马尔尼菲青霉病8例临床分析

目的分析总结马尔尼菲青霉病的临床及实验室特征。方法以回顾性分析的方法 ,对我科2003～2009年期间诊治的8名明确诊断为马尔尼菲青霉感染患者的临床特征、皮损组织病理学特点及其皮损组织、骨髓或咽拭子等进行真菌培养的菌落及真菌形态等实验室检查结果进行分析。结果①马尔尼菲青霉感染多发生于HIV感染患者或AIDS患者。②皮损可与全身症状同时或先后出现,皮损表现为淡红色丘疱疹、坏死性丘疹、传染性软疣样丘疹、皮肤溃疡及血痂。③多伴有多系统损害。④37℃培养呈酵母相,25℃呈菌丝相,皮损组织病理可以看到典型的"桑葚样"改变。结论马尔尼菲青霉感染好发于HIV感染患者或其他免疫功能低下的患者,常表现为特征性皮疹,皮损组织、骨髓或分泌物于25℃、37℃真菌培养结合皮肤病理是明确诊断的关键。     

交织帚枝孢霉致透明丝孢霉病国内首例

目的报道国内首例交织帚枝孢霉致透明丝孢霉病。方法患者女,69岁,主因"左前臂结节9个月"就诊。取结节边缘组织进行病理学和真菌学检查。将组织接种于沙氏培养基、脑心培养基,并对培养物进行形态学及rDNA序列测定。结果组织病理下可见组织细胞肉芽肿及透明真菌孢子和菌丝。组织真菌培养阳性,菌落生长中等速度,为浅灰色绒毛状菌落。镜下见无色分枝分隔菌丝,分生孢子梗侧生,分生孢子长圆形,薄壁,1~2个隔。经DNA序列分析,与Sarocladium implicatum相似性100%。诊断为透明丝孢霉病,予硝酸舍他康唑乳膏外用及10%碘化钾溶液口服治疗,3个月后患者治愈。结论根据其形态学特点和DNA序列分析,菌株被鉴定为Sarocladium implicatum。该菌引起的透明丝孢霉病为国内首例报道,舍他康唑及碘化钾联合治疗本病效果显著。     

淡紫拟青霉所致皮肤及皮下感染:临床和实验研究

目的 报道国内首见淡紫拟青霉所致皮肤及皮下感染患者1例。方法 取患者皮损标本多次直接镜检,真菌培养,根据真菌培养的菌落特点和镜下形态、扫描电镜及DNA序列测定鉴定致病菌种,并对分离的致病菌进行体外药敏试验。结果 患者为6个月患儿,出生12d时面部出现粟粒大暗红丘疹,脓疱,并缓慢扩大。刮取脓疱直接镜检见无色细长分支菌丝。多次培养见灰紫色羊毛状菌落生长,光镜及电镜下可见无色细长分支、分隔菌丝、直立的分生孢子梗、顶端单轮生帚状支及大量柱状或分散柱状排列椭圆形、近球形小分生孢子。根据其菌落颜色及瓶梗形态等形态学特征鉴定为淡紫拟青霉,其rRNA基因DNA测序分析证实其表型分类。动物实验证实该菌的致病性,组织相中胶质性损害为其主要病理特征。体外药敏试验提示其对特比萘芬、酮康唑较敏感,对伊曲康唑、氟康唑敏感陛较差。结论 该病例被确定为淡紫拟青霉所致的皮肤及皮下感染,为国内首例。     

