Lima J S, Figueiredo J G, Gomes R G, Stringari D, Goulin E H, Adamoski D, Kava-Cordeiro V, Galli-Terasawa L V, Glienke C
Department of Genetics, Centro Politécnico, Universidade Federal do Paraná (UFPR), Box 19071, 81531-990 Curitiba, PR, Brazil.
ISRN Microbiol. 2012 May 30;2012:215716. doi: 10.5402/2012/215716. Print 2012.
In this study, we reported thirty-nine endophytic fungi identified as Colletotrichum spp. associated with Brazilian pepper tree or aroeira (Schinus terebinthifolius Raddi. Anacardiaceae) in Paraná state, Brazil. These endophytes were identified by morphological and molecular methods, using PCR taxon-specific with CaInt/ITS4, CgInt/ITS4, and Col1/ITS4 primers, which amplify specific bands in C. acutatum, C. gloeosporioides lato sensu, and Colletotrichum boninensis, respectively, and by DNA sequence analysis of the nrDNA internal transcribed spacer region (ITS1, 5.8S, ITS2). We also assayed the presence of dsRNA particles in Colletotrichum spp. isolates. Combining both morphological characters and molecular data, we identified the species C. gloeosporioides, C. boninense, and C. simmondsii. However, we found a high genetic variability intraspecific in C. gloeosporioides which suggests the existence of several other species. Bands of double-stranded RNA (dsRNA) were detected in three of thirty-nine isolates. Identity of these bands was confirmed by RNAse, DNAse, and S1 nuclease treatments for the isolates LGMF633, LGMF726, and LGMF729. This is the first study reporting these particles of dsRNA in C. gloeosporioides.
在本研究中，我们报告了39种被鉴定为炭疽菌属的内生真菌，它们与巴西巴拉那州的巴西胡椒树或胡椒木（Schinus terebinthifolius Raddi. 漆树科）相关。这些内生真菌通过形态学和分子方法进行鉴定，使用CaInt/ITS4、CgInt/ITS4和Col1/ITS4引物进行PCR分类特异性扩增，这些引物分别在尖孢炭疽菌、广义盘长孢炭疽菌和博宁炭疽菌中扩增出特异性条带，并通过对nrDNA内部转录间隔区（ITS1、5.8S、ITS2）进行DNA序列分析。我们还检测了炭疽菌属分离物中双链RNA颗粒的存在情况。结合形态特征和分子数据，我们鉴定出了盘长孢炭疽菌、博宁炭疽菌和西蒙兹炭疽菌。然而，我们发现盘长孢炭疽菌种内存在高度的遗传变异性，这表明可能还存在其他几种物种。在39个分离物中的3个中检测到了双链RNA（dsRNA）条带。通过对分离物LGMF633、LGMF726和LGMF729进行RNA酶、DNA酶和S1核酸酶处理，确认了这些条带的身份。这是首次报道在盘长孢炭疽菌中发现这些双链RNA颗粒的研究。
