Bacterial biofilms in patients with chronic rhinosinusitis

The possible presence of biofilms was examined in mucosal specimens of 15 patients, undergoing functional endoscopic sinus surgery or a modified Caldwell-Luc approach for chronic rhinosinusitis (CRS). Biofilms were found in 7 of the 15 patients, positive cultures being obtained in most samples, which supports the role of biofilms as an important factor in the pathogenesis of CRS.     

Pathophysiological classification of chronic rhinosinusitis

Background

Recent consensus statements demonstrate the breadth of the chronic rhinosinusitis (CRS) differential diagnosis. However, the classification and mechanisms of different CRS phenotypes remains problematic.

Method

Statistical patterns of subjective and objective findings were assessed by retrospective chart review.

Results

CRS patients were readily divided into those with (50/99) and without (49/99) polyposis. Aspirin sensitivity was limited to 17/50 polyp subjects. They had peripheral blood eosinophilia and small airways obstruction. Allergy skin tests were positive in 71% of the remaining polyp subjects. IgE was<10 IU/ml in 8/38 polyp and 20/45 nonpolyp subjects (p = 0.015, Fisher''s Exact test). CT scans of the CRS without polyp group showed sinus mucosal thickening (probable glandular hypertrophy) in 28/49, and nasal osteomeatal disease in 21/49. Immunoglobulin isotype deficiencies were more prevalent in nonpolyp than polyp subjects (p < 0.05).

Conclusion

CRS subjects were retrospectively classified in to 4 categories using the algorithm of (1) polyp vs. nonpolyp disease, (2) aspirin sensitivity in polyposis, and (3) sinus mucosal thickening vs. nasal osteomeatal disease (CT scan extent of disease) for nonpolypoid subjects. We propose that the pathogenic mechanisms responsible for polyposis, aspirin sensitivity, humoral immunodeficiency, glandular hypertrophy, eosinophilia and atopy are primary mechanisms underlying these CRS phenotypes. The influence of microbial disease and other factors remain to be examined in this framework. We predict that future clinical studies and treatment decisions will be more logical when these interactive disease mechanisms are used to stratify CRS patients.     

Hydrophobic and adhesive capacity of Candida strains isolated from patients with fungal infection

The aim of this study was to evaluate hydrophobicity of fungal strains of Candida sp. isolated from clinical materials of patients with fungal infections. Two hundred and eighty one strains of C. albicans and 29 strains belonging to the other species of Candida were tested in salt aggregation test (SAT). Strong hydrophobicity (autoaggregation in the test) was found in 29.4% of tested strains. The majority of them was isolated from vagina. In 20 randomly selected strains hydrophobicity was measured not only by salt aggregation test but also by hydrophobic interaction chromatography (MIC). The ability to attach to buccal epithelial cells in vitro was compared between 50 strongly hydrophobic strains and 30 with low hydrophobicity. The former strains attached significantly stronger (p less than 0.001).     

Identification and inoculation of fungal strains from Cedrus deodara rhizosphere involve in growth and alleviation of high nitrogen stress

Cedrus deodara is economically and ethnobotanically an important forest tree and is shown to be at decline in Northern areas of Pakistan in recent years mainly due to high concentration of Nitrogen in forests. Ectomycorrhizal (ECM) association forming fungi enables the forest trees to develop optimally by absorbing water from the rhizosphere through their absorptive hyphae and by making available the nutrients by mobilization of N and P from the organic substrates. This study was conducted to identify the ECM strains from C. deodara rhizosphere and to analyse the impact of high N load on the C. deodara seedlings to establish N critical load value for coniferous forests of Pakistan. Six new fungal strains were identified from the rhizosphere of C. deodara and were registered at GenBank (NCBI) as Emmia latemarginata strain ACE1, Aspergillus terreus strain ACE2, Purpureocillium lilacinum strain ACE3, Talaromyces pinophilus strain ACE4, A. fumigatus strain ACE5 and T. pinophilus strain ACE6 with accession numbers MH145426, MH145427, MH145428, MH145429, MH145430 and MH547115. Four out of six isolated strains were inoculated with seedlings of C. deodara singly and in consortium (CN) in combination with nitrogen load of 0 (C), 25 (T1), 50 (T2), 100 kg N ha⁻¹ yr⁻¹ (T3). Agronomic, physiological and gene expression studies for ExpansinA4 (EXPA4) and Cystatins (Cys) were made to analyse the impact of fungal strains in relation to high N stress. This study suggests a positive impact of T1 (25 kg N ha⁻¹ yr⁻¹) Nitrogen load and a negative impact of T3 (100 kg N ha⁻¹ yr⁻¹) on growth parameters and expression patterns of EXPA4 and Cys genes. Peroxidase (POX) activity decreased in the order ACE5 > ACE2 > C > ACE3 > ACE1 > CN. However, the results of Superoxide dismutase (SOD) showed decreasing trend in the order ACE5 > C > CN > ACE1 > ACE2 > ACE3. Strain ACE3 was shown to have a positive impact on the seedlings in terms of growth, physiology and expression of genes. Present study suggests that newly identified fungal strains showing positive impact on the growth and physiology of C. deodara could be used for the propagation of this economically important plant in Pakistan after pathogenicity test.     