马尔尼菲青霉的分类学研讨

马尔尼菲青霉能引起人和鼠类的马尔尼菲青霉病,1956年由Capponi等分离自越南的竹鼠,后由Segretain (1959)正式发表为新种.DiSalvo等(1973)首次报道此菌对人的自然感染,也分离了菌种。Pitt (1980'1979')承认Segretain的菌株为马尔尼菲青霉,但把DiSalvo的菌株(ATCC 24100)由于具明显的黄色菌丝而与岐生青霉金黄色变种(ATCC 10438)合并,成立了樱草黄青霉新种。近年来一些中国的研究者对广西的竹鼠作了大量的调查,证明竹鼠普遍携带此菌,并分离了大量菌株。作者对来自广西的34株分离物(2株来自患者,32株来自银星竹鼠)根据形态和培养特征作了鉴定,并用IMI 68794(来自模式),ATCC 24100和ATCC 10438作了对比。全部34株分离物尽管在菌落外观有明显不同(许多菌株产生黄色具饰菌丝,影响菌落外观)但根据其帚状枝及瓶梗的特征,产生紫红色素以及在370C形成酵母状细胞诸特点,作者认为都是马尔尼菲青霉。ATCC 24100也同样具有马尔尼菲青霉的特征。而ATCC 10438虽然也产生明显的黄色菌丝,但其帚状枝和瓶梗与马尔尼菲青霉不同,不产生紫红色素,在370C不生长,因此不是同一个种。ATCC 24100应保持原来的种名马尔尼菲青霉。     

AIDS合并马尔尼菲青霉病36例分析

目的探讨艾滋病合并马尔尼菲青霉病的临床表现,治疗与转归。方法回顾分析本院2008年8月～2009年8月收治的艾滋病合并马尔尼菲青霉36例。结果 36例艾滋病合并马尔尼菲青霉病平均年龄35.8岁。马尔尼菲青霉感染临床表现呈现非特异性,其中发热86.11%,贫血94.44%,GGT升高69.44%,AST升高63.89%,淋巴结肿大88.89%,脾大63.89%,低蛋白血症83.33%,咳嗽36.11%,皮损30.56%(其中典型改变仅5例,占13.89%),CD4+50cells/mm388.89%,骨髓培养(27/27)及皮损活检培养(2/2)阳性率100%,血液培养阳性率69.44%(25/36)。36例经抗真菌治疗,其中29例给予HAART治疗,28例治愈,7例好转,1例死亡。结论马尔尼菲青霉是艾滋病常见的机会性感染,早发现,早治疗,长程敏感抗真菌药物联合治疗可提高治愈率,减少复发。     

裂褶菌培养基形态学观察和DNA序列分析

目的通过观察裂褶菌在5种培养基上的生长状态、扫描电镜及DNA序列分析,了解该菌形态学及分子生物学等方面的特征。方法菌落转种于沙氏培养基(SDA),麦芽浸膏琼脂(MEA),马铃薯葡萄糖琼脂(PDA),玉米粉琼脂(CMA)和察氏琼脂(CZA)平皿培养基,27℃和37℃培养2周,观察菌落生长情况,进行扫描电镜检测及DNA序列分析。结果菌落在SDA,MEA和PDA上生长状态较好,呈蓬松白色羊毛状;尿素酶试验阳性,放线菌酮耐受试验阴性。光镜下见分支分隔菌丝、侧生的钉状突起及类水母体变异子实体。扫描电镜见菌丝分隔处闭锁联合、侧生钉状突起和泪滴状球形分泌物。经26S rDNAD1/D2区序列分析证实该菌株为裂褶菌。结论裂褶菌只有丝状型一种菌落形态;分支分隔菌丝及分隔处闭锁联合,侧生钉状突起和泪滴状球形分泌为其形态学特征;孢子由类水母状子实体产生。     

