candida albicans was the main blood stream infection species in our hospital. Most Candida strains had high sensitivity to antifungal drugs, but the resistance rate of Candida tropicalis was relatively high.