慢性重型乙肝继发侵袭性真菌感染的研究

目的明确侵袭性真菌感染(invasive fungal infections,IFI)在慢性重型乙肝患者中发病情况及主要病因。方法依据IFI和乙肝诊断标准,筛选上海长征医院感染科慢性重症乙肝患者,60例IFI者为病例组,66例未发生IFI为对照组,进行回顾性分析。结果 IFI患病率47.62%,病死率40%。乙肝病毒DNA水平是最主要的危险因素,当DNA高于3.16×103copies/mL,IFI发病可能性增大。结论慢重肝患者IFI的患病率和病死率超出临床预期,降低DNA拷贝可延缓乙肝进展,在预防及治疗IFI中亦具积极意义。     

白细胞介素-33在慢性鼻-鼻窦炎伴鼻息肉患者黏膜组织中的表达及意义

目的观察慢性鼻-鼻窦炎伴鼻息肉(CRSwNP)患者鼻黏膜组织中白细胞介素-33(IL-33)的表达与嗜酸性粒细胞(Eos)的浸润情况,探讨IL-33与Eos在其发病中作用及相关性。方法采用免疫组织化学LSAB法分别检测CRSwNP患者(观察组)32例和单纯鼻中隔偏曲患者(对照组)24例的鼻黏膜组织中IL-33的表达情况;苏木精-伊红染色下观察炎性细胞浸润程度并对Eos计数;全血细胞分析法测定两组患者血液中嗜酸性粒细胞含量。结果 (1)CRSwNP组织中上皮层及间质内Eos、中心粒细胞为主的炎性细胞及部分腺体内有IL-33的表达;(2)免疫组织化学检测显示IL-33阳性细胞数、苏木精-伊红染色Eos计数均高于对照组,差异具有统计学意义(P0.05)。Eos计数与IL-33阳性细胞数在CRSwNP中存在正相关性(P0.05);(3)血液中Eos含量无统计学意义。结论 IL-33是一种新型的细胞因子,在CRSwNP患者体内高表达,可能与嗜酸性粒细胞浸润互相促进,在CRSwNP发生发展中具有重要的作用。     

应用糖皮质激素进行鼻腔冲洗对慢性鼻窦炎患者鼻腔菌群多样性和生物膜的影响

目的 分析应用糖皮质激素进行鼻腔冲洗对慢性鼻窦炎(CRS)患者鼻腔菌群多样性及生物膜的影响。 方法 将2018年1月至2018年6月于我院治疗的40例慢性鼻窦炎患者随机分成两组,每组各20例,一组给予0.9%氯化钠注射液进行鼻腔冲洗(NS组),另一组给予0.9%氯化钠注射液及布地奈德进行鼻腔冲洗(NS+BUD组),对两组患者治疗前后的菌群结构差异、多样性及细菌生物膜生成情况进行比较。 结果 治疗前两组患者菌群多样性指标(Chao1指数、Ace指数、Simpson指数及Shannon指数)比较,差异均无统计学意义(均P>0.05),治疗后两组患者的Chao1指数、Ace指数及Shannon指数均明显下降(均P结论 应用糖皮质激素对慢性鼻窦炎患者进行鼻腔冲洗可降低患者鼻腔微生物多样性,减少菌群的异常增殖,降低主要致病菌的生物膜表达,提高临床疗效。     

Differences in proteolytic activity and gene profiles of fungal strains isolated from the total parenteral nutrition patients

