Identification and Isolation of Two Ascomycete Fungi from Spores of the Arbuscular Mycorrhizal Fungus Scutellospora castanea

Abstract

Two filamentous fungi with different phenotypes were isolated from crushed healthy spores or perforated dead spores of the arbuscular mycorrhizal fungus (AMF) Scutellospora castanea. Based on comparative sequence analysis of 5.8S ribosomal DNA and internal transcribed spacer fragments, one isolate, obtained from perforated dead spores only, was assigned to the genus Nectria, and the second, obtained from both healthy and dead spores, was assigned to Leptosphaeria, a genus that also contains pathogens of plants in the Brassicaceae. PCR and randomly amplified polymorphic DNA-PCR analyses, however, did not indicate similarities between pathogens and the isolate. The presence of the two isolates in both healthy spores and perforated dead spores of S. castanea was finally confirmed by transmission electron microscopy by using distinctive characteristics of the isolates and S. castanea. The role of this fungus in S. castanea spores remains unclear, but the results serve as a strong warning that sequences obtained from apparently healthy AMF spores cannot be presumed to be of glomalean origin and that this could present problems for studies on AMF genes.The order Glomales (class Zygomycota) comprises arbuscular mycorrhizal fungi (AMF) that are able to form mutualistic symbioses with roots of approximately 60% of all plant species (25). AMF are important for plants because they assist plants in absorbing nutrients, especially phosphate (14), and have a protective role against plant-pathogenic fungi (10, 17). It has also been shown that the diversity of AMF is an important determinant of plant diversity and productivity (26,27,28). AMF are obligate biotrophs. Their hyphae are coenocytic, and asexual spores form on the termini of hyphae (8). These spores contain hundreds to thousands of nuclei per spore (1,11,29). In the species Scutellospora castanea, one spore contains approximately 800 nuclei (11). The spores are the only form by which individual species can be identified on the basis of morphological characteristics (33).Because of the symbiotic function of AMF, there is great interest in identifying AMF genes that are involved in the establishment and functioning of the symbiosis. However, at present, very few genes other than rRNA genes have been studied in these fungi. Several studies have shown that genetic diversity of the internal transcribed spacer (ITS) exists among and within single spores (9,16,22). Six different ITS1-5.8S-ITS2 types (T1 to T6) from S. castanea were reported (8). The 5.8S sequences published by Hijri et al. (9) were subsequently used in a phylogenetic analysis of Glomales and other fungi (20). This analysis showed that the highly divergent sequences T1, T3, T5, and T6 clustered within the ascomycetes (with Phoma-Leptosphaeria as the closest relatives to T1 and T3). Other studies have also reported sequences of rRNA genes obtained from glomalean spores that clustered in the ascomycetes in phylogenetic analyses (4,12,18). The question therefore arises as to what is the true origin of these s...

Endomycorrhizal Fungi and <i>Opuntia ficus-indica</i> Seed Germination on a Lunar Regolith Simulant

“…AM fungi are Glomeromycota [4]. Their hyphae are coenocytic and the spores form terminal on them [19], which we could demonstrate in SEM studies (Figure 5(a)). Endomycorrhizal fungi penetrate the root of the plant and enter the cells where they will live as intracellular symbionts.…”

“…Similar to Phoma, the Bipolaris fungus might play a role in establishing the seedling. In agreement with Hijri et al (2002) [19], these fungi might serve as a possible weapon against non-mycorrhizal competitors. T. viridae is presently on the market as an antagonistic fungus to protect crops from soil borne diseases.…”

“…Hijri et al (2002) [19] found spores of pathogenic fungi inside the spores of an arbuscular mycorrhizal fungus without causing symptoms, which they interpreted as a possible weapon of the fungus against non-mycorrhizal competitors. The cultivation of a Glomus sp.…”

Detection of arbuscular mycorrhizal fungi,archaeospora leptoticha, and related species coloning plant roots by specific PCR primer

“…3. cation may be rare, but it is inevitable in PCR detection of AM fungi (Redecker et al 1999;Hijri et al 2002). Since the present optimization of PCR increased the sensitivity of the detection of target DNA, careful purification of template DNA and the use of a relatively small amount of template DNA were required to avoid these artifacts.…”

Endophytic fungi associated withTaxus fuana(West Himalayan Yew) of Pakistan: potential bio-resources for cancer chemopreventive agents

“…Next, the samples were cut into 5-6 pieces (2-6 mm in size) and placed on water agar plates (distilled water, 1.5% agar) and potato dextrose agar (PDA Oxoid) plates supplemented with penicillin G (100 U/mL) and streptomycin (100 lg/mL). Plates were incubated at 28 C for 2-4 weeks until growth was initiated (Hijri et al 2002). The hyphal tips emerging from the plant part were selected, purified and maintained on PDA plates for further studies.…”