3株不同表型马尔尼菲青霉外分泌蛋白酶活性比较

目的探讨如何有效诱导马尔尼菲青霉蛋白酶分泌及比较3株不同表型的马尔尼菲青霉菌株之间外分泌蛋白酶活性差异。方法选用临床分离的马尔尼菲青霉B-6323株(P株)及其两个不同表型的突变株M/M株和M/Y株,用以牛血红蛋白为底物的酵母碳基琼脂培养基(蛋白酶诱导培养琼脂基)诱导其外分泌蛋白酶分泌。将pH值为4.0、5.6及7.2的蛋白酶诱导培养琼脂基平板打孔后分别等量接种上述3株菌,分别置于25℃及37℃培养6 d后,行考马斯亮兰G-250染色,测定各菌株周围的透亮圈(蛋白分解环)直径。结果以牛血红蛋白为底物的酵母碳基琼脂培养基可有效诱导马尔尼菲青霉蛋白酶分泌;3株马尔尼菲青霉在任意3种pH值培养基中培养,37℃培养下菌周透亮圈直径大于25℃培养下,差异经t检验有统计学意义。经方差分析,37℃,pH4.0、5.6、7.2培养条件下马尔尼菲青霉菌周透亮圈直径之间差异有统计学意义(F值为97.198,P=0.000 1),用SNK法进行两两比较发现pH4.0与pH7.2菌周透亮圈直径及pH5.6与pH7.2菌周透亮圈直径差异均有统计学意义。当温度为37℃或25℃时,在任意一个pH值下,均为M/M株的菌周透亮圈直径均数最大,其次为P株,M/Y株最小。结论马尔尼菲青霉在37℃,酸性培养条件(pH4.0、5.6)下外分泌蛋白酶活性大,且不同表型株之间外分泌蛋白酶活性有差异,M/M株及P株为高外分泌蛋白酶活性株,M/Y株为低外分泌蛋白酶活性株。     

马尔尼菲青霉菌的鉴定

为了提高马尔尼菲青霉菌的分离率和鉴定的正确率,通过培养检查特征性的菌丝和观察菌落形态,通过形态学观察其在骨髓和外周血以及PAS染色的形态。马尔尼菲青霉菌为双相菌,霉菌型菌落可发现特征性的扫帚状菌丝,沙保罗培养3 d后产生水溶性玫瑰色素;骨髓和外周血细胞内外均可发现孢子;PAS染色可见菌体呈圆形、椭圆形或腊肠状,大小不一,约为2~8μm,胞壁染红色且清楚连续,在腊肠状的细胞内可见一明显的横隔,胞质不易着色。鉴定结论:特征性的扫帚状菌丝;骨髓和外周血细胞内外均可发现孢子;水溶性玫瑰色素;PAS染色对其有鉴定价值。     

红树林免疫增强真菌的筛选及菌株PH0016的鉴定探讨

目的:探讨红树林免疫增强真菌的筛选及菌株PH0016鉴定。方法:对采集获得的红树林土壤样本进行分离和培养,选取单独的菌落群,使用分区划线法进行纯化。将选取的单独菌落群接种在海水马丁氏的培养基平板上,对接种的培养基进行环绕式烧灼以达到灭菌目的。对纯化后的菌株进行编号,置于实验菌种库中进行保存,恒温4℃。取单独的菌落,置于发酵培养基中,以每分钟250转的速度进行离心,置于恒温28℃的摇床中进行连续5天的培养。采取四甲基偶氮唑盐方法对PH0016菌株的免疫活性进行检测。观察菌株的菌落情况、形态以及r DNA ITS序列,对菌株PH0016进行鉴定。结果:本次分离出325株真菌,其中12株真菌具有免疫增强作用。通过r DNA ITS序列分析联合同源性检索,PH0016与拟青霉属的相似性达到100%。结论:菌株PH0016具有较强的免疫增强活性,能够促进外周血单个核细胞的增殖。菌株PH0016为拟青霉属丛梗孢科菌株,能够有效增强免疫功能。     

血涂片检出淋巴瘤合并马尔尼菲蓝状菌感染1例

目的通过分析淋巴瘤合并马尔尼菲蓝状菌感染的血涂片特征,探讨漏诊和误诊原因,提高诊断率。方法对1例血常规检出淋巴瘤合并马尔尼菲蓝状菌感染的血涂片、骨髓及微生物学检查结果进行分析。结果血涂片及骨髓均见中性粒细胞吞噬腊肠样物,微生物学检查呈霉菌相和酵母相双相型菌落。结论血常规检查时需注意观察仪器的报警提示并关注散点图,仔细观察血涂片,可及早发现马尔尼菲蓝状菌感染,避免漏诊和误诊。     