Fungal infections constitute a serious clinical problem in the group of patients receiving total parenteral nutrition. The majority of species isolated from infections of the total parenteral nutrition patients belong to Candida genus. The most important factors of Candida spp. virulence are the phenomenon of “phenotypic switching,” adhesins, dimorphism of fungal cells and the secretion of hydrolytic enzymes such as proteinases and lipases, including aspartyl proteinases. We determined the proteolytic activity of yeast-like fungal strains cultured from the clinical materials of patients receiving total parenteral nutrition and detected genes encoding aspartyl proteinases in predominant species Candida glabrata—YPS2, YPS4, and YPS6, and Candida albicans—SAP1–3, SAP4, SAP5, and SAP6. C. albicans released proteinases on the various activity levels. All C. glabrata strains obtained from the clinical materials of examined and control groups exhibited secretion of the proteinases. All 13 isolates of C. albicans possessed genes SAP1–3. Gene SAP4 was detected in genome of 11 C. albicans strains, SAP5 in 6, and SAP6 in 11. Twenty-six among 31 of C. glabrata isolates contained YPS2 gene, 21 the YPS4 gene, and 28 the YPS6 gene. We observed that clinical isolates of C. albicans and C. glabrata differed in SAPs and YPSs gene profiles, respectively, and displayed differentiated proteolytic activity. We suppose that different sets of aspartyl proteinases genes as well as various proteinase-activity levels would have the influence on strains virulence.     

Diagnosis of invasive fungal disease in hospitalized patients with chronic obstructive pulmonary disease

Background

The role of culture-independent techniques (galactomannan, (1-3)-β-d-glucan) in the early diagnosis of invasive fungal diseases (IFD) is well assessed in hematological patients, but there are no clear conclusions in patients with chronic obstructive pulmonary disease (COPD).

Biomodification and biodeterioration of carbon coatings by fungal strains

Over the past years, surface protection by hard carbon coatings has been increasingly applied in many fields. This study aimed to characterize changes in the properties of carbon coatings brought about by the growth of fungi (the first report about these processes was published by Kaczorowska et al., Nanodiam, PWN, 2006, 99–116). Alterations in the structure of diamond-like carbon (DLC) and nanocrystalline diamond (NCD) coatings, examined by Raman spectroscopy, XRD, XPS, and FTIR, were found to be caused by the processes of oxidation (incorporation of oxygen atoms) and reduction (hydrogenation) occurring within graphite carbon and amorphous carbon domains (components of the surface layer of the tested coatings along with the diamond domain). Determination of the activity of selected enzymes synthesized by the studied strains of filamentous fungi revealed that the observed biomodification and biodeterioration processes involved, among others, laccase, Mn-dependent peroxidase, two catechol dioxygenases, and esterases. The biosynthesis of these enzymes by fungi was enhanced when graphite was added to their culture media. Because of the high risk of fungal infections, the quality of carbon coatings should be routinely controlled by standard microbiological tests employing suitable strains of filamentous fungi (e.g., Aspergillus niger) and yeasts synthesizing enzymes that attack carbon materials.     

Novel strategies for inhibition of bacterial biofilm in chronic rhinosinusitis

An important role has been recently reported for bacterial biofilm in the pathophysiology of chronic diseases, such as chronic rhinosinusitis (CRS). CRS, affecting sinonasal mucosa, is a persistent inflammatory condition with a high prevalence around the world. Although the exact pathological mechanism of this disease has not been elicited yet, biofilm formation is known to lead to a more significant symptom burden and major objective clinical indicators. The high prevalence of multidrug-resistant bacteria has severely restricted the application of antibiotics in recent years. Furthermore, systemic antibiotic therapy, on top of its insufficient concentration to eradicate bacteria in the sinonasal biofilm, often causes toxicity, antibiotic resistance, and an effect on the natural microbiota, in patients. Thus, coming up with alternative therapeutic options instead of systemic antibiotic therapy is emphasized in the treatment of bacterial biofilm in CRS patients. The use of topical antibiotic therapy and antibiotic eluting sinus stents that induce higher antibiotic concentration, and decrease side effects could be helpful. Besides, recent research recognized that various natural products, nitric oxide, and bacteriophage therapy, in addition to the hindered biofilm formation, could degrade the established bacterial biofilm. However, despite these improvements, new antibacterial agents and CRS biofilm interactions are complicated and need extensive research. Finally, most studies were performed in vitro, and more preclinical animal models and human studies are required to confirm the collected data. The present review is specifically discussing potential therapeutic strategies for the treatment of bacterial biofilm in CRS patients.     

