Effect of de-phosphorylation of linearized pAN7-1 and of addition of restriction enzyme on plasmid integration in Penicillium paxilli

As part of a study to investigate the pathways of plasmid pAN7-1 integration in Penicillium paxilli, a molecular analysis of 90 different integration events was carried out. Twenty out of forty five integration events analyzed from transformants obtained without the addition of restriction enzyme to the transformation reaction mixture were single-copy integrations, whereas the remaining 25 were tandem-repeat integrations. The addition of restriction enzyme resulted in a shift in this ratio in favour of single-copy integration events. Analysis of the 33 tandem-repeat integration events showed that the orientation of the plasmid copies was not random, with 88% organized as tandem head-to-tail arrays. De-phosphorylation of linearized pAN7-1 did not affect the frequency with which multiple copies were integrated. This suggests that the predominant mechanism for the generation of tandem repeats in P. paxilli is by homologous recombination rather than in vivo ligation of linearized plasmids. Received: 11 March / 6 May 1997     

产黄青霉penDE基因的克隆及其在带小棒链霉菌中的表达

penDE基因编码40k的酰基-CoA：6-APA酰基转移酶（IAT），可催化产黄青霉中青霉素生物合成途径的最后一步酶反应。利用RT-PCR法从产黄青霉中扩增得1．1kbp的不含内含子的penDE基因，并将其克隆至大肠杆菌-链霉菌穿梭质粒pUWL201，使PenDE基因位于ermEp启动子的下游，再将其转化带小棒链霉菌，获得了能够表达IAT的转化子。PDAB法和抗菌活性测定表明，所表达的IAT蛋白对6-氨基青霉烷酸和苯乙酰COA有明显的活性。     

马尔尼菲青霉病的实验室诊断研究进展

马尔尼菲青霉菌是一种双相型致病真菌,可致人类感染,多器官受累,引起系统播散性马尔尼菲青霉病(PSM)。病情凶险,如未能及时有效诊断和治疗,病死率极高。由于其临床表现复杂无特异性,诊断比较困难。为了提高PSM的早期诊断水平,近年来各国学者对PSM的实验室诊断进行了深入研究并取得了可喜进展。作者就PSM的实验室诊断进展予以综述。     

Disseminated Penicillium marneffei infection in acquired immunodeficiency syndrome: a case report

Penicillium marneffei(P.marneffei) is a facultative intracellular pathogen and the only therrmally dimorphic fugus. This fungal intection is cormmonly found in Southeast Asian,     

Hemolysin chrysolysin from Penicillium chrysogenum promotes inflammatory response

Some strains of Penicillium chrysogenum produce a proteinaceous hemolysin, chrysolysinTM, when incubated on sheep's blood agar at 37 degrees C but not at 23 degrees C. However, 92% (11/12) of the indoor air isolates produced hemolysis but only 43% (3/7) of the non-indoor air isolates did so. Chrysolysin is an aggregating protein composed of approximately 2kDa monomers, contains one cysteine amino acid, and has an isoelectric point of 4.85. Treatment of murine macrophage cell line RAW 264.7 with purified chrysolysin caused statistically significant (T-test, p < 0.05) increased production of macrophage inflammatory protein-2 (MIP-2) in a dose dependent manner after 6 h treatment. This suggests that chrysolysin might act to promote the host's inflammatory response after P. chrysogenum exposures.     

Genome Organization and Expression of the <Emphasis Type="Italic">Penicillium stoloniferum</Emphasis> Virus F

The complete sequences of three double-stranded (ds) RNAs (referred to F1, F2 and F3) of Penicillium stoloniferum virus F (PsV-F) were established. The F1 dsRNA was 1677 bp in length, and it contained one open reading frame (ORF) of 538 amino acids (molecular weight of 63 kDa, referred to P63), The F2 dsRNA was 1500 by in length, and also it contained one ORF of 420 amino acids (molecular weight of 46 kDa, referred to P46). The F3 dsRNA was 677 bp in length, but contained a small ORF with unknown function. A sequence motif of (5′-CGTAAAA-3′) was found only at the 5′ termini of the F1 and F2 dsRNAs, and a sequence motif of (5′-TAAAAAAAAA-3′) was found at the 3′ termini of all three dsRNA segments. The predicted amino acid sequence of F1 showed 38–48% sequence homology with the putative dsRNA-dependent RNA polymerases (RdRp) of dsRNA viruses, but the predicted amino acid of F2 showed no homology. Phylogenetic analysis using the RdRp sequences of the various Partitiviruses and Alphacryptoviruses revealed that PsV-F clustered well with Partitiviruses, but showed remote relationship with PsV-S. Near full-length and positive-sense single-stranded (ss) RNAs derived from the Fl, F2 and F3 dsRNAs were detected from the PsV-infected host cell. The expressed proteins of P63 and P46 showed a positive reaction against PsV-F antiserum, indicating P63 and P46 as RdRp and capsid protein, respectively. These results suggest that PsV-F can be a member of Partitivirus, but it is quite distinct from PsV-S electrophoretically, serologically and genetically, though both viruses coexist in the same cell.     

Disseminated Penicillium marneffei infection in an HIV-positive female from Thailand in Germany

We report the case of a 33 year old Thai female, who was married in Germany for eight years and used to travel to Thailand every year for several weeks. She presented with abdominal and back pain, prolonged fever, generalized lymphadenopathy, and a recent history of oral thrush. She was diagnosed HIV positive with initial CD4 counts of 18/microliter and an HI virus load of 59,000 copies/ml. Antiviral therapy was installed with zidovudin, lamivudin, and efavirenz. Abdominal CT scans revealed greatly enlarged abdominal lymph nodes. Fine needle aspirates of cervical and retroperitoneal lymph nodes, sputum samples, blood samples, and a bone marrow biopsy were microscopically positive for Penicillium marneffei and grew P. marneffei. The isolates were sensitive to amphotericin B, flucytosine, itraconazole, and fluconazole. Both universal and specific fungal polymerase chain reaction assays were positive in various samples. Serum Aspergillus galactomannan antigen, which is known to crossreact with P. marneffei, was elevated and subsequently used for monitoring of therapy. With antifungal treatment (intravenous amphotericin B 0.6 mg/kg/d for two weeks, oral itraconazole 400 mg/d for 10 weeks and 200 mg/d as maintenance therapy), the fever declined in 6 days, the size of the enlarged lymph nodes gradually decreased in the CT scans, and the initial abdominal and back pain vanished.     

真菌Penicillium sp. DT-F27次级代谢产物及其活性研究

摘要：目的 一株海洋来源真菌Penicillium sp. DT-F27次级代谢产物及其抗肿瘤活性的研究。方法 采用硅胶柱色谱、反相高效液相制备色谱等方法,对该真菌发酵产物的乙酸乙酯提取物进行分离纯化;采用波普解析方法对化合物的结构进行鉴定;采用MTT法测定化合物对人前列腺癌PC-3细胞的抗肿瘤作用。结果 从真菌Penicillium sp. DT-F27的发酵物中分离得到15个化合物,分别为麦角甾醇（1）,dehydrocyclopeptine（2）,3-O-methylviridicatin（3）,verrucofortine（4）,cyclopenin（5）,cyclo(dehydroala-L-Leu)（6）,cyclopenol（7）,4-hydroxy-2-methoxy acetanilide（8）,trans-(3RS,4RS)-3,4-dihydro-3,4,8-trihydroxynaphthalen-1(2H)-one（9）,cyclo(D-Pro-D-Val)（10）,brevianamide F（11）,cyclo(L-Pro-L-Val)（12）,anacine（13）,cyclo(L-Orn-L-Tyr-L-Orn-L-Ala)（14）,cyclo(L-Pro-L-Tyr)（15）。结论 从真菌Penicillium sp. DT-F27的次级代谢产物中分离15化合物,其中麦角甾醇（1）和3-O-methylviridicatin（3）具有较强的抗肿瘤活性,抑制率分别为45.6%和34.8%。     

海绵来源真菌黄灰青霉Sp-19中的抗肿瘤活性成分研究

目的对一株来源于羽毛山海绵Mycale plumose的真菌黄灰青霉Penicillium auratiogriseumSp-19发酵产物中的抗肿瘤活性成分进行分离和鉴定。方法采用溶剂萃取、硅胶柱色谱及制备HPLC等分离手段对该菌株发酵产物的活性部位进行了活性追踪分离,通过理化性质及波谱学手段进行化学结构鉴定,以SRB法评价了化合物的抗肿瘤活性。结果与讨论从发酵产物中分离得到5个生物碱类化合物,其结构分别鉴定为fructigenines A(1),去乙酰化fructigenines A(2),fructigeninesB(3),1,4-benzodiazepine-2,5-diones(4)和cyclopenin(5);化合物4和5在3μg.mL-1时对小鼠乳腺癌tsFT210细胞具有强的细胞毒活性。