非人类免疫缺陷病毒感染马尔尼菲青霉病9例临床特征分析及文献复习

为探讨和总结非人类免疫缺陷病毒(human immunodeficiency virus,HIV)感染者发生马尔尼菲青霉病的临床特点,回顾性研究复旦大学附属华山医院感染科2007年1月-2017年8月收治的9例及同期发表文献中的马尔尼菲青霉病病例,分析其临床表现、实验室检查、治疗及转归。9例马尔尼菲青霉病患者的HIV检测均为阴性,5例CD4^＋ T细胞计数正常。非HIV感染马尔尼菲青霉病的起病较缓,临床表现与经典马尔尼菲青霉病类似,真菌血症较少见,病理特点以肉芽肿改变及化脓性炎症为主,诊断方法仍以培养为主(8/9),1例通过病理及二代测序技术诊断。目前,非HIV感染马尔尼菲青霉病发病率有升高趋势,部分发生于免疫正常人群,需引起临床医师的重视。     

艾滋病合并咽喉马尔尼菲青霉菌病的研究

目的研究艾滋病(AIDS)患者合并咽喉马尔尼菲青霉菌病(Penicilliosis marneffei,PSM)的咽喉表现和诊疗方法。方法收集2002年5月~2007年3月,经真菌学、血清学和组织病理学证实的5例AIDS合并咽喉PSM患者。应用抗逆转录病毒药物治疗AIDS,用两性霉素B联合氟康唑静滴治疗PSM。结果咽痛、声嘶及粘膜溃疡是AIDS合并PSM的咽喉部症状体征。5例病变组织病理学检查发现细胞内PM菌体。真菌培养分离出双相性马尔尼菲青霉菌(Penicillium marneffei,PM)。经抗真菌治疗,2例体温下降,咽喉疼痛减轻,声嘶改善,扁桃体炎症消退,溃疡面缩小,全身症状减轻。最终4例机会性感染死亡,1例仍在随访中。结论AIDS合并咽喉PSM是根据病史、临床表现和实验检查三方面作出诊断;HIV/AIDS用抗逆转录酶病毒治疗,PM用两性霉素B联合氟康唑静滴治疗。     

RNA干扰Fus3基因对二态性真菌马尔尼菲青霉菌的孢子形成和细胞壁组分合成功能的影响

马尔尼菲青霉菌Penicillium marneffei是一种重要的条件致病真菌,可致艾滋病患者产生严重的系统性霉菌病并发症。基因组学的研究和RNA干扰技术,为马尔尼菲青霉菌致病基因和致病机制的深入探讨提供了可能。我们研究马尔尼菲青霉菌的一个新基因Fus3,它是丝氨酸/苏氨酸-特异性激酶丝裂原活化蛋白激酶家族的成员。为了研究Fus3的功能,我们利用根癌农杆菌介导的双链RNA干扰技术构建了Fus3 RNA干扰菌株(Fus3-i)。RNA干扰的活性是由木糖诱导的启动子xylP控制的。Fus3基因活性下降后影响了马尔尼菲青霉菌生长,包括孢子的生成,细胞壁组分的合成。实验表明,Fus3基因对于马尔尼菲青霉菌细胞生长发育起着重要的调控作用,为研究真菌疾病提供了帮助。     

Imported penicilliosis marneffei in the United States: report of a second human infection

The first natural infection due to Penicillium marneffei in a human was reported in the United States in 1973. We describe a second case of penicilliosis marneffei that was diagnosed in Florida. In both instances, the patients had a history of travel in Southeast Asia where P. marneffei is endemic. The Florida patient had recurrent episodes of hemoptysis attributed to bronchitis and bronchiectasis. In spite of therapy with various antibacterial antibiotics for tuberculosis, the granulomatous lesions in the left upper lobe of the lungs persisted. The diagnosis of penicilliosis marneffei was established by isolating and identifying the dimorphic species of Penicillium, P. marneffei. The histopathologic features of the lung tissue included granulomata with central areas of necrosis and neutrophilic infiltration with many yeast-like, tissue-form cells of P. marneffei, which multiplied by a fission rather than a budding process.     