7株野生虫草菌的鉴定及其菌丝体醇提取物对HepG2细胞的抑制活性

结合形态学与ITS序列分析对7株野生虫草真菌进行分类鉴定。MTT法分析它们的菌丝体醇提取物对肝癌HepG2细胞增殖的抑制活性。鉴定结果表明菌株MF7、MF9、MF14为细脚棒束孢Isaria tenuipes,菌株MF11、MF12、MF13为蝉棒束孢Isaria cicadae,菌株MF10为球孢白僵菌Beauveria bassiana;MTT结果显示分离到的3株细脚棒束孢和3株蝉棒束孢的菌丝体醇提取物对HepG2的抑制活性较差,IC₅₀均大于500μg/mL;球孢白僵菌MF10对HepG2细胞有一定抑制作用,IC₅₀值为221.6μg/mL,略强于蝙蝠蛾拟青霉发酵菌丝粉产品金水宝胶囊（IC₅₀=364μg/mL）和中华被毛孢发酵菌丝粉产品百令胶囊（IC₅₀=268.7μg/mL）。另外,发现供对比试验的3株蛹虫草菌株（MF1、MF5、MF15）对HepG2细胞均有较好的抑制作用,其中MF15的发酵菌丝体醇提取物活性最强,IC₅₀为55.56μg/mL,暗示蛹虫草发酵菌丝体具有重要的研究价值。     

Caspofungin reduces the incidence of fungal contamination in cell culture

Fungal contamination is a major problem in cell culture, and the antifungal compounds currently in use can affect cultured cells. Echinocandins are antifungal drugs that inhibit fungal cell wall synthesis by targeting an enzyme that has no counterpart in mammalian cells. We evaluated whether the echinocandin caspofungin affected the growth or morphology of six murine cell lines (a macrophage-like cell line (J774.16) and five hybridoma lines), or primary human endothelial cells. The antifungal did not influence cellular characteristics at concentrations less than 512 μg/ml, while effectively reducing the incidence of fungal contamination. Also, caspofungin did not affect the production of antibody by hybridoma cells, or alter the cytokine production of J774.16 cells, although modest increases in IL-4 and IFN-γ occurred upon LPS stimulation. Hence, echinocandins appear to be relatively non-toxic, and protect against fungal contamination in cell culture.     

Contamination by uranium mine drainages affects fungal growth and interactions between fungal species and strains

The presence of aquatic hyphomycetes has been reported for several heavy metal-contaminated waters. Tolerance probably is one adaptation to coping with heavy metals. To help clarify this issue strains of two species of aquatic hyphomycetes (Tricladium splendens Ingold and Varicosporium elodeae Kegel) were isolated from a reference stream and a stream contaminated with heavy metals and grown on malt extract agar prepared with reference and contaminated water to characterize colony morphology, growth rate, growth inhibition and interaction among species and strains. In V. elodeae the morphology of colonies differed between strains. Colony diameter increased linearly over time with growth rates being lower for strains isolated from contaminated than from reference streams (mostly for V. elodeae). Strains from the contaminated stream grew faster in medium prepared with contaminated water than in medium prepared with reference water, while for strains from the reference stream there was no significant difference in growth rates on the two media. In interacting isolates radial growth toward the opposing colony was generally lower than toward the dish edge. Percentage growth inhibition was higher for isolates in intraspecific interactions (13-37%) than in interspecific interactions (3-27%). However differences in growth inhibition experienced by interacting isolates were observed only in three cases out of 16. The difference between the percentage inhibition caused and experienced by a given isolate was highest in interactions involving isolates with distinct growth rates. Our results suggest that strains from the reference stream tolerate heavy metals while strains from the contaminated stream seem to be adapted to contaminated waters. We hypothesize that in natural environments fungal species-specific limits of tolerance to metal contamination might determine an abrupt or gradual response of the original fungal community to mine pollution giving origin to a poorer fungal community dominated by adapted strains with distinct functional efficiency.     

