Botany and Microbiology Department, Faculty of Science, Mansoura University, Mansoura, Egypt.
Enzymology and Fungal Biotechnology Lab, Botany and Microbiology, Faculty of Science, Zagazig University, Zagazig, Egypt.
Microb Cell Fact. 2023 Aug 3;22(1):143. doi: 10.1186/s12934-023-02158-3.
Fungal producing potency of camptothecin (CPT) raise the hope for their usage to be a platform for industrial production of CPT, nevertheless, attenuation of their productivity of CPT with the subculturing and preservation is the challenge. So, screening for novel endophytic fungal isolates with a reliable CPT-biosynthetic stability was the objective. Among the isolated endophytic fungi from the tested medicinal plants, Aspergillus terreus OQ642314.1, endophyte of Cinnamomum camphora, exhibits the highest yield of CPT (89.4 μg/l). From the NMR, FT-IR and LC-MS/MS analyses, the extracted CPT from A. terreus gave the same structure and molecular mass fragmentation pattern of authentic CPT (349 m/z). The putative CPT had a significant activity against MCF7 (0.27 µM) and HEPG-2 (0.8 µM), with a strong affinity to inhibits the human Topoisomerase 1 activity (IC 0.362 μg/ml) as revealed from the Gel-based DNA relaxation assay. The purified CPT displayed a strong antimicrobial activity for various bacterial (E. coli and B. cereus) and fungal (A. flavus and A. parasiticus) isolates, ensuring the unique tertiary, and stereo-structure of A. terreus for penetrating the microbial cell walls and targeting the topoisomerase I. The higher dual activity of the purified CPT as antimicrobial and antitumor, emphasize their therapeutic efficiency, especially with growth of the opportunistic microorganisms due to the suppression of human immune system with the CPT uses in vivo. The putative CPT had an obvious activity against the tumor cell (MCF7) metastasis, and migration as revealed from the wound healing assay. The overall yield of A. terreus CPT was maximized with the Blackett-Burman design by twofolds increment (164.8 μg/l). The CPT yield by A. terreus was successively diminished with the multiple fungal subculturing, otherwise, the CPT productivity of A. terreus was restored, and increased over the zero culture upon coculturing with C. camphora microbiome (1.5% w/v), ensuring the restoring of CPT biosynthetic potency of A. terreus by the plant microbiome-derived chemical signals "microbial communication". This is the first report exploring the feasibility of A. terreus "endophyte of C. camphora" to be a preliminary platform for commercial production of CPT with a reliable sustainability upon uses of indigenous C. camphora microbiome.
真菌产生喜树碱（CPT）的能力提高了它们被用作 CPT 工业生产平台的希望，然而，随着传代和保存，它们 CPT 生产力的衰减是一个挑战。因此，筛选具有可靠 CPT 生物合成稳定性的新型内生真菌分离物是目标。在测试药用植物中分离的内生真菌中，内生真菌 Aspergillus terreus OQ642314.1，来自樟科植物，表现出最高的 CPT 产量（89.4μg/l）。从 NMR、FT-IR 和 LC-MS/MS 分析中，从 A. terreus 中提取的 CPT 具有与天然 CPT 相同的结构和分子质量碎片模式（349m/z）。从凝胶 DNA 松弛测定中揭示，所得到的 CPT 对 MCF7（0.27µM）和 HEPG-2（0.8µM）具有显著的活性，并且对抑制人拓扑异构酶 1 活性具有很强的亲和力（IC0.362μg/ml）。从基于凝胶的 DNA 松弛测定中揭示，从凝胶 DNA 松弛测定中揭示，纯化的 CPT 对各种细菌（大肠杆菌和蜡样芽孢杆菌）和真菌（黄曲霉和寄生曲霉）分离物表现出很强的抗菌活性，确保了 A. terreus 的独特的三级和立体结构，使其能够穿透微生物细胞壁并靶向拓扑异构酶 I。纯化的 CPT 作为抗菌和抗肿瘤的双重活性，强调了它们的治疗效率，特别是在 CPT 用于体内时，由于抑制了人体免疫系统，导致机会性微生物的生长。从伤口愈合试验中可以看出，所得到的 CPT 对肿瘤细胞（MCF7）转移和迁移有明显的活性。通过两倍的增量（164.8μg/l），利用 Blackett-Burman 设计使 A. terreus CPT 的总产率最大化。通过多次真菌传代，CPT 的产率会逐渐减少，但是通过与樟科植物微生物组（1.5%w/v）共培养，A. terreus 的 CPT 生产力得到了恢复，并超过了零培养水平，确保了 A. terreus 中 CPT 生物合成能力的恢复，这是由植物微生物组衍生的化学信号“微生物通讯”所导致的。这是第一个探索 A. terreus“樟科植物内生菌”作为具有可靠可持续性的 CPT 商业生产初步平台的可行性的报告，该平台可利用本土樟科植物微生物组。