Factors associated with lack of awareness of HIV infection before diagnosis of AIDS.

OBJECTIVE--To identify characteristics of people likely to be unaware of their HIV infection before diagnosis of AIDS defining disease. DESIGN--Survey of continuing surveillance of voluntarily reported AIDS cases. SUBJECTS--4127 adults with AIDS diagnosed during 1989-92 and reported to the Public Health Laboratory Service AIDS Centre. SETTING--England and Wales. MAIN OUTCOME MEASURE--Lack of prolonged awareness of infection before diagnosis of AIDS, defined as an interval of nine months or less between first positive test result and diagnosis of AIDS. RESULTS--Of 3556 adults with known dates of first positive HIV test result and AIDS diagnosis, 1742 (49%) had been unaware of their infection for up to nine months before AIDS was diagnosed. Lack of awareness was independently and positively associated with infection through heterosexual contact (odds ratio 4.46, 95% confidence interval 3.15 to 6.33), AIDS reported outside the Thames regions (1.64, 1.38 to 1.96), and being non-white (1.99, 1.51 to 2.61). Women were less likely to be unaware than men (0.50, 0.33 to 0.76), and people diagnosed in 1992 were least likely to be unaware (0.48, 0.39 to 0.60). Those aged 25-49 years at diagnosis were less likely to be unaware than those aged 15-24 years and those aged 50 and over. CONCLUSIONS--People with certain characteristics are more likely than others to be unaware of their HIV infection before AIDS is diagnosed and are therefore less likely to receive prophylaxis. Methods for educating this heterogeneous group need to be investigated.     

艾滋病治疗药物-HIV抑制剂作用机制及研究进展

艾滋病（AIDS）是由人类免疫缺陷病毒（HIV）感染而引起的慢性进行性致死性传染病,又称获得性免疫缺陷综合症,目前无有效治愈的方法,严重危害着人类的健康。现今,艾滋病治疗药物主要包括逆转录酶抑制剂、蛋白酶抑制剂、进入抑制剂、整合酶抑制剂四大类化学药物和一些中草药制剂。抗HIV药物虽然不能完全治愈艾滋病,但可以控制艾滋病病情的发展,延长患者的无病生存期,提高患者的生活质量。本文就艾滋病发病机制、HIV抑制药物的抗病机制、副作用及其研究进展做一综述。     

Fifteen Cases of Penicilliosis in Guangdong,China

We analyzed the occurrence, clinical and physical features, laboratory findings, diagnostic criteria, reaction to therapy and prognosis of 15 cases Penecilliosis marneffei. Four were localized infection on skin, eleven were systemic infection, all cases were found in Guangdong province of China. We found that males were predominant in infection compared with females in 15 patients, and all patients observed had different occupations. The patients with AIDS as underlying disease were commonly drivers or unemployed. Thirteen of 15 patients had underlying diseases, i.e., connective tissue disease, aplastic anemia, kidney transplant, AIDS. Clinical features showed different symptoms, including weight loss, high fever, skin lesions, and respiratory system symptoms. Seven of 15 patients received antifungal therapy, the remaining eight patients did not receive therapy for different reasons. Nine of the patients died five recovered. Fifteen isolates were cultured from skin lesions, blood, bone marrow, peritoneum dialysis and pleural effusion and were confirmed to be Penicillium marneffei by morphology, thermal dimorphism, pathology and DNA sequence analysis. This report indicates that Penicillium marneffei may exist in nature in Guangdong province of SE China.     

Fine needle aspiration diagnosis of Penicillium marneffei infection

A disseminated infection with Penicillium marneffei, a rare human pathogen that may infect both healthy and immunocompromised patients, was diagnosed by fine needle aspiration cytology in a patient infected with the human immunodeficiency virus. The presence of yeast-form organisms with an eccentric or central dot and occasional septate and elongated forms highly suggested the diagnosis, which was confirmed on culture. Establishment of the diagnosis is important because this infection is potentially curable.