Production of extracellular enzymes by Antarctic fungal strains

 Thirty-three fungal strains, isolated from different sites on Victoria Land (continental Antarctica), were plate-screened for their ability to produce twelve extracellular enzymes. Lipases were generally present and in high quantities in almost all the strains. Polygalacturonase, as well as amylase and phosphatase, was common. Glucose oxidase, protease and DNAase appeared to be generally low or absent. Many strains, producing a limited number of enzymes, appeared to have a low eco-nutritional versatility while a few, such as Verticillium cfr. lecanii no. 1, V. cfr. lecanii no. 3, Aspergillus versicolor and Phoma sp. no. 2, showing a diversified enzymatic competence, are probably advantaged in extreme terrestrial environments characterized by low competition. The possibility of utilizing the enzyme-producing ability of these fungi in applied research is also discussed. Received: 3 November 1995/Accepted: 29 May 1996     

慢性阻塞性肺疾病患者继发肺部真菌感染的临床分析

目的探讨慢性阻塞性肺疾病(COPD)患者继发肺部真菌感染的临床特点。方法回顾性分析2008年1月到2010年12月安徽医科大学第一附属医院收治的COPD继发肺部真菌感染患者病例,并对其耐药情况进行比较。结果本组199例COPD患者检出白色念珠菌137例(68.84%),光滑念珠菌32例(16.08%),热带念珠菌17例(8.54%),克柔念珠菌9例(4.52%),毛霉菌3例(1.51%),清酒假丝酵母菌1例(0.50%);白色念珠菌检出率有下降趋势,热带念珠菌有上升趋势;196例真菌对伏立康唑、氟康唑、两性霉素B、伊曲康唑、氟胞嘧啶的耐药率分别为3.6%、5.1%、1.0%、8.7%和0;2008年至2010年白色念珠菌和光滑念珠菌耐药率变化差异无统计学意义。结论 COPD患者继发肺部真菌感染病原菌仍以白色念珠菌为主,其次为光滑念珠菌和热带念珠菌;白色念珠菌和光滑念珠菌耐药率无明显改变。     

Systemic expression of inflammatory mediators in patients with chronic rhinosinusitis and nasal polyps with and without Aspirin Exacerbated Respiratory Disease

BackgroundSystemic reactions are related to the pathogenesis of Aspirin Exacerbated Respiratory Disease (AERD). With this work we wanted to study the changes in the systemic levels of inflammatory mediators in both baseline and after oral aspirin challenge in patients with and without AERD.MethodsPatients with nasal polyposis and asthma with AERD (n = 20) and without (n = 18) were orally challenged with aspirin in a single-blind placebo controlled study. Serum samples and urine were collected before and 6 h after placebo and aspirin oral challenges. Serum levels of inflammatory mediators were assayed by using the Luminex technology and ELISA. The concentrations of 9-alpha, 11-beta prostaglandin F₂, and leukotriene E₄ (uLTE₄) were measured in urine samples by ELISA. The expression of T-cell surface markers was analyzed in peripheral blood mononuclear cells isolated before and after the challenges.ResultsAERD patients showed significantly higher baseline levels of s-IL-5R-alpha, uLTE₄ and percentage of CD4⁺CD25⁺CD127^pos and CD4⁺CD45RA⁻CD45RO⁺ but decreased levels of TGF-β₁ and number of CD4⁺CD25⁺CD127^neg cells. Aspirin challenge induced the release of uLTE₄, IL-6 and increased the number of CD4⁺CD45RA⁻CD45RO⁺ memory T-cells only in AERD patients but failed to reduce the levels of sCD40L as observed in non-AERD subjects. Further, IL-8 and sIL-5R-alpha levels directly correlated with the PD₂₀ASA and the effects of aspirin on IL-6 and number of memory T-cells was more pronounced in subjects showing more strong reaction (bronchial and nasal).ConclusionsAERD patients have a differential baseline inflammatory pattern that supports the role inflammation as underlying mechanism of the disease. Systemic response to oral aspirin challenge was related to an increase in serum IL-6 and the number of circulating memory T-cells in AERD patients.     

2648例股癣病原菌分离鉴定结果分析

股癣是由皮肤丝状菌所引起的一种慢性接触性传染性皮肤病。为了解股癣病原菌的流行分布情况,我们对1998—2004年来我院门诊就诊的2648例股癣患者进行了病原菌的分析,现将分析结果报